L. Robertson

Hepta-, hexa-, tetra- and dichloronaphthalene congeners as inducers of hepatic microsomal drug-metabolizing enzymes.

Pretreatment of immature male Wistar rats with 1,2,3,4,5,6,7-hepta-, 1,2,3,4,5,6,8-hepta- and 1,2,3,4,5,6-hexachloronaphthalene resulted in the induction of several hepatic microsomal drug-metabolizing enzymes. The enzymic activities, reduced cytochrome P-450:CO and ethylisocyanide binding difference spectra and electrophoretic mobilities of the induced microsomal proteins were comparable to those observed after administration of the classical inducer of microsomal aryl hydrocarbon hydroxylase, 3-methylcholanthrene. The 1,2,3,4,5,6,7-heptachloronaphthalene congener, which is fully substituted in the lateral 2,3,6 and 7 positions, was more potent than the 1,2,3,4,5,6,8-hepta- and the 1,2,3,4,5,6-hexachloronaphthalene congeners which contain only 3 lateral chloro substituents. 1,2,3,4-Tetra- and several lower chlorinated naphthalenes were inactive as inducers of microsomal aryl hydrocarbon hydroxylase. The effects of structure on the induction activities of the polychlorinated naphthalenes were similar to those observed for other halogenated aryl hydrocarbons.

Octachloronaphthalene induction of hepatic microsomal aryl hydrocarbon hydroxylase activity in the immature male rat

Administration of octachloronaphthalene to immature male Wistar rats resulted in a dose-dependent increase in several enzymic, electrophoretic and spectral parameters associated with induction of the hepatic microsomal enzymes. Compared to corn-oil (control)treated animals octachloronaphthalene (150 mumol . kg-1) induced hepatic cytochrome P-450 (1.5-fold), benzo [alpha] pyrene hydroxylase (18-fold) and 4-chlorobiphenyl hydroxylase (18-fold) enzyme activities. In addition to increases in the relative peak intensities of the reduced microsomal cytochrome P-450 : CO and ethylisocyanide (EIC) difference spectra the peak maxima were observed at 448.5 and 452.2/428.0 nm, respectively. The effects of administering octachloronaphthalene to the rat were similar to those observed after pretreatment with 3-methylcholanthrene (MC) and electrophoresis of the induced microsomal proteins showed that both compounds enhance heme-staining peptides with comparable electrophoretic mobilities. Moreover coadministration of MC (3 x 100 mumol . kg-1) and octachloronaphthalene (2 x 150 mumol . kg-1) indicated that their inductive effects were not additive. It was concluded that octachloronaphthalene was an MC-type inducer of hepatic microsomal enzymes.

PCBs AS AHH INDUCERS

ABSTRACT The structure-activity rules for polychlorinated biphenyls (PCBs) as inducers of microsomal aryl hydrocarbon hydroxylase (AHH) activity have been determined by assessing the effects of synthetic PCB isomers and congeners on the immature male rat. PCBs which are AHH inducers must be substituted at both para positions, at least two meta positions, but not necessarily on the same ring; the introduction of one ortho chloro substituent into the nucleus of an AHH inducer does not eliminate this type of activity. The addition of a second ortho substituent eliminates the AHH activity in all the PCBs with exception of the 2,2′,3,3′,4,4′-hexa-, 2,2f,3′,4,4′,5-hexa-, 2,3,3′,4,4′,6-hexa-, 2,3,4,4′,5,6-hexa- and 2,2′,3,3′,4,4′,5-heptachlorobiphenyl congeners.

