Microsomal Mixed-function Oxidase Research Articles

Liver Microsomal Mixed-Function Oxidases in Response to Dietary n-3 Fatty Acid Levels in Rats

Phenobarbital (PB)-induced activities of the liver microsomal mixed-function oxidase (MFO) system in response to dietary levels of n-3 fatty acids were investigated in rats. The MFO activities were assessed by the contents of cytochromes P-450 and b5, and the activities of NADPH-cytochrome c reductase, aminopyrine N-demethylase and aniline hydroxylase. A linoleic acid (C18:2, n-6; LA) diet (2.0 energy % as LA) and n-3 fatty acid diets (0.3, 1.0, 1.9 or 4.8 energy % as n-3 fatty acids) were fed to rats for 16 days. The activities of MFOs except that of NADPH-cytochrome c reductase increased, as compared with the LA diet, with increasing the dietary n-3 fatty acid level, concomitant with the increase in n-3 fatty acids and decrease in n-6 fatty acids in liver microsomal lipids. The dietary LA over 1.8 energy % up to 13.8% did not increase the activities of PB-induced MFOs. Consequently, dietary n-3 fatty acids further elevated the PB-induced MFO activities effectively. These results suggest a differential response of dietary n-6 and n-3 fatty acids to the PB-induced MFO activities.

Liver microsomal mixed-function oxidases in response to polyunsaturated/saturated and n-6/n-3 fatty acid ratios of dietary lipids in rats.

The effects of various polyunsaturated/saturated (P/S; 0.29-2.74) and n-6/n-3 (0.49-11.21) ratios of dietary fatty acids and of phenobarbital (PB) stimulation on the activity of liver microsomal mixed-function oxidase (MFO) system were studied in rats using a combination of palm oil, safflower oil and fish oil concentrate. When the n-6/n-3 ratio was kept constant (4.6-4.9) without induction by PB, the highest P/S ratio (2.74) tended to increase the basal PB-uninduced MFO activities; while the PB-induced MFO activities were elevated as the dietary P/S ratio increased. This definitely indicates the role of both n-6 and n-3 polyunsaturated fatty acids (PUFAs) in the elevation of MFO activities with and without PB stimulation. Whereas the n-6/n-3 ratio was varied while maintaining a constant P/S ratio (1.3), the PB-induced MFO activities were significantly elevated by decreasing the n-6/n-3 ratio although the MFO activities were not affected without PB stimulation. The MFO activities induced by PB were directly related to the n-3 PUFA content and inversely related to the n-6 PUFA content of the diets with n-6/n-3 ratios less than 4.55. These results obtained herein suggest that there are significant differences in the effects of n-6 and n-3 PUFAs on liver microsomal MFO activities, especially in relation to the induction of MFOs by PB.

Effect of Propylthiouracil on Hepatic Microsomal Mixed Function Oxidase and Lipid Deposition in Broiler Chicks Fed Diets with Different Fat Content.

Effect of Propylthiouracil on Hepatic Microsomal Mixed Function Oxidase and Lipid Deposition in Broiler Chicks Fed Diets with Different Fat Content.

Effect of Dietary Alpha-Linolenic, Eicosapentaenoic and Docosahexaenoic Acids on Liver Microsomal Mixed-Function Oxidases in Rats

