Hyperhomocysteinemia induced by folic acid deficiency and methionine load — applications of a modified HPLC method

• Published in: Clinica chimica acta Vol. 252; no. 1; pp. 83 - 93
• Main Authors: Durand, P., Fortin, L.J., Lussier-Cacan, S., Davignon, J., Blache, D.
• Format: Journal Article
• Language: English
• Published: Shannon Elsevier B.V 15.08.1996; Elsevier
• Subjects: Adult; Animals; Arteriosclerosis - complications; Atherosclerosis; Atherosclerosis (general aspects, experimental research); Biological and medical sciences; Blood and lymphatic vessels; Cardiology. Vascular system; Chromatography, High Pressure Liquid - methods; Cricetinae; Cysteine - blood; Female; Folic acid deficiency; Folic Acid Deficiency - blood; Folic Acid Deficiency - complications; Glutathione - blood; Homocysteine; Homocysteine - blood; Humans; Male; Medical sciences; Methionine - toxicity; Methionine load; Middle Aged; Rats; Sensitivity and Specificity; GSH, glutathione; HCYS, homocysteine; Atherosclerosis; CYS, cysteine, CYSGLY, cysteinyl-glycine; Folic acid deficiency; Homocysteine; Methionine load; SBD-F, ammonium 7-fluorobenzo-2-oxa-1,3-diazole-4-sulphonate; Vascular disease; Human; Biological fluid; Deficiency; Clinical biology; HPLC chromatography; Cardiovascular disease; Exploration; Folate; Quantitative analysis; Blood plasma
• ISSN: 0009-8981; 1873-3492
• DOI: 10.1016/0009-8981(96)06325-5

The increasing possibility that homocysteine might be involved in atherosclerosis in non-homocysteinuric subjects has required the measurement of low concentrations of this aminothiol in biological samples. The procedure described here represents an improvement of different HPLC methods. We utilized an isocratic HPLC system with fluorescence detection of plasma total homocysteine derivatized after reaction with ammonium 7-fluoro-benzo-2-oxa-1,3-diazole-4-sulphonate. With the help of the rapidly eluting internal standard N-acetyl-cysteine, the method ensures very good recovery (∼100%), reproducibility and precision (within-assay: 2.31%; day-to-day: 2.8%) in the physiological concentration range. This procedure allowed us to validate various animal models of hyperhomocysteinemia such as dietary folic acid deficiency in rat and acute methionine loads in rat and hamster. Using this method, we also confirmed that men have higher plasma total homocysteine levels than women. Due to its simplicity and reliability, our procedure is suitable for routine analysis of total homocysteine and other aminothiols (cysteine, cysteinyl-glycine and glutathione) in biological samples, as required in clinical and research laboratories.