The Australian Wine Research Institute Annual Report 2001

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Registered office

Waite Road, Urrbrae, SA 5064Postal Address:PO Box 197, Glen Osmond, SA 5064t (08) 83 03 66 00 f (08) 83 03 66 01

The Company

The Australian Wine Research Institute was incorporated underthe South Australian Companies Act on 27 April 1955. It isa company limited by guarantee, it does not have a sharecapital and it has been permitted, under licence, to omit theword ‘limited’ from its registered name.

The Memorandum of Association of The Australian WineResearch Institute sets out in broad terms the aims of theInstitute and the Report of the Committee of Review for theInstitute published in March 1977 identified the followingspecific aims:1. To carry out applied research in the field of oenology.2. To service the extension needs of the winemakers of Australia.3 To be involved in the teaching of oenology at both

undergraduate and postgraduate levels.4. To assume responsibility for the co-ordination of oenological

activities, and the collection, collation and disseminationof information on oenological and viticultural research tothe benefit of the Australian wine industry.

The Institute’s laboratories and offices are located on theWaite Campus of The University of Adelaide at Urrbrae inthe Adelaide foothills, on land leased from the University.The original lease is for a term of 99 years, with a right ofrenewal clause for a further 99 years. The Instituteformally affiliated with The University of Adelaide in 1990.The first buildings were erected and opened in 1957 andalterations and extensions were completed in 1976. Thebuildings have been extensively modified and refurbishedsince that time with major extensions being undertaken in1994 and 1999.

The Institute is adjacent to the Faculty of Agricultural andNatural Resource Sciences of The University of Adelaide,three Divisions of the CSIRO and the South AustralianResearch and Development Institute.

Council Members

Mr D.J. McWilliam, BSc, MScChairman—Elected amember under Clause 6(e)of the Articles of Association

Mr R.E. Day, BAgSc, BAppScElected a member underClause 6(e) of the Articles of Association

Mr R.L. Gibson, BAppSc, BAppScElected a member underClause 6(e) of the Articles of Association

Mr P.F. Hayes, BSc, BAppSc, MSc, DipEdElected a member underClause 6(e) of the Articles of Association

Professor P.B. Høj, MSc, PhDEx officio under Clause 6(d)of the Articles of Associationas Director of the Institute

Mr T.W.B. James,AssDip(WineProd)Elected a member underClause 6(e) of the Articles of Association

Mr G.R. Linton,BAppSc(AppChem),GradDip(SysAnal)Elected a member underClause 6(e) of the Articles of Association

Professor G.R. Scollary,MSc, PhD, BEd, BAppSc(Wine Science), FRACICharles Sturt UniversityRepresentative under Clause 6(c) of the Articles of Association

Professor M. Sedgley, PhDThe University of AdelaideRepresentative under Clause 6(b) of the Articles of Association

Dr R.R. Walker,BAgSc(Hons) PhDCSIRO Representative underClause 6(a) of the Articles of Association

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Chairman’s report

Council notes

Director’s report

Staff

Highlights of the year

Staff activities

Research Teams’ reports

Appendixes 1–4

Abridged financial report

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46th Annual Report

Presented to the

Australian Wine Industry

30th June 2001

Table of contents

It is my pleasure to again present my Chairman'sReport for the past year's activities of TheAustralian Wine Research Institute. This pastyear has been full of activity and outstandingprogress with results now issuing from suchprojects as the cork closure trial, the potentialenhancement of oak flavour during bottleageing and the ability of NIR to rapidlymeasure berry colour and other qualityparameters. This last project has generatedenormous industry interest and has not onlyseen generous industry support from some ofour major companies but has now progressedfar enough to be trialled with confidence atthe winery weighbridge. The NIR projectstarted as a collaborative project between theInstitute and BRL Hardy but has now beenincorporated in the new Cooperative ResearchCentre for Viticulture's programs and alongwith the direct industry investment has ledto a strong industry desire to progress thedevelopment of affordable instrumentationthat can be readily taken up by industry. Thecommercialisation of the IP associated withthis technique needs to be rapidly implementedand it is hoped to obtain prompt and satisfactoryagreement with all vested interests.

The closure trial, now more than 18 monthsold, has revealed significant differences in theperformance of different closures as outlinedin the publication which was due to appearin the July issue of the Australian Journal ofGrape and Wine Research. What is mostimportant to recognise is that the testingprotocol and the individual results have to beevaluated from each winemaker's perspectiveof what performance factors they deemimportant for their individual situation. TheInstitute trial was a massive undertakinginvolving considerable cost and resourcesand only provides a ‘snapshot in time’ of the

rapidly changing and advancing technologyof closure design.

One of our aims has been to strengthen thechemistry side of our activities and we havebeen fortunate to have Dr Markus Herderichjoin our Institute staff. Markus undertook postdoctoral studies at our Institute in 1994-95and has strengths in flavour and phenolicchemistry. Other major staff changes are theretirement, after 14 years service, of ourSecretary, Janet Currie. She not only kept atight reign on the financial affairs of ourInstitute but was a counsel and confidant toall of the staff. We now have a most suitablereplacement and welcome Hans Muhlack tofill the vacancy created by Janet’s retirement.

The new extensions to the Institute are fullyoccupied and Analytical Services has continuedto expand their range of services offered tothe industry during the last year. The Instituteurgently needs to address the renovation andextensions of the old chemistry lab. With astronger emphasis on flavour and phenolicchemistry, these facilities are inadequate forthe future needs of both equipment and staff.

It is pleasing to see that the Grape and WineResearch and Development Corporation isproposing to move to a ‘rolling’ funding cyclewhich will remove some of the uncertaintyof the current annual funding. It has been ofmutual benefit to have their representativeattend our Council meetings and I thank themfor their advice, interest and involvement.

I also thank all members of Council for theirdiligence and efforts with a special thank youto Professor Margaret Sedgley from TheUniversity of Adelaide who retires in favourof Professor Steve Tyerman, the newly

appointed Professor of Viticulture. Thank youto all the staff who carry the Institute's messagesto industry so well via the forums of roadshows,seminars, workshops, lectures and ourpublication, Technical Review. Congratulationsto our Communication and Publicity Managerand the Library staff for the implementationof the on-line searchable database of theJohn Fornachon Memorial Library.

Finally, I offer my thanks to the Director,Peter Høj, and all of his staff for a successfulyear. It has indeed been a pleasure for me tohave been associated with the Institute andits employees in my capacity as Councilmember and Chair. I urge all members ofour industry to not only continue, butincrease, their interest and support for thiswonderful asset we all share.

Doug McWilliamChairman of Council

Chairman’s Report

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Doug McWilliam, Chairman of Council

ChairmanAt the Council Meeting held on 30October 2000, Mr D.J. McWilliam wasre-elected Chairman of Council.

Members of the Executive CommitteeMr D.J. McWilliamMr R.L. GibsonMr P.F. HayesProfessor P.B. HøjMr T.W.B. JamesProfessor M. Sedgley

Deputy Members of CouncilMr N.P. BlieschkeMr L.P. DeansDr P.R. DryMr J. NortheyDr N.S. ScottDr A.M. SmithMr P.J. Wall

MeetingsOrdinary General MeetingThe 46th Ordinary (Annual) GeneralMeeting was held on 30 October 2000.

CouncilThe Council of the Institute met on the following dates: 25 July 2000, 30October 2000, 6 March 2001, 1 May2001. Executive members of Councilmet on 5 December 2000.

FundingThe Council of the Institute acknowledgesthe continuing financial support of theGrape and Wine Research andDevelopment Corporation.

AppreciationThe Institute acknowledges the assistanceand cooperation of the followingorganizations throughout the year:

Adelaide UniversityAustralian Wine and Brandy CorporationAustralian Wine FoundationCharles Sturt UniversityCommonwealth Scientific and IndustrialResearch Organization (CSIRO)Cooperative Research Centre forViticultureDepartment of Agriculture, Forestry and Fisheries AustraliaSouth Australian Wine and Brandy Industry AssociationState Departments of AgricultureWinegrape Growers’ Council ofAustralia, Inc.Winemakers’ Federation of Australia Incorporated

Council notes

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Left to right:

Peter Hayes, Robin Day, Geoff Linton, Doug McWilliam, Margaret Sedgley,Richard Gibson, Steve Tyerman, Tim James, Peter Høj.

The continued growth of the Australian wineindustry and the increase in research levies, witheffect from the 1999 vintage, has allowedfor an expansion in the total research portfoliomanaged by the Grape and Wine Researchand Development Corporation (GWRDC) and ,in turn, for an expansion of funding to awide range of different research organisations.The funds allocated directly and indirectly tothe Institute have also increased in dollar-terms and now constitute about 35% of thetotal matched levy funds administered by theGWRDC and an estimated 16-18% of publicexpenditure on grape and wine R&D inAustralia. We are appreciative of this fundingas it has widened the scope of our activitiesand further reduced our already very low percapita running costs to service the needs ofindustry better. These activities will, in turn,help underpin sustained growth of wineindustry activities with consequential flow-onbenefits to local communities.

Tangible outcomes, about which much iswritten in the remainder of this AnnualReport, include the conduct, data collection,analyses, writing up and acceptance forpublication in the Australian Journal of Grapeand Wine Research of the Institute’s seminalclosure trial, ably led by our Manager ofIndustry Services (Peter Godden) with strongsupport from our sensory science specialist(Leigh Francis) and the dedicated members of

our trained sensory panel. This event precipitatedunprecedented national and internationalmedia interest. Throughout this process,relevant Institute staff conducted themselveswith a high degree of professionalism in anarea which, due to its commercial significance,had to be navigated with a great deal ofdiscipline and thought. Another highlightrelates to the much awaited installation ofsix computer-controlled Potter fermentorsand six rotofermentors in the recently builtHickinbotham-Roseworthy Wine ScienceLaboratory at the Waite Campus of AdelaideUniversity. This facility and its eventualcompletion is a prime example of howrelentless belief in a cause and distinctcontributions by a range of determinedindividuals can lead to long term gains whereothers might have given up due to lack ofsupport. The Institute is most appreciative ofthe University of Adelaide’s decision to buildthis facility and indeed helped start the ballrolling by pledging a $250,000 contributionover five years starting in July 1997; acommitment we now have met fully due tothe strong performance of our own AnalyticalService and its fourteen staff. In return forthis significant contribution, we have beengranted access to the Facility for 25 years.The first such substantial access occurredthrough a collaborative effort in our tanninproject where eight tonnes of fruit werefermented in triplicate with different

hardware and different yeasts. The wines arenow subjected to detailed sensory analysesand outcomes will be communicated toindustry upon its completion and presentedto industry in workshops conducted at the11th Australian Wine Industry TechnicalConference in October 2001.

The 11th AWITC will be the largest andprobably most diverse ever conducted. TheConference Planning Committee is Chairedby the Institute Director, the Program PlanningCommittee by Southcorp’s NationalViticulturist (Peter Hayes) while the detailedmanagement is undertaken in a highlyprofessional manner by the Institute’s Rae Blairand her Secretariat Staff. This triennial eventhas grown to become a major Institute activityin the area of knowledge dissemination andadoption. It involves detailed planning,coordination and management of nearly1700 delegates, the staging of 72workshops (coordinated by our IndustryServices Personnel) and arranging some10,000 m2 of trade exhibition comprising 178 individual exhibitors.

The selected examples of Institute activitiescited above illustrate the continuedcommitment of our staff to the wineindustry which, in many cases, extendsbeyond what can reasonably be expectedand sustained in the longer term without

Director’s Report

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Peter Høj

increased funding to employ more staff andupgrade infrastructure significantly. TheInstitute is working actively to help itself inthis regard. One such example relates to thelarge extension of Institute facilities weundertook in 1998. That $1.1 million extensionwas facilitated by an injection of $375,000over three years from the GWRDC andotherwise funded by savings as well as a largecommercial loan, the latter on track to bepaid back during the financial year 2001/2002.

Notwithstanding our past Institute expansionsand the construction of the HickinbothamRoseworthy Wine Science Laboratory byAdelaide University, more paint is needed tocomplete the picture, chiefly due to the needto accommodate staff and an increasingnumber of overseas visitors as well asacquiring new equipment for grape andwine research. The Institute, with very strongsupport from CSIRO Plant Industry, therefore,acted as a proponent agent in what laterturned out to be a successful bid to theFederal Government for funding of $4.5million under the new and once off ‘MajorNational Research Facility Schemes’ (MNRF).This will lead to a $7,000,000 upgrade offacilities at both the Waite Campus of theUniversity of Adelaide and, to a lesserextent, at the Ron Potter Centre of CharlesSturt University. Throughout the bid process,we enjoyed strong support from the South

Australian Government (including a$700,000 cash contribution) and fromCouncil members who endorsed the Directorto commit to a further commercial loan inorder to be in the race. A special mentionshould go to Mr Robin Day who, on severaloccasions, gave up his own time tocontribute so strongly to our bid as anindustry representative. The conduct and pursuit of the MNRF bidwas a heartening exercise where institutionscooperated with good-will and a minimumof fuss to generate an outcome not possibleto achieve by any agent alone. Clearly, wehave a challenge to put in place areasonable MNRF management structurewhich is effective and user-friendly, but weare confident this will happen usingstructures which have been proven over thelong term. In parallel with our quest tosecure a better funding base for ourresearch, we must also identify costs (andthose potentially hidden) associated withvarious models for its distribution,management and execution. ThroughoutAustralian research management, thereappears to be a case for identifyingexcellence, fund it appropriately and then letthe individuals get on with the job within aframework defined robustly by majorstakeholders, in our case the GWRDC, theAustralian wine industry and thecommunities benefiting from its success.

On a more positive note, the Institute, inaddition to the outcomes cited above, againlooks back on a year with achievements in adiverse range of areas including ourtrendsetting NIRS work, the protein basedfingerprinting of grape varieties and theapplication of our unique tannin mouthfeelwheel to the desciption of the mouthfeelproperties of chemically defined tannins.Only time will tell if any of thesedevelopments rate against the impressivework of past illustrious Institute employeessuch as Fornachon.

Present and past achievements would nothave been possible without the strongsupport of dedicated wine industryindividuals, staff and collaborators. TheInstitute is most appreciative of this muchneeded support and is looking forward to along partnership in our quest to serve theAustralian wine industry’s future increased

R&D needs. A special thanks must go to JanetCurrie, our long-serving Company Secretarywho, over a period of 14 years, steered thefinancial activities of the Institute without aflinch and allowed us to expand the Institutein a sustainable fashion through several phases.We wish Janet all the best in her retirement.

We must never forget that our endeavor toremain innovative is dependent on ourcapacity to recruit and further developtalented individuals within all aspects of thewine industry. The Institute is committed topartake in this undertaking with the wineindustry as our partner and joint success asthe outcome.

Peter HøjDirector

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Peter Bordier Høj, MSc, PhD, Copenhagen, Director

Research team members

Paul Anthony Henschke, BSc(Hons) PhDUAdel., Principal Research Microbiologist

Mark Aidan Sefton, BSc(Hons) PhD UWA,Principal Research Chemist

Elizabeth Joy Waters, BSc PhD UAdel.,Principal Research Biochemist

Ian Leigh Francis, BSc(Hons) Monash, PhDUAdel., Senior Research Chemist

Robert George Dambergs, BSc(Hons)UAdel., PhD UQld, Senior ResearchChemist (commenced 6 February 2001)

Markus Johannes Herderich, PhDUWuertzberg, Senior Research Chemist(commenced 9/4/01)

Yoji Hayasaka, DipEng(IndChem) Tokyo I.T.,MPharm Vic. Col. Pharm., CertIntBusMgtMonash, Manager—Mass Spectrometry Facility

Miguel Antonio de Barros Lopes, BScOregon, PhD UC (Santa Barbara),Molecular Biologist

Eveline Jutta Charlotte Bartowsky,BSc(Hons) PhD UAdel., Research Microbiologist

Zhong Kui Peng, BSc MSc, Zhejiang, PhDUAdel., Research Chemist

Alan Percy Pollnitz, BSc(Hons), PhD UAdel.,Research Chemist

Gordon Michael Elsey, BSc(Hons), PhD,Flinders, Postdoctoral Research Fellow

Michael Brian Esler, BSc(Hons) USydney, PhDUWollongong, Postdoctoral Research Fellow

George Kyriakos Skouroumounis,BSc(Hons), Flinders, PhD UAdel.,Postdoctoral Research Fellow

Stéphane Vidal, Eng Dip. (Biochem), Nat.Instit. App. Sc. Lyon, MBiochem, UClaudeBernard Lyon, PhD UJoseph FourierGrenoble, Postdoctoral Research Fellow

Kenneth Frank Pocock, BAppSc UAdel.,FAIFST, Senior Chemist

Michael Raunkjaer, MSc Copenhagen, VisitingResearch Chemist (concluded 1/12/00)

Anders Håkansson, MSc Copenhagen,Visiting Research Chemist (commenced

Dr Encarnacion Gomez Plaza, Chem, PhDMurcia Visiting Research Chemist (twomonth stay, commenced 23/10/00)

Tracey Ellen Siebert, BSc UAdel., Chemist(commenced 14/3/01)

Peter James Costello, BSc (Hons) MScUNSW, PhD UAdel., Microbiologist

Holger Gockowiak, BSc(Hons) UAdel.,Microbiologist (concluded 13/11/00) Part timeLaboratory Manager (commenced 16/4/01)

Jeffrey Mark Eglinton, BSc(Hons) UAdel.,Microbiologist/Senior Computer Systems Officer

Dimitra Capone, AssDip(Chem) USA,Laboratory Technician

Maria Jolanta Kwiatkowski, MSc Gliwice,Laboratory Technician

Simon Dillon, BSc(Hons) Flinders, ResearchAssistant (commenced 11/9/00)

Bianca van Wegen, BEnvSc(Hons) Flinders,Part-time Research Assistant (commenced 21/11/00)

Wieslawa Cynkar, BSc PhD Wroclaw,Technical Officer

Gayle Ann Baldock, BSc(Hons) Guelph,Technical Officer/Casual Analyst

Jennifer Bellon, Part-time Laboratory AssistantJane Melissa McCarthy, AdCertMedLabSc

USA, CertVetNurs, CertAnimHand TAFE,Technical Assistant (commenced 21November 2000).

Shauna Liam Brown, BBiotech(Hons)Flinders, Postgraduate Student

Anthony John Heinrich, BBiotech(Hons)Flinders, Postgraduate Student

Anita Oberholster, BSc(Hons), Stellenbosch,Postgraduate Student (19/4/01)

Kate Susan Howell, BSc(Hons) UNSW,Postgraduate Student

Kerry Leigh Wilkinson, BSc(Hons) Flinders,Postgraduate Student

Carolyn Jane Puglisi, BSc Flinders,BSc(Hons) UAdel., Postgraduate Student

Agnieszka Janusz, BSc(Hons) Flinders,Postgraduate Student (commenced 26/3/01)

Heather Eunice Smyth (nee Pain),BSc(Hons) UAdel., Postgraduate Student(commenced 2/4/01)

Nadia D’Incecco, BAgSc(Food Tech)(Hons)Padova, Visiting Italian PostgraduateStudent (concluded 6/2/01)

Paolo Sivilotti, BAgSc Udine, Visiting ItalianPostgraduate Student (commenced8/11/00, concluded 31/5/01)

Teresa Girbau Solà, BPharm, DipOphthamBarcelona, Visiting Spanish PostgraduateStudent (commenced 14/2/01, concluded3/8/01)

Sandra Hoffman, Visiting GermanPostgraduate Student (commenced 18/6/01)

Industry Services team members*

Peter William Godden, BAppSc (WineScience) UAdel., Winemaker/ManagerIndustry Services

Mark Gishen, BE(Chem)(Hons) MEngSc(Chem)UMelb., Quality Liaison Manager

Sally-Jean Bell, BSc(Hons) UWA, GradDip(Wine)Roseworthy, PhD UWA, Viticulturist

Adrian Dermott Coulter, BSc Flinders,GradDipOenol, UAdel., Oenologist

Peter Valente, BSc(Hons) Flinders, ChemistElla Margaret Clare Robinson, BA,

BSc(Hons) UAdel., ChemistRaelene Joan Blair,

CertAppMgt(Marketing) AIM,Communication and PublicityManager/Personal Assistant to the Director

Creina Standish Stockley, BSc(Hons)UAdel., MSc Flinders, MBA USA, Healthand Regulatory Information Manager

Catherine Grace Daniel, BA ANU,GradDip(Lib) RMIT, Librarian

Ingrid Betty-Maud Oats, DipLibInfo Adel.Tafe, Library Technician

Jodi-Ann Slade, Library Assistant(concluded 20/2/01)

Anna Capone, Library Assistant(commenced 20/2/01)

*Several members of the Industry ServicesTeam lead or take part in a number ofresearch projects.

Analytical Service

Donald Robert Buick, BSc UAdel., GradDip(BusAdmin) USA, AIFST, Manager –Analytical Service

John Benjamin Hughes, DipWineMrktgUAdel., Analytical Service Supervisor –Administration

Matthew Grant Holdstock, BSc Flinders,Analytical Service Supervisor – Laboratory

Gregory Andrew Ruediger, BAppSc SAIT,GradDipOenol, UAdel., Trace AnalysisLaboratory Supervisor

Kevin Herbert Pardon, AssDip(AppChem)SAIT, Analyst

Andrea Dale Kemp, AssDip(Farm Mgmt),Roseworthy, BA, UAdel., Casual Analyst

Randell Leith Taylor, BSc(Hons) UAdel.,Casual Analyst

David Rolfe Boehm, BSc UAdel., Casual AnalystAmanda Louise Cook, AdvCert (Lab Tech)

Mackay, Casual Laboratory TechnicianRadka Kolouch, AssDip(Food Tech) Czech

Republic, Casual Laboratory TechnicianMatthew James Cream, Casual

Laboratory TechnicianJeremy Crispin Hack, Casual

Laboratory TechnicianBao Tran, AssocDipChemTech USA, Casual

Laboratory TechnicianSandra Lloyd-Davies, BA Flinders, Casual

Laboratory Administration Assistant

Administration

Janet Currie Currie, BA Glasgow, MAICD,Company Secretary (retired 15/1/01)

Hans Engelbert Muhlack, BEc Adelaide,CPA Aust., Company Secretary(commenced 3/10/00)

Sita Soetratma, B.Bus(Mgmt) USA,Accountant (concluded 8/12/00)

Rachel Lee Edwards, Accountant(commenced 13/12/00)

Heather Margaret Donnell, Secretary tothe Director

Carolyn Debra Grant, AssDip(Acc)Panorama TAFE, Secretary to the CompanySecretary/Conference Secretariat

Narelle Elizabeth D’Costa,Administration Officer

Emma-Kate Darling, ReceptionistMaria Concettina Mills, Casual ReceptionistElizabeth Morgan, Administration SupportBranka Zepina, Cleaner (commenced 12/3/01)

Staff

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The outcomes of the Institute’s majorwine closure trial were analysed 20months post-bottling and submitted tothe Australian Journal of Grape and WineResearch for publication with expectedpublication of the 42 page article in midJuly 2001.

Flavour scalping studies showed a very largedifferential in the rate of disappearanceof known wine aroma compounds whenstored in ‘bag in box’ containers.

The terminology developed by our sensorypanel and collaborators to characterisethe mouthfeel of red wines is gainingacceptance as a ‘mouthfeel wheel’ byresearchers and winemakers worldwide.

Gram quantities of nine different tanninfractions from grape seeds and grapeskins for sensory evaluation were isolatedand characterised using HPLC andelectrospray mass spectrometry followedby sensorial assessment by a tastingpanel over an eight week period.

Combined laboratory and industry-basedwinemaking trials, coupled with chemicaland quantitative sensory descriptiveanalysis, continued to demonstrate thepotential of S. bayanus, Candida andnew hybrid yeast as positive modulatorsof wine sensory characteristics.

A laboratory model system for predictingthe effects of fermentation yeast onmalolactic bacteria has been developedto enable selection of the most appropriatecombination of strains to provide greaterreliability of malolactic fermentation.

The Institute secured strong privatefinancial support from Lallemand toboost our research on the effect of wineyeast on red wine phenolic compounds.

The Institute paid its final instalment of its$250,000 donation to the HickinbothamRoseworthy Wine Science Laboratory andconducted a series of triplicated fermentson a commercial scale to study the effectof winemaking practices and yeast strainson colour, mouthfeel, flavour and aromaof both white and red wines.

Successful organic synthesis of a high-yielding precursor to the ubiquitous wine flavor compund ß-damascenonewas achieved as a first step to understandits formation from non odoriferous grape precursors.

Organic synthesis disproved the structuralassignment of an oak lactone precursorwhich has been assumed for more than20 years.

Our Analytical Service increased itsturnover and recorded an increase incontract research as well as a very largedemand for analyses for 4-ethylphenol,an indicator of Brettanomyces activitywhich may be more widespread thaninitially assumed.

A double modification of a laboratoryyeast by means of molecular biologygives a remarkable demonstration of thepower of the technology and it isdemonstrated that the genomes ofcommercial wine strains are quitedifferent to those of laboratory strains.

Formal ‘Roadshow’ visits were made toSA and Vic in November and December2000 (see Appendix 1). Senior Institutestaff presented seven full-day seminars,each seminar consisting of a minimum oftwelve presentations focussing on currentareas of Institute research, or topicalissues in the wine industry.

Institute staff authored/co-authored 37publications; gave 35 seminars/talks(excluding the Roadshow seminarsabove); presented 11 workshops; presented3 posters; gave 38 lectures and supervised36 students during the year.

Institute staff hosted more than 113international visitors to the Instituteduring the year including five visitingscientists on extended stay.

The new website of The Australian WineResearch Institute @ www.awri.com.auwas launched including an extensiveeducational/troubleshooting section onwine hazes and deposits.

Requests for information from the JohnFornachon Memorial Library staffincreased by 108% over requestsreceived in 1999/00.

The 14th Advanced Wine AssessmentCourse was held in July 2000, givinganother 30 participants the opportunityto develop and test their sensoryevaluation performance. This was thesecond Course presented under a four-day format, which includes over 40 hoursof activities over the four days and acontribution by more than ten seniorAustralian wine show judges.

Institute staff responded to more than4,400 enquiries in relation to wineindustry matters.

The human capital of the Institute is furtherenhanced by qualification of one additionalstaff member for the PhD degree, byqualification of one additional staff memberfor the Postgraduate Diploma in Oenology,by recruitment of a distinguished Germanscientist to lead our tannin project and bythe recruitment of a PhD qualified chemistwith almost two decades of experience inthe wine industry.

Highlights of the year

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In addition to undertaking research and otherprojects described in this report, the Instituteperforms a large number of external activitiesin support of the Australian wine industry.

Information on seminars, talks and posterpapers given to outside organizations,academic lectures delivered, graduatestudents supervised, and the papers publishedis tabulated and can be found in Appendices1–4 of the Annual Report. Activities in additionto those in the Appendices are described below.

Peter Høj is a member of the following:Prime Minister’s Science, Engineering andInnovation CouncilSouth Australian Innovation, Science andTechnology CouncilInternational Trade and Technical AdvisoryCommittee (AWBC)Compliance and Technical AdvisoryCommittee (AWBC)Technical Committee (Winemakers’Federation of Australia)Wine Committee (Royal Agricultural andHorticultural Society of South Australia)Waite Campus Management CommitteeCommittee of Management, ViticulturalPublishing, publisher of Australian Journalof Grape and Wine ResearchEditorial board of the Journal Internationaldes Sciences de la Vigne et du VinConference Planning Committee of theEleventh Australian Wine IndustryTechnical Conference (7-11 October2001, Adelaide) (Chair)Australian Organising Committee of the26th World Congress and 81st GeneralAssembly of the Office International de laVigne et du Vin (OIV) (11-18 October2001, Adelaide) and is Section Presidentof the Oenology Sub-Commission for theAustralian OIV Congress.

He is also the Institute’s representative onthe following Adelaide University’sBoards/Committees:

Faculty BoardDepartment of Horticulture, Viticultureand Oenology Advisory CommitteeManagement Committee of theDepartment of Horticulture, Viticultureand Oenology

Hans Muhlack is the Public Officer of theAustralian Wine Industry TechnicalConference Inc.

Rae Blair is a member of the ConferencePlanning Committee and is the Treasurer andConference Manager of the EleventhAustralian Wine Industry TechnicalConference. She is also a member of theAustralian Organising Committee of the26th World Congress and 81st GeneralAssembly of the OIV.

Creina Stockley is a member of theAustralian Wine and Brandy Corporation’sLegislative Review Committee, a member ofthe Technical Committee of the Winemakers’Federation of Australia, a member of theNational Reference Committee –Environment Strategy Development (SouthAustralian Wine and Brandy IndustryAssociation) and was elected Vice-Presidentof the Nutrition and Wine Expert Group forthe Australian OIV Congress.

Elizabeth Waters is Leader of Program 1 of theCooperative Research Centre for Viticulture II.

Mark Sefton is on the editorial reviewboard of the International Journal of Vineand Wine Sciences and is the project leaderof project 1.3 of the Cooperative ResearchCentre for Viticulture II.

Leigh Francis is an Editorial Board memberof the Journal of the Science of Food andAgriculture, Editorial Review Board memberof the American Journal of Enology andViticulture, Chairman of the Royal AustralianChemical Institute (SA Branch), Medicinaland Agricultural Chemistry Group.

Ken Pocock serves on the Waite CampusOccupational Health and SafetyCoordinating Committee.

Paul Henschke serves on the EditorialReview Board of the following journals:Australian Journal of Grape and WineResearch; American Journal of Enology andViticulture; and Australia and New ZealandWine Industry Journal.

Peter Godden is a member of theConference Planning and Program sub-Committees and is the WorkshopCoordinator of a program of 72 workshopsto be held at the Eleventh Australian WineIndustry Technical Conference.

Matthew Holdstock is a member of theInterwinery Analysis Group Inc.

