microalgal bioprocessing - introduction
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MicroalgalBiotechnology:
CultivationandProcessing
MBT Fall2015
October29th,2015 HectorDelaHozSiegler
DepartmentofChemicalandPetroleumEngineering
UniversityofCalgary
HectorDelaHozSiegler,Ph.D.,P.Eng.
Outlineoftodaystalk
I. Introduc tion to microalga e
What and why
Applications
II. Renewable energy from microalga e
Motivation
Biofuel portfolio
Biodiesel
III. Culturing techniques
Medium requirements
Open ponds and photobioreactors
Phototrophic and heterotrophic
IV. Optimization of heterotrophic culturesV. Summary
INTRODUCTIONTOMICROALGAL
BIOTECHNOLOGY
PartI
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Microalgae:whatarethey?
Microalgaeareplantlikeunicellularorganisms:apolyphyletic group
ofphotosyntheticeukaryotes.
Sunlight-driven cell factories able to convert carbon dioxide to
potential biofuels, food, and high-value products
4
ProductsfromMicroalgae
Image source: Rosenberg, J.N., Oyler, G.A., Wilkinson, L., Bet enbaugh, M.J.Agreen light for engineeredalgae: redirecting metabolismto fuel abiotechnology
revolution ,Current Opinion in Biotechnology, 19 (5), pp. 430-436 (2008)
Applications
6
Microalgae
Biofuels
Finechemicals:
e.g.antioxidants
Wastewater
treatment/
Remediation
Pharmaand
nutraceuticals
Humanand
animalfood
CO2Capture
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Microalgaeascellfactories
Benefits:
Highlyefficient
Simplenutritional
requirements
Easilyadaptabletoenvironmentalstresses
Produceandstorehighamountsofoil
Othervaluablebyproducts
Eukaryotic!
Challenges
Lowculturedensity
Slowgrowth:lowproductivity(comparedtobacteriaandyeast)
Highproductioncosts
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SomeCommercialApplications
S pe ci es/ gr ou p Pro du ct A pp li ca ti on a re as Pr od uc ti on
facilities
References
Haematococcus
pluvialis /
Chlorophyta
Carotenoids,
astaxanthin
Health food, feed additives
and pharmaceuticals
Open ponds,
PBR
Del Campo et al. (2007)
Odontella aurita
/ Bacillariophyta
Fat ty acids Pha rm aceu ti ca ls ,
cosmetics, baby food
Open ponds Pulz and Gross (2004)
Isochrysis
galbana /
Chlorophyta
Fatty acids Animal nutrition Open ponds,
PBR
Molina Grima et al.
(1994); Pulz and Gross(2004)
Phaedactylum
tricornutum /
Bacillariophyta
Lipids, fatty
acids
Nutrition, fuel production Open ponds,
basins, PBR
Yongmanitchaiand Ward(1991); Acien-
Fernandez et al. (2003)
Muriellopsis sp.
/ Chlorophyta
Carotenoids,
Lutein
Health food, food
supplement, feed
Open ponds,
PBR
Blanco et al. (2007); Del
Campo et al. (2007)
Crypthecodinium
cohnii
DHA Food additive Fermenters
(heterotrophic)
Carvalho et al.
(2006)
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Currently,applications
of
microalgal
biotechnology
are
limited
to
niche
(small)
markets.
Though
highvalue!Weexpecttomoveintolargescalemarkets.
ALGAEASASOURCEOF
RENEWABLEENERGY
PartII
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Energy,economy,andGlobalwarming
Poland
Uzbekistan
Chad
Ghana
Vanuatu
India
China
Russia
Brazil
UK
France
US
Norway
Japan
South
Korea
Canada
China (2002)
India(2002)
US(2002)
0.01
0.1
1
10
100
100 1,000 10,000 100,000
PrimaryEnergyConsumption(kWyear/p
erson)
GDP($/person/year)
Energyreserves/Energyconsumption
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Climatechangedebate
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Thenaturalcarboncycle
G. A. Olahand colleagues, J.Am.Chem.Soc, 133, 12881-12898, 2011
Carbon CaptureandUtilisation(CCU)
T. Shirvani, X. Yan, O. R. Inderwildi, P. P. Edwards and D. A. King, Energy Environ. Sci., 2011, 4, 37733778
Biofuels
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1stGeneration: derived from food-crops, i.e.ethanol from sugar cane or corn, biodieselfrom canola or soybeans.
