Rapid quantitative evaluation of Amphotericin B in human plasma, by validated HPLC method

Abstract

A simple, rapid, and sensitive HPLC method was developed and validated to quantify Amphotericin B (AmB) in human plasma. AmB was extracted from spiked plasma by simple protein precipitation with methanol. The separation was performed on an XBridge™ C18 (150 × 4.6 mm, 3.5 μm) column, with a mobile phase of acetic acid (0.73%) - acetonitrile (60:40, v/v) and at a flow rate of 1 mL/min. The eluted peak of AmB was monitored at 408 nm with photo-diode array detector (PDA) detector. The calibration curve was found linear in the AmB concentration range of 1000 - 50 ng/mL (r2>0.99). The inter- and intra-day precisions (%CV) were less than 11.2%. The extraction recoveries were 85-91%. The method developed and validated is simple, rapid (<3 min per injection), sensitive, and reproducible. It potentially can be used for the pharmacokinetic, bioequivalence, and toxicokinetic studies of AmB. © 2013 Balabathula P, et al.