BIOTECHNOLOGY - V. H. Gerasymenko - 2006

Part II. Specialized Biotechnologies

CHAPTER 11. BIOTECHNOLOGY OF ANTIBIOTIC PRODUCTION

11.1. PRODUCTION OF β-LACTAM ANTIBIOTICS

Production of Penicillins. The antibacterial action of penicillin and its potential as a therapeutic agent were first established in 1940 by A. Fleming, H. Florey, E. Chain, and their colleagues at Oxford University. At that time, the productivity of the laboratory mold strain was a mere 2 mg of the drug per 1 liter of culture broth, which was far from sufficient for industrial-scale antibiotic manufacturing. Researchers managed to significantly boost the fungus's productivity through systematic, repeated exposure to mutagens such as X-rays and ultraviolet radiation, as well as nitrogen mustard, combined with spontaneous Mutations and the Selection of superior producers. As a result, the penicillin concentration in the culture broth was raised to 2%, representing a 10,000-fold increase (20 g/L of culture broth).

Despite being labor-intensive and time-consuming, the method of enhancing antibiotic-producing strains based on random mutagenesis is still in use today. This is because antibiotic Biosynthesis results from the coordinated action of 10 to 30 different Enzymes encoded by a corresponding number of genes. Due to this polygenic mechanism underlying antibiotic biosynthesis, modifications of individual genes rarely yield the desired outcomes. Furthermore, the MOLECULAR MECHANISMS OF biosynthesis remain elusive for many commercially produced Antibiotics.

For large-scale production of penicillin G, a high-yield industrial strain of Penicillium chrysogenum is employed, which was developed through successive cycles of mutagenesis and selection. It is cultivated using the submerged Fermentation method in bioreactors in the presence of phenylacetic acid, which serves as the precursor for the benzyl side chain of the antibiotic molecule. Intensive penicillin synthesis begins once a high density of mycelial biomass is reached, following the complete depletion of glucose and lactic acid in the medium, at a near-neutral pH. The microbiological phase of penicillin production lasts approximately 200 hours. Upon completion of fermentation, the culture broth is filtered, and the mold Cells are washed. Using butanol and potassium ions in specialized crystallizer units, crystalline potassium penicillin G (with a purity of 99.5%) is recovered from the filtrate and washings, serving as the Starting Material for subsequent chemical modifications.

To this day, penicillin antibiotics constitute the most crucial group of chemotherapeutic agents. The core Structure of penicillins is 6-aminopenicillanic acid, or 6-APA, which is utilized to produce Semisynthetic penicillins (Fig. 11.1).

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Fig. 11.1. STRUCTURE OF THE penicillin molecule

Production of cephalosporin antibiotics.

Cephalosporins represent the second major group of extracellular β-lactam antibiotics, characterized by a six-membered dihydrothiazine ring fused to a β-lactam ring. Consequently, the core structure of cephalosporins is 7-aminocephalosporanic acid, or 7-ACA (the cephem Nucleus), which was first isolated during the purification of cephalosporin C.

Fig. 11.2. 7-Aminocephalosporanic acid (7-ACA)

The commercial production of cephalosporin C combines microbiological Methods with subsequent chemical modification. Cephalosporium acremonium is used as the antibiotic producer. This antibiotic exhibits antibacterial activity against both Gram-positive and Gram-negative microorganisms. However, achieving a therapeutic effect requires high concentrations of cephalosporin C, making it an essential intermediate for further chemical modifications.