Download essential amino acid

Amino acid
Productions
Amino acid
 Amino Acids Are Molecules Containing An Amine Group, A Carboxylic
Acid Group And A Side-chain That Varies Between Different Amino
Acids. The Key Elements Of An Amino Acid Are Carbon , Hydrogen ,
Oxygen , And Nitrogen An Alpha-amino Acid Has The Generic
Formula H2NCHRCOOH, Where R Is An Organic Substituent. Amino
Acids Are Critical To Life, And Have Many Functions In Metabolism.
 One Particularly Important Function Is To Serve As The Building Blocks
Of Proteins , Which Are Linear Chains Of Amino Acids. Amino Acids
Can Be Linked Together In Varying Sequences To Form A Vast Variety
Of Proteins.
Essential and Nonessential AAs
Histidine Alanine Isoleucine Arginine *
Leucine Asparagine Lysine Aspartic acid
Methionine Cysteine * Phenylalanine
Glutamic acid Threonine Glutamine *
Tryptophan Glycine Valine Ornithine *
Proline * Selenocysteine * Serine * Taurine
* Tyrosine * (*) Essential only in certain
cases
EAAs
• An essential amino acid or indispensable
amino acid is an amino acid that cannot be
synthesized de novo by the organism being
considered, and therefore must be supplied in
its diet. The nine amino acids humans cannot
synthesize are phenylalanine, valine,
threonine, tryptophan, methionine, leucine,
isoleucine, lysine, and histidine.
Non EAAs
• Six amino acids are considered conditionally
essential in the human diet, meaning their
synthesis can be limited under special
pathophysiological conditions, such as
prematurity in the infant or individuals in severe
catabolic distress. These six are arginine,
cysteine, glycine, glutamine, proline and
tyrosine .
• Five amino acids are dispensable in humans,
meaning they can be synthesized in the body.
These five are alanine, aspartic acid,
asparagine, glutamic acid and serine.
Amino Acid by Fermentation
• NOW A DAYS THREE MAJOR AMINO ACIDS ARE
BEING PRODUCED ON LARGE SCALE.
GLUTAMIC ACID
LYSINE
METHIONINE
• All PRODUCED BY PROCESS OF FERMENTATION
USED TO BE PRODUCED BY CHEMICAL
SYNTHESIS
Why Produced on a Large Scale?
• The amino acid business is a multi-billion dollar
enterprise, All twenty amino acids are sold,
although each in greatly different quantities
• Amino acids are used as animal feed additives
(lysine, methionine , threonine ), flavor enhancers
(monosodium glutamic, serine, aspartic acid) and
as specialty nutrients in the medical field.
• Glutamic acid, lysine and methionine account for
the majority, by weight, of amino acids sold .
• GLUTAMATE IS MEDICALY USED AS A
NEUROTRANSMITTER.
Glutamic Acid Production
• Discovered GLUTAMIC ACID (L-glutamate) after acid hydrolysis
and fractionation of kelp and neutralization with caustic soda.
• These treatments enhance the taste of kelp
• Gave rise to the birth of:
MONO SODIUM GLUTAMATE (MSG), flavor enhancing compound.
It was extracted from soy and wheat. Now micro-organisms
(Corynebacterium glutamicum) are used for MSG production.
Commercial production of MSG is the largest and biggest industries
world over.
Commercial Production
Glutamic acid > lysine > methionine > threonine > Aspartic acid
The market is growing steadily by about 5–10% per year.
USES OF AMINO ACIDS IN INDUSTRIAL APPLICATIONS
Food industry: 65%
Feed Additives: 30%
Pharmaceutical: 5%
FOOD INDUSTRY
• Flavor enhancers, MSG, glycine, alanine. Tryptophan and histidine
act as antioxidants to preserve milk powder. For fruit juices
cysteine is used as an antioxidant.
• Aspartame, dipeptide (aspartyl-phenylalanine-methyl ester)
produced by combination of asp and Phe is 200 times sweeter
than sucrose. Used as low calorie artificial sweetener in soft
drink industry
• Essential amino acids are those deficient in plant based foods
like lys, met, thr, Trp improves nutritional quality of food and
feed additives (animal). Bread: lysine, soy products or soyabean
meal (pigs/animals): methionine
PHARMACEUTICAL INDUSTRY
Used as medicines, infusions to patients with post operative treatment
CHEMICAL INDUSTRY
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Used as a precursor for production of several cpds
Glycine used to manufacture GLYPHOSATE
Threonine used for AZTHREONAM (herbicide)
Poly methyl glutamate: manufac. Of synthetic leather
N-acyl derivatives of amino acids used for making cosmetics
METHODS FOR PRODUCTION OF AMINO ACIDS
1.
