4. The strain
Penicillium chrysogenum Penicillium notatum
Mold conidiophores, fruiting structures, sporangia, conidia, and asexual spores of Penicillium notatum, also known as Penicillium chrysogenum.
The mold is commonly found in homes, is used in the production of green- and blue-mold cheese, and is used to produce the antibiotic
penicillin. Penicillin was the first antibiotic to be discovered by Alexander Fleming. Magnification of X600
5.
6. • More than fifty years have passed since penicillin was first produced in
volume
10. • large fermenting vats which can hold 10,000 gallons of liquid. The
network of pipes keep the temperatures constant during the process of
making Penicillin
11. Outline of Presentation
1. Media
2. Inocula preparation
3. Process and control parameters (ph, Aeration , Agitation , Temp , D.O.
etc)
4. Downstream processing (Recovery and Purification)
13. • Total Composition of Typical Media:
• Solids (40-60%), lactic acid (12-27%),total nitrogen(7.4-7.8%) , amino
nitrogen(2.6-3.3%),reducing sugars as glucose (1.5-
14%)magnmasum(18-20%).
• Carbon source: Lactose in concentration of 6% satisfactory.
Cornsteep (greatly enhances the yeield of penicillin ). And/or one of
the protein rich oil cakes like cotton seed and groundnut.
• One or more sugars like lactose, sucrose and glucose and glucose
along with a vegetable oil like soybean oil, groundnut oil
• Nitrogen source : Sodium nitrate,ammonium sulphate,ammonium
acetate, ammonium lactate,cornsteep liquor etc.serve as.
14. • Amino acids such as L-cystine,L-cysteine and valine are important in
the synthesis of the b-lactum thiazolidine ring system of the penicillins.
• Mineral salts including sources of sulphate and phosphate
• Precursors are used to increase the yield of penicillin by the
fermentation The requisite precursor, eg. phenylacetic acid,phenoxy
acetic acid and phenylacetamide are commonly used as precursors.
15. Inoculation methods
• Various media employed in the manufacture of penicillin can be
inoculated by several methods, like ..
surface culture : surface of the medium is inoculated with dry
spores. the spore material is applied in such a way as to cover the
surface as uniformly as possible.
submerged culture : production medium is inoculated with dry
spores,by pellet inocula or by ungerminated spores can be prepared in
sterile 0.1% soap solutions, in sterile water containing 100 ppm of
sodium lauryl sulphonate.pellet inoculation saves time in the production
stage. pellet inocula are prepared by growing mycelium from mold
spores under submerged conditions.
18. Conditions of fermentation
• Optimum Temperature – 25 OC.
• Optimum ph Range: 5-7.5, lower ph range yield penicillin substantially
lower
• Buffering agent : Calcium carbonate , however it is not suitable in
surface culture production as it decreases the growth of the molds and
the yield of penicillin.P.chrysogenum being strictly aerobic,
• Rate of Aeration: adequate aeration of the fermenter is essential, rate
vary from around 0.5 volume of air/volume of liquid/minute.,
Effectiveness may be enhanced by increase in pressure of abt 20lb/sq
inch.
• Aeration rate is also attained by the use of proper type of stirrer and at
correct speed.
• Antifoam agents such as tributyl citrate, octadecanol, and lard oil,
prevent excessive foam formation during the production of penicillin by
submerged culture method.
• Prevention of contamination during the production of penicillin is
essential because contamination usually causes rapid destruction of
penicillin.
• Sterilization of facilities and media are easily achieved through steam.
19.
20. • lag phase: when a suspension of bacteria is inoculated into a fresh
medium growth may not occur immediately. The length of the delay, or
lag, in growth depends on the organism, the growth phase of the
inoculum, the composition of the new medium and other factors. During
this period the viable bacteria in the inoculum undergo physiological
readjustment enabling them to utilise substrates in the new medium.
• exponential phase: bacteria enter a period of balanced growth, during
which all constituents of cells increase by the same proportion over the
same time interval and the population doubles in a definite and
constant time (the generation time).
• stationary phase: eventually the bacteria stop growing, because some
nutrient becomes exhausted or because a metabolic end product
reaches toxic concentrations. Balanced growth is not possible and cells
no longer have a constant composition. Also, cells in the stationary
phase are smaller than those in exponential phase because cell division
continues after the synthesis of most macromolecules has slowed
down.
21. • What does the term productive mean?
producing large quantities of something useful (antibiotic)
What is meant by an isolate?
a single strain (pure culture) of organism from a particular source
How might mutations be encouraged?
radiation (ultra-violet, X-rays etc) or chemicals
What is meant by a screening programme?
Many experiments, testing lots of strains to see if they work or not
22. Isolation and Purification
The first step is the recovery process is the removal of mycelium or cells by
filtration or centrifuging.
Second step is to remove the antibiotic from the spent production medium
by solvent extraction, adsorption or precipitation.
Additional solvent extraction,distillation,sublimation, column
chromatography or other methods accomplish purification.
Semi-synthetic penicillins.
Semi synthetic such as penicillin such as Ampicillin, Methicillin, Oxocillin,
Propicllin are prepared by chemical acylation of 6-aminopenicillanic
acid.
23. • The instability of the penicillin molecule under acidic conditions
and its low concentration in the fermentation broth required the
development of extraction equipment that could efficiently contact the
aqueous penicillin-containing broth with the water-immiscible extraction
solvent, and then rapidly separate the two phases.
• This permitted rapid extraction of the penicillin from the acidified
aqueous phase and rapid neutralization of the penicillin-rich
solvent so as to minimize acid degradation of the penicillin. The multi-
stage counter-current contactor developed by Walter Podbielniak for
this purpose enjoyed wide usage for penicillin production, and for other
systems as well.
