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This study assessed the effect of food restriction on the metabolism of model monooxygenase substrates in the perfused rat liver. Female Sprague-Dawley rats has access ad lib. to a Purina 5001 nonpurified diet (control) or were given 65% of the intake of controls for 3 weeks. Livers were perfused with oxygenated Krebs-Henseleit buffer using a non-recirculating system, and the rates of monooxygenation of p-nitroanisole and 7-ethoxycoumarin were measured. The results indicate that food restriction stimulated p-nitroanisole O-demethylation from 2.9 +/- 0.2 to 4.6 +/- 0.5 mumol/(g.hr) when saturating concentrations of p-nitroanisole were infused. Concomitantly, the ratio of beta-hydroxybutyrate to acetoacetate (B/A) and the rates of ketogenesis (B + A) were increased significantly by food restriction. Further, p-nitroanisole (200 mumol/L) increased hepatic malate concentration nearly 3-fold in liver extracts from food-restricted rats. However, infusion of either a low concentration of p-nitroanisole (50 mumol/L) or 7-ethoxycoumarin (200 mumol/L) did not alter these parameters. On the other hand, food restriction did not alter rates of monooxygenation in isolated microsomes supplemented with excess NADPH. Taken together, these data support the hypothesis that high concentrations of p-nitroanisole increased monooxygenation in food-restricted rats by stimulating fatty acid oxidation, which elevates the mitochondrial NADH/NAD+ ratio. This, in turn, increases the availability of reducing equivalents in the form of NADPH by a malate-pyruvate exchange system, leading to increased drug metabolism.

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1. We have examined the effects of (S)-(+)-6-(2-chlorophenyl)-3-cyclopropanecarbonyl-8, 11-dimethyl-2,3,4,5-tetrahydro-8H-pyrido[4',3':4,5]thieno[3, 2-f][1, 2, 4]triazolo[4, 3-a][1, 4]diazepine (E-6123), a novel thienodiazepine platelet-activating factor antagonist, on drug-oxidizing capacity in beagle dog, using antipyrine (AP) and trimethadione (TMO) as two model substrates. 2. The plasma half-life (t1/2) and area under the curve (AUC) of AP (0.5 mg/kg, i.v. injection) increased in a dose-dependent manner after a single oral dose of E-6123 (0.2, 1 or 10 mg/kg), whereas the total body clearance (Cl) of AP was decreased, and the apparent volume of distribution (Vd) was unchanged. 3. The pharmacokinetic parameters (t1/2, Cl and AUC) of the metabolism of TMO (4 mg/kg, i.v.) after repeated oral administration of E-6123 (10 mg/kg for 7 days) were not significantly changed in comparison with findings in control dog. The ratio of dimethadione (DMO), being the only TMO metabolite, to TMO in plasma after i.v. administration of TMO in E-6123-treated dog was increased only 5 and 15 min after the final dose, but was not changed at other sampling times (0.5, 1, 2 4, 6, 8 and 12 h). 4. The content of b5, the activity of p-nitroanisole O-demethylase and benzphetamine N-demethylase were significantly increased, compared with controls, by repeated E-6123 treatment. However, aniline hydroxylase activity was not significantly changed. 5. Content of P450 2B was significantly increased in E-6123 treated dog, while that of 3A was not.(ABSTRACT TRUNCATED AT 250 WORDS)

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1. We examined the effects of N-methoxy-3-(3,5-di-tert-butyl-4- hydroxybenzylidene pyrrolidin-2-one (E-5110), a novel non-steroidal anti-inflammatory drug, on the pharmacokinetics of trimethadione (TMO) and characterized the P450 isozymes involved in the metabolism of TMO in beagle dog. 2. In the E-5110-treated dog (50 mg/kg/day for 7 days: oral) the plasma half-life (t1/2) and the area under the curve (AUC) of TMO (4 mg/kg, i.v.) in vivo were decreased, and total body clearance (CL) was increased; the apparent volume of distribution (Vd) was relatively unchanged. 3. Contents of P450 and b5, and the activity of p-nitroanisole O-demethylase and benzphetamine N-demethylase in vitro were significantly increased compared with controls by repeated E-5110 treatment in dog. 4. Contents of CYP2B and 3A were increased by E-5110 pretreatment in dog. 5. TMO N-demethylation was inhibited by the anti-CYP2B and 3A IgG fractions in liver microsomes obtained from the E-5110-treated dog. 6. Results of both the in vivo and in vitro studies of the effects of E-5110 treatment in dog on TMO indicate that these effects may be attributed to the induction of CYP2B and 3A.

