This document discusses the production of vitamin B12 through fermentation using Streptomyces olivaceus bacteria. It provides details on the history and structure of vitamin B12, as well as its sources, biosynthesis, functions, and fermentation process. The fermentation process involves preparing an inoculum and production medium, maintaining optimal aeration, temperature, and pH levels during fermentation, and recovering the vitamin B12 through purification and crystallization techniques. Vitamin B12 production yields using this method are typically between 1-2 mg per liter of fermented broth.

1. INDUSTRIAL MICROBIOLOGY
SRI PARAMAKALYANI COLLEGE
DR . VISHWANATHAN.S., Ph.D,
Head of the department,
Sri paramakalyani college ,
Alwarkuruchi.
PETCHIAMMAL .G,
II MSC Microbiology,
Sri paramakalyani college,
Alwarkuruchi.
REACCREDIATED WITH A+ GRADE WITH CGPA OF 3.39 IN THE III CYCLE BY NACC
AFFILIATED TO MANONMANIUM SUNDHARANAR UNIVERSITY, THIRUNELVELI.
ALWARKURICHI - 627 412,TAMILNADU, INDIA.
POST GRADUATE & RESEARCH CENTER - DEPARMENT OF MICROBIOLOGY
( GOVERNMENT AIDED)
IV SEM CORE
PRODUCTION OF VITAMIN B12

2. SYNOPSIS
① INTRODUCTION
② HISTORY OF VITAMIN B12
③ STRUCTURE OF VITAMIN B12
④ TYPES
⑤ SOURCES
⑥ BIOSYNTHESIS
⑦ FERMENTATIVE PRODUCTION
VITAMIN B12
⑧ FUNCTIONS OF VITAMIN B12
⑨ REFERENCE

3. VITAMINS
 Organic compounds,essential nutrient that are organisms
requires in limitted amount.
 Play a vital role in the growth and metabolism.
 13 vitamins are universally recognised at present by
biological and chemical activity.
 Classified as, Fat soluble and Water soluble vitamins.
 Casimir Funk - Father of vitamin Therapy

6. INTRODUCTION
 Vitamin B12, also called as cobalamin, is synthesized in nature exclusively by
microorganisms and it is required to animals for growth and metabolism.
 It is present in every animal tissue in a very low concentration (1 ppm in the liver cells).
 It remains in the form of acoenzyme (adenosyl or methyl cobalmine) in the animal tissue.
 Animals get the required vitamin B12 through feed or by absorption of it from the intestinal
wall as it is produced in the intestine by the intestinal microorganisms.
 However, human beings obtain vitamin B12 only from food, since the vitamin B12 secreted
in the intestine by microorganisms cannot be assimilated.
 The daily requirement of human beings is 0.001 mg.
 Helps in the utilization of vegetable proteins.
 used to treat pernicious anaemia.

7. HISTORICAL MILESTONE

8. STRUCTURE OF COBALAMINE

9.  Pure cyanocobalamin
possess deep pink
colour associated with
most octahedral cobalt
complexexs.
 It is used to treat
pernicious
anaemia ,vitamin B12
deficiency.
TYPES
 Methylcobalamine
(mecobalamine,mecbl or
meB12,)
CYANOCOBALAMINE a form
of vitamin B12.
 It differs from
cyanocobalamin ,in that the
cyanide group is replaced by
methyl group.
 It is used in the treatment of
pheripheral neurophathy.
 Sometimes.denoted by vit
amin B12,it has an avid
affinity for cyanide ion
 Has been use antitode for
cyanide poisoning.
 It is supplied typically in
water solution for injection
METHYLCOBALAMINE

10. SOURCES

11. BIOSYNTHESIS

12. FUNCTIONS OF VITAMIN B12

13. FERMENTATION PROCESSING

14. FERMENTATIVE PRODUCTION OF VITAMIN B12

15. MICROBES INVOLVED IN THE PRODUCTION OF VITAMIN B12
Microorganisms Yield ( mg / l )
Bacillus megaterium 0.51
Streptomyces olivaceaus 3.31
Butyribacterium rettgeri 5.0
Microspora spp 11.5
Propionibacterium feeudenerichii 19.0
Propio bacterium shermani 35.0
Pseudomonas denitrification 60.0
Rho pseudomonas protamicus 135.0

