Mixotrophic continuous cultivation of
Chlorella protothecoides for sustainable
lipid production
Algal Biomass Summit 2013
October 1st, Orlando Florida, US
Yanming Wang, Heiko Rischer and Marilyn Wiebe
VTT Technical Research Centre of Finland
07/10/2013
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VTT in brief
VTT Technical Research Centre of Finland is the largest multi-technological
applied research organisation in Northern Europe.
VTT Group
Turnover: 316 M€ (31.12.2012)
Personnel: 3,000
President & CEO: Erkki KM Leppävuori
Established: 1942
Headquarter: Otaniemi, Espoo, Finland
Part of Otaniemi bird view (www.futureblock.fi)
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VTT group on the map
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VTT Industrial Biotechnology’s interests in algae
Metabolic
engineering
Algal
physiology
Cultivation
process
Mixotrophic continuous cultivation of Chlorella
protothecoides for sustainable lipid production
Algal
biomass
production
Algal lipid
production
Mixotrophic
cultivation
Biorefinery
Waste water
treatment
Bioactive
compound
discovery
Sustainable
process
Wang, Y., Rischer, H., Eriksen, N.T. & Wiebe, M.G. Mixotrophic continuous flow cultivation of
Chlorella protothecoides for lipids. Bioresource technology 2013, Vol. 144, pp. 608.
07/10/2013
Mixotrophy, an alternative solution?
Autotrophy
•
•
•
•
CO2 consuming
Low cell density
Low productivity
High operational cost
Heterotrophy
• High cell density
• High productivity
• Expensive substrate
• What is the scenario of mixotrophy?
• Will mixotrophy bring any benefit?
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Experiments
Batch cultivations:
• Mixotrophic
• Autotrophic
Glucose as organic
carbon source
Different nitrogen
concentration
tested
Continuous cultivations:
• Mixotrophic
• Heterotrophic
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Mixotrophy improves the cell density and productivity
Autotrophy Mixotrophy
x (g L-1)
5.9
8.5
rx (g L-1d-1)
0.3
1.8
These results clearly
showed the contribution of
heterotrophy in a
mixotrophic culture.
Batch cultures of C. protothecoides in defined medium with
(mixotrophic) and without (autotrophic) glucose (16 g L -1), in
a stirred tank reactor with external illumination.
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Applying nitrogen limitation in mixotrophic batch cultures
24 mM N
14 mM N
C/N
>22
>38
x (g L-1)
8.5
8.5
L (g L-1)
2.2
2.9
rx (g L-1d-1)
1.80
1.80
rL (g L-1d-1)
0.46
0.61
L% (% of x)
25.3%
34.1%
Yx/Glu (g g-1)
0.50
0.53
Wang et al. 2013
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Biomass and lipid production at different C/N
Biomass yield was
not affected by C/N
Lipid content and
yield increase with the
decreasing nitrogen
57% total lipid content
observed
Mixotrophic steady states,
dilution rate of 0.44 d-1
Wang et al. 2013
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Mixotrophy in extreme nitrogen limitation conditions
Choice of energy
source:
Light or glucose?
6 mM nitrogen, dilution rate of 0.44 d-1
Wang et al. 2013
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Mixotrophy in moderate nitrogen limitation conditions
Difference of
product profile:
Biomass and CO2
14 mM nitrogen, dilution rate of 0.44 d-1
Wang et al. 2013
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Carbon fluxes in mixotrophic chemostat steady state
Glucose
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Carbon flux calculation
Feed N
Carbon
Light
Glucosein
CO2 in
Glucoseout
CO2 out
Biomass out
(mM)
balance
14
+
94.5
5.5
0
32.7
62.6
0.95
14
-
94.4
5.6
0
50.2
40.2
0.90
6
+
94.3
5.7
7.5
37.8
53.2
0.98
6
-
94.3
5.7
0
43.5
53.7
0.97
assuming a formula of CH1.775O0.459N0.148P0.010S0.006 (50.7% of biomass is carbon) for Chlorella.
Wang et al. 2013
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CO2 and O2 internal circulation, during mixotrophy
H2O + light
energy
CO2
autotrophy
biomass
biomass
heterotrophy
O2
glucose
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Conclusions
 Autotrophy and heterotrophy are additive or synergistic in
mixotrophic C. protothecoides culture.
 Continuous mixotrophic algal cultivation is a good way to
maintain high algal lipid content and production rate.
 C. protothecoides produced up to 57% lipid in its biomass in
mixotrophic N-limited culture.
 When growth was limited by nitrogen source, C. protothecoides
preferred CO2 as carbon source or light as energy source.
07/10/2013
Microalgal research at VTT – contact persons
Heiko Rischer, Plant Biotechnology
[email protected]
Laura Ruohonen, Metabolic Engineering
[email protected]
Marilyn Wiebe, Bioprocess Technology
[email protected]
Jaakko Peres, Renewable Materials
[email protected]
Mona Arnold, Green Solutions for Water and Waste
[email protected]
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VTT creates business from technology
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