Ion-pair reversed-phase and low-molecular-weight aqueous gel permeation high-performance liquid chromatography methods for the determination of amoxicillin oligomers

Abstract

Polymeric substances formed from concentrated sodium amoxicillin in an aqueous solution have been separated using two high-performance liquid chromatographic methods. We used a C18 reversed-phase column with tetrabutylammonium chloride as an ion-pairing agent with an acetonitrile gradient and a TSKgel G2500PWxl column with water as the solvent for gel permeation chromatography. The separated materials, ranging in size from the monomer to the tetramer, have been characterized by functional-group chemical analysis, while the identification of the piperazine-2,5-dione was done using a pure standard. A greater number of peaks which were also better defined were obtained using the ion-pair reversed-phase method, and open and closed beta-lactam ring polymer forms could be distinguished. Using the gel permeation method, only a few monomer, piperazine-2,5-dione, dimer, trimer and combined amoxicillin trimer and tetramer peaks were obtained with water, although those obtained were quite well defined. The data on the time-course of formation of the oligomers and the amoxicillin degradation product were virtually identical by both methods.