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Biomedical subjects

L S Kaminsky

Publications and source records attributed to L S Kaminsky.

At least 19 recordsLinked to original sources
PubMed1995

Characterization of human cytochromes P450 involved in theophylline 8-hydroxylation.

Studies were undertaken to determine which human P450 enzymes catalyze the metabolism of theophylline to 1,3-dimethyluric acid (1,3-DU), to facilitate predictions of theophylline drug-drug interactions, and to develop a noninvasive test for human P4501A2. Microsomes from a human cell line transfected individually with human P450 cDNAs for P4501A1, 1A2, 2A6, 2B6, 2C9, 2D6, 2E1, or 3A4 were used to demonstrate that only P4501A2 exhibited catalytic activity for theophylline metabolism to 1,3-DU with high affinity and low capacity (Km = 0.6 mM, Vmax = 37.8, pmol/min/mg), while P4502D6, 2E1, and 3A4 (Km = 14.4, 19.9, and 25.1 mM, respectively, and Vmax = 219.8, 646.4, and 20.8 pmol/min/mg, respectively) exhibited activities with low affinity and variable capacities. Correlations of rates of theophylline 8-hydroxylation to 1,3-DU with other P450 form-specific activities, in a series of ten human liver microsomal preparations, at 5 and 40 mM theophylline concentrations, revealed that at low concentrations the metabolism was catalyzed primarily by P4501A2, while at high substrate concentrations P4502E1 was primarily responsible for catalysis. The results with individually expressed P450s and hepatic microsomal preparations were consistent, indicating that the former system provides a qualitatively accurate reflection of the function of the heterogeneously expressed liver P450s. At pharmacologic theophylline concentrations achieved in vivo, its metabolism must thus be catalyzed primarily by P4501A2.

Cell Line↗
PubMed1995

Determination of theophylline and its metabolites in rat liver microsomes and human urine by capillary electrophoresis.

A capillary electrophoretic (CE) method has been developed for the determination of theophylline and all of its identified and potential metabolites. The method is rapid, resolves all metabolites to baseline, and requires extraction of only some biological fluids. It has been applied to the analysis of theophylline metabolism by hepatic microsomes from rats treated with a variety of inducing agents for different forms of P450 enzymes which metabolize theophylline, and to human urine spiked with theophylline and its metabolites, and concentrated by solid-phase extraction.

Animals↗
PubMed1995

Quantitative RNA-polymerase chain reaction-DNA analysis by capillary electrophoresis and laser-induced fluorescence.

Quantitative RNA-polymerase chain reaction (RNA-PCR) is an extremely powerful analytical tool owing to its specificity and high level of sensitivity. Quantitative RNA-PCR is, however, highly labor intensive. No analytical method currently exists that can accurately and rapidly quantitate the small quantities of DNA in RNA-PCR reaction mixtures. We have developed a method using capillary electrophoresis and laser-induced fluorescence to detect YOYO-1 complexes of DNA produced by PCR. RNA-PCR mixtures can be analyzed either directly (without primer and protein removal) or by electrokinetic injection following desalting. Modified competitive and multiplex competitive RNA-PCR assays for glyceraldehyde-3-phosphate dehydrogenase and P4501A1 were tested in a series of mixtures containing equal concentrations, but different proportions, of RNA from untreated (essentially no P4501A1 mRNA) and 2,3,7,8-tetrachlorodibenzo-p-dioxin-treated (high levels of P4501A1 mRNA) HepG2 cells. Twofold differences in concentrations between two P4501A1 mRNA solutions could be detected by competitive RNA-PCR. Glyceraldehyde-3-phosphate dehydrogenase concentrations were constant throughout. Multiplex competitive PCR produced more variable results due to the presence of contaminating peaks, which hindered accurate area integration. These data demonstrate the potential usefulness of capillary electrophoresis in a variety of quantitative PCR applications.

Base Sequence↗
PubMed1994

Site-directed mutagenesis of putative substrate recognition sites in cytochrome P450 2B11: importance of amino acid residues 114, 290, and 363 for substrate specificity.

