Determination of 3-amino-5-mercapto-1,2,4-triazole in serum

• Published in: Journal of chromatography. B, Analytical technologies in the biomedical and life sciences Analytical technologies in the biomedical and life sciences, 2004-03, Vol.801 (2), p.359-362
• Main Authors: Depree, G.J., Siegel, P.D.
• Format: Article
• Language: English
• Subjects: 3-Amino-5-mercapto-1,2,4-triazole; Acetonitriles; Analysis; Analytical, structural and metabolic biochemistry; Animals; Biological and medical sciences; Chemical Precipitation; Chromatography, High Pressure Liquid - methods; Fluorescent Dyes; Fundamental and applied biological sciences. Psychology; General pharmacology; Humans; Male; Medical sciences; Pharmacology. Drug treatments; Quality Control; Rats; Rats, Inbred F344; Sensitivity and Specificity; Solutions; Triazoles - blood; Water
• ISSN: 1570-0232; 1873-376X
• DOI: 10.1016/j.jchromb.2003.11.031

A high performance liquid chromatography (HPLC) method using fluorescence detection to determine 3-amino-5-mercapto-1,2,4-triazole (AMT) levels in serum has been developed. Sample preparation involved treatment with tributylphosphine (TBP) to reduce disulfides formed during storage, precipitation of proteins with acetonitrile (ACN), and precolumn derivatization using the thiol reactive fluorescent probe monobromobimane (MBB). The conjugate (AMT-MBB) was resolved by gradient elution from a C 18 reversed-phase column. The assay method was linear over a concentration range of 0.78–50 μg/ml and had a limit of detection (LOD) of 0.05 μg/ml AMT (10 μl injection). This method provides a sensitive and specific tool for the determination of AMT in serum and may have potential industrial hygiene application.