KINETIC ANALYSIS OF BENZYLPENICILLIN DEGRADATION IN ALKALINE MEDIA AND ELUCIDATION OF THE EPIMERIZATION MECHANISM OF BENZYLPENICILLOIC ACID

The problems of allergy associated with penicillin therapy have been partially explained by the interaction of tissue macromolecules and various degradation products of penicillin. Consequently, extensive research has been conducted in order to identify and quantify the decomposition products. This study describes the development of an ion-pair reversed-phase high-pressure liquid chromatographic method for the separation and quantitation of benzylpenicillin and its decomposition products in aqueous solutions. It also describes the characterization of certain breakdown products of benzylpenicillin in alkaline media using liquid chromatography, ultraviolet spectroscopy, differential pulse polarography and nuclear magnetic resonance spectroscopy. The rate and mechanism of epimerization of benzylpenicilloic acid as well as the kinetic analysis of benzylpenicillin degradation at pH 7.4 and pH 10.0 are also reported. The major degradation product of benzylpenicillin in alkaline media was found to be benzylpenicilloic acid, a compound which exists in solution as mixture of isomers. The direct hydrolytic product was 5R,6R-benzylpenicilloic acid, which, later, isomerized slowly to the 5S,6R-epimer until equilibrium was established. Epimerization proceeded via the imine rather than the enamine tautomer of penamaldic acid as had been suggested, and, at equilibrium, 5S,6R-benzylpenicilloic acid was found to be the favored product. The conversion process was monitored using reversed-phase high-pressure liquid chromatography and nuclear magnetic resonance spectroscopy. The rate constants for the forward and reverse reactions were also determined by assuming that the rate of change of the concentration of the intermediate approaches zero after a finite time, and, consequently, the overall reaction was treated as a reversible reaction. Finally, benzylpenicillin was allowed to degrade at pH 7.4 and pH 10.0 under different experimental conditions and the sequence of formation of the various degradation products was determined. Based on the results obtained, two degradation schemes for benzylpenicillin degradation at pH 7.4 and pH 10.0 were proposed