Download Fatty acid productivity of Scenedesmus obliquus under nitrogen

Fatty acid productivity of Scenedesmus obliquus
under nitrogen starvation in mixotrophic cultivation exceeds the
combination of autotrophic and heterotrophic cultivations
Xiao-Fei SHEN (Ph.D. Candidate)
Department of Chemistry
University of Science and Technology of China
[email protected]
Content
1. Background
2. Materials and Methods
3. Results and discussion
4. Conclusions
Content
1. Background
2. Materials and Methods
3. Results and discussion
4. Conclusions
1.Background-Comercial production obstacle
Background
Nutrient, light, water, carbon source
Select species
Cultivation
Harvest
Oil extraction and
transesterification
Biodiesel
Lipid productivity
Biomass productivity
Lipid content
University of Science and Technology of China
Nitrogen
starvation
Lipid
content
Biomass
productivity
Lipid
productivity
decrease
Role of
phosphorus
The effect of P
concentration
on biomass
and lipid
productivities
of microalgae
under nitrogen
starvation
conditions
University of Science and Technology of China
-1
-1
FAME productivity (mg L day )
Autotrophic cultivation of S. obliquus
28
N ＆P
N ＆P-lim
N ＆P-
24
N-＆P
N-＆P-lim
N-＆P-
20
FAME productivity:
16
 N starvation conditions > N sufficient
12
conditions;
8
 N-&P > N-&P-lim > N-&P-
4
0
4th
8th
12th
Time (days)
4th
16th
Bioresource Technology . 2014, 152: 241–246
Heterotrophic cultivation of S. obliquus
with acetate
55.9mg/L/d
 Both the highest FAME
productivity and FAME
yield were also obtained
under N-&P.
 FAME productivity under
nitrogen starved
conditions increased
fourfold than that under
nitrogen sufficient
conditions.
Applied Energy. 2015, 158: 348–354
Whether oil production from mixotrophic culture can exceed the sum of
those from autotrophic and heterotrophic culture？
Contradictory conclusions
from previous studies
Influence factors?
Algae species and carbon source
Nitrogen starvation can significantly increase the lipid content and the lipid productivity of
microalgae in both autotrophic and heterotrophic systems
Mixotrophic cultivation
The cell concentration and lipid production of
Scenedesmus
obliquus
The lipid production of Chlorella vulgaris under
mixotrophic Chlorella sp. was even higher
Acetate
mixotrophic cultivation was lower than under
than the sum of those from photoautotrophic
Nitrogen
starvation
heterotrophic cultivation due to the unsatisfactory
and heterotrophic culture.
lipid content (13.8%) of mixotrophic culture
Content
1. Background
2. Materials and Methods
3. Results and discussion
4. Conclusions
Materials and Methods
 Algae species: Axenic Scenedesmus obliquus NIES-2280
 Basic medium: BG-11 medium
 5 g·L-1 sodium acetate was added as organic carbon source
 Nitrogen source was removed from the media
 Phosphorus concentration: 40 mg·L-1 (Sufficient)
 Proteomics analysis: the isobaric tags for the relative and
absolute quantitation technique (iTRAQ)
Calculation of biomass productivity
and fatty acid productivity
Biomass productivity was calculated based on:
1
−1
Biomass productivity 𝑚𝑔 · 𝐿
2
·𝑑
−1
=
𝐵𝑖𝑜𝑚𝑎𝑠𝑠 𝑚𝑔 ·𝐿−1 𝑡1 −𝐵𝑖𝑜𝑚𝑎𝑠𝑠 𝑚𝑔 ·𝐿−1 𝑡0
𝑐𝑢𝑙𝑡𝑖𝑣𝑎𝑡𝑖𝑜𝑛 𝑡𝑖𝑚𝑒 𝑑
1
FA (fatty acid) productivity was calculated based on:
2
FA productivity 𝑚𝑔 · 𝐿−1 · 𝑑−1
𝐵𝑖𝑜𝑚𝑎𝑠𝑠 𝑚𝑔 · 𝐿−1 × 𝐹𝐴 𝑐𝑜𝑛𝑡𝑒𝑛𝑡 𝑚𝑔 · 𝑔−1 𝑡1 − 𝐵𝑖𝑜𝑚𝑎𝑠𝑠 𝑚𝑔 · 𝐿−1 × 𝐹𝐴 𝑐𝑜𝑛𝑡𝑒𝑛𝑡 𝑚𝑔 · 𝑔−1
=
𝑐𝑢𝑙𝑡𝑖𝑣𝑎𝑡𝑖𝑜𝑛 𝑡𝑖𝑚𝑒 𝑑
3
𝑡0
Calculation of fatty acid yield
1
The fatty acid yield was calculated based on COD coefficient (the amount of O2
2
needed to oxide the COD, O2 g/ COD g). COD coefficient of acetate and fatty acids
3
(CxHyOz) could be calculated by Eq. (1):
4
5
6
𝐶𝑂𝐷 𝑐𝑜𝑒𝑓𝑓𝑖𝑐𝑖𝑒𝑛𝑡 𝐶𝑥 𝐻𝑦 𝑂𝑧 =
2𝑥+𝑦 2−𝑧 ×16
12𝑥+𝑦+16𝑧
(1)
Then the fatty acid yield could be obtained by Eq. (2):
𝐶𝑂𝐷 𝑓𝑎𝑡𝑡𝑦 𝑎𝑐𝑖𝑑 ×𝑀𝑎𝑠𝑠 𝑓𝑎𝑡𝑡𝑦 𝑎𝑐𝑖𝑑 𝑔
𝑎𝑐𝑒𝑡𝑎𝑡𝑒 ×𝑀𝑎𝑠𝑠 𝑐𝑜𝑛𝑠𝑢𝑚𝑒𝑑 𝑎𝑐𝑒𝑡𝑎𝑡𝑒 𝑔
Fatty acid yield = 𝐶𝑂𝐷
(2)
Content
1. Background
2. Materials and Methods
3. Results and discussion
4. Conclusions
During the whole cultivation period, the
biomass productivities of the mixotrophic
culture exceeded the combination of the
autotrophic and heterotrophic cultures.
