Prepared By : Renu Juneja

Date: 28/02/2017

PRESENTATION ON COMMERCIAL PRODUCTION OF ANTIBIOTICS & PENICILLIN-G

1. Introduction of Antibiotics.

2. Commercial Production of Antibiotics.

3. Process Description

4. Introduction of Penicillin G (Benzyl-Penicillin)

5. Biosynthesis of Benzyl-Penicillin

6. Production of Penicillin G (Benzyl-Penicillin)

7. Process Description

8. PFD

INDEX

Antibiotics are chemical substances that can inhibit the growth of, and even destroy, harmful microorganisms. They are derived from special microorganisms or other living systems, and are produced on an industrial scale using a fermentation process.

The first antibiotic was discovered in 1896 by Ernest-Duchesne and "rediscovered" by Alexander-Flemming in 1928 from the filamentous fungus  Penicilium-notatum. 

ANTIBIOTICS

Antibiotics are used in many forms:

For bacterial infections on the skin surface, eye, or ear, an antibiotic may be applied as an ointment or cream.

If the infection is internal, the antibiotic can be swallowed or injected directly into the body. The antibiotic is delivered throughout the body by absorption into the bloodstream.

APPLICATION MODE

ANTIBIOTICS

Inoculum: A small amount of material containing bacteria,

viruses, or other microorganisms that is used to start a culture.

A high yielding strain (Bacteria) is a prerequisite for antibiotic production.

The Inoculum is prepared usually in the form of a spore suspension, which is transferred into the fermenter.

COMMERCIAL PRODUCTION OF ANTIBIOTICS

Fermenter / Bioreactor: Antibiotic are generally produced in stainless steel fermenters

used in the batch or fed mode, see in figure below.

Water cooling is often used to maintain the temperature between 24-26° C.

Generally, the fermenter is maintained at above atmospheric pressure which reduces contamination risk and enhances O2 supply.

The final stage fermenter is preferably used for antibiotic production for the longest period.

BATCH FERMENTER

Initial stages of fermentation are designed for considerable microbial growth; these are carried out in seed stage fermenters of smaller size.

One or more seed stages may be used, depending on the process and the strain, to produce the maximum amount of biomass.

Flow Diagram

Production Medium: Antibiotic production employs a variety of media, a

different one for each stage of operation. In some cases, a suitable precursor for the antibiotic is

also provided.List of Media for different stages

Process Description: The production fermenter is run in a fed batch

mode in which a nutrient e.g., glucose is added continuously throughout the fermentation to enhance the duration of antibiotic production.

This is accompanied by withdrawal of small volumes of broth to check increase in volume of the broth in the fermenter.

Antifoaming agents are added at the appropriate stage of fermentation.

Selected microbiological, physical and chemical features are monitored during the fermentation in order to achieve proper control.

At the end of final production stage incubation, the broth contains only a low concentration (3-35% If the total solutes in the broth) of the antibiotic.

Antibiotic recovery is done by separation of cells from the broth by filtration or centrifugation which is followed by purification.

PENICILLIN G What Is Penicillin?: A class of antibiotics that comes from mold (fungi). Discovered by accident in 1928 by Alexander Fleming, is

the first antibiotic called Penicillin.

Benzyl-penicillin, also known as penicillin G, is an antibiotic used to treat a number of bacterial infections.

Penicillin (PCN or pen) is a group of antibiotics which include penicillin G (intravenous use), penicillin V (use by mouth), procaine penicillin, and benzathine penicillin (intramuscular use).

Formula:C16H18N2O4S Mol. Wt: 334.39 g/mol

The sulfur and nitrogen of the five-membered thiazolidine ring are shown in yellow and blue respectively.

The image shows that the thiazolidine ring and fused four-membered β-lactam are not in the same plane.

The key structural feature of the penicillins is the four-membered β-lactam ring; this structural moiety is essential for penicillin's antibacterial activity.

BIOSYNTHESIS OF PENICILLIN-G Three main and important steps to the biosynthesis

of Penicillin G (benzyl-penicillin).

The first step is the condensation of three amino acids L-α-aminoadipic acid,L-cysteine L-valine into a tripeptide.

Before condensing into the tripeptide, the amino acid L-valine must undergo epimerization to become D-valine.

The condensed tripeptide is named δ-(L-α-aminoadipyl)-L-cysteine-D-valine (ACV).

The second step in the biosynthesis of penicillin G is the oxidative conversion of linear ACV into the bicyclic

intermediate isopenicillin N.

Isopenicillin N is a very weak intermediate, because it does not show strong antibiotic activity.

The final step is a transamidation by isopenicillin N N-acyltransferase, in which the α-aminoadipyl side-chain of isopenicillin N is removed and exchanged for a phenylacetyl  side-chain.

RAW MATERIALS:FUNGUS (MO): Penicillum chrysogenum

CARBON SOURCES: Lactose-concentration of 6%.Others such as glucose & sucrose may be used.

NITROGEN SOURCES: Corn steep liquor (CSL), Ammonium sulphate and ammonium acetate, yeast extract etc

MINERAL SOURCES: Elements potassium, phosphorus, magnesium, sulphur, zinc and copper are essential for penicillin. e.g: Corn steep liquor

PRECURSOR:Phenylacetic acid or Phenoxyacetic acid

PRODUCTION OF PENICILLIN G

Like all antibiotics, penicillin is a secondary  metabolite.

Primary metabolites are produced during active cell growth, and secondary metabolites are produced near the onset of stationary phase.

The industrial production of penicillin was broadly classified in to two processes namely, Upstream processing Downstream processing

UPSTREAM PROCESSING : It encompasses any technology that leads to the

synthesis of a product. Upstream includes the exploration, development and production.

The medium is designed to provide the organism with all the nutrients that it requires. Inoculation method- submerged technique Spores -major source of inoculum.

DOWNSTREAM PROCESSING: The extraction and purification of a biotechnological

product from fermentation is referred to as downstream processing.

PROCESS DESCRIPTION The medium is inoculated with a suspension of conidia

of Penicillium chrysogenum. The medium is constantly aerated and agitated, and the mould grows throughout as pellets.

After about seven days, growth is complete, the pH rises to 8.0 or above, and penicillin production ceases.

Downstream processing is relatively easy since penicillin is secreted into the medium.

Removal of cells: The first step in product recovery is the separation of

whole cells and other insoluble ingredients from the culture broth by technique such as filtration and centrifugation.

The product needs to be very pure, since it being used as a therapeutic medical drug, so it is dissolved and then precipitated as a potassium salt to separate it from other substances in the medium.

Commercial PFD

ISOLATION OF BENZYL PENICILLIN: The PH is adjusted to 2-2.5 with the help of phosphoric or

sulphuric acids. In aqueous solution at low PH values there is a partition

coefficient in favor of certain organic solvents such as butyl acetate.

This step has to be carried out quickly for penicillin is very unstable at low PH values.

Antibiotic is then extracted back into an aqueous buffer at a PH of 7.5, the partition coefficient now being strongly in favor of the aqueous phase.

The resulting aqueous solution is again acidified & re-extracted with an organic solvent.

These shifts between the water and solvent help in the purification of penicillin.

Penicillum chrysogenum

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