Determination of amphotericin B, liposomal amphotericin B, and amphotericin B colloidal dispersion in plasma by high-performance liquid chromatography

• Published in: Journal of chromatography. B, Biomedical sciences and applications Vol. 760; no. 2; pp. 307 - 313
• Main Authors: Egger, Petra, Bellmann, Romuald, Wiedermann, Christian J
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 05.09.2001; Elsevier Science
• Subjects: Amphotericin B; Amphotericin B - blood; Amphotericin B - pharmacokinetics; Analysis; Antibiotics; Antibiotics. Antiinfectious agents. Antiparasitic agents; Antifungal agents; Antifungal Agents - blood; Antifungal Agents - pharmacokinetics; Area Under Curve; Biological and medical sciences; Chromatography, High Pressure Liquid - methods; Critical Care; Drug Monitoring; General pharmacology; Humans; Liposomes; Medical sciences; Pharmacology. Drug treatments; Reproducibility of Results; Sensitivity and Specificity; Spectrophotometry, Ultraviolet; Amphotericin B; Antibiotics; Human; Validation; Solid phase extraction; Biological fluid; Intravenous administration; HPLC chromatography; Liposome; Colloidal dispersion; Blood plasma; Antifungal agent; Ultraviolet detector; Polyenic compound; Dosage form; Pharmacokinetics; Quantitative analysis
• ISSN: 0378-4347; 1387-2273
• DOI: 10.1016/S0378-4347(01)00292-4

Amphotericin B is a potent polyene antifungal drug for intravenous treatment of severe infections. It is used as amphotericin B-deoxycholate and in order to reduce amphotericin B toxicity as lipid-formulated complex (liposomal or colloidal dispersion). A sensitive and specific analytical method is presented for the separation of lipid-complexed and plasma protein-bound amphotericin B in human heparinized plasma. This separation, which is required for pharmacokinetic studies, is achieved by solid-phase extraction (SPE) via Bond Elut C 18. The protein-bound amphotericin B has a higher affinity to the SPE material and is therefore retained, whereas the lipid-complexed amphotericin B is eluted in the first step. The recovery of the SPE was >75% for high concentrations and >95% for low concentrations. Quantification was performed by reversed-phase HPLC using a LiChrosorb-RP-8 column, UV detection ( λ=405 nm) and a mixture of acetonitrile–methanol–0.010 M NaH 2PO 4 buffer (41:10:49, v/v) as mobile phase. The retention time for amphotericin B under the given conditions was 6.7 min. The calibration curves were found to be linear ( r≥0.999) in two different ranges (5.0–0.50 μg/ml and 0.50–0.005 μg/ml). Intra- and inter-day precision and accuracy fulfilled the international requirements. No interference from other drugs (typical broad medication for intensive-care patients) or common plasma components was detected in >400 samples analyzed.