Eleven amino acid residues unique to dog cytochrome P450 (P450) 2B11, compared with rat 2B1 and 2B2, rabbit 2B4 and 2B5, and mouse 2B10, in the putative substrate recognition sites [J. Biol. Chem. 267:83-90 (1992)] were mutated to the residues found in 2B1 or 2B5. The mutants were expressed initially in COS cells and screened for activity toward androstenedione and 2,2',4,4',5,5'-hexachlorobiphenyl (245-HCB). P450 2B11 mutants V107I, M199L-N200E-V204R, V234I, A292L, Q473R, and I475S showed no differences from wild-type P450 2B11 in metabolite profiles with either substrate. Mutants V114I, D290I, and L363V exhibited altered androstenedione metabolite profiles and were expressed in Escherichia coli for further study with androstenedione, testosterone, 7-ethoxycoumarin, (R)- and (S)-warfarin, and 245-HCB. With V114I, hydroxylation of steroids and warfarin and 2-hydroxylation of 245-HCB were decreased, whereas 7-ethoxycoumarin O-dealkylation and 3-hydroxylation of 245-HCB were unaltered. For D290I, activities toward all substrates were decreased, except for 16 beta-hydroxylation of testosterone. The activity of L363V was increased 5-6-fold for 16 alpha-hydroxylation of androstenedione and testosterone but was decreased to 40-50% of wild-type activity with 7-ethoxycoumarin and warfarin and to 6-8% of control for 2-hydroxylation of 245-HCB. Alignment of P450 2B11 with P450 101 and super-imposition of the 11 mutated 2B11 residues on a P450 101 three-dimensional model suggest that only residues 114, 290, and 363 represent substrate contact residues, in excellent agreement with the experimental results. The data indicate the importance of the three residues 114, 290, and 363 in substrate specificity and regio- and stereoselectivity of P450 2B11 and also demonstrate that the effects of the mutations vary considerably with different substrates.

Amino Acids↗
PubMed1992

Rat liver metabolism and toxicity of 2,2,2-trifluoroethanol.

2,2,2-Trifluoroethanol (TFE) is a metabolite of anesthetic agents and chlorofluorocarbon alternatives. Its toxicity in rats is a consequence of its metabolism to 2,2,2-trifluoroacetaldehyde (TFAld) and then to trifluoroacetic acid (TFAA). The enzymes involved in the toxic metabolic pathway have been investigated in this study. For the reaction of TFE to TFAld, the major hepatic metabolism associated with toxicity (as assessed by pyrazole-inhibitability) was NADPH dependent and occurred in the microsomes, whereas for TFAld conversion to TFAA, NADPH-dependent microsomal metabolism was significant, but mitochondrial and cytosolic metabolism in the presence of NADPH were also major contributors. NADPH-dependent hepatic microsomal metabolism of TFE to TFAld and TFAld to TFAA was inhibited by carbon monoxide, 2-allyl-2-isopropylacetamide, SKF-525A, metyrapone, imidazole, and pyrazole, and both reactions were oxygen dependent. The metabolism of TFE to TFAld was inhibited by diethyldithiocarbamate, a specific inhibitor of cytochrome P450E1, and by a monoclonal antibody to P4502E1, whereas the metabolism of TFAld was inhibited by neither. Ethanol pretreatment of rats enhanced the Vmax for hepatic microsomal metabolism of TFE to TFAld from 5.3 to 9.7 nmol/mg protein/min, while for TFAld to TFAA the Vmax was increased from 4.3 to 6.5 and the Km was unaffected for both reactions. Phenobarbital pretreatment of the rats did not affect any of these kinetic parameters. Coadministration of ethanol and a lethal dose of TFE very markedly decreased the lethality. Both the lethality (LD50 0.21 to 0.44 g/kg) and the metabolic kinetic parameters [(Vmax/Km)H(Vmax/Km)D = 4.2] were affected markedly when deuterated TFE replaced TFE. In contrast, deuteration of TFAld did not affect its lethality or rates of metabolism, but did affect its Km. Taken together these results indicate that P4502E1 catalyzed toxicity-associated hepatic metabolism of TFE to TFAld, while TFAld metabolism was catalyzed by a P450 which was not P4502E1. The hepatic metabolism of TFAld was not associated with its toxicity, which has been determined previously to be associated with its intestinal metabolism.

