Simultaneous determination of purity and potency of amphotericin B by HPLC

Currently, the quality-control strategy of amphotericin B in US Pharmacopoeia, British Pharmacopoeia, European Pharmacopoeia and Pharmacopoeia of the People’s Republic of China all adopt the combination of purity measurement by HPLC and potency measurement by microbiological assay. In this study, we prepared pure amphotericin B and quantified the relationship between amphotericin B content and potency values using the mass-balance method and microbiological assay. The potency of amphotericin B with an absolute purity of 100% was determined to be 1048.63 U mg –1 . An HPLC method was then established to simultaneously determine the content and potency of amphotericin B, which unified the quality-control procedure for amphotericin B. A good linear relationship was observed between the peak area and the concentration, which could be expressed as y =113074 x +4196.5, R 2 =0.9999. The lower limit of quantification was 0.4473 ng. The HPLC method is expected to become the routine quality-control method and replace the current laborious quality-control procedure in pharmacopoeias.