IMPROVED ASSAY FOR SERUM AMPHOTERICIN-B BY FAST HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

A selective and sensitive method for the determination of amphotericin B in dog serum, after deproteinization, using high performance liquid chromatography HPLC with UV detection at 382 nm was developed. After protein precipitation with methanol, 5 mu l of the supernatant was inj ected onto a reversed phase C-18 column (1.5 mu m, 33 x 4.6 mm ID), wi th a mobile phase composed of 0.05 M sodium acetate - acetonitrile - m ethanol (40:30:30, v/v). The analytical recovery of amphotericin B in serum was 92.30%. The linearity of the assay method was verified up to 15 mu g/mL of amphotericin B. The assay was reproducible with satisfa ctory intra-day and inter-day coefficients of variations (CV < 3%). Th e sensitivity of the method was less than or equal to 0.625 ng of amph otericin B for serum. The method was successfully employed for the pha rmacokinetic analysis of amphotericin B in dogs following intravenous administration of liposomal amphotericin B (AmBisome).