Multixenobiotic resistance mechanism monitoring: standardization of fluorescence emmited by Rhodamine B

Abstract

Many aquatic organisms express the multixenobiotic resistance mechanism (MXR) mediated by a membrane protein denominated P-glycoprotein (Pgp), which reduce the accumulation of xenobiotics by active transport to out of cells. In order to establish fluorescence microscopy as a quantitative method to monitor MXR activity, the kinetic of fluorescence decay emitted by rhodamine B (RB) was determined. Rhodamine B (1 at 1000 nmoles L-1) were spotted on silica gel plate and fluorescence decay recorded and determined as exposition time (ET, sec) using a fluorescence microscope’s photo sensor. The ET at zero time was obtained from a linear equation of plotting ET(s) against the respective rhodamine B concentrations. The resulting mathematical model (RB = (28/ET) -1 -Blank, r2 = 0.9945), allowed the quantitative determination of rhodamine B intracellular accumulation (nmoles L-1This transport activity quantitation is consequence of the MXR mechanism activity operating in viable sections of gills of the mussel Perna perna and allows its measurement as a molecular biomarker.