Acetaldehyde↗
PubMed1992

Azidowarfarin photoaffinity probes of purified rat liver cytochrome P4501A1.

Substrate specificity differences between various forms of cytochrome P450 (P450) are governed by substrate binding site amino acid residue differences. To determine the identities of these residues, four analogs of warfarin, a thoroughly investigated anticoagulant drug which is regio- and stereoselectively metabolized by many P450s, have been synthesized as photoaffinity probes. The probes 4'-, 6-, 7-, and 8-azidowarfarin were readily photolyzed in neutral solution by 254-nm light, with half-lives of less than 15 s. When the azidowarfarins were photolyzed in the presence of beta-naphthoflavone-inducible P4501A1 (2.5 microM) at -196 degrees C and the P450 was subsequently reconstituted for warfarin metabolism, 50% inactivation was achieved with 160 microM 4'-azidowarfarin, 64 microM 6-azidowarfarin, 127 microM 7-azidowarfarin, and 29 microM 8-azidowarfarin. This inactivation is irreversible. When these concentrations of the azidowarfarins were photolyzed prior to addition to P4501A1, less inhibition of P450 activity was detected and the inhibition was reversible. The CO-ferrous P450 spectrum of P4501A1 at 448 nm was diminished when photoactivated azidowarfarins bound to and inactivated the enzyme, with essentially no formation of P420 except in the case of 4'-azidowarfarin. The inactivation of P4501A1 by photoactivated 4'-azidowarfarin was prevented by 50% by 1.2 mM R-warfarin or 0.3 mM 4'-nitrowarfarin, consistent with the latter being a better P4501A1 substrate than R-warfarin. The photoinactivation of P4501A1 by each of the azidowarfarins was prevented to variable extents by R-warfarin or by 4'-, 6-, 7-, or 8-nitrowarfarin. Taken together these results demonstrate that all four azidowarfarins are potentially useful photoaffinity probes of the substrate binding site amino acid residues of P450s.

Affinity Labels↗
PubMed1992

17 beta-estradiol hydroxylation catalyzed by human cytochrome P450 1A1: a comparison of the activities induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin in MCF-7 cells with those from heterologous expression of the cDNA.

Exposure of MCF-7 breast cancer cells to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) causes an elevated cytochrome P450 content and a marked increase in the microsomal hydroxylation of 17 beta-estradiol (E2) at the C-2, C-4, C-15 alpha, and C-6 alpha positions. In this study we investigated the involvement of cytochromes P450 of the 1A gene subfamily in this metabolism of E2. Hydroxylation at each of these four positions of E2 was inhibited by P450 1A-subfamily inhibitors, alpha-naphthoflavone, benzo[a]pyrene, and 7-ethoxyresorufin. Northern blots showed that treatment of MCF-7 cells with TCDD resulted in production of the 2.6-kb CYP1A1 mRNA, but not the 3.0-kb CYP1A2 mRNA. Immunoblot analyses with anti-P450 1A antibodies confirmed the production of P450 1A1 protein in TCDD-treated MCF-7 cells. Anti-rat P450 1A IgG inhibited the hydroxylation of E2 at C-2, C-15 alpha, and C-6 alpha, but not hydroxylation at C-4. E2 hydroxylation by human cytochromes P450 1A1 and P450 1A2 was assessed in experiments with microsomes from Saccharomyces cerevisiae after transformation with cDNAs encoding the two cytochromes. The major hydroxylase activities of expressed human P450 1A1 were at the C-2, C-15 alpha, and C-6 alpha positions of E2; expressed human P450 1A2 catalyzed hydroxylation predominately at C-2. While both expressed P450s 1A1 and 1A2 had minor hydroxylase activities at the C-4 position, neither catalyzed a low-Km hydroxylation at C-4 similar to that observed with microsomes from TCDD-treated MCF-7 cells. These results provide strong evidence that P450 1A1 catalyzes the hydroxylations of E2 at the C-2, C-15 alpha, and C-6 alpha in incubations with microsomes from TCDD-treated MCF-7 cells, but suggest TCDD may also induce a cytochrome P450 E2 4-hydroxylase that is distinct from P450 1A1 or P450 1A2.

