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Updated: Apr 23, 2026
Quantifying the Modulation of Elastase Enzyme Activity Through Colorimetric Analysis
Published on: January 17, 2025
Fluorescence polarization enzyme assay: a novel approach for the quantification of quercetin based on its interaction
Svetlana M Filimonova1, Evgeniy S Melnikov2, Liliya I Mukhametova3
1A.P. Nelubin Institute of Pharmacy, Federal State Autonomous Educational Institution of Higher Education I.M. Sechenov First Moscow State Medical, University of Ministry of Health of the Russian Federation (Sechenovskiy University), Prospect Vernadskogo Str. 96/1, 119311, Moscow, Russia. filimonova.svetl@mail.ru.
Abstract:
Traditional fluorescence polarization immunoassay (FPIA) requires costly monoclonal antibodies. This work develops a cost-effective fluorescence polarization enzyme assay (FPEA) for quercetin, employing α-amylase as a significantly cheaper recognition agent. Based on competitive binding with a fluorescent zearalenone tracer, the assay achieves a detection limit of 1.7 mg/mL, a working range of 2.3-6.4 mg/mL, and completes analysis within 5 min, offering a substantial time saving versus typical HPLC runs (~ 20 min). It demonstrates high specificity, with only minimal cross-reactivity to rutin (0.1%) and none to dihydroquercetin, gallic acid, and acarbose. FPEA analysis yielded a quercetin*α-amylase binding affinity constant of 3.9 × 103 L/mol. Successful application to onion peel extract and a dietary supplement yielded results consistent with liquid chromatography with diode-array detection (LC-DAD). The proposed FPEA provides a rapid, simple, and economical alternative for quality control of sources with high quercetin content.
