Rapid determination of intramacrophagic amphotericin B by direct injection HPLC

Abstract

Key WardsColumn liquid chromatography Cell cultures Amphotericin B SummaryThe uptake and kinetic release of the antifungal amphotericin B by phagocytic cells appears to be important in the toxicily-activily mechanism of the drug at the cellular level. An analytical HPLC method has been developed, that is selective, sensitive and reproducible for the determination of amphotericin B uptake by J774.2 macrophage-like cells. Chromatography was on a reversed-phase, end-capped column (Lichrospher Select B Cs) and samples were eluted with a mobile phase of acetonitrile, water, ammonium acetate, acetic acid and 0.02 M triethylamine. The limit of quantitation of amphotericin B was 1 ng ml 1. Precision and accuracywas <5%.

<p>Synthesis, characterization, and mechanistic studies of a gold nanoparticle–amphotericin B covalent conjugate with enhanced antileishmanial efficacy and reduced cytotoxicity</p>

“…RAW 264.1 (1×10 6 cells/ml) cells were treated with 300 µM of AmB and GL-AmB for 1,2, 4, 6, 12 and 24 h. Cells were harvested, washed five times in PBS, and the cell pellet lysed with 0.5 mL of 1% Triton-X-100 (in PBS) for 20 min at 37 °C. Supernatant (0.2 mL) obtained after centrifugation was mixed with 0.2 mL DMSO, mixed by shaking, and then the top layer (0.2 mL) was taken after centrifugation for HPLC analysis 54,55. Samples (10 μL) were analyzed by HPLC as described earlier.…”