Hepatic Mixed Function Oxidase Activity Research Articles

Variation of Hepatic Ethoxycoumarin O-de-ethylase Activity with Body Weight and Other Factors in Brook Trout (Salvelinus fontinalis)

Data from several previous experimental studies of hepatic mixed function oxidase (MFO) activity were reexamined to study the possible relationship between ethoxycoumarin O-de-ethylase activity and body weight and other factors in "uninduced" brook trout (Salvelinus fontinalis). Weight-specific O-de-ethylase activity was inversely related to body weight; regression of the log transformation of the total hepatic O-de-ethylase activity on log liver weight yielded a regression coefficient of 0.850 ± 0.047 close to the value of ~0.75 expected for a relationship based on overall metabolic rate. The regression coefficient was significantly higher in males than in females. Multiple regression analysis showed O-de-ethylase activity to be positively correlated with both microsomal protein and cytochrome P-450 content. The data confirm that size (or age) is a factor to be eliminated in any biological effects monitoring program based on MFO assays.Key words: mixed function oxidases, ethoxycoumarin O-de-ethylase, brook trout, Salvelinus fontinalis; hepatic mono-oxygenases

Induction of hepatic mixed function oxidase activity in trout ( Salvelinus fontinalis) by Aroclor 1254 and some aromatic hydrocarbon PCB replacements

Groups of brook trout ( Salvelinus fontinalis) were fed a polychlorinated biphenyl (PCB) mixture (Aroclor 1254) or three PCB replacements based on phenylxylylethane or diisopropylnaphthalene mixtures at dose levels of 30 and 100 μg·g −1. Both dose levels of all the materials tested induced hepatic ethoxyresorufin O-deethylase activity; ethoxycoumarin O-deethylase activity was induced by Aroclor 1254 and one of the phenylxylylethane mixtures. Benzphetamine N-demethylase activity was not induced by any treatment. Effects on other indices of MFO induction, such as protein content and cytochrome P-450 content, were variable. The fish accumulated from 13 to 46% of the Aroclor 1254 dose but only 2 to 7% of the PCB replacements. All components of the phenylxylylethane mixtures were accumulated but some major components of the diisopropylnaphthalenes were not.

Effects of Dietary Selenium and Vitamin E on Hepatic Mixed-Function Oxidase Activities and In Vivo Covalent Binding of Aflatoxin B1 in Rats

Male weanling Fischer-344 rats were fed a selenium (Se)-vitamin E (VE) deficient Torula ycast basal dict or that dict supplemented with graded levels of Se (0.2–6.0 ppm as Na2SeO3) or VE (100 iu/kg as all-rac-2-tocopheryl acetate), or both, for 4 or 6 weeks. Se deficiency and excess (6.0 ppm) markedly depressed in vivo covalent binding of aflatoxin (AFB1) to macromolecules in livers of rats killed 2 hours after an i.p. dose of 1 mg/kg tritiated AFB1. VE supplementation had no effect. Prior phenobarbital (PB) treatment generally decreased adducts without changing diet-related trends. Some hepatic enzyme capabilities were also measured. Cytochrome b3 content and cytochrome c reductase activity were unaffected by diet. VE increased cytochrome P-450 content, ethylmorphine N-demethylase and benz(α)pyrene hydroxylase activities; all these were unaffected by Se levels. Se deficiency and excess (but not VE deficiency) increased glucuronyl transferase. PB induction affected all diet groups and was more in agreement with MFO activity than transferase. Adduct formation was more consistently related to transferase activity than to MFO activities. The contrasting effects of SE and VE on AFB1 adducts in rats and chicks are discussed.selenium vitamin E aflatoxin adducts metabolism

Hepatic mixed-function oxidase activity in mice infected with Trypanosoma brucei gambiense or treated with trypanocides

Three days after mice were inoculated with Trypanosoma brucei gambiense, total hepatic cytochrome P-450 levels were decreased 14% from control values, while the specific mixed-function oxidase activity (per nmole of cytochrome P-450) was inhibited almost 40% in infected animals. Furthermore, drugs used to treat trypanosomiasis (Suramin and Melarsoprol B) are themselves potent in vitro inhibitors of hepatic mixed-function oxidase activity. These two trypanocides also produced a decrease in mixed-function oxidations and an increase in pentobarbital sleeping time when administered intraperitoneally in vivo. These results demonstrate that mice with trypanosomiasis or undergoing trypanosome chemotherapy have a significantly impaired capacity to metabolize foreign compounds.