Aryl hydrocarbon hydroxylase (AHH) induction by polybrominated biphenyls (PBBs): Enhancement by photolysis

Abstract Irradiation of the commercial polybrominated biphenyl (PBB) mixture, fireMaster BP-6, in cyclohexane solution at 300 nm for 930 min resulted in a marked diminution of the major components of the mixture. Administration of the photolyzed PBB mixture or fireMaster BP-6 to immature mall Wistar rats caused dose-related decreases in thymus weight and increases in hepatic microsomal benzo[a]pyrene hydroxylase (AHH), 4-dimethylaminoantypyrine N-demethylase and NADPH-cytochrome c reductase activities and cytochrome P-450 content. The dose effecting half-maximal AHH induction for the photolyzed PBBs (9 mg · kg−1) was approximately 6 times lower than that of fireMaster Bp-6 (50 mg · kg−1). Futhermore, the concentration of photolyzed PBBs (2 μM) required to displace 50% of the specifically-bound [3H]TCDD from its high-affinity cytosolic Ah receptor was approximately 150 times lower than that required for fireMaster BP-6 (300 μM), as measured by sucrose density gradient centrifugation analysis. The results suggest that the photolysis of the commercial PBB mixture yields products which possess increased biologic activity.

POLYBROMINATED BIPHENYLS, POLYCHLORINATED NAPHTHALENES AND POLYCHLORINATED TERPHENYLS AS MICROSOMAL ENZYME INDUCERS

ABSTRACT The commercial polybrominated biphenyl mixture, fireMaster BP-6, is a mixed-type inducer of the rat hepatic microsomal drug-metabolizing enzymes. Several congeneric components of this mixture, including 2,3′,4,4′,5-pentabromobiphenyl, 2,3,3′,4,4′,5-hexabromobiphenyl and 2,2′,3,4,4′,5′-hexabromobiphenyl have been synthesized and also exhibited mixed-type inducing activity. A comparison of the structure-activity rules for polybrominated and polychlorinated biphenyls as microsomal enzyme inducers shows both differences and similarities and emphasizes the need for further research in this area. The commercial polychlorinated terphenyl, Aroclor 5440, elicited a mixed-type pattern of microsomal enzyme induction, however, the more highly chlorinated product, Aroclor 5460 (60% by weight C1) was inactive. The effects of the commercial polychlorinated naphthalenes were dependent on their degree of chlorination and three of the more highly chlorinated products, Halowaxes 1013, 1014 and 1051 were classified as mixed-type inducers. Octachloro-naphthalene, the major component of Halowax 1051, induced microsomal benzo[a]pyrene hydroxylase (X15) and dimethylaminoantipyrine N-demethylase (X1.3) activities and intensified a Mr 55,000 protein-staining band on SDS polyacrylamide electrophoresis. This band was also the major hepatic microsomal protein induced by pretreatment of rats with 3-methylcholanthrene.

Synthesis, characterization and biologic effects of polybrominated naphthalenes

Although polybrominated naphthalenes (PBNs) are contaminants of the commercial fire retardant fireMaster BP-6, the individual PBN isomers have not been identified. In order to study PBNs possessing an analogous level of bromination to those found in fireMaster BP-6, three synthetic PBN mixtures, averaging 5.0, 5.3, and 5.6 bromines per naphthalene were synthetized and partially characterized. The PBN mixtures were administered to immature male Wistar rats and found to be potent inducers of cytochrome P-450-dependent monooxygenases. At the lowest dose tested, 30 μmol · kg−1, each PBN mixture caused maximal induction of benzo[a]pyrene hydroxylase activity. On the basis of enzyme activities, ligand-binding spectra and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, the PBN mixtures were determined to be 3-methylcholanthrene-type inducers of cytochrome P-450 (P-448), resembling qualitatively the most toxic polyhalogenated biphenyls, dibenzofurans, and dioxins in this respect. Liver weights were significantly increased and thymus weights diminished by PBN treatment. Light microscopy revealed proliferation of the smooth endoplasmic reticulum in periportal hepatocytes as a consistent change; some rats also had mild fatty changes in centrilobular hepatocytes. Thymuses displayed mild to marked depletion of cortical lymphocytes. The PBN mixtures were much more potent than fireMaster BP-6 in causing these effects; raising the possibility that PBNs are among the minor components of fireMaster BP-6 that contribute significantly to the toxicity of this environmental contaminant.

