Development of HPLC method for identification and quantification of genistein substance

INTRODUCTION. low-toxic polyphenolic compound with significant antioxidant activity. Genistein Genistein (5,7-dihydroxy-3-(4-hydroxyphenyl)chromene-4-on) is in combination with chemoradiotherapy can prevent and reduce radiation damage. However, in order to be included in the prevention and treatment regimens of radiation injuries, genistein should be authorised as a medicinal product. This warrants assessment methods of genistein content that would meet pharmacopoeial requirements for analytical methods.

AIM. This study aimed to develop a method of high performance liquid chromatography (HPLC) for identification and quantitation of the potential active pharmaceutical substance genistein.

MATERIALS AND METHODS. Samples of active pharmaceutical substance genistein were synthesised in St. Petersburg State Chemical and Pharmaceutical University. All samples were analysed using Prominence LC-20 Shimadzu HPLC system with diode array detection (Japan). The temperature in the column compartment was 40  °C, the injection volume 100 μL, the detection wavelength 261 nm. A Shimadzu C18 column (4.6 mm×250 cm, 5 μm particle size) was selected for analysis.

RESULTS. A method was developed for identification and quantitation of a potentially active pharmaceutical substance by HPLC with diode array detection using a mixture of acetonitrile and purified water (75:25 v.:v.) as a mobile phase. The flow rate was 0.7 mL/min and the column temperature 40  °C. Applicability of the developed system was evaluated. During optimisation of the conditions, the flow rate of the mobile phase was set to 0.7 mL/min. The suitability of the developed system was evaluated. The method was validated according to the following parameters: specificity, linearity (0.001–0.01%; r=0.997), correctness (RSD%=1.26), reproducibility (RSD%=0.43), intraday precision (RSD%<2), and robustness.

CONCLUSIONS. The developed method of genistein identification and quantitation by HPLC with diode array detection meets pharmacopoeial requirements for analytical methods and can be proposed for inclusion in the draft pharmacopoeial monograph “Genistein”.

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