Polyhalogenated biphenyls and phenobarbital: Evaluation as inducers of drug metabolizing enzymes in the sheepshead, Archosargus probatocephalus

Several doses of Aroclor 1254 (polychlorinated biphenyl (PCB) mixture), Firemaster FF1 (polybrominated biphenyl (PBB) mixture), 2,2′,4,4′,5,5′-hexabromobiphenyl (HBB), 3,3′,4,4′,5,5′-hexachlorobiphenyl (HCB) and phenobarbital (PB) were administered to the marine fish sheepshead ( Archosargus probatocephalus). The PCB and PBB mixtures caused induction of hepatic microsomal benzo[ a]pyrene hydroxylase (AHH), 7-ethoxycoumarin O-deethylase (7-EC) and 7-ethoxyresorufin O-deethylase (ERF) activities, but not benzphetamine N-demethylase (BND), epoxide hydrolase (EH) or glutathione S-transferase (GSH-T) activities. This induction pattern is typical of that caused by polycyclic aromatic hydrocarbons (PAH) in fish and mammals or by tetrachlorodibenzo- p-dioxin (TCDD) in mammals. The extent of induction of AHH-activity and cytochrome P-450 content was higher when experiments were carried out in summer (water temperature 25 ± 4°C) than in winter (water temperature 11 ± 3°C). Firemaster FF1 (15 mg/kg) induced fish for at least 56 days in both summer and winter at which time the liver concentrations of PBB were in the ppm range. PCB concentrations in the ppm range have been found in fish from polluted lakes and seas, thus we may expect that environmental exposure to PCB is sufficient to induce hepatic mixed function oxidase (MFO) activities. The PCB isomer 3,3′,4,4′,5,5′-HCB, which induces the same spectrum of hepatic drug-metabolizing activities as TCDD and PAH in rats, had a broadly similar effect in the sheepshead. Another purified isomer, 2,2′,4,4′,5,5′-HBB, which induces the same enzymes as PB in rats, had no effect on drug-metabolizing activities in sheepshead. PB was also without effect on sheepshead hepatic drug-metabolizing enzymes, although a typical narcotic effect was produced in PB-treated sheepshead. Our studies provide further evidence that drug-metabolizing activities in fish liver are readily induced by chemicals like TCDD or PAH, but we fail to demonstrate induction after treatment of sheepshead with inducers of the PB type.

Time- and dose-dependence of hepatic mixed function oxidase activity in brook trout Salvelinus fontinalis on polychlorinated biphenyl residues: Implications for ‘biological effects monitoring’

Brook trout Salvelinus fontinalis which had been fed a single dose of polychlorinated biphenyls (PCB: Aroclor 1254) had elevated hepatic mixed function oxidase (MFO) activity detectable between 2 and 20 days after feeding, as shown by ethoxycoumarin O-de-ethylase activity. O-de-ethylase activity was well correlated ( r = 0 · 74) with whole body PCB residues; both variables declined slowly during the period after feeding. In two dose-response experiments, hepatic O-de-ethylase activity increased with increasing PCB residue concentration deposited from a single feeding; the relationship was best described by a power function of the form: ln y = 0 · 23 ln x + 5·8 (where y is O-de-ethylase activity as nanomoles of product formed per gramme of liver per hour; and x is PCB concentration in fillet as μg g −1 wet weight. Despite the relatively insensitive response of O-de-ethylase activity to changes in PCB concentrations, the enzyme assay offers a potential screening method for the detection of ‘hot-spots’ of contamination by PCB, and possibly other MFO inducers.