2ndGeneration
: produced from lignocellulosicmaterials, i.e. ethanol from wood chips,switch grass.
3rdGeneration: fuels from microalgae
4thGeneration: from crops designed for fuelsin combination with highly efficient microbes.
Timetorealworldapplication
Landrequiredforsatisfydemand
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Themicroalgalbiofuelsportfolio
Algal Biomass:
- Oil/Lipids
- Sugars/Starch
- Lignocellulose
Excreted products:
- Hydrogen
- Alcohols
Sugars
Bio-oil
SynGas
Biodiesel
Green Diesel
Gasoline
Hydrogen
Alcohols
CO2
Water
Sunlight
Trace elements
Feedstocks Photosynthe sis Inte rme diates Fu els
Pyrolysis
Hydrolysis
Hydrodeoxygenation
Hydrotreating
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BiodieselfromMicroalgae
Crop Oil yield
(L/Ha)
Land area needed
(M Ha)
% of existing US
cropping area
Corn 172 1540 846
Soybean 446 594 326
Canola 1190 223 122
Oil Palm 5950 45 24
Microalgae(70% oil w/w)
136900 2 1.1
Microalgae(30% oil w/w)
58700 4.5 2.5
Biodieselderivedfromoilcropsisapotentialrenewableand
carbonneutralalternativetopetroleumfuels.
Biodieselfromoilcrops,wastecookingoilandanimalfatcannot
realisticallysatisfythedemandfortransportfuels.
Croplandrequirement
bydifferentoilcropsto
replace50%ofall
transportfuelneedsof
theUS.Chisti(2007).
Toooptimistic
to
be
true!
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Algaeasasourceofoil
Species Oil content
(% dw)
Reference
Ankistrodesmus TR-87 28 40 Ben-Amotz and Tornabene (1985)
Botryococcus braunii 25 75 Sheehan et al. (1998); Banerjee et al. (2002); Metzger and Largeau (2005)
Chlorella sp. 28 32 Sheehan et al. (1998), Chisti (2007)
Chlorella protothecoides 15 55 Xu et al. (2006)
Cyclotella DI-35 42 Sheehan et al. (1998)
Dunaliella tertiolecta 36 42 Kishimoto et al. (1994); Tsukahara and Sawayama (2005)
Hantzschia DI-160 66 Sheehan et al. (1998)
Isochrysis sp. 7 33 Sheehan et al. (1998); Valenzuela-Espinoza et al. (2002)
Nannochloris 20 -35 (6 -63) Ben-Amotz and Tornabene (1985); Negoro et al. (1991); Sheehan et al. (1998)
Nannochloropsis 46 (31 -68) Sheehan et al. (1998); Hu et al. (2006)
Nitzschia TR-114 28 50 Kyle DJ, Gladue RM. WO 91/14427 (Patent)
Phaeodactylum tricornutum 20 31 Sheehan et al. (1998), Chisti (2007)
Scenedesmus TR-84 45 Sheehan et al. (1998)
Stichococcus 33 (9 -59) Sheehan et al. (1998)
Tetraselmis suecica 15 32 Sheehan et al. (1998); Zittelli et al. (2006); Chisti (2007)
Thalassiosira pseudonana (21 -31) Brown et al. (1996)18
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OilandBiodiesel
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Triglycerides:
BiodieselProduction:
Glycerol
FattyAcids
PolyunsaturatedFattyAcids(PUFA)
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Fattyacidswithmultipledoublebonds
C18:3andlongerareessential:mammalscannotsynthesize
C18:3.Needtotakethemfromtheirdiet
Multiplebiologicalfunctionsassignallingmoleculesorbuilding
blocks
DHA: C22:6
EPA: C20:5
MicroalgaeasaSourceof3PUFA
FishoilhasbeenusedforthecommercialproductionofEPAandDHA.
Factorsthatlimitfishoilasasourceof3fattyacidsinclude:taste,odourandstabilityproblems.Highpurificationcost.
Fishobtain3fattyacidsfromtheirdiet.
SeveralspeciesofmicroalgaeareprimaryproducersoflongchainPUFA.
US$1.5billion/yeargeneratedfromtheproductionofDHA(Pulz andGross,2004).