This study was undertaken to compare the effects of three major dietary n-3 fatty acids, i.e. alpha-linolenic acid (C18:3, n-3; LN), eicosapentaenoic acid (C20:5, n-3; EPA) and docosahexaenoic acid (C22:6, n-3; DHA), on liver microsomal mixed-function oxidase activities with and without phenobarbital (PB) induction. Male Sprague-Dawley rats were fed a 5% lipid diet containing either LN, EPA or DHA as major constituent fatty acid (about 40%) of dietary lipids for 15 days. The fatty acid compositions of liver microsomal lipids of each n-3 fatty acid group were characteristically different, particularly in the n-3 fatty acid compositions, but were nearly the same with and without PB treatment. Dietary LN, EPA and DHA did not affect the levels of PB-inducible cytochrome P-450 species nor the activities of aminopyrine N-demethylase and aniline hydroxylase. However, the PB-uninduced constitutive levels of cytochromes P-450 increased in the following order with a slight but significant difference between LN and DHA groups: LN group < EPA group < DHA group. Both the PB-uninduced activities of aminopyrine N-demethylase and aniline hydroxylase changed in a very similar manner to the constitutive levels of cytochromes P-450. These results suggest that the uninduced constitutive level of certain cytochrome P-450 species is modulated differentially by dietary LN, EPA and DHA, whereas dietary LN, EPA and DHA do not affect the PB-inducible levels of cytochrome P-450 species.

Dehydroepiandrosterone pretreatment protects rats against the pro-oxidant and necrogenic effects of carbon tetrachloride

A single intraperitoneal injection of dehydroepiandrosterone (3β-hydroxy-5-androsten-17-one, DHEA) 17 hr before carbon tetrachloride (CCl 4) poisoning protects rats against liver injury induced by the haloalkane. In liver homogenates, both the increase in malondialdehyde production and the formation of fluorescent lipid peroxidation products are significantly reduced. Also, liver microsomes obtained from DHEA-pretreated rats incubated in vitro with CCl 4 are less susceptible to lipid peroxidation than microsomes from normal animals. The release of liver enzymes into the blood is much reduced in DHEA-pretreated rats, confirming a cause-effect relationship between lipid peroxidation and hepatocyte death. Treatment with DHEA inhibits neither glucose-6-phosphate dehydrogenase activity in the cytosol, nor the microsomal mixed function oxidase system (cytochrome P450 content, aminopyrine demethylase and ethoxycoumarine de-ethylase activities). In animals treated with DHEA, the liver content of total glutathione and vitamin E is not modified. These results support the hypothesis that DHEA protects against CCL 4-induced liver injury through its own antioxidant activity, rather than by interfering with the metabolism of the toxin or with the tissue level of primary antioxidants.

Effect of 4-(diethylamino)benzaldehyde on ethanol metabolism in mice.

The compound 4-(diethylamino)benzaldehyde (DEAB) is a potent inhibitor of cytosolic (class 1) aldehyde dehydrogenase (ALDH) in vitro and can overcome cyclophosphamide resistance in murine leukemia cells characterized by their high content of ALDH. In this study, we examined the in vivo effect of DEAB in mice on ethanol metabolism and on antipyrine clearance as a measure of the microsomal mixed function oxidase activity. DEAB administered in doses of 50 and 100 mg/kg increased the blood acetaldehyde concentration and decreased the plasma acetate concentration in mice treated with ethanol. A pharmacokinetic approach demonstrated that DEAB in doses of 50 and 100 mg/kg inhibited the fraction of ethanol converted to acetate by 32.5 and 67.5%, respectively. This inhibition was comparable with that produced by disulfiram. DEAB produced optimal inhibition of ALDH 10-15 min after administration. DEAB did not change the half-life or the total clearance of antipyrine. We conclude that DEAB is a potent inhibitor of ALDH in vivo and has no effect on the mixed function oxidase activity as determined by antipyrine clearance.

Sex-related differences of hepatic microsomal mixed function oxidase system and antibody production in broilers implanted with corticosterone and/or fed ascorbic acid

1. Most of the components of the mixed function oxidase (MFO) in hepatic microsomes were reduced by corticosterone implants, and the degree of the reduction in females and at an older age was greater than those in males and at a younger age. 2. Ascorbic acid (AA) prevented the reduction in the MFO caused by corticosterone implants. 3. The activities of aniline hydroxylase and aminopyrine N-demethylase were enhanced by corticosterone implants regardless of AA supplementation. 4. The activity of NADPH-cytochrome c reductase in male broiler was greater than that in females under normal conditions. 5. Corticosterone implants and dietary AA had less influence on the antibody production, especially to T-cell dependent antigen.