Breast Neoplasms↗
PubMed1992

Phase II metabolism of warfarin in primary culture of adult rat hepatocytes.

We used adult rat hepatocytes in primary culture (HPC) as a model system to study the hepatic phase II metabolism of the anticoagulant warfarin. Hepatocytes were isolated by a collagenase perfusion technique and maintained for 24 hr in Waymouth's medium containing 0.1 mM (R)-warfarin. When HPC medium was analyzed by reverse phase high performance liquid chromatography with diode-array detection, 4'-, 6-, and 7-hydroxywarfarin were identified. Several putative conjugates were observed eluting between 13 and 18 min. Treatment of hepatocyte medium with beta-glucuronidase and sulfatase resulted in the loss of five putative conjugates and concomitant increases in 4'-, 6-, and 7-hydroxywarfarin and warfarin, suggesting that these metabolites and warfarin were conjugated. Use of the beta-glucuronidase inhibitor saccharic acid 1,4-lactone enabled the determination of the relative extents of conjugation of each metabolite by glucuronic acid and sulfate. Glucuronidation was the predominant pathway for 4'-hydroxywarfarin, whereas 6-hydroxywarfarin and warfarin occurred mainly as sulfate conjugates. In contrast, 7-hydroxywarfarin was converted to both glucuronide and sulfate conjugates. Exposure of HPC to phenobarbital resulted in a decrease in cytochrome P-450-mediated production of hydroxylated warfarin metabolites; however, an increase in the production of 8-hydroxywarfarin was observed when HPC were exposed to beta-naphthoflavone. Unique conjugation patterns were found when hydroxylated warfarins were substituted for warfarin in HPC medium. Both 7- and 8-hydroxywarfarin were converted to one sulfate and two glucuronide conjugates, whereas 4'-hydroxywarfarin was converted to a single glucuronide conjugate. A spectral library of these conjugates was used to identify the major conjugates of warfarin formed by rat HPC.

Animals↗
PubMed1991

Small intestinal cytochromes P450.

Small intestinal cytochromes P450 (P450) provide the principal, initial source of biotransformation of ingested xenobiotics. The consequences of such biotransformation are detoxification by facilitating excretion, or toxification by bioactivation. P450s occur at highest concentrations in the duodenum, near the pylorus, and at decreasing concentrations distally--being lowest in the ileum. Highest concentrations occur from midvillus to villous tip, with little or none occurring in the crypts of Lieberkuehn. Microsomal P4503A, 2C8-10, and 2D6 forms have been identified in human small intestine, and P450s 2B1, possibly 2B2, 2A1, and 3A1/2 were located in endoplasmic reticulum of rodent small intestine, while P4502B4 has been purified to electrophoretic homogeneity from rabbit intestine. Some evidence indicates a differential distribution of P450 forms along the length of the small intestine and even along the villus. Rat intestinal P450s are inducible by xenobiotics--with phenobarbital (PB) inducing P4502B1, 3-methylcholanthrene (3-MC) inducing P4501A1, and dexamethasone inducing two forms of P4503A. Induction is most effectively achieved by oral administration of the agents, and is rapid--aryl hydrocarbon hydroxylase (AHH) was increased within 1 h of administration of, for example, 3-MC. AHH, 7-ethoxycoumarin O-deethylase (ECOD), and 7-ethoxyresorufin O-deethylase (EROD) have been used most frequently as substrates to characterize intestinal P450s. Dietary factors affect intestinal P450s markedly--iron restriction rapidly decreased intestinal P450 to beneath detectable values; selenium deficiency acted similarly but was less effective; Brussels sprouts increased intestinal AHH activity 9.8-fold, ECOD activity 3.2-fold, and P450 1.9-fold; fried meat and dietary fat significantly increased intestinal EROD activity; a vitamin A-deficient diet increased, and a vitamin A-rich diet decreased intestinal P450 activities; and excess cholesterol in the diet increased intestinal P450 activity. The role of intestinal P450 in toxifying or detoxifying specific xenobiotics has been clearly demonstrated to only a limited extent. However, elevated intestinal P450 levels have been indirectly linked to gastrointestinal cancer. Intestinal metabolism of 2,2,2-trifluoroethanol produces intestinal lesions with consequent systemic bacterial infection.

