Microbial Production of

Oils and Fats

Course: Food Microbiology 2014
Lecturer: Dr. Vu Thi Lam An
Group 5: Nguyen Quynh Hai Yen BTFTIU11009
Ho Quang Loc BTFTIU12011
 Contents
1. Introduction to Single Cell Oils (SCO)

2. SCO production process

3. Applications

4. The significance and safetyness of SCO

1. Introduction to Single Cell Oils.
» What is it?

» Condition for lipid accumulation

» Principles of lipid accumulation

 What is Single Cell Oils?
• Single Cell Proteins?

• Single Cell Oils (SCO): triglyceride fats generated by

microorganisms.
– Lipid components are similar to those found in plant and animal.

→ Applications: animal feeds, aqua feeds, biodiesel.

What is Single Cell Oils?
Is it safe for human consumption?
• Microorganism → a human food source from
ancient time.
– Can you name some?
 Oleaginous microorganism
• Microorganism that gathers >20-25% of their
biomass as oil.

• Many yeast, fungi and several algae are maximum

producers.
– Algae → biodiesel
– Yeast and fungi → edible oil
• They are also capable of producing high levels of
nutritionally important polyunsaturated fatty acids
(PUFAs).
 Advantages of Oleaginous
microorganism

• Growth on various substrates

→ utilizing by-product = reduce cost

• Ability to synthesize a divert

array of fatty products
→ many useful applications.

• Able to be genetic manipulation

→ selection for highest productivity.
 Condition for lipid accumulation
• Lipid accumulation is trigger when
– Nutrients in the growth medium (usually Nitrogen
source) is exhausted
– Surplus of Carbon source (usually glucose)

→ Cells stop multiplying.

→ Cells convert Carbon source to Storage oils or fats.
Condition for lipid accumulation
Stages of Lipid accumulation
in Batch culture
Prepare medium: high Carbon, low Nitrogen

Microorganisms grow and multiply until a certain time

Exhaustion of N, cells stop divide

Cells convert C → storage lipid (intracellular)

Lipid accumulation → cells expand

Cells reach limit of obesity → stop accumulating.

...
 Biochemistry of Lipid accumulation

TCA

Cellular respiration of microorganism

Biochemistry of Lipid accumulation
Exhaustion of N

[AMP] is diminished;
Isocitrate dehydrogenase in mitochondrion stop working

TCA is disrupted

Citric acid in mitochondrion → cytosol

Enzyme ATP-citrate lyase: Citric acid → acetyl-CoA

(Oleaginous microorganisms only)

Acetyl-CoA →(fatty acid biosynthesis)→ triacylglycerols

Biochemistry of Lipid accumulation
2. Production of Commercial SCO.
 Production of commercial oils and fats
from SCO
• Only if cost of production of SCO could compete against
plant oil.
• In fact, production cost of SCO is much higher.

Plant oil = 400-

800$/ton

- 4 tons of sugar (300$ each)

→ 1 ton SCO + biomass
- Cost of extraction and refining
→ food graded oil
- Cost of selling, waste disposal...
 Production of commercial oils and fats
from SCO

• High production cost

→ Apply for production of specialty oils and fats only.

→ Choose fast growing microorganisms, high oil yield, simple

growth requirements.

→ Focusing on the use of by-products of agriculture and industry

as substrates
 Production Process

Harvesting Drying the

Cultivation Extraction
the cells cells
Production Process
 Commercial Production Process
• It is important to harvest SCO before Carbon substrate is
completely depleted.
• As the cells will use the lipid that they have accumulated for
their own survival.

• Consequences:
1. Low Oil yield
2. Cells forming lipase → degrade quality of SCO
• Alter flavor, color, functional characteristics.
– How to avoid lipase?
→ Answer: heat denaturation <60ᴼC
 Commercial Production Process
• SCO usually contain natural antioxidant
→ A clear, bright oil with a minimum of coloring → favorable.

→ Very stable to oxidation SCO by Mortierella alpina

→ Good for consumption

SCO by Cryptococcus curvatus

3. Applications of Single Cell Oils.
» Cocoa butter equivalent fat

» Polyunsaturated fatty acids (PUFAs)

• DHA

• ARA
 Cocoa Butter Equivalent Fat
• Cocoa Butter: triacylglycerol, consists of:
- palmitic acid (16:0)
- oleic acid (18:1)
- stearic acid (18:0)
 Cocoa Butter Equivalent Fat

• Cocoa Butter: very expensive

• SCO = Cocoa Butter Equivalent Fat = cheaper substitute

• However, SCO are low in stearic acid

→ genetic modification is necessary

Cacao Butter Substitutes

 Cocoa Butter Equivalent Fat
* Henk Smit (Netherlands):
– Using the yeast Cryptococcus curvatus D
– Delete the gene that codes for the Δ9−desaturase, which converts
stearate into oleate
→ Fail, not stable products

* Julian Davies (New Zealand)

– Culturing in O2 deficiency → stop enzyme desaturase from working →
more stearate
→ But uneconomical

* Chance for you!