These two absorption curves are very
similar, and no significant difference was
found between the assimilation rates
from the heterotrophic and mixotrophic
cultures
1. At the end of the experiment, the fatty
acid contents of the autotrophic,
heterotrophic, and mixotrophic cultures
were 19.7, 46.7, and 58.3%, respectively.
2. The highest fatty acid content was
obtained in the mixotrophic culture.
3. The fatty acid content of all the three
groups rose steadily and significantly.
 C18:1 was the most predominant composition, accounting for 52.8% and 66% of the
total fatty acids in the mixotrophic and heterotrophic cultures, respectively.
 The highest unsaturated fatty acid content was obtained in mixotrophic culture
 The protein contents of all the three
systems decreased significantly during
the cultivation period
 The lowest protein content was
obtained in the mixotrophic culture; it
declined sharply from 55.6% to 9.1%
during the nine-day cultivation period.
 No significant difference was observed
between the results for the
heterotrophic and mixotrophic cultures
Fatty acid productivity
(mg/L/d)
Fatty acid yield
Autotrophic
14.7±2.1
-----
Mixotrophic
118.4±6.4
0.45±0.04*
Heterotrophic
57.5±5.3
0.23±0.02
Note: the fatty acid produced through autotrophic process has been deducted.
More assimilated acetate is directed to lipid synthesis rather than protein
and starch accumulation with the presence of light and supply of CO2.
Auto-
Hetero-
Mixo-
Proteomics analysis
1,065 proteins
were identified
Isobaric tags
for the
relative and
absolute
quantitation
technique
(iTRAQ)
Compare the
expression
levels of
proteins from
mixotrophic and
heterotrophic
cultures
Identify key
proteins of
lipid
synthesis
299 proteins had
significant changes in
the expression level
Some proteins
participated in
growth and lipid
synthesis
processes
Accession
Biological process
Protein name
Peptides (95%)
Fold change
CV
Expression
A8IWA6
Growth
Glutamate synthase, NADH-dependent
15
1.71
0.10
Up-regulated
A8HNQ7
Lipid metabolism
Thioredoxin reductase
1
2.42
0.37
Up-regulated
A8IRQ1
Lipid metabolism
Ribose-5-phosphate isomerase
7
1.77
0.01
Up-regulated
A8JGJ6
Lipid metabolism
Mg protoporphyrin IX S-adenosyl methionine
O-methyl transferase
2
1.77
0.04
Up-regulated
A8J2S0
Small molecular metabolism
Citrate synthase
3
2.74
0.10
Up-regulated
A8J0R7
Generation of precursor
metabolites and energy
Isocitrate dehydrogenase
1
2.60
0.14
Up-regulated
A0A0D2K714
Carbohydrate metabolism
Pyruvate kinase
15
2.17
0.09
Up-regulated
B6E5W6
Carbohydrate metabolism
Glucose-6-phosphate isomerase
9
1.71
0.10
Up-regulated
A0A0D2NR30
TCA cycle
The activity of the TCA cycle is improved in mixotrophic culture,
Catabolic process
Glycerol-3-phosphate dehydrogenase
resulting in more
fatty acid synthesis in2S. obliquus2.52cells. 0.23
Up-regulated
A8I8Z4
Lipid metabolism
Ribosomal protein
2
0.59
0.10
Down-regulated
A8IKQ0
Lipid metabolism
Fructose-1,6-bisphosphatase
4
0.52
0.22
Down-regulated
D8TTF7
Lipid metabolism
Plastid acyl-ACP desaturase
2
0.15
0.22
Down-regulated
A0A0D2JX51
Small molecular metabolism
Malate dehydrogenase
16
0.65
0.19
Down-regulated
Q4U1D9
Biosynthetic process
Soluble starch synthase III
3
0.58
0.10
Down-regulated
Q8VXQ9
Carbohydrate metabolism
Glyceraldehyde-3-phosphate dehydrogenase
A, chloroplast
20
0.24
0.48
Down-regulated
Content
1. Background
2. Materials and Methods
3. Results and discussion
4. Conclusions
Conclusions
Under nitrogen starvation, the biomass and biodiesel productivities of
mixotrophic S. obliquus exceeded the combination of autotrophic and
heterotrophic cells when using acetate as carbon source.
The fatty acid yield from mixotrophic culture (0.45) was almost two times
greater than for heterotrophic culture (0.23).
Proteomics analysis revealed that the activity of the TCA cycle was
improved in mixotrophic culture when compared with heterotrophic culture.