Animals↗
PubMed1991

Selective inhibition of warfarin metabolism by diltiazem in humans.

Healthy men were administered i.v. racemic warfarin, S-warfarin or R-warfarin on two occasions, once in the drug-free state and once during concomitant administration of oral diltiazem 120 mg three times daily. Blood samples for determination of plasma warfarin and diltiazem concentrations, the prothrombin time and complete urine collections were obtained for determination of urinary excretion of dehydrowarfarin, 4'-hydroxywarfarin, 6-hydroxywarfarin, 7-hydroxywarfarin, 8-hydroxywarfarin and unchanged parent drug for 216 h after each dose. Concurrent diltiazem administration decreased total racemic warfarin clearance (+/- S.D.) [3.60 +/- 0.99 (control) versus 3.19 +/- 0.86 (diltiazem) ml/min; P less than .05]. Disposition of the more potent anticoagulant S-warfarin was not altered by diltiazem treatment. In contrast total clearance of R-warfarin was decreased [3.47 +/- 1.10 (control) versus 2.77 +/- 0.92 ml/min; P less than .05]. No change in the area under the prothrombin time versus time after warfarin administration curve was associated with diltiazem treatment for any warfarin trial. Inhibition of R-warfarin by diltiazem was regioselective for metabolites as determined by fractional clearance of urinary metabolites. R-6-hydroxywarfarin clearance (+/- S.E.) [29.5 +/- 7.2 (control) versus 15.7 +/- 2.8 ml/h; P less than 0.05] and R-8-hydroxywarfarin clearance [21.1 +/- 3.6 (control) versus 11.2 +/- 2.0 ml/h; P less than .05] were selectively decreased with no significant change in the urinary clearance of R-dehydro, R-4-hydroxy and R-7-hydroxywarfarin. Urinary clearance of unchanged R-warfarin was also decreased during diltiazem administration. Diltiazem inhibits warfarin disposition in humans in a stereospecific and regiospecific manner.

Adult↗
PubMed1990

Catalytic activities of human liver cytochrome P-450 IIIA4 expressed in Saccharomyces cerevisiae.

