investigating the factors influencing hop aroma in...

IBD Conference, Wellington, 19-23 March 2018

Investigating the factors influencing

hop aroma in beer

Graham Eyres, Tobias Richter, Jamie Scrimgeour,

Pat Silcock and Phil Bremer

Department of Food Science University of Otago, Dunedin, New Zealand

Graham Eyres, Factors Influencing Hop Aroma in Beer, IBD Conference, Wellington, 19-23 March 2018

Outline

• Introduction: Hop aroma in beer

• Research program at University of Otago

– Research objectives

• Study 1: Effect of yeast strain on

hop-derived compounds in beer

• Study 2: Effect of yeast pitch rate

• Study 3: Online measures during

fermentation

• Summary and next steps


Introduction: Hop Aroma in Beer

• Consumers crave novel aroma and flavour

characters and seeking greater diversity.

• Hop aroma and flavour in beer is a major

driver for this diversity.

– Wide diversity of sensory attributes.



• New aroma cultivars being released

around the world.

– E.g. “Hops with a difference” breeding program

• Selected for novel aroma characters

– But the compounds responsible in beer are not well

characterised.



Hop aroma in beer is not well understood

Opportunity

• Current resurgence of interest in aroma hops and

hop chemistry due to growth of the craft

beer industry.

• Add value to the industry through understanding

of hop aroma in beer.


Research Objectives

Long term goals

• Understand the aroma compounds responsible for

hop derived aroma in beer.

• Understand the factors that influence hop flavour in

beer.

Current research objective

• Investigate the effect of yeast strain and

fermentation parameters on hop compounds in beer.

Target Outcome: Control and optimise hop flavour in beer


Study 1: Effect of yeast strain on hop flavour

Research Question What is the effect of yeast strain on hop

volatile compounds in beer?

+ =

+

+ +


Study 1: Effect of yeast strain on

hop-derived compounds in beer

Introduction

• Evidence that yeast strain and fermentation factors

have a dramatic effect on hop flavour in beer.

• Limited information in the literature of these interactions

under controlled conditions.

Hypotheses:

– Adsorption and loss during fermentation

– Biotransformation of hop compounds

– Release of glycosidically bound compounds

– Sensory interactions with fermentation derived compounds

• The fundamental mechanisms and relative magnitude of

these different effects are not well understood.

Beer fermented in 1L bottles

With aroma hops

No aroma hops

Hot water aroma hop extract

Key parameters:

• 10P wort

• 20 IBU (with Waimea)

• 12 day fermentation at

20C

• 5 day maturation

at 2C

• Yeast: US-05

(1x107cell/mL)

GC-

MS

• 90C for

5 min

First Step: Identification of target compounds

Aroma Hop:

Motueka (5g/L)

90C for 5 min

41 Compounds Significantly Differed

Hop-derived positively influenced by fermentation

Hop-derived negatively influenced by fermentation

Yeast-derived positively influenced by hops

Yeast-derived negatively influenced by hops

Hop-derived not influenced

Hot Water Aroma Hop Extract Beer without Aroma Hops Beer with Aroma Hops

130 Compounds Detected

Main Study Design – Yeast Selection

Strain Details

US-05 Fermentis ale strain

S-33 Fermentis ale strain

BE-256 Fermentis ale strain

WB-06 Fermentis wheat beer

OTA 79 Ale strain

WLP045 Scotch whiskey yeast

WLP730 Chardonnay wine yeast

Exotics SPH Anchor Wine yeast hybrid

OTA 48 Schizosaccharomyces pombe

Fermented @ 20C Fermented @ 15C

Strain Details

W-34/70 Fermentis lager strain

S-23 Fermentis lager strain

S-189 Fermentis lager strain

OTA 19 European lager yeast

VIN 13 Anchor Wine yeast hybrid

OTA 29 Champagne yeast, S. bayanus

15 Yeast strains

Methods

• Wort (10P, 20 IBU) Late Hopped with Motueka (5g/L).

• Each strain: 2 fermentation replicates x 2 yeast cultures.

• Headspace sampling in duplicate with Gerstel Twister (PDMS) and

analysed using GC/MS.

