Deconjugation Characteristics of Ethylparaben Conjugates in Human Urine by Indirect Liquid Chromatography Tandem-mass Spectrometry

Abstract

Summar y An indirect quantitation method based on liquid chromatography tandem-mass spectrometry (LC/MS/MS) has been developed for the analysis of glucuronic acid and sulfuric acid conjugates of ethylparaben, for which standard reference reagents are not available. This method was then used for determining both the ethylparaben conjugates in human urine. The decay of these two conjugates in urine samples containing β-glucuronidase/arylsulfatase appeared to follow the first-order-like reaction kinetics; 4-hydroxybenzoic acid was not produced in the reaction. In addition, a multiple regression (MR) equation was obtained (R 2 adj ＝0.99, P＜0.01) for the increase in ethylparaben concentration and decrease in the peak areas of its glucuronic acid and sulfuric acid conjugates. In the case of methylparaben and its glucuronic acid and sulfuric acid conjugates, a higher P-value was obtained in the MR equation for the glucuronic acid conjugate, owing to the low urinary concentration of methylparaben. Furthermore, this method was applied to determine the concentration of ethylparaben and its two conjugates in urine samples. Following the ingestion of an ethylparaben-containing supplement drink, ~16% of ethylparaben was excreted as a mixture of ethylparaben and its glucuronic acid and sulfuric acid conjugates in the first morning urine of the subject on the following day.