Effect of stannous fluoride and sodium fluoride on hepatic mixed-function oxidase activities in the rat

Alterations in rat hepatic mixed-function oxidase activities were measured in vivo and in vitro 20 hr after pretreatment with a single dose of stannous fluoride or sodium fluoride. SnF 2 pretreatment significantly decreased the N-demethylation of aminopyrine and O-demethylation of p-nitroanisole; whereas, NaF in an equimolar dose did not affect N-demethylation but did decrease O-demethylation. Hexobarbital sleeping time was significantly prolonged by SnF 2 but not by NaF. Furthermore, the elimination half-life of antipyrine was increased by SnF 2 but not by NaF. These results demonstrate that the inhibition by SnF 2 of hepatic mixed-function oxidase activities in the rat is primarily due to the stannous ion and that fluoride ion has very little affect on these enzyme activities.

Depression of hepatic mixed-function oxidase activity by B. pertussis in splenectomized and athymic nude mice

Administration of Bordetella pertussis (B. pertussis), Corynebacterium parvum (C. parvum), and several other immunoactive substances is known to cause a marked decrease in the activity of the hepatic microsomal mixed-function oxidase system. The effect of C parvum has been reported to involve the reticuloendothelial system. In the present study, the effect of B. pertussis administration to decrease hepatic microsomal drug metabolism was studied in unoperated, sham-operated, and splenectomized mice as well as in athymic nude (nu/nu) mice and their phenotypically heterozygous (+/nu) littermates. Administration of B. pertussis to the splenectomized, sham-operated, and unoperated mice resulted in a decrease in the activity of the microsomal enzyme system that was approximately the same for each of the three groups of animals. Administration of B. pertussis to nu/nu mice and the +/nu mice also decreased the microsomal enzyme activity measured 24 hr after injection. However, at 7 days after B. pertussis administration, the hepatic drug-metabolizing activity of the nu/nu mice was not significantly different from control values, whereas the activity of the +/nu mice was still significantly depressed. The failure of splenectomy to prevent the decrease in microsomal mixed-function oxidase activity caused by B. pertussis indicated that the effect of this agent differs from that of C. parvum, whose effect was prevented by splenectomy. Indeed, the results obtained with the athymic nude mouse suggests that the depression of hepatic mixed-function oxidase activity by B. pertussis may involve T-cell dependent responses.

Alterations in hepatic mixed-function oxidase activity of rainbow trout after acute treatment with polybrominated biphenyl isomers and FireMaster BP-6.

The effects of FireMaster BP-6 and several pure isomers on hepatic microsomal mixed-function oxidase (MFO) activity in rainbow trout have been investigated. After one parenteral injection of these agents at 150 mg/kg and sacrifice 5 d later, there was no elevation of hepatic cytochrome P-450. The noncoplanar isomers 2,4,5,2',4',5'-hexa and 2,3,4,5,2',4',5'-heptabromobiphenyl did not increase any MFO activity generally associated with an increase in cytochrome P-450 levels, including the dealkylations of ethoxycoumarin and benzphetamine. The coplanar isomer 3,4,5,3',4',5'-hexabromobiphenyl produced an increase in O-deethylation of both ethoxycoumarin and ethoxyresorufin, suggesting enhancement of cytochrome P-448 associated activity, but produced no increase in benzphetamine N-dealkylation. The "mixed inducer" 2,4,5,3',4',5'-hexabromobiphenyl, produced a small but insignificant elevation of cytochrome P-448 associated MFO activities and no increase in cytochrome P-450 associated activities. At both 150 and 500 mg/kg, the commercial mixture of polybrominated biphenyls FireMaster BP-6 produced marked elevations of both ethoxycoumarin and ethoxy-resorufin dealkylations but had no effect on demethylation of benzphetamine. When microsomes from rainbow trout treated with FireMaster BP-6 were examined electrophoretically, A coomassie Blue-staining band at 57,000 daltons was intensified, as seen after treatment of fish with the cytochrome P-448 inducer beta-naphthoflavone. It is concluded that coplanar polybrominated isomers produce an induction of hepatic cytochrome P-448 associated activity, while noncoplanar isomers are ineffective as inducers of cytochrome P-450 related MFO activities in rainbow trout.

