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1 Acta Chromatographica

1 2008

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An isocratic UV HPLC assay for analysis of total and free melphalan concentrations in human plasma

Nath C. E., Zeng L., Eslick A., Trotman J., Earl W.

Acta Chromatographica

2008

Vol. 20, no. 3

383-398

A simple isocratic HPLC-UV assay for measurement of total and free melphalan concentrations in human plasma is described. Samples were prepared by methanol precipitation (total melphalan assay) and ultrafiltration (free melphalan assay). On a 25 cm × 4.6 mm C column with 0.016 M mixed soldium phosphate citrate buffer (pH 3.75)-acetonitrile 87:13 as mobile phase, at a flow rate of 1 mL min -1, the retention time of melphalan was 11.5 min. Detection was at 254 nm. For total melphalan assay response was a linear function of concentration up to 40 µg mL -1, with excellent interday precision (<6% for 0.5-40 µg mL -1 melphalan), accuracy (<2% deviation from the true concentration), and recovery (91-110%). For free melphalan assay response was a linear function of concentration up to 2.5 ?g mL -1, with good precision (<11% for 0.7-2.5 µg mL -1 melphalan) and recovery (89-93%). Detection limits were 0.1 µg mL -1 and 0.05 µg mL -1 for total and free melphalan assays, respectively. The assays were clinically applied in a study of myeloma patients.