EXTRACTION: hydrolysis of proteins to isolate amino acids like
cys, tyr, leu
2. CHEMICAL SYNTHESIS: can result in racemic mixture (D and L
amino acids), most applications are for L-form sometime DL or D
maybe required.
3. MICROBIOLOGICAL SYNTHESIS
a. Direct fermentation: MO use carbon sources and produce aa.
Carbon like glu, fructose, alkanes, ethanol, glycerol, molasses,
starch, methanol etc.
b. Conversion of metabolic intermediates to amino acids:
c. Use of enzymes (microbial) or immobilized cells: resting cells,
crude cell extracts, immobilized cells can be used.
STRAIN IMPROVEMENT METHODS FOR AA PRODUCTION
Because of regulatory control of metabolic reactions natural over
production is rare
Regulatory control has to be removed
Mutagenesis and screening for mutants are done
1. Auxotrophic mutants: lack of formation of regulatory end
product (repressor or effector molecule). Intermediates
accumulate and get excreted.
2. Genetic recombination: for overproduction (recombinant
molecules created) or protoplast fusion to develop hybrids
3. Recombinant DNA Technology: gene cloning, gene engineering
4. Functional genomics: whole chromosome sequencing data
L-GLUTAMIC ACID
Corynebacterium glutamicum, is a short, aerobic, Gram-positive
rod capable of growing on a simple mineral salt medium with
glucose, provided that biotin is also added.
Production of L-glutamic acid by C. glutamicum is maximal at a
critical biotin concentration of 0.5 mg g-1 of dry cells, which is
suboptimal for growth
Detergents like Tween-40,
addition of penicillin,
use of Glucose,
Glucose-6P,
CO2,
fatty acid auxotrophic strains, or
addition of ethambutol- inhibiting
synthesis.
arabinogalactan
L-GLUTAMIC ACID
Regulatory control:
Good supply of glucose and efficient conversion of phosphoenol
pyruvate to oxaloacetate
Phosphoenol pyruvate carboxylase and pyruvate carboxylase, pyruvate
dehydrogenase
a-ketoglutarate dehydrogenase (low activity by adding penicillin,
surfactants)
Glutamate dehydrogenase (high activity)
1 mole of glucose should produce 1 mole of glu
In practice, efficiency is 70%
• Glu is synthesized intracellularly
• Carrier mediated processess
• Biotin is essential co factor (for Acetyl CoA carboxylase),
deficiency of biotin affects fatty acid biosynthesis, membrane
formation alters, permeability is affected and intracellular
export of glu is altered
FACTORS INFLUENCING PRODUCTION
1.
2.
3.
4.
Carbon sources
Nitrogen source: ammonia for carbon to glu pH control
Growth factors :biotin
O2 supply: high conc inhibits growth and low O2 leads to lactic
acid production and succinic acid, Afftects Glu production in both
cases
Nutrients
Glucose (12%)
Dissolving tank
38oC
30-35h
Sterlizer
Buffer tank
Inoculum
Sterile air
FERMENTER
Cell separator
Ammonia, pH control (7.8)
(ammonium acetate 0.5%)
Eluted in NaOH
Anion exchanger
Evaporation
Crystallization
100g/L
Bakteri asam laktat juga memproduksi asam format, asetat,
propionat, kaproat, kaprilat, kaprat, butirat, dan isovalerat dari
metabolisme fermentasi dan transformasi enzimatik asam amino.
Protease merupakan enzim yang mendegradasi protein. Protease
tergolong enzim hidrolase yang mengkatalisis reaksi degradasi
substrat dengan pengolahan air. Pengertian protease menurut
Winarno (1985) ada dua yaitu proteinase yang mengkatalisis
reaksi hidrolisis molekul protein menjadi fragmen besar polipeptida,
dan peptidase yang menghidrolisis fragmen besar polipeptida
menjadi asam amino.
Kasein pada yogurt sebagai substrat untuk uji aktivitas protease
dengan metode Lowry. Kasein akan terhidrolisis menjadi peptida
dan asam amino dengan bantuan air oleh enzim protease.