16. VITAMIN B12, production by streptomyces olivaceeus

17. • A pure agar slant culture of Streptomyces olivaces NRRL B-1125 is inoculated and cultured in 100 to
250 ml of inoculum medium housed in Erlenmeyer flasks during the establishment of the inoculum.
• During incubation, these flasks should be placed on the platform of a mechanical shaker to aerate the
media.
• The flask cultures are then used to inoculate greater quantities of inoculum media stored in a
succession of inoculum tanks.
• Typically, two or three subsequent transfers are necessary to obtain the necessary quantity of
inoculum culture.
• It has been demonstrated that inoculating the production tank with approximately 5% of the production
medium volume is adequate.
PREPARATION OF INOCULUM

18. PREPARATION OF INOCULUM

19. • Typical production media for this fermentation include carbohydrates, proteinaceous material, a
cobalt supply, and other salts.
• In Table, a typical production medium is provided. Under optimal conditions, the majority of
distillers’ solubles, soya bean meal, yeast, casein, etc. are deemed good.
• Cobalt must be added to the medium in order to maximise cobalamin yields.
• Cobalt has nothing to do with the growth of S. olivaceus, it should be mentioned.
• In certain instances, it is necessary to add cyanide to the medium in order to facilitate the
conversion of other cobalamins to vitamin B12.
• By- The medium can be sterilised in batches or in a continuous manner.
PRODUCTION MEDIUM

20. PRODUCTION MEDIUM
• In batchwise sterilisation, the production tank’s medium is heated at 250°F for one hour.
• In the latter method, the production tank is charged (e.g., at 330 degrees Fahrenheit for thirteen
minutes) by mixing it directly with live steam.
• Steam is blown into apertures during sterilisation, and all transfer lines are filled with live steam
while not in use to ensure sterility.

21. CARBON & NITROGEN SOURCES
NITROGEN SOURCES
• Ammonium phosphate
• Ammonium Hydroxide

22. • The streptomycete strain’s growth rate is dependent on
the aeration and agitation rates.
• Greater than optimal aeration rates result in excessive
foaming.
• The optimal aeration rate is around 0.5 volume
air/volume medium per minute.
• Activated charcoal is typically used to sterilise air by
passing it through column.
AERATION AND AGITATION

23. • Foam formation is a significant issue with this fermentation, particularly at the beginning and end of
the process.
• There are numerous antifoaming compounds that can be used to prevent the creation of foam.
• Soybean oil, corn oil, lard oil, and silicones are significant defaming agents.
• Depending on the need, a defaming agent in its sterile form is given to the medium during foaming.
• In some circumstances, an antifoaming agent is included during the production medium formulation
process.
ANTIFOAM AGENTS

24. • Sterility must be maintained until the fermentation is complete, as contamination invariably
results in extremely low yields.
• All equipment must therefore be sterile.
• Moreover, transfers are conducted under aseptic circumstances.
PREVENTION OF CONTAMINATION
Yields
The yields of cobalamin in
fermented broth are
typically between 1 and 2
mg per litre.

25. • During fermentation, a temperature of 80°F in the production tanks is acceptable.
• During the first 24 hours of fermentation, there is a pH drop of a few tenths of a unit and a
rapid consumption of sugar.
• After two to four days, mycelium lysis commences, resulting in an increase in pH.
• To stabilise the mash, sulfuric acid is used to lower the pH to about 5 and a little amount of a
reducing agent is added (e.g. sodium sulphite).
TEMPERATURE
pH

26. PRODUTION OF VITAMIN B12

27. RECOVERY
Purification by
adsorption ,
clarification,filteration
Chromatography
(Al2O3,XAD)
Crystallization

28.  Food preservative
 Cofactor
 Protective medicine
USES

29. RECOMMENTTED DIEATRY ALLOWANCE

30. BIBLIOGRAPHY

31. SKILLS GAINED BY SEMINAR

34. Thank you !
SRI PARAMAKALYANI COLLEGE
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