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The content of cytochrome P-450 in liver microsomes from guinea pigs was decreased by ascorbic acid-deficiency. Since ascorbic acid is an antioxidant in vivo, the possible involvement of lipid peroxidation in this phenomenon was investigated. In fact, the level of lipid peroxides in liver homogenates of guinea pigs was increased by ascorbic acid deficiency. The level was significantly decreased when the animals were given tocopherol acetate (25 mg/kg/day, s.c.) with an ascorbic acid-free diet. The activities of aminopyrine N-demethylase, aniline hydroxylase, p-nitroanisole O-demethylase and 7-ethoxycoumarin O-deethylase, and the content of cytochrome P-450 spectrally determined did not restore the control level by the administration of tocopherol acetate to the ascorbic acid-deficient animals. Western blot analysis of liver microsomes with antibodies to rat P-450IA2 (P-448-H), P-450IIB1 (P-450b) and human P-450IIIA4 (P-450NF) showed that ascorbic acid-deficiency resulted in a decrease in the amount of cytochrome P-450 immunochemically related to P-450IA2, but not the amounts of the forms of cytochrome P-450 cross-reactive with antibodies to P-450IIB1 and P-450IIIA4. The reduced amounts of cytochrome P-450 cross-reactive with antibodies to rat P-450IA2 in liver microsomes of ascorbic acid-deficient animals remained unchanged even when lipid peroxidation was inhibited by tocopherol acetate, suggesting that there is a mechanism(s) other than lipid peroxidation involved in the reduction of amounts of cytochrome P-450 by ascorbic acid deficiency.

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The effects of pyrethroid insecticides on hepatic microsomal enzymes were studied in rats. Animals were treated orally with cypermethrin (80 mg/kg), deltamethrin (15 mg/kg), and permethrin (100 mg/kg), as a solution in soyabean oil, for 1 to 20 days. The content of cytochromes P-450 and b5, activity of NADPH cytochrome P-450 reductase, glutathione S-transferase, aniline 4-hydroxylase, p-nitroanisole O-demethylase in microsomes, the activity of glutathione S-transferase, and the level of sulfhydryl groups in cytosol were determined. Also the relative liver weight was measured. Only few changes in the investigated parameters were ascertained. These changes have an irregular and transient character. On the whole, the action of pyrethroids on microsomal enzymes results in a slight induction.

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A 2-year toxicity/oncogenicity study was done to evaluate the potential effects of the leukotriene antagonist LY171883 in B6C3F1 mice. Dietary concentrations of LY171883 during the initial 7 months of the study were 0.0, 0.005, 0.015, or 0.05% but were increased to 0.0, 0.0075, 0.0225, or 0.075% during Months 7 through 24. The estimated average daily compound intake was 0.0, 7.3, 22.5, or 80.5 mg/kg for males and 0.0, 9.2, 27.5, or 95.9 mg/kg for females. Survival was not adversely affected by treatment, however, body weight of males and females in the high dose group was significantly lower than that of controls. The chronic toxicity was localized primarily to the liver. Liver weights were increased in males in the high dose group and in females in the mid and high dose groups. Microsomal p-nitroanisole-O-demethylase activity was increased in mid and high dose females. Hepatic peroxisomal beta-oxidation was increased approximately twofold in both sexes in the high dose group only. Centrilobular eosinophilic granular change of hepatocytes was a common histopathologic finding in male and female mice in the high dose group, with the incidence and severity being greater in females. An increased incidence of hepatocellular carcinomas was observed in female mice in the mid and high dose groups. The number of male mice in the high dose group with hepatocellular carcinomas was higher than that of controls but the change was not statistically significant. Hepatocellular adenomas were increased in females in the high dose group but not in males. All groups of treated females had increased nodular hepatocellular hyperplasia.(ABSTRACT TRUNCATED AT 250 WORDS)

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Five benzodiazepines (diazepam, oxazepam, clonazepam, nitrazepam and chlordiazepoxide) were compared with respect to their capacity to affect microsomal aniline hydroxylase, aminopyrine N-demethylase and 4-nitroanisole O-demethylase of rats exposed to the high ambient temperature of 28 degrees C or 35 degrees C. We have stated that the highest effect on the microsomal enzyme activities was observed after chlordiazepoxide, clonazepam or nitrazepam treatments. These results indicate that the presence of nitro group at position 7 or N-oxide at position 4 of benzodiazepine is important for the ability of benzodiazepines to affect the hepatic microsomal enzymes. High environmental temperature modifies the effect of benzodiazepines on tested microsomal enzymes.

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The aim of this study was to investigate the activity of the mixed function oxidases in the liver of the rat during the first days of extrahepatic cholestasis. In particular, contradictory results of previous studies of the metabolism of type I- and type II-substrates should be examined. The activities of the aminopyrine demethylase, aniline hydroxylase and nitroanisole-O-desalkylase were measured in the liver microsomes after sham operation and on the first and third day after ligature of the common bile duct. On the first day of cholestasis both the metabolism of the type I-substrate (aminopyrine) and of the type II-substrates (aniline, nitroanisole) was significantly reduced. On the third day the metabolism of type II-substrates reached nearly normal levels, whereas the metabolism of aminopyrine was even further reduced. Contradictory results of previous studies seem to be due to different times of measurement.

