title: A versatile method for analysis of saliva, plasma and urine for total thiols using HPLC with UV detection.
abstract: A simple and rapid HPLC method using 2-chloro-1-methyllepidinium tetrafluoroborate (CMLT) as a derivatization reagent was developed for simultaneous determination of homocysteine (Hcy), glutathione (GSH), γ-glutamylcysteine (γ-GluCys), cysteinylglycine (CysGly), N-acetylcysteine (NACys) and cysteine (Cys) in human saliva, plasma and urine. Separation of the analytes was achieved in just 7min using an HPLC, followed by UV detection at 355nm. Chromatographic separation was accomplished on Aeris PEPTIDE XB-C18 (150mm×4.6mm, 3.6µm) column from Phenomenex with a gradient elution: 0-4.0min, 7-30% B; 4.0-5.5min, 30-7% B; 5.5-7.5min, 7% B; (A: B, v/v); (A) 0.5% CH3COOH and (B) EtOH. Mobile phase was delivered at a flow rate 1.0mLmin(-1). Linearity in detector response for total thiols was observed over the range of 0.1-20μmolL(-1) for Hcy, GSH and γ-GluCys, 0.25-50μmolL(-1) for NACys and CysGly and 5-300 for Cys. The LOQ values for Hcy, GSH, γ-GluCys, NACys, CysGly and Cys were 0.05, 0.05, 0.10, 0.06, 0.12 and 0.08μmolL(-1), respectively. The method was successfully implemented to analysis of the samples donated by 15 apparently healthy volunteers and 10 patients.
authors.affiliation: University of Łódź, Faculty of Chemistry, Department of Environmental Chemistry, 163 Pomorska Str., 90-236 Łódź, Poland.
authors.forename: Justyna
authors.initials: J
authors.lastname: Stachniuk
authors.affiliation: University of Łódź, Faculty of Chemistry, Department of Environmental Chemistry, 163 Pomorska Str., 90-236 Łódź, Poland.
authors.forename: Paweł
authors.initials: P
authors.lastname: Kubalczyk
authors.affiliation: University of Łódź, Faculty of Chemistry, Department of Environmental Chemistry, 163 Pomorska Str., 90-236 Łódź, Poland.
authors.forename: Paulina
authors.initials: P
authors.lastname: Furmaniak
authors.affiliation: University of Łódź, Faculty of Chemistry, Department of Environmental Chemistry, 163 Pomorska Str., 90-236 Łódź, Poland. Electronic address: glowar@uni.lodz.pl.
authors.forename: Rafał
authors.initials: R
authors.lastname: Głowacki
mesh_terms: D000328:Adult; D001774:Blood Chemical Analysis / Q000379:methods*; D002138:Calibration; D002851:Chromatography, High Pressure Liquid; D005260:Female; D006801:Humans; D057230:Limit of Detection; D008297:Male; D008875:Middle Aged; D012463:Saliva / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet; D013438:Sulfhydryl Compounds / Q000032:analysis* / Q000097:blood / Q000652:urine; D016482:Urinalysis / Q000379:methods*; D055815:Young Adult
chemical_list: D013438:Sulfhydryl Compounds
keywords: HPLC; Plasma; Saliva; Thiols; Urine