Effects of a beta-adrenergic agonist (clenbuterol) on performance, carcase composition, hepatic microsomal mixed function oxidase and antibody production in female broilers treated with or without corticosterone.

1. Beta-adrenergic agonist (Clenbuterol, 0.33 mg/kg) and corticosterone (10 mg/kg) were incorporated into a diet based on maize and soybean meal. Their effects on performance, carcase composition, hepatic microsomal mixed function oxidase and antibody production were investigated in female broilers. 2. Dietary corticosterone reduced the titre to sheep red blood cells, while it was unchanged by clenbuterol. 3. Clenbuterol exerted a promoting effect on gain-to-food ratio, carcase protein and hepatic microsomal cytochrome P-450 content. 4. Addition of clenbuterol to the corticosterone-containing diet prevented the increase in abdominal fat weight and uric acid excretion induced by corticosterone, but did not affect total fat mass. 5. The results showed that clenbuterol reduced abdominal rather than carcase fat and prevented protein degradation in the body when chicks were treated with corticosterone. Clenbuterol also influenced the content of cytochrome P-450, but not the humoral immunity.

Response of liver microsomal mixed-function oxidases to dietary linoleic acid levels in rats.

Responses of the activities of liver microsomal mixed-function oxidases (MFOs) induced by phenobarbital (PB) to dietary linoleic acid (LA) levels were investigated in rats. Diets varied in LA content (0, 1.8, 4.1, 7.0, or 13.8 energy%) were fed to rats for 16 days. The activities of MFOs did not change significantly with increasing the dietary LA level, where they reached a plateau even at 1.8 energy %, although the fatty acid composition of liver microsomal lipids reflected well that of dietary lipids. Accordingly, 2 energy % of LA as essential fatty acid seemed to be sufficient to supply the dietary requirement as assessed by the activities of PB-induced MFOs.

Effect of anti-basic liver protein antibody-induced liver injury on hepatic drug-metabolizing enzymes in C57 BL/6J mice.

We attempted to produce a model mouse with a liver injury resulting from an immunological mechanism in C57BL/6J mice, and the effect of hepatitis on the hepatic microsomal mixed-function oxidase system was studied. An experimental immunological liver injury model was caused by the intravenous injection of an anti-basic liver protein (BLP) antibody in mice which had been previously immunized with normal rabbit IgG (RGG) and complete Freund's adjuvant. C57BL/6J strain mice showed the highest susceptibility to the immunological liver injury. Typical histopathological changes in the liver included submassive hepatocellular necrosis and infiltration of lymphocytes into the portal tract and sinusoid area in a necrotic lesion. The liver injury in this model was markedly inhibited by the administration of prednisolone (20 mg/kg, p.o.), cyclophosphamide (15 mg/kg, i.p.), levamisole (10 mg/kg, p.o.), glycyrrhizin (50 mg/kg, i.p.) and cepharanthine (10 mg/kg, i.p.), which act on the immune system. Twenty-four hours after the injection of anti-BLP antibody, the activities of aminopyrine N-demethylase, aniline hydroxylase and NADPH-cytochrome c reductase and the content of cytochrome P-450 were mostly reduced, whereas cytochrome b5 and NADH-ferricyanide reductase were not. These results suggest that the experimental liver injury model in C57BL/6J mice is useful as a model of liver injury model, and its hepatitis was shown to inhibit the cytochrome P-450-dependent biotransformation of drugs in the mouse.

Toxicity studies of a synthetic antioxidant, 2,2'-methylenebis (4-ethyl-6-tert-butylphenol) in rats. 2. Uncoupling effect on oxidative phosphorylation of liver mitochondria.