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1. A study of the sex- and age- (4 to 103 weeks) related changes in liver microsomal mixed-function oxidase components of Sprague-Dawley rats, showed that total cytochrome P-450 ranged from 12.0 to 34.0 nmol/g, peaking between weeks 26 and 39; males had higher levels than females at weeks 4 to 51. Cytochrome b5 ranged from 5.0 to 15.0 nmol/g with inconsistent age- and sex-related differences. 2. NADPH:cytochrome C reductase ranged from 2.6 to 4.8 mumol/min per g, with maximal activity at week 39; males had greater activity at weeks 12 to 78. NADH:cytochrome C reductase ranged from 5.2 to 26.9 mumol/min per g, peaking between weeks 51 and 78; females had greater activity at weeks 12 to 51. 3. This age- and sex-related pattern of changes in these components is specific to the Sprague-Dawley strain. There are slight but significant differences between sexes only in rats less than 1 year old. There was no significant correlation between rates of p-nitroanisole or aniline metabolism and cytochrome P-450 concentrations.

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In a chronic study by the National Toxicology Program (NTP), dimethyl hydrogen phosphite (DMHP) caused neoplastic and nonneoplastic changes in the lungs and forestomach of F344/N rats following gavage administration for 2 years. The current investigation was designed to study the effect of a short-term exposure on a series of biochemical systems in target and nontarget tissues which may be involved in the metabolism and/or the manifestation of DMHP toxicity. Rats were treated daily with a dose similar to that used in the NTP study (200 mg/kg) for 4, 5, or 6 weeks. Two groups of animals were also treated for 4 weeks and then treatment was discontinued and the rats were allowed to recover for 1 or 2 weeks. An equal number of animals was treated similarly with the vehicle and used as control. The microsomal and soluble fractions were separated from liver, lungs, kidneys, forestomach, and glandular stomach from the 6-week treatment group. Another group of rats treated for 6 weeks was prepared for pathology examination of the lungs, forestomach, and glandular stomach. There was a significant increase in the weight of the forestomach of rats treated for 4, 5, or 6 weeks relative to control animals, while no significant difference was observed in the weight of liver, lungs, kidneys, and glandular stomach. The forestomach weight of rats treated for 4 weeks returned to the control value after 1 week of recovery. Microscopic examination of the forestomach of rats treated for 6 weeks revealed a thickened stratified squamous epithelium characterized by hyperplasia, hyperkeratosis, and subepithelial inflammation and edema. There were no microscopic changes in the lungs or glandular stomach of animals treated for 6 weeks. The activity of angiotensin converting enzyme in the serum of rats treated for 4, 5, or 6 weeks was significantly increased over that of control animals. The activity of this enzyme returned to near levels seen in the control animals after 1 week of recovery following 4 weeks of treatment. No treatment-related effect was observed in the activities of the microsomal p-nitroanisole demethylase, soluble glutathione S-transferase, and soluble superoxide dismutase in the five tissues studied. There was a significant increase in the level of nonprotein soluble sulfhydryls in the forestomach but in no other tissue of rats treated for 6 weeks. Also the activity of soluble carboxylesterase was significantly reduced in the lungs and forestomach, but not in any other tissue of the 6-week-treated rats.(ABSTRACT TRUNCATED AT 400 WORDS)

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The effects of delta 8-tetrahydrocannabinol (delta 8-THC) and its major and active metabolite, 11-hydroxy-delta 8-tetrahydrocannabinol (11-OH-delta 8-THC), on the hepatic microsomal drug-metabolizing enzyme system were studied in mice. The repeated administration of 11-OH-delta 8-THC (5 mg/kg/day, i.v.) for 3 or 7 days increased significantly the activities of aniline hydroxylase and p-nitroanisole O-demethylase. By the same treatment, cytochrome P-450 content (3 days) or NADPH-cytochrome c reductase activity (7 days) was also increased significantly. The treatment with delta 8-THC for 7 days (5 mg/kg/day, i.v.) significantly increased aniline hydroxylase only. 11-OH-delta 8-THC increased the Vmax, but not the Km, values for both drug-metabolizing enzymes, whereas delta 8-THC decreases significantly the Km value (270 microM) for p-nitroanisole O-demethylase as compared with the control (398 microM). Repeated administration of these cannabinoids for 7 days also increased the metabolism of delta 8-THC by hepatic microsomes; this was attributed to an enhanced formation of 11-OH-delta 8-THC. In contrast, microsomal formation of 7 alpha-OH-delta 8-THC was decreased significantly by treatment with delta 8-THC. 11-OH-delta 8-THC, but not delta 8-THC, treatment increased the metabolism of 11-OH-delta 8-THC by hepatic microsomes. These findings indicate that delta 8-THC and 11-OH-delta 8-THC treatment can induce hepatic microsomal drug-metabolizing enzymes and affect differently the catalytic properties of the enzymes.

