This document discusses the process of fermentation, with a focus on penicillin production. It begins with an overview of fermentation and its importance in various industries like wine, beer, and food production. It then describes the fermentation process, equipment used (fermentors), and different types of fermentation (aerobic and anaerobic). A large portion of the document details the production of penicillin, including the microorganism used (Penicillium chrysogenum), inoculum preparation, nutrient media formulation, fermentation process, extraction of penicillin from the media, and purification steps. It also discusses semi-synthetic penicillins derived from 6-aminopenicillanic acid.

1. FERMENTATION
PHARMACEUTICAL PROCESS CHEMISTRY
PRESENTED BY
Mr. Darshan N U B Pharm ( M Pharm)
M Pharmacy II Semester
Dept. of Pharmaceutical Chemistry
SACCP
PRESENTING TO
Mr. Purushotham K N M Pharm ( Ph.D)
ASST. Professor
Dept. of Pharmaceutical Chemistry
SACCP
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3. Products of Fermentation
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Wine Beer Vinegar Kimchi
Pepperoni Sauerkraut Yogurts cheese

5. Importance of Fermentation
It allows the cells to get two ATP gain from one molecule of glucose, even without oxygen
Fermentation takes away the end products of glycolysis
Fermentation is important to the Baking industry
Fermentation is important in wineries and Breweries because yeast uses fermentation to
produce alcohol
Fermentation is important in muscles because it allows the muscle to keep getting a little
energy from glucose even when the oxygen supply cannot keep up with the demand
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6. Fermentation is carried out in an equipment called as fermentor. Fermentor can be defined
as a vessel in which sterile nutrient media and pure culture of microorganism are mixed and
fermentation process is carried out under aseptic and optimum condition. The fermenter
provides sterile environment , an optimum condition that are important for micro-organism to
grow.
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7. Aerobic Fermentation
Aerobic fermentation is a metabolic process by which cells metabolize sugars via
fermentation in the presence of oxygen and occurs through the repression of normal
respiratory metabolism.
Anaerobic Fermentation
Anaerobic fermentation occurs in the fermentation vessel once the oxygen is discharged and
replaced with N2, CO2, or another by-product of the fermentation process. Anaerobic
fermentation is usually a slower process.
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8. PENCILLIN
Penicillin refers to a group of beta-lactam antibiotics used in the treatment
of bacterial infections caused by susceptible, usually Gram-positive
organism.
The chemical structure of penicillin was determined by Dorothy Crowfoot
Hodgkin in the early 1940 s, enabling synthethic production.
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9. The structure of penicillin composed of 4-membered beta lactam ring with
thiazolidine nucleus. It contains three chiral carbon atoms ( C3,C5,C6).
6-Amino penicillanic acid (PAP) is the basic skeleton structure of all
penicillins.
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6-(APA)

10. General Structure of Penicillin
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Generic Name Chemical Name R Group
Penicillin G Benzyl penicillin
Penicillin V Phenoxy-methyl penicillin
Penicillin X P-Hydroxy benzyl penicillin
Penicillin F 2-pentenyl penicillin
Penicillin K n-Heptyl Penicillin CH3(CH2)7-

11. Production of Penicillin
P. Chrysogenum is high yielding strain and therefore most widely used as production strain
Inoculum Preparation:
Purpose is to develop a pure inoculum in an adequate amount.
To do so various sequential steps are necessary like:
1.A starter culture is needed for inoculation.
2.After getting growth on solid media, one or two growth stages should allowed in shaken flask
cultures to create a suspension, which can be transferred to seed tanks for further growth.
3.After about 24-28 hours, the content of the seed tanks is transferred to the primary fermentation
tanks. 11

12. 4. All the bio parameters like temperature, pH , aeration, agitation etc. Should be
properly maintained
Bio parameters
pH : near 6.5
Temperature : 26 to 28 0 C
Aeration : a continuous stream of sterilized air is pumped into it.
Agitation : have baffles which allow constant agitation (200 rpm)
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13. Nutrient Media
• Fermentation broth contains all necessary elements required for the proliferation of the
microorganism.
• Generally, it contains a carbon source, nitrogen source, mineral source. Precursors and
antifoam agents.
Carbon Source
 Lactose in a concentation of 6 %.
 Other carbohydrates like glucose & sucrose.
 Complex as well as cheap source like molasses, or soya meal can also be used which are
made up of lactose and glucose sugars.
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14. Nitrogen Source
 Ammonium salts such as ammonium sulphate, ammonium acetate, ammonium lactate or ammonia
gas are used for this reason
 Sometime corn steep liquor may be used.
Mineral Source
 These elements include phosphorus, sulphur, magnesium, zinc, iron, and copper which generally
added in the form of water soluble salts.
Precursors
 Various types of precursors are added into production medium to produce specific type of penicillin
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15. For example, if phenyl acetic acid is provided then only Penicillin-G will
be produced but if hydroxy phenyl acetic acid is provided then then
penicillin –X will be produced.
Phenoxy acetic acid is provided as precursor for Penicillin V production
When corn steep liquor is provided as nitrogen source, it also provides
Phenyl acetic acid derivatives; therefore it is widely used in the
production of penicillin-G
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16. Anti-foam agents
Anti-foaming agents such as lard oil, octadecanol and silicones are used
to prevent foaming during fermentation
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17. Penicillin is commercially produced in the industry by culturing the
fungus penicillium notatum or penicillin chrysoenum.
The fungus can be cultured in two methods:
• Surface culture method
• Submerged culture method
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18. Surface culture method:
• Fungus is cultured on the surface of the liquid media without agitation
• After appropriate an incubation period the penicillin is extracted from the
media.
Submerged culture method:
• Fungus is cultured in a liquid media which is vigorously aerated and agitated.
• After appropriate an incubation period the penicillin is extracted from the
media.
• Now day, penicillin is produced by this method
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Media Formulation
• Lactose: 1%
• Calcium Carbonate: 1%
• Cornsteep Liquor: 8.5% Corn steep liquor is a by-product of corn wet-milling. A
viscous concentrate of corn solubles which contains amino acids, vitamins and
minerals, it is an important constituent of some growth media.
• Glucose: 1%
• Phenyl acetic acid: 0.5g
• Sodium hydrogen phosphate: 0.4%
• Antifoaming Agent: Vegetable oil

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Biosynthesis of Penicillin

21. Master stock of P. chrysogenum
Sporulation (flask culture)
Spore suspension
Germination /seed tank
Production fermenter
Filtration
Filtrate
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Flow diagram for Penicillin G Production

22. Filtrate
pot. bicarbonate/acetate
Rich solvent
Aqueous extraction
Rich solvent solution
Active carbon treatment
Azeotropic distillation
Solvent wash
Crystalline mass
Drying /Blending
Potassium penicillin G
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23. Semi Synthetic Penicillins
Semi-synthetic penicillins such as Ampicillin, Methicillin, oxocillin, propicillin are
prepared by chemical degradation of 6-Aminopenicillanic acid.
Few of semi-synthetic penicillins are known and some of them have proved to be
definitely superior to Penicillin G in clinical application
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Ampicillin Amoxicillin
Methicillin Nafacillin
Oxacillin

25. Reference
Pharmaceutical Microbiology by N.K. Jain
Medicinal Chemistry volume II by ILANGO and P. Valentina
Wikipedia Date 28/07/2022
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26. THANK YOU
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