Heterotrophic algae

1. Biofuels production from Microalgae
after heterotrophic growth
(藻类异养转化制备生物质液体燃料)
Qingyu Wu (吴 庆 余) Xiaoling Miao (缪晓玲)
Department of Biological Sciences and Biotechnology, Tsinghua
University, Beijing 100084, P.R.China
(清华大学生物科学与技术系）

2. BIOMASS -BIOFUEL
higher plants or woody materials :
pine wood, rice and cotton straw and
stalk, energy crops
Microalgae advantages:
Higher photosynthetic efficiency, larger
biomass, faster growth, easy industry
culture 5days
10 years cell culture engineering

3. 1. Materials and biotechnolgy
Chlorella protothecoides 小球藻
a b
light microscope electron microscope

4. photoautotrophic heterotrophic photoautotrophic heterotrophic
(in liquid medium) (on agar plates)
By cell biotechnology to change the nutrition metabolism
pathway, green autotrophic cells (AC cells) were
transformed to heterotrophic growth.

5. Yellow heterotrophic cells (HC cells) under light
microscope (10×40) lipid vesicles

6. Fig. 7 Differences in cell organization between HC and AC cells.
（图7 小球藻异养和自养转化细胞结构变化示意图）

7. The contents of main chemical components in cells of
AC and HC.
（表1 AC、HC细胞主要生化成分含量）
Strain Protein Lipid Carbohydrate Ash Moisture Others
(%) (%) (%) (%) (%) (%) (%)
AC 52.64 14.57 10.62 7.38 8.74 6.05
HC 10.28 54.70 15.19 7.24 5.40 7.19

8. 1 2
The culture process of the HC cells by “Two Step”
method.

9. 12 12
无菌培养
开放式培养
9 9
OD 540
OD540
6 6
10L下口瓶半无菌培养
10L下口瓶补料半无菌培养
3 3
20L下口瓶半无菌培养
20L下口瓶补料半无菌培养
0 0
0 24 48 72 96 120 0 24 48 72 96 120
Time (h) Time (h)
Comparison of the HC cell growth under axenic,
half-axenic and no-axenic culture conditions

10. 2. Fast pyrolysis of HC cell
AC HC Microcystis
Dry cells powder

11. Fast pyrolysis system and conditions
Pyrolysis system: the fluid bed reactor
Temperature ：500℃
Heating rate ：600 ℃/s
Vapor residence time：2-3 s
biomass-feeding rate ：4 g/min
particle size < 0.5 mm

12. The diagram of fast pyrolysis system of fluid bed

13. Yields of fast pyrolysis at temperature of 500 ℃,
heating rate of 600 ℃ /s and the sweeping gas flow
rate of 0.4m3/h.
Strain Gas Bio-Oil Char
(%) (%) (%)
AC 29.6 16.6 53.8
HC 31.6 57.2 11.2
Microcystis 55.2 23.7 21.1
All the yields were expressed on the basis of the dry
weight of samples.

14. 60
Product Yield (%) 50
40
30
20
10 Char
Gas
0
AC Bio-oil
HC
M
Product yields of microalgae by fast pyrolysis at
temperature of 500 ℃, heating rate of 600 ℃ /s and the
sweeping gas flow rate of 0.4m3/h.

15. AC HC Microcystis
Bio-Oil products of AC, HC and Microcystis by fast
pyrolysis.

16. Comparison of typical properties of fossil oil and
fast pyrolysis oil of wood and HC.
Typical value
Properties Bio-oils Fossil oil
Wood AC HC
C 56.4% 62.07% 76.22% 83.0-87.0%
H 6.2% 8.76% 11.61% 10.0-14.0%
O 37.3% 19.43% 11.24% 0.05-1.5%
N 0.1% 9.74% 0.93% 0.01-0.7%
S n.d. n.d. n.d. 0.05-5.0%
Density (kg l-1) 1.2 1.06 0.92 0.75-1.0
Viscosity (Pa s) 0.04-0.20 0.10 0.02 2-1000
(at 40oC) (at 40oC) (at 40oC)
Heating value 21 30 41 42
(MJ kg-1)
Stability Not as stable as Not as stable as fossil fuels, but more
fossil fuels stable than the bio-oil from wood

17. 3. Biodiesel production from microalgal oil
(利用微藻油脂制备生物柴油)
1. Biodiesel （生物柴油）
2. The preparation of microalgal oil
（微藻油脂的制备）
3. Acidic transesterification of microalgal oil
（酸催化酯交换法制备微藻生物柴油）
4. Characteristics of biodiesel from microalgal oil
（微藻生物柴油的特性）

18. Biodiesel (生物柴油)
Fatty acid methyl esters originating from vegetable oils
and animal fats are known as biodiessel.
Biodiesel from transesterification can be used directly
or as blends with diesel fuel in diesel engine
催化剂
甘油三酯 甲醇 脂肪酸甲酯 甘油
生物柴油

19. Oil from microalgae (微藻油脂的提取)
Growth of HC cells (A), powder of HC cells (B) and microalgal oil
from HC cells (C).

20. Method of Acidic transesterification for biodiesel preparation from
microalgal oil: (微藻生物柴油制备: 酸催化酯交换技术)
The reaction mixture consisted
of oil, methanol and
concentrated sulfuric acid The separation of the reaction
mixture (A) and the biodiesel from
microalgal oil (B,C)

21. Comparison of biodiesel from microalgal oil and diesel fuel
(微藻生物柴油与石化柴油的特性比较)
Propertis Biodiesel from Diesel fuel
(特性) microalgal oil (石化柴油)
(微藻生物柴油)
Density 密度（kg l-1） 0.864 0.838
Visicosity 粘度（Pa s） 5.2×10-4 (40 °C ) 1.9-4.1 ×10-4 (40 °C )
Flash point 闪点（ °C ） 115 75
Solidifying point 凝点（ °C ） -12 -50 – 10
Cold filter plugging point -11 -3.0 (-6.7 max)
冷滤点（ °C ）
Acid value 酸值 (mg KOH g-1) 0.374 0.5 max
Heating value 热值（MJ kg-1） 41 40-45
H/C ratio 1.18 1.81

22. Summery
1 Heterotrophic growth of Chlorella protothecoides resulted
in the accumulation of a large amount of lipid in cells,
which is about 4 times of that in control. By using HC cells
rich in lipids as biomass for pyrolysis and biodiesel
preparation, we got high yield and high quanlity bio-fuel.
2 The results suggests that the new process, which combines
biotechnology with biofuel production, is a feasible and
effective method for the production of high yield and high
quality fuel oils from microalgae.
3 The study could contribute to the creation of a system to
produce biofuels from microalgae which might have great
commercial potential for liquid fuel production.

23. Thank you !