A human liver cytochrome P-450 (P-450) IIIA4 cDNA clone was inserted behind an alcohol dehydrogenase promoter in the plasmid vector pAAH5 and expressed in Saccharomyces cerevisiae (D12 and AH22 strains). A cytochrome P-450 with typical spectral properties was expressed at a level of approximately 8 x 10(5) molecules/cell in either strain of yeast. The expressed P-450 IIIA4 had the same apparent monomeric Mr as the corresponding protein in human liver microsomes (P-450NF) and could be isolated from yeast microsomes. Catalytic activity of the yeast microsomes toward putative P-450 IIIA4 substrates was seen in the reactions supported by cumene hydroperoxide but was often lower and variable when supported by the physiological donor NADPH. The catalytic activity of purified P-450 IIIA4 was also poor in some systems reconstituted with rabbit liver NADPH-P-450 reductase and best when both the detergent 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate and a lipid extract (from liver or yeast microsomes) or L-alpha-1,2-dilauroyl-sn-glycero-3-phosphocholine were present. Under these conditions the expressed P-450 IIIA4 was an efficient catalyst for nifedipine oxidation, 6 beta-hydroxylation of testosterone and cortisol, 2-hydroxylation of 17 beta-estradiol and 17 alpha-ethynylestradiol, N-oxygenation and 3-hydroxylation of quinidine, 16 alpha-hydroxylation of dehydroepiandrosterone 3-sulfate, erythromycin N-demethylation, the 10-hydroxylation of (R)-warfarin, the formation of 9,10-dehydrowarfarin from (S)-warfarin, and the activation of aflatoxins B1 and G1, sterigmatocystin, 7,8-dihydroxy-7,8-dihydrobenzo[a]pyrene (both + and - diastereomers), 3,4-dihydroxy-3,4-dihydrobenz[a]anthracene, 3,4-dihydroxy-3,4-dihydro-7, 12-dimethylbenz[a]anthracene, 9,10-dihydroxy-9,10-dihydrobenzo[b]fluoranthene, 6-aminochrysene, and tris(2,3-dibromopropyl) phosphate to products genotoxic in a Salmonella typhimurium TA1535/pSK1002 system where a chimeric umuC' 'lacZ plasmid is responsive to DNA alkylation. Reaction rates were stimulated by 7,8-benzoflavone and inhibited by rabbit anti-P-450 IIIA (anti-P-450NF), troleandomycin, gestodene, and cimetidine. Evidence was obtained that rates of reduction of ferric P-450 IIIA4 in yeast microsomes and the reconstituted systems are slow and at least partially responsible for the lower rates of catalysis seen in these systems (relative to liver microsomes). The results of these studies with a defined protein clearly demonstrate the ability of P-450 IIIA4 to catalyze regio- and stereoselective oxidations with a diverse group of substrates, and this enzyme appears to be one of the most versatile catalysts in the P-450 family.

Alcohol Dehydrogenase↗
PubMed1990

Fluoride: benefits and risks of exposure.

This summarizes current knowledge of the benefits and risks of fluoride ingestion. The preponderance of evidence indicates that fluoride can reduce the incidence of dental caries and that fluoridation of drinking water can provide such protection. Due to the ubiquitous nature of exposures to fluoride sources other than drinking water, it is currently impossible to draw firm conclusions regarding the independent effect of fluoride in drinking water on caries prevalence using an ecologic study design. Moderate dental fluorosis occurs in 1 to 2% of the population exposed to fluoride at 1 mg/l in drinking water and in about 10% of the population at 2 mg/l; moderate/severe fluorosis occurs in variable percentages ranging up to 33% of the population exposed to fluoride at 2.4 to 4.1 mg/l in drinking water. The issue of whether moderate or severe dental fluorosis represents an adverse health effect is still controversial. There is no evidence of skeletal fluorosis among the general U.S. population exposed to drinking water fluoride concentrations lower than 4 mg/l. Radiographically detected osteosclerosis after chronic exposure to fluoride in drinking water at 8 mg/l was not associated with clinical symptoms. Reports of crippling skeletal fluorosis associated with low concentrations of fluoride in drinking water in tropical countries have been attributed to other dietary factors. The available data suggest that some individuals may experience hypersensitivity to fluoride-containing agents. Further studies on hypersensitivity are required. There is no evidence of increased incidence of renal disease or renal dysfunction in humans exposed to up to 8 mg fluoride per liter in drinking water. Structural changes in kidneys of experimental animals have been detected at doses exceeding 1 to 5 mg fluoride per kilogram per day. Based on four case reports, individuals with renal insufficiency who consume large volumes of naturally fluoridated water at 2 to 8 mg/l are possibly at increased risk of developing skeletal fluorosis. Studies on the effects of fluoride in individuals with renal insufficiency are needed. There is no evidence that chronic exposure to concentrations of fluoride reported to be greater than 2 mg/l in drinking water increases human cancer mortality or incidence. A study of lifetime exposure to fluoride on cancer incidence in rats and mice has been completed, but assessment for cancer has not been completed. There is no evidence that fluoride is genotoxic except in some in vitro assays at cytotoxic concentrations. There is no in vivo evidence that fluoride affects human cellular enzyme activities.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1989

Analysis of oxidative warfarin metabolites by thermospray high-performance liquid chromatography/mass spectrometry.