Visitors to the InstituteAustralia

Eric Parnis and Neil Grant, SouthAustralian Department of Industry andTrade (1 August 2000)Linda Lee, Hong Kong Trade DevelopmentCouncil, Sydney (2 August 2000)Matthew Wingfield, International SouthAustralia (2 August 2000)Dr Erika Winter, Agriculture Victoria (3August 2000 and other occasions)Group of 30 Advanced Viticulture studentsfrom South Eastern TAFE, accompaniedby Denis Vice (3 August 2000)Linda Bowes and Brian Smedley, SAWBIA(18 August 2000, and other occasions)

Senator Judith Troeth, ParliamentarySecretary to the Minister for Agriculture,Fisheries and Forestry, Australia (25August 2000)Michael Alder, Agriculture, Fisheries andForestry, Australia (25 August 2000)David Monck, South Australian Centrefor Manufacturing, Marilyn Arnold,Flavour South Australia, The FoodIndustry Association Inc. (28 August 2000)Sandy Grieve, Australian Council ofAgricultural Journalists Inc (4 September 2000)Jenny Eichler and four Year 11 ChemistryStudents from Murrayville College (6 September 2000)Members of the Executive Committee of theAustralian Academy of Technological Sciencesand Engineering (6 September 2000)Ross Dobinson and Tim Hoban,Technology Structuring Limited (7 September 2000)David Hall and Dr Deann Glenn, Grapeand Wine Research and DevelopmentCorporation (on several occasions)Richard Heinemann, The Business Centre(14 September 2000)H-Michael Schwandt, Consul-General,Federal Republic of Germany, Pat Vaughan,Commonwealth visit co-ordinator,Department of the Prime Minister andCabinet, Nektarios Tsirbas, Policy Officer,Department of Agriculture, Fisheries andForestry, Thomas Kruckemeyer,Interpreter, SA, Dean Dempsey, DeputyManager, Protocol, Department of thePremier and Cabinet, Peter Hayes,Rosemount Estates (3 November, 2000)Juan Bournas, Trade Commissioner ofChile and Kees van Haasteren, AgTour(15 November 2000)Noel Hayes, Amorim (20 November 2000)Olga Kostic, International MarketingManager and Brian Dolling, ManagingDirector, NuKorc (22 November 2000)Peter Flinn, Agriculture Victoria (22November 2000)Tom Angove, Angove’s Wines (2 January 2001)Andrew McPherson, Andrew Dean, HamishMacGowan, Leigh Clarnette, Guida Vazzoler,McPherson Wines; Alister Purbrick, AllanGeorge, Neil Larson, Chateau Tahbilk;Geoff Merrill, Scott Heidrich, Joe DiFabio,Geoff Merrill Wines (2 February 2001)Tony Royal, Seguin Moreau (12 February 2001)Michael Burleigh, Foss Pacific (28 February 2001 and 24 April 2001)Cecil Camilleri, Yalumba Wines; Alex Sas,BRL Hardy; Wendy Allan, PrudenceHonner, Southcorp Wines; Peter Hayes,Rosemount Estate; Prue Henschke, C.A.Henschke & Co; David Bruer, TempleBruer; Russell Johnstone, OrlandoWyndham Group; Vic Patrick, Syd Kyloh,Blass Beringer; John Sabine; AndrewBenger; Lou Cremasco; Geoff Hardy,

Staff Activities

8

Pertaringa Vineyards; James Altmann,Fruit Doctors; Nic Fazekas, IAMA; BillMurray, IHD, Leon Redunz, AvCare,Bayer; Keith Jones, South AustralianWine and Brandy Industry Association;DeAnn Glenn, Grape and Wine Researchand Development Corporation; MattCahill, Dow AgroSciences; StephenStrachan, Winemakers’ Federation ofAustralia; David Baker, David Braybrook,Cooperative Research Centre forViticulture; Robyn van Heeswijck, AnuKuma, Department of Horticulture,Viticulture and Oenology; Trevor Wicks,South Australian Research andDevelopment Institute; Ary Hoffman,LaTrobe University; Peter Merriman,Alison MacGregor, Rob Sward, MarkKrstic, M. Mollah, Department of NaturalResources and Energy; Rai Kookana, RayCorrell, CSIRO (27 February 2001)Natasha Mitchell, ABC radio (12 March 2001)Imre Lele, Bottle Magic and Ken Craig, SACentre for Manufacturing (13 March 2001)Duncan MacDonald, Rod Pemberthy,Sanjay Sinha, Burns Philp (14 March 2001)A/Professor Graham Bell, Director,University of New South Wales, Centre forChemoSensory Research (15 March 2001)Rob Lewis, South Australian Research andDevelopment Institute and Nigel Scott CSIRODivision of Plant Industry (26 March 2001)Dr John Radcliffe, CSIRO (27 May 2001)Jennifer Gardiner, Adelaide University (27 May 2001)Libby Boschen, WINETAC (25 May 2001)B. Sara, C. Belperio, T. Mitani, K.Ringwood, M. Wong, L. Pattison, R.Elfenbein, M. Trewartha, FoodTechnology students from the RegencyInstitute of TAFE (31 May 2001)Robert Druscetta and Anne Patterson,Australian Wine and Brandy Corporation(28 June 2001)Don Woolman and Paul Clancy,Australian Wine Industry Suppliers’Association (29 June 2001)Cooperative Research Centre for ViticultureBoard members: G. Scollary, D. Gall, D.McWilliam, P. Barnes, T. Martin, J. Dal Bro,P. Kriedemann, P. Mansfield (4 July 2001)Professor Edwina Cornish, AdelaideUniversity (5 July 2001)

ChileAgapito Garcia Vidal; Ximena Torti Solar,Agricola San Diego de Puquillay Ltda;Guillermo Achurra Montes, AgricolaTunquelen Ltda; Jorge Luis Ollé Cordero,Agricola y Comercial Crucesillas Ltda;Carlos Valenzuela Olave; Patricio AzocarMonasterio, Cooperativa AgricolaPisquera Elqui Ltda; Carlos Alfredo TerresSilva, Cooperativa Agricola Vitivinicola deCurico Ltda; Diego Garcia de la Huerta,Juan Sutil Servoin; Maria Adriana CerdaArriagada, Nevada Export S.A.; Roberto

Echeverria P. De V; Claudio SalgadoVillablanca, Sergio Salgado Segura,Armando Fuenzalida Salgado, SociedadAgricola Chequen Ltda, Ana Maria FloresLopez, Sociedad Agricola Ganadera yForestal Arboleda Ltda; Santiago AchurraHernandez, Sociedad Agricola RequinguaLtda, Javier Eduardo Ramirez Vodnizza,Sociedad Eduardo Ramirez Letelier y CiaLtda; Luis Antonio Paredes Nuñes,Sociedad Nuñez y Cia Ltda; FernandoAlmeda Ollé, Sociedad Vinicola MiguelTorres S.A.; Gonzalo Bertelsen Diaz,Terramater S.A.; José Antonio EsturilloCastaño, Vinos del Sur S.A.; Dario PolloniSch, Viña Cantera S.A.; Roberto I.Echeverria Z, Viña Echeverria Ltda;Eugenia del Carmen Diaz Aedo, ViñaMorandé S.A.; Teresa Medina Fernández,Viña Santa Mónica Ltda; Max Undurraga,Viña Undurraga S.A.; Jorge Rojas Garcés,Viña ValdiviesoS.A.; Eugenio Maffei Rojo,Viñedos y Bodegas Corpora S.A.; AngelDomper Pons, Viñedos y BodegasDomper S.A.; Sara Cabrera Gajardo,Fontec; Tonchi Tomic Jakaf; Carmen SutilServoin; Manuel Madrid Baeza,Corporación Chilena del Vino; Chile (15November 2000)

DenmarkThorvald Akselsen, Foss Electric, Denmark (26 April 2001)Mai Nygaard, Chr. Hansen, Denmark (17 November 2000)

FranceMounir Saoma, Michel Picard, France (2 February 2001)Laurent Lebrun, Seguin Moreau, France(12 February 2001)Marc Sabate, Michel De Tapon, SabatéDiosos, France (12 February 2001)

GermanyHis Excellency, Mr Karl-Heinz Funke,Federal Minister of Food, Agriculture andForestries, Mr Jürgen Detken, Head ofDivision, Generlal EU Policy, Mr HellmutAltpeter, Director, Economic Relations tonon-European Industrial Countries, MsSigrun Neuwerth, Head of the PressOffice, Mr Wolfgang Jaeger, ExecutiveOfficer, Minister’s Office, Ms Ines Mann,Interpreter, Germany (3 November 2000)

GreeceYanis Vogiatzis, John Boutari & Son –Wineries S.A., Greece (12 March 2001)

Hong KongAntonio Y.L. Koon, Ponti Trading Limited,Kowloon, Hong Kong (2 August 2000)Johnny Wan, Hong Kong TradeDevelopment Council, Wanchai, HongKong (2 August 2000)

IndiaDr K.K. Jindal, Dr YS Parmar University ofHorticulture and Forestry, Nauni, Solan,India (3 August 2000)

IsraelAmos Mizrach, Volcani Institute, BetDagan, Israel (23 February 2001)

ItalyElio Novello, Gianpalo Vaona, Bolla, Italy(2 February 2001)

JapanDr Yoshio Kodama, Meiji Seika Ltd, Japan(15 December 2000)

PortugalAntonio Amorim and Professor MiguelChabral, Amorim, Portugal (20 November 2000)

United KingdomDr Steven Walker, Campden & Chorleywood Food Association Group, UnitedKingdom (31 August 2000)Mr John Gumbley, Senior Producer, SkyNews, United Kingdom (14 October 2000)Eddie Deaville, ADAS Nutritional SciencesResearch Unit, United Kingdom (22 November 2000)Carmel Kilcline, Sainsbury’s, UnitedKingdom (3 April 2001)Professor Ian R Peterson , CoventryUniversity, United Kingdom (22 May, 2001)

United States of AmericaDr Charles Edwards, Washington StateUniversity, USA (17 November 2000)Paul Dolan, Dennis Martin, John White,Bob Blue, Nancy Walker, Heath Dolan, BillLeair, Cara Hane, Jill Jepson, Fetzer Vineyards,Hopland, CA, USA (2 February 2001)Rick Boyer, Marc Imbert, Jekel Vineyards,USA (2 February 2001)Paul Ahvenainen, Korbel, USA (2 February 2001)Lois Levitan, Cornell University,Environmental Risk Analysis ProgramCentre for the Environment, USA (27February 2001)Members of the University of CaliforniaCooperative Extension Service: RhondaSmith – Sonoma, Larry Bettiga –Monterey, George Leavitt – Madera,Steve Vasquez – Fresno, Christian Butzke– Enologist, UC Davis, Chuck Ingles –Sacramento, Jennifer Hashim – Kern, EdWeber – Napa, Paul Verdegaal – SanJoaquin, USA (12 March 2001)Paul Heck, Scott Laboratories, USA (27 March 2001)

OtherThirty participants from the InternationalFederation of Agricultural Journalists’ NinthWorld Congress (4 September 2000)

9

Sample changer for autotitrator

Sevenh

ill Win

ery: 1900Photograph courtesy of The State Library of South A

ustralia

Wine grape tannin and colour specification

Staff: Dr Markus Herderich, Dr ElizabethWaters, Dr Stéphane Vidal, AnitaOberholster, Dr Z Peng, Stella Kassara,Mariola Kwiatkowski, Dr Mark Sefton,Dr Leigh Francis, Yoji Hayasaka, GayleBaldock, Dr George Skouroumounis,Heather Donnell, Richard GawelCollaborators: Adelaide University -Dr James Kennedy, Dr RobertAsenstorfer, David Lee, StephanieLambert, Dr Graham Jones, RenateRistic, Dr Patrick Iland, Stephen Clarke

This project was initiated as collaborationbetween The Australian Wine ResearchInstitute and The University of Adelaide at therequest of industry; an Industry ReferenceGroup composed of twelve senior winemakersadvises it. Funding for the tannin project isprovided by the GWRDC as well as by the CRCV.

The ambitious objectives of this challengingmulti-disciplinary project are:• To gain an understanding of the types of

compounds important to red wine colourand mouthfeel and of conditionsfavouring their presence in wine.

• To determine whether the interaction ofproanthocyanidins (tannins) with othergrape and wine compounds modifies theirsensory properties.

• To establish viticultural and oenologicaltechniques that enhance wine colour anddesirable mouthfeel.

General observationsThe new Leader of the Tannin Project, DrMarkus Herderich, commenced at the Instituteon 9 April 2001. Dr Jim Kennedy, post-doctoral fellow at the University of Adelaide,has accepted a tenure track position as anAssistant Professor in the department ofFood Science and Technology at OregonState University and left the project in May2001. Other fluctuations of the involvedstaff include Robert Asenstorfer, student atthe University of Adelaide, who qualified forthe award of the degree of Doctor ofPhilosophy; Anita Oberholster, PhD studentat the AWRI, who finished the experimentalpart of her PhD research; and Stella Kassara,chemist at the AWRI, who left for maternityleave in July 2000 and recently resigned.

Despite these changes good progress hasbeen made and the appointment of DrMarkus Herderich as new leader of theTannin project has further focussed to ouractivities. Dr Liz Waters, the precedingproject leader, is thanked for her significantpast contribution which, in the longer term,was not sustainable due to her demandingtask as Manager of Program 1 of theCooperative Research Centre for Viticulture.Besides other measures of success andprogress, it is very pleasing to see that the

outcomes from this project are continuouslybeing communicated in peer-reviewedjournals. In this reporting period, threepapers have been published, four have beensubmitted for publication, and at least afurther four are in draft form. Just asimportant is the public and internationalrecognition of the achievements obtained bythe Tannin project. A fine example is theterminology developed by our sensory panelto characterise the mouthfeel of red wines,which - after publication in 2000 - instantlywas accepted and identified as ‘mouthfeelwheel’ by researchers and winemakers allaround the world. The internationalsignificance of the outcomes of the Tanninproject is also reflected by various invitedpresentations, for example, the Institute wasinvited to give a 45 minute plenarypresentation entitled ‘Red wine colour andmouthfeel: Analysis of phenolic pigmentsand tannins’ at the Annual Meeting of theAmerican Society for Enology and Viticulture,held in San Diego from 27-30 June 2001.This task was admirably performed by thenew project leader and appeared togenerate much interest and follow-upenquiries from parties interested inestablishing collaborations.

A similar reception was given when the tanninproject and some of its recent outcomeswere presented by Dr Vidal at an internationalconference in Bordeaux, where much praisewas given for our achievements in a difficultarea of research. More specific achievementsrelate to outcomes listed below.

Method developmentMethods to quantify the levels of tannins inseed extracts and in wine have been developed,and the identity of polymeric proanthocyanidins(tannins) has been rigorously demonstratedby mass spectrometric analysis.

The results have been published (Z. Peng,Y. Hayasaka, P.G. Iland, M. Sefton, P. Højand E.J. Waters, J. Agric. Food Chem., 49,2001, Institute publication #660) andanother manuscript was submitted forpublication (Z. Peng, P.G. Iland, A.Oberholster, M.A. Sefton and E.J. Waters).The main outcomes are:

• Reversed phase HPLC methods have beendeveloped for quantitative analysis ofpolymeric procyanidins (tannins) in grapeseed extracts and for quantitative analysisof polymeric pigments in wines. Therecovery of polymeric pigments with thismethod was 95%.

• Results obtained with the HPLC methodcorrelated well with spectrophotometricmeasures of total pigmented polymersand indices of wine age.

• Using this technology, a study of 120wines has confirmed that the proportion

of pigmented material incorporated intopolymeric proanthocyanins increases withwine age.

• At present, these analytical protocols areadopted and validated as routine methodsfor the determination of oligomeric andpolymeric polyphenols in grape, seeds, andwine samples from numerous viticultural andwinemaking trials. Libraries summarizingUV- and mass spectral data will facilitatethe identification of key polyphenols inthe future. These initiatives will result insignificantly reduced analysis time andsecure higher through-put of samples.

Wine pigmentsRobert Asenstorfer’s PhD thesis was passed.• The findings from Robert Asenstorfer’s

thesis have now been prepared forpublication by Robert and Dr Graham Jones.

In a collaboration with Robert, Yoji Hayasakadeveloped and optimised methods for thedetection of stable wine pigments byElectrospray ionastion mass spectrometry(ESI-MS/MS). Applying tandem massspectrometric experiments numerouscomplex red pigments could be detected inwine samples. With this approach we alsoidentified four novel anthocyanin polyphenolderivatives for the first time as naturalproducts in wine.

A paper outlining this methodology hasbeen submitted for publication.

Analytical methods to identify the existenceof new wine compounds is a necessary butnot sufficient tool to advise winemakers onhow to modulate wine composition to meetset quality specifications. Once identified,the relative importance of such compoundsmust be established. The next step is thusto synthesize some of the newly discoveredcompounds in sufficient quantities todescribe their chemical and sensorialproperties. To this end, David Lee, a PhDstudent funded by CRCV, and Mr AndersHaakansson, a visiting synthetic chemistfrom Denmark, has initiated synthesis ofthese new wine pigments.

CopigmentationCopigmentation reactions are stronglyinfluenced by pH, with its relative importancebeing greatest at higher pH values. Ethanoldecreases the copigmentation interaction.

Of the common simple phenolics in wine,quercetin has been shown to form the moststable intermolecular complex with malvidin-3-glucose.

Self-association of malvidin-3 glucose alsoappears to be a major cause of colourenhancement. As model experiments showthat the extent of intermolecular copigmentationdepends on the concentration of anthocyanins,

Team Reports

11

more is likely better in young red wines.Conversely, aged products feature significantlyreduced concentrations of anthocyanins and thecontribution of intermolecular copigmentationto the color of older red wines remains to beestablished. Stephanie Lambert, an AdelaideUniversity PhD Scholarship holder, presenteda poster on her physical chemistry studies atIn Vino Analytica Scientia, held in Bordeauxfrom 14-16 June.

Grape seed and skin proanthocyanidinsAs reported previously, adding extra seeds toa red wine ferment produced wines withincreased levels of phenolics, includingtannins and pigmented polymers, andenhanced colour density.

After ageing in bottle for two years, winesmade from must with extra seedscontained more colour and were higher inastringency than wines madeconventionally. Importantly, the winesappeared not to be more bitter.

This material is being drafted forpublication (Z. Peng, P.G. Iland, I.L. Francis,Y. Hayasaka and E.J. Waters).

A method for characterising the compositionof grape and wine tannins using acidhydrolysis in presence of phloroglucinol hasbeen developed.

The analytical protocol has been published(J.A. Kennedy and G.P Jones, J. Agric.Food Chem., 49, 2001, 1740.).

For the first time, nano electrosprayionisation in combination with massspectrometry allowed us to detectindividual tannins on a molecular level.This method has been developed by YojiHayasaka and represents a major progressfor our studies aiming to understand thechemical properties of tannins.

Application of this methodology enabledus to characterize the composition andpurity of several different tannin fractionsfrom grape seeds and grape skins. This isessential when attempting to ascribespecific sensorial properties to individualclasses of isolated tannins.

An investigation on structural changes ofgrape skin tannins and pigmentedpolymers at different stages of berrydevelopment has been conducted. Again,the methods developed proved to beessential for this study on the compositionof grape skin proanthocyanins.

The results of the latter experiments havebeen submitted for publication (J.A.Kennedy, Y. Hayasaka, S. Vidal, E.J. Watersand G.P. Jones – manuscript in review).

Shiraz grapes were collected during theperiod of berry development from a range ofviticultural regions (Riverland, Barossa andCoonawarra). In some cases samples werecollected from trials where differentirrigation schedules had been applied. Thesestudies showed that levels of catechin,epicatechin and extractable tannins(procyanidins) in seeds increased up untilveraison, where they were at theirmaximum, and then declined. Thedifferences in seed composition of grapessourced from the range of viticultural regionswere less than the differences in skin colour(anthocyanin levels).

When the seed colour of berries fromdifferent vineyards within one region arecompared at the same stage of ripeness(e.g. 24 ºBrix) there is little difference inthe degree of seed colour, indicating thatonce berries have ripened to a particularpoint, assessment of seed colour is not adiscriminating factor between thepotential quality of different vineyards.

A viticultural trial to investigate the differencesbetween seed physical characteristics (seednumber, size and weight) and seed chemicalcomposition of grapes that develop inshaded canopy conditions versus that ofgrapes that develop in moderately and wellexposed canopy conditions has beenestablished by Pat Iland and Renata Ristic atThe University of Adelaide.

The extractable tannin content of seedswas maximal in the three weeks prior toveraison and then decreased as the fruitripened. The seed content of the flavan-3-ol monomers also decreased after veraison.

Visually seeds were initially green, plumpand had pliable seed coats. Beginningfrom veraison, the seeds browned incolour, became desiccated and the seedcoats hardened.

These changes in content and visualaspects can be explained by oxidativeevents occurring in the seed coat duringfruit ripening. This information has beenpublished (J.A. Kennedy, G.J. Troup, J.R.Pilbrow, D.R. Hutton, D. Hewitt, C.R.Hunter, R. Ristic, P.G. Iland and G.P. Jones,Aust. J. Grape Wine Research 6, 2000, 244).

There were no differences in totalamounts (monomers and procyanidins),subunit composition (amount of terminaland extension units) or mean degree ofpolymerisation, of seeds of differentcanopy treatments.

Linking the astringency and mouthfeelproperties of wines to proanthocyanidinand or other polymer fractions in wineFunded by the CRCV and employed by the

Institute, Stéphane Vidal commenced workin Véronique Cheynier’s laboratory at INRA-IPV in Montpellier on 6 June 2000, where heprepared tannin fractions from seeds andskins for sensory assessment.

Stéphane has isolated and purified gramquantities of ten different tannin fractionsfrom grape seeds and grape skins.

A panel of tasters has been convened andtrained by Richard Gawel over the last fewmonths. In late March these tasterscompleted a very extensive tasting of thetannin fractions isolated by Stéphane.

This is the first time that such fractionshave been tasted and profiled for theirastringency sub-qualities. The data willallow us to understand how the chemicalproperties of tannins relate to theirsensory properties and are currently beingsubjected to extensive statistical analysis.

Stéphane presented this work at In VinoAnalytica Scientia, held in Bordeaux from14-16 June.

Winemaking practices that effectproanthocyanidin levels and sensoryproperties of wine

To study the effect of winemaking practiceson colour and mouthfeel the first commercialscale winemaking trials have beenconducted at the Hickinbotham RoseworthyWine Science Laboratory located on theWaite Campus with support from ourIndustry Services Manager and continuedinput by the facility Manager, Mr StephenClarke, a winemaker with 20 yearscommercial experience.

Winemaking was performed according to theguidelines of the industry reference group.

The objectives of these initial 20fermentations are to analyze the influenceof red grape varieties (Coonawarra Shirazand Adelaide Hills Cabernet Sauvignon),fermentation tanks (rotary tanks versusstationary fermenters), and fermentationtemperatures (18°C, 25°C) on red winephenolic compounds. A complementarytrial conducted by our Wine Microbiologyteam assessed the influence of yeaststrains on tannin profiles.

In addition, we aim to study the variationbetween replicate fermentations and toidentify key parameters which contributeto variation.

Reference samples of juice and skins havebeen collected on a daily basis duringfermentations, and are continuously kepton a weekly basis prior to bottling.

Team Reports

12

Comprehensive chemical analysis ofseveral hundred samples for polyphenoland tannin content has been initiated. Itwill be complemented with sensoryanalysis after short term bottle storage,and several of the wines produced will bepresented to industry in workshopsconducted at the Eleventh AustralianWine Industry Technical Conference.

An analysis of compositional differencesbetween grape marc derived tanninmaterial and tannins extracted from skinsduring fermentation, indicates differencesin the relative proportions of polymeric vsmonomeric pigments and the ratio ofmalvidin-3-glucoside-6’ acetates tomalvidin-e-glucoside-6’-p-coumarateanthocyanins. For example, the proportionof these natural polymeric pigment in youngwines is approximately 15% but canconstitute greater than 60% in grape marcextracts. Similarly, the proportion ofmalvidin-3-glucoside-6’-p-coumarate is higherin marc extracts than in most young wines.

These characteristics can be determinedby a single HPLC analysis using themethods developed by Peng.

Further analysis of a liquid grape marcextract indicated that dextrins were notdetectable in the preparations analysed.This contrasts with previously publishedresults on the composition of solid grapemarc extracts.

Rapid instrumental techniques

Staff: Dr Leigh Francis, Dr Bob Dambergs, DrMichael Esler, Dr Bruce Kambouris, DrWies Cynkar, David Boehm, ProfessorPeter Høj, Dr Sally-Jean Bell, Peter GoddenCollaborators: Adelaide University -Dr Graham Jones, Dr Patrick IlandOrlando Wyndham Group - RussellJohnstone, Ben Zander, Inca LeeSouthcorp Wines - Dr Andrew Kleinig,Eric Wilkes

A comprehensive suite of accurate and precisecalibrations has been developed for themeasurement of colour, pH, and TSS in redgrapes on three distinctly different types ofNIR instruments. These calibrations range inspecificity from general 'pan-Australian' tomore precise region and variety specific.Greater than 2,000 samples have been collectedthis vintage by collaborating companies andsome of these have been delivered to theInstitute and analysis has commenced.

As part of our obligations to the CooperativeResearch Centre for Viticulture and to theCommonwealth, publication of this methodhad earlier been held over pendingfinalisation of intellectual property issues.

These matters have now been largelyresolved and the potential forcommercialisation of the technology hasbeen examined more fully. While it wasdeemed timely to delay publication earlier toallow the Institute to secure the advantageof being an early entrant into the field bynot disclosing information to potentialcommercial competitors or imitators, it issuggested that this delay is no longerwarranted. The publication of the NIR methodfor colour, pH and total soluble solids, willnow be accelerated.

The sample collection program providedmore than 2000 samples from the 2001vintage from grape growing regions inNSW, VIC, WA as well as SA. It isanticipated that by the end of 2001 thechemical and spectroscopic analysis of thisdata set will be completed and the dataused to refine earlier NIR calibrationsbased on 1999 and 2000 data anddevelop something approaching a pan-

Australian calibration or set ofcalibrations. The significance of regional,varietal and especially seasonal variationfor NIR analysis will be further investigated.At this point in time, it appears that region-specific and variety-specific calibrationswill be slightly better than pan-Australiancalibrations. Whether the differences havereal practical consequences is yet to beresolved (see Figure 1).

Negotiations with the CRCV and thecollaborating wine companies have takenplace in order to clarify the mode ofcommercialisation and provide a mandatefor the management of such activity.

The transfer of calibrations between well-matched or standardised instrumentsappears likely to be successful and will beexamined in detail with samples from the2001 vintage. It should be notedhowever that this is not a trivial exerciseand may require significant effort. While

13

Figure 1: NIR calibrations for total anthocyanin concentration for a population of ~900 red wine grape samples and one of itssub-populations. A. ‘Global’ calibration; 909 samples across 10 growing regions, 10 grape varieties and 2 seasons, colouredby variety. B. Subpopulation, localised for both a single grape variety, Cabernet Sauvignon, and a single growing region,the Riverland. Observe that NIR measurement precision (SECV) improves by focusing on a particular region and variety.

Tota

l ant

hocy

anin

s by

NIR

(m

g g-1

)

Total anthocyanins by reference method (mg g-1)

A B2.0

1.5

1.0

0.5

0.5 1.0 1.5 2.0 0.5 1.0 1.5 2.0

N = 909 samples10 varieties10 regions

R2 = 0.90SECV = +/-0.14 mg g-1

N = 78 samples1 variety (Cab.Sauv.)1 region (Riverland)

R2 = 0.94SECV = +/-0.05 mg g-1

Dr Leigh Francis and Dr Bob Dambergs

the direct calibration of a singleinstrument has proven to be relativelystraightforward, the simultaneousstandardisation of a population of severalor many instruments to the one calibrationis a more complex issue. In particular, itappears that not all types of instrumentsare equally capable of being standardised.On the other hand, there now existsrecognised protocols for thestandardisation of more expensiveresearch grade instruments, such as theNIRSystems6500 spectrophotometer.Commercially available software that isdesigned for calibration transfer andinstrument standardisation is to beevaluated in standardisation of theInstitute’s NIRSystems6500 spectrometerwith BRL Hardy’s based on 2000 and2001 vintage data.

Investigations with instruments capable ofscanning whole fruit are continuing inassociation with two wine companies, butno data has yet been received. The PertenDA7000 instrument offers the possibilityof being able to rapidly analyse samples ofintact whole berries. The investigation isproceeding on a large data set alreadyprovided by BRL Hardy using samples fromthe 2001 vintage.

Trials with an existing prototype bin probehave commenced and data is being collected.Initial trials have focussed on off-site evaluationof reproducibility of spectral collection.

Trials with prototype bin probes continuedduring the 2001 vintage to furtherexamine the feasibility of the concept.Whilst the trials have demonstrated thepromise of this approach of in-situscanning, there is some indication thatcommercial development of a NIR ‘bin-probe’ may be ambitious at present giventhe current state of commercially availableNIR technology. Institute efforts may bebetter focused on addressing the moretractable issues encountered indevelopment of a compact laboratory-based instrument. The latter is likely to bedeliverable to industry earlier than theformer.

Three of the relatively-inexpensiveprototype NIRTech instruments will betested in parallel at the Institute in orderto evaluate instrument effects in theabsence of all other experimentalvariables. The successful standardisationof cheaper diode-array based instrumentsin particular (such as this prototype), is stilla subject of considerable research inquiryinternationally.

It was planned to expand the trial of theseprototype instruments to a small numberof companies for the 2001 vintage.

Disappointingly, there have been unforeseendelays and only one instrument is likely tobe supplied to industry in 2001.

Considerable effort has been expended ininvestigating previously identified variation inthe glucosyl-glucose (G-G) assay calibrations(particularly for white grapes). Detailedscrutiny of the reference method has led tosome refinements of the analytical techniques.In particular, the use of a 'stomacher' for gentlymacerating white grapes in a reproduciblemanner that is also more relevant to realwinemaking practices, has improved thedegree of uncertainty in the measurement.

Further samples of Chardonnay in particularare being collected this season withcooperation from BRL Hardy. These sampleswill be tracked from juice right through towine and linked with the subsequent qualityrating determined at the in-house allocationtastings. Other wines assessed at theallocation tastings will also be scanned andthen discrimination models will be tested.

Studies using commercial tasting dataindicate the approach of using NIR forallocation tastings and measurement of G-G in white fruit may have some value andthis will be addressed more fully in futurework. Samples from the 2001 vintagehave now been received at the Instituteand work will commence shortly.

In some recent preliminary studies, grape andother tannin fractions of various sizes, chemicalcomposition and degree of polymerisationthat have been prepared by Dr StèphaneVidal as part of project AWR 96/1 (Winegrape tannin and colour specification) havebeen scanned by NIRS. The spectra appearto show significant differences and suggestthat the technique may be useful indistinguishing these tannin fractions eitherquantitatively, qualitatively, or both.

Yeast flavour and fermentation activity

Staff: Dr Paul Henschke, Dr Eveline Bartowsky,Dimitra Capone, Dr Peter Costello, NadiaD'Incecco, Jeff Eglinton, Dr Leigh Francis,Holger Gockowiak, Kate Howell, JaneMcCarthy, Dr Mark Sefton, TraceySiebert, Professor Peter Høj Collaborators: Professor Graham Fleet,The University of New South Wales,Stephen Clarke, The University of Adelaide

This GWRDC-funded project has as a primaryobjective, to investigate microbiologicalinterventions which can be used to generatea wider spectrum of wine sensory propertiesthan currently achieved with conventional useof well-characterised Saccharomyces cerevisiaestrains and well-known malolactic bacteria.Three current main activities are related to:

• Development of new isotopic dilutionassays for quantifying the key impactcompounds produced by yeast, as a toolfor determining the importance of yeaststrain and fermentation conditions,

• Evaluation of the oenological properties ofnon-conventional yeasts with an emphasison correlating chemical composition andorganoleptic attributes of wines madeunder winery conditions,

• Laboratory and winery studies concerningthe use of mixed yeast strain startercultures, compared with conventionalsingle strain cultures, as a strategy forsimulating 'natural ferments', and

Quantification of yeast key impact aromacompounds—methods developmentAs stated in the previous annual report, the aimof this collaboration with Drs Mark Sefton andLeigh Francis and their teams, is to developstable isotope dilution assay methods forquantifying the key impact compounds formedduring the yeast alcoholic fermentation. Workhas focused on the principal esters produced byyeast which are an important contributor to thefruity/floral/estery character of wine, which isoften referred to as the 'fermentation bouquet'of wine. Good progress has again been madewith the synthesis of ten isotopically-labelledester standards being completed. Thedevelopment of validated assays for the ethylesters has commenced. Ms Tracey Siebert, whohas recently joined the team, will focus onapplying the assays to wines produced throughour fermentation trials; a series of Chardonnayand Cabernet Sauvignon wines are currentlyundergoing extensive chemical analysis andsensory evaluation. This analytical approach willallow us to relate the ester profile of winedetermined chemically and by sensorytechniques and to determine the relativeimportance of yeast strain and fermentationconditions on the development of thefermentation bouquet of wine. In additionto assays on esters, several other stableisotope dilution assays for important winearoma and flavour compounds is underway.

Industry evaluation of selected novelflavour yeastsReplicated winery scale fermentation trialswere undertaken during the 2001 vintagewith the aim to consolidate the preliminarytrials undertaken previously on a laboratoryscale and to focus on the key attributes thatthese non-conventional yeasts can offer winemakers in search of increasing the flavourdiversity of wines.

