Penicillin - Biosynthesis, Media preparation, fermentation

 Penicillin Biosynthesis

Penicillin is a modified tripeptide made from three amino acids aminoadipic acid, cysteine and valine. The principle enzyme involved in penicillin biosynthesis is ACV-synthetase. The β-lactam ring is formed  by isopenicillin-N synthetase; finally, side chains are added

 

Penicllin Fermentation- Medium preparation

Raw materials are primary requirement to design the fermentation broth. Fermentation broth contains all the necessary elements required for the proliferation of the microorganisms.  Generally, it contains a carbon source, nitrogen source, mineral source, precursors and antifoam agents if necessary.

Basic composition of inoculation/fermentation media

Corn steep liquor (3.5%)

Lactose (3.5%)

Glucose (1%)

CaCO₃ (1%)

KH₂PO₄ (0.4%)

Edible Oil (0.25%)

and Penicillin precursor in fermentation media (absent during inoculum preparation)

Inoculum preparation

Aim is to develop a pure inoculum in an adequate amounts. Inoculum is built up sequentially from small flasks to small fermenters. All the parameters like temperature, pH, aeration, agitation etc. should be properly maintained.  

pH: near 5.5 to 6.5

 Temperature: 26°C to 28°C

Aeration: a continuous stream of sterilized air is pumped into it.

Agitation: have baffles which allow constant agitation. 

Carbon Source

Lactose acts as a very satisfactory carbon compound for fermentation. Other carbohydrates like glucose is used during inoculum preparation. Lactose is preferred in fermentation media to provide slow feeding rate and starvation conditions to induce antibiotic formation. Sucrose may also be used. Complex as well as cheap sources like molasses, or soy meal can also be used.

Nitrogen Source

Another essential compound for metabolism of organisms is nitrogen. Ammonium salts such as ammonium sulfate, ammonium acetate, ammonium lactate or ammonia gas are used. Corn steep liquor also may also be used.

Mineral Source

Additionally, some minerals are necessary for the proper growth of these organisms like phosphorus, sulfur, magnesium, zinc, iron, and copper which are 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. The most important naturally occurring penicillin is Penicillin-G. Depending upon the precursors added, the type of penicillin going to produced can be changed.

For example, if phenyl acetic acid is provided then  penicillin-G will be produced but if hydroxy phenyl acetic acid is provided 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.

Anti-foam agents

Anti-foaming agents such as lard oil, octadecanol and silicones are used to prevent foaming during fermentation. 

Following three points should be kept in mind before choosing raw materials for manufacture of penicillin,

1. An abundant growth of mycelium 

2. Maximum accumulation of penicillin 

3. Ease of extraction and purification of antibiotics.

 (contd..)