The effects of ingested petroleum on the naphthalene-metabolizing properties of liver tissue in seawater-adapted mallard ducks ( Anas platyrhynchos)

Hepatic mixed function oxidase activities were estimated in seawater-adapted mallard ducks ( Anas platyrhynchos) that had been consuming food contaminated with one of five different types of crude oil. After 50 days of exposure to contaminated food, enzyme activities of liver microsomal preparations were assessed in terms of their naphthalenemetabolizing properties in vitro. Although dose-dependent increases in the total hepatic enzyme activities (nmole naphthalene metabolized per minute per unit mass body weight) were observed in birds consuming food contaminated with each type of crude oil, three patterns of response were apparent. Crude oils from South Louisiana and Kuwait stimulated large and significant increases in the specific activity of the enzyme system (nmole naphthalene metabolized per minute per unit mass microsomal protein), whereas little or no increase in either microsomal protein content or relative liver weight were observed. In contrast, two crude oils from Santa Barbara, Calif., induced only small increases in specific activity but significant increases occurred in hepatic microsomal protein concentration and relative liver weight. The crude oil from Prudhoe Bay, Ala., evoked intermediate patterns of response. The possible significance of these data is discussed in relation to the survival of seabirds consuming petroleum-contaminated food and drinking water.

Toxicological response of rats fed Lake Ontario or Pacific Coho salmon for 28 days.

Abstract The present study was undertaken to determine if the feeding of Lake Ontario salmon containing a mixture of chemical contaminants, could produce a toxicological response in rats. Male and female rats were fed freeze‐dried Lake Ontario or Pacific Coho salmon for 28 days at levels of 1.45%, 2.90% and 5.80% of their normal diet. Body weight changes, organ weights, food intake, routine hematology, biochemical parameters, residue levels and gross and microscopic pathological changes were determined. Body weight gain was decreased in all groups of male rats fed Lake Ontario or Pacific salmon but this was attributed to the unpalatibility of the diet since the food intake in these groups was decreased during the first week of the study. No effects were observed on body weight gain or food intake in female rats. Hematological parameters including hemoglobin, hematocrit, white blood count and red blood count were not affected by treatment. Serum potassium levels were significantly decreased in all three groups of male rats receiving Pacific salmon and in the two highest groups of female rats receiving the Pacific fish. Hepatic mixed function oxidase activity was increased in both males and females of the two highest Lake Ontario treatment groups. There appeared to be an altered steroid hormone metabolism in female adrenals which occurred only in Pacific Coho groups. Altered steroid metabolism in male adrenals was also observed but this occurred in both Lake Ontario and Pacific groups. Minor histological changes were observed in thyroid and liver of males and females fed Lake Ontario salmon and were attributed to the contaminants found in this fish source. Lesions of the kidney were observed in all female groups receiving either source of salmon but these could not be attributed to any specific contaminants. The results obtained here indicate that the feeding of Lake Ontario salmon results in certain biochemical and histological changes in male and female rats which are attributed to chemical residues found in those fish.

Effects of induction of R- and S-warfarin and metabolite concentrations in rat plasma

The plasma half-lives of R- and S-warfarin were statistically indistinguishable ( p < 0.05) for uninduced male Wistar rats, although plasma concentrations of S-warfarin were invariably higher at equivalent doses. Induction of hepatic mixed-function oxidase activity by phenobarbital significantly shortened the plasma half-lives and lowered the plasma zero-time concentrations of both enantiomers relative to uninduced rats. In contrast, induction with 3-methylcholanthrene produced a significantly lower than normal zero-time concentration of R-warfarin but not of S-warfarin. R- and S-warfarin plasma half-lives did not differ significantly from one another in phenobarbital or 3-methylcholanthrene-induced rats, but the half-lives differed significantly between the two induced groups. Concentrations of S-warfarin metabolites exceeded those of R-warfarin metabolites in the plasma of uninduced or induced rats. The major plasma metabolites were 4′-hydroxywarfarin from S-warfarin and, to a lesser extent, from R-warfarin. Plasma R- and S-warfarin metabolite patterns were markedly different from in vitro patterns catalyzed by hepatic microsomal enzymes of uninduced and phenobarbital- or 3-methylcholanthrene-induced rats. In uninduced rats a dose of 1 mg of S-warfarin/kg produced an anticoagulant response equivalent to 10 mg of R-warfarin/kg. The intrinsic activity of S warfarin was calculated to be 4.9 ± 0.4 times that of R-warfarin. Induction of rats with phenobarbital or 3-methylcholanthrene diminshed the magnitude of the normal anticoagulant response to R- and S-warfarin.