Saccharomyces bayanus trialsIn addition to the barrel fermentation trialsreferred to in the past annual report, awinery scale trial using S. bayanus, and S.cerevisiae as a reference wine yeast, withChardonnay and Cabernet Sauvignon hasbeen undertaken in the HickinbothamRoseworthy Wine Science Laboratory in

Team Reports

14

Atomic absortion spectrometer

Sepp

eltsfield W

ine C

ellar: 1945Photograph courtesy of The State Library of South A

ustralia

collaboration with Mr Stephen Clarke duringthe 2001 vintage. The Wine ScienceLaboratory offers the opportunity to havethe wines contract made essentially underwinery conditions but the close proximity tothe Institute also allows us to closely monitorfermentation progress. We are grateful toBRL Hardy for generously providing 4000L ofChardonnay juice and 4 tonnes of CabernetSauvignon grapes for use in these trials.

The main purpose of the Chardonnay trialwas to evaluate the performance and todetermine the organoleptic properties of S.bayanus AWRI 1375 under larger scaleconditions using 260-280L fermentors. As inprevious trials S. cerevisiae AWRI 838 wasthe reference yeast. In addition, treatmentswere included which were designed toevaluate different methods for ensuring thatwine with low residual sugar was achievedafter fermentation with non-conventionalyeasts, in this case S. bayanus 1375. Thetreatments involved:

• Sequential inoculation with S. cerevisiaeAWRI 838 when the S. bayanus fermentwas approximately 70% completed,

• Aerobic pumping-over of a stuck S.bayanus ferment, and

• Inoculation of a stuck S. bayanus fermentwith an S. bayanus AWRI 1375 rescueculture prepared by the Instituterecommended procedure.

The reference wine yeast, S. cerevisiae AWRI838, completed fermentation at 15°C in 18-24 days whereas S. bayanus AWRI 1375ceased fermentation with a residual sugar of40 g/L after 35 days fermentation.Sequential inoculation of the S. bayanusferment with S. cerevisiae AWRI 838prevented lagging of the S. bayanus fermentwhich completed in 39 days. Both theaerobic pumping over treatment and the useof the S. bayanus rescue culture successfullyremoved the residual sugar from the stuck S.bayanus ferment. The latter procedure gavean especially interesting result because therescue procedure not only worked well withthe same yeast that caused the stuckfermentation but that the Institute'srecommended rescue procedure wassuccessful with the less robust S. bayanusfermentation yeast compared with thenormally recommended S. cerevisiae. Theeffect of these treatments on wine chemicalcomposition and sensory characteristics isbeing determined as a guide torecommending treatment preference.

Based on encouraging results obtained withS. bayanus AWRI 1375 in a previouslaboratory-scale Shiraz trial, a larger scaletrial using six 800 kg capacity rotaryfermentors located in the WSL wasundertaken during the 2001 vintage. S.cerevisiae AWRI 838 was the reference yeast.

Mechanically harvested Cabernet Sauvignonfruit, which had been treated with SO2during harvest, was crushed and inoculatedwith S. bayanus or S. cerevisiae. Problem-free ferments with both yeast completed in13-14 days, however, starter culture-inducedMLF was delayed in the wines fermentedwith S. bayanus compared with S. cerevisiae.

All trial wines are currently being filtered andbottled in readiness for extensive chemical,instrumental and sensory analysis. In particular,we plan to quantify the main yeast-derivedflavour compounds of these wines with GC-MSanalysis combined with the newly developed

stable isotope dilution assays developed by theteam of Dr Mark Sefton and Dr Leigh Francisto establish the chemical basis for the flavourattributes of this yeast and the impact offermentation conditions on their formation.

Several of the winery trial wines will beselected for use in the Winemaking withnon-conventional yeasts workshop beingplanned in conjunction with the EleventhAustralian Wine Industry Technical Conferenceto be held in Adelaide in October 2001. Theteam is quietly confident that the soon to beperformed rigorous analysis and results willpoint to the AWRI 838 and AWRI 1375strains giving very different wines withdiffering positive attributes.

Candida stellata and Candida krusei trialsDue to tank limitations in the HickinbothamRoseworthy Wine Science Laboratory, 18LChardonnay (BRL Hardy) fermentation trials wereperformed in glass with the experimentalCandida strains, C. stellata AWRI 861, 1159 andC. krusei 863, at the Institute. The Chardonnayjuice provided by BRL Hardy was also used forthese trials. The ferments were superinoculatedwith S. cerevisiae AWRI 838 at approximately50% fermentation progress to ensure theproduction of wines with low residual sugar. TheS. cerevisiae AWRI 838 ferment failed tocomplete and was rescued by reinoculation withAWRI 838. The wines were cold-settled, racked,

treated with SO2, membrane filtered and bottled.Formal sensory descriptive evaluation andchemical analysis has commenced.Preliminary analysis confirms our previousfindings (Institute staff publications #614and #625) that the wines made withdifferent yeast have marked differencesespecially in the ester profile (floral versusbanana versus fruity versus ethyl acetate).Chemical analysis is recorded in the Table 1.

The chemical and sensory data confirm thepotential of novel yeasts for developingwines with novel attributes.

Team Reports

16

Dr Peter Costello and

Optimisation of C. stellata inoculation forinducing fermentationSome non-Saccharomyces yeasts are knownto exhibit greater sensitivity than S. cerevisiaeto growth inhibition by SO2, which is oftenadded to grape juice/must prior to inoculationwith a selected wine yeast to inhibit indigenousyeasts and to prevent browning; this observationwas noted in the Institute staff publication #625in relation to C. stellata. Since this propertymay impact on the effective deployment ofthis strain by industry, the sensitivity of C.stellata to SO2 has been quantified.

The inhibitory affect of SO2 on yeast growthwas determined in a freshly prepared grapejuice by monitoring the delay in the onset offermentation. C. stellata 1159 showed anextended lag phase of 2-4 days wheninoculated into sultana juice containingfree/total SO2 of at least 32/50 mg/L (pH3.2) whereas S. cerevisiae commencednoticeable growth within one day. By contrastwith C. stellata, C. krusei was more tolerantand commenced good growth with aminimal lag phase. The low tolerance of C.stellata to SO2 is likely related to low productionof SO2 and acetaldehyde by this yeast.

In addition to the use of H2O2 to reduce theSO2-induced growth lag by oxidation of theSO2, the lag phase in grape juice was alsoabolished by the addition of acetaldehyde,which presumably acted by binding the toxicfree SO2 , even at a concentration of 65mg/L SO2. Recent research from Prof.Pamment’s laboratory, University ofMelbourne, has shown acetaldehyde tostimulate fermentation independent ofreaction with SO2 so it is possible that bothmechanisms have stimulated yeast growth.

These data confirmed that C. stellata, likemost indigenous yeasts, is more susceptiblethan S. cerevisiae to SO2 requiring eitherreduced SO2 addition before inoculation,prolonged storage of juice at low temperatureto ensure maximal binding of the free SO2or a reduction in free SO2 content by

oxidation with H2O2 or aeration, or byacetaldehyde treatment. The treatment withacetaldehyde is experimental and is notpermitted under existing wine regulations.

Yeast impact on wine flavour andaroma—mixed culture versusmonoculture fermentationIt is universal practice to induce the alcoholicfermentation of grape juice/must with aselected wine strain of Saccharomycescerevisiae in order to produce a reliablefermentation and a wine with expectedchemical and organoleptic properties.However, a relatively small, but increasingproportion of wines are made withoutinoculation thereby allowing the indigenousyeasts to proliferate, to give a so-called‘natural ferment.’ This practice, which somewinemakers believe gives wines preferredsensory characteristics, can be unreliable andits application is limited because the winemaking properties of the indigenous yeastsare not known in advance.

A key difference between inoculated andnon-induced fermentation relates to theinitial population of S. cerevisiae present.Non-induced ferments typically initiallycontain a mixture of indigenous non-Saccharomyces yeasts which dominate theearly stage of fermentation but then giveway to a mixture of S. cerevisiae strainswhich complete the fermentation of thesugars whereas induced ferments, while alsohaving a similar population of indigenousyeast species are generally dominated by theinoculated S. cerevisiae yeast culture fromthe outset. This project was undertaken toevaluate the use of a mixed culture strategyto simulate ‘natural’ ferments and toinvestigate the interaction between strains.As a simplified model, the mixed cultureferments were performed with three strainsof S. cerevisiae wine yeast.

Five different strains of S. cerevisiae in threedifferent combinations of three strains weretested under nutrient limiting and non-

nutrient limiting conditions in a Chardonnayjuice. Wines were also made with each ofthe five strains. These treatments resulted inthree wines made by mixed culture ferments(M1-M3) and five wines made by monocultureferments (T1-T5) under the two nutrientconditions. Three blended wines were alsoprepared by mixing in equal proportion themonoculture wines which corresponded tothe yeasts used in the mixed culture ferments(B1-B3). The wines were tested for organolepticdifferences by duo-trio aroma testing.

The proportion of each yeast strain in themixed culture ferments was monitored bymicrosatellite-PCR analysis. Themicrosatellite sequences, present in the yeastchromosomal DNA, were demonstrated tobe sufficiently polymorphic to permit reliablestrain identification.

Duo-trio testing of the resultant winesshowed that in some cases multiple strainsof S. cerevisiae interacted to produce winesthat were sensorially different to:• Any of the wines produced by

monoculture fermentation with thecorresponding yeasts, or

• A wine produced by blending in equalproportion the three wines produced bythe corresponding monoculturefermentations.

These preliminary results need verification, butnevertheless suggest that wine with an aromaprofile different from wines of single yeastferments, or their blends, can be producedby mixed culture fermentation only whenthe yeast strains interact and no-one straindominates, as appears to normally be thecase for 'natural' ferments. This suggeststhat mixed cultures of S. cerevisiae in winefermentations could be used to modify thearoma of wine reminiscent of naturalferments but with a minimal risk typical ofinoculated monoculture fermentations.

17

Juice 3.24 7.5 13 35

S. cerevisiae 838 13.5 5.9 3.39 7.5 0.41 0.5 22 112

C. stellata 1159 + Sc 838 13.4 14.8 3.28 9.5 0.94 0.8 24 107

C. stellata 861 + Sc 838 13.4 10.0 3.39 7.6 0.21 1.0 26 131

C. krusei 863 + Sc 838 (13.8) 6.6 3.37 7.3 0.16 0.8 28 102

Range for malate = 2.3-2.5 g/L, lactate <0.1 g/L, tartrate=4.0 g/L, and citrate=0.3-0.4 g/L

Table 1. Comparison of the chemical composition of Chardonnay wine made by fermentation

with S. cerevisiae AWRI 838, C. stellata AWRI 861, 1159 and C. krusei 863.

Treatment Alcohol Glycerol pH TA VA Succinate free SO2 total SO2(% v/v) (g/L) (g/L) (g/L) (g/L) (mg/L) (mg/L)

Orbital shaking water bath withsmall scale fermentations

Photograph courtesy of The State Library of South Australia

Ms Kate Howell has performed this work atthe Institute as part of her PhD studiesundertaken through the University of NSWwith supervision by Dr Eveline Bartowsky,Professor Graham Fleet (University of NewSouth Wales) and Dr Paul Henschke.

Role of malolactic bacterial glycosidaseenzymes in the development of wine flavourNadia D’Incecco, a PhD student from TheUniversity of Padua, Italy, working at theInstitute under the supervision of Dr EvelineBartowsky and Dr Paul Henschke has completedher studies on determining the role of ß-glucosidase activity, produced by malolacticbacteria, on the release of glycosylatedflavour precursor compounds isolated fromChardonnay wine. This study complementsher work in Italy where she is characterisingthe ß-glucosidase activities of yeast.

The role of malolactic bacteria in thedevelopment of wine flavour is not clear butis believed to involve at least threemechanisms, i) deacidification, by which L-malic acid is decarboxylated to L-lactic acidand CO2, ii) metabolism of wine substratesto form either flavoured or flavourlessproducts, such as the metabolism of citricacid to form diacetyl with a 'buttery' flavour,acetic acid, and 2,3-butanediol which has noflavour significance in wine (reviewed byBartowsky and Henschke 2000, staff reprint#637), and iii) the hypothesised release ofgrape derived glycosylated flavour precursorcompounds by excreted glycosidic enzymes(GWRDC project UCS 92/4).

An experiment was designed and conductedto establish whether or not malolacticbacteria have the ability to hydrolyse andrelease glycosylated flavour precursorcompounds isolated from Chardonnay wine.The flavour precursor compounds present inwine are typically ß-glucosides ordiglycosides in which an arabinose,rhamnose, apiose or xylose residue is linkedto the inner glucose residue (Institutepublication #458). An extract of glycosideswas made from a Chardonnay wine, addedto a model wine, and after MLF, the glucosecontent of the wine was measured by theglycosyl-glucose (GG) assay to indicate theextent of hydrolysis and a GC-MS analysisperformed to reveal the extent of liberatedflavour aglycons. Control experimentsincluded no bacterial addition to test foracidic hydrolysis, and addition of variouscombinations of glycosidic enzymes asreference positive controls.

Using synthetic substrates, the two strains ofOenococcus oeni studied were shown toexhibit low �-L-arabinofuranosidase and �-L-rhamnosidase activities, endoglycosidases,which are necessary to cleave the outersugar, often arabinose or rhamnose, fromthe diglycoside. These strains had significant

ß-glucosidase activity, which cleavesglucosides formed by the action of thepreviously mentioned endoglycosidaseenzymes or are directly derived from thegrape, to release the flavour aglycon.

Monitoring the total a-a content of themodel wine containing the glycosidic extractshowed that purified ß-glucosidase onlyprovided limited hydrolysis of the wineglycosides and confirmed that �-L-arabinofuranosidase and �-L-rhamnosidasewere required for efficient hydrolysis of theglycosides. MLF conducted by the O. oenistrain only led to limited hydrolysis of thewine glycosides, suggesting that thisbacterium does not elaborate sufficientactivity of the enzymes necessary for thehydrolysis of disaccharides. Addition of �-L-arabinofuranosidase and �-L-rhamnosidaseto the wine during MLF demonstrated thatthese enzymes are required for efficienthydrolysis of grape glycosides during MLF.

This work has provided important newinformation on the somewhat uncertain roleof MLF in wine flavour development. Thisinvestigation, which requires confirmationwith additional strains, suggests that theability of ML bacteria to release glycosylatedflavour precursor molecules is relativelyrestricted and probably confined to the ß-glucosides, unless yeast derived or extraneousglycosidases, such as rhamnosidases andarabinofuranosidases, are present.

Malolactic fermentation and wine flavour

Staff: Dr Paul Henschke, Dr EvelineBartowsky, Dr Peter Costello, HolgerGockwiak, Jane McCarthyCollaborators: Dr Leigh Francis,AWRI, and Dr Andrew Markides,Adelaide University

Sensory attributes associated with themalolactic fermentation of wine - diacetylformation by commercial MLF bacteria in wineThe flavour compound, diacetyl, is a majorcontributor to the ‘buttery’ attribute of winewhich has undergone malolactic fermentation(MLF). The ML bacterial strain can affect thediacetyl content of wine (Institute publications#546 and #637), however, limited comparativeinformation on commercially available strainsis available to winemakers. This project aims tosystematically characterise these strains undersimilar conditions in a red and white wine(alcohol 12.5-13.0% v/v, malic acid 2.5 g/L,citric acid 0.5 g/L, pH 3.4, SO2 negligible). Thepresent report relates to diacetyl production byfour ML bacteria cultures, Chr. Hansen Vinifloraoenos, Lallemand EQ54, Lallemand OSU, andCondimenta/Lallemand Bitec/Viacell proVino, ina 2000 Chardonnay wine, the results pertainingto the red wine (1998 Cabernet Sauvignon) trialwere summarised in the previous annual report.

Completion of MLF ranged 2 - 8 weeks forthe four strains in the Chardonnay winewhereas the same four strains completedMLF within two weeks in the CabernetSauvignon wine, suggesting a nutrientlimitation or an inhibitory metabolite mayhave selectively delayed the growth andfermentation activity of two of the strains inthe Chardonnay wine. The peak diacetylconcentrations were lower and of greaterrange in the white wines (1-4.5 mg/L)compared with those obtained with the redwines (8-10 mg/L). As for the red wines(Figure 2), diacetyl peaked after theconsumption of malic acid and whenapprox. 75% of the citric acid had beenmetabolised (Figure 2). After the completeremoval of citric acid, diacetyl had fallen toless than 50% of the peak value.

These data agree with our bottled wine surveythat red wines carry a higher diacetylconcentration than whites (Institute staffpublication #546). These data also showed acorrelation between peak diacetyl concentrationand duration of MLF, although further workis needed to confirm this observation. Thekinetic pattern of malic and citric aciddegradation and diacetyl evolution wasqualitatively similar for all strains in each wine.

In conclusion, these results suggest thatalthough strains vary in ability to producediacetyl, other factors, such as duration ofMLF, and the stage of MLF at which thewine is stabilised (SO2 addition or membranefiltration) appear to give relatively greaterdifferences in diacetyl concentration. Thechoice of ML bacterial strain is important tothe extent that growth response in a particularwine is variable, thereby significantlyinfluencing the final diacetyl concentration.

Most importantly, these data confirm theresult obtained previously with red wine thatdiacetyl is a transient metabolite and thatthe timing of wine stabilisation (e.g. sulfiteaddition) for controlling the desired level ofdiacetyl in wine is important.

Preliminary evaluation of a chemical sensorwith wines having undergone MLFA small project has been undertaken to evaluatethe ability of a chemical sensor to determinethe aroma profile of wines which hadundergone MLF. The chemical sensorinstrument, a Hewlett Packard model HP4440,located in the Department of HVO ofAdelaide University, is a mass spectrometrybased ‘electronic nose’ for thecharacterisation of wine volatiles, where thefragmentation patterns may provide afingerprint of the volatile compounds presentin the headspace of the wine sample. Thisinstrument comprises an autosampler (capacityfor 44 samples) and a mass sensor wherethe wine volatiles are ionised and fragmentsdetected, and the abundance of ions

Team Reports

19

determined. The instrument has a majoradvantage over conventional instruments inthat it can rapidly analyse many samples.

Two wine types (Chardonnay and CabernetSauvignon) from three different viticulturalregions (Chardonnay – Eden Valley, Sunraysiaand Padthaway; Cabernet Sauvignon – EdenValley, Coonawarra and Swan Hill) weresubjected to MLF with two commercial bacteriapreparations (Viniflora oenos and LalvinMCW). Malolactic fermentation was carriedout at 20ºC in an oxygen-free atmosphere.

The chemical sensor could distinguishbetween both the red and white wines, andpre and post MLF wines, and in some caseswas also able to distinguish the wines on thebasis of origin, possibly on the basis offermentation esters. There was evidencethat the bacterial strain used to conduct theMLF could be distinguished. However, thechemical sensor was unable to distinguishwines on the basis of the flavour compound,diacetyl, due to the diacetyl ion fragmentsoverlapping those of ethanol. Further workis required to establish the identity of thevolatile compounds which allowed for thedifferentiation of wines pre and post MLFwith a particular bacteria strain. Tounequivocally assess the use of the chemicalsensor for discriminating wines fromdifferent regions, further work wouldrequired, for example, standardised winemaking of several examples of fruit fromseveral viticultural regions.

This project was undertaken by Ms HelenMcCarthy as an Honours project in theDepartment of Horiculture, Viticulture andOenology, with supervision by Drs EvelineBartowsky and Leigh Francis. Helen wasawarded a first class honours degree, andcommenced as Graduate Winemaker withSouthcorp Wines at Karadoc in December 2000.

This work has provided an insight into thepotential use of the chemical sensor as ameans to analyse wines, and may proveuseful as a quick method for the initialexamination of wines for MLF character,and the technique could give information

on the factors which allow the wines tobe distinguished. However, this sensorcannot detect diacetyl, the maincompound conferring the 'buttery'character, in the presence of alcohol.

Model studies on the effect of timing of O. oeni inoculation on the induction of MLFBased on the chemically defined grapejuice/wine medium reported on in the 1999Annual Report, a trial has been undertakento evaluate this medium for the ability tomodel aspects of the MLF. This investigationfocused on the effect of timing of O. oeniinoculation on the initiation and duration ofMLF and diacetyl production. A compatibleyeast (Lalvin CY-3079) and ML bacteria strain(AWRI Lc5p) were chosen. The ML bacteriawere inoculated at different stages duringalcoholic fermentation (AF) (mid exponentialyeast phase and at approximately 5 ºBrixsugar), immediate post AF and 8 weeks postAF, in the presence or absence of yeast, toexamine the effect on MLF duration and theeffect of yeast lees on diacetyl content.

It was found that the timing of O. oeniinoculation affected the initiation andduration of the MLF as reported by othersfor wines. MLF was found to initiate andcomplete more rapidly when O. oeni wasinoculated towards the end of the stationaryphase and during the decline phase of yeastgrowth, particularly if the yeast was removedbefore inoculation. When O. oeni wasinoculated in the presence of fermentingyeast, the MLF was delayed until near thecompletion of the alcoholic fermentation.Insufficient diacetyl data have been obtainedto date to show any clear association withinoculation for MLF and diacetyl content.

This project was undertaken by Ms MariePearce as an Honours project in theDepartment of Horticulture, Viticulture andOenology, with supervision by Drs EvelineBartowsky and Andrew Markides, TheUniversity of Adelaide. Marie was awardeda first class honours degree, and commencedas Graduate Winemaker with SouthcorpWines at Coonawarra in January 2001.

This work has demonstrated that therecently defined model grape juice mediumcan support yeast growth and completionof alcoholic fermentation and the modelwine supports MLF and completion in atimely fashion which is reflective of a ‘real’grape juice/wine. The model alsoreflected grape juice studies with respectto the effect of timing of inoculation withML bacteria on the time course of MLF.

A test system for predicting the stimulatelyand inhibitory affects of wine yeast onmalolactic bacteriaAs stated previously, the aim of this project isto develop a test system for detectingstimulatory and inhibitory interactionsbetween wine microorganisms, and to usethis system to characterise the interactionbetween pairs of popular wine yeast and MLbacteria so that winemakers can choose themost appropriate combination for their wineprocess. Previously, we have described thegrowth response of one Oenococcus oenistrain to four fermentative yeast strains usingthe model test system. Because this testsystem uses a chemically defined medium toreproducibly simulate the importantnutritional and chemical properties of grapejuice, work is in progress to validate this testsystem with several different grape juices.

Juice prepared from hand-picked, disease free,undamaged grapes has been ameliorated toclosely match the chemical composition of themodel medium, with respect to sugar, acidity,nitrogen and so on. Wines have been producedfrom one juice with the test yeasts and thegrowth response of the test ML bacteriumhas been determined. Quantitatively, thegrowth response was lower in terms ofbiomass yields when compared with themodel medium, but qualitatively similarresults were obtained, indicating that thetest system acts as a grape juice model,based on these results derived from a singlejuice. Following validation with the additionalgrape juices, the model system will be usedto screen various combinations of popularcommercial wine yeasts and ML bacteria.

Aspects of this work have recently beencommunicated to industry via i) InstituteRoadshows to winemaker groups in SouthAustralia and North-western Victoria duringNovember/December 2000, ii) presentation tothe Department of Horticulture, Viticulture andOenology Grape and Wine Industry OutreachSeminar, University of Adelaide, November2000, and iii) ‘Can interactions between wineyeast and lactic acid bacteria be morepredictable?’ formed part of an article calledClearer answers sought for old questionswhich appeared in the Aust. N.Z. Wine Ind.J., Nov./Dec. 2000. This project wasundertaken as a collaboration with DrAndrew Markides of The University ofAdelaide (GWRDC Project UA 92/3).

Team Reports

20 20

Days after inoculation

Ace

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)

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citric acid

L - lactic acid

malic acid

diacetyl

acetic acid*

Figure 2. Relationship between diacetyl and organic acid metabolism by Oenococcus oeni in Cabernet Sauvignon wine

The model test system shows promise forthe relatively rapid laboratory assessmentof the inhibitory, neutral or stimulatoryaffect of wine yeasts on the growth andfermentation of a ML bacterial strain basedon the results obtained with a single grapejuice sample. Validation of the test systemwith additional juices is in progress.

Preliminary characterisation of an MLFstimulatory fraction from wineBased on recent reports from the literaturethat a yeast mannoprotein fraction could bestimulatory for MLF, a small project wasundertaken to investigate this hypothesisusing a chemically defined grape juicemedium to simplify analysis. Wines weremade by fermentation with AWRI 838, anMLF stimulatory yeast, and AWRI 839, ayeast with neutral affect on MLF. The wineswere clarified by filtration, and contained <5mg/L total SO2. Various factions of thesewines were prepared with 30,000 and 3,000molecular weight cut-off ultrafiltermembranes. The effect of these fractions onthe growth of a test strain of O. oeni hasbeen assessed by supplementation of asynthetic wine medium.

Growth and malic acid degradation wasgreatest in the unfractionated wine,followed by the 3K eluate, 3K retentate,30K retentate, and synthetic wine medium.This result suggested that the low molecularweight material was most stimulatory tobacterial growth, more so than the highmolecular weight fraction, known to contain(manno)proteins. HPLC analysis of thefractions revealed that the yeast-derivedprotein content was highest in theunfractionated wine and the 30K retentate.With the exception of the unfractionatedwine, the protein content of the fractionsgave a negative relationship to bacterialgrowth and malolactic activity.

This study was undertaken by Ms NinaViergutz as an Honours project in theDepartment of Horticulture, Viticultureand Oenology, with supervision by DrsPeter Costello and Andrew Markides, TheUniversity of Adelaide. Nina has beenawarded an honours degree, andcommenced as an Assistant Winemakerwith Andrew Peace Wines, Victoria, inearly February 2001.

These results suggest that stimulation ofmalolactic bacterial growth may be relatedmore to yeast produced growth factors, suchas vitamins, amino acids and peptides thanto the macromolecular fraction. Our futurework will aim to characterise which factors,if any, are responsible for MLF stimulation.Such knowledge could be used to managermalolactic fermentation better.

Microbiological analysis of industrytechnical problemsStaff: Dr Paul Henschke, Dr Eveline

Bartowsky, Dr Peter Costello, JeffEglinton, Holger Gockowiak, JaneMcCarthy, Dr Leigh Francis, Mark Gishen,Dr Michael Esler, Dr Bob Dambergs

In addition to investigating specific winerymicrobiological problems this project alsoprovides resources for maintaining theInstitute's culture collection which providesyeast and bacteria for teaching and research,performing advanced wine microbiologicalanalysis, undertaking dried yeast qualitytesting, and providing microbiologicalconsultation to industry personnel. Aworkshop on the identification andoenological characteristics of wine spoilagemicroorganisms is being planned for the11th Australian Wine Industry TechnicalConference in October 2001.

The suitability of Near Infrared Spectroscopyfor measurement of yeast assimilablenitrogen content of grape juiceAs reported previously, this project, which isbeing pursued in collaboration with GWRDCAWR 98/2, has the objective to develop andevaluate near infrared spectroscopy (NIRS)for estimating the yeast assimilable nitrogencontent (YAN) of grape juice as measured byestablished reference chemical methods.The poor correlations previously observedbetween NIRS and chemical measures ofYAN were thought to relate to low signaldue to the low relative concentration of freeamino acids in grape juice. Work onevaluating several techniques to enhance theweak NIRS signals has recommenced with DrBob Dambergs recently joining the Institute'sNIRS project team.

Restarting stuck fermentsThis subproject, detailed in previous annualreports, has been concluded with acommunication to industry (Eglinton andHenschke [2001] Aust. N.Z. Wine Ind. J. 16:77–81. Institute staff publication #657).This paper relates to the effect of acetic acidconcentration on the ability to restart a stuckfermentation. Aspects of this work withflavour implications have been taken up inproject GWRDC AWR 2. A S. bayanusChardonnay ferment which stuck in theHickinbotham Roseworthy Wine ScienceLaboratory was successfully restarted with arescue culture of S. bayanus, prepared by theInstitute's recommended method, indicatingthat yeast other than certain strains of S.cerevisiae can be used to restart stuckferments under appropriate conditions. Thiswine, and others rescued by aeration/stirringor with S. cerevisiae are undergoing sensoryevaluation to determine the effect of therescue procedure on wine flavour.

Mousy off-flavour occurrence andformation in wineTwo manuscripts communicating themicrobiology of mousy off-flavour formationin wine to industry have been published:• Grbin and Henschke (2000) Mousy off-

flavour production in grape juice and wineby Dekkera and Brettanomyces yeasts.Aust. J. Grape Wine Res. 6: 255–262(Institute publication #650);

• Costello, Lee and Henschke (2001) Abilityof lactic acid bacteria to produce N-heterocycles causing mousy off-flavour inwine. Aust. J. Grape Wine Res. (in press).

The main points of these papers are that allspecies of Dekkera/Brettanomyces yeast andmany species of the heterofermentativelactic acid bacteria, including some strains ofthe wine malolactic bacterium Oenococcusoeni, are capable of producing mousy off-flavour N-heterocyclic compounds undercertain conditions. Aspects ofBrettanomyces/Dekkera management in

21

Cu

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Time (days)

Yeast 1

Yeast 2

Yeast 3ReferenceYeast 4

}

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Neutral

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60

40

20

00 5 10 15

Figure 3. Growth of Oenococcus oeni in model wines made by fermentation with four different yeast strains and theunfermented reference model wine. Relative to bacterial growth in the model wine, yeast 1 and yeast 2 are designated to bestimulatory, yeast 3 neutral and yeast 4 inhibitory.

winemaking in relation to mousy off-flavourand formation of volatile phenols waspresented to the Southern Pinot NoirWorkshop held in Methven, Canterbury,New Zealand, 18-21 January 2001.

Yeast interaction with red grapephenolics and effect on wine sensory properties

Staff: Dr Eveline Bartowsky, Simon Dillon, Dr Paul Henschke

A collaboration between the Institute andLallemand has been established to investigatethe interactions between yeast and grapephenolics during red wine fermentation.

Lallemand is contributing a substantial sumof money for our investigations, for whichwe are most appreciative.

Red grape must contains a variety of phenoliccompounds which contribute to the colour,aroma, flavour, texture and stability of wine.The principal groups of phenolic compoundsare the monomeric (flavanols, proanthocyanidins,anthocyanidins and flavonols) and polymericphenolics (tannins). Fermentation yeasts arebelieved to interact with many of thesecompounds in a complex manner, involvingphysical, chemical and enzymatic mechanisms.These interactions may include: liberation ofphenolics by chemical/enzymatic macerationof grape tissues; loss from wine byadsorption to the yeast cell wall; intracellularmetabolism (uptake, enzymatic modification,

excretion); extracellular metabolism(enzymatic); and production of chemicallyand physically reactive metabolites (such as,carbonyls and polysaccharides). Many ofthese reactions and their implications forwine properties are not well researched, andmore specifically at the applied level, thespecific properties of individual wine strainsare not well characterised.

A laboratory scale fermentation procedureusing replicated quantities of red grape must(0.25-1.0 kg) has been shown to givestatistically reproducible fermentationprofiles and wine chemical composition.Wine colour density measurements were alsofound to reproduce well.

Comparison of 17 commercial wine yeast fortheir ability to effect extraction and or retentionof various grape phenolic compounds intowine was examined using triplicated 1 kglots of Shiraz grapes (Clare Valley). Thegrape must was adjusted to pH 3.5, 25 ºBrixsoluble solids, 250 mg/L YAN and 25 mg/LSO2, and fermentation was conducted at25ºC with no protection from the air duringcap plunging. All 17 yeast strains completedalcoholic fermentation within 23 days. Onthe basis of wine colour density analysis, theyeast strains were ranked into three groupswhich generally reflects industryobservations, as reported by Lallemand.

Colour density comparisons of wines madewith grapes obtained from three viticulturalregions (Clare Valley, Langhorne Creek andAdelaide Hills) and fermented with six yeast

strains showed a similar ranking for eachyeast. One yeast consistently demonstratedan ability to allow the production of wineswith low colour density whereas anotherconsistently enabled the elaboration ofwines with high colour density. These resultssuggest that yeast strains have a significantinfluence on the phenolic content of wines.Studies are currently in progress to betterunderstand the nature of this observationwith the view to make recommendationsabout the suitability of individual yeasts forspecified fermentation outcomes.