Oxidative metabolites of the anticoagulant, warfarin [4-hydroxy-3-(3-oxo-1-phenylbutyl)-2H-1-benzopyran-2-one], produced by the actions of cytochromes P450 were analyzed by thermospray high-performance liquid chromatography/mass spectrometry. Warfarin, dehydrowarfarin, and the 6-, 7-, 8-, and 4'-hydroxy derivatives of warfarin were found to ionize well by the thermospray process in the presence of ammonium acetate. Thermospray mass spectra of these compounds were generally dominated by the protonated molecule, (M + H)+, and ions formed by the loss of water from the protonated molecule, (M + H - H2O)+. Fragment ions arising from the hydroxycoumarin, benzylhydroxycoumarin, and phenylbutanone portions of the molecules were observed, and the relative intensity of these fragment ions was greatly increased with filament ionization and application of a high repeller potential (100-130 V). Selected-ion monitoring of the (M + H)+ and (M + H - H2O)+ ions provided sensitivities for these compounds in the 2 to 10 ng range. A method employing thermospray HPLC/MS with selected-ion monitoring and internal standard quantitation for the analysis of the oxidative metabolites of warfarin is described.

Animals↗
PubMed1989

Teratology of 2,3,7,8-tetrachlorodibenzo-p-dioxin in a complex environmental mixture from the love canal.

The organic phase of a leachate (OPL) from the Love Canal chemical dump site contains more than 100 organic compounds including 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD). The teratogenic potential of OPL was determined in two inbred and one hybrid mouse strain which differ in their sensitivity to aromatic hydrocarbon (Ah) receptor-mediated toxicity. OPL was orally administered in corn oil on Days 6-15 of gestation to C57BL/6J mice (Ahb/Ahb) at doses of 0, 0.1, 0.3, 0.5, and 0.7 g kg-1 day-1 and to DBA/2J (Ahd/Ahd) females, which were mated with either DBA/2J or C57BL/6J males, at 0, 0.5, 1, and 2.0 g kg-1 day-1. In C57BL/6J mice, which express a high-affinity Ah receptor that avidly binds TCDD, the ED50's of OPL for cleft palate and hydronephrosis were 0.44 and 0.11 g OPL kg-1 day-1, respectively. Maternal mortality was 5% at the highest dose. In DBA/2J fetuses, which express a low-affinity receptor, neither treatment-related cleft palate nor hydronephrosis was induced by dose levels that caused 36% maternal mortality. In hybrid D2B6F1 fetuses, the incidence of cleft palate reached only 8% at 2 g OPL kg-1 day-1 but the ED50 for hydronephrosis was 0.76 g OPL kg-1 day-1. TCDD was similarly administered to pregnant C57BL/6J mice at 0, 0.5, 1, 2, and 4 micrograms kg-1 day-1 and to DBA/2J mice at 0, 0.5, 2, 4, and 8 micrograms kg-1 day-1. In C57BL/6J fetuses, the ED50's for cleft palate and hydronephrosis were 4.6 and 0.73 microgram TCDD kg-1 day-1, respectively. In DBA/2J fetuses the ED50's for cleft palate and hydronephrosis were 15.0 and 6.4 micrograms TCDD kg-1 day-1, respectively. Both the OPL and TCDD caused maternal hepatomegaly and thymic atrophy in all strains, but increased only C57BL/6J fetal weights. OPL decreased the number of fetuses per C57BL/6J dam at the two highest doses but there were no other reproductive effects in any of the groups. It was concluded that the OPL is teratogenic and that hydronephrosis is a sensitive measure of TCDD toxicity in a complex organic mixture. Based on the ED50's of OPL- and TCDD-induced cleft palate and hydronephrosis in the C57BL/6J strain, the OPL had TCDD equivalence of 6.6 and 10.5 ppm, respectively. These values compare closely with the chemical analysis of 3 ppm.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