Paper-based sol-gel thin films immobilized cytochrome P450 for enzyme activity measurement.

Paper-based sol-gel thin films immobilized cytochrome P450 for enzyme activity measurement. Anal Chim Acta. 2020 Feb 15;1098:86-93 Authors: Nuchtavorn N, Leanpolchareanchai J, Suntornsuk L, Macka M Abstract Cytochrome P450 (CYP450), and in particular CYP3A4, is the most abundantly expressed CYP450 isozyme implicated in many drug-drug and medicinal plant-drug interactions. Therefore, incorporation of CYP3A4 enzyme screening at an early stage of drug discovery is preferable in order to avoid enzymatic interactions. Here we present for the first time a paper-based CYP3A4 immobilized sol-gel-derived a platform using resorufin benzyl ether as a fluorogenic enzyme substrate used to investigate enzyme activity. The fluorescence intensity of the product can be simply quantified by using a handheld digital microscope and an image analysis software. The limit of quantitation was 0.35 μM with good precision (RSDs < 4.1%). Furthermore, the assay of CYP3A4 activity on the developed paper-based device provided comparable results with those obtained from conventional well-plates (p > 0.05), while offering simplicity and lower cost. Kinetic parameters of the immobilized CYP3A4 in sol-gel coated paper were calculated from the Lineweaver-Burk plot, including Michaelis constant (Km) and maximum velocity (Vmax), which were 2.71 ± 0.35 μM and 0.43 ± 0.05 μM/min, respectively. Moreover, a functional test of these devices w...
Source: Analytica Chimica Acta - Category: Chemistry Authors: Tags: Anal Chim Acta Source Type: research