Selection and improvement of wineyeasts by application of molecular biology

Staff: Dr Miguel de Barros Lopes, Dr PaulHenschke, Jeff Eglinton, AnthonyHeinrich, Jenny Bellon and ProfessorPeter Høj

In the past few years a number of moleculargenetic projects studying different aspects ofwine yeast have been carried out, mostlywithin the program of the previous CRCV-1.These included studies on hydrogen sulfideproduction, killer yeasts, nitrogen utilisation,acetic acid formation and glycerol production.Many of these projects have been concluded,and the students researching these areashave recently completed their thesis. Duringthe past year Chris Smyl received his PhDand is currently doing post-doctoral researchon tannins at The University of Adelaide.Nicholas Yap also received his PhD and ispresently employed in the Australian wineindustry. Two of the projects, acetic acidformation and glycerol production, havebeen combined in order to successfullyproduce a yeast that accumulates lessethanol during fermentation.

Construction of a wine strain producingless ethanolIn previous annual reports a commercial wineyeast overexpressing the glycerol synthesis gene(GPD2) has been described. These yeasts(GPD2-OP) accumulate two to three fold moreglycerol than non-modified strains during grapejuice fermentation, and divert sugar awayfrom ethanol production. These modified strains,however, also increase the acetic acidconcentration to amounts generallyunacceptable for winemaking (Institutepublication #649). Several strategies have beentested to decrease the amount of acetic acid.The strategy that appears to be the mostsuccessful is the modification of a secondgene ALD6, which encodes for an aldehydedehydrogenase. Deletion of the ALD6 gene(ald6∆) in an otherwise wild-type straindecreased the amount of acetic acid morethan three-fold, demonstrating that this is aneffective strategy of decreasing aceticaccumulation during fermentation. Importantly,the concentration of acetic acid produced

Team Reports

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Simon Dillon and Dr Eveline Bartowsky

when the cells overexpress the GPD2 gene wasalso diminished when the ALD6 gene wasdeleted (from 1.42 g/L to 0.36 g/L). In fact, theaccumulation of acetic acid with the strainmodified at both genes (GPD2-OP / ald6∆)was lower than that of a non-modified strain(0.66 g/L). The biosynthesis of glycerolremained elevated (increasing from 6.0 g/Lto 16.3 g/L), with a simultaneous decrease inthe concentration of ethanol (from 31.1 g/Lto 27.3) (Note: these ferments were donewith an initial sugar concentration of 80 g/L).

Gas chromatography/mass spectrometry(GC/MS) was used to demonstrate thatGPD2/ALD6 modification affected thebiosynthesis of a number of secondarymetabolites of fermentation, including acids,esters, aldehydes and higher alcohols, manyof which are flavour-active. The concentrationof acetaldehyde, acetoin and 2,3-butanediolare the compounds most affected by theincrease in GPD2 expression. Also, theconcentration of a number of ethylideneglycerols (acetals of glycerol and acetaldehyde),that have been previously found in otherwines that contain high concentrations ofacetaldehyde such as sherry, increased morethan ten-fold in a GPD2 overproducing strain.The formation of secondary metabolites wasinfluenced by the inclusion of oxygen, whichalso affected the growth rate, final cell number,and sugar consumption of the culture. Theresults indicate that modification of GPD2and ALD6 expression represents an effectivestrategy to increase glycerol concentrationand decrease ethanol and acetic acidconcentrations in wine. These modificationsalter the chemical composition of the winesuch that, potentially, novel flavour diversityis possible. A double modification of a yeastby means of molecular biology gives aremarkable demonstration of the power ofthe technology. A paper describing some ofthis work has been published (M. de BarrosLopes, A.U. Rehman, H. Gockowiak, A.J.Heinrich, P. Langridge, P.A. Henschke.Australian Journal of Grape and WineResearch, 2000, Institute publication #649).A second manuscript describing the morerecent work has been submitted for publication.

Use of non-molecular techniques for theimprovement of wine strainsIn order to produce novel wine yeasts withunique characteristics, hybrids between

commercial wine strains and other specieswith winemaking potential are being made.Hybrids between strains of S. cerevisiae andother Saccharomyces species have beensuccessful. These hybrids generally shareproperties of both parents, including ethanoltolerance and the ability to grow in mediacontaining high levels of sugar from the wineyeast, and improved growth at lowtemperature from the second parent. Acommercial wine yeast X S. paradoxus hybridhas been used in a winemaking trial. The hybridfermented grape juice at the same rate as thecommercial wine yeast parent, even thoughthe S. paradoxus parent was unable to fermentgrape juice at all. Results indicate that thegenome of these hybrid yeasts is stable duringfermentation. Chemical analysis demonstratesthat the composition of the wine is changedwhen the hybrid yeast is used. For example,a significant increase in the glycerolconcentration, without a correspondingincrease in the acetic acid concentration,was seen in the wine made with the hybridyeast. Further chemical studies and sensoryanalysis are necessary to evaluate thewinemaking properties of the strain. It is alsobeing tested if this hybrid yeast fermentsmore efficiently at low temperature, acharacteristic derived from the non-S.

cerevisiae parent. These results demonstratethat producing hybrids between S. cerevisiaewine strains and other species is a usefulstrategy for acquiring yeasts with novelwinemaking characteristics from species thatare less suited for grape juice fermentation.One of the main advantages of this methodis that it is considered safe and not classifiedas genetic modification.

Identifying genes for wine yeastimprovementSaccharomyces cerevisiae is a model biologicalorganism and the first eukaryote to have itschromosomal DNA completely sequenced. Thishas been followed by a systematic approachto study the function of each of its morethan 6000 genes. The strains used in thesestudies are laboratory strains of S. cerevisiae,which have important physiological differenceswhen compared to commercial wine strains.For example, phenotypic studies have confirmedthat wine strains tolerate an increased sugarand alcohol concentration, and generallymultiply more rapidly. Also, wine yeast producesecondary metabolites during fermentationthat are positive for wine flavour. The geneand regulatory sequences that specify thesekey wine yeast properties are unknown. ADNA approach (amplified fragment lengthpolymorphism - AFLP) has been used toidentify genes that differ between wine andlaboratory strains. These results demonstratethat the genome of commercial wine strainsare quite different to those of laboratorystrains with several genes duplicated andnovel sequences present.

To further understand the mechanismsbehind the phenotypic differences betweenyeasts, and hence to understand whatmakes a wine yeast perform well underdifferent fermentation conditions, two-

23

Jeff Eglinton, Dr Miguel de Barros Lopes and Alan Pollnitz

Glucose (g/L)1 3.3 4.25 10.95 2.9

Glycerol (g/L)1 5.1 13.4 6.0 16.3

Ethanol (g/L)1 34.1 26.0 31.1 27.3

Acetic acid (g/L)1 0.66 1.42 0.20 0.36

Compound GPD2 GPD2-OP GPD2 GPD2-OPALD6 ALD6 ald6∆ ald6∆

Table 2. Concentration of some metabolites for wild-type and modified strains

dimensional gel electrophoresis (2D gels) hasbeen employed. This system allows proteomecomparisons of different yeast strains underdifferent growth conditions. The wine andlaboratory strains display distinct protein maps,which supports the AFLP results showing thatthere are important differences between thetwo types of strains. Initially, this technique isbeing used to identify proteins that may provideimproved ethanol tolerance of commercialstrains. The results show that the expressionof several proteins are increased in response toethanol stress. A subset of proteins also appearto be destabilised by ethanol. Proteins of interestare being isolated from the gel, digested withspecific proteases, and the resulting peptidesanalysed by matrix-assisted laser ionisation/desorption time-of-flight spectrometry(MALDI-TOF) in collaboration with Dr GraemeCurrie at the University of Melbourne. Theproteins can then be identified by comparisonto databases that exist for the well studiedlaboratory yeast. Characterising proteins thatare altered during wine fermentation will bebeneficial for tracking wine fermentations,predicting strain attributes and selecting andproducing new improved strains.

Grape composition and wine flavour

Staff: Dr Mark Sefton, Dr Leigh Francis, YojiHayasaka, Stella Kassara, GayleBaldock, Professor Peter Høj, WiesCynkar, Dr George Skouroumounis,Kevin Pardon, Dimitra Capone, DrMarkus Herderich, Dr Gordon Elsey,Carolyn Puglisi

This project has the long-term objective ofunderstanding how the combination ofviticultural and winemaking practicesdetermines the aroma and flavourcharacteristics of wine. While some aromaand flavour compounds present in finishedwine are derived unchanged from the grape,the majority are formed by biological andchemical processes from flavourless grapemetabolites. To achieve the aims of thisproject thus requires not only anunderstanding of the compounds that areresponsible for wine flavour but also theprocesses by which they are formed. Onlythen can viticultural practices be linked toflavour outcomes in finished wine.

The program is divided into three broad areas:the identification of grape-derived winecomponents that affect wine aroma and flavour;studies of the formation and degradation ofthese wine flavour constituents including theidentification of their grape-derived precursors;and, ultimately, studies on how viticulturalvariables affect the production of wine flavourcompounds and their precursors in the grape.In order to achieve these aims, it is necessaryto have available routine, sensitive and accurateanalytical methods for determining theconcentration of these grape and winecomponents in various matrices. Thedevelopment of such methods is, and continuesto be, a high priority of the program.

Black pepper aroma in Shiraz winesEarlier work on this project had shown thatthe black pepper character can be tastedand scored in Shiraz grapes from the 1999vintage. Analytical data had indicated thatcertain compounds, that are reported to beinvolved in the aroma of peppercorns, occurin higher concentration in the high-pepperShiraz samples, than in the low-pepperShiraz samples.

Further grape lots were obtained fromvineyards identified from historical data asproducing wines with pepperycharacteristics. Fruit from the 2000 seasonfrom four separate vineyards in Coonawarra,and two lots of grapes from a centralVictorian vineyard, have been studied.Following informal tasting of the berrysamples, only one lot of fruit (from theCentral Victoria vineyard) was characterisedas high in pepper flavour, and this plus a lowpepper sample has been analysed by GC/MS.From data collected from the 1999 samplesanalysed, it was intended to confirmwhether the compounds identified previouslywere also present in the 2000 seasonsamples, and carry out more detailedinvestigations. The project is now on hold asa result of the key researcher, Ms StellaKassara, having taken maternity leave andsubsequently resigned. This project willresume once a suitable replacement forStella is recruited. With the consent of theGWRDC, salary savings from this projectwere employed to further boost theInstitute’s NIR-research by securing theservices of Dr Bob Dambergs.

Synthesis of potential wine flavourcompounds formed from grape-derived glycosidesIn the past, we have tentatively identifiedseveral compounds (four megastigmatrienonesand four maleimides) in mild-acid hydrolysatesof grape-derived glycosides. These mild-acidhydrolysates have been demonstrated, byformal sensory studies, to confer honey, limeand woody characteristics to base wines towhich they have been added (Institutepublication #420). Enzyme hydrolysates

Team Reports

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Kevin Pardon, Dr Mark Sefton and Carolyn Puglisi

GC/MS with tray samples

Nich

olas B

roo

ks at ‘Bro

oksid

e’: 1930Photograph courtesy of The State Library of South A

ustralia

which do not confer these characteristics, do not contain these compounds. They arepatented as flavour additives for otherproducts, but are not commercially available.In order to assess the impact of the individualcompounds on wine flavour it has beennecessary to produce them by synthesis.

The four megastigmatrienones has beensuccessfully prepared, while the synthesis ofthe four substituted maleimides is welladvanced. Following some difficulties withthe final step of the first two sequences thatwere attempted, an alternative route wassuccessful, and two of the targetcompounds were obtained. Thesecompounds will be prepared in sufficientquantities for sensory investigation. Progresstowards the synthesis of the remainingtarget compounds, based on the successfulstrategy towards preparing the first two, iswell advanced, and this task should becompleted shortly. A sensory evaluation ofall of these compounds will then beundertaken – to ascertain their importancefor wine sensory characters. Thecompounds that are found to have verystrong sensory characteristics atconcentrations typically found in wine will bethe target of future viticultural andoenological experimentation.

Precursors to damascenoneß-Damascenone is one of the mostimportant flavour compounds known toscience; it is found in many fruit andvegetable products, including wine where itimparts ‘stewed apple’, ‘fruity’ and honey-like characters, and tonne quantities areproduced commercially for the perfume andflavouring industries. Recent studies by ushave indicated that ß-damascenone in wineresults from the hydrolytic breakdown ofcomplex grape-derived secondarymetabolites formed from carotenoids such asneoxanthin (Institute publication #630).

Two synthetic sequences towards an acetylenicdamascenone precursor were developed inparallel, and the target compound has nowbeen obtained. Consequently, we have beenable to identify this precursor as an intermediatein the conversion of a grape-derivednorisoprenoid to damascenone in fruit, anda preliminary hydrolytic study in model wine hasshown that this precursor gives damascenonein high yield. Once grape samples from thisyear’s vintage have been processed, extractsof these samples will be examined to determinewhether glycoconjugates of this precursorare components of ripe fruit, and whether itwill be desireable to develop techniques formeasuring these compounds in fruit asquality parameters. Further material is beingprepared for detailed hydrolytic studies atroom temperature. The more efficient of thetwo synthetic sequences is now being

employed towards the synthesis of a glycosidicderivative of this acetylenic precursor. Thereactivity of this glucoside at wine pH willthen be determined.

Analysis of other wine flavour compoundsand their grape-derived precursorsPrecise, sensitive and relatively simpleanalytical methods, employing stableisotope-labelled internal standards, arecontinuing to be developed for quantifyingknown key grape-derived wine flavourcompounds. These methods once completedwill enable future research on viticultural andoenological controls of flavour formation,and will be added to over time with furthercompounds identified from the literature orfrom on-going research. Following thecompletion of stability studies of deuterium-labelled analogues, fully validated methodshave been developed for measuring ß-damascenone (fruity and honey likecharacter) �- and ß-ionone (berry, violets),isobutylmethoxypyrazine (capsicum/vegetative),nerol, geraniol and linalool (floral/citrus),1,1,6-trimethyl-1,2-dihydronaphthalene(TDN, kerosine character in aged Rieslingwines) and cis-rose oxide (lychee character,particularly in Traminer wines). Deuterium-labelled ß-phenylethanol, whichhas a floral/rose-like aroma, and labelled ß-phenylethyl acetate (fruity) have beensuccessfully prepared.

Two grape-derived precursors to theimportant wine aroma compound known as‘wine lactone,’ which has woody andcoconut-like aromas in concentrated form,have been synthesised on a large scale.These compounds will be studied todetermine the rate at which they can beconverted to wine lactone under typicalbottle storage conditions. A parallelsynthesis of deuterium-labelled analogues ofthese compounds has also been completed.The deuterium-labelled analogues will beused in future viticultural trials for measuringthe variability in concentration of theseprecursors in grapes.

Ms Agnieszka Janusz from Flinders Universityhas recently taken up an offer of a CRCVpostgraduate scholarship to work on theidentification of new wine flavour compoundsformed from glycosides derived from Shirazand Cabernet Sauvignon grapes. Ms HeatherSmyth (ne Pain) has taken up an Institutepostgraduate scholarship to study therelationship between wine composition andflavour in young Riesling and Chardonnaywines.

The influence of oak cooperage on winecomposition

Staff: Dr Mark Sefton, Dr Alan Pollnitz, Kevin Pardon

Several experiments on shelf-life andextraction rates have been conducted withcommercial sized French and American oakchips, and also with fine oak shavings. Nosignificant change in amounts of extractableoak lactones, vanillin, guaiacol or 4-methylguaiacol were observed after sixmonths storage of the commercial-sizedchips in plastic bags.

Extraction of volatile oak componentsfrom finer shavings was faster than fromchips over the first week of extraction, butthe amount of volatile compoundsextracted over a longer period (28 days)was similar. Soaking for this longer periodis necessary to extract most of the flavourfrom commercial sized chips.

Earlier work in the Institute had shown thatthe presence of oxygen can influence theeffects of heating on oakwood composition.Lignin breakdown by heat is more extensivewhen oxygen is present and this results ingreater amounts of vanillin, guaiacol, and 4-methylguaiacol as well as cis- and trans-oaklactone being formed in the oak comparedto when oak is heated in an inertatmosphere. This indicated that thecomposition of heated shavings, which havea high surface area to volume ratio, will bequite different to that of larger pieces ofheated oak, a supposition supported byrecent experiments.

When oak which had been chipped orshaved to different sizes was heated in air,the finer fragments (shavings) were richer inextractable volatile constituents than werethe coarser fragments. The effect of heatingthe finer shavings in the absence, comparedto the presence of air, was similar to that ofheating coarser versus finer shavings. Theeffects of air and chip size on thecomposition of heated oak pieces was notthe same for all compounds.

Thus the relative proportions as well asabsolute amounts of oak volatiles wasaffected by these parameters, andsmoky/toasty aromas are more likely to bepredominant when smaller pieces of woodare heated, especially if heated in air.

As previously stated, when oak volatiles aretotally extracted into a wine from oak shavingsand the shavings are then removed, thesevolatiles can continue to be generatedhydrolytically, presumably from precursor forms,also extracted from the oak samples.Further studies on exhaustively extracted oakshavings have shown that when these shavings

Team Reports

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are resuspended in a fresh sample model wine,vanillin can continue to be formed over time.The formation of vanillin was greater whenair was present. This demonstrates that oakcontains non-extractable precursors of vanillin aswell as precursors which can be extracted intowine. The accumulation of oak componentsin wine during barrel ageing is therefore notonly a function of extraction rates but is alsodependent on this hydrolytic generation.This new insight has implications for thepotential development of ‘oaky’ charactersin wine following bottling.

Precursors to oak lactoneDespite their sensory importance, the originof cis- and trans-oak lactone remainsunclear. These compounds are alreadypresent in green oakwood, but additionalquantities of these compounds can begenerated in the wood during the drying(seasoning) and coopering processes, in oakextracts heated to 50ºC, and even in theinjector block of a gas chromatographduring analysis of oak extracts. Theseobservations indicate the presence of one ormore precursor forms of oak lactone in oak.Several potential precursors have beenobserved. More than 20 years ago, scientistsfrom the whisky industry isolated, from oakwood powder, a compound which theytermed a ‘precursor’ to oak lactone. On thebasis of rather limited degradation studies,they assigned a structure (a methylatedgallate ester) to this compound. Despite thelimited evidence presented, this compoundas a precursor to oak lactone has gainedcredence through literature citation.

We, therefore, set out to synthesise thismethylated gallate ester in an unambiguousfashion. Because of the possibility that themethylation on the aromatic ring might alsohave occurred as an artefact of the originalisolation, the corresponding straight gallateester was also synthesised. Monomethylgallicacid derivatives are relatively uncommon innature, whereas gallate esters are majorconstituents of oak and other woods.

Both the gallic acid ester of cis-oak lactoneand its monomethyl analogue have nowbeen successfully synthesised. The work hasshown that the monomethylgallate esterstructure originally proposed for the oaklactone precursor isolated and studied 20years ago and accepted in the literaturethereafter is in fact in error. A French group,with whom we have established acollaboration, are using the sample of thestraight gallate ester we have synthesised asa reference to determine whether thiscompound is an oak component and,therefore, potential oak lactone precursor.Understanding of the precursor pool in oakis an important prerequisite for optimal useof oak in cooperage and winemaking.

4-ethylphenol in red wines – implicationsfor microbial problemsIn previous staff publications (Institutepublications #624, #627), we pointed outthat a survey of Australian red winesindicated a relatively high concentration of4-ethylphenol, a product of Brettanomycesactivity, was found in some Australian redwines at concentrations above 500-600mg/L. 4-ethylphenol can impart anundesirable ‘medicinal’/ ‘bandaid’ character.Using our stable isotope dilution assay, ourresearch group measured the concentrationof 4-ethylphenol in wine stored in 44American and 47 French new and used oakbarrels from several suppliers. Wine storedin shaved and refired oak barrels containedup to 85% less 4-ethylphenol and 4-ethylguaiacol than wine stored in normalbarrels of the same age that were notshaved. The concentration of 4-ethylphenolfound in 61 bottled commercial Australianred wines of various ages ranged from 2µg/L in a Merlot up to 2660 µg/L in a Shiraz,with a mean concentration of 795 µg/L. Thehighest level of 4-ethylphenol observed in a‘problem wine’ submitted to the Institutewas 8000 µg/L! Further analyses by ourAnalytical Service of more than 700 differentsamples submitted by industry, paints a verysimilar picture. It appears that winerieswould be well advised to analyse theirproducts for the presence of 4-ethylphenolin order to pinpoint any microbial problemsthat might occur, and to track the efficiencyof corrective measures. Our experienceshows that the ‘problem’ is widespread andthat it can occur suddenly in wineries whichpreviously have not had such problems.

Studies on unstable wine proteinsinvolved in haze formation

Staff: Dr Elizabeth Waters, Dr Leigh Francis,Ken Pocock, Dr Miguel de BarrosLopes, Yoji Hayasaka, Shauna Brown,Professor Peter Høj

Previous work in this project focussed oncharacterising the proteins responsible forwine haze and the effect of irrigation andmechanical harvesting on the proteinconcentration of juice and has beencommunicated in many publications. Theproject has now entered its final stage ofdeveloping alternative methods to preventhaze formation and of exploiting theproperties of the proteins to discriminatejuices and wines varietally.

Use of mass spectrometry todifferentiate varietiesSince subtle differences between themolecular weights of proteins are readilydetectable by electrospray massspectrometry, it is theoretically possible toidentify cultivars by the protein profile of

berries, must and wine. Such anidentification technique would complementand extend the identification servicecurrently available, since DNA fingerprintingis valid for berries and must only. This hasbeen explored and led to a novel approachfor varietal discrimination of fruits.

Proteins with a wide range of masses (13 to 33kDa) were found in the juices of 19 differentvarieties of grape and were identified as mostlythaumatin-like proteins and chitinases. Smallconsistent differences in molecular masseswere noted when otherwise identical proteinswere compared from different varieties. Thesedifferences persisted through different harvestyears and in fruit grown in different locations.Based on the definition of 4 different massesfor thaumatin-like proteins and 12 differentmasses for the chitinases and using statisticalanalysis, the methods developed could be usedfor varietal differentiation of grapes on thebasis of the protein composition of the juice.A refereed paper describing this work hasbeen published (Institute publication #663).The next step is to determine whether themethod can be applied to fined wines wherethe concentration of proteins is much lower.

Proteolytic enzymes and/or heat asalternatives to bentoniteAs reported previously, work at the Instituteand elsewhere has shown that juice andwine proteins are resistant to proteolyticattack at temperatures below 25°C. At highertemperatures, other groups have reportedlosses of proteins and reduced bentoniterequirements for juices and wines treated withproteases. Nevertheless, the use of proteasescombined with heat treatment is not widespreadin industry, possibly due to the perception thatheating under any conditions is detrimentalto wine quality. This appears to be unfoundedbecause previous work at the Institute(Institute publications #444, #462) showedthat short time (2 or 10 minutes)/hightemperature (~90°C) treatment had no clear sensory effect on wine.

Two unstable wines were processed with twodifferent enzymes on industrial scale heattreatment equipment at the Highett facilityof Food Science Australia. The enzymes wereadded to the wines immediately beforeheating at 90ºC for 1 minute. The wineswere rapidly cooled to 18ºC after heating.

This combined heat and enzyme treatmentreduced the protein levels in both wines toabout 40% of the original levels (see anexample in Figure 5). Heat treatment alonereduced the protein levels to about 75%. Thebentonite fining requirement was reduced toapproximately 35% in the combined heatand enzyme treated wines, compared to theheat alone treatment that reduced therequirement to approximately 60%, of thebentonite requirement for the untreated wines.

27

Importantly, and in line with previousInstitute work, sensory evaluation of thewines have shown that the treatmentsresulting in reduced bentonite finingrequirements had negligible effect onaroma and palate in the timeframes andstorage period employed in this study.

The long-term stability of the treated wineslooks promising. After six months:• All wines subsequently fined with

bentonite after heat and or enzymetreatment were stable after storage attypical cellar temperatures (15ºC) andtypical retail temperatures (25ºC).

• Hazes were not detected in any winesafter storage at 15ºC.

• At 25ºC hazes were only detected inuntreated wines and wines treated withone of the enzymes. The heated winesand wines treated with the other enzymehave not given hazes so far even thoughthese wines contain unstable protein.

These experiments were repeated in June 2001with a Yarra Valley Sauvignon Blanc from thecurrent vintage to test whether the proteinreductions are reproducible and whether thesensory impact of the treatment is negligible forpremium wines as well as for the commercialwines tested in the previous vintage.

Treatment of an unstable Sauvignon Blancwine with and without enzymes at 45ºC forlonger time periods has also been assessed.Unlike short term high temperaturetreatment, this process has been shown byothers to have a significant sensory impact.After seven hours (the first sampling time),the combined heat (45ºC) and enzymetreatment reduced the protein level to about30% of the original levels. Heat treatmentalone reduced the protein levels to about80%. After two days, the combinedtreatment had reduced the protein levels to10% with the heat alone reducing the levelto 70% of the original. With one enzyme

there was no protein left after treatment for14 days. The sensory impact of thistreatment is not yet known.

Our results show that for some wines, itis, in principle, possible to reducebentonite fining requirements drastically,apparently without gross short-termsensory impacts.

A poster describing these experiments willbe presented at 11AWITC to communicatethese results to industry.

Haze protective mannoproteinsHaze protective mannoproteins or factors(HPFs) were discovered at the Institute in1991 and are yeast cell wall mannoproteinspresent in wine (Institute publications #439,443, 460, 464, and 471). HPFs have thepotential to prevent haze formation.Previous work has examined methods toextract HPFs from yeast cells or fermentationmedia, and two manuscripts describing thiswork have been published in this reportingperiod (Institute publications #624 and#639). Due to low yields these appear not tobe commercially viable. The project is nowfocused on producing yeast strains that overexpress HPFs.

HPFs have the potential to prevent hazeformation through their ability to change theparticle size of haze. In order to understandmechanism better a bid to augment the HPFproject through collaboration within the CRCfor Bioproducts was mounted and has beensuccessful. Dr Filomena Pettolino wasappointed as a postdoctoral fellow to workwith Professor Tony Bacic (University ofMelbourne) on the physiochemical basis forHPF action. Apart from the Institute’s in-kindcontribution the funds to employ DrPettolino are derived from CRC forBioproducts and Dr Bacic’s group is ofoutstanding quality.

Vanessa Stockdale submitted her PhD thesisin April 2000 and qualified for the award ofthe degree of Doctor of Philosophy inNovember 2001. The thesis will not be

Team Reports

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2000 Chardonnay 2000 Semillon

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Figure 4. The effect of heat and or enzyme addition on the concentration of protein and the bentonite required to stabilize a Chardonnay and Semillon wine.

Dr Elizabeth Waters and Tracey Siebert

available in the public domain until the IPregarding the identity of the HPF genes hasbeen protected.

Shauna Brown has cloned the yeast HPFgenes (HPF1, HPF1’ and HPF2) and successfullydeleted a single copy of all three in haploidand diploid lab yeast. Shauna is nowassessing the relative importance of eachgene for the production of HPF activity.Preliminary phenotypic analyses havesuggested that the HPF mannoproteins mayhave some impact on the ability of yeast totolerate certain fermentation stresses. If thissurvives closer scrutiny, this is an unexpectedbonus from the HPF project and illustratesthe serendipitous nature of research.

Studies on random oxidation of bottledwines

Staff: Dr Elizabeth Waters, Dr GeorgeSkouroumounis, Dr Mark Sefton, DrLeigh Francis, Dr Zhong Kui Peng,Mariola Kwiatkowski

Random post bottling oxidation shows itselfin white wines as an obvious browning in aproportion of the bottled wine typically aftersix to 18 months’ storage, and isaccompanied by loss of SO2 and ascorbateas well as the development of oxidisedflavour. The estimated costs to the industry is$160 million per year in spoiled wine. Wehave worked in this area since 1995 and,excluding oxygenation of wine due to poorbottling procedures, have identified variableoxygen permeability of the closure as amajor cause of the problem. It is possiblethat other winemaking techniques such asuse of ascorbic acid, and upright storage,contribute to the manifestation of randomoxidation (see e.g. Institute publications#577 and #595). This aspect and also anunderstanding of why closures arepermeable have been the focus of theproject during the past 12 months.

The effect of ascorbic acid, bottleposition and wine type on oxidationA Riesling and a wooded Chardonnay wine,for an experiment undertaken withassistance from Southcorp Wines to evaluatethe effect of ascorbic acid, bottle positionand wine type on the extent of oxidation,were bottled in late August 1999. The winesare stored under relatively stabletemperature and humidity conditions in theHickinbotham Roseworthy Wine ScienceLaboratory (HRWSL, annual temperaturerange from 16.9 to 22.1ºC and humidityrange from 45 to 84%). Browningmeasurements on at least 30 replicatebottles for each treatment have been takenthroughout their storage period and willcontinue for another 6 months at least.Other analyses have and will be done at

selected times. After 18 months storage ofthe bottled wines at HWSL the followingconclusions were drawn:• The phenomenon of random oxidation

was not yet evident. • Storage position (upright versus inverted)

had generally no impact on the level ofbrowning in either wine type under thestable temperature and humidity storageconditions used in this experiment. Forone of the natural cork closures there wasgreater browning if bottles were storedupright. The difference in browningbetween bottles stored upright or invertedwas small (maximum of 0.006 at 420 nmafter 18 months) and likely to beundetectable by eye. In the invertedstorage position, this closure is showingconsiderable seepage. It is possible thatthe higher moisture levels in the corkand/or in the cork/bottle interface in theinverted storage position relative to theupright position is contributing to theseclosures in the inverted storage positionbeing a more effective barrier to oxygen.

• For both wines, and at all analysis times,there was greater browning if ascorbicacid had been added to the wine (see an

example in Figure 6). The difference inbrowning between wines with andwithout ascorbic acid was small (maximumof 0.014 AU at 420 nm after 18 months)and was expected to be difficult to pick byeye. This is to be confirmed.

A much smaller and incomplete set ofbottles is stored in an office at AWRI. Thetemperature is controlled at 23ºC duringbusiness hours but uncontrolled at othertimes, and there is no humidity control.These wines are consistently browner thanthe equivalent wines stored at HRWSL (forexample see Figure 6). Similar to the winesin HRWSL, the difference in browningbetween wines with and without ascorbicacid was small (maximum of 0.017 AU at420 nm after 18 months). The relativeimportance of discussed variation andabsolute temperature to the enhancedbrowning of the office-stored wine, is yet tobe elucidated. It should be noted that thebrown colour in the office-stored winewould now be judged too intense by mostindividuals (Institute publication #577).

In summary, this trial has demonstrated that • Upright or inverted storage position had

only a small or little impact on wineoxidation for either wine style whenbottles are stored under ‘best practice’conditions (humidity and temperaturecontrol) for 18 months.

• Addition of ascorbic acid to both winestyles increased the level of browning.

The sensory impact of ascorbic acid additionafter two years storage will be determined inAugust 2001. Immediately following bottlingand after six months storage there was nodiscernible difference in nose or palateattributes for wines with and withoutascorbic acid addition.

It has not been possible to determine theimpact of ascorbic acid addition, wine styleand storage position on the incidence of

29

Storage time (months)

Bro

wn

ing

(A

vera

ged

OD

420

(a.

u.)

)

0.220

0.200

0.180

0.160

0.140

0.120

0.1000 2 4 6 8 10 12 14 16 18 20

Temperature � 15ºC

Temperature (uncontrolled)� 22ºC during business hours

without ascorbic, office

with ascorbic, office

without ascorbic, HWSL

with ascorbic, HWSL

Figure 5. The effect of ascorbic acid addition and storage location on the browning of a Wooded Chardonnay wine sealed with a synthetic closure.

Figure 6. View of the top of the natural cork ‘window’bottles. Araldite was applied to the top of the bottle tocover the closure bottle interface. A circle of approximately30% of the original cork surface area was left exposed inthe centre of the closure.

