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Biomedical subjects

D E Moody

Publications and source records attributed to D E Moody.

At least 19 recordsLinked to original sources
PubMed1996

Correlations of the induction of microsomal epoxide hydrolase activity with phase II drug conjugating enzyme activities in rat liver.

Within the selective induction of phase II enzymes following treatment with dipyridyls or N-heterocyclic analogs of phenanthrene, strong correlations (r > or = 0.70) are observed between the increase of microsomal epoxide hydrolase (mEH) activity and UDP-glucuronosyltransferase (UGT) activities towards 4-nitrophenol, 1-naphthol and morphine. The present study investigates whether this correlation is maintained with inducing agents known to also increase phase I enzyme activities. Rats were treated with beta-naphthoflavone, isosafrole, phenobarbital, ethanol, dexamethasone and clofibric acid regimens in which P450 isozyme induction could be confirmed. Comparisons between the responses of mEH, UGT and glutathione S-transferase (GST) activities were made. mEH activity was increased by beta-naphthoflavone, isosafrole, phenobarbital and clofibric acid. The elevation in mEH activity by these agents showed modest but significant correlations with GST activities toward all the substrates monitored (r values range between 0.49 and 0.65) and a strong correlation with UGT activity towards only one substrate, morphine (r = 0.70). This study suggests that induction of mEH activity correlates with the increases in select phase II enzyme activities whether it is accompanied by P450 induction or not.

Animals↗
PubMed1995

A radiometric TLC assay of liver microsomal dextromethorphan O-demethylation.

A simple and sensitive assay for in vitro analysis of dextromethorphan O-demethylation, a marker for P450 2D deficiency in both humans (2D6) and rats (2D1), has been devised. Commercially available [N-methyl-3H]-dextromethorphan was used to develop a radiometric TLC assay for dextromethorphan O-demethylation. Hexane-triethylamine efficiently extracted dextromethorphan and metabolites from rat liver microsomes, and a solvent system of cyclohexane-toluene-diethylamine (65:15:20, v/v/v) provided sufficient separation (approximately 2 cm) between the two radioactive bands, dextromethorphan and dextrorphan, and no interference from the unlabeled N-demethylation products, 3-methoxymorphinan and 3-hydroxymorphinan. The recovery of dextrorphan from TLC plates increases with microsomal protein and incubation time. An eight-fold decrease in activity was noted in female Dark Agouti relative to the male Sprague-Dawley rats, respective models for poor and extensive P450 2D metabolizers. The assay, even with an approximately 100-fold dilution of radiolabeled substrate, had an approximate limit of detection of 100 pmol. Within- and between-run imprecision was 12.4% and 7.2%, respectively. The radiometric TLC assay for dextromethorphan O-demethylation was sensitive and easy, and used readily available equipment.

Animals↗
PubMed1995

Assignment of the growth hormone receptor gene to bovine chromosome 20 using linkage analysis and somatic cell mapping.

A polymorphism was identified in the bovine growth hormone receptor (GHR) gene by digesting polymerase chain reaction (PCR) products with the restriction enzyme Alul. Two alleles were segregating in cattle of Bos indicus descent, but one allele appears to be fixed in Bos taurus cattle. GHR was localized to bovine chromosome 20 using bovine-rodent hybrid cell lines and linkage analysis.

Animals↗
PubMed1994

Urinary excretion of amphetamine and 4'-hydroxyamphetamine by Sprague Dawley and dark Agouti rats.