 Polyunsaturated fatty acids
• Omega-3 and omega-6: cannot synthesize by
human body.
– Omega-6: from plant food.
– Omega-3 (EPA, DHA):
• Are long chain PUFAs
• Most plants are unable to synthesize
• Must obtain from animal food.
 Dietetic significant of PUFAs
• Long chain PUFAs:
– Key components of cell
membrane
– Inflammation response

• Arachidonic acid (n-6)

and DHA (n-3):
essential for brain and
neural development

• In general, our diet

contains too much n-6
and too little n-3
 SCO as source of Omega-3
• Animal source of Omega-3:
– Oily fish → distinct fishy taste
– Animal organs (esp. liver)
→ Unfavorable + shortage of supply + extensive purification
→ Costly
→ Why not SCO?
• In this presentation, we will also briefly introduce to
you:
– Arachidonic Acid (ARA) production process.
– Docosahexaenoic acid (DHA) production process.
Processes for the production of various PUFAs
Arachidonic Acid (ARA)

Arachidonic Acid is a long chain

polyunsaturated fatty acid with twenty carbon
atoms and four double bonds. Its systematic
name is (all-cis)- 5,8,11,14-eicosatetraenoic
acid(ETA)
Processes for the production of various PUFAs
Arachidonic Acid (ARA)

Biosynthetic pathway of polyunsaturated fatty

acids in Mortierella alpina

ARA is synthesized through

many enzymatic steps of fatty acid synthesis,
elongation of fatty acid and desaturation of
fatty acid
 Arachidonic Acid (ARA)

Microorganism ARA Scale The fatty acid

yield/cultivation composition
period of 50 Mortierella
Submerged culture subgenus isolates
Mortierella alpina 1S-4 13 g/L/10d 10-kL fermentor was analyzed Amano
et al
M. alpina ATCC 32222 11 g/L/11d 250-mL flask
M. alpina ATCC 32221 11 g/L/16d 500-L fermentor They found that ARA
M. alpina UW-1 5.5 g/L/6d 20-L fermentor composition of alpina
M. alpina LPM 301 4.5 g/L/8d 30-L fermentor was higher than
those of any
Mortierella alliacea YN-15 7.1 g/L/6d 50-L fermentor
Mortierella schmuckeri S12 2.3 g/L/3d 14-L fermentor
Mortierella sp. S-17 0.96 g/L/7d 1-L flask
Mortierella elongata SC-208 0.49 g/L/5d 250-mL flask
Pythium irregulae ATCC 3.1 g/L/8d 250-mL flask
10951
Solid-state culture
M. alpina IFO 8568 13 g/kg-medium/20d
M. alpina CCF 185 36 g/kg-medium/21d
Processes for the production of various PUFAs
Docosahexaenoic acid (DHA)

Docosahexaenoic acid (DHA,

22:6) is a particularly important
ω ‐3
PUFA, with a 22‐carbon chain
and six double bonds.
Processes for the production of various PUFAs
Docosahexaenoic acid (DHA)
 Microbial PUFAs on market

ARA for muscle

ARA-rich SCO and DHA-
stimulation
rich SCO in infant
formulae in many
countries of Europe,
Australia, and the Far East
Advantages and Disadvantages of SCO
ADVANTAGES:
1. Simple fatty acid profile → easy for extraction and
purification.

2. Both quality and quantity of the product can be guaranteed.

3. SCO is not affected by geographical or environmetal factors

(e.g pollution etc.)

4. Rapid production: increase cells in a short time and

independence from climatic conditions.

5. SCO act as a novel source of polyunsaturated fatty acids,

such as DHA and AA for nutritional supplementation.
Advantages and Disadvantages of SCO
DISADVANTAGES:

1. Limited production capacity.

2. Consumer acceptance.
3. DHA from fish oil are linked to growth retardation in infant
→ also DHA of SCO
 Understanding 2 routes for safety
evaluation of a food ingredient

1. Petition for FDA approval as a food additive.

– FDA will review before products are available on the market.
– FDA is responsible for their decision.

2. Manufacturer can determine that a substance is GRAS if

there is scientific consensus among qualified experts
about its safety under the intended condition of use.

→ SCO is only FDA approval.

 Safety of SCO
• SCO is only FDA approval. Why?
→ For some years, high level of SCO-ARA in the human diet was
thought to induce potential adverse effects on blood
clotting.

• However, later intensive and prolong studies demonstrate

the lack of evidence of toxicity in volunteers.

• Recently, microbial oils have been shown to be no more toxic

than oils from traditional sources.
• More researches are needed!
 Summary
• SCO are now used to produce high value oils rich in
arachidonic acid (ARA) and docosahexaenoic acid
(DHA).
• These oils are used in infant nutrition and also as
nutriceuticals and food additives appropriate for adults.
• Potential therapeutic applications are very promising.

• Production of SCO:
– SCO can be produced in large scale.
– To ensure stabilization of SCO, must harvest the cells before
substrates are totally depleted.
– SCO extraction is much the same as used for plant seeds.
 References
• James, P.W., Colin, R. Microbial production of fats and oils. Food Biotechnology.
2006, pp. 443-472

• http://www.ukessays.com/essays/education/microbial-production-of-fats-and-
oils.php
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