Effects of 2,2'-methylenebis (4-ethyl-6-tert-butylphenol) (MBEBP) on hepatic mitochondrial oxidative phosphorylation in vitro, and on hepatic peroxisomal enzymes activities and microsomal mixed-function oxidase activities were studied. 1. A low concentration of MBEBP, less than 50 microM, increased state 4 respiration and decreased state 3 respiration. However, a higher concentration of MBEBP, greater than 100 microM, acted as a respiratory inhibitor. Therefore, MBEBP was found to act as an uncoupler of oxidative phosphorylation in rat liver mitochondria. 2. MBEBP significantly decreased peroxisomal enzymes, cyanide-insensitive palmitoyl-CoA oxidizing activity and catalase activity in the livers of rats fed 0.2, 1.0 or 5.0% MBEBP for 4 weeks. 3. In microsomal enzyme assay, NADPH cytochrome c reductase activity was significantly increased, however, cytochrome P-450, cytochrome b5 levels, aminopyrine N-demethylase and benzo [a] pyrene hydroxylase activities were not significantly increased in the livers of rats fed 1.0 or 5.0% MBEBP for 4 weeks. The weight loss and the decrease of serum triglyceride level observed in the MBEBP-treated rats seemed to be caused by its uncoupling effects, which might also be the cause of the testicular damage induced by MBEBP.

Enhancement of the toxicity of methyl bromide toTribolium castaneum adults by its admixture with carbon Monoxide

The effect of methyl bromide in admixture with 20%, 30% and 40% carbon monoxide (CO) onTribolium castaneum adults was tested. The combination of methyl bromide with 20% CO increased the toxicity of CH3Br 1.4-fold, with 30% CO-1.5-fold, and with 40% CO-1.7-fold. It is supposed that the inhibitory effect of CO on the insect’s microsomal mixed-function oxidase and on other enzymatic systems causes this effect.

Inhibition of rat liver microsomal lipid peroxidation by boldine

The alkaloid boldine, found in the leaves and bark of boldo, was an effective inhibitor of rat liver microsomal lipid peroxidation under a variety of conditions. The following systems all displayed a similar sensitivity to boldine: non-enzymatic peroxidation initiated by ferrous ammonium sulfate; iron-dependent peroxidation produced by ferric-ATP with either NADPH or NADH as cofactor; organic hydroperoxide-catalyzed peroxidation; and carbon tetrachloride plus NADPH-dependent peroxidation. Boldine inhibited the excess oxygen uptake associated with microsomal lipid peroxidation. Thus, boldine was effective in inhibiting iron-dependent and iron-independent microsomal lipid peroxidation, with 50% inhibition occurring at a concentration of about 0.015 mM. Boldine did not appear to react efficiently with Superoxide radical or hydrogen peroxide, but was effective in competing for hydroxyl radicals with chemical scavengers. Concentrations of boldine which produced nearly total inhibition of lipid peroxidation had no effect on microsomal mixed-function oxidase activity nor did boldine appear to direct electrons from NADPH-cytochrome P450 reductase away from cytochrome P450. Boldine completely protected microsomal mixed-function oxidase activity against inactivation produced by lipid peroxidation. The effectiveness of boldine as an anti-oxidant under various conditions, and its low toxicity, suggest that this alkaloid may be an attractive agent for further evaluation as a clinically useful anti-oxidant.

L-Tryptophan alleviates fatty liver and modifies hepatic microsomal mixed function oxidase in laying hens

1. 1. Three experiments were conducted to study the effect of dietary l-tryptophan supplementation (250–1000 ppm) on lipid accumulation, an occurrence of hemorrhages and microsomal mixed function oxidase in the liver of laying hens. 2. 2. Dietary l-tryptophan supplementation resulted in significant decreases in hepatic lipids, in particular triglyceride, and occurrence of hemorrhage in laying hens. 3. 3. Hepatic lipid accumulation by estrogen injection in starved-refed growing chicks decreased as dietary tryptophan content increased. 4. 4. Supplementation of l-tryptophan at 1000 mg/kg diet enhanced alanine aminotransferase activity in the hepatic tissue and at 500 mg/kg diet, increased cytochrome b5, a component of the mixed function oxidase, in the hepatic microsomes. 5. 5. These results demonstrate that l-tryptophan alleviates fatty liver in laying hens and modifies microsomal mixed function oxidase in the liver.