TSQ Mass spectrometer

Win

e Cellars, M

agill: 1920

Photograph courtesy of The State Library of South Australia

random oxidation because randomoxidation, as judged by browning, has notbeen evident in this trial.

The reader should note that the amount ofascorbic acid added at bottling to thesewines was relatively low (60 ppm) and theconcentration of SO2 relatively high (29ppm). This is particularly important to bearin mind when comparisons to earlierexperiments are made. In these studies (e.g. Institute publication #595) the mostdramatic pro-oxidative effects of ascorbicacid were seen at 200ppm addition levelswith low concentrations of SO2.

The inherent permeability of corks to oxygenAs discussed previously, a large effort hasgone into developing a method that wouldallow us to reliably measure permeability ofoxygen through closures in wine bottles, in areasonable time. A new water-soluble trapof singlet oxygen has been synthesised byKevin Pardon under the supervision of DrGeorge Skouroumounis. This compoundshould be appropriate to measure oxygenpermeating through closures into modelwine. George Skouroumounis and KevinPardon have improved the publishedsynthetic method and currently have morethan one gram of the material. Due todelays in delivery of equipment, no furtherprogress has been made on assessing thecompound in this period.

A small subset of wines from the bottlingtrial described above have been used in apilot experiment to determine whether theclosure or the closure glass interface is themain route for oxygen permeation. Araldite,a material well known for its oxygenimpermeability, was applied to the tops ofbottles over the glass and the closure toeither completely cover the closures of tworeplicate bottles, or to partially cover theclosure so that a ‘window’ remained in thecentre of the closure for four replicatebottles (Figure 7). The amount of closuresurface exposed in these ‘windows’ wasbetween 20% and 30% of the controlclosures and was in the centre of theclosure. These samples have been stored atHRWSL for nine months. In general thehighest level of browning was shown by theuncovered controls and the lowest level bythe covered samples (Figure 8). These resultsare further evidence that wine oxidation is atleast partially due to oxygen permeationthrough closures.

In addition these data suggest thatcoverage of the tops of the bottles andclosures with an easily removed materialwith good oxygen barrier propertiescould reduce wine oxidation. Thebrowning of wines in bottles with closureswith the ‘windows’ was similar to that in the

bottles with the fully covered closures for thenatural closures. For the synthetic closures,browning in bottles with a window washigher than that shown by wines in bottleswith the fully covered closures (Figure 8).Data interpretation must be made withcaution due to the small sample size,however, the results suggest that the relativeimportance of distinct routes for oxygenpermeation into wine might differ betweennatural corks and synthetics. Theseconclusions need to be confirmed with alarger sample set in a custom designedexperiment.

Chemical analysis of industry technicalproblems

Staff: Dr Mark Sefton, Yoji Hayasaka, Dr AlanPollnitz, Gayle Baldock, Dimitra Capone

Problem solving work carried out by thegroup is carried out in collaboration with,and under the direction of the IndustryServices section. Accordingly, the results ofthis work are given in the report by IndustryServices as are extensive studies into theadsorption of flavour active constituents bybag in box material.

A breakdown of industry technical problemsinvestigated by the group over the past 12years has been carried out in order to prioritisetargets for the development of new analyticalmethods for wine taints and othercontaminants. Nearly half of all such problemsare associated with chloroanisoles, and theseare now dealt with using routine analyticalmethods developed earlier by the group forboth problem-solving and research purposes.

Other than chloroanisoles, the mostcommon contaminants identified in wine aresimple aromatic compounds, accounting forapproximately one quarter of the problemsinvestigated. These are mainly associatedwith contamination caused by paints, resinsused for tank lining, and some instances ofoil contamination. Rapid and accurateanalytical methods using solid phasemicroextraction techniques are thereforebeing developed for these aromaticcompounds, which include styrene,ethylbenzene, isomers of xylene,trimethylbenzene, tetramethylbenzene anddiethylbenzene, as well as benzyl alcohol,benzaldehyde and isomeric monomethyl-and dimethylnaphthalenes. Once thesemethods have been fully validated, thegroup will be able to conduct a survey oflocal wines to determine the ‘background’level of these compounds in wine.

General activities of the IndustryServices team

Staff: Peter Godden, Mark Gishen, AdrianCoulter, Peter Valente, Ella Robinson,Greg Ruediger, Heather Donnell

The continuing growth of the Australian wineindustry is reflected in the level of demandfor the services provided by the Institute'sIndustry Services team (Tables 3 and 4).However, following several years of rapidgrowth, the level of demand has stabilised tosome extent over the last twelve months, albeitat what could be considered as historicallyhigh levels. The Institute is convinced that theprovision of these services add value and acompetitive edge to the industry, but alsorecognise that maintenance of a quality

Team Reports

31

natural synthetic stelvin

no coverage

70-80% covered

fully covered

0.17

0.165

0.16

0.155

0.15

0.145

0.14

Figure 7. The effect of Araldite coverage of two closure types on the browning of wooded Chardonnay wines containingascorbic acid after 18 months storage at HRWSL. Closures were covered with Araldite after nine months initial storage. Half ofthe bottles from each coverage and closure type were stored upright, the remainder were stored inverted. Data from bothstorage positions were pooled. Sample size of no coverage for natural and synthetic closures was 60 and for the Stelvin closurewas 12, sample size of 70-80% covered for naturals and synthetic closures was 8, and sample size of fully covered closures forboth natural and synthetic closures was 4.

service will require increased resources, inorder to maintain its value and responsiveness,and therefore relevance to the industry andits development of human capital. Inaddition to the direct services provided towinemakers on an individual basis, theIndustry Services team is conducting severalprojects as described below, and alsocontribute to many other Institute projectsdescribed elsewhere in this report.

Technical problem solving and consulting

Staff: Peter Godden, Adrian Coulter, PeterValente, Ella Robinson

The provision of technical support service tothe Australian wine industry, primarily in theform of an advisory service which disseminatesa wide range of technical information, and aproblem solving and analysis service,represents a significant proportion of theworkload of the Industry Services team. Inaddition, the team manages a major researchtrial that is examining the technicalperformance of various types of wineclosure, and a project which is developing a web-based technical reference manual forthe Australian wine industry, with associatedworkshops which are presented inconjunction with Institute Roadshow seminars.During the year, the first sections of thisweb-based manual went live, providing levypaying wineries with a comprehensivetrouble shooting and diagnostic guide forwine instability problems (www.awri.com.au).

The Industry Services Laboratory analysesseveral hundred samples each year (Table 3),using a wide range of routine and sophisticatedanalytical techniques. The majority of samplesare wine, the analysis of which is supplementedby detailed sensory evaluation by a panel ofexperienced tasters. Of the remaining samplesanalysed the majority consist of wine additives,closures, or compounds which are suspectedto have caused taints and or deposits in wine.The aim of the service is to offer remedial andpreventative advice based on the cumulativeproblem solving experience of the staff, and thepractical winemaking experience of the teamManager, Oenologist and other staff members,rather than providing a simple diagnosis of thecause of the problem. Increasingly, staffregard their role as educational, seeking todisseminate information in a variety of ways inorder to prevent the recurrence of particulartypes of problem. The Industry Servicesteam also provides technical support to theInstitute's Analytical Service, particularly inthe maintenance and auditing of the qualitymanagement system, contract trial-winemaking services, and the interpretationof analytical results. The Analytical Servicealso supplies chemical analysis on problemsolving and research samples to the IndustryServices team, on a contractual basis.

Confidentiality is an important aspect of theservices provided, and is strictly maintainedat all times. Assisted by lawyers, strict Termsand Conditions for the provision of problemsolving services have been developed duringthe year, which reinforces the confidentialityof the work performed. This facilitates afrank exchange of information between theInstitute and its clients, which in turn allowsthe maximisation of the knowledge gainedfrom the provision of these services. Whena particular problem is considered to be ofinterest to the wider industry, the results ofinvestigative work are made availablethrough relevant publications. The IndustryServices team has contributed six TechnicalNotes and six other articles to TechnicalReview during the past year. When preparingmaterial for publication, great care is takento ensure that, under no circumstances is thename of the client, or any possibleidentifying reference, revealed.

As outlined in the previous Annual Report,sensory evaluation is an important analyticaland research tool, and is commonly used inproblem solving investigations, and commercialwork conducted by the Analytical Service.The quantity of sensory analysis performedhas increased substantially during the last year,and the team manager and the Institute'ssensory chemist have continued to developprotocols for the training and assessment of anenlarged sensory evaluation panel. A summaryof the number and type of investigationsconducted by the Industry Services team overthe past three financial years is presented inTable 3. The manner in which problemsolving investigations are reported has beenchanged from previous years, in order tomore accurately reflect the nature ofinvestigations conducted, and the workloadof Industry Services staff. The total numberof samples analysed increased by 33% from

the previous year, while the number ofinvestigations (a figure not previouslyreported) actually declined slightly.The number of samples which were found toexhibit the combined and related problemsof hazes and deposits and microbiologicalinstabilities is disappointing, and as withprevious years, the number of these winesthat have a high pH, a low concentration ofsulfur dioxide, and in some cases incompleteprimary or malolactic fermentations, remainsunacceptably high. The issues related tothese wine instability problems are the firstto have been addressed by GWRDC-fundedproject, Targeted training of wine industrypersonnel: Compilation of a technical referencemanual and delivery of complementaryworkshops, which is discussed in more detaillater in this report. In addition, the issues werealso discussed in an article written by theteam manager in December 2000: Persistentwine instability issues in the AustralianGrapegrower & Winemaker (Institutepublication #645), a seminar paper presentedin Melbourne in July 2000, and lecturesgiven to Oenology students at the Universityof Adelaide in October 2000 and May 2001.

The types of investigations recorded in Table3 as 'other investigative analyses' are varied,and some particularly interesting and unusualcases have been investigated this year.

• two separate investigations (oneconducted for an overseas company on afee-for-service basis) concluded that thedeposits in bottled white wines consistedof a protein-phenolic complex, thephenolic material having originated fromthe corks and the protein from the wines.In both cases the deposit had formed atthe cork/wine interface. It should benoted that it would be considered as quiteusual for phenolic compounds to be

Team Reports

32

Identification of hazes and deposits 79 114 135

Microbiological investigations 48 85 50

Sensory assessments 52 29 51

Taint problems 117 186 66b

Other investigative analyses 245 252 128

Closure-related investigations 154 122 24b

Total 695 788 454c

Samples received (98/99 and 99/00) and investigations conducted (00/01)a

1998/99 1999/00 2000/01

Table 3. Summary of the number and type of problem solving investigations conducted by Industry Services during the past three years

athe manner in which sample numbers have been calculated for Table 3 has changed from previous reports. Someinvestigations require the analysis of multiple samples, while, in some situations, one investigation is conducted onseveral different wines from the same winery which all exhibit the same problem, and all become the subject of asingle report. Therefore, in future, the actual number of investigations conducted will be listed in the body of theTable, while the number of samples analysed will be provided as a footnote.bmuch of the decrease in the number of 'taint problems' and 'closure related investigations', many of which involvethe analysis of multiple samples, can be attributed to a portion of this work now being conducted by the Institute'sAnalytical Service on a commercial contractual basis.ca total of 1048 separate wine samples were analysed as part of these investigations

extracted from corks into wine, andtherefore the issue lies with the need forwinemakers to take this factor intoaccount when protein stabilising theirwines. Some older winemaking textsadvocate the addition of 'tannic acid'when conducting heat stability tests,presumably to simulate the conditions inwhich wine may be placed once it comesinto contact with corks after bottling.

• the previous Annual Report stated thatthree cases of oxidative pinking had beeninvestigated, two of which were in bottledwine. In each case, the wineriesconcerned had not previously experienceda pinking problem in wines made fromfruit from the same vineyards, as thosethat were being investigated. It wasascertained that, in each case, the onlychange in winemaking procedures thathad occurred was the reduction orelimination of the use of ascorbic acid.The incidence of this problem increasedduring the remainder of 2000, with atotal of ten such cases being reported,equal to the total number of casesinvestigated during the previous nineyears. In seven of these cases the wineriesindicated that they had reduced oreliminated the use of ascorbic acid in thevintage in which the wines were made.While only two cases have been reportedin the first half of 2001, previousexperience suggests that oxidative pinkingis a seasonal (and perhaps vineyardspecific) problem and is more likely to bereported in bottled white wines from the2001 vintage in the second half of theyear. Winemakers are, therefore,encouraged to note when bottling theirwines, that the conclusions to be drawnfrom the Institute's previous research intoascorbic acid use (Institute publications#577 and #595), are not that the use ofascorbic acid necessarily needs to beeliminated. Rather, everything else beingequal, the use of ascorbic acid requires ahigher concentration of SO2 to be presentin the wine, if accelerated long-termbrowning is to be avoided.

• the previous Annual Report also describeda Riesling wine that was found to containa concentration of 4-ethylphenol, themajor spoilage compound formed byBrettanomyces yeast, of 2050 microgramsper litre (parts per billion). This wasconsidered an unusually highconcentration, especially in a white wine,based on previous work conducted at theInstitute (Institute publication #623). Thistype of spoilage is poorly documented inwhite wine, and is almost exclusivelyassociated with red wines. During the lastyear the same compound has been foundin a batch of white, bottle fermentedsparkling wine, the concentration in most

samples analysed being between 350 and400 micrograms per litre, with the highestconcentration being approximately 600micrograms per litre. Theseconcentrations would almost certainly beabove the sensory detection threshold forthis wine type.

• the number of wines that are found to becontaminated with unintentional substancesthat are not permitted additives to wine isof concern. During the first six moths of2001, nine cases of refrigerant brinecontamination and six of hydraulic oilcontamination have been investigated.Once again, the issues related to theminimisation of the risks of this type ofcontamination are addressed in theworkshops described above, and at leasttwo of the wineries affected by the recentcases had personnel who attended theworkshops held in November and December2000. These wineries have since reportedthat they have now implemented theadvice given in the workshops, and that ifthey had taken this action sooner, then theirparticular cases of contamination wouldnot have eventuated. As discussed below,a new gas chromatographic analyticalmethod for the identification andquantification in wine of the concentrationof propylene glycol, a common coolant usedin winery refrigeration systems was developedduring the year, to aid these investigations.

• it is pleasing to note that the number ofstuck fermentations reported to theInstitute has declined dramatically overrecent years. However, stuck or sluggishfermentation is still considered animportant issue in the industry, and it isinteresting to note that five out of sevenregions chose this as a topic for discussionat the Roadshow seminars presentedduring November and December 2000.This was in spite of the fact that similarpresentations had been made at theprevious two seminars to be held in eachof these regions.

The first case of stuck fermentationreported in 2001 was particularlyinteresting. A Shiraz fermentation hadvirtually stopped after approximately 80grams of sugar had been fermented, andthe must at that stage contained a

concentration of 1.91 g/L of volatileacidity (VA) expressed as acetic acid.Analysis of the wine further revealed thepresence of numerous, viable, catalasepositive, rod-shaped bacteria. The cellmorphology and catalase activity of thebacteria was consistent with thebehaviour of acetic acid bacteria, but it isdifficult to distinguish betweengluconobacter sp. and acetobacter sp.However, since acetobacter are moretolerant of ethanol than gluconobacter,and since gluconobacter are normallyisolated from grapes and must, have apreference for sugar and appear to beintolerant of the concentration of ethanolpresent in the wine in question, it is likelythat the bacteria isolated from the winewere acetobacter sp. It is further possiblethat that both gluconobacter andacetobacter species were present on thegrapes, and that gluconobacterpredominated early in the fermentationcausing the high VA, and thenacetobacter became dominant

• another case of high VA investigatedduring the year was also of interest, and aTechnical Note based on the investigationwas published in the June 2001 issue ofTechnical Review (issue #132). This caseinvolved a Shiraz wine, one barrel of 25 ofwhich had developed a concentration ofVA above 3 g/L. The wines in ten otherbarrels that were sampled all contained aconcentration of approximately 0.6 g/L.The barrels in which the wine was storedwere predominantly new, and when thewine had last been racked out of barreland blended, approximately four monthsearlier, the winemaker had not noted anunusually high concentration of VA.Numerous viable acetic acid bacteria, thebacteria most usually associated with VAproduction in wine, were isolated fromeach of the eleven barrels which weresampled. Measurements of stavethickness, the 'ovality' of the bungholeand the degree of ullage in each barrelwere also made, and the degree of taperof each bunghole was also noted, but notmeasured. It was found that the problembarrel differed from the others inspectedonly by the ovality of the bunghole, whichwas also noted to exhibit the greatesttaper. A small crack in the bung-stave on

33

Wineries 980 1152 1155

Government organisations 70 99 117

Other 419 413 359

Students 35 36 26

Total 1504 1700 1657

1998/99 1999/00 2000/01

Table 4. Enquiries received by advisory staff during the past three years

one side of the bunghole was also noted,but the ullage in the barrel was less thanthat in many of the other barrels. It was,therefore, speculated that thecombination of imperfections of thebunghole, and the crack in the bung-stave, had allowed a greater concentrationof oxygen to enter this barrel, and thusallow the acetic acid bacteria to becomemore active. Importantly, theimperfections in the barrel would not havebeen identified by casual inspection.

• three cases of cork-boring insects wereinvestigated during the year, bringing thetotal number of cases of this problem tofour in the last eighteen months. In eachcase the insects were found to beinfesting the corks in bottled wines thatwere being stored without capsules orlabels, in bins, in what could beconsidered as standard warehousingconditions. In one case, a brand newpurpose-built warehouse was involved.The four cases involved three differentinsect species, which were identified withthe assistance of CSIRO Entomology.

• an investigation was conducted for awinery which produces sweet wines fromgrapes infected with the mould Botrytiscinerea, in order to ascertain if precipitatesthat tended to appear in the bottledwines over time, and which were thoughtto consist of calcium tartrate, could berelated to the concentration of calcium inthe wines. If this had been found to bethe case, then those batches of wine withhigh calcium levels and thus a greaterpropensity to form a deposit could beidentified, and processed in a way toreduce the risk of deposit formation. A 1993 Semillon wine made frombotrytised Semillon grapes was analysedas part of this investigation. The winecontained a large amount of a whitedeposit that appeared to consist ofcrystals, but existed in clumps rather thanthe more usual crystalline shape of saltssuch as calcium tartrate or potassiumbitartrate. After isolation, microscopyrevealed the deposit did consist of crystalswith a rhombic-like morphology, and thecrystals left a white-coloured residue whenignited in the flame of a Bunsen burner,indicating the presence of calcium.However, the infra-red (IR) spectrum ofthe material did not match that of any ofthe calcium salts contained in theInstitute’s IR spectral library. As the winewas made from botrytised grapes, it wassuspected that the crystals might havebeen calcium mucate. Mucic acid is a by-product of botrytis mould growth andreacts with calcium to form insolubleprecipitates, that deposit slowly. Toconfirm that the crystals were indeedcalcium mucate, a sample of mucic acid

was obtained and reacted with calciumcarbonate to form calcium mucate. TheIR spectrum of the artificially inducedcalcium mucate precipitate was identicalto that of the crystals from the wine.

• during the year there were two caseswhere winemakers were concerned aboutthe possibility of metal fragments fromwinemaking equipment entering theirwines, and subsequently dissolving andcausing irreversible contamination.

In the first case, small fragments offoreign matter from a sulfitometer (apressurised container from which sulfurdioxide, a commonly used preservative,can be added to wine), were isolated ongauze and submitted to the Institute foridentification. The winemaker wasconcerned that the fragments mightcontain heavy metals. The isolatedmaterial was analysed with the assistanceof staff from the Inductively CoupledPlasma Atomic Emission Spectrometry(ICPAES) unit, at the Waite Campus of theUniversity of Adelaide. The resultsindicated that the material contained alarge proportion of iron and some nickeland molybdenum, all components ofstainless steel. No heavy metals weredetected. Unfortunately, the ICPAES scandoes not detect chromium, anotherimportant component of stainless steel.However, the results do suggest thefragments originated from stainless steel.It is possible that this could occur if waterwere to come into contact with the insideof a sulfitometer; causing the sulfurdioxide to react to form sulfurous acid,which could cause corrosion of thestainless steel.

In the second case, metal fragments werefound to be flaking off nickel-coatedcomponents of a bottle-filling machine.Identification of the fragments wasrequested, as well as analysis of somewines that had recently been bottledusing the filling machine, in order todetermine if similar fragments hadcontaminated any of these wines. Thewine samples and a portion of thefragments were also analysed for thepresence of metals at the University ofAdelaide ICPAES facility, and the staff arethanked for their help in theseinvestigations. The fragments were foundto contain approximately 85% nickel and10% phosphorus, which indicates that thematerial was likely to be a type of nickelalloy. The range of concentrations ofnickel in wine reported in the literaturefrom various countries is large, and all ofthe wines analysed contained nickelconcentrations within this range.However, one of the samples contained aconcentration of nickel approximately

seven times higher than the averageconcentration of nickel in the othersamples. It was concluded that this mayhave been due to contamination byfragments from the filling machine, ormay have been due to differences invariety, soil and viticultural practicesbetween the different vineyards fromwhich the grapes were sourced, eventhough these were neighbouringvineyards owned and managed by thesame company.

Winemaking consultation

Staff: Peter Godden, Adrian Coulter, MarkGishen, Greg Ruediger

The Industry Services team provides awinemaking consultancy service principallythrough the Manager, Peter Godden, aqualified and experienced winemaker, AdrianCoulter, a Graduate in the Diploma inOenology from The University of Adelaide,and Mark Gishen. Greg Ruediger, the TraceAnalysis Laboratory Supervisor, also joinedthe Industry Services team on a 0.25 basisduring the year, and also holds the Graduatein the Diploma in Oenology from TheUniversity of Adelaide. Adrian Coultergained nine weeks of practical winemakingexperience during the 2001 vintage, workingat a winery close to Adelaide. GregRuediger has gained similar experience inboth the 1999 and 2000 vintages.

Most queries received are technical in natureand arise predominantly from Australianwinemakers. However, many queries arealso received from wine industry suppliersand Government bodies, as well as arelatively small number from the generalpublic and secondary and tertiary students.Where appropriate, the query is answeredover the telephone, by facsimile or e-mail, andIndustry Services staff supply approximatelyfive hundred technical papers or other piecesof relevant literature to callers each year.Similarly, the support facilities provided byresearch and library staff are important insupplying relevant information to callers, andthe analytical capacity of the Industry ServicesLaboratory plays an important role in respondingto many of these enquiries. In addition,most of the investigations recorded in Table 3result in a full written report being preparedfor the client. These reports contain detailedtechnical information relating to the problembeing investigated and are written in a waywhich seeks to explain the underlying causesof the problems encountered, and provideadvice on how to prevent them re-occurring.The reports are often accompanied by anumber of technical references relating tothe area of investigation.

Team Reports

34

The Institute often acts as a referral service,increasingly for Government bodies, wineindustry suppliers and wine journalists, havinglinks to Australian and international wineresearch and political bodies. The vast storeof information, both formal (in the JohnFornachon Memorial Library) and informal, is a valuable resource to the wider industry.

During the year, the Institute providedassistance to the developers of publicdisplays for the new National Wine Centre.In previous years the Institute's Director,Sensory Scientist (Dr Francis) andwinemaking staff have provided advice tocompanies involved in this project, with aview to ensuring that the displays areaccurate from a technical perspective.During the current year, Greg Ruediger hasadvised on the principles and practices ofwinemaking for typical white and red winestyles, and has reviewed and edited flowcharts for white and red winemakingprocesses. Many of the displays at theCentre will be interactive, and the public willhave the opportunity to make their own'virtual wine.' To this end, multiple choicequestions, along with appropriate weightingfor the importance of various winemakingtechniques for the creation of a range ofwine styles, have been reviewed and edited.In addition, digital images of typical winemicroorganisms have been supplied for usein the displays.

A summary of the enquiries received byPeter Godden, Adrian Coulter, Mark Gishenand Greg Ruediger during 2000/2001 ispresented in Table 4. The figures show a 2.5%decrease in the number of enquiries receivedin the previous year, following steady increasestotalling 35% over the previous five years.Thus, the number of enquiries has apparentlystabilised at a very high level, with theincreased demand for this service clearlyreflecting the expansion of the Australian wineindustry over the same period. The proportionof enquiries received from wineries increasedto 70%, continuing a trend that has beenhighlighted in previous Annual Reports.

The Consultation and Investigative andAdvisory Services are supported by vineyardand winery visits and seminar tours to allmajor wine growing regions, generallyorganised in conjunction with localvignerons’ associations. The Institute aims tovisit each major Australian viticultural regionthrough such formal visits and tours everysecond, or in some cases third year, as thenumber of wine growing regions continuesto increase. Routine shorter visits by keystaff are also made, as opportunities arise—frequently in conjunction with industryevents such as capital city wine shows, andseminars held by other industry bodies.

Formal ‘Roadshow’ visits were made toSouth Australian and Victorian regions inNovember and December 2000 (seeAppendix 1). Up to six senior Institute staffpresented seven full-day seminars, eachseminar consisting of a minimum of twelvepresentations focussing on current areas ofInstitute research, or topical issues in thewine industry. Each regional winemaker'sassociation was asked to select thepresentations to be made at their seminar,from a list of approximately forty areas ofcurrent Institute activity, in order that eachseminar was closely tailored to the interestsof the audience in each region. In addition,a workshop developed under GWRDCproject 99/1 Targeted training of industrypersonnel: compilation of a technical referencemanual and delivery of complementaryworkshops was delivered by Industry Servicesstaff in five regions on the day following theseminar, thus a total of 12 days ofRoadshow activities were presented.The 14th Advanced Wine Assessment Coursewas held in July 2000, giving another 30participants the opportunity to develop andtest their sensory evaluation performance.This was the second Course presented undera four-day format, which includes over 40hours of activities over the four days. Fifteenleading wine show judges, journalists andwinemakers, assisted in the presentation ofthe Course. The demand for the Coursecontinues to be extremely strong, with manyrepeat customers, despite the fact that it hasnot been promoted or advertised, which isan indication of the need for practical trainingof industry personnel away from their individualwork environment. As in the past, severalAssociate Judges for the 2000 Adelaide WineShow were selected from the most successfulrecent participants in the Course, and othershows have expressed interest using Courseresults as part of their selection criteria fornew Associate Judges. The Course continuesto attract interest from the production,marketing, sales and educational arms of theindustry, from all States of Australia, NewZealand, South Africa and the United Kingdom.

Evaluation of new analytical techniquesand of processing aids for winemaking

Staff: Peter Godden, Mark Gishen, AdrianCoulter, Peter Valente, Ella Robinson

The Industry Services Laboratory maintains aGWRDC funded project for the improvement,development and evaluation of methods ofwine analysis, and the evaluation ofwinemaking processing aids and additives.The evaluations take one of two forms: therelative performance of commerciallyavailable products and the evaluation of newmaterials marketed to the industry. IndustryServices staff also provide advice to theInstitute's Analytical Service on the development

of protocols which relate to trials beingconducted on a fee for service basis forvarious wine companies and industry suppliers.

During the year, the vast majority ofresources allocated to this project were usedfor the Closure Trial, a wide-ranging trialthat is examining the technical performanceof 14 different closure types.

Various aspects of theperformance of the closures are beingexamined, including those relating to eachclosure's physical characteristics andextraction from the bottle, chemical analysisof the wine in order to examine apparentsealing performance, and sensory analysis.In addition, Dr Mark Sefton's team willperform chemical analysis using GC/MS, toexamine any effect the closures may have onwine aroma and flavour.

As demonstrated by Table 3, problemsassociated with the use of both natural corkor synthetic closures continue to represent alarge proportion of the problem solvinginvestigations conducted by the IndustryServices team, and an increasing amount ofcommercial work performed by theAnalytical Service. However, until now littlepublished information has been availableregarding the performance of syntheticclosures, or little data available regarding therelative importance of various parameters ofclosure performance, by which natural cork,screw caps, currently available syntheticclosures and closures which may becomeavailable in the future, can be reliablycompared. The broad aim of this trial is,therefore, to elucidate these factors, byexamining a large number of parameters ofclosure performance for a broad crosssection of currently available closure types,on an ongoing basis for up to ten years.From this data, the relative importance ofvarious performance parameters will bedetermined, and this knowledge can then beapplied to the development of testingprotocols by which the future performanceof closures can be rapidly and reliablyevaluated and possibly predicted. Inaddition, the trial will also produce data onparameters relating to bottling and storage,which may then be used to elucidate theimportance of those parameters.

35

Robotic arm with needle

Photograph courtesy of Orlando W

yndham

Senior personnel from the Australian wineindustry with extensive prior experience inthe evaluation and purchase of wine bottleclosures have had significant involvement inthe design of the trial. In addition, extensiveconsultation with closure suppliers ensuredthat the closures examined were representativeof those being used by the wine industry,and ensured that the bottling procedureswere optimal for each closure, as far aspractical. The wine was bottled undercontrolled conditions at an ISO 9002-certified commercial contract bottling facility,with personnel who had prior experiencewith the use of many of the closures. Thedesign of the experiment was motivated bythe need to generate data of practicalrelevance, with all aspects of the protocolmatching as closely as possible currentindustry practice, with due regard for carefulexperimental design and appropriatereplication. The wine used for the trial wasproduced during the 1999 vintage, fromsound Semillon grapes grown in the ClareValley of South Australia, was fermented todryness and was made utilising commonAustralian winemaking practices for wines ofthis style. Importantly, measured variables ofcomposition at bottling indicate that the winehad a low pH (3.1), and contained adequateSO2 (free 30 mg/L total 95 mg/L) to preventpremature oxidation. In addition, the presenceof laccase, an enzyme which can causeaccelerated oxidation of wine, was not detected.

The study is ongoing, and sufficient wine wasbottled so that the testing of many variablesmay continue for up to ten years. Results willbe published periodically, and testing willcontinue for as long as it is considered thatuseful data is being generated.

The published testing protocol (AustralianGrapegrower & Winemaker [425] p 59-64,1999, Institute publication #593), wasexpanded at the 24 months post-bottlingtime point to include the testing of a highernumber of replicates for measurements ofwine composition (SO2, ascorbic acidconcentration and browning), and forsensory analysis. In addition, sensory analysiswas conducted on bottles that had beenstored in both an upright, and invertedorientation, and SO2, ascorbic acid andbrowning were also measured in these samebottles. Testing was also conducted at 18months post-bottling (concentration of freeSO2) and at 21 months (free and total SO2and sensory evaluation on the same bottles),which was additional to the testingforeshadowed in the protocol. This extratesting was conducted, because it appearedfrom previous results that the developmentof the wine had reached a crucial stage.

The large amount of work that has beeninvolved in all aspects of the planning andimplementation of this project culminated inthe submission of a large publication late inthe reporting year. The paper, whichreported the results obtained up to the 20month time point after bottling, wasscheduled to appear on July 12, 2001published as the entire contents of theVolume 7 issue #2) of the Australian Journalof Grape and Wine Research (AJGWR)(Institute publication #666). This trial hasbeen a major team effort, and IndustryServices staff acknowledges the assistance ofthe many members of Institute staff, andpersonnel from other organisations, whohave been involved in the project. Dr LeighFrancis, the Institute's Senior Research

Chemist and sensory evaluation expert, hasbeen responsible for the sensory evaluationwork conducted as part of the trial, and isthe second named author on the publishedpaper – in recognition of his very strong andsustained contribution to the project.

Because the testing at the 24 month pointof the trial was conducted at the very end ofthe reporting period, there was insufficienttime for a full analysis of the data. Howeverdata obtained at the 21 month time pointconfirm the trends of relative closureperformance which are discussed below, andpublished in detail in the AJGWR [Vol 7, #2],which the reader is recommended to consultin order to be fully informed of all relevanttrial details. Inspection of the 24 month datasuggests that these trends also continued tothat time point.