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The drug-metabolizing activities (aminopyrine N-demethylase and p-nitroanisole O-demethylase activities) have been measured at monthly intervals throughout the year in liver microsomes of male and female Japanese bullfrogs, Rana catesbeiana. The aminopyrine N-demethylase activity based on cytochrome P-450 of both sexes was significantly higher in May-July (spring-summer) than in August-October (summer-autumn). On the contrary, the p-nitroanisole O-demethylase activity based on cytochrome P-450 of males was significantly higher in July-October (summer-autumn) than in May-June (spring). However, there was no seasonal changes in the O-demethylase activity in females. There were significant differences between males and females in the N-demethylase activity in August-October and in the O-demethylase activity in May-June (or July-October).

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Hepatic monooxygenase activities of the mountain vole, Microtus montanus, were measured after i.p. injections of phenobarbital, B-naphthoflavone and Aroclor 1254 at doses ranging from 5 to 80 mg/kg. The results showed that mountain voles differed in their induction of hepatic monooxygenase activity relative to other rodents. The results also suggest substrate specificity in the detection of enzymatic induction and the importance of considering effects of varying inducer doses on hepatic monooxygenases.

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High-performance liquid chromatographic techniques were developed for the simultaneous detection of metabolites in a cytochrome P-450 model system composed of NADH, haemoglobin and methylene blue. Monohydroxylated metabolites were determined following aniline, acetanilide and phenol hydroxylations. 4-Aminoantipyrine, 7-hydroxycoumarin and p-nitrophenol were determined after dealkylation of 4-N,N-dimethylamino-antipyrine, 7-ethoxycoumarin and p-nitroanisole. These substrates are commonly used for measuring cytochrome P-450 activities. Treatment of the samples was minimal, consisting of a simple deproteinization, and did not involve any organic extraction. Separations were carried out on reversed-phase columns and the products were detected by UV adsorption. Separations were completed in less than 15 min and the detection limits were between 0.5 and 4 microM.

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Rats were pretreated with phenobarbital to induce hepatic cytochrome P-450. Compared to noninduced rats, a similar relation between the dose of acetaminophen and mortality, and between dose and changes in liver function (prothrombin index) and identical time courses, was found. The urinary excretion of acetaminophen mercapturate and acetaminophen cysteine was identical in induced and noninduced rats. The metabolism of acetaminophen in terms of blood levels and excreted metabolites was not influenced by phenobarbital induction. At the same dose level, hepatic necrosis was accelerated (maximum 24 h) compared to noninduced animals (maximum 72 h), but no difference in the maximum extent was found. These data cannot support the concept that induction of cytochrome P-450 leads to greater formation of the hypothetical toxic metabolite of acetaminophen, or that induction enhances its hepatotoxicity, in the rat. Several factors may contribute to accelerate the necrotic changes which make it possible to histologically identify cell damage and death. In that case, functional studies are more relevant than morphological evaluation in quantitative assessment of liver damage.

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Intraperitoneal injection of rats with 2 mg/kg ring labelled 14C AFB1 (spec. act. 110 mCi/mM/nmole) showed a higher level of radioactivity in the urine of test animals on diets containing 600 mg/kg vit. E 24 h after pretreatment. Analysis of the urine by chloroform extraction, thin layer chromatography and liquid scintillation counting of the various fractions showed less aflatoxin M1 (AFM1) and less unmetabolized AFB1 in test samples than in controls. Incubation of ring labelled 14C AFB1 with hepatic 10,000 g supernatant fractions, however, showed increased metabolism of AFB1 by fractions from test animals as compared with the controls. Rate of disappearance of 14C AFB1 and the consequent formation of AFM1 was greater in the test fractions than in the controls. At 30 days all test animals showed higher levels of serum vitamin E than the controls. Hepatic aniline hydroxylase and ethyl morphine N-demethylase activities of the liver fractions and blood glutathione reductase activity were greater in the tests. P-nitroanisole-O-demethylase activity was reduced while hepatic and serum reduced glutathione levels remained basically unaltered.

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A study was made of the effect of X-radiation of different doses on the content of P-450 cytochrome in a microsomal fraction of rat liver. When the haemoprotein level markedly decreased an increase in Km and a decrease in Vmax were noted in the reaction of O-demethylation of para-nitroanisole by microsomes of the irradiated rat liver. It is suggested that one of the cause of the effect observed is the postirradiation change in the composition of cytochrome P-450 pool resulting from a selective decrease in the level of the radiosensitive forms of haemoprotein.

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