Quality Testing for Aprotinin Preparation by High-Performance Liquid Chromatography

Abstract

The analytical method for aprotinin preparation has been established using reversed phase high performance liquid chromatography, and the characteristics of the four contaminating unknown compounds in this preparation were studied. Aprotinin and the four contaminants were separated by using C18 column (CHEMCOSORB 5-ODS-H) and 13% acetonitrile containing 0.1 M potassium dihydrogenphosphate adjusted to pH 3.0 with phosphoric acid as a mobile phase, or by using a C4 column (CHEMCOSORB 300-7C4) and 10% acetonitrile solution containing the same phosphate buffer as the mobile phase.Five commercially available aprotinin injection forms were found to be composed of aprotinin and identical ratios of the four contaminants. In addition, the four contaminants had weaker inhibitory activity than aprotinin. These four contaminants were considered to be originated from aprotinin as their amino acid compositions of these contaminants were the same as that of aprotinin. Therefore, the production of these four contaminants may occur through an irreversible or extremely biased equilibrium process.