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Binding and metabolism of tetrachlorobiphenyls by Cytochrome P450 enzymes : structure-activity relationships

University of British Columbia

Polychlorinated biphenyls (PCBs) are a class of persistent organic pollutants that bioaccumulate in humans and wildlife. Cytochrome P450 (CYP) enzymes play a critical role in the biotransformation of PCBs. The objective of this study was to explore structure-activity relationships between the chlorine substitution pattern of PCBs and their interaction with CYP enzymes. Benzyloxy-, ethoxy- and methoxyresorufin 0-dealkylase (BROD, EROD and MROD, respectively) activities were determined in the presence of four symmetrically substituted PCBs: 2,2’,4,4’-tetrachlorobiphenyl (PCB 47), 2,2’,5,5’-tetrachlorobiphenyl (PCB 52), 2,2’,6,6’-tetrachlorobiphenyl( PCB 54) and 3,3’,4,4’-tetrachlorobiphenyl (PCB 77). The most potent inhibitor of MROD and EROD activities was PCB 77 with approximate Ki values of 10 nM and 46 nM, respectively. PCB 47 and PCB 52 inhibited MROD activity with Ki values of 15 μM and 9 μM, respectively, and inhibited EROD activity with Ki values of 5.5 μM and 6.5 μM, respectively. PCB 54 was unable to inhibit EROD or MROD activity. PCB 47, 52 and 54 inhibited BROD activity with Ki values of 440 nM, 60 nM and 100 nM, respectively. BROD activity was unaffected by PCB 77. A validated and optimized method to analyze PCB 54 biotransformation was developed using liquid chromatography-electrospray mass spectrometry. The major metabolite produced was a monohydroxyl metabolite. The hydroxylation position on the biphenyl ring could not be determined due to a lack of PCB 54 metabolite standards. The rate of formation of the monohydroxyl metabolite was compared in hepatic microsomes from corn oil-, 3-methylcholanthrene (MC)-, phenobarbital (PB)- and dexamethasone (DEX)-treated rats. No metabolite was detectable when PCB 54 was incubated with hepatic microsomes from MC- or corn oil-treated rats. Metabolite formation was greater using hepatic microsomes from PB-treated rats than DEX-treated rats. The involvement of CYP enzymes was investigated further using a panel of recombinant rat CYP enzymes. The rate of monohydroxyl metabolite formation was 5.9 pmol/min/pmol CYP in SUPERSOMES expressing CYP2B1. The only other recombinant CYP enzyme that metabolized PCB 54 was CYP3A1, which had a rate of formation of 0.03 pmol/min/pmol CYP. In summary, the results demonstrate the involvement of the CYP2B enzymes and CYP3A enzymes in the biotransformation of PCB 54.

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2010-01-08

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http://hdl.handle.net/2429/17929

Pharmaceutical Sciences

University of British Columbia

UBCV

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