PCDDs and Related Compounds: Metabolism and Biochemistry

Polychlorinated dibenzo-p-dioxins and related halogenated aromatic compounds are a group of chemicals which exhibit similar physical and environmental properties and elicit a number of common toxic and biologic responses. Moreover within each group of halogenated aromatics, there area multiplicity of isomers and congeners and their metabolism and biochemical effects are remarkable dependent on structure. The metabolism of individual halogenated aromatics thus depends on their degree of halogenation and the specific halogen substitution pattern. Complementary results obtained for the poly-chlorinated dibenzo-p-dioxins and biphenyls show that the toxic congeners are approximate isostereomers of 2,3,7,8-tetrachlorodibenzo- p-dioxin; furthermore, there exists a correlation between the toxicity of halogenated aromatics and their binding activities to a cytosolic receptor protein and potencies as inducers of microsomal aryl hydrocarbon hydroxylase.

INDUCTION OF CYTOCHROME P-448 AND P-450 BY PCB ISOMERS AND CONGENERS

The effects of several synthetic PCB isomers and congeners as inducers of microsomal cytochrome P-450- and P-448-dependent monooxygenases indicated that several changes in the proposed structure-activity relationships were required. With the exception of one compound, 2,3′, 4, 4′,5,5′-hexachlorobiphenyl, all the PCB isomers and congeners which are substituted in the para and meta positions of both rings and at one ortho position were mixed (PB+MC) inducers. 2,3,4,4′,5-Pentachlorobiphenyl was also a mixed inducer. In addition PCBs which contained only one (2,3,3′,4′,5,6-hexachlorobiphenyl) or no (2,3,3′,5,5′,6- hexachlorobiphenyl) para chloro substituents were also PB-type inducers. The results greatly expand the structure-activity rules for induction of PB- and MC-type microsomal enzyme activity.

PCBs AS MICROSOMAL ENZYME INDUCERS: STRUCTURE-ACTIVITY RULES

ABSTRACT A number of highly purified synthetic polychlorinated biphenyl (PCB) isomers and congeners were administered to male Wistar rats to test previously reported structure-activity rules for PCBs as hepatic microsomal drug-metabolising enzyme inducers. By comparing the mode of induction of each PCB with that of the classical inducers phenobarbitone (PB) and 3-methylcholanthrene (MC), the following structure-activity rules were apparent. (1) Structure-activity rules for the induction of PB-type activity could not be clearly defined due to the structurally-diverse PCB isomers and congeners which fall into this category. (2) PCBs which induce MC-type activity must be substituted in both para positions, at least two meta positions (but not necessarily on different phenyl rings) and can also contain one or two ortho chlorines particularly if one phenyl ring contains a 2,3,4-trichloro substitution pattern. Twelve PCB congeners (namely 2′,3,4,4′,5-penta-, 2,3,4,4′,5-penta-, 2,3,3′,4,4′-penta-, 2,3′,4,4′,5-penta-, 2′,3,3′,4,4′,5-hexa-, 2,3,3′,4,4′,5-hexa, 2,3,4,4′,5,6-hexa-, 2,2′,3,3′,4,4′-hexa-, 2,2′,3′,4,4′,5-hexa-, 2,3,3′,4,4′,6-hexa-, 2,3,3′,4,4′, 5,5′-hepta- and 2,2′,3,3′,4,4′,5-heptachlorobiphenyl) elicited both PB- and MC-type responses producing a pattern of mixed (PB + MC) induction. Unlike the previously identified MC-type inducers (namely 3,3′,4,4′-tetra-, 3,3′,4,4′,5-penta-and 3,3′,4,4′,5,5′-hexachlorobiphenyl), many of the mixed-type inducers are major or minor components of the commercial PCB preparations (e.g., Aroclor 1254) and must contribute to their activity as MC-type inducers. Since many of the mixed-type PCB inducers exhibit unusually high toxicity, it is possible that these components are major contributors to both the biologic and toxicologic properties of commercial PCB mixtures.

Further characterization and applications of the 4-chlorobiphenyl hydroxylase assay.

Studies of the metabolism-mediated covalent binding of halobiphenyls to cellular macromolecules revealed that the in vitro metabolism of 4-chlorobiphenyl (4-CBP) by hepatic microsomes is greatly enhanced by in vivo pretreatment of rats with 3-methylcholanthrene (MC) but not phenobarbitone (PB) (Crawford and Safe, 1979). This differential response enables these two classes of inducers to be readily distinguished (Parkinson et al., 1980a).