HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC ASSAY FOR MELANOTAN-1([NLE(4)-DPHE(7)]ALPHA-MELANOCYTE-STIMULATING HORMONE) IN BIOLOGICAL MATRICES

The overall objective of this research was to develop a sensitive, specific, and stability-indicating HPLC assay for the determination of the [Nle(4)-DPhe(7)]alpha-melanocyte-stimulating hormone analog known as Melanotan-1 (MT-1) in biological matrices, i.e., cell culture transport media and human plasma. Separation was accomplished isocratically within 8.0 min using a C-8 reversed-phase column. The mobile phase consisted of 0.1 M phosphate buffer-acetonitrile (80:20, v/v) with 18 mu l/1 triethylamine at pH 2.50. The flow-rate was 1 ml/min with detection at 214 nm. Standard curves (n = 5) were linear over the concentration range 100-1000 ng/ml. The precision, accuracy, intra- and inter-day variations were good with C.V.s typically within 8.7% for concentrations greater than 100 ng/ml. This method was applied to a study of the transport of MT-1 in the Caco-2 cell monolayer model.