Black soybean has been widely utilized as foods and oriental medicinal materials. The pigmentation in the seed coat of black soybean is due to accumulate antho-cyanins in the epidermis palisade layer. The anthocyanins of black soybean seed coat are considered as a parameter of quality evaluation of black soybean. Therefore, the pur-pose of this study was to investigate the most suitable HPLC condition for simultaneous determination of antho-cyanins in black soybean seed coats extracts. The efficient HPLC analytical condition of D3G, C3G, and Pt3G con-tained extracts of black soybean seed coats was developed. The gradient elution employed a 250 mm × 4.6 mm i.d. YMC-pak ODS-AM 303 column. The gradient system was used two mobile phases. A gradient elution was performed with mobile phase A, consisting of 5% aqueous formic acid, and mobile phase B, comprising 5% formic acid - acetonitrile, and delivered at a flow rate of 0.7 mL/min as follows: 0~35 min, 90% A ~ 60% A; 36 min, 90% A; 46 min, 90% A. The UV-VIS. detection wavelength was set at 520 nm. The limit of detection (LOD) for D3G, C3G, and Pt3G were under 10 ng/mL.
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