Urinary excretion of amphetamine and 4'-hydroxyamphetamine has been studied in male and female Sprague Dawley (SD) and Dark Agouti (DA) rats. The DA rat is an animal model for the cytochrome P450 (P450) 2D poor metabolizer. Rats were given d-amphetamine sulfate (5 mg/kg, i. p.) and urines were collected at 12 hour intervals for extraction and analysis of the amphetamines by HPLC. There was no significant difference between the sexes of either SD and DA rats in urinary 4'-hydroxyamphetamine and amphetamine excretion, but significant differences were seen between the two strains. The percentage of dose per ml urine recovered as 4'-hydroxyamphetamine from the urine over 24 hours was 11.1 and 9.1 in the SD male and female rats, and 2.3 and 2.5 in DA male and female rats, respectively. The percentage of dose per ml urine recovered as amphetamine was correspondingly lower in the SD male and female rats, 1.1 and 1.0, than that of the DA male and female rats, 5.9 and 5.0. These results support our hypothesis that P450 2D is involved in hepatic 4'-hydroxylation of amphetamine in rats.

Amphetamine↗
PubMed1993

Determination of tolbutamide hydroxylation in rat liver microsomes by high-performance liquid chromatography: effect of psychoactive drugs on in vitro activity.

A simplified HPLC method for tolbutamide metabolism to hydroxytolbutamide has been used to screen sixty psychoactive drugs for their ability to inhibit rat liver microsomal tolbutamide hydroxylation. One-step extraction with diethyl ether was followed by reconstitution and isocratic HPLC analysis with a binary mobile phase (ammonium phosphate:methanol, 45:55, v/v). Nanogram amounts of hydroxytolbutamide formation were estimated with UV detection at 240 nm. Hydroxytolbutamide formation was linear with incubation times of 40-120 min, but specific activity increased with increases in microsomal protein (0.15-1.10 mg). A differential inhibitory response was demonstrated for tolbutamide and debrisoquine hydroxylation to 5 psychoactive drugs, suggesting that tolbutamide hydroxylation is not dependent on P4502D1. Sixty psychoactive drugs, or drug metabolites, (at 33 microM) were then co-incubated with tolbutamide (at 2.5 and 10.2 microM). Tolbutamide hydroxylation was refractory (< 25% inhibition) to twenty-four of the drugs and only mildly inhibited (25-50% inhibition) by twenty-eight. Two compounds, trans-3-methylfentanyl and flurazepam, produced > 50% inhibition that was independent of tolbutamide concentration. Five of the drugs (methadone, chlorpheniramine, meperidine, 6-monoacetylmorphine and methylphenidate), however, caused greater than 50% inhibition in a competitive manner which suggests these drugs may share an affinity for the substrate binding site for tolbutamide.

Animals↗
PubMed1993

Immunoassay detection of nordiazepam, triazolam, lorazepam, and alprazolam in blood.

The ability of commercial benzodiazepine immunoassays to detect nordiazepam, lorazepam, alprazolam, and triazolam in blood samples was investigated. Two radioimmunoassays (RIA) (Abuscreen [RIA-a] and the Diagnostic Products Corporation serum kit [RIA-d]), two enzyme immunoassays (EIA) (Emit d.a.u. [EIA-u] and Emit TOX serum assay [EIA-s]), and two fluorescence polarization immunoassays (FPIA) (X-systems urine [FPIA-u] and X-systems serum assay [FPIA-s]) were evaluated for their ability to detect benzodiazepines in fortified drug-free human or bovine blood. Prior extraction of the blood was necessary for analysis on the equipment used for EIA and FPIA. Extraction with an organic solvent, such as butyl chloride used in this study, was preferable to precipitation with methanol or zinc sulfate. For all these assays, extraction eliminated matrix effects and offered the possibility of increased sensitivity by reconstitution of the extract in a smaller volume. Extraction was necessary in bovine blood, as there is some nonextractable substance(s) in this matrix that increases non-specific binding. Using extraction with reconstitution in one-half the original volume, the apparent limit of detection for nordiazepam in blood ranged from 3 ng/mL, with RIA-d, to 30 ng/mL, with EIA-s and FPIA-s. These limits of detection were improved by further reduction of the reconstitution volume. RIA-a, EIA-s, and FPIA-s had cross-reactivity for alprazolam that was equivalent to or slightly better than nordiazepam. RIA-d had enhanced cross-reactivity for alprazolam at 30-300 ng/mL.(ABSTRACT TRUNCATED AT 250 WORDS)

Alprazolam↗
PubMed1993

Selective induction of rat liver phase II enzymes by N-heterocycle analogues of phenanthrene: a response exhibiting high correlation between UDP-glucuronosyltransferase and microsomal epoxide hydrolase activities.

