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Preparation and application of a monoclonal antibody against the isoflavone glycoside daidzin using a mannich reaction‐derived hapten conjugate
Author(s) -
Yusakul Gorawit,
Sakamoto Seiichi,
Juengwatanatrakul Thaweesak,
Putalun Waraporn,
Tanaka Hiroyuki,
Morimoto Satoshi
Publication year - 2015
Publication title -
phytochemical analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.574
H-Index - 72
eISSN - 1099-1565
pISSN - 0958-0344
DOI - 10.1002/pca.2604
Subject(s) - daidzin , chemistry , daidzein , pueraria , equol , hapten , chromatography , kudzu , puerarin , conjugate , aglycone , monoclonal antibody , isoflavones , glycoside , biochemistry , genistein , antibody , organic chemistry , immunology , medicine , mathematical analysis , alternative medicine , mathematics , pathology , traditional chinese medicine , biology
Daidzin and its aglycone daidzein are major pharmacologically active compounds of soybean ( Glycine max ), kudzu ( Pueraria lobata ), and kwao kruea khao ( P . mirifica ). Pharmacological activities of daidzin are mediated by its more potent metabolites daidzein and equol; however, daidzin is the predominant compound found in these medicinal plants, and the efficacy and safety of equol depend on the amount of daidzin consumed. Objective To develop a specific monoclonal antibody (MAb)‐based indirect competitive enzyme‐linked immunosorbent assay (icELISA) for standardisation of daidzin content in herbal medicines or nutraceuticals. Methodology The Mannich reaction was used for the synthesis of a highly immunogenic conjugate between daidzin and a cationised carrier protein. Splenocytes of hyperimmunised mice were fused with myeloma cells to generate a hybridoma secreting antibody against daidzin. Results The icELISA showed high selectivity and acceptable sensitivity for daidzin determination (1.56–100 ng/mL) with high reproducibility (coefficients of variation were < 5%). The icELISA was a reliable analytical method for daidzin in Glycine max , Pueraria lobata and P . mirifica , for which daidzin recoveries from spiked samples were 98.99–104.94%. Daidzin content of these plant‐derived products determined using the icELISA were in close agreement with those determined by a HPLC‐UV method. Conclusion The icELISA is useful for specific daidzin determination because of its reliability, low cost, speed and high throughput. Copyright © 2015 John Wiley & Sons, Ltd.