Extrapulmonary effects of NO 2 as reflected by pentobarbital-induced sleeping time in mice

The influence of nitrogen dioxide (NO 2) on pentobarbital (PEN)-induced sleeping time (S.T.) was investigated in mice. Acute exposure to concentrations as low as 470 μg NO 2 m 3 (0.25 ppm) caused a significant increase in PEN-induced S.T. No significant effects on PEN-induced S.T. were detected after 1 or 2 days (3 h/day) exposure to 235 μg NO 2 m 3 (0.125 ppm). None of the exposure regimens affected the time required for the drug to induce sleep. When the effects of repeated daily exposures (3 h/day) to 9400 μg NO 2 m 3 (5 ppm) were compared in male and female mice, the females had a significantly increased PEN-induced S.T. after 1 and 2 days of exposure. However, an increase in PEN-induced S.T. was not observed in males until the 3rd day of exposure. Since the duration of PEN-induced S.T. is primarily determined by hepatic mixed function oxidase activity, it is possible that NO 2 may alter some aspects of xenobiotic metabolism.

Functional and biochemical correlates of chlordecone exposure and its enhancement of CCl 4 hepatotoxicity

Animals pretreated with chlordecone exhibit a greatly increased hepatotoxic response to CCl 4 challenge. Possible mechanisms underlying this interaction were examined. A single p.o. administration of chlordecone (5 mg/kg) was followed by CCl 4 (200 μl/kg) administered i.p. 48 h later. Twenty-four hours later, animals treated with chlordecone + CCl 4 had decreased hepatic excretory function (20% of controls) and elevated plasma transaminase activities and bilirubin. Hepatic mixed function oxidase activity was assessed as pentobarbital sleeping time and was not affected by chlordecone pretreatment. Irreversible binding of label from 14CCl 4 to hepatic protein or lipid was not different in the chlordecone group compared to vehicle controls. Hepatic and renal glutathione concentrations were not affected by chlordecone alone (at 6 h, 1, 2, 3, 5 and 7 days) or by a combination of chlordecone (48 h) and CCl 4 (24 h). CCl 4-induced lipid peroxidation of liver tissue, measured in vitro or in vivo, was not increased by chlordecone treatment. Thus, while the mechanism for the enhanced toxicity remains to be elucidated, these results suggest that the interaction between chlordecone and CCl 4 is a subtle one, not causally involving increased covalent binding of the toxin, increased susceptibility of tissue lipids to peroxidative damage or decreased hepatic GSH.

Hepatic mixed function oxidase (MFO) activities during the seasonal life cycle of the frog, Rana temporaria

1. A single dose of DDT (5 mg/kg) did not affect the activities of hepatic mixed function oxidase (MFO) components in fed summer frogs, Rana temporaria. 2. The activity of MFO system showed remarkable seasonal variation with its higher activity level during the summer period. The MFO activity tended to be lower in the females than in the males at the beginning of spawning time while at the end of the spawning the reverse tendency was observable. 4. In frogs overwintered in laboratory the activity levels of MFOs were lower than in the frogs undergone a natural wintering. The activity levels in an immature female resembled those observable in mature males.

