Steps for Commercial Production of Antibodies,Inoculum and Media Preparation,Solid Surfaces,Liquid Culture,Source of Energy,Carbon

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Home >> Industrial and Microbial Biotechnology >> Microbes and Microbial Genomics for Industry >> Steps of Commercial Production of Antibodies

Steps for Commercial Production of Antibiotics
Inoculum and media preparations. A small sample of the cultured stock is taken and the amount of material is amplified by growth on solid surfaces or in liquid culture. Spores of the micro-organism are collected and retained in a suspension form. This suspension is transferred into a container suitable for attachment to the fermenter.

Different media are required at different stages of antibiotic production.

A typical Antibiotic Fermentation Plant

A Typical Antibiotic Fermentation Plant

1. Motor	2. Motor	3. Motor	4. Motor
5. Motor	6. Batching vessel	7. Feed vessel	8. Feed vessel
9. Feed vessel	10.Seed stage fermenter	11. Motor	12. Final stage fermenter
13. Air compressor	14. Batch sterilizer	15.Steam-in	16. Steam-in
17. Motor	18. Rotary drum filter	19. Belt Filter	20. Steam-out
21. Steam-out	22. Cooling tower	23. Centrifuge

Manufactures spend considerable amount of time and money in developing media both for increasing antibiotic productivity and for decreasing costs. The medium must supply to the organism, source of energy, carbon, nitrogen, phosphate, trace elements and in some cases specific precursors for biosynthesis of antibiotics (e.g. phenyl acetic acid for the production of penicillin G). Composition of a typical medium used in the final stage of penicillin production is shown in the table. In many cases, small amounts of yeast extracts or similar substances are added to increase the rate of fermentation.

The fermenter. A typical antibiotic fermentation plant is illustrated in the diagram. The medium may be batched (made up) and sterilized in the fermenter itself. However, most of the fermenters are attached to a batching vessel and to sterilizers.

Composition of fermentation medium for the production of penicillin.

Components	Per cent (approximate, w/v)
Sugar (e.g. glucose/sucrose/lactose)	5 – 10
Maize steep liquor	5 – 9
Pharma media	3 – 5
Chalk	0 – 1.0
(NH4)2S04	0 – 1.0
Phosphate	0 – 0.5
Edible oil	0.3 – 0.8
Phenyl (or phenoxy) acetic acid	03 – 0.6

The feed vessels connected to the fermenter are used to supply nutrients or precursors during fermentation process. The seed and the final stage fermenters must be capable of operating under aseptic conditions. The fermenter vessels are usually stainless steel containers with a capacity of 30 to 300 m3. Agitation of the broth is facilitated via two or three flat bedded impellers. Air which is sterile, filtered and compressed is passed into the vessel immediately below the bottom impeller. Heat generated by the agitation system and by the metabolic activities of the micro-organisms is dissipated by using cooling water columns, since optimum temperature for most of the antibiotic producing organisms is 24-28°C. For continuous sterilization of the system, steam is used. The steam delivery system is fitted in such a way that only the desired section of the system can be sterilized.

The production stage
All antibiotics are produced by batch or fed batch fermentation methods. Fed batch techniques are used to extend the duration and productivity of fermentation. By continuous addition of glucose or other sugars, an antibiotic can be produced with the same system for a period extending up to two weeks. Addition of sugars and of other nutrients (if required) may be achieved by metering pumps, flow meters, or burettes. Sometimes, considerable amount of foam is generated during fermentation, specially when the media are proteinaceous and they are vigorously agitated and aerated. The foam reduces the vessel capacity and, therefore, it is undesirable. To prevent it, often anti-foam chemicals such as vegetable oils are added in the media. The fermentation is generally carried out at 24-28°C.

Recovery of antibiotic
A variety of physico-chemical methods are used for the removal of antibiotic from the fermenter. When penicillin is the antibiotic, most of it (about 99%) is excreted out by the fungus into the medium. In such a case, the mycelium is separated by filtration and the antibiotic is recovered from the medium by two stage continuous counter-current extraction of the fermenter broth with amyl or butyl acetate. Streptomycin and many other amino glycoside antibiotics are also secreted out and are removed by ion exchange columns. Cell bound amino glycosides such as gentamycin, sisamicin and fortimicin are released from the cell by acidification of the culture broth with H2SO4 to achieve a pH of 2.0-2.5.

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