Age-related changes in hepatic microsomal mixed function oxidase in three strains of rats

Age-related changes in hepatic microsomal mixed function oxidase in three strains of rats

Antimutagenicity of ellagic acid towards the food mutagen IQ: Investigation into possible mechanisms of action

The ability of the plant phenol ellagic acid to inhibit the mutagenicity of the food mutagen IQ was evaluated using Salmonella typhimurium strain TA98 in the Ames mutagenicity test. Ellagic acid caused a concentration-dependent decrease in the S-9- and microsome-mediated mutagenicity of IQ. The plant phenol did not interact directly with the IQ-derived mutagenic species and did not modify the cytosol-mediated activation of the promutagen. At the concentrations used in the mutagenicity studies, ellagic acid failed to inhibit microsomal mixed-function oxidase activity, including that mediated by the P450I family responsible for the bioactivation of IQ, despite being an essentially planar molecule as indicated by computer-graphic analysis. The inhibitory effect of ellagic acid was independent of its ability to chelate Mg 2+. However, pre-incubation of ellagic acid with the bacteria, followed by removal of the plant phenol, did not completely prevent the inhibitory effect of the phenol on the mutagenicity of IQ. Intraperitoneal administration of ellagic acid to rats caused a decrease in total cytochrome P-450 levels and related activities as well as in cytosolic glutathione S-transferase activity. Finally, the possibility that the reported anticarcinogenic action of ellagic acid reflects nothing more than non-selective destruction of hepatic cytochromes P-450, and thus reduced chemical activation, is considered.

The effects of direct and microsomal activated aflatoxin B 1 on chicken peritoneal macrophages in vitro

Sephadex-elicited peritoneal exudate cells were cultured on glass coverslips in order to determine the effects of aflatoxin B 1 (AFB 1) on chicken macrophages. Adherent macrophage monolayers were exposed for 1 h to 5, 10, and 20 μg ml −1 of AFB 1, directly or to 0.01, 0.1, 0.5, 1, and 5 μg ml −1 of AFB 1 in the presence of a chicken microsomal mixed function oxidase system (MFO). After exposure, the macrophage cultures were washed and allowed to recover for 2 h in fresh culture medium. Parameters measured at 2 h post recovery period were the substrate adherence potential, morphological alterations, phagocytic ability, and number of sheep red blood cells (SRBC) internalized per phagocytic macrophage. Direct in vitro exposure to AFB 1 resulted in a dose-dependent decrease in macrophage adherence potential, and an increase in cell damage as determined by nuclear disintegration and cytoplasmic blebbing, but no detrimental effects were observed on percent phagocytic cells or the number of internalized SRBC. However, significant reductions in adherence potential, increased morphological alterations, and reduced phagocytosis and internalization of SRBC were observed when MFOs were added to cultures treated with much lower doses of AFB 1. Addition of piperonyl butoxide (a P-450 inhibitor) abrogated AFB 1-MFO induced alterations. This study suggests that microsomal activated AFB 1 causes significant alterations in chicken macrophage functions.

Effects of Dietary Protein and Corticosterone on Mixed Function Oxidase in Liver and Immunity in Male Broilers.