1. Among nitrogen heterocycles based on the planar phenanthrene structure are three (1,7- and 4,7-phenanthroline and phenanthridine) which selectively increase rat hepatic phase II drug metabolizing enzyme activities without increasing cytochrome P450 concentration. Of six monooxygenase activities investigated, only ethoxyresorufin dealkylase was raised but this was only minor. 2. The detergent-activated UDP-glucuronosyltransferase activities towards morphine, 4-nitrophenol, and 1-naphthol were increased up to five-, three- and two-fold of control respectively. Microsomal epoxide hydrolase activity towards cis-stilbene oxide was increased up to three-fold and cytosolic glutathione S-transferase activity towards 1-chloro-2, 4-dinitrobenzene reached twice the control value. 3. Cytosolic 4-nitrophenol sulphotransferase activity was not increased by any compound and like some monooxygenase reactions, was decreased by 4,7- and 1,7-phenanthrolines. 4. 1,10-Phenanthroline and two compounds which lack a heterocyclic nitrogen atom, (phenanthrene and 9-phenanthrol), failed to elicit any induction of enzyme activities. 5. Changes in microsomal epoxide hydrolase activity showed high correlation (r = 0.97) with changes in UDP-glucuronosyltransferase (4-nitrophenol) activity.

Animals↗
PubMed1992

Gas chromatography/mass spectrometry assays for the determination of debrisoquine and sparteine metabolites in microsomal fractions of rat liver.

Debrisoquine and sparteine are prototype substrates of a genetic deficiency in cytochrome P450-dependent drug metabolism. Sensitive assays of in vitro oxidation of sparteine and debrisoquine are required for evaluation of this polymorphism. The activities were measured by quantitative analysis of 2-dehydrosparteine and 4-hydroxydebrisoquine production, respectively, using capillary column gas chromatography coupled with mass selective ion detection. With a single extraction, separation of parent drug, metabolite, and a suitable internal standard was readily achievable. Time-dependent production of both metabolites could be detected from as little as 40 micrograms of microsomal protein. Both activities showed a maximal activity with a 240-min incubation period. The ability to simultaneously quantify the parent drug and its metabolite suggests it would also be useful for evaluation of in vivo metabolism.

Animals↗
PubMed1992

Effect of phenobarbital treatment on carbon tetrachloride-mediated cytochrome P-450 loss and diene conjugate formation.

The effect of phenobarbital treatment on the linkage between carbon tetrachloride-mediated cytochrome P-450 loss and lipid peroxidation in rat liver microsomes was studied. Male Sprague-Dawley rats, pretreated with 3 daily i.p. doses of phenobarbital (50 mg/kg) or saline, were orally dosed with carbon tetrachloride (0.01-2.5 ml/kg), with liver microsomes prepared at 7.5-180 min after carbon tetrachloride treatment. In vivo cytochrome P-450 loss displayed pseudo-first-order kinetics, and the initial rates of diene conjugate formation were saturable with dose. Phenobarbital pretreatment decreased the in vivo t0.5,max from 27.0 to 15.6 min, and increased the Kd,app from 0.78 to 1.30 ml/kg for carbon tetrachloride mediated cytochrome P-450 loss. Phenobarbital had no effect on the in vivo Vmax (1.03 to 1.04 delta OD232 nm/min/mg phospholipid) for carbon tetrachloride mediated diene conjugate formation, but decreased the Km,app from 0.22 to 0.10 ml/kg. These results are consistent with destruction of cytochrome P-450 heme resulting from a metabolite which does not leave the site of generation, and with phenobarbital pretreatment enhancing the initiation of lipid peroxidation.

