Analytical Sciences

Sequential Injection Analysis with Chemiluminescence Detection for the Antioxidative Activity against Singlet Oxygen

Aoi MIYAMOTO, Kuniko NAKAMURA, Yoshihito OHBA, Naoya KISHIKAWA, Kenichiro NAKASHIMA, and Naotaka KURODA

Graduate School of Biomedical Sciences, Nagasaki University, 1-14 Bunkyo-machi, Nagasaki 852-8521, Japan

A sequential injection analysis (SIA) with chemiluminescence (CL) detection was developed for the measurement of antioxidative activity against singlet oxygen (¹O₂). Lactoperoxidase-hydrogen peroxide-bromide ion system was used for the generation of ¹O₂. When a 100 mM sodium acetate buffer (pH 4.5) was used as a carrier solution, the SIA-CL system could be optimized with respect to the flow-rate of the carrier, concentration of reagents and their aspiration order. The antioxidative activity was expressed as an attenuation of luminol CL due to the quenching of ¹O₂ by an antioxidant. The relative standard deviations of antioxidative activity (n = 3) against ¹O₂ for within- and between-day analyses were ≤ 1.6% (20 µM Trolox). The system was successfully applied to the assay of antioxidative activities of various antioxidants including vitamin supplements at a rate of 10 samples within 15 min. The proposed SIA-CL system was rapid and reproducible with minimum consumption of the sample and of reagents, and thus was useful for the screening of compounds possessing antioxidative activity against ¹O₂.

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