Direct analysis of benzo[a]pyrene metabolites with strong overlapping in both the spectral and lifetime domains

Abstract

The ultimate goal of the present study is to develop screening methodology for the urine analysis of metabolites of polycyclic aromatic hydrocarbons. The investigated approach is based on surface matrix fluorescence spectroscopy, where octadecyl silica membranes serve the dual purpose of metabolite extraction and solid substrate for fluorescence measurements. One of the main challenges faced by this approach is the interference of concomitants in the sample matrix. The present study focuses on three metabolites with strong spectral and lifetime overlapping, namely benzo[a]pyrene-trans-9,10-dihydrodiol, benzo[a]pyrene-r-7,t-8,c-9-tetrahydrotriol and benzo[a]pyrene-r-7,t-8,c-9,c-10-tetrahydrotetrol. As an attempt to improve spectral and lifetime resolution, time-resolved excitation emission matrices are recorded at 77K with the aid of a cryogenic fiber optic probe and laser-based instrumentation. In comparison to conventional spectrofluorimeters, the use of laser-based instrumentation improves the limits of detection by approximately two-orders of magnitude. The fiber optic probe facilitates the collection of time-resolved excitation emission matrices for the detection of the three metabolites at the pgmL−1 concentration level. Their accurate determination in urine samples of unknwon composition is only possible with the aid of unfolded-partial least squares/residual tri-linearization. This algorithm demonstrated to be well-equipped to handle strong overlapping in both the spectral and time domains.