A rapid HPLC method for the measurement of ultra-low plasma methionine concentrations applicable to methionine depletion therapy.

A rapid, sensitive and specific high-performance liquid chromatographic (HPLC) method for the separation and quantification of L-methionine in plasma has been developed. After derivatization of plasma amino acids with o-phthalaldehyde (OPA), a 50 microl sample was loaded on a reversed-phase Supelcosil LC-18-DB column (particle size 5 microm, 25 cm x 4.6 mm, 120A pores). A customized gradient program using tetrahydrofuran/methanol/0.1 M sodium acetate, pH 7.0, v/v/v=5/95/900 and methanol was used with detection by fluorescence. The elution time was 15 minutes, a 3-fold improvement over existing methods. The linearity was 1-100 microM. The limit of detection was 0.5 micromol/L, a 10-fold improvement over existing methods. The inter-assay CVs were 2-5%, and the intra-assays CVs were 4-8%. The sensitivity and rapidity of this HPLC method is particularly applicable to determine the efficacy of methionine depletion therapy of cancer.