• Wine under the screw cap closure retainedthe greatest concentration of sulfur dioxide(SO2) and ascorbic acid and had the slowestrate of browning, at all testing intervals(including 21 and 24 months). For otherclosures the trend of SO2 loss relative tothe screw cap closure was apparent from anearly stage of testing, and was most evidentin the group of synthetic closures,intermediate in the conventional corks, andleast evident in the technical cork closures.For several closures upright storage tendedto accelerate loss of SO2 from the wine,but in many cases this effect was marginalup to and including 18 months. However,at 12 months there was greater variabilityin the data for bottles that had beenstored upright, as opposed to inverted.

The loss of SO2 was in general highlycorrelated with an increase in wine browning(OD420) and the concentration of SO2 inthe wine at six months was a strongpredictor of future browning in the wine,particularly after eighteen months. Neitherthe concentration of dissolved oxygen atbottling (0.6–3.1 mg/L), nor the physicalclosure measures were predictors of futurebrowning, and the concentration of dissolvedoxygen at bottling was not a strongpredictor of later SO2 loss from the wine.

Seven of the closures displayed a strongcorrelation between the concentrations offree and total SO2 and ascorbic acid (P <0.001) at 12 months, but with the otherclosures the correlation was weak, or didnot exist at all. In addition, wine sealedwith many of the closures contained ahigher concentration of ascorbic acidwhen stored upright as opposed toinverted, although there was only astatistically significant difference for threeof the closures. These observations mayimply that some closures themselves playa role in the degradation of ascorbic acid.

37

Team Reports

Peter Godden and Adrian Coulter

• The closures differed widely in regard tophysical characteristics. All but one of thesynthetic closures, and one of the technicalcorks, were found to elongate to a greaterextent than the natural corks after insertioninto the bottle. One of the syntheticclosures was found to contract uponinsertion. The elongation observed raisespossible issues related to achieving a suitableheadspace (the space between the closuresand the wine) when bottling wine.

In general, synthetic corks appeared least‘consumer-friendly’ in terms of the higherextraction forces and energies required toremove them from the bottle, and ease ofclosure re-insertion into the bottleneckafter extraction. In addition, three of thesynthetic closures required a torque toremove them from a corkscrew that wasapproximately double that required for thenatural corks (while some synthetic closuresrequired less torque than the naturalcorks). However, there was a trend fornatural cork closures to exhibit largervariability in these physical characteristicsthan technical cork and synthetic closures.

All of the closures were found to expandin diameter by a similar amount duringthe 30 minutes following extraction, andthere was no relationship between degreeof post extraction expansion or thechemical and sensory properties of thewine sealed with each of the closures.Likewise, there was no relationshipbetween the physical measurements ofmaximum extraction force, total extractionenergy, and the torque to remove theclosure from the corkscrew, and thechemical and sensory analysis of the winessealed with each of the closures, whendata for a closure which displayed notablyhigh extraction force, and notably poorSO2 retention, (and lowest density) wasremoved from this analysis. In addition,when data from another closure whichdisplayed notably high torque, extractionforce and density was also removed, therewas found to be no relationship betweenthe density of the material from which theclosure was manufactured, and thechemical or sensory analysis of the winessealed with each of the closures.

• Sensory analysis indicated large differencesin wine flavour properties, with closureswhich tended to result in the bestretention of free SO2 having wine sensoryscores for citrus that were generally highwhilst scores for the attributes developed /oxidised were low. The situation wasreversed for wine under closures thatperformed poorly in the retention of freeSO2. It was found that below a criticallevel of free SO2 remaining in the wine(10 mg/L) closures exhibited substantially

higher oxidised aroma. These trendscontinued, and tended to be reinforcedfor all closures, at 21 and 24 months.

Trichloroanisole (TCA)-type taint was anoticeable problem for some cork andtechnical cork closures, at all testingintervals up to and including 24 months.Any plastic-type taint appeared not to bea problem with most synthetic closures upto 18 months, but ratings for the attributeglue-like / plastic by our highly skilledpanel, were notably higher for many ofthe synthetic closures at 24 months.

Whilst the ROTE closure was rated ashighest for the fruit attributes at alltesting intervals up to and including 24months, a reduced (rubbery or sulfidic)character began to be evident in winesealed with some closures (in generalthose which had tended to retain thegreatest concentration of SO2) at 18months, and was significantly higher inwine sealed with the ROTE closurecompared to all other closures. Thesetrends continued at the 21 and 24 monthtime points, with the intensity of thereduced attribute increasing.Closures that scored highest for theoxidised attribute at 18 months also tendedto display the greatest bottle-to-bottlevariation in the rating of this attribute.

ConclusionsDifferences in bottled wine composition thatresult from the use of closures of differenttypes have been assessed, and importantly,the study has allowed the compositionalmeasures of SO2 and degree of browning tobe directly related to the sensory propertiesof the wine. It is apparent that many of theclosures included in the study are suited toshort term (approximately 6 to 12 months)storage of wines, but for longer time periodsthere is doubt over the abilities of variousclosure to act as an adequate seal, withconsequent effects on antioxidant levels,browning and on oxidised aroma. It wasapparent early in the trial that the closuresshowing lowest SO2 concentration wererated as sufficiently high in oxidised aromato consider the wine from these bottles asmarkedly lower in quality. These issuesbecome particularly important when itcomes to managing stock on shelves.

The apparent importance found in this workof a critical concentration of free SO2(approximately 10 mg/L) below which winesare perceived as substantially affected byoxidised aroma is of considerable interest.This observation requires confirmation todetermine whether this might be a generallyapplicable cut-off value, or at the least mightbe appropriate as a guide for white wines ofthe style used in this study. If this observationwere confirmed, the periodic measurement

of SO2 in bottled wine during storage couldbe used by wine companies, distributors andretailers, to identify batches of wine that areapproaching this critical level, and thus takethe appropriate action in the marketing ofthose wines. It is possible that by increasingthe concentration of SO2 in wines at bottlingto compensate for the losses over timeidentified in this study, winemakers couldincrease the longevity of their bottled wines.

For a broad range of performance measures,variability among bottles from the sameclosure was not a serious issue, except forthe TCA-affected samples, but there wassome variability evident in the SO2concentration, where several closures hadrelatively large variation. Whilst no closurescould be considered to be showingunacceptable variation in SO2 concentration,indicating that so-called 'random bottleoxidation' was not apparent up to 18months of testing, there was greatervariability in the SO2 concentration in bottlesstored upright. In addition, the closures thatdid exhibit higher oxidised aroma at 18months also tended to display greater bottle-to-bottle variation on the rating for oxidised.

It will be of interest to note if this observationis manifested as greater variability in winedevelopment at later time points.

Overall, technical cork closures were foundto exhibit less variability than syntheticclosures, which in turn tended to be lessvariable than natural cork closures, whenassessed for a number of the physical andwine composition variables. However, theredid not appear to be a trend for eithermoulded or extruded synthetic closures,when considered as groups, to be more orless variable than each other.

One objective of this study was to identifythe most important variables of closureperformance, and then apply this knowledgeto the development of protocols that can beused by the wine and closure industries forthe rapid and reliable testing of new orexisting closures. Sulfur dioxide concentrationwas found to be a particularly importantmeasurement when assessing the comparativeperformance of closures, and short-termstorage followed by simple SO2 or browningmeasurements provided an excellentindication of later sensory performance.

No single closure tested in this study wasconsidered entirely suitable by all criteriaassessed, for the long term storage of wine,although the ROTE was the best performingclosure in terms of chemical measures ofwine composition, and sensory evaluation. Itis possible that the reduced character whichwas particularly identified in wine sealedwith this closure, was at least in partexacerbated by the manner in which this

Team Reports

38

closure was used which, while still complyingwith the closure's specifications of use, didseek to exclude oxygen from the headspace.

The incidence of ‘cork taint’ (TCAcontamination of the wine) was consideredan important factor with all four of the cork-based closures tested. Wine sealed with oneclosure was affected by a styrene-like off-flavour when assessed at the six month timepoint. However, due to the very poorretention of SO2 displayed by this closure, itwas excluded from further testing.

Note: Readers are asked to keep in mindthat the current study is ongoing, and thatthe results reported here generally representthe performance of the closures underexamination only up to 20 months postbottling. It is possible that, over time, therelative performance of various closures maychange, or that closures which haveapparently performed well to this point maybe found to perform less well in futuretesting. Readers of the Annual Reportshould also refer to the full publication ofthe closure study (Institute publication #666)to understand the full context in which thisstudy was performed.

Analytical method development and evaluation

Staff: Peter Godden, Mark Gishen, AdrianCoulter, Peter Valente, Ella Robinson

Although the majority of resources allocatedto the evaluation of methods and processingaids project has been spent on the closuretrial during the last year, some developmentof methods has also been conducted.

During the year, the Industry Services teamcommissioned a new Perkin Elmer Fouriertransform - infra-red (FT-IR) spectrometer, anew Agilent 1100 Series High PerformanceLiquid Chromatography (HPLC) system, anda reconditioned Hewlett Packard 5890 SeriesII gas chromatograph (GC). The assistanceof the GWRDC in providing capital funding forthis equipment is acknowledged as is the verysubstantial contribution of Analytical Service.

The Institute’s internal Quality System (basedon NATA requirements) required that theanalytical assays previously performed usingthe old HPLC and GC systems, were validatedfor the new equipment. Validations of keyassays required for the closure trial andproblem-solving investigations, ascorbic acidand the 'organic acid profile,' were the firstvalidated for the HPLC, whilst the methodsfor methanol, ethanol and ethyl acetatewere validated for the new GC system, andvalidation of the GC method foracetaldehyde has commenced.

In response to the high number of cases ofrefrigerant brine contamination reported tothe Institute, as described above, a methodfor the determination of propylene glycol inwine, using gas chromatography (GC) hasbeen developed and validated. Previously,the chief analysis available to Industry Servicesstaff when investigating cases of this sortwas the concentration of methanol, a commoncomponent in refrigerant brine solutions.However, this was considered unreliable inidentifying low levels of contamination. Twoof the five cases of suspected contaminationanalysed since the method was developedwere found to contain concentrations ofpropylene glycol far in excess of the normallevels reported in the literature.

Good progress has been made on thedevelopment of an assay for the identificationof isovaleric acid in wine. Isovaleric acid isreported in the literature as an importantspoilage compound produced byBrettanomyces yeast, and a number of winessubmitted to the problem solving service arethought to possibly contain this compound.

Many wines are also submitted to the problemsolving service that apparently contain sulphurcompounds which cannot be removed bythe addition of copper sulphate, a legal andrelatively common technique employedworldwide for the removal of hydrogen sulfideand mercaptans from wine. Therefore, thedevelopment of methods for the identificationand quantification of these compoundswould be of great benefit. During the year aliterature review has been conducted on theanalysis of sulphur containing compounds inwines, ranging from the simple volatilecompounds such as sulphides andmercaptans to more complex and lessvolatile thiol containing compounds, with aview to the development of such methods.

The superiority of the new FT-IR spectrometercompared to the previously used dispersiveIR instrument has been demonstrable. Spectraare obtained in a matter of seconds usingthe new FT-IR and are stored as digitalinformation. This facilitates a range ofinterpretive techniques including matchingwith reference spectra from spectral libraries,identification of mixtures of dissimilarmaterials, and quantification programs. TheIndustry Services team is continuing to compileits own spectral library using the newinstrument, which has been useful inidentifying a number of deposits and increasingthe confidence of positive identification ofsamples, compared with the previous system.

In conjunction with the Institute's AnalyticalService, the following analytical methodshave been validated during the year, to thestandards required by the Institute's internalQuality System, which is based on NATArequirements and ISO Guide 25: Malathion

and Fenitrothion in grapes and marc by GC-MS; Fludioxonil, pyrimethanil, fenhexamid,tebufenozide and cyprodinil in wine, juice,grapes and marc by solid phase extractionfollowed by HPLC, Eugenol in wine andwood products by GC-MS; Spiroxamine,indoxacarb, trifloxystrobin, axoystrobin inwine, juice, grapes and marc by GC-MS;Trifloxystrobin, axoystrobin in wine, juice,and grapes by GC-MS; Fludioxonil in juiceand wine by GC-MS, and, Cyprodinil in wineand juice by liquid-liquid extraction followedby HPLC. In addition, NATA accreditationwas obtained for the following methods:Determination of ochratoxin A in wine byHPLC, and, Malathion and Fenitrothion inwine and juice (pending final NATA approval).This work is conducted by the Institute'sQuality Liaison Manager, Mr Mark Gishen,who is part of the Industry Services team.

Flavour scalping

Staff: Dr Mark Sefton, Alan Pollnitz, Dimitra Capone

A detailed study on the capacity of two-litrebag-in-box packaging to absorb wine flavourcompounds has been completed. Changes inwine composition resulted both fromhydrolytic reactions in the wine (determined bymeasuring changes to the wine stored in sealedglass ampoules) and from contact with thepackaging. Some wine components wereabsorbed rapidly and near-completely fromwine by the plastic components of thepackaging, while others were essentiallyunaffected by this process.

The volatile wine components that were themost efficiently removed were the leastpolar. Thus, 1,1,6-trimethyl-1,2-dihydronaphthalene (TDN), which has akerosene-like aroma and can be animportant contributor to the aroma andflavour of Riesling wines in particular, wasslowly generated from precursor forms in aSemillon wine stored in the sealed glassampoules for 32 days, but more than 97%of this compound was absorbed when thesame wine was stored under identicalconditions in bag-in-box packaging (figure 8).Seventy five percent of this absorption tookplace following the first 24 hours of contact.Naphthalene, which is occasionally found asa trace taint compound in wine and has astructure similar to that of TDN was lessefficiently absorbed. Nevertheless, some 70% ofthis compound was removed by the packaging,again, mostly within the first 24 hours.

A series of ethyl esters produced duringfermentation and including several which aremajor contributors to the fruity character ofwine were included in the study. Theconcentration of these compounds changedduring storage of the Semillon wine in sealed

39

glass containers for 32 days, but not in asystematic way – most decreased slightly inconcentration during glass storage, while theconcentration of ethyl octanoate increasedslightly. For the most part, more substantialdecreases in concentration resulted fromstorage in the bag-in-box packaging, andthese decreases were greater for the longerchain (and therefore less polar) esters. In thecase of ethyl decanoate, a combination of a50% reduction in concentration resultingfrom hydrolysis and an 85% absorption of

the remaining ester by the packagingresulted in most of this compound beingremoved from the wine over 32 days.Decreases in concentration, attributable toabsorption of ethyl octanoate, ethylhexanoate and ethyl butanoate were 55%,30% and 10% respectively.

Absorption played less of a role in thechanging concentration of several other winecomponents. Thirty-five percent of ß-ionone,which can give an aroma of violets to red

wine and is found in highest concentrationin Pinot Noir wines, was absorbed over 32days, and 27% of cis-rose oxide, which isassociated with a lychee aroma, particularlyin Traminer wines, was absorbed over thesame period. A more than 50% decrease inß-damascenone took place in both glass andbag-in-box packaging. In this case absorptionphenomena were not responsible for thechange in concentration.

A variety of flavour compounds associatedwith oak products were unaffected bystorage in the bag in box packaging. Thesewere the cis- and trans-isomers of oaklactone which, depending on concentration,give coconut, vanilla and woody charactersto wines, vanillin which is the principalaroma component of natural vanilla,guaiacol and 4-methylguaiacol (smokyaromas), and 4-ethylphenol and 4-ethylguaiacol which are commonly formedby Brettanomyces yeasts during barrelmaturation and which are associated withelastoplasts, medicinal and clove-like aromas.

This study with the bag-in-box packagingclearly demonstrates the capacity of someplastics to absorb flavour compounds fromwine in a relatively short time. Earlierresearch had also shown that natural cork-bark closures could similarly absorbchloroanisoles from wine, albeit slowly(Institute publication #616). This indicatedthat wine bottle closures, be they naturalcork bark or synthetic, might be able tomodify wine flavour by absorption, and thatsuch absorption could contribute to flavour

Team Reports

40

Dimitra Capone and Andres Hakkensson

Differential behavior of known aroma compounds in bag-in-box

CHO

OCH3

OH

TDN

(u

g/L

)

Time (days)

0

10

20

30

40

50

60

70

0 1 2 4 8 16 32

Van

illin

(u

g/L

)

Time (days)

0

100

200

300

400

500

600

0 1 2 4 8 16 32

Figure 8. Drastic differences in the behaviour of known aroma compounds is observed when stored in bag-in-box containers. Known amounts of trimethyldihydopthalene(TDN) responsible for the ‘aged Riesling/kerosene’ character, and vanillin (a known volatile compound extracted from oak) were added to a 1999 Clare Valley Semillon andtheir concentration followed over time as indicated. The published aroma thresholds for the two compounds are indicated with the red line.

Cork extraction test

Photograph courtesy of Orlando W

yndham

changes associated with bottle-ageing. Along term study of flavour absorption frombottled wine by various types of closure hastherefore been undertaken and the winesare approaching the end of the second yearof storage. A report on changes duringthese first two years of storage will be madein the following Annual Report.

Links between viticultural andoenological research

Staff: Dr Sally-Jean Bell

During 2000/2001, the Viticulturistresponded to 592 enquiries, 86 more than inthe previous 12 months. The majority ofthese enquiries related to agrochemicals.The Viticulturist participated in severalResearch to Practice workshops, in differentstates, and Institute Roadshows during theyear (details provided in Appendix I).

Nine thousand copies of the Institute’s annualpublication, Agrochemicals registered for usein Australian viticulture 2001/2002 wereproduced and the information duplicated onthe Institute’s website. The booklet wasdistributed with the Australian Grapegrower& Winemaker, Technical Review and in theResearch to Practice™: IPM and SprayApplication manuals. The tables are featuredin Australian Viticulture. The 2001/2002MRLs for Australian export markets was alsoupdated for the Institute’s website.

As a direct result of the Institute’sagrochemical publication and its ResidueAnalysis Service, the Viticulturist and GregRuediger have been devoting an increasingamount of time in liaising with major chemicalcompanies. Companies are increasingly awareof the importance the wine industry placeson meeting export market specifications. Thus,they are keen to work more closely with theindustry and the Institute to ensure that theirproducts can be used in viticultural pest anddisease control programs in such a way thatthe maximum residue levels set by Australianexport markets will not be exceeded.

With short notice, the Viticulturist coordinatedand prepared the application for NationalRegistration Authority (NRA) approvedemergency use permits for four insecticideson behalf of the wine industry for the controlof locusts, and these permits are held by theWinegrape Growers’ Council of Australia Inc.

The investigation into the effect of plant waterstatus and canopy management on thephenolic profiles of grapes and wine incollaboration with Dr Patrick Iland and Dr PeterDry (University of Adelaide), OrlandoWyndham (Langhorne Creek) and ProfessorEnrico Peterlunger (Universita di Udine, Italy),commenced in October 2001. This trial was

conducted by the Viticulturist and a visitingPhD student Mr Paolo Sivilotti (visiting forseven months from Universita di Udine). Todate it has been confirmed that water stressin both VSP and Scott Henry vines resulted inreduced leaf water potential, stomatalconductance, leaf area, canopy density, shootlength, pruning weight and berry weight.Fruit exposure consequently increased andpreliminary assessment indicates that skinsand seeds from exposed fruit had lowerquantities of tannin and anthocyanin thanskins and seeds from less exposed fruit.Wine was made from the fruit and the firststage in the sensory analysis is scheduled tostart towards the end of November 2002.

Preparation of information on wine andhealth issues

Staff: Creina Stockley

This project has been funded since 1990.Ms Creina Stockley, a clinical pharmacologist,assumed this position in 1991 as part of herresponsibilities as Health and RegulatoryInformation Manager. A database of researchon the beneficial and detrimental healtheffects of alcohol and in particular, wine,was established on the internal computernetwork of the Institute and as part of theJohn Fornachon Memorial Library. This wasfacilitated by the subscription to relevantmedical and scientific journals, and by theformal and informal exchange of informationbetween complementary organisations, bothnational and international. During2000/2001, 74 independent informationrequests were received on wine and healthissues from industry, government and thegeneral public by the Health and RegulatoryInformation Manager.

Subscription to relevant medical and otherjournals has continued. The journals havebeen regularly scanned, the database ofresearch on the health effects of wine hasbeen added to and articles have beenprepared for inclusion in the Institute’spublication, Technical Review and for otherAustralian wine industry and internationalalcohol industry newsletters. Articles andother material have also been prepared forthe electronic and print media, for example,six articles have been prepared for thebimonthly international publication, AIM—Alcohol in moderation, and four articles forthe quarterly newsletter of the AustralianSociety of Wine Education. The Health andRegulatory Information Manager is also amember of the editorial board of AIM—Alcohol in moderation. A national radiointerview and a television interview were alsoundertaken related to the issue of potentialblood products in winemaking. A review ofthe draft Training Procedures Manual for theNational Wine Education and Training Centre

was also undertaken, as well as preparingmaterial to support the WFA-funded Costsand benefits of drinking wine–A discussionand review of Collins and Lapsley by theCentre for International Economics.

Submissions prepared in support of theAustralian wine industry include:• Draft Australian Alcohol Drinking

Guidelines of the National Health andMedical Research Council of Australia;

• National Alcohol Research Agenda of theFederal Department of Health and AgedCare;

• Foetal Alcohol Syndrome Draft ScopingPaper of the National Expert AdvisoryCommittee on Alcohol; and

• PRELIMINARY DRAFT RESOLUTIONECO/REGL/96/23/Stage 3 on ingredientlabelling and PRELIMINARY DRAFTRESOLUTION OENO/ASP/99/129/Stage 3on a health warning for the of theNutrition and Wine Expert Group of theOffice International de la Vigne et du Vin(OIV) (Creina Stockley is a member of theAustralian OIV delegation—Nutrition andHealth Sub-commission).

The Health and Regulatory InformationManager has been instrumental in organisingand developing the program for the wineand health symposium session of the 26th

World Congress and 81st General Assemblyof the Office International de la Vigne et duVin to be held 11–18 October 2001. Thetheme of this session is "Current wine andhealth information: the issues in perspective".There are also three sub-themes: 1) Thecurrent and future status of wine and healthinformation; 2) The need for consumers torelate this information to their diet andlifestyle; and 3) Responsible communicationof wine and health information.

Institute and industry consumer health andsafety information and support systemThe Health and Regulatory Information Managerhas also prepared the Institute’s and industry’sconsumer health and safety information andsupport system, which is accessible on theInstitute’s web-site www.awri.com.au.

Project coordinationThrough Creina Stockley, the Institute hasplayed a coordinating and a participatingrole in three GWRDC-funded researchprojects on medical aspects of wineconsumption entitled AWR97/2 Potentialcardio- and cancer-protective effects andmechanisms of wine. GWRDC pays theHeart Research Institute, The University ofWestern Australia and CSIRO Health Sciencesand Nutrition directly. The respective projecttitles are: Potential cardioprotective activitiesof wine components based on synergisticinteraction with vitamin E; Grape antioxidantphenolics: absorption and inhibition of lipidperoxidation in humans; and Reduction of

Team Reports

42

damage to LDL and DNA from oxidative freeradicals by the regular and moderateconsumption of wine. All three projects havecompleted their third year of funding.

Technical and regulatory support to theAustralian wine industry

Staff: Professor Peter Høj, Creina Stockleyand Rae Blair

Information requestsOne of the activities of the Institute has beento provide legal/regulatory and technicaladvice and assistance to the Australian wineindustry, through the Health and RegulatoryInformation Manager, the Industry Servicesteam and the Director. During 2000/2001,110 independent information requests ontechnical and regulatory issues from thegovernment and industry were fielded by theHealth and Regulatory Information Manager.

Industry committee membershipAdditional support to the industry is derivedfrom the Director’s membership on the AWBCCompliance and Technical Advisory Committee,International Trade and Technical AdvisoryCommittee and the WFA Technical Committee.One of the important aspects of the Institute’ssupport of the Australian wine industry is itspivotal role in facilitating the triennialAustralian Wine Industry Technical Conference(AWITC), in conjunction with the AustralianSociety of Viticulture and Oenology. TheDirector is the Chair, and the PersonalAssistant to the Director is the ConferenceManager and Treasurer. Professor Høj andMs Blair are also members of the AustralianOrganising Committee of the 26th WorldCongress and 81st General Assembly of theOffice International de la Vigne et du Vin tobe held 11-18 October 2001.

At present, the Health and RegulatoryInformation Manager is a member of thefollowing committees: the AWBC LegislativeReview Committee; the WFA TechnicalCommittee; and the National ReferenceCommittee-Environment Strategy Development(SAWBIA). Ms Stockley was also electedVice-President of the Nutrition and WineExpert Group in March 2001.

Reviews and publicationsDuring 2000/2001, supplementary to theStandard 2.7.4 for wine and wine product ofthe [joint] Australia New Zealand FoodStandards Code that was gazetted inDecember 2000, a draft industry standardfor wine, sparkling wine, fortified wine, wineproducts, reduced and low alcohol wineswas prepared by the Health and RegulatoryInformation Manager in conjunction withthe AWBC and WFA, for potential inclusionin the Australian Wine and BrandyCorporation Act 1980 and Regulations.

Significant efforts, energies and resources havealso been directed towards ensuring theestablishment of realistic and reasonablemaximum limits for the additives and processingaids approved for use in winemaking by theCodex Alimentarius Commission, on behalfof the AWBC, WFA and AFFA (Wine, Vineand Citrus Section). Technical informationand/or issues that have been reviewed, anddiscussion or position papers prepared orcontributed to during 2000/2001 include: thenational system of gene technology regulationof the Therapeutics Goods Administrationand Australia New Zealand Food Authority;ochratoxin A; silver nitrate; tannin andtannin-like additives; current status ofcountry of origin labelling; acidification withlactic, malic and tartaric acids; wine as asource of dietary sodium; BSE and theAustralian wine industry; and factorsinfluencing the sodium and potassiumcontent of Australian wines.

Technical Information is also disseminated tothe Australian wine industry, including allgrape and levy payers, by the institute’sbimonthly publication, Technical Review, ofwhich the Health and Regulatory InformationManager is editor, and by the Institute’sAnnual Report edited by the Director andCommunication and Publicity Manager.Australian Wine Industry Technical ConferenceSeveral Institute staff members, namely,Professor Peter Høj, Peter Godden and RaeBlair, serve on the Conference PlanningCommittee of the Eleventh Australian WineIndustry Technical Conference, and ProfessorHøj and Mr Godden served on the ProgramSub-Committee. The formal program hasbeen developed and will cover eight twohour sessions from Sunday, 7 October toWednesday, 10 October. Some 74workshops are being planned, and theworkshop program, being convened by MrGodden, will run from Sunday, 7 October toThursday, 11 October. Institute staff willconduct several workshops and preparemore than 30 posters. Jeremy Hack acts asthe poster coordinator.

This project of technical and regulatory supportto the Australian wine industry is ongoing,as technical and regulatory issues are regularlyraised by the government or by industry, bothin Australia and internationally. Furthermore,these issues often span several years.

Provision of technical information

Staff: Rae Blair, Catherine Daniel, Ingrid Oats

The John Fornachon Memorial Library holdsthe largest collection of wine technicalliterature in Australia. The Library’s principalresponsibility is to provide technicalinformation to the Australian wine industry

and to the researchers of the Institute. TheLibrary is also used extensively by othergroups such as students, government bodiesand private companies (see Table 5).

Information and document delivery servicesThe Library has excellent access to internationaldatabases, particularly in the fields of science,technology and medicine. If requested, theLibrarian, Catherine Daniel, will carry outonline searches on commercial databases onany appropriate topic (on a fee-for-servicebasis). Alternatively, Library staff can provide,free of charge, a report of relevant articlesindexed on the Library’s in-house databases.Requests for information from Library staffcontinue to rise significantly with a 108%increase in information requests for2000/2001 over 1999/2000. Most of theLibrary's services showed increases indemand, with the exception of online searchrequests and requests for articles fromTechnical Review (-3.8%). This minordecrease in requests for articles fromTechnical Review has been more thancompensated for by a 40.5% increase inother articles forwarded over 1999/2000figures. Database searches undertaken oninternet and CD-ROM networked databases,now recorded as Information requests, offsetthe continuing decline in online databasesearches. These specialist databases are nowonly used to access information not availablein the public domain and to deliver specialistcurrent awareness services. The fundingapproved for the third Library staff member(whilst primarily to assist with the loading ofrecords for the web-accessible database) hasfacilitated an appropriate level of service andresponse time to the steady increase inrequests received by the Library.

Document deliveryThe Library can supply either books orphotocopies from its collection or obtainsuch items for wine industry clients throughthe interlibrary system. Patents or standardscan also be ordered. Electronic ordering anddelivery services mean that most interlibraryrequests are fulfilled within five days.Charges apply for the supply of some items.

Specialised information servicesThe Library staff continue to be activelyinvolved in the production of specialisedinformation products for the benefit of thewine industry, such as the annual and web-based editions of the Agrochemicalsregistered for use in Australian viticulture,the bimonthly Technical Review (where theLibrarian is Acting Editor for editions #133and #134), and several in-house technicalinformation databases.

Library collectionA total of 79 monographs and sevenconference proceedings were added to thelibrary collection in 2000/2001. The Library

43

subscribes to 54 journals and receivesapproximately 70 annual reports, journalsand newsletters through exchange anddonation. The Library also maintains acollection of over 19,000 reprints.

Library databasesApart from a computer-based catalogue ofbooks and journal holdings, the Library hasseveral specialist in-house databases, whichindex over 38,700 scientific and technical reprintarticles; over 2,000 articles on the medicalaspects of alcohol consumption; and thebibliographic details of the Library’s collectionof the European Union wine legislation.

The Librarian, Catherine Daniel, providesreports, either on particular subjects orauthors, listing the records retrieved fromany of the Library’s in-house databases. Asummary of the size of the Library’scatalogue and information databases isgiven in Table 6.

The Library provides access to its databases,24 hours a day, via the internet, to Wine andGrape Levy payers only. The restriction inaccess is enforced to comply with copyrightapprovals obtained from the various publishers.Edited database records continue to beposted onto the Library's web database,with the total number of searchable recordsavailable in excess of 10,000.

Agrochemicals GridAs reported elsewhere in this Annual Report,Dr Sally Bell and Catherine Daniel preparedthe twelfth edition of the Agrochemicalsregistered for use in Australian viticulture.

All levy payers receive a printed copy of therevised edition automatically, and the web-based edition (accessible on www.awri.com.au)is updated on a regular basis.

Technical ReviewTechnical Review is received by all Wine Levypaying wineries in Australia and, throughsubscription, by government and otherorganisations and individuals, both in Australiaand overseas. Technical Review providesprogress reports to the industry on theInstitute’s research as well as updates on relevantconferences, regulatory amendments andmedical issues. Technical Review’s ‘Current

Literature’ section provides citation detailsand abstracts of recently published technicaland scientific articles. Recipients of TechnicalReview may order articles featured in the‘Current Literature’ section via a request formavailable within each issue. The Librarian,Catherine Daniel, was Acting Editor of TechnicalReview’s June issue, #133. Dr Barbara HardyAO continues to support the publication ofTechnical Review through financial contributionto the Thomas Walter Hardy Memorial Trust.

Email serviceThe Email advice and information ontechnical issues service continues to be a fastand cost-efficient way of disseminatingimportant technical information to interestedmembers of the Australian wine industry.There are over 530 email addresses recordedto receive the email bulletins to date. Twelveemail bulletins were issued during the yearand are shown in Table 7.

Team Reports

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Rae Blair, Catherine Daniel and Heather Donnell

Wine Staff Other Total % inc/(dec)industry over 1999/2000

Information requests 515 484 1155 2154 108%Online database searches 5 14 19 (47.2%)Interlibrary loans• requests sent1 159 496 655 2.8%• requests received2 159 159 117%Technical Review requests3 167 (21%)Technical Review articles forwarded4 907 (3.8%)Articles forwarded5 662 40.5%Number of Institute publicationsforwarded 440 18%Articles photocopied 3795 n/a

1 Staff at the JFML sent a request to another library for an article.2 Requests received by the JFML from other libraries for articles from our collection.3 Number of requests received for articles published in the Technical Review.4 Number of articles forwarded (usually more than one article is requested).5 Number of articles forwarded, excluding staff publications (usually more than one article is requested).