Study 1: Preliminary Results PCA of all yeast strains – selected compounds

BE-256

BE-256

Exotics

Exotics

OTA19

OTA19

OTA29

OTA29

OTA48

OTA48

OTA79

OTA79

S-189

S-189

W-34/70 W-34/70

S-23

S-23

S-33

S-33

US-05

US-05

VIN 13

VIN 13

WB-06

WB-06

WLP045

WLP045

WLP730

WLP730

-6

-4

-2

0

2

4

6

8

-6 -4 -2 0 2 4 6 8 10 12

PC

2 (

16 %

)

PC 1 (25 %)

• Spread of differences in volatile

compounds between yeast

strains

• No distinct groups according to

Ale, Lager or wine yeast types

Wheat beer yeast

Champagne Yeast

Chardonnay Yeast

European Lager

-6

-4

-2

0

2

4

6

8

-6 -4 -2 0 2 4 6 8 10 12

PC

2 (

16 %

)

PC 1 (25 %)

myrcene

linalool

terpinen-4-ol

citronellol

citronellol acetate

geraniol

geranyl acetate

nerol

neryl acetate

methyl geranate

Study 1: Preliminary Results PCA of all yeast strains – selected compounds

Ale Yeasts – Selected Compounds

Study 1: Conclusions

• Different yeast strains resulted in a variety of volatile

compound profiles.

• Evidence in differences in biotransformation of hop

derived compounds.

• Expected to have impacts on hop character and intensity

in beer.

– Ongoing research to look at Sensory Evaluation.


Study 2: Yeast pitch rate on hop compounds in beer

Low Pitch

2 M cells / mL Standard Pitch 10 M cells / mL

High Pitch 50 M cells / mL

Late Hop LH_LP LH_SP LH_HP

No Late Hop NH_SP

Research Question What is the effect of pitch rate on hop

volatile compounds in beer?

• Three pitch rates with

Fermentis SafAle US-05 • Late hopping with

Nelson Sauvin (5g/L)


Sample preparation and analysis

• Wort (10P, 20 IBU) were fermented at 20C to

constant gravity in EBC fermentation tubes.

• Conditioned at 3C for 10 days.

• Fermentations performed in duplicate.

• Samples taken for volatile analysis using GC/MS

using a headspace SPME method (n=3).


Fermentation Profiles

0

5

10

15

20

25

30

35

40

45

0 50 100 150 200 250

YE

AST

CE

LLS

PE

R M

L (M

ILLI

ON

S)

TIME (HOURS)

NH_SP 1 NH_SP 2

LH_LP 1 LH_LP 2

LH_SP 1 LH_SP 2

LH_HP 1 LH_HP 2

0

2

4

6

8

10

12

0 50 100 150 200 250

YE

AST

CE

LLS

PE

R M

L (M

ILLI

ON

S)

TIME (HOURS)

NH_SP 1 NH_SP 2

LH_LP 1 LH_LP 2

LH_SP 1 LH_SP 2

LH_HP 1 LH_HP 2

High Pitch

High Pitch

Std Pitch

Low Pitch

Low Pitch

Std Pitch


Results

• Fermentation esters decreased with increasing pitch rate.

Low Std High Low Std High No Late

Hop

No Late

Hop

• Significant effects of pitch rate on hop-derived compounds



• Concentrations of hop derived compounds varied

with yeast pitch rate.

• Compounds followed different trends.

– Potential for different hop flavour balance.

• Biotransformation pathways require further

investigation (not clear correlations).

• Large differences in fermentation compounds

(e.g. esters) expected to have an impact on hop

flavour through sensory interactions.