Animals↗
PubMed1992

Concentration of plasma cholesterol in beef cows and calves, milk production and calf gain.

Milk yield of 59 beef cows that calved in late September through November was measured monthly in early and late lactation and biweekly during midlactation. Milk yield was estimated by milking with a machine after over-night separation of cows from calves. Concentration of plasma cholesterol of cows and calves was measured when calves averaged 44, 93, 136, and 178 d of age (SD = 17 d). Cholesterol of calves also was measured 2 wk after weaning, when they averaged 220 +/- 2.3 d of age. Cholesterol of calves was highest at second and third samplings and dropped after weaning. The estimated intake of milkfat by calves, and to a lesser extent their intake of milk, was related positively to their plasma cholesterol as they approached weaning age. The relationship was not strong enough, however, for differences among calves in concentration of plasma cholesterol to identify accurately differences in milk yield of their dams. Within breed group, age of cow, and stage of lactation, the regression coefficients of milk yield on plasma cholesterol of cows were close to zero. The concentrations of plasma cholesterol in both cows and calves were highly repeatable, with the exception of samples that were collected when calves averaged 44 d of age. Although plasma cholesterol of calves was related positively to milk yield and milkfat yield in late lactation, the former trait was not an accurate indicator of the two latter traits.

Animals↗
PubMed1991

Effects of environmentally encountered epoxides on mouse liver epoxide-metabolizing enzymes.

Male mice were treated (i.p.) for 3 days with 15 different environmentally encountered epoxides, and the effects of these compounds on liver microsomal and cytosolic epoxide hydrolase (mEH and cEH), glutathione S-transferase (mGST and cGST) and carboxylesterase (mCE) activities were determined. The epoxides included the pesticides: heptachlor epoxide, dieldrin, tridiphane, and juvenoid R-20458; the natural products: disparlure, limonin, nomilin, and epoxymethyloleate; the endogenous steroids: lanosterol epoxide, cholesterol-alpha-epoxide, and progesterone epoxide; and the industrial or synthetic epoxides: epichlorohydrin, araldite, trans-stilbene oxide, and 4'-phenylchalcone oxide. The pesticide epoxides were the most effective inducers of liver weight, microsomal protein, and the enzyme activities measured, with mEH and cEH activities towards cis-stilbene oxide (mEHcso and cEHcso), cGST activities towards four of five substrates, and mCE towards clofibrate (mCEclof) and p-nitrophenylacetate (mCEpna) increased following treatment with most of the pesticides. The synthetic epoxides increased some of the same activities, while the natural products, except for increases in cGST activities, and endogenous steroid epoxides were generally not inductive. cEH activity towards trans-stilbene oxide (cEHtso) was increased only following treatment with the peroxisome proliferator, tridiphane, but decreased following treatment with several of the epoxides, while microsomal cholesterol epoxide hydrolase (mEHchol) was increased only moderately by disparlure. Microsomes could effectively conjugate glutathione to chlorodinitrobenzene (mGSTcdnb) and cis-stilbene oxide (mGSTcso). These two activities were differentially induced by a few of the epoxides, suggesting that they may be selective substrates for different isozymes of mGST. Correlation coefficients were determined for the relative response of liver weight, subfraction protein, and enzyme activities. A relatively high correlation was found between the response of liver weight and cytosolic hydrolysis of trans-stilbene oxide (r = 0.73) and cis-stilbene oxide (r = 0.62), and cytosolic glutathione conjugation of dichloronitrobenzene (r = 0.66) and trans-stilbene oxide (r = 0.75). In addition, relatively high correlations were found between the different cGST activities, in particular for dichloronitrobenzene with trans-stilbene oxide (r = 0.89). These studies show that there exists a wide variation in the response of xenobiotic-metabolizing enzymes to environmentally encountered epoxides and that a fairly strong correlation exists between the increases in liver size and increases in certain cytosolic enzyme activities; they also suggest further studies concerning the possibility of an additional isozyme of mGST.

