Biological thiols are one of the most important anti-oxidant defense mechanism in protecting cells against oxidative stress. Their determination in biological materials is carried out mainly by different spectrophotometric assays. In this study; one of the commonly used spectrophotometric assay was adapted to Hitachi 911 Clinical Chemistry Analyzer. Precision and accuracy studies were performed. Inter-assay precision was between 1.8-2.6% (n=15) while intra-assay precision was found to be between 1.4-1.9% (n=15). The obtained recoveries were between 94.6-97.5%. The linearity of the assay was up to 9000 mol/L. The automatized assay correlates well to spectrophotometric assay and gave regression equation of y=0.817x+58.75 (r 2=0.95). In healthy volunteers, total sulfhydryl levels were found to be ranging from 606-757 mol/L. This assay is fully automated and assay time is 5 min. As it is time-saving and reproducible, large number of samples could be processed and this assay could aid the follow up of different pathologic conditions.