Applications of Microorganisms in the Partial Synthesis of Pharmaceuticals

APPLICATIONS OF MICROORGANISMS IN THE PARTIAL SYNTHESIS OF PHARMACEUTICALS

Microorganisms and microbially derived enzymes continue to play a significant role in the production of novel antibiotics. The potential of microorganisms as chemical catalysts, however, was a later development and first realized in the synthesis of industrially important steroids. These reactions assumed increasing importance following the discovery that certain steroids could be formulated as potent therapeutics, e.g. hydrocortisone has anti-inflammatory activity, and derivatives of the steroidal sex hormones are useful as oral contraceptive agents. More recently, chiral inversion of non-steroidal anti-inflammatory drugs (NSAIDs) has also been demonstrated.

A)  Production Of Antibiotics

In the antibiotics industry, the hydrolysis of benzylpenicillin to give 6-aminopenicillanic acid by the enzyme penicillin acylase is an important stage in the synthesis of many clinically useful penicillins. The combination of genetic engineering techniques to produce hybrid microorganisms with significantly higher acylase levels, together with their entrapment in gel matrices, which appears to improve the stability of the hybrids, has resulted in considerable increases in 6-aminopenicillanic acid yields.

A second example is provided by the production by fermentation of cephalosporin C, which is used solely for the subsequent preparation of semisynthetic cephalosporins.

Furthermore, antibiotics produced by fermentation of various moulds and particularly Streptomyces spp., can be utilized by medicinal chemists as starting blocks in the production of what might be more effective antimicrobial compounds.

B)    Steroid Biotransformations

Previously, steroid hormones could only be obtained in small quantities directly from mammals and therefore attempts were made to synthesize them from plant sterols, which can be obtained economically in large quantities. However, adrenocortical steroids are characterized by the presence of an oxygen molecule at position 11 in the steroid nucleus and although it is relatively easy to hydroxylate a steroidal compound it is extremely difficult to achieve site-specific hydroxylation, such that many of the routes used for synthesizing the desired steroid are lengthy, complex and consequently expensive. This problem was overcome when it was realized that many microorganisms are capable of performing limited oxidations with both stereo-and regio-specificity. By simply adding a steroid to growing cultures of the appropriate microorganism, specific site-directed chemical changes can be introduced into the molecule. In 1952, the first commercially employed process involving the conversion of progesterone to 11 α-hydroxyprogesterone by the fungus Rhizopus nigricans was introduced (Figure 26.4). This reaction is an important stage in the manufacture of cortisone and hydrocortisone from more readily available steroids. Table 26.4 gives several other examples of microbially directed oxidations that have been or are employed in the manufacture of steroidal drugs.

More recently, microorganisms utilized for biotransformation reactions have been immobilized by entrapment in a polymer gel matrix to avoid the often costly and time-consuming enzyme extraction steps that can result in enzyme inactivation. Immobilization also serves to increase the stability of membrane-associated enzymes that are unstable in the solubilized state, as well as permitting the conversion of water-insoluble compounds like steroids in two-phase water–organic solvent systems.

C)  Chiral Inversion

Several clinically relevant drugs including salbutamol (a β-adrenoceptor agonist), propanolol (a β-adrenoceptor antagonist) and the 2-arylpropionic acids (NSAIDs) are administered in their racemic form but undergo in vivo chiral inversion through metabolic transformations by microorganisms that mimic phase I metabolic processes, i.e. functionalization reactions. For example, the activity of NSAIDs (e.g. ibuprofen), resides almost exclusively in the S(+) isomers. However, unidirectional chiral inversion from R (–) to S(+) (Figure 26.5) occurs in vivo over a 3-hour period. The S(+) form is a more effective inhibitor of prostaglandin synthesis, and enzymes from some fungal species (e.g. Verticillium lecanii) convert a racemic mixture into the S(+) isomer in vitro. Another example is the biotransformation of ±propranolol to S(+) propranolol by R. arrhizus and Geotrichum candidum with around 83% efficiency.