An Enzyme-linked Immunosorbent Assay for Genistein 7-O-[α-rhamnopyranosyl-(1→6)]-ß-glucopyranoside Determination in Derris scandens using a Polyclonal Antibody.

ABSTRACT

INTRODUCTION: Genistein 7-O-[α-rhamnopyranosyl-(1→6)]-ß-glucopyranoside (GTG) is a major bioactive compound in Derris scandens. It is responsible for anti-inflammatory activity by inhibition of cyclooxygenase and lipoxygenase. There are many commercial products of D. scandens available in Thailand. OBJECTIVE: To develop an enzyme-linked immunosorbent assay (ELISA) for the quantitative analysis of GTG in plant material and derived products using a polyclonal antibody. METHODS: An immunogen was synthesised by conjugating GTG with a carrier protein. The polyclonal antibody against GTG (GTG-PAb) was produced in New Zealand white rabbits. The ELISA method was validated for specificity, sensitivity, accuracy, precision and correlation with HPLC. RESULTS: The polyclonal antibody was specific to GTG and genistin within the range of compounds tested. The GTG ELISA was applied in the range 0.04-10.00 µg/mL with a limit of detection of 0.03 µg/mL. The recovery of GTG in spiked Derris scandens extracts ranged from 100.7 to 107.0%, with a coefficient of variation less than 7.0%. The intra- and inter-assay variations were less than 5.0%. The ELISA showed a good correlation with HPLC-UV analysis for GTG determination in samples, with a coefficient of determination (r2 ) of 0.9880. CONCLUSION: An ELISA was established for GTG determination in Derris scandens. The GTG-PAb can react with GTG and genistin, but genistin has not been found in the plant. Therefore, the ELISA can be used for high throughput quality control of GTG content in D. scandens and its products. Copyright © 2016 John Wiley & Sons, Ltd.