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PUFAproportionsinMicroalgae(%TFA)
Organ is ms ARA ( 20 :4 ) EPA ( 20 :5 ) DHA ( 22 :6 )
Gymnodinium splendens 8 30
Cricosphaera elongata 2 28
Isochrysis galbana 15 7.5
Monodus subterraneus 4.7 33
Nannochloropsis sp. 35
Schizochytrium sp. 1.0 2.3 40.9
Chlorella minutissima 5.7 45
Hetermastrix rotundra 1 28 7
Chromonas sp. 12.0 6.6
Cryptomonas sp. 16 10
Rhodomonas sp. 8.7 4.6
Asterionella japonica 11 20
Biddulphia sinensis 24 1
Crypthecodinium cohnii 30
Nitzschia laevis 6.2 19.1
Phaeodactylum Tricornutum 34.5
Skeletonema costatum 29.2 22
GeneralProcessDiagram
Harvesting
Dryer
CultureExtraction
Crude
Product
debris
S/L Separator
Solvent
recovery
Cell disrupter
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MICROALGALCULTURING
TECHNIQUES
PartIII
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Nutritionalrequirements
Dependsonapplication
Foodorhealthoils:foodgradechemicals
Otherwiseindustrial
chemicals
or
seawater
/wastewater
Carbonsource:CO2,sugars,acetate,ethanol
Macronutrients:Nitrogenandphosphorus
Micronutrients:Fe,Mg,Si,S,K
Traces:Ca,Mn,Zn,Co,Se,Cu,Mo
Vitamins:B1,B12,B6,B2
Seawater:Na,K,Mg,Ca,Cl,SO4,HCO3,BO3
Br,F,IO3,Li,Rb,Sr,Ba,Mo,V,Cr,As,Se
NO3,PO4,Fe,Zn,Mn,Cu,Co,Si,Ni25
Multiplewaysofgrowing
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Mixotrophic
FixCO2
Fast growth
Productivealldaylong
Complexand
unknowncellregulation
CO2 capture
Waste
water
treatment Highvalueproducts
Proteins
EnergySource
Applications
Phototrophic
FixCO2
Slowgrowth
Noactivityduringnight
CO2capture
Heterotrophic
Fast growth
Productivealldaylong
Dontfixgreenhousegases
ProduceCO2
Wastewatertreatment
Highvalue
products
Proteins
SolarradiationinCanada
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SolarradiationinAlberta
FortMcMurray: 4 181MJ/m2y
Edmonton: 4510MJ/m2y
MedicineHat: 5221MJ/m2y
Munich(GE): 4044MJ/m2y
Naples(IT): 5293MJ/m2y
KualaLumpur: 5622MJ/m2y
Orlando(FL): 5922MJ/m2y
Acapulco(MX): 7261MJ/m2y
Phoenix(AZ): 7621MJ/m2y
Solarradiation datatakenfrom:U.S.Department ofEnergyEnergyPlusWeather Data.
http://apps1.eere.energy.gov/buildings/energyplus/cfm/weather_data.cfm
Culturingtechniques:OpenPonds
Byfar,themostcommonproductionsystem.
Lowinstallationcost
Lagoonsorartificialponds
Highriskofcontamination: Unwantedalgae
Grazers
Applicationlimitedtofewspecies(extremophiles).
Unmixedponds:arearangefrom1200Ha,depth2030cm
Racewaypondsareupto1Ha.
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Culturing:ClosePondsandTanks
Simplerdesignssimilartoopen
ponds,withacover
(greenhouses).
Aimtoreducecontamination
risks. ControlCO2looses.
Tanksareusuallymixedby
aeration.
Deeptanksareinefficient.Bad
lighttransmission.
Easytooperate,lowcost.
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Culturing:Photobioreactors
TubularPhotobioreactor AlgaeandBiofuelsFacility, SouthAustralianResearchandDevelopmentInstitute
FlatPanelphotobioreactor
ArizonaCenterforAlgalTechnology andInnovation
FlexibleplasticfilmPhotobioreactorAlgenol,Florida 31
Culturing:Photobioreactors
Betterculturecontrol
Higherproductivity,andculture
density
Minimalcontaminationrisk
Wellmixed
Excellenttemperaturecontrol
Oxygencontrolisanissue
Highcapitalinvestment
Frequentcleaningrequired
Coolingrequired
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HeterotrophicProductionofAlgae
Somealgaespeciescangrow
usinganorganiccarbonsource.
