Biological monitoring of the five major metabolites of di-(2-ethylhexyl)phthalate (DEHP) in human urine using column-switching liquid chromatography–tandem mass spectrometry

Abstract

We present a fast and reliable on-line clean-up HPLC-method for the simultaneous determination of the five major urinary metabolites of di-(2-ethylhexyl)phthalate (DEHP) namely mono-(2-ethyl-5-carboxypentyl)phthalate (5carboxy-MEPP), mono-[2-(carboxymethyl)hexyl]phthalate (2carboxy-MMHP), mono-(2-ethyl-5-hydroxyhexyl)phthalate (5OH-MEHP), mono-(2-ethyl-5-oxohexyl)phthalate (5oxo-MEHP) and mono-(2-ethylhexyl)phthalate (MEHP). These metabolites represent about 70% of an oral DEHP dose. We for the first time succeeded to reliably quantify 5carboxy-MEPP and to identify 2carboxy-MMHP as major metabolites in native urines of the general population. The analytical procedure consists of an enzymatic hydrolysis, on-line extraction of the analytes from urinary matrix by a restricted access material column (RAM), back-flush transfer onto the analytical column (betasil phenylhexyl), detection by ESI–tandem mass spectrometry and quantification by isotope dilution (limit of detection (LOD) 0.25 μg/l). Median concentrations of a small collective taken from the general population ( n = 19) were 85.5 μg/l (5carboxy-MEPP), 47.5 μg/l (5OH-MEHP), 39.7 μg/l (5oxo-MEHP), 9.8 μg/l (MEHP) and about 37 μg/l (2carboxy-MMHP). The presented method can provide insights into the actual internal burden of the general population and certain risk groups. It will help to further explore the human metabolism of DEHP—an occupational and environmental toxicant of great concern.