Three experiments were undertaken to determine if dietary protein and corticosterone affect hepatic microsomal mixed function oxidase (MFO) system and immunoresponses in male broiler chicks. In Experiments 1 and 2, chicks (14 days of age) were fed on 11.5 or 23% protein diet supplemented with 0, 5 or 50 ppm corticosterone for 14 days. In Experiment 3, chicks (20 days of age) were given 10, 20 or 40% protein diet supplemented with 0 or 10 ppm corticosterone for 12 days. Dietary corticosterone increased the content of cytochrome P-450 in chicks fed on the 11.5% protein diet, but not on the 23% protein diet. Cytochrome P-450 in chicks fed the 40% protein diet with 10 ppm corticosterone is lower as compared to that in chicks fed the diet without corticosterone. The activity of NADPH-cytochrome c reductase increased linearly with dietary corticosterone or protein. Antibody titer to Sheep Red Blood Cell in chicks fed on the 40% protein diet was lower than that in chicks fed on the 10 or 20% protein diet. The corticosterone administration did not affect the titer. The titer to Brucella canis was decreased by the administration of corticosterone, but not by an increase in dietary protein contents. The results suggest that dietary corticosterone activates the MFO system in hepatic microsomes, while the degree of the activation of the MFO is dependent on dietary protein contents, and also suggest that effects of dietary protein and corticosterone on antibody production may be different from type of antigen used though decreasing protein and increasing corticosterone in the diets reduces organ weight related to immunity.

Effect of the fungicide benomyl on xenobiotic metabolism in rats

The effect of benomyl administered orally (p.o.) and intraperitoneally (i.p.) on the activity of hepatic microsomal mixed-function oxidases (MFOs) was studied in rats. A dose of 100 mg/kg given i.p. reduced the activities of several hepatic drug-metabolizing enzymes 24 h following the treatment. A similar reduction in the activities of the MFOs was also noted 24 h following oral benomyl administration at a dose of 500 mg/kg. Furthermore, in vivo inhibition of drug metabolism by benomyl was demonstrated by increased pentobarbital sleeping-time 24 h after p.o. as well as i.p. dosing. No alterations were found in the serum sorbitol dehydrogenase (SDH) at 24 h after i.p. or oral benomyl indicating a lack of hepatotoxic effect. These results indicate that benomyl shows a route-independent effect on MFOs and is not toxic to the liver.

Potentiation of bromobenzene-induced pneumotoxicity by phenobarbital as determined by bronchoalveolar lavage fluid analysis.

Bromobenzene produces pulmonary, renal and hepatic damage in the rat. Phenobarbital potentiates bromobenzene-induced hepatotoxicity. Studies were initiated to determine if phenobarbital potentiates the pulmonary damage produced by bromobenzene. In a dose ranging study, adult male rats were treated daily for 4 days with phenobarbital (80 mg/kg, ip) and on the fifth day were dosed with 0, 2, 3, or 4 mmoles/kg, ip, bromobenzene. In a larger study phenobarbital was given for 4 days (80 mg/kg, ip), and bromobenzene (3.3 mmoles/kg, ip) was given on the fifth day. Pulmonary damage was assessed 24 hours later in the first study and 12 hours later in the second study by bronchoalveolar lavage fluid analysis (BALF), microsomal mixed function oxidase measurements and histopathological evaluations. BALF biochemical markers employed were gamma-glutamyl transpeptidase (GGT), lactate dehydrogenase (LDH) and total protein. Phenobarbital treatment greatly enhanced bromobenzene-induced GGT and LDH release into the lavage fluid in a dose-dependent manner. Phenobarbital treatment increased microsomal 7-O-dealkylation of both ethoxy- and pentoxyresorufin in the liver without affecting these activities in the lung. Bromobenzene treatment decreased the hepatic microsomal dealkylation of decreased the hepatic microsomal dealkylation of pentoxyresorufin in a dose-dependent manner. Since known rat pulmonary P450 isozymes have been reported to be insensitive to phenobarbital induction, it may be that the toxicity is due to transport of a reactive metabolite(s) formed in the liver to the lung or bromobenzene is activated by some other pulmonary P450 isozyme responsive to phenobarbital.