title: Validation and clinical application of an UHPLC method for simultaneous analysis of total homocysteine and cysteine in human plasma.
abstract: Several studies indicate that high levels of homocysteine (Hcy) and L-cysteine (L-Cys) are independent risk factors for cardiovascular disease. The validation and clinical application of an ultra HPLC method for analysis of Hcy and L-Cys is described. The reported method is simple, sensitive, rapid, precise, and less aggressive than other previously reported methods. The effect of the derivatization reaction time, pH, and organic solvent contents in the mobile phase are described and discussed. Optimized conditions resulted in excellent peak shapes. Results of method validation showed a good linearity (r(2) ≥ 0.993) over the investigated concentration ranges and were observed for both compounds. The LOD and LOQ were 0.05 μM and 0.15 μM for Hcy and 0.24 μM and 0.80 μM for L-Cys, respectively. Validation results proved that the method precision was good and the accuracy was satisfactory. This validated method was successfully applied in an epidemiological study to measure and compare the prevalence of Hcy and L-Cys high levels in plasma of Portuguese type 2 diabetic patients with and without angiopathy. The study results showed that prevalence of hyperhomocysteinemia and hypercysteinemia were at least two times higher in diabetic patients with angiopathy compared to diabetics without angiopathy.
authors.affiliation: Department of Food and Nutrition, National Institute of Health Dr. Ricardo Jorge, Lisbon, Portugal.
authors.forename: Ana
authors.initials: A
authors.lastname: Valente
authors.forename: Maria Rosário
authors.initials: MR
authors.lastname: Bronze
authors.forename: Manuel
authors.initials: M
authors.lastname: Bicho
authors.forename: Rui
authors.initials: R
authors.lastname: Duarte
authors.forename: Helena Soares
authors.initials: HS
authors.lastname: Costa
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003545:Cysteine / Q000097:blood*; D003924:Diabetes Mellitus, Type 2 / Q000097:blood; D006710:Homocysteine / Q000097:blood*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D057230:Limit of Detection; D015203:Reproducibility of Results
chemical_list: D006710:Homocysteine; D003545:Cysteine