Table 5. Summary of information requests during 2000/2001

Library catalogue databasesBOOKFILE: Books, conferences and theses 3,304

ARTICLES: scientific papers 38,737

MEDIC: medical papers 2,304

JOURNALS: journals, newsletters,statistics and annual reports 428

Library information databasesREGS: European Community wine legislation 372

ISYS – full text retrieval database coveringUnited States of America Federal Register 689

Web accessible database 10,309

Table 6. Number of records on the Library’s catalogue, information and web-accessible databases

The John Fornachon Memorial LibraryEndowment FundThe Institute acts as the Trustee of this fund,which was established in 1969 by donationsfrom winemakers and friends of the lateJohn Fornachon, the first Director of Researchof the Institute. The Library is funded by anannual grant from the Grape and WineResearch and Development Corporation,together with the income generated frominvestment of the Endowment Fund.

AcknowledgmentsThe Institute wishes to thank all individualsand companies who contribute to thecollection through donations or exchangeagreements. The support of the followingpersons and organisations that have donatedbooks or journals is acknowledged:Australian Bureau of Agriculture andResearch Economics, Australian Dried FruitsCorporation, Australian Wine and BrandyCorporation, Australian Wine and BrandyProducers’ Association, CommonwealthScientific and Industrial ResearchOrganisation, K.F. Pocock, Dr B.C. Rankine,Viticultural Publishing Inc., Winemakers’Federation of Australia Incorporated,Yalumba Winery.

Targeted training of wine industrypersonnel: compilation of a technicalreference manual and delivery ofcomplementary workshops.

Staff: Peter Godden, Mark Gishen, AdrianCoulter, Peter Valente, Ella Robinson

The primary aim of this project is to produceflexible and updateable informationpackages on selected technical subjects,which will be delivered to winegrape levypayers via the world-wide-web, andworkshops to be held in all major grapegrowing areas of Australia in conjunctionwith the existing Roadshow seminars.

The Institute has a considerable amount ofcollective knowledge pertaining to grapeand wine production, much of it generatedover the years by research and IndustryServices' projects that have been supportedby industry research-levy funding. Althoughthis research has produced many technicalpublications, there is also a great deal ofinformation generated and recorded in amore informal manner by the staff concerned.This project, therefore, seeks to make arecord of this information, so that it can bedelivered to the industry in a manner in which itis both readily useable, and relevant to thoseinvolved in day-to-day wine production. It isenvisaged that the project will be ongoing, andthat many areas of interest to winery technicalpersonnel will be addressed in due course.During the year, the first two major objectivesof this project were achieved; the posting oftechnical information and associated detailedproblem solving guide onto the Institute'sweb-site, and the delivery of the firstworkshops developed under this project, infive wine production areas in South Australiaand Victoria. Both the web-based informationand the workshops relate to wine instabilityissues. As discussed elsewhere in this reportthe combined and related area of hazes anddeposits and microbiological instabilitiescontinue to represent a significant proportionof the problem wine sample investigationsconducted by the Industry Services team. Inmany cases it is apparent that the problemscould have been avoided if winemakers weremore aware of the causes of many types ofinstability, and were able to access practicaland relevant tools with which to trouble-shoot the problems, before significant winespoilage had occurred. The web site andworkshops have addressed these issues,providing practical trouble shooting informationand simple diagnostic tests, with which toisolate and identify a wide range of hazes anddeposits in wine. The text is supported by agallery of images of deposits in wine that havebeen generated from routine problem solvinginvestigations. In addition the web siteprovides extensive background informationon the causes of a wide range of instabilities,written by a contracted technical writer and

based on the collective information generatedby various teams within the Institute. Theworkshops also contain a large amount ofpractical winemaking advice regarding notonly the causes of problems, but also theiravoidance, and remedial advice on how todeal with them if they occur.

During the year, work commenced on newsections of the web site, particularly thosewhich will deal with wine microbiology inmore detail, with particular emphasis on theidentification of both wanted and unwantedwine microflora, and good laboratory andwinery practice with regard to winemicrobiology issues. It is envisaged that thissection will be expanded to encompass theissue of stuck fermentation, thus addressinganother cause of potential wine instability, asdiscussed elsewhere in this report. A furthersection on which work has commenced isthe design of a winery laboratory, an areathat has obvious synergies with thosealready addressed, and the subject of anincreasing number of advice calls to theIndustry Services team.

The information on the web site(www.awri.com.au) is password-protected, foruse only by Australian grape and wine researchlevy payers. The password has been supplied toall levy payers via personal letter. Levy payerswho are unsure of the password may obtainit by contacting either the Institute's Librarianor Communications and Publicity Manager.The posting of the technical informationgenerated by this project onto a web-sitewas a large undertaking, and required asubstantial amount of design anddevelopment of the website in order to makeit both as practical and relevant as possible,while at the same time aesthetically pleasing.It was, therefore, decided that concurrentdevelopment of the Institute's overall sitewould generate substantial efficiencies,rather than addressing this as a separateissue at a later date. Industry Services staff,therefore, had a high level of involvement inthe development of a new web site for theInstitute, in conjunction with the Institute'sCommunications and Publicity Manager,Director, and two external design and web-site development companies. The templatedesign developed for material generatedunder this project has been successfullyapplied to many other areas of the site.

Waite Campus Mass Spectrometry Facility

Staff: Yoji Hayasaka, Gayle Baldock

The four important roles of the WaiteCampus Mass Spectrometry Facility are toact 1) as a leader in the application of massspectrometry to grape and wine research; 2)as an investigator to solve the problemsfacing the wine industry and individual

45

18/7/00 Launch of new Agrochemical booklet

18/7/00 Use of Shirlan for control of Phomopsis

19/9/00 Notice of First NationalEnvironment Conference

3/10/00 Switch Fungicide registered forcontrol of Botrytis

4/10/00 Chemical Control of AustralianPlague Locusts in Viticulture

6/11/00 Fenitrothion emergency use permit

6/11/00 Flint fungicide

6/2/01 AWRI denies claims ofendorsement of NuKorc

16/2/01 Blood products for sale fromYeruva SA, Argentina

2/5/01 Non thermal food processingseminar advice

10/5/01 Eleventh Australian Wine IndustryTechnical Conference – updatesand early registration closingdate notification

16/5/01 Eleventh Australian Wine IndustryTechnical Conference – advice ofnew workshop added to program

Table 7. Email bulletins sent during 2000/2001

winemakers, using mass spectrometrictechniques; 3) as a collaborator with TheUniversity of Adelaide in the research andteaching activities involving massspectrometry; and 4) as a provider ofversatile and advanced mass spectrometrictechniques and expertise to the scientificcommunity including public as well asprivate institutions. Much of our work is,therefore, conducted on a fee-for-service basis.

The Facility’s instrumentsThe Facility accommodates four instrumentsnamely a ThermoQuest TSQ gaschromatography tandem mass spectrometer(GC-MS/MS) installed in February 2000, a PESciex ionspray tandem mass spectrometer(LC-MS/MS) installed in October 1995 andtwo Agilant gas chromatographs with massselective detectors (GC-MSD) installed inMarch 1997 and January 2001, respectively.The GC-MSDs are used mainly for grape andwine flavour projects under the supervisionof Dr Alan Pollnitz, and one has been fittedrecently with a Gerstel MPS2 autosamplerwhich also has solid phase micro extractionand headspace capability. The GC-MS/MSand LC-MS/MS are used for various purposesand appropriate financial arrangements forall users are in place to recover the runningcost of the Facility. Table 8 details the timeusage of the GC-MS/MS and LC-MS/MS.

Problem solving workDuring the reporting period, thirteen casesof chemical analysis of either juice or winewere conducted using GC-MS, in collaborationwith staff of the industry services.

As mentioned in the previous annual report,the development of a taint screening methodhas commenced using GC-MS. Thisdevelopment is conducted for the projects of AWR 19 Chemical analysis of industrytechnical problems and AWR 10 Technicalproblem solving and consulting. In order toinvestigate the common contaminantsfrequently encountered in the wine industry,the problem solving work investigated by the Institute using GC-MS in the past 12years was reviewed.

The review revealed that toluene, xylene,styrene, alkyl (C3-C4) benzene,naphthalene and alkyl (C1 and C2)naphthalene were common ascontaminants in the rare faulty juices andwines submitted. These compounds arenot normal constituents of grape juiceand wine but they are volatilecomponents of some petroleumproducts. Therefore, these compoundswere nominated as target compounds ofthe taint screening of juice and wine. Themethod development is in progress.

Research activitiesInvolvement in the research activity of theInstitute is one of the most important rolesof the Facility and its demand has beenincreasing in not only classic applications likevolatile flavour analysis but alsocharacterisation of macro, involatile andlabile compounds like proteins, tannins andglycosidic conjugates. The following researchactivities highlight this function.

• The use of mass spectrometry todifferentiate varieties of juice wasinvestigated under project AWR 8 ‘Studieson unstable wine proteins involved in hazeformation.’ Varietal differentiation of juicewas successfully achieved based on thecomposition of PR-proteins determined byelectrospray mass spectrometry. This workwas published in the Journal ofAgricultural and Food Chemistry (Institutepublication #663) and an oralpresentation was made by Yoji Hayasakaat the 18th Conference for the Australiaand New Zealand Society for MassSpectrometry (4-9 February 2001). Theapplication of this technique for wine isunder investigation.

• Search for wine pigments in red wineusing nanoelectrospray tandem massspectrometry was conducted as part ofproject AWR 96/1 ‘Wine grape tannin andcolour specification,’ in collaboration withDr Robert Asenstorfer of the University ofAdelaide. Wine pigments such as apyruvic acid adduct (vitisin A), a 4-vinylphenol adduct (pigment-A) and avinylcatechin adduct of malvidin-3-monoglucoside have been reported in thepast five years. All wine pigments areformed during and after vinification. Theyare substituted at the C-4 position ofmalvidin-3-glucoside and thought to bemore chemically stable than malvidin-3-glucoside. Nanoelectrospray was used forthe screening of wine pigments(anthocyanin derivatives) in red wine. Theadvantages of this technique overconventional electrospray are the bettersensitivity, the smaller size of a samplerequired (1 µL) and that sufficient time forthe application of various tandem massspectrometric techniques – is available dueto very low flow-rates.

The screening method for wine pigmentswas successfully developed usingnanoelectrospray tandem mass spectrometry.This method was applied to search for newpossible wine pigments in red wine extracts.Apart from the wine pigments reportedabove, the possible presence of malvidin-3-monoglucoside hydroxy, vinylalcohol, 4-vinylcatechol and vinylsyringol adducts weredemonstrated. A manuscript outlining thisstudy is well progressed.

Characterization of highly polymerised seedtannin by electrospray mass spectrometrywas conducted under the projects of AWR96/1 and CRCV 1.2 in collaboration with DrStephane Vidal. The characterization ofhighly polymerised tannin (proanthocyanidins)is a challenging task and has been attemptedusing chromatographic and analyticaltechniques including mass spectrometry. Todate, the sizes of proanthocyanidinsdetermined by mass spectrometry werepentamer in grape seeds by liquid secondaryion mass spectrometry (Vivas 1996),heptadecamer in apple (Guyot, 1997) anddocosamer in litchi (Roux, 1998) byelectrospray mass spectromery and undecamerin grape seed (Kruger, 2000) by matrix assistedlaser desorption ionisation mass spectrometry.In our studies, grape seed tannins werefractionated into two fractions by columnchromatography according to their molecularsizes. The two fractions, LDPS (lower degreeof polymerisation) and HDPS (higher degreeof polymerisation), were analysed byelectrospray mass spectrometry in the negativemode. The LDPS and HDPS fractions werewell characterised and the presence of morethan pentacosamer of procyanidin gallates inthe HDPS fraction was demonstrated. Theseanalytical techniques are sophisticated butmuch needed to understand better thetannin properties of wine. A manuscriptoutlining this study is in progress.

The considerable contribution of the Facilityto the CRCV tannin projects (Program 1.2)was made in collaboration with staff of TheUniversity of Adelaide. The followingoutcomes were achieved.- Asenstorfer, R.E.; Hayasaka, Y.; Iland, P.G.;

Lambert, S.G.; Jones, G.P. Wine phenolics:the development of pigments in red wine.Steans, G., ed. Proceedings of the 1999conference of the New Zealand Society forViticulture and Oenology; 4-5 November1999; Auckland, New Zealand Society forViticulture and Oenology; 1999: 83-78.

- Asenstorfer, R.E.; Hayasaka, Y.; Jones, G.P.Isolation and structures of oligomeric winepigments by bisulphite-mediate ionexchange chromatography. J. Agric FoodChem. Submitted.

- Kennedy, J.A; Hayasaka, Y.; Vidal, S.; Waters,E.J.; Jones, P.J. Composition of grape skinproanthocyanidins at different stages of berrydevelopment. Manuscript in preparation.

Team Reports

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Staff/students of AWRI 90% 82%

Adelaide University 10% 15%

Other Institutes 0% 3%

GC- LC-MS/MS MS/MS

Table 8. Time usage of the GC-MS/MS and LC-MS/MSfor 2000/2001

HPLC instumentation

Grape pressing contest: 1973Photograph courtesy of The State Library of South Australia

Quality Liaison Manager

Staff: Mark Gishen

The major output of the Institute’s activities inthe provision of advice on quality managementtechniques to industry remains the FromGrapes to Glass program jointly run with theSouth Australian Wine and Brandy IndustryAssociation. The program was published inAugust 1997 and enhanced with a simpleHACCP module in 1999. Industry interestremains greatest in the HACCP module — asimple program delivered in a one-day coursethat incorporates an HACCP-type (hazardanalysis and critical control point) food safetyplan. This module was designed to satisfythe requirements of the proposed (nowdelayed) changes to the food hygieneregulations, and meet the needs of thesmaller scale businesses in the industry.Three courses for the HACCP module wereconducted throughout the year, adding 20more to the growing list of companieshaving attended (now totalling 73). TheFrom Grapes to Glass program provides asimple and relatively cheap program thatuses a staged approach in the attainment ofinternationally recognised standards, startingfrom the Codex HACCP principles andleading to the full ISO 9000 qualitymanagement standard. The program maysoon be upgraded and simplified to reflectthe changes brought in with the new year-2000 version of the ISO 9000 standard.

The Code of good manufacturing practicethat was developed during the developmentof the HACCP module of the From Grapesto Glass program, was also published on the

Institute's new web site to enhance itscirculation throughout the industry. Mark Gishen is assisting in the staging ofevents of interest to the wine industryincluding workshops on objective grapequality measurement, quality managementoptions, HACCP plan development, and NIRspectroscopy calibration for the 11th

Australian Wine Industry TechnicalConference (Adelaide, October 2001), and iscontributing as part of the speaker programfor the same conference.

Mark Gishen takes primary responsibility forthe internal quality management systems ofthe Analytical Service, overseeing managementreviews, documentation, auditing, andcorrective actions. An external reassessment ofthe laboraotry's quality system was conductedby NATA during this year. The method for thedetermination of ochratoxin A was successfulin gaining accreditation, and several changesto signatory status of staff were approved. Itwas pleasing to note that improvements tothe system that have been implementedrecently meant that the system already complieswith many of the elements of the revisedstandard, ISO 17025, ahead of therequirements of NATA. The Analytical Servicecontinues to participate and excel in bothnational and international proficiency testingprograms for routine wine analysis and foragrochemical residue testing. The AnalyticalService continues to manage its qualitymanagement system with the aid of theParadigm Quality software package and will beupgrading to a newer version release of thesoftware in the coming year. Following theupgrade, the software will be introducedthroughout the Institute, primarily as a means

of assisting control and availability of policyand procedural documentation, as part of ageneral move toward quality managementsystems implementation.

As reported elsewhere, Mark Gishen remainsheavily involved in the collaborative researchproject evaluating the use of near infraredspectroscopy (NIRS) for the rapid determinationof a number of compositional parameters ingrapes, must, wine and grape spirit, andrecently assumed primary responsibility forthe project. The details of this project arereported elsewhere in the Annual Report.

Publications and seminars given by theQuality Liaison Manager and staff workingon the NIR project are shown in Appendix 1.

Communications and publicity

Staff: Rae Blair

The Communication and Publicity Manager’srole is a part-time position for Rae Blair, whois also the Personal Assistant to the Directorand Conference Manager of the EleventhAustralian Wine Industry Technical Conference.As Communication and Publicity Manager,she is responsible for ensuring that industryand stakeholder groups clearly comprehendthe positioning (value) of the Institute. Thispositioning is developed in line with theInstitute’s mission statement and businessplan objectives. Part of her role is tocoordinate the Institute’s printed materialand other non-technical communications,and to act as a conduit for media. TheCommunication and Publicity Manager isalso responsible for the performance andoutput of the John Fornachon MemorialLibrary. The report of the activities of theLibrary can be found elsewhere in this Report.

Technical ReviewThe Communication and Publicity Managerhas overall fiscal responsibility for theproduction and supply of Technical Review,and the pricing structure of Technical Reviewwas assessed in line with the introduction ofGST as at 1 July 2000. During the reportingperiod the cost of subscribing to TechnicalReview was further considered and no pricerise was deemed necessary, although thecost structure of recouping postage costswas streamlined. The subscription periodwas also set at expiring on 30 June each year.The Communication and Publicity Managermet with the Director, Health and RegulatoryInformation Manager (as Editor of TechnicalReview) and the Librarian to discusschanging the format of part of the TechnicalReview to ensure its copyright-compliance.

Coordination of mediaInstitute staff were interviewed byrepresentatives from various forms of media

Team Reports

48

Gayle Baldock and Yoji Hayasaka

over the year as shown in Table 9.On Monday and Tuesday, 5 and 6 February,the Communication and Publicity Managercirculated a press release (prepared by theManager Industry Services and theCommunication and Publicity Manager) tocorrect some information relevant toNuKorc, to 62 wine industry organisationsand wine media Australia-wide, and to 358email addresses via the Institute's emailbulletin. The release was also published inthe February edition of Technical Review.

EditingThe Communication and Publicity Managerassisted the Analytical Service staff byproofing their 2000-2001 Analytical FeeSchedule and with the Director she alsocompiled and edited the 2000 AnnualReport. The Communication and PublicityManager coordinated the production ofAgrochemicals registered for use inAustralian viticulture 2001/2002.

Institute’s websiteMuch of the year was spent on theevaluation and implementation of the designof the new Institute website and preparationof information to be loaded onto thewebsite. It was decided that the Instituteneeded a separate domain from theAdelaide University’s server, and accordingly,the Communication and Publicity Managerarranged for the Institute to purchase serverspace from Internode, and has registered itsown domain name awri.com.au

Accordingly, the address for the new websiteis www.awri.com.au

The sitemap was designed using four simple,user-friendly boxes: ‘Research,’ ‘Services,’‘About Us’ and ‘Information.’ Each boxcontains various sub-sections and can beaccessed by rolling the cursor over the boxto reveal a slide-out menu of sub-sections.

The Director had strong input into the finalproduct, along with the Communication andPublicity Manager, the Manager – IndustryServices and his staff. The site was designedby Geoffrey Reed Communications, and wasfacilitated by WineBiz.Website maintenanceVarious staff within the Institute are chargedwith ensuring the website is kept up to date.The ‘Research’ section is maintained by LeighFrancis, Paul Henschke, Michael Esler, BobDambergs and Miguel de Barros Lopes. PeterGodden (and his team), Don Buick and SallyBell maintain the ‘Services’ section. Rae Blairmaintains the ‘About Us’ section, and CreinaStockley is responsible for the ‘Information’section. Dr Alan Pollnitz undertakes theactual updating of the www files in his roleas part-time Computer Systems Officer.

Eleventh Australian Wine IndustryTechnical ConferenceFull details of the Eleventh Australian WineIndustry Technical Conference will be providedin the 2002 Annual Report. However,during the year, Rae Blair, in her role asConference Manager and Treasurer ofAustralian Wine Industry Technical ConferenceInc.; spent much time liaising withinternational speakers, venue operators,sponsors, graphic designers and exhibitionmanagement. She is ably assisted by theSecretariat of the Conference, Carolyn Grant,

who is responsible for the registrationprocess, and who had processed a recordnumber of ‘early registrations’ to the eventby 8 June 2001.

The Institute’s Director, Peter Høj is Chair ofthe Conference Planning Committee; PeterGodden, Trudy Goode and Ella Robinson arecoordinating 74 workshops and Jeremy Hackis coordinating the poster display. Excellentbackup is provided by our Accountant andCompany Secretary.

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3/8/00 PWG Brettanomyces Tim White/AustralianFinancial Review

13/10/00 PWG Addition of tannin to wine Dan Sogg/Wine Spectator

Several media interviews were conducted by Peter Høj and Mark Gishen as a result of astand-alone press release issued by BRL Hardy regarding the Near Infrared Spectroscopy project

14/10/00 PBH Research in the Australian Wine Industry Sky TV (UK)

8/8/00 MG Grape quality/NIR ABC Regional SA(Country Hour)

9/8/00 MG Grape quality/NIR ABC Tropical North (Drive)

11/10/00 SJB Chemical control of locusts ABC 5CK the North & West Report

27/10/00 ADC ‘Sour sob’ character in Sauvignon Blanc wine Philip White/The Advertiser

6/11/00 ILF AWRI Stefan Maus/Wein-Wissenschaft

8/11/00 ILF Flavour chemistry Paul Clancy/AustralianViticulture and AustralianNew Zealand WineIndustry Journal

10/11/00 PBH Silver nitrate in wine Jeni Port/The Age

12/12/00 PBH Pesticide spray and residues Philip Satchel/ABC 5AN

22/2/01 CSS Blood products in wine Natalie Larkin/ABC Radio National

22/2/01 CSS Blood products in wine ABC TV News

18/1/01 PWG Closure trial and AWRI Mark Stean/The Advertiser

27/1/01 SJB Conventional versus organic viticulture Nick Galvin/ABC’s Organic Gardener

1/2/01 PBH Impact odorants in wine Max Allen/The Australian

15/2/01 PWG Closure trial Eric Cummins/National Grapegrower

2/3/01 PWG Closure trial Laura Lee Madonna/www.WorldWineTrade.com

9/3/01 ILF Mouthfeel wheel Mark Steene/The Advertiser

12/3/01 PBH/MGNear Infrared Spectroscopy project Natasha Mitchell/ABC

16/3/01 ILF Mouthfeel wheel Cindie Lange/National GrapeGrowers

20/3/01 PBH Genetically modified organisms in vines/yeasts. Florence Kennell/La Revuedu Vine de France andwww.magnumvinum.fr

16/4/01 PBH AWRI Channel 9 Directions

16/4/01 PWG Closure trial Channel 9 Directions

31/5/01 PBH Chemicals used in vineyards Ken Dickin 5DN

31/5/01 PWG Closure trial Rose Murray Brown

Date Staff Discussed Media

Table 9. Staff interviews by media 2000/2001

11AWITC websiteDuring the year, the Conference Managerpurchased server space and coordinated theproduction and uploading of information forthe website of the Australian Wine IndustryTechnical Conference. The website addressis www.awitc.com.au.

26th World Congress and 81st GeneralAssembly of the Office International dela Vigne et du VinThe Communication and Publicity Manageralso manages the Secretariat of the OIVCongress, Marcia Burnett. The InstituteDirector and the Communication andPublicity Manager are both members of theAustralian OIV Planning Committee. TheCommunication and Publicity Manager securedaccommodation for delegates; produced thelogo, letterhead, first announcement andposter announcing the event in Australia,the preliminary programme (in threelanguages) and established a skeletonbudget for the event. Further details of theInstitute’s contribution to the OIV Congressand General Assembly held in Australia willbe shown in the 2002 Annual Report.

Analytical Service

Staff: Don Buick, John Hughes, MatthewHoldstock, Greg Ruediger, GayleBaldock, Amanda Cook, MatthewCream, Jeremy Hack, Radka Kalouch,Andrea Kemp, Adam Loveys, KevinPardon, Randall Taylor, Sandra Lloyd-Davies, Bao Tran and David Boehm

The Analytical Service is a commercial facilityoperating independently on a fully-costedbasis within the Institute with the Manager,Mr Don Buick, reporting to the Director. It isfunded by the income generated from feeearning work and contributes significantly tothe financial well being of the Institute. TheAnalytical Service provides services to theAustralian wine industry including use of thepractical outcomes of the Institute’spublished research. Due to the nature ofsome of its work, the Analytical Service,under appropriate cross-chargingarrangements, makes use of the expertiseexisting within the Industry Service team andwithin the Research groups to interpret oradvise on results or project design.

Don Buick and John Hughes have visitedclients in the McLaren Vale region and in theMargaret River region during the year as wellas attending Wine Australia in Melbourne inNovember and the Margaret River Field Dayin May. These visits plus direct mail and theplacement of advertisements in industryjournals and publications have assisted in thedevelopment of improved client relationshipsand the generation of several new clients.As a consequence, a significant increase in

workload particularly from the McLaren Valeregion has occurred during the 2001 vintageperiod. This is evident from the increasednumbers of tests for malic acid performedduring the year. The personal contact withwinemakers, technical staff and export staffhas been successful and will be continued inthe new year. For the first time, theAnalytical Service sponsored a trophy for ‘thebest tawny port’ at the Adelaide Wine showin September 2000.

In October 2000, a new fee scheduleincorporating details and pricing for theservices offered was mailed to all clients.The terms and conditions of our serviceshave been brought to the attention of clientsthrough the fee schedule, the web site andin formal quotations. Formal quotations areissued for larger or non-routine jobsincluding clear identification of the servicesto be provided. Most incoming work of aroutine nature has client requirementsidentified by letter or sample labels.

The services provided include analysesconsistent with the requirements of exportcertification and support for the quality controlactivities of winemaking and viticulture oftenrequiring more specialised skills than thosecommonly found in production laboratories.The Analytical Service conducted over46,000 individual analyses on wine andgrapes in 2000/2001 and covering simpleanalyses such as malic acid to more complexanalyses such as the multi-residue screen foragricultural chemicals. The aim is to providea range of quality analytical services that areimportant to industry and which can beprovided in a reliable, price competitive,accurate and timely manner.

The number of tests performed by theAnalytical Service increased by 3% from theprevious year with significant increases in someareas of testing being offset by a lowernumber of tests performed as part of theInstitute led closure trial during the year.Significant increases occurred in AWBCcertificates, EU certificates, malic acid, sensoryassessments, multi-residue assays, colour inred grapes, glycosyl-glucose in grapes and 4-ethyl phenol analyses. The number of datapoints produced increased by 7% providinga better measure of the increase in workload.

Analytical methods for organic acids,ascorbic acid, and methanol have been re-validated following the purchase of a newhigh pressure liquid chromatograph and therefurbishment of a gas chromatograph.

The sensory assessment service has been asignificant growth area with large numbersof samples assessed for taints, faults orgeneral quality parameters. New softwarepurchased by the Institute has streamlinedthe recording of comments, the scoring andthe reporting of sensory work. Much of thesensory work has been conducted inconjunction with chemical testing and usedfor dispute resolution or insurance purposes.

Staff have worked closely with the IndustryServices’ team to analyse the 18 month and24 month samples from the closure trial (seereport under Industry Services activities). Theanalysis of large numbers of free and totalsulphur dioxide, dissolved oxygen andOD420 were performed with staff workingin shifts to complete the work within two tothree days of commencement.

John Hughes successfully completed theAdvanced Wine Assessment Course conductedin July 2000 and has contributed to theplanning of the July 2001 course to beconducted by Industry Services. OtherAnalytical Service staff contributed to the courseincluding pouring of wine, cleaning up andrecording of the trainee’s results for evaluation.

The Analytical Service was re-assessed byNATA in September of 2000 and complimentedon the quality of its documentation andprocedures. The method for ochratoxin Awas approved and the inclusion of malathionand fenitrothion in the multiresidue assay isunder review. Don Buick and Randell Taylorwere approved as NATA signatories. NATAaccreditation previously required compliancewith ISO Guide 25. This internationalstandard has been replaced by ISO17025with laboratories required to meet the newstandard by 2002. Compliance withISO17025 ensures the laboratory operates inaccordance with ISO9000. The assessmentprovided a check against the new standardand confirmed good progress towardscompliance with the new standard.

Trace Analysis LaboratoryThe Trace Analysis Laboratory, operatingwithin the Analytical Service, performs morespecialised testing including a multi-residuescreen using gas chromatography/massspectrometry including 30 residues, coveringover 50 brand name agrochemicals. Severalpesticides given special approval for use incontrolling plague locusts have beenincluded in the multi-residue screenincluding chlorpyrifos, carbaryl, dimethoate,malathion and fenitrothion. A multi-residuescreen for several compounds unable to be

Team Reports

50

97/98 98/99 99/00 00/01

Total number of tests 35852 39087 44846 46037

Table 10: Comparison of selected tests performed by the Analytical service by period.

analysed by gas chromatography has beendeveloped using high pressure liquidchromatography and includes cyprodinil,fludioxinil, pyrimethanil, fenhexamid andtebufenozide. Other methods have beendeveloped and validated for spiroxamine,trifloxystrobin, axoystrobin and indoxacarb.

Several extended residue trials have beenperformed for chemical companies duringthe year to establish the levels of residuepresent or absent in grapes or winefollowing the application to vines of newlydeveloped agrochemicals. These trialsinvolve testing of grapes, small scalewinemaking and analysis followingdevelopment and validation of methods ofanalysis. The data produced is used by thechemical companies for further developmentof agrochemicals and for application to theNational Registration Authority forregistration approval. From 2003 this workwill require Good Laboratory Practice (GLP)accreditation from NATA. The Trace AnalysisLaboratory has begun to prepare foraccreditation and Greg Ruediger is to attenda training course on GLP in the near future.

Large numbers of samples have been analysedduring the year for chloroanisoles, the principlecontributors to cork taint in wine and for 4-ethylphenol which is an indicator ofBrettanoyces/Dekkera yeast spoilage activity.4-ethylphenol is included in the oak flavourcompound assay but is now offered separatelywhen only 4-ethylphenol is required.Eugenol is now included in the oak flavourcompounds assay. The relatively highconcentrations of 4-ethylphenol in many

samples indicates that industry practitionersseriously should consider performing thisrelatively cheap test on a more routine basis.

The validation of a method for thedetermination of histamine in wine usinghigh pressure liquid chromatography hascommenced following limited success usingan ELISA kit method for this purpose. Themeasurement of histamine is necessary tomeet the requirements for importation intoSwitzerland and will allow a survey of Australianwine to be performed in the new year.Little change in staffing occurred during theyear. Adam Loveys formerly employed as acasual Laboratory Technician graduated fromthe University of Adelaide with an HonoursDegree in Agricultural Science andsubsequently resigned to take up aviticultural position on Kangaroo Island.Adam has worked at the Institute for severalyears and made a valuable contribution tothe Analytical Service whilst continuing hisstudies. John Hughes took long serviceleave during the year including travelling inthe winemaking regions of Germany andFrance. Matt Holdstock, Analytical ServiceSupervisor, has commenced part-time studiesto complete the Graduate Diploma inOenology. This will mean that the AnalyticalService will have two degree qualifiedoenologists able to service industry’s needs.Several other staff have attended specialisedinstrument training courses to ensure newequipment is well maintained and used to itsmaximum capability. The Analytical Serviceaims to support the continual upgrading ofstaff skills as an important element inexpanding our portfolio of services.

