HISTORY
Sir Alexander Fleming (1881-1955), a British bacteriologist observed that on a bacterial culture plate which had become contaminated by a mould, a bacterial growth in the vicinity of the mould colony was inhibited. He carried on his experiments at St. Mary's Hospital, London in 1928. His results were published in the Journal of Experimental Pathology in 1929. Since the mould was a strain of Penicillin notatum, he named the antibacterial substance , Penicillin. A decade later, two other English men, Chain and Florey engaged themselves in furthering Fleming's investigations. Thus, an antibiotic preparation was developed for human use.
Source:
From Penicillium chrysogenum (ancestral fungus), high yielding strains have been developed for the commercial production of penicillin.
Mode of action of enzyme:
Penicillin inhibits the formation of peptidoglycan cross links in the bacterial cell wall, which is achieved through binding of the 4 membered beta- lactum ring of penicillin to the enzyme DD- transpeptidase. As a consequence, DD- transpeptidase cannot catalyze formation of these cross-links cell walls without intact peptidoglycan cross- links are structurally weak, prone to collapse and disintegrate when the bacteria attempts to divide.
Type of fermentation:
It is a fed batch process that is carried out aseptically in stainless steel tank reactors with a capacity of 30-100 thousand gallon. The fermentation involves 2-3 initial seed growth phases, followed by a fermentation production phase with a duration ranging from 120-200 hours.
Production parameters:
High yielding strains of P. chrysogenum are genetically unstable. Hence, production strains are stored in a dormant form by either of the following techniques:
⦁ Spore suspensions can be lyophilized in appropriate media
⦁ Spore suspension can be stored under liquid nitrogen i.e., in a frozen state.
All manipulations are carried out in laminar flow cabinets in rooms which contains the filtered air.
Personnel wear sterilized clothing and they are careful in their aseptic technique.
There is strict necessity for pure culture in antibiotic fermentations.
Inoculum preparation: the aim is to develop a pure inoculum in sufficient volume and in the fast growing phase for the production fermenter. The time taken for each stage is measured in days, and it decreases as the sequence progresses.
⦁ The medium is designed to provide the organism with all the nutrients that it requires.
⦁ The medium can be solidified by increasing the amount of agar.
⦁ Adequate oxygen is supplied in the form of sterile air.
⦁ Temperature (24 C) is controlled.
Principle criteria for transfer to next stage in progression are:
Freedom from contamination
Growth to a predetermined cell density.
The volume of culture increases approximately 10 fold with each successive stage. The mould, P. chrysogenum grows in filamentous form.
Primary source of spores stored on soil - One or more growth stages on solid media - one or more growth stages in shaken flask culture- 0.5-1.0 m3 seed storage - 10-20 m3 seed storage- 125- 250 m3 production stage
Inoculation methods: Any one of the following methods may be used to inoculate the fermentation medium in the submerged culture production of penicillin:
⦁ Dry spores may be used to seed the fermentation medium
⦁ Inoculation by a suspension of ungerminated mould spores
⦁ The fermentation medium may be seeded by pellet inoculation.
Raw materials: three points should be kept in mind in choosing raw materials for the manufacture of penicillin.
⦁ An abundant growth of mycelium.
⦁ Maximum accumulation of penicillin
⦁ Ease of extraction and purification of antibiotic.
⦁ Carbon sources: lactose, glucose or sucrose
⦁ Nitrogen sources: ammonium sulphate, ammonium acetate, ammonium lactate, corn steep liquor.
⦁ Mineral sources: potassium, phosphorus, magnesium, sulphur, zinc and copper.
⦁ Precursor: phenylacetic acid (PAA).
Component. Percent
Corn steep liquor solid. 3.5
Lactose. 3.5
Glucose. 1.0
Calcium carbonate. 1.0
Potassium dihydrogen phosphate 0.4
Edible oil. 0.25
Penicillin precursor with slow feeding rate
DOWNSTREAM PROCESSING (Extraction and Purification)
⦁ Removal of mycelium: in the first step, mycelium is removed by filtration using a rotary vacuum filter.
⦁ Countercurrent solvent extraction of penicillin: the pH of the filtrate is adjusted to 2-2.5 with the help of phosphoric acid. The antibiotic is then extracted back into an aqueous buffer at pH 7-7.5 through countercurrent solvent extractor. The partition coefficient shifts in the favor of aqueous phase. The resulting solution is again acidified and reextracted with an organic solvent. These shifts between water and solvent help in purification of penicillin.
⦁ Treatment of crude extract: penicillin is manufactured as various salts according to the intended use (e.g. for administration to humans and/or animals), usually sodium penicillin.
APPLICATIONS:
⦁ Penicillin was the first antibiotic to be discovered. This antibiotic was extensively used to treat American soldiers wounded in world war II.
⦁ Penicillin is used to treat bacterial infections as it either inhibits bacterial growth or kills it. It does this by preventing bacterial enzymes from creating cell wall growth.
⦁ Other than bacterial infections, penicillin can also be used to treat other medical problems like leptospirosis, chlamydia in pregnant women, helicobacter-pylori associated gastritis or peptic ulcer disease, gas gangrene and typhoid fever.
⦁ Penicillin has its medical significance in treatment of throat infections, meningitis and syphilis.
