Determination of four metabolites of the plasticizer di(2-ethylhexyl)phthalate in human urine samples

Abstract

A method for biological monitoring of exposure to the plasticizer di(2-ethylhexyl)phthalate (DEHP) is described. In this method the four main metabolites of DEHP [i.e., mono(2-ethylhexyl)phthalate (MEHP), mono(5-carboxy-2-ethylpentyl)phthalate, mono(2-ethyl-5-oxohexyl)phthalate, and mono(2-ethyl-5-hydroxyhexyl)-phthalate] are determined in urine samples. The procedure includes enzymatic hydrolysis, ether extraction, and derivatization with triethyloxonium tetrafluoroborate. Analysis is performed by gas chromatography-electron impact mass spectrometry. The detection limit for all four metabolites is less than 25 micrograms/l urine. The coefficient of variation based on duplicate determinations of urine samples of workers occupationally exposed to DEHP was 16% for MEHP (mean concentration 0.157 mg/l) and 6%-9% for the other three metabolites (mean concentrations 0.130-0.175 mg/l). The method described here was used to study DEHP metabolism in man. Most persons excrete mono(2-ethyl-5-oxohexyl)-phthalate and mono(2-ethyl-5-hydroxyhexyl)phthalate as a (glucuronide) conjugate. Mono(5-carboxy-2-ethylpentyl)phthalate is mainly excreted in free form, while for MEHP a large interindividual variation in conjugation status was observed. Of the four metabolites quantified, 52% are products of a (omega-1)-hydroxylation reaction of MEHP [i.e., mono(2-ethyl-5-oxohexyl)phthalate and mono(2-ethyl-5-hydroxylation reaction of MEHP [i.e., mono(5-carboxy-2-ethylpentyl)phthalate], and 26% is not oxidized further (i.e., MEHP). A good correlation is obtained when the amount of MEHP omega-hydroxylation products is compared with the amount of MEHP (omega-1)-hydroxylation products in urine samples. When the internal dose of DEHP has to be established we recommend that the levels of all four metabolites of DEHP be studied in urine samples.