Animals↗
PubMed1991

The effect of structurally divergent herbicides on mouse liver xenobiotic-metabolizing enzymes (P-450-dependent mono-oxygenases, epoxide hydrolases and glutathione S-transferases) and carnitine acetyltransferase.

Male mice were treated with structurally diverse herbicides to study their effect on liver xenobiotic-metabolizing enzymes. Chlorfiurecol, trifluralin, alachlor, propham, MCPP and 2,4-DP caused increases in phase I (cytochrome P-450, ethoxycoumarin O-deethylase, and/or aminopyrine N-demethylase) and phase II (microsomal epoxide hydrolase and cytosolic glutathione S-transferase) activities. MCPP and 2,4-DP also increased cytosolic epoxide hydrolase and carnitine acetyltransferase activities suggestive of peroxisome proliferation. Benthiocarb and molinate increased only some phase II enzyme activities. Dicamba, at the dose employed, caused mortality and decreases in some of the enzymes monitored. Most of the herbicides tested induced xenobiotic-metabolizing enzyme activities, the pattern of induction being dependent on herbicide structure.

Animals↗
PubMed1991

The effect of tetrahydrofuran on biological systems: does a hepatotoxic potential exist?

Tetrahydrofuran (THF) is a widely used solvent in industry and research. THF is a weak toxin, with approximate acute LD50s in the range of 2 to 3 g/kg, 8 to 20 mg/L, and 800 mg/kg following oral, inhalation, and i.v. routes, respectively. The two primary signs of intoxication are narcosis and hepatocellular dysfunction, both occurring at doses of approximately one-half of the lethal dose. Little is known concerning the pharmacokinetics of THF. No evidence exists for genotoxicity due to THF. Ongoing carcinogenicity bioassay tests sponsored by the National Toxicology Program have not yet been completed. Two significant interactions of THF with cellular components have been studied; first, THF inhibits a number of enzymatic reactions at concentrations ranging from 10 to 100 mM. Most notably, THF is an inhibitor of a number of cytochrome P450 (P450) dependent mixed function oxidase activities, with a particular affinity for the alcohol-induced isozyme (P450IIE1). Second, THF has been noted to enhance the toxic action of a number of compounds (i.e. solvent effect), in particular stimulating the more rapid absorption of reactive metabolites. A third factor to be considered is THF's ability to form peroxides upon standing. Little is known concerning the toxicity of THF peroxides. Therefore, while there is little evidence to suggest that THF would be a direct (Type I) hepatotoxin at relatively low doses, its ability to inhibit drug-metabolizing reactions (perhaps more so in alcohol users), and enhance the absorbance of reactive metabolites, are relatively unexplored avenues for THF to contribute to a hepatotoxic response. As a great deal of the human hepatotoxicity (Type II) is "idiosyncratic" and cannot be readily reproduced in experimental animals, this potential route of hepatotoxicity should be considered in future evaluations of human exposure to THF.

Administration, Inhalation↗
PubMed1991

Drug and alcohol involvement in railroad accidents.

Postaccident testing of railroad employees for drugs and alcohol was performed in 175 qualifying accidents or incidents (events) from April 1987 through March 1988. Initial tests for drugs were performed on urine, and for alcohol on blood. Presumptive positives were confirmed and quantitated using both blood and urine when available. In 42 of these events (24%), at least 1 employee tested positive (6.7% of 736 employees) for drugs or alcohol. A higher proportion of drug or alcohol-positive events, or both, 40 versus 21%, was found when a fatality was involved. In 11 of the 32 fully investigated drug or alcohol-positive events (involving 14 employees), the investigating agencies determined that substance use was determined to be a probable cause of, or a factor relating to, the accident. Cannabinoids, ethanol, cocaine, or multiple-drug use were found in 5, 3, 3, and 3 of those employees, respectively. Detectable drug and alcohol use occurs among railroad employees; occasionally it has resulted in accidents.

Accidents↗