HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC ASSAY FOR [5-(HYPOXANTHIN-9-YL)PENTYLOXYCARBONYL]-L-ARGININE (ST 789) IN PLASMA BY CYCLIZATION WITHBENZOIN AND FLUOROMETRIC DETECTION

This paper describes a new highly sensitive assay for 5-(hypoxanthin-9 -yl)pentyloxycarbonyl]-L-arginine, an immunomodulatory agent, required for clinical pharmacokinetic investigation. A pre-column derivatizati on by cyclization with benzoin in aqueous medium produces the fluoresc ent 2-substituted amino-4,5-diphenylimidazole fluorescing at 450 nm (e xcitation wavelength 310 nm). L-Arginine-acetyl-L-carnitinamide chlori de (ST 857, II), another arginine derivative, was used as an internal standard. A C18 DB column (5 mum, 250 mm x 4.6 mm I.D.) and a 45:55 (v /v) mixture of 0.05 M ammonium phosphate at pH 7.2 and methanol as mob ile phase were used. Linearity was ascertained in the range 5-100 ng. Extraction recovery from plasma proved to be higher than 90% in the ra nge 5-50 ng/ml. Intra-day precision, expressed as coefficient of varia tion, was in the range 4.7-6.0%. The limit of quantification proved to be 5 ng/ml and the limit of detection 2 ng/ml at a signal-to-noise ra tio of 5. The method is specific.