Anaerobic Dehalogenation of Hydroxylated Polychlorinated Biphenyls by Desulfitobacterium dehalogenans

Amendments in 1996 to the Safe Drinking Water and Food Quality Protection Act require the monitoring of estrogenic substances in drinking water. Hydroxylated polychlorinated biphenyls (HO-PCBs), the hydroxylated and excreted metabolites of PCBs, are among the pollutants which have been shown to exhibit both estrogenic and antiestrogenic effects (4, 7, 8, 10, 16, 19). PCBs are widespread, potentially toxic, and carcinogenic pollutants that persist in soil and aquatic sediments (6, 13, 27) and bioaccumulate in the food chain. Despite a ban on the production of PCBs in the United States in 1977, leakage from existing transformers due to corrosion or lightning strikes continues to result in the release of PCB-laden fluids into the environment, and furthermore, dredging and other processes that disturb contaminated sediments result in the mobilization of PCBs. Thus, PCB pollution of the environment remains a potentially serious health threat. As a result of bioconcentration, PCBs can reach high levels, especially in fish-eating birds and mammals (2, 25). The major route of PCB metabolism in these organisms is via monohydroxylation by mixed-function oxidases in the microsomal cytochrome P-450 system (for examples, see references 7, 20, and 28). HO-PCBs have also been detected in fish, wildlife, and human organs, blood, fatty tissues, and milk. HO-PCB congeners are relatively stable in mammalian systems (5, 28). Apparently they are eliminated mainly by excretion via urine and droppings and can be detected as residues in the environment (14, 15, 26). The elimination half-life of the metabolite from the most toxic coplanar PCB congener (3,3′4,4′-Cl-BP), 3,3′,4′,5-tetrachloro-4-hydroxybiphenyl (3,3′,4′,5-Cl-4-HO-BP), administered to pregnant mice was 69 h for the liver and 13 h for serum. Urine from rabbits dosed with 4,4′-dichlorobiphenyl contains 4,4′-Cl-3-HO-BP, 3,4′-Cl-4-HO-BP, and 4-Cl-4′-HO-BP as major metabolites (24). A study of 3,3′,4,4′-[14C]Cl-BP in the fetus in pregnant rats and with nonpregnant rats showed a significantly higher retention of the radioactivity in the pregnant rats (20), which was due to the accumulation of the hydroxylated metabolite 3,3′,4′,5-Cl-4-HO-BP. This metabolite has been shown to be relatively nontoxic (compared to the parent PCB) in adult rats and chicken embryos, but it has a high affinity for transthyretin, the major thyroid binding hormone in rats. HO-PCBs, beside having the potential for estrogenic and antiestrogenic effects, have been shown to act as inhibitors of vitamin A and thyroxin transport (2, 4, 7–11, 16, 19, 20, 25, 29). Morse et al. (20) speculated that HO-PCBs might adversely affect development in utero. Differently chlorinated HO-PCBs have been shown to cause different effects (reference 7 and references therein). Thus, bioremediation and toxicological studies of PCBs need to focus on specific congeners and their metabolism for a valid risk assessment. Practically nothing, however, is known about the degradation of these HO-PCBs in the anaerobic environment. We were interested in the fate of the estrogenic and antiestrogenic 3,5-dichloro-4-hydroxybiphenyl (3,5-Cl-4-HO-BP) and 3,3′5,5′-tetrachloro-4,4′-dihydroxybiphenyl congeners (3,′3,5,5′-Cl-4,4′-diHO-BP). The latter compound has been shown to be a metabolite of the most toxic coplanar PCB congener, 3,3′,4,4′-Cl-BP, in mice (22, 30). Since we are not aware of existing information on the influence of substitutions on the dehalogenation and mineralization of HO-PCBs, we assume that properties similar to those observed with PCBs determine the degradation of HO-PCB under aerobic conditions; since congener 3,3′,5,5′-Cl-4,4′-diHO-BP lacks two suitable free carbons for the aerobic hydroxylation, it should not be readily degraded aerobically and should therefore persist under aerobic conditions (1, 12). More highly chlorinated PCB congeners and, as we show in this report, hydroxylated derivatives can be degraded relatively quickly under anaerobic conditions via reductive dehalogenation. Reductive dehalogenation of PCBs in an anaerobic environment was first demonstrated unequivocally in 1988 (23), based on a previous report (6), but 10 years later and despite many attempts, no pure cultures of PCB-dehalogenating anaerobic prokaryotes have been isolated (33); neither was an HO-PCB-dehalogenating anaerobic culture described before this report. We report here that cell suspensions of an axenic culture of the chlorophenol-dehalogenating anaerobic bacterium Desulfitobacterium dehalogenans (32) can reductively dehalogenate para-hydroxylated PCB derivatives, 4-hydroxylated and 4,4′-dihydroxylated PCBs containing chlorine substituents adjacent to the hydroxyl groups in the absence of any soil or sediment-like matrixes. D. dehalogenans exhibits a high substrate specificity and dehalogenates only chlorines from phenols and hydroxylated aromatic compounds when they are positioned adjacent to the hydroxyl group. In contrast to this substrate specificity, D. dehalogenans is an anaerobe which can utilize a wide spectrum of traditional electron acceptors (the reduced product is given in parentheses), such as sulfite (→ sulfide), fumarate (→ succinate) (31), nitrate (→ nitrite or ammonium) (reference 31 and unpublished results), Fe(III) [→ Fe(II)], and perchloroethylene (→ dichloroethane) (18, 21), but not sulfate.