51

Greg Ruediger, Don Buick and John Hughes

Statement of Financial Performance

52

For the year ended 30 June 20012001 2000

$ $Revenue from operating activities

Grape and Wine Research and Development Corporation

Project funds 3,569,186 3,191,013Capital grants

Building 0 250,000Equipment 426,000 722,514

Other project funds 545,348 409,742

Analytical Service 1,167,825 1,130,203

Sundry income 193,853 161,180

Expenses from operating activities

Employee benefit expense 3,437,532 2,961,477

Consumables used 514,417 418,478

Depreciation and amortisation expense 551,179 338,130

Repairs and maintenance 99,282 72,429

Travel expenses 96,512 98,800

Borrowing cost expense 12,406 24,447

Other expenses from ordinary activities 959,196 1,092,163

Profit from operating activities 231,688 883,175

Net gain (loss) on disposal of assetsMotor vehicles 14,935 0Equipment 397 (495)

Profit from ordinary activities 247,020 882,680

Write back of asset revaluation reserve onreversion to measuring buildings at cost (603,067) 0

Transfer of accumulated amortisation on buildings 125,536 0

Total changes in equity (230,511) 882,680

Hans Muhlack

Statement of Financial Position

53

As at 30 June 20012001 2000

$ $Current assets

Cash assets 920,798 120,020

Receivables 307,584 346,413

Commercial bills 670,000 648,000

Other current assets 52,326 25,621

Total current assets 1,950,708 1,140,054

Non current assets

Leasehold buildings 1,596,247 2,102,021

Plant and equipment 1,918,127 1,960,983

Australian Wine Industry Chair of Oenology 840,000 840,000

Total non current assets 4,354,374 4,903,004

Total Assets 6,305,082 6,043,058

Current liabilities

Payables and other accruals 924,288 436,328

Project funds not expended

GWRDC 37,150 51,877CRCV 13,936 0

Interest bearing liabilities 120,000 170,000

Provisions 391,490 362,822

Total current liabilities 1,486,864 1,021,027

Non current liabilities

Provisions 90,970 64,272

Total non current liabilities 90,970 64,272

Total Liabilities 1,577,834 1,085,299

Net Assets 4,727,248 4,957,759

Equity

Reserves 966,750 1,569,817

Retained profits 3,760,498 3,387,942

Total Equity 4,727,248 4,957,759

Statement of Cash Flows

54

For the year ended 30 June 20012001 2000

$ $

Cash flows from operating activities

Grants and other income 5,874,426 5,929,599

Interest received 68,845 67,825

Payments to suppliers and employees (4,605,745) (4,701,320)

Net cash provided by operating activities 1,337,526 1,296,104

Cash flows from investing activities

Payment for commercial bills (22,000) (27,000)

Payments for building improvements (8,858) (684,620)

Payments for plant and equipment (507,390) (911,819)

Proceeds from sale of plant and equipment 51,500 49,283

Net cash used in investing activities (486748) (1,574,156)

Cash flows from financing activities

Repayment of loans (50,000) 0

Proceeds from loans 0 170,000

Net cash provided by financing activities (50,000) 170,000

Net increase (decrease) in cash held 800,778 (108,052)

Cash at 1 July 120,020 228,072

Cash at 30 June 920,798 120,020

Reconciliation of net cash provided byordinary activities with ordinary profit

Profit from ordinary activities 247,020 882,680

Non cash flows in operating profit

Amortisation and depreciation 551,179 338,130(Profit) loss on the sale of plant and equipment (15,332) 495Charges to (reduction in) provisions 55,366 45,351

Changes in assets and liabilities

(Increase) decrease in inventories (29,332) (12,000)(Increase) decrease in receivables and prepayments 41,456 (68,043)Increase (decrease) in sundry creditors and accruals 487,169 109,491

Net cash provided by ordinary activities 1,337,526 1,296,104

External seminars and talks

K.F. Pocock, Effects of winemaking and Lincoln University Grape and Wine 24 July 2000D. Tattersall2, viticultural practices on protein School, Christchurch, New ZealandY. Hayasaka, instability in wineI. Dupin1,2, B.McKinnon1,2,R. van Heeswijck2,P. Høj, E.J. Waters

Z.K. Peng, P.J. Iland2, The influence of winemaking Lincoln University Grape and Wine 24 July 2000R. Ristic2, R. Gawel2,5, and viticultural practices on School, Christchurch, New ZealandA. Oberholster1,2, colour and tannins in red wineI.L. Francis, E.J. Waters

P.W. Godden Bunch rots: understanding the ASVO Managing Bunch Rots seminar Mildura Vic 28 July 2000winemaker's dilemma

P.W. Godden Persistent wine instability issues, New Horizons for Wine Quality, Victorian Wine 10 August 2000and, Towards better barrel Industry Association’s Seminar. Melbourne Vicmanagement – a review ofthe AWRI oak research

P.B. Høj What happened in the wine Committee for Economic Development of Australia’s 6 September 2000industry in the last 10 years that seminar ‘Building of the knowledge economy’has transformed it from a shadow Hyatt Regency Adelaideof Europe to the vibrant player inthe global area it is today?

G.M. Elsey C13-Norisoprenoids: their generation Flinders University weekly seminar 8 September 2000and contribution to aroma program, Flinders University, SA

P.A. Henschke Fermentation management and E. & J. Gallo Winery, Sonoma, 10 September 2000importance of yeast nutrition California, USA

P.B. Høj Genetic engineering, the First National Environmental Conference Adelaide SA 1 November 2000environment and the wine industry - a personal perspective

P.W. Godden, Roadshow seminars McLaren Vale, SA 8 November 2000P.J. Costello, S.J.Bell, E.J. Bartowsky,A.D. Coulter

P.B. Høj Current trends in Australian South African Society for Enology and Viticulture’s 8-10 November 2000oenological research: the quest 2nd International Viticulture and Enology Congress,to produce grapes and wine to Cape Town South Africaa predetermined specification

P.W. Godden, Roadshow seminars Adelaide Hills, SA 9 November 2000S.J. Bell, M. Gishen,I.L. Francis, G.A.Ruediger, A.D. Coulter

P.B. Høj, P.W. Roadshow seminars Barossa Valley, SA 13 November 2000Godden, S.J. Bell,M.A. Sefton,E.J. Bartowsky, P.A.Henschke, I.L. Francis,M. Gishen

Appendix

55

Author Title Organization/Location Date

Appendix 1. External seminars, talks and poster papers presented by institute staff during 2000/01

Appendix

56

P.B. Høj, P.W. Roadshow seminars Clare Valley SA 16 November 2000Godden, S.J. Bell,M.A. Sefton, E.J.Bartowsky, P.A.Henschke, I.L. Francis,M. Gishen

P.B. Høj An update of the use of Near Grape and Wine Industry Outreach 17 November 2000Infrared Spectroscopy for Seminar, Adelaide University, Plantmeasurement in grape quality Research Centre, Urrbrae SAparameters

P.J. Costello, Towards understanding the Grape and Wine Industry Outreach 17 November 2000P.A. Henschke, interactions between wine yeast Seminar, Adelaide University, PlantA.J. Markides and lactic acid bacteria Research Centre, Urrbrae SA

P.A. Henschke Wine microbiology and Lallemand meeting, AWRI Urrbrae, SA 20 November 2000biotechnology projects overview

J.M. Eglinton The search for alternative wine Lallemand meeting, AWRI Urrbrae, SA 20 November 2000yeasts. Flavour and aroma diversityusing Saccharomyces bayanus

E.J. Bartowsky Microbial spoilage of bottled red Lallemand meeting, AWRI Urrbrae, SA 20 November 2000wine by acetic acid bacteria, and,Diacetyl ‘buttery’ attribute ofwine and MLF?

P.J. Costello Mousy off-flavour spoilage of Lallemand meeting, AWRI Urrbrae, SA 20 November 2000wine, and, Interaction of wineyeast and lactic acid bacteria

M. Esler New technologies for rapid Interwinery Analysis Group meeting, 24 November 2000measurement (NIRS and FT-IR) Renmark, SA

P.B. Høj, P.W. Roadshow seminars Berri, SA 4 December 2000Godden, S.J. Bell,P.A. Henschke, I.L.Francis, M. Gishen

M.A. Sefton, M. Precursors to oak lactone: Annual Adelaide Organic Chemistry 4 December 2000Raunkjaer, G.K. synthesis of gallate ester Symposium, Flinders University, SASkouroumounis, derivates of 3-methyl-4-G.M. Elsey hydroxyoctanoic acid

P.W. Godden, S.J. Roadshow seminars Mildura, Vic 6 December 2000Bell, P.A. Henschke,M. Gishen, A.D. Coulter

I.L. Francis Tannins and mouthfeel Southcorp Winemakers’ Forum Mildura, Vic 7 December 2000

P.B. Høj, P.W. Roadshow seminars Geelong, Vic 15 December 2000Godden, A.D. Coulter

P.A. Henschke Brettanomyces - winemaking Southern Pinot Noir Workshop, 18 January 2001significance and prevention Methven, Canterbury, New Zealand

A.P. Pollnitz, The analysis of wine volatiles Australian and New Zealand Society 5 February 2001G.K. Skouroumounis, derived from oak for Mass Spectrometry, 18th Conference,K.H. Pardon, D. Capone, Coolangatta, QldM.A. Sefton

Author Title Organization/Location Date

Appendix 1. External seminars, talks and poster papers presented by institute staff during 2000/01

57

Y. Hayasaka, K.S. Use of electrospray mass Australian and New Zealand Society for Mass 6 February 2001Adams2, K.F. spectrometry for mass Spectrometry, 18th Conference, Coolangatta, QldPocock, G.A. determination of grape (VitisBaldock, E.J. Waters, vinifera) juice pathogenesisP.B. Høj related proteins: potential tool

for varietal differentiation

E.J. Bartowsky Wine microbiology Non-clinical meeting of the Australian 28 March 2001Society for Microbiology, Plant Research Centre, Urrbrae, SA

I.L. Francis Relating grape composition and Grape Expectations: University of 11 April 2001wine quality, and, Key grape- California, Davis, California, USAderived wine flavour compounds: can we define flavour specifications?

C.S. Stockley A role for wine in the diet Department of Clinical Dietetics’ continuing education 11 April 2001program, The Royal Adelaide Hospital

I.L. Francis Near infrared for rapid grape analysis Grape Expectations: University of California, 12 April 2001Davis, California, USA

E.J. Waters Program 1: Vineyard management Cooperative Research Centre Board, 9 May 2001to meet grape quality specifications Plant Research Centre, Urrbrae, SA

M.J. Herderich Winegrape tannin and CRCV Second Year Review, 22 May 2001colour specification Plant Research Centre, Urrbrae, SA

P.B. Høj Advice to young scientists, a Adelaide University Science Scholars 29 May 2001personal perspective

I.L. Francis Wine flavour and sensory analysis Australian Institute of Food Science and Technology 30 May 2001(Victoria) Sensory Evaluation Group, Werribee, Vic

P.B. Høj Use of research outcomes for BRL Hardy Vintage Conference, 4 June 2001the management of problems Lake Crackenback, NSWand opportunities

E.J. Waters, K.F. Pocock, The unstable proteins of wine: Unité Mixte de Recherche Sciences pour l'Oenologie, 13 June 2001D. Tattersall2, Y. Hayasaka, a summary of their properties INRA, Montpellier, FranceI. Dupin1,2, B. McKinnon1,2, and of factors influencing theR. van Heeswijck2, P.B. Høj1,2 levels of these proteins in wine

P.B. Høj A general introduction to The Unité Mixte de Recherche Sciences pour l'Oenologie, 13 June 2001Australian Wine Research INRA, Montpellier, FranceInstitute and an outline ofselected research activities

S. Vidal Sensory features of grape tannins In Vino Analytica Scientia, Bordeaux, France 15 June 2001

E.J. Waters, K.F. Pocock, The unstable proteins of wine: In Vino Analytica Scientia, Bordeaux, France 15 June 2001D. Tattersall2, Y. Hayasaka, a summary of their propertiesI. Dupin1,2, B. McKinnon1,2, and of factors influencing theR. van Heeswijck2, P.B. Høj1,2 levels of these proteins in wine

M.J. Herderich Red wine colour and mouthfeel: American Society of Enology and Viticulture 29 June 2001analysis of phenolic pigments Annual General Meeting, San Diego, USAand tannins

Author Title Organization/Location Date

Appendix 1. External seminars, talks and poster papers presented by institute staff during 2000/01

Appendix

58

Posters

M.B. Esler, M. Effects of variety, region and 10th International Conference on Near Infrared 10-15 June 2000Gishen, I.L. Francis, season on near infrared Spectroscopy, Kyongju, South KoreaR.G. Dambergs, A. reflectance spectroscopicKambouris, W.U. analysis of quality parametersCynkar, D.R. Boehm in red wine grapes

R.G. Dambergs, Wine quality grading by near 10th International Conference on Near Infrared 10-15 June 2000A. Kambouris, N. infrared spectroscopy Spectroscopy, Kyongju, South KoreaSchumacher, I.L. Francis,M.B. Esler, M. Gishen

Workshops

S-J. Bell Spray application in viticulture Research to Practice™, McLaren Vale, SA 15 August 2000

S-J. Bell Spray application in viticulture Research to Practice™, McLaren Flat, SA 17 August 2000

S-J. Bell Grapevine nutrition Research to Practice™, Langhorne Creek, SA 26 September 2000

S-J. Bell Integrated Pest Management Research to Practice™, McLaren Vale, SA 7 November 2000

P.W. Godden, A.D. Identification and management McLaren Vale, SA 8 November 2000Coulter, P. Valente, of wine instabilitiesE.M. Robinson, M. Gishen

S-J. Bell Grapevine Nutrition Research to Practice™, Barossa Valley, SA 11 November 2000

S-J. Bell Grapevine Nutrition Research to Practice™, McLaren Vale, SA 13 November 2000

P.W. Godden, A.D. Identification and management Barossa Valley, SA 14 November 2000Coulter, P. Valente, of wine instabilitiesE.M. Robinson

P.W. Godden, A.D. Identification and management Clare Valley, SA 17 November 2000Coulter, P. Valente, of wine instabilitiesE.M. Robinson

P.W. Godden, A.D. Identification and management Berri, SA 5 December 2000Coulter, P. Valente, of wine instabilitiesE.M. Robinson

P.W. Godden, A.D. Identification and management Mildura, Vic 7 December 2000Coulter, P. Valente, of wine instabilitiesE.M. Robinson

Advanced Wine Assessment Course

P.W. Godden, E. Robinson Adelaide SA 18-21 July 2000J.B. Hughes, M. Gishen,A. Loveys, A.D. CoulterI.L. Francis

Author Title Organization/Location Date

Appendix 1. External seminars, talks and poster papers presented by institute staff during 2000/01

59

Subject Number of lectures Institute staff

Appendix 2. Teaching responsibilities of Institute staff during 2000/01

2000—Semester 2

Adelaide University

9685 Advances in Oenology 2 Dr M.A. Sefton

9685 Advances in Oenology 2 Dr I.L. Francis

9685 Advances in Oenology/

9086 Advances in Oenology (supplemented) 3 Dr P.A. Henschke

9685 Advances in Oenology/

9086 Advances in Oenology (supplemented) 2 Dr E.J. Bartowsky

9685 Advances in Oenology/

9086 Advances in Oenology (supplemented) 1 Dr P.J. Costello

9685 Advances in Oenology 1 P.W. Godden

1958 Wine packaging and quality management 2 C.S. Stockley

1958 Wine packaging and quality management 1 M. Gishen

The Flinders University of South Australia

MMED 3921 Industrial and Pharmaceutical Microbiology 1 Dr P.A. Henschke

2001—Semester 1

Adelaide University

2580 Stabilisation and Clarification 3 Dr E.J. Waters

2580 Stabilisation and Clarification 1 P.W. Godden

1005/3113 Winemaking 2 Dr P.A. Henschke

1958 Wine Packaging and Quality Management 1 M. Gishen

2213 Grape Industry Practice Policy and Communication ~ 40 hours Professor P.B. Høj and(subject coordination and delivery for five weeks) C.S. Stockley

5693 Wine and Marketing in Society 6 C.S. Stockley

Honours

C. Day P.B. Høj, S.J. Clarke2 Adelaide University

A. Matthews E.J. Bartowsky, I.L. Francis Adelaide University

M. Mercurio G.K. Skouroumounis, G.M. Elsey Flinders University

M.A. Smith G.M. Elsey Flinders University

A. Walkenhorst I.L. Francis, R.G. Dambergs Adelaide University

Masters

D. Coates E.J. Bartowsky, R. Gawel2,5 Adelaide University

PhD

S.L. Brown M.A. de Barros Lopes, E.J. Waters, P.B. Høj GWRDC

F. Carrau P.A. Henschke, E. Dellacassa8 University of the Republic of Uruguay staff member

N. D’Incecco E.J. Bartowsky, P.A. Henschke Italian PhD Scholarship

J.M. Eglinton P.A. Henschke, P.R. Langridge6 Institute staff

R. Gawel I.L. Francis, A.J. Markides2,4 DHVO staff

W. Greenrod M. Fenech9, M.Abbey9, M.A. Sefton, P. Burcham10 GWRDC

S. Hoffmann P. Winterhalter, C.S. Stockley University of Braunschweig

K.S. Howell P.A. Henschke, E.J. Bartowsky, G.H. Fleet3 APRA scholarship

A. Janusz G.M. Elsey, M.A. Sefton, M. Perkins7 CRCV

D. Lee G.P. Jones2, E.J. Waters CRCV

A. Oberholster E.J. Waters, I.L. Francis, G.P. Jones2, P.G. Iland2 GWRDC

C.J. Puglisi G.M. Elsey, M.A. Sefton, R. Prager7 CRCV

R. Ristic P.J. Iland, I.L. Francis Adelaide University

P. Sivilotti S.J. Bell Italian Government and University of Udine

H. Smyth (nee Pain) I.L. Francis, M.A. Sefton GWRDC

K.L. Wilkinson M.A. Sefton, R. Prager7, G.M. Elsey GWRDC

Appendix

60

Student Supervisor/s Source of funds

Appendix 3. Graduate and Honours student supervision responsibilities of Institute staff for 2000/01

61

Robert Asenstorfer PhD Isolation, structures and properties of Dr G.P. Jones2, Dr P.J. Iland2, Dr E.J. Watersanthocyanins and oligmeric pigments in wine

Patrick Iland PhD A study of glycosides in grapes and wines Dr B. Coombe2, Dr P.J. Williams, Professor P.B. Høj

Patrick Jones PhD Glucosyl transferases involved in the biosynthesis Professor P.B. Høj, Dr B.L. Møller11

of cyanogenic glucosides in sorghum bicolor

Alan Pollnitz PhD The analysis of volatile wine components derived Dr M.A. Sefton, Dr G.P. Jones2

from oak products during winemaking and storage

Christopher Smyl PhD L-Proline: a potential nitrogen source for yeast Dr P.A. Henschke, Professor P. Langridge6,Saccharomyces cerevisiae under anaerobic conditions Dr M.A. de Barros Lopes

Vanessa Stockdale PhD Biochemical analysis of mannoproteins associated Dr E.J. Waters, Dr P.J. Williams,with haze protection in white wine Dr G.B. Fincher6

Nicholas Yap PhD The sensitivity of yeasts to killer yeast toxin, Dr P.A. Henschke, Dr M.A. de Barros Lopes,with focus on the killer yeast Pichia membranifaciens Professor P. Langridge6

Julie Campbell Honours The influence of chip size, heating processes Dr A.P. Pollnitz, Dr M.A. Seftonand other factors on the impact of oak chipson wine composition

David Dowden Honours Effect of polysaccharides on Grenache wine instability Dr G.P. Jones2, Dr E.J. Waters

Freya Hohnen Honours Changes in seed phenolic composition during berry Dr P.J. Iland2, Dr Z.K. Pengmaturation and throughout fermentation/extendedmaceration of Vitis vinifera cv. Cabernet Sauvignon

Helen McCarthy Honours Evaluation of the ability of the chemical sensory Dr E.J. Bartowsky, Dr I.L. Francisnose to determine the aroma profiles of wineswhich have undergone malolactic fermentation

Marie Pearce Honours Optimising diacetyl levels in wine Dr E.J. Bartowsky, Dr A.J. Markides2,4

Nina Viergutz Honours The interaction between wine yeast and Dr P.J. Costello, Dr A.J. Markides2,4

lactic acid bacteria

1 The Australian Wine Research Institute, 2 Department of Horticulture, Viticulture and Oenology, Adelaide University; 3 Department of Food Science, University of New South Wales; 4 now with Lallemand Australia; 5 now with Roseworthy Wine Tasting Programs Pty Ltd; 6 Department of Plant Science, Adelaide University; 7 Flinders University; 8 University of the Republic of Uruguay, 9 CSIRO Health Sciences and Nutrition, 10 Department of Clinical and Experimental Pharmacology, Adelaide University, 11 Royal Agricultural University of Copenhagen

Student PhD/Honours Title Supervisor/s

Theses completed

629 Carrau, F.M.; Henschke, P.A.; Medina, K.; Gioia, O.; Dellacassa, E. Assimilable nitrogen addition can cause stuck wine fermentations with mixed cultures of Saccharomyces cerevisiae killer and sensitive yeast. VII Congreso Latino Americano de viticultura y enología; 28 de Noviembre al 3 de Diciembre de 1999; Mendoza, Argentina: Instituto Nacional de Vitivinicultura; 2000: 149–154.

630 Skouroumounis, G.K.; Sefton, M.A. Acid-catalyzed hydrolysis of alcohols and their ß-D-glucopyranosides. J. Agric. Food Chem. 48: 2033–2039; 2000.

631 Waters, E.J.; Dupin, I.; Stockdale, V. A review of current knowledge on polysaccharides which ‘protect’ against protein haze in white wine. Aust. Grapegrower Winemaker (438a): 13–16; 2000.

632 Eglinton, J.; Fogarty, M.; McWilliam, S.; Francis, L.; Kwiatkowski, M.; Høj, P.; Henschke, P. Using Saccharomyces bayanus to modify the chemical and sensory profile of wine. Aust. Grapegrower Winemaker (438a): 28–32; 2000.

633 Todd, B.E.N.; Fleet, G.H.; Henschke, P.A. Promotion of autolysis through the interaction of killer and sensitive yeasts: potential application in sparkling wine production. Am. J. Enol. Vitic. 51: 65–72; 2000.

634 Stockley, C.S. Ochratoxin A–a metabolite on the agenda for the global wine industry. Aust. Grapegrower Winemaker (438a): 111–112; 2000.

635 Stockley, C.S. Advances in Australian research into the potential cardioprotective properties of wine. XXVth World Congress of the OIV;19–23 June 2000; Paris, France: Office International de la Vigne et du Vin; 2000: 33–41.

636 Gishen, M.; Holdstock, M. Preliminary evaluation of the performance of the Foss WineScan FT120 instrument for the simultaneousdetermination of several wine analyses. Aust. Grapegrower Winemaker (438a): 75–81; 2000.

637 Bartowsky, E.J.; Henschke, P.A. Management of malolactic fermentation for the ‘buttery’ diacetyl flavour in wine. Aust. Grapegrower Winemaker (438a): 58–67; 2000.

638 Pollnitz, A.P.; Pardon, K.H.; Sefton, M.A. 4-ethylphenol, 4-ethylguaiacol and oak lactones in Australian red wines. Aust. Grapegrower Winemaker (438): 45–52; 2000.

639 Dupin, I.V.S.; McKinnon, B.M.; Ryan, C.; Boulay, M.; Markides, A.J.; Jones, G.P.; Williams, P.J.; Waters, E.J. Saccharomyces cerevisiaemannoproteins that protect wine from protein haze: their release during fermentation and lees contact and a proposal for theirmechanism of action. J. Agric. Food Chem. 48: 3098–3105; 2000.

640 Haselgrove, L.; Botting, D.; van Heeswijck, R.; Høj, P.B.; Dry, P.R.; Ford, C.; Iland, P.G. Canopy microclimate and berry composition: the effect of bunch exposure on the phenolic composition of Vitis vinifera L cv. Shiraz grape berries. Aust. J. Grape Wine Res. 6: 141–149; 2000.

641 Stines, A.P.; Grubb, J.; Gockowiak, H.; Henschke, P.A.; Høj, P.B.; van Heeswijck, R. Proline and arginine accumulation in developing berries ofVitis vinifera L. in Australian vineyards: influence of vine cultivar, berry maturity and tissue type. Aust. J. Grape Wine Res. 6: 150–158; 2000.

642 Godden, P. The use of copper sulphate in winemaking. Aust. N.Z. Wine Ind. J. 15: 66–67; 2000.

643 Asenstorfer, R.E.; Hayasaka, Y.; Iland, P.G.; Lambert, S.G.; Jones, G.P. Wine phenolics: the development of pigments in red wine. Steans,G., ed. Proceedings of the 1999 conference of the New Zealand Society for Viticulture and Oenology; 4-5 November 1999; Auckland,New Zealand Society for Viticulture and Oenology; 1999: 83-78.

644 Yap, N.A.; de Barros Lopes, M.; Langridge, P.; Henschke, P.A. The incidence of killer activity of non-Saccharomyces yeasts towardsindigenous yeast species of grape must: potential application in wine fermentation. J. Appl. Microbiol. 89: 381–389; 2000.

645 Godden, P. Persistent wine instability issues. Aust. Grapegrower Winemaker (443): 10–14; 2000.

646 West, S.; Bell, S.-J.; Henry, K. Some facts about locust control in vineyards. Aust. Grapegrower Winemaker (443): 20–22; 2000.

647 Eglinton, J.M.; McWilliam, S.J.; Fogarty, M.W.; Francis, I.L.; Kwiatkowski, M.J.; Høj, P.B.; Henschke, P.A. The effect of Saccharomyces bayanusmediated fermentation on the chemical composition and aroma profile of Chardonnay wine. Aust. J. Grape Wine Res. 6: 190–196; 2000.

648 Gawel, R.; Oberholster, A.; Francis, I.L. A ‘Mouth-feel Wheel’: terminology for communicating the mouth-feel characteristics of red wine.Aust. J. Grape Wine Res. 6: 203–207; 2000.

649 de Barros Lopes, M.; Rehman, A.-U.; Gockowiak, H.; Heinrich, A.J.; Langridge, P.; Henschke, P.A. Fermentation properties of a wine yeastover-expressing the Saccharomyces cerevisiae glycerol 3-phosphate dehydrogenase gene (GPD2). Aust. J. Grape Wine Res. 6: 208–215; 2000.

650 Grbin, P.R.; Henschke, P.A. Mousy off-flavour production in grape juice and wine by Dekkera and Brettanomyces yeasts. Aust. J. Grape Wine Res. 6: 255–262; 2000.

651 Gishen, M.; Dambergs, R.G.; Kambouris, A.; Kwiatkowski, M.; Cynkar, W.U.; Høj, P.B.; Francis, I.L. Application of near infraredspectroscopy for quality assessment of grapes, wine and spirits. Davies, A.M.C.; Giangiacomo, R., eds. Near Infrared Spectroscopy:proceedings of the 9th international conference; Chichester, UK. NIR Publications; 2000: 917–920.

652 Ganter, P.F.; de Barros Lopes, M. The use of anonymous DNA markers in assessing worldwide relatedness in the yeast species Pichia kluyveri Bedford and Kudrjavzev. Can. J. Microbiol. 46: 967–980; 2000.

653 Stockley, C. A review of recent findings on the relationship between wine and health. Aust. N.Z. Wine Ind. J. 15: 36–43; 2000.

654 Godden, P. Bunch rots–understanding the winemaker’s dilemma. Davies, C.; Dundon, C.; Johnstone, R., eds. Managing bunch rots:proceedings of a seminar; 28 July 2000, Mildura Arts Centre, Mildura, Victoria. Adelaide, S.A.: Australian Society of Viticulture and Oenology; 2000: 52–54.

655 Jones, P.R.; Andersen, M.D.; Nielsen, J.S.; Høj, P.B.; Møller, B.L. The biosynthesis, degradation, transport and possible function ofcyanogenic glucosides. Romeo, J.T.; Ibrahim, R.; Varin, L.; de Luca, V., eds. Evolution of metabolic pathways. Oxford: Pergamon; 2000:191–247. (Recent advances in phytochemistry; 34).

Appendix

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Appendix 4. Institute staff publications 2000/01

656 Gawel, R.; Iland, P.G.; Francis, I.L. Characterizing the astringency of red wine: a case study. Food Qual. Pref. 12: 83–94; 2001.

657 Eglinton, J.; Henschke, P.A. The effect of a high concentration of acetic acid on the restarting of a stuck ferment. Aust. N.Z. Wine Ind. J. 16: 77–81; 2001.

658 Stockley, C.S. Evidence of the specific benefits of wine consumption on human health – an update on the cardiovascular and other physiological mechanisms. Aust. Grapegrower Winemaker (446): 72–77; 2001.

659 Stockley, C.S. Advances in Australian research into the potential cardioprotective properties of wine. Bull. OIV 73: 660–675; 2000.

660 Peng, Z.; Hayasaka, Y.; Iland, P.G.; Sefton, M.; Høj, P.; Waters, E.J. Quantitative analysis of polymeric procyanidins (tannins) from grape(Vitis vinifera) seeds by reverse phase high-performance liquid chromatography. J. Agric. Food Chem. 49: 26–31; 2001.

661 Stewart, R.J; Varghese, J.N; Garrett, T.P.J; Høj, P.B.; Fincher, G.B. Mutant barley (1,3;1,4)-ß-D-glucan endohydrolases with enhancedthermostability. Protein Engineering 14: 245-253; 2001.

662 Barker, D.A.; Capone, D.L.; Pollnitz, A.P.; McLean, H.J.; Francis, I.L.; Oakey, H.; Sefton, M.A. Absorption of 2,4,6-trichloroanisole by wine corks via the vapour phase in an enclosed environment. Aust. J. Grape Wine Res. 7:40-46; 2001.

663 Hayasaka, Y.; Adams, K.S.; Pocock, K.F.; Baldock, G.A.; Waters, E.J.; Høj, P.B. Use of electrospray mass spectrometry for massdetermination of grape (Vitis vinifera) juice pathogenesis-related proteins: a potential tool for varietal differentiation. J. Agric. Food Chem. 49:1830-1839; 2001.

664 Williams, P.J.; Francis, I.L. Wine flavor research-experience from the past offer a guide to the future. Rantz, J.M., ed. Proceedings of theASEV 50th Anniversary Annual Meeting; June 19-23, 2000; Davis, California. American Society for Enology and Viticulture; 2001: 191-195.

665 Høj, P.B.; Tattersall, D.B.; Adams, K.; Pocock, K.F.; Hayasaka, Y.; van Heeswijck, R.; Waters, E.J. The ‘haze proteins’ of wine – a summaryof properties, factors affecting their accumulation in grapes, and the amount of bentonite required for their removal from wine. Rantz,J.M., ed. Proceedings of the ASEV 50th Anniversary Annual Meeting; June 19-23, 2000; Davis, California. American Society for Enologyand Viticulture; 2001: 149-154.

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Appendix 5. Institute Committees

Staff member Management Research Industry Communication Analytical Quality Information Biosafety Occupational Super- Staff Advisory Steering Services Steering Service System Technology Health and annuation Code

Steering Steering Review Safety Negotiation

Peter Høj C C X X X X X

Eveline Bartowsky X X X X

Sally-Jean Bell X X X X

Rae Blair X C

Shauna Brown X

Don Buick X X X X

Dimitra Capone X

Adrian Coulter X X X X

Catherine Daniel X

Miguel de Barros Lopes X X X

Jeff Eglinton X C C

Leigh Francis X X X X

Mark Gishen X X X C C

Peter Godden X X C X X X

Jeremy Hack X

Yoji Hayasaka X X X

Paul Henschke X X X C

Markus Herderich X X

Matthew Holdstock X

John Hughes X X

Hans Muhlack X X X

Ingrid Oats X

Ken Pocock X C

Alan Pollnitz X

Greg Ruediger X

Mark Sefton X X X

Creina Stockley X X

Elizabeth Waters X X

C = denotes holder of Chair

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Acknowledgement

Compiled and edited by Professor Peter Høj and Rae Blair

Art direction, design and typography by Geoffrey Reed Communications

Photography by Robert Geh

Vintage photographs courtesy of The State Library of South Australia

Printing by Five Star Press

The staff of The Australian Wine Research Institute

The Australian Wine Research Institute