Simultaneous determination of multiple phthalate metabolites and bisphenol-A in human urine by liquid chromatography–tandem mass spectrometry

Phthalates and bisphenol A are environmental endocrine-disrupting chemicals used widely in common consumer products. There is increasing concern about human exposure to phthalates and bisphenol A due to the potential adverse effects related to the anti-androgenic activity of phthalates and estrogenic activity of bisphenol A. In assessing environmental exposure to phthalates and bisphenol A, it is essential to have a validated analytical method that can quantify trace concentrations of phthalate metabolites and bisphenol A in humans. In this study, we developed and validated an accurate, sensitive, and robust LC–MS/MS method to simultaneously quantify 5 phthalate monoester metabolites, including mono-methyl phthalate, mono-ethyl phthalate, mono-butyl phthalate, mono-benzyl phthalate, mono-2-ethylhexyl phthalate, and bisphenol A in human urine. In this method, the phthalate metabolites and bisphenol A, along with their isotope labeled internal standards, were extracted from 200 μl of human urine using automated off-line solid phase extraction. The analytes were quantitatively determined using LC–MS/MS operated in negative electrospray ionization multiple reaction-monitoring mode. The limit of quantification was 0.3 ng/ml for mono-methyl phthalate, mono-ethyl phthalate, mono-benzyl phthalate and bisphenol A, and 1 ng/ml for mono-butyl phthalate and mono-2-ethylhexyl phthalate. The precision and accuracy were well within the acceptable 15% range. This validated method has been used successfully in assessing exposure to phthalates and bisphenol A in humans.