クメンヒドロペルオキシドを用いたヒト血漿抗酸化能の測定

摘要

We describe a new method using cumene hydroperoxide to determine antioxidant activity (AO) in human plasma.We used a kit (Determiner LPO:Kyowa Medex Co., LTD. Tokyo Japan) for the determination of lipid peroxides in plasma or serum.30mul of sample was mixed with 70mul of cumene hydroperoxide (50nmol/ml) and incubated at 30degC for 120min before analysis. Samples were mixed with 1.0ml of reagent-I (Determiner LPO) and incubated at 30deg for 5min.Then 2.0ml of reagent-II (Determiner LPO) was added and incubated at 30degC for l0min, at which time the absorbance at 675nm was measured. AO were calculated using the following formula:AO nmol/ml=35nmol/ml-(EsEb)/(Estd-Eb)X35nmol/ml(Es=sample abs., Eb=blank abs.,Estd=standard abs.).Within-run precision for plasma AO was 2.3%.AO in plasma samples stored for 4h at 4deg was decreased by Inmol/ml.After 3h at room temperature, AO was decreased by the same amount.Because this method measured ascorbic acid, a-tocopherol,glutathione peroxidase and quercetin as antioxidant compounds,we were able to measure antioxidant activity in human plasma.Our reference values were calculated from the volunteers group which consisted of 172 students and 82 soldiers. The reference intervals for plasma AO by this procedure were 15.4~20.9nmol/ml.