Conventionalbioreactorscanbe
used.
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Phototrophicvs.Heterotrophic?
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SpecieOil content
(%)
Cell conc.
(g/L)
Oil Prod.
(mg/L d)References
Ettlia oleoabundans 36 42 2.9 164 Griffiths et al (2009);
Li et al. (2008)
Nannochloropsis sp. 31 68 2.1 204 Rodolfi et al. (2009)
Amphora 40 51 - 593 Sheehan et al. (1998)
Chlorella sp. 28 32 1.1 139 Hsieh and Wu (2009)
Chlorella vulgaris 25 42 1.7 54 Liang et al. (2009)
Chlorella zofingiensis 25.8 1.9 35 Liu et al. (2010)
Chlorella zofingiensis 51.1 9.6 354 Liu et al. (2010)
Nitzschia laevis 16.5 22.1 914 Wen and Chen (2003)
S. Limacinum (DHA) 17.3 37.9 656 Chi et al. (2009)
A. protothecoides 38.3 53.0 8.4 820 Cheng et al. (2009)
A. protothecoides 50.3 57.8 51.1 3320 Xiong et al. (2008)
Phototrophic
Heterotrophic
MODELBASEDOPTIMIZATIONOF
HETEROTROPHICALGALCULTURES
PartIV
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BioprocessOptimization
36
Strainselection
Mediaformulation
Processconditions
Continuous/
Realtime
Geneticmodification
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TheObjectiveforOptimization
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StressOil sto ring is a metabol ic response to stress,
particularly nitrogen def iciency. At nitrogen
deficient conditions, algal cells over-accumulatelipids.
The challenge is to maximize biomass production
while keeping a high oil content. It is necessary to
determine the nitrogen supplementation strategy
to achieve this.
Nitrogen
As nitrogen is required for protein synthesis, its
deficiency negatively affects growth and cel l
functioning. Therefore, conditions that favored oil
accumulation constraint productivity.
Understandingalgalgrowth
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Nitrogen uptake
Lipid production
Cellular growth
Analgalgrowthmodel
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Cellular growth
Nitrogen uptake
Oil production
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Macroscopicbalances
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Optimization:Problemformulation
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Subject to:
Simulationresults
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Biomassproductivityin
continuouscultures
Lipidproductivityin
continuouscultures
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Experimentalresults
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Biomassproductivityandgrowthrate
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Lipidproductivityandproductionrate
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Comparativestudy:growthonglucose
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Specie Lipid content
(%, w/w)
Oil Productivity
(g/L h)
References
E. coli (gen. modified) 25.4 0.246 Elbahoul et al (2010)
R. opacus PD630 38.4 0.171 Kurosawa et al (2010)
M. ramanniana 67.7 0.17 Hiruta et al (1997)
C. echinulata 26.9 0.07 Kosa et al (2011)
R. toruloides 67.5 0.54 Li et al. (2007)
L. starkeyi 56.0 0.04 Kosa et al. (2011)
C. curvatus 82.7 0.47 Zhang et al. (2011)
Schizochytrium sp. 30 0.096 Ganuza et al (2007)
C. vulgaris 9.7 0.12 Doucha et al. (2011)
A. protothecoides 50.3 0.14 Xiong et al. (2008)
A. protothecoides 49.4 0.43 0.84 De la Hoz et al (2012)
Bacteria
Molds
Yeasts
Microalgae
Optimization:closingremarks
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Modelbased optimization of heterotrophic microalgal cultures
allowed to reach very high densities, with biomass productivity
greater than 30 g/L d, and as high as 70 g/L d.
High oil content (4060% w/w) can be sustained with a lipid
productivity around 20 g/L d.
High quality monitoring and control is essential to achieve high
productivities.
Better control / sensors = higher productivity.
Summary
Algaearepromisingorganisms:highlyefficient
Goodsourceofoil:PUFA,biodieselprecursor
Algaecan
growth
on
simple
inexpensive
media
Severalreactortypesandgeometry.Applicationwilllimit
reactorchoice
Severalsuccessfulcommercialapplicationscurrentlyworking.
Alotofresearchisstillneeded!
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