High-performance liquid chromatographic assay for N2-[5-(hypoxanthin-9-yl)pentyloxycarbonyl]-L-arginine (ST 789) in plasma by cyclization with benzoin and fluorimetric detection.
This paper describes a new highly sensitive assay for N2-[5-(hypoxanthin-9-yl)pentyloxycarbonyl]-L-arginine, an immunomodulatory agent, required for clinical pharmacokinetic investigation. A pre-column derivatization by cyclization with benzoin in aqueous medium produces the fluorescent 2-substituted amino-4,5-diphenylimidazole fluorescing at 450 nm (excitation wavelength 310 nm). L-Arginine-acetyl-L-carnitinamide chloride (ST 857, II), another arginine derivative, was used as an internal standard. A C18 DB column (5 microns, 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 mobile phase were used. Linearity was ascertained in the range 5-100 ng. Extraction recovery from plasma proved to be higher than 90% in the range 5-50 ng/ml. Intra-day precision, expressed as coefficient of variation, 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 ratio of 5. The method is specific.
Bruno
G
G
Department of Analytical Methods, Sigma-Tau S.p.A., Rome, Italy.
Curti
S
S
Longo
A
A
Marzo
A
A
Arrigoni Martelli
E
E
D000276:Adjuvants, Immunologic / Q000097:blood* / Q000493:pharmacokinetics; D001120:Arginine / Q000031:analogs & derivatives* / Q000097:blood / Q000493:pharmacokinetics; D001573:Benzoin; D002851:Chromatography, High Pressure Liquid; D003500:Cyclization; D006801:Humans; D007042:Hypoxanthines / Q000097:blood* / Q000493:pharmacokinetics; D013050:Spectrometry, Fluorescence
D000276:Adjuvants, Immunologic; D007042:Hypoxanthines; C054420:PCF 39; D001120:Arginine; D001573:Benzoin