Study 3: Tracking aroma compounds during fermentation

Beer prepared with different treatments

Yeast\Hop No aroma hop

Motueka Hops

Nelson Sauvin Hops

No yeast Control MT NS

California Ale Yeast

CA CA / MT CA / NS

Scottish Ale Yeast

SA SA / MT SA / NS

3 ml fermentation in HS vials

Key parameters: • 10P wort • 20 IBU (with Waimea) • Flame-out hop addition: 5g/L • Pitching: ~1x107cell/mL • Volume: 6 replicates with 3mL • 5 day fermentation at 20°C

PTR-TOF-MS Proton transfer reaction-time of flight-mass spectrometer

Richter et al. (2018) Submitted to Food Research International

Ethyl hexanoate

Note the changes in hop related, yeast related and hop / yeast related compounds over time

0

2

4

6

8

10

12

14

16

18

0 6 12 18 24 30 36 42 48 54 60 66 72 78

concentration

[pptv]

fermentation time [hours]

Scottish Ale with Nelson Sauvin

Scottish Ale with Motueka

Scottish Ale without hop

no yeast/no aroma hops

Motueka without yeast

Nelson Sauvin without yeast

0

2

4

6

8

10

12

14

16

18

0 6 12 18 24 30 36 42 48 54 60 66 72 78

concentration

[pptv]

fermentation time [hours]

California Ale with Nelson Sauvin

California Ale with Motueka

California Ale without hop

Motueka without yeast

Nelson Sauvin without yeast

no yeast/no aroma hops

Octanoic acid / Methyl heptanoate

Scottish Ale Yeast California Ale Yeast

NS NS

MT MT

No yeast Control

m/z 145.121

Tracking Fermentation with PTR-TOF-MS

0

500

1000

1500

2000

2500

0

5

10

15

20

25

30

0 6 12 18 24 30 36 42 48 54 60 66 72 78

Concentration EtOH [ppbv]

Concentration CO2 [au]

Fermentation time [h]

Control CO2

Fermentation CO2

Control EtOH

Fermentation EtOH

0 h 42 h

6 h 48 h

12 h 54 h

18 h 60 h

24 h 66 h

30 h 72 h

36 h 78 h

Volatile profile differentiation: At the start of fermentation is due to the hop cultivar’s characteristics At end of fermentation is due to differences in the metabolism of the yeast strains.

631 m/z total 168 m/z Sign Diff

CO2 and ethanol curves show rates of metabolism and changes in metabolism


-8

-6

-4

-2

0

2

4

6

8

-10 -5 0 5 10 15

PC

3 (

9%

)

PC1 (40%)

California Ale with Motueka

California with Nelson Sauvin

Scottish Ale with Motueka

Scottish Ale with Nelson Sauvin

ms43.017 ms61.027 ms62.030

ms73.063

ms75.042

ms89.057 ms90.061

ms103.073 ms117.090 ms118.093

ms131.105

ms145.122 ms146.125

ms173.153

ms81.068

ms87.078

ms95.084

ms129.126

ms137.132 ms138.135

ms75.077

ms101.094

ms231.974

Loadings

Hop effects and Yeast / Hop interaction effects are evident. Esters and Terpenes are mainly associated with the differences between the beers

Possible to differentiate the beers based on the last 4 time

points (time points 60-78h)

SA

CA




• Volatile generation during fermentation is

complex and a dynamic process.

• PTR-ToF-MS can successfully differentiate and

monitor the change in compounds during

fermentation.

– Including hop derived compounds.

• Real-time monitoring provides temporal

information that will enhance understanding of

the biochemical pathways and chemical

processes occurring during fermentation.


Ongoing Research

• Continuing to mine data for hop-yeast

interactions.

• Explore biotransformation pathways and

mechanisms for the differences.

• Link to sensory evaluation to determine the

impact on perceived hop flavour and

aroma.


Summary

• Apply advanced analytical tools to investigate hop

flavour in beer.

– Effect of yeast strain and fermentation.

– Understand changes over time.

• Gain a mechanistic understanding of the hop-yeast

related chemical changes.

• Enable brewers to select and manage yeasts

cultivars to enhance hop aroma in beer.

– Achieve target flavour profile with consistency.

Acknowledgements

• Dr Franco Biasioli, FEMS, San Michele all’Adige

• Tobias Richter acknowledges Emerson’s JP Scholarship to support his studies.

• Jo Pitt (Fermentis) for providing yeast.

• Ron Beatson (Plant and Food Research) and

Doug Donelan (NZ Hops Ltd) for provision of

hop samples.

THANK YOU

Dr Graham Eyres Sensory Science Research Centre Department of Food Science University of Otago Dunedin, New Zealand [email protected] (03) 479 7661

investigating the factors influencing hop aroma in...

Documents