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Abstract
In the present study, we developed and validated an analytical method using ultra performance liquid chromatography-mass spectrometry (UPLC/MS) for the quantitative determination of 2,2',4,4'-tetrabromodiphenyl ether (BDE-47) metabolism by rat hepatic microsomes. BDE-47 is a brominated flame retardant that was widely used in a variety of consumer products and has subsequently been identified as a ubiquitous environmental contaminant. Hydroxy-bromodiphenyl ethers (OH-BDEs) were isolated from rat hepatic microsomes by liquid-liquid extraction. Chromatographic separation was achieved by UPLC on a C18 column with gradient elution using a mobile phase consisting of methanol and water, each containing 0.1% formic acid, at a flow rate of 0.2 mL/min. Detection and quantification were performed using a mass spectrometer in single ion recording mode with negative electrospray ionization. The UPLC/MS method was validated for linearity, limit of quantification (LOQ), accuracy, precision and recovery. The weighted calibration curves (1/X2) were linear over a concentration range of 5 - 250 nM with LOQ values between 5 nM and 50 nM for the individual OH-BDEs. Intra- and inter- day accuracy (%DEV) and precision (%RSD) values ranged from –11.7% to 9.5% and 5.9% to 16.5%, respectively. Recovery values of 70% to 90% were obtained for all OH-BDEs. The validated method allowed us to successfully analyze metabolite formation following incubation of BDE-47 with hepatic microsomes prepared from phenobarbital-treated rats. Results demonstrate that the UPLC/MS method has sufficient sensitivity and reproducibility to fully characterize the in vitro metabolism of BDE-47 and possibly other PBDEs.
Highlights
Polybrominated diphenyl ethers (PBDEs) are halogenated aromatic hydrocarbons that have been used as additive flame retardants on a variety of consumer products since 1965 [1]
APCI and electrospray ionization mode (ESI) in negative and positive modes were compared for the detection of OH-BDE standards by mass spectrometry using selected ion recording (SIR)
OH-BDE standards were not detected in positive ion mode
Summary
Polybrominated diphenyl ethers (PBDEs) are halogenated aromatic hydrocarbons that have been used as additive flame retardants on a variety of consumer products since 1965 [1]. PBDEs were marketed as commercial mixtures containing a limited number of the 209 possible brominated diphenyl ether (BDE) congeners [2]. PBDEs are not chemically bound to the polymer components of the products to which they are applied and can be released into the environment during manufacture [5], use [7] and disposal [8] of these products. This factor, together with the high lipophilicity, chemical stability and the widespread use of BDE mixtures has resulted
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