Studies On Metabolism Of Low Substituted Polybrominated Diphenyl Ethers (PBDEs)and Their Hydroxylated Derivatives By Pig Liver Microsomes In Vitro

As a class of additive brominated flame retardants, polybrominated diphenyl ethers (PBDEs), especially the penta-, octa-and deca-BDEs, are widely applied to a number of different commercial products. As a result of their persistence, bioaccumulation and potential biological toxicity such as reproductive toxicity, neurotoxicity, estrogenicity and throid disruption capacity, penta-BDEs and octa-BDEs were classified as persistent organic pollutants (POPs) by Stockholm Convention. To date, PBDEs have been detected in biotia (marine mammals, fish, birds and mollusks) and human tissues (breast milk, blood and adipose tissue). BDE-15, BDE-28and BDE-47were the predominant substituted PBDEs, and were ubiquitous in the environment. In addition, higher substituted PBDEs can undergo debromination and to form lower substituted PBDEs. However, further biotransformation of these generated low substituted PBDEs in organism were still unclear.Metabolic metabolites of PBDEs in mammals-hydroxylated PBDEs (HO-PBDEs) attract considerable concern with more attention to PBDEs. More studies demonstrated HO-PBDEs were also detected in various media (abiota environment, biota and human tissues), and HO-PBDEs have greater biological toxicity effect compared to parental PBDEs. Occurrence of HO-PBDEs in organism and human illustrated bioaccumulation of HO-PBDEs. However, the stability and further metabolism of HO-PBDEs in biota was unclear. Given toxicity of HO-PBDEs, to study in vitro biotransformation of HO-PBDEs is an important significance to evalue thoroughly the potential biological threatIn the present study,2,4,4’-tribromophenyl ether (BDE-28) and three HO-PBDE homologs, namely3’-OH-2,4-diBDE (3’-OH-BDE-7),4’-OH-2,2’,4-triBDE (4’-OH-BDE-17),3-OH-2,2’,4,4’-tetraBDE (3-OH-BDE-47), were target compounds, pig liver microsomes were reaction media and biotransformation of low substituted PBDEs and HO-PBDEs in vitro was studies. Here, metabolic kinetic of the four compounds in pig liver microsomes were investigated, Metabolites formed in the processes were identified, and yields of metabolites of the three HO-PBDEs were quantified. The key enzymes responsible for the metabolism of HO-PBDEs were determined with prototypical Cytochrome P450s (CYPs) inhibitors. In addition, in order to explain the molecular mechanism of interaction of HO-PBDEs with correspondent isoenzymes, molecular docking was used. The results are summarized as follows:(1) BDE-28,3’-OH-BDE-7,4’-OH-BDE-17and3-OH-BDE-47was able to be metabolized by pig liver microsomes significantly. Metabolic rates of homologous compounds decreased with the increasing the number of bromine atoms. Compared one compound with its corresponding hydroxylated derivative, the hydroxylated derivatives seemed to be more easily biotransformed by pig liver microsomes.Large amounts of studies demonstrated PBDEs could be biotransformed to HO-PBDEs via hydroxylation, and HO-PBDEs have greater biological toxicity effect compared to parental PBDEs. However, in the present study, high biotransformation rate of HO-PBDEs in organism might reduce the substantial bioaccumulation potential, which could provide important basis for evaluating potential toxicity level of HO-PBDEs in biota.(2) BDE-28have ten metabolites in pig liver microsomes, namely one4-bromophenol (4-BP), one2,4-dibromophenol (2,4-DBP) and eight mono-OH-triBDEs, and some mono-OH-triBDEs appear to result from an NIH shift (intramolecular migration of a hydrogen atom primarily during hydroxylation) of a bromine atom. Main metabolic pathway of BDE-28in pig liver microsomes might be hydroxylation and cleavage of the diphenyl ether bond. On the other hand,2,4-bromophenol (2,4-DBP),2-bromophenol (2-BP) and4-bromophenol (4-BP) were metabolites of3’-OH-BDE-7,4’-OH-BDE-17and3-OH-BDE-47by pig liver microsomes, which demonstrated that cleavage of the diphenyl ether and debromination were main biotransformation pathway of the three HO-PBDEs.(3) The key enzymes potentially involved in vitro biotransformation of BDE-28and HO-PBDEs by pig liver microsomes were investigated. We observed CYP1A and CYP3A4isoforms of pig liver microsomes participated in metabolism of PBDE congeners. However, metabolism of OH-PBDEs was susceptible to influence of CYP3A4in pig liver microsomes. All three HO-PBDEs were able to be docked into the active site of CYP3A4through molecular docking.3’-OH-BDE-7and4’-OH-BDE-17other than3-OH-BDE-47with ALA305residue to form hydrogen bonds which make substrate further approach to the heme center and the hydroxylation more readily to occur in benzene rings.