Impairment of hepatic mixed-function oxidase activity by α- and β-naphthylisothiocyanate: Relationship to hepatotoxicity

Since administration of α-naphthylisothiocyanate (ANIT) to rats inhibits hepatic mixed-function oxidase (MFO) activity in vivo and in vitro, the nature of this inhibition was investigated; its role in the pathogenesis of ANIT-cholestasis was also examined. Male rats were treated (150 mg/kg, ip) with ANIT or β-naphthylisothiocyanate (BNIT), a noncholestatic isomer. ANIT was also given to hamsters (a resistant species) and to rabbits (a nonsusceptible species). The animals were killed at different intervals (2–72 hr); MFO activity was compared to plasma biochemical alterations (glutamic-pyruvic transaminase, bilirubin, 5′-nucleotidase) indicative of liver injury. In the rat, reductions in cytochrome P-450, aniline oxidase, and aminopyrine demethylase were evident before the development of hyperbilirubinemia. After 24 hr, cytochrome b 5, total heme, and NADPH-cytochrome c reductase were also reduced, but glucuronyl transferase activity was increased. BNIT caused no elevation in plasma bilirubin or 5′-nucleotidase activity but decreased MFO activity in rats. Reductions in cytochrome P-450 and microsomal metabolism were also observed in hamsters and rabbits treated with ANIT, although only a moderate hyperbilirubinemia was observed in hamsters and none in rabbits. Both ANIT and BNIT caused reductions in cytochrome P-450 when incubated in vitro with microsomes. Loss of cytochrome P-450 due to increased degradation is suggested. We conclude that the metabolic lesion induced by ANIT is primary and does not result from the accumulation of bile acids within the hepatocyte. It appears, however, that this lesion has no causal relationship to the subsequent development of ANIT-induced cholestasis.

Nutritional and toxicological studies with growing pigs fed aquatic plant rations.

Aquatic weeds from Cayuga Lake in New York were included as 10% or 20% dry weight of rations which were fed to growing pigs for 69 days. The animals gained weight at a rate comparable to those on a basal diet or a diet in which an equivalent weight of alfalfa replaced the aquatic plants. The feed efficiency (kg ration consumed/kg wt gained) of the aquatic rations was significantly lower, probably because of the lower nutritive value of the aquatic plants. Polychlorinated biphenyls (PCBs) were found in the aquatic plants (0.59 ppm, dry weight) and in the pig fat of those fed the aquatic ration. Hepatic microsomal mixed function oxidase activity was not significantly different in the animals fed the various treatment rations. Histopathologic examination of animal tissues revealed no lesions or abnormalities which could be attributed to the diets.

The Effect of Dietary Protein Depletion and Repletion on Rat Hepatic Mixed Function Oxidase Activities

The Effect of Dietary Protein Depletion and Repletion on Rat Hepatic Mixed Function Oxidase Activities

Hepatic mixed-function oxidase activity in rainbow trout exposed to several polycyclic aromatic compounds

The effects of polycyclic aromatic hydrocarbons (PAHs) on microsomal liver enzymes were examined in rainbow trout. Various PAHs (naphthalene through benzo( a)pyrene [B( aP]) were injected ip to screen for mixed-function oxidase (MFO) activity. Chrysene, B( a)P, and Aroclor 1254 caused MFO induction. When fish were also exposed to solubilized pyrene, fluoranthene, and B( a)P in water, bioaccumulation of B( a)P resulted in MFO induction, whereas bioaccumulation of pyrene and fluoranthene did not. Based on water and injection exposure to B( a)P, it was predicted that tissue concentrations in excess of 300 μg/kg B( a)P would be accompanied by MFO induction in rainbow trout.

Effect of alteration of rat hepatic mixed-function oxidase (MFO) activity on the toxicity of 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD)

The effect of alteration of the activity of the hepatic mixed-function oxidase (MFO) enzyme systems on the toxicity of 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) in rats has been examined. These experiments have indicated the naturally occurring age and sex-related differences in hepatic MFO activity in rats are inversely correlated with the 20-day LD50 of TCDD in these animals. The effect of administering inducers and inhibitors of the hepatic MFO enzyme systems on the 20-day LD50 of TCDD in rats has also been examined. In all cases there was an inverse relationship between the activity of the hepatic MFO system at the time of TCDD administration and the toxicity of TCDD as determined by the 20-day LD50.