My thesis proposal

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Thesis Proposal
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Extractive fermentation of succinic acid
using an external precipitation technique
การแยกกรดซัก ซิน ิก ควบคู่ก ับ กระบวนการหมัก
ด้ว ยเทคนิค การตกตะกอนนอกถัง หมัก
Asst. Prof. Apichat Boontawan
Jiraphorn Lubsungnoen M5430116

2. Content 01
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•Background
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•Research Objective
•Research Hypothesis
•Experimental
- Material & Method
- Experimental setup
•Expected Result
•Research Plan

3. Background 02
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- Currently, the interest on development of
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biodegradable plastic since global warming
problem has come.
- Succinic acid (SA) is promising chemical
which has a wide range of applications
in many industries.

4. Background 03
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5. Background 04
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- SA can be used as a precursor for synthesis
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biodegradable plastic such as polybutyrate
succinate (PBS) and polyamide to produce
bioplastics.

6. Background 05
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-Thailand is a large producer of raw material
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to produce SA such as casava and sugar
cane
- Therefore, the country has high potential for
development of SA production in order to
support bioplastic industry.

7. Background 06
- Commercial scale production of SA is
petroleum-derived.
-However, Fermentative production of SA
from renewable resource has received
increasing interest .
- SA can be produced by microbial
fermentation using various microorganism.

8. Background 07
-SA is a common intermediate in the metabolic
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path way of several anaerobic and facultative
microorganism .
-Those organisms suitable for the efficient
production of SA can be categorized into
bacteria.

9. Background 08
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10. Background 09
- Many different microorganism have been
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screened and studied for SA such as ;
- Actinobacillus succinogenes,
- Anaerobiospirillum succiniciproducens,
- Mannheimia succiniciproducens and
- Recombinant Escherichia coli.

11. Background 10
-Bio-based SA is potential substitute for
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the current petroleum-based process.
-However, compared with petrochemical-
derived SA , biological SA production is still
not economically competitive .
-Because of its major draw backs:
low product concentration in fermentation
broth and difficult product recovery.

12. Background 11
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-In addition, product inhibition is important
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factor for bioprocess development .
-The product inhibition effect, which can be
described by specific growth rate equation
ai
 S   P 
v = v max  '  ∏ 1 − ' 
 S (
 K + S + S2 /K'
i ) 
 

 P 
m 


13. Background 12
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- The cost of downstream processing can
make a very high Template total production
portion in the
cost, more than 50% .
- It is necessary to reduce the production cost
of fermentation process in order to increase
its competitiveness with that of petrochemical
process.

14. Background 13
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-The separation methods studied for SA
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recovery mainly include ;
-Direct crystallization,
-Precipitation,
-Membrane separation,
-Extraction,
-Chromatography,
-In situ separation.
.

15. Background 14
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16. Background 15
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-Formation of byproduct is a major problem
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that has to be solved because it reduces
SA yield and productivity, while increases
the complexity and cost of SA recovery .
-In situ product removal or extractive
fermentation can be solved the
aforementioned problem.
.

17. Background 16
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- The advantage of in situ process
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separation that can be improved the final
SA
. concentration and reduces end-product
inhibition.
-Precipitation process is a very interesting
technique due to its simplicity, and no does
not require additional chemical or unit
operation.

18. Background 17
-This work, fermentation of SA will used
commercial strain of SA producing bacteria
and using precipitation technique couple with
fermentation SA .
-Calcium carbonate will be used to control the
pH of the broth.

19. Background 18
- Submerge ceramic microfiltration membrane
will be employed for take out the clarified broth
prior to precipitation.
-As a result, Calcium succinate will be
precipitate out , the remaining will then be
re-circulated back into the fermenter.
-The precipitate will be harvested ,washed
and dried to produce a high purity calcium
succinate

20. Research Objectives 19
-To study the precipitation process of
calcium succinate by using synthetic
solution.
-To compare SA production performance in
term of yield and volumetric productivity
between this work and conventional batch
fermentation.

21. Research Objectives 20
-To obtain a high purity of calcium
succinate powder which facilitates
subsequent downstream processing.

22. Research Hypothesis 21
- SA can be produced by using
Actinobacillus succinogenes ATCC
55618.
- Precipitation of calcium succinate can
be achieved in the precipitation tank.
- Product inhibition can be minimized and
SA production is significantly increased.

23. Materials 22
-Actinobacillus succinogenes
ATCC 55618
- Glucose - Calcium carbonate
- Autoclave - 5 L Bioreactor
- HPLC - Glucose analyzer

24. Methods 23
- Fermentation will be carried out in the absence of oxygen by
flushing CO2 gas at the beginning of fermentation

25. Methods 24

26. Methods/ Batch fermentation 25

27. Methods/ Fed batch fermentation 26
- Fed batch fermentation will be started with
1.2 L of culture medium and the concentrated
culture medium will be gradually added.
- pH of broth will be controlled at 6.8
- Batch fermentation will be carried out first,
feeding of the concentrated culture medium will be
started when amount of glucose is nearly depleted
or lower than 20 g.L-1.

28. Experimental setup 27
Experimental setup for extractive fermentation
of SA using external precipitation method.
Calcium succinate

29. Analytical procedure 28
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30. Analytical procedure 29
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-Reducing sugar will be measured by
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Dinitrosalicylic (DNS) method according
to Miller (1995)
- Cell concentration will be determined by
using UV–VIS Spectrometer. Cell viability
will be accomplished using fluorescence
microscopy.
- Quantitative analysis of organic acids will
be analyzed by HPLC .

31. Expected results 30
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1.High yield and volumetric productivity
of succinic acid will be achieved using
the external precipitation method.
2.This simple high efficiency method
can be applied to pilot scale production
of succinic acid.

32. Research plan
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33. WINTER
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THANK YOU VERY MUCH
Jiraphorn Lubsungnoen M5430116