Quantitation of Melphalan in Human Plasma by Liquid Chromatography and Solid Phase Extraction: Application to Pharmacokinetic Study

• Published in: Journal of liquid chromatography & related technologies Vol. 27; no. 2; pp. 301 - 313
• Main Authors: Poujol, S., Pinguet, F., Mougenot, P., Malosse, F., Fabbro, M., Culine, S., Astre, C., Bressolle, F.
• Format: Journal Article
• Language: English
• Published: Colchester Taylor & Francis Group 01.01.2004; Taylor & Francis
• Subjects: Analysis; Biological and medical sciences; General pharmacology; HPLC; Human plasma; Medical sciences; Melphalan; Pharmacology. Drug treatments; SPE; Antineoplastic agent; Human; Solid phase extraction; Biological fluid; Chemical analysis; HPLC chromatography; Blood; Chemical enrichment; Blood plasma; Alkylating agent; Reversed phase chromatography; Ultraviolet detector; Sample preparation; Melphalan; Pharmacokinetics; Quantitative analysis
• ISSN: 1082-6076; 1520-572X
• DOI: 10.1081/JLC-120027101

A high-performance liquid chromatographic (HPLC) method for the determination of melphalan in human plasma was developed. This method involved a solid phase extraction (SPE) of melphalan and the internal standard (propylparaben) from plasma, using bond Elut C2 SPE columns with an elution solvent of 0.5 mL of acetonitrile-purified water (50:50, v/v). Separation of the two analytes was achieved within 15 min, using a reversed-phase Kromasil C18 analytical column (150 × 4.6 mm I.D., 5 µm particle size) with a mobile phase of methanol-water-acetic acid (48:51:1, v/v). An ultraviolet detector operated at 261 nm was used, with a linear response observed from 10 to 250 ng mL −1 . Obtained from the method validation, inter-assay precision was below 6% and accuracy is near 100%. The extraction efficiency of the assay was approximately 71% and was constant across the calibration range. The lower limit of quantitation was 10 ng mL −1 ; at this level, precision was 5% and accuracy was 101%. The applicability of this method has been demonstrated by the successful analysis of clinical plasma samples. The SPE procedure developed in this paper, to quantify melphalan in biological samples requiring three steps for sample loading, clean-up, and elution, can easily be automated by using either a robot or an automated sample preparation system.