Sir Alexander Fleming (1881-1955), a British bacteriologist observed that on a bacterial culture plate which had become contaminated by a mould, a bacterial growth in the vicinity of the mould colony was inhibited. He carried on his experiments at St. Mary's Hospital, London in 1928. His results were published in the Journal of Experimental Pathology in 1929. Since the mould was a strain of Penicillin notatum, he named the antibacterial substance , Penicillin. A decade later, two other English men, Chain and Florey engaged themselves in furthering Fleming's investigations. Thus, an antibiotic preparation was developed for human use.
Source:
From Penicillium chrysogenum (ancestral fungus), high yielding strains have been developed for the commercial production of penicillin.
Mode of action of enzyme:
Penicillin inhibits the formation of peptidoglycan cross links in the bacterial cell wall, which is achieved through binding of the 4 membered beta- lactum ring of penicillin to the enzyme DD- transpeptidase. As a consequence, DD- transpeptidase cannot catalyze formation of these cross-links cell walls without intact peptidoglycan cross- links are structurally weak, prone to collapse and disintegrate when the bacteria attempts to divide.
Type of fermentation:
It is a fed batch process that is carried out aseptically in stainless steel tank reactors with a capacity of 30-100 thousand gallon. The fermentation involves 2-3 initial seed growth phases, followed by a fermentation production phase with a duration ranging from 120-200 hours.
Production parameters:
High yielding strains of P. chrysogenum are genetically unstable. Hence, production strains are stored in a dormant form by either of the following techniques:
⦁ Spore suspensions can be lyophilized in appropriate media
⦁ Spore suspension can be stored under liquid nitrogen i.e., in a frozen state.
All manipulations are carried out in laminar flow cabinets in rooms which contains the filtered air.
Personnel wear sterilized clothing and they are careful in their aseptic technique.
There is strict necessity for pure culture in antibiotic fermentations.
Inoculum preparation: the aim is to develop a pure inoculum in sufficient volume and in the fast growing phase for the production fermenter. The time taken for each stage is measured in days, and it decreases as the sequence progresses.
⦁ The medium is designed to provide the organism with all the nutrients that it requires.
⦁ The medium can be solidified by increasing the amount of agar.
⦁ Adequate oxygen is supplied in the form of sterile air.
⦁ Temperature (24 C) is controlled.
Principle criteria for transfer to next stage in progression are:
Freedom from contamination
Growth to a predetermined cell density.
The volume of culture increases approximately 10 fold with each successive stage. The mould, P. chrysogenum grows in filamentous form.
Primary source of spores stored on soil - One or more growth stages on solid media - one or more growth stages in shaken flask culture- 0.5-1.0 m3 seed storage - 10-20 m3 seed storage- 125- 250 m3 production stage
Inoculation methods: Any one of the following methods may be used to inoculate the fermentation medium in the submerged culture production of penicillin:
⦁ Dry spores may be used to seed the fermentation medium
⦁ Inoculation by a suspension of ungerminated mould spores
⦁ The fermentation medium may be seeded by pellet inoculation.
Raw materials: three points should be kept in mind in choosing raw materials for the manufacture of penicillin.
⦁ An abundant growth of mycelium.
⦁ Maximum accumulation of penicillin
⦁ Ease of extraction and purification of antibiotic.
⦁ Carbon sources: lactose, glucose or sucrose
⦁ Nitrogen sources: ammonium sulphate, ammonium acetate, ammonium lactate, corn steep liquor.
⦁ Mineral sources: potassium, phosphorus, magnesium, sulphur, zinc and copper.
⦁ Precursor: phenylacetic acid (PAA).
Component. Percent
Corn steep liquor solid. 3.5
Lactose. 3.5
Glucose. 1.0
Calcium carbonate. 1.0
Potassium dihydrogen phosphate 0.4
Edible oil. 0.25
Penicillin precursor with slow feeding rate
DOWNSTREAM PROCESSING (Extraction and Purification)
⦁ Removal of mycelium: in the first step, mycelium is removed by filtration using a rotary vacuum filter.
⦁ Countercurrent solvent extraction of penicillin: the pH of the filtrate is adjusted to 2-2.5 with the help of phosphoric acid. The antibiotic is then extracted back into an aqueous buffer at pH 7-7.5 through countercurrent solvent extractor. The partition coefficient shifts in the favor of aqueous phase. The resulting solution is again acidified and reextracted with an organic solvent. These shifts between water and solvent help in purification of penicillin.
⦁ Treatment of crude extract: penicillin is manufactured as various salts according to the intended use (e.g. for administration to humans and/or animals), usually sodium penicillin.
APPLICATIONS:
⦁ Penicillin was the first antibiotic to be discovered. This antibiotic was extensively used to treat American soldiers wounded in world war II.
⦁ Penicillin is used to treat bacterial infections as it either inhibits bacterial growth or kills it. It does this by preventing bacterial enzymes from creating cell wall growth.
⦁ Other than bacterial infections, penicillin can also be used to treat other medical problems like leptospirosis, chlamydia in pregnant women, helicobacter-pylori associated gastritis or peptic ulcer disease, gas gangrene and typhoid fever.
⦁ Penicillin has its medical significance in treatment of throat infections, meningitis and syphilis.
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