P-nitroanisole O-demethylase Research Articles

Effects of an immunosuppressive agent, tacrolimus (FK-506), on the activities of cytochrome P-450-linked monooxygenase systems in rat liver microsomes

The effects of an immunosuppressive agent, tacrolimus (FK-506), on the activities of cytochrome P-450-linked monooxygenase systems with respect to three cytochrome P-450 isozymes in rat liver microsomes were investigated. FK-506 non-competitively inhibited the aniline p-hydroxylase, p-nitroanisole O-demethylase and lidocaine N-deethylase activities of cytochrome P-450-linked monooxygenase systems, these activities being mainly catalyzed by cytochromes P-450 CYP2E1, CYP2C11 and CYP3A4, respectively, and the Ki values of the activities for FK-506 were determined to be 605, 491 and 97 μM, respectively. The inhibition of cytochrome P-450-linked monooxygenase systems by FK-506 seemed to involve the direct inhibition of cytochromes P-450 because the NADPH-cytochrome c reductase and NADPH-ferricyanide reductase activities of NADPH-cytochrome P-450 reductase were not affected by the presence of 1 mM FK-506 at all. A spectrophotometric study showed that a reverse type I spectral change was induced on the addition of FK-506 to rat liver microsomes, and the Ks value was apparently 125 μM. On the other hand, the EPR spectra of cytochromes P-450 in rat liver microsomes were not affected by 1 mM FK-506. These results suggest direct interaction between FK-506 and cytochrome P-450 apoproteins, except for the heme iron regions of cytochromes P-450, resulting in inhibition of the drug-metabolism activities catalyzed by cytochromes P-450.

Influence of partial hepatectomy in rats on the activity of hepatic microsomal enzymatic systems.

The influence of partial hepatectomy on the activity of the hepatic microsomal enzymatic systems was determined in rats. Cytochrome P-450, cytochrome b5, four mixed functional oxidase (MFO) activities (microsomal aniline hydroxylase, p-nitroanisole O-demethylase, aminopyrine N-demethylase and NADPH cytochrome c reductase) and glutathione levels were measured in unhepatectomized rats (control group) and in hepatectomized rats 12 h, 24 h, 3 days and 6 days after 70% hepatectomy. Following surgery the remaining lobes of the liver grow rapidly in order to restore the original liver mass. Partial hepatectomy significantly reduces cytochrome P-450 and b5 content in the remaining liver as well as the four MFO activities studied. But when the enzymatic systems are expressed as nmoles/mg microsomal protein, only cytochrome P-450 shows statistical differences. The hepatic biotransformation capacity of drugs and xenobiotics decreases during the regeneration period due to the reduction of hepatic mass rather than because of a reduction of their metabolic capacity. Glutathione levels are increased after partial hepatectomy but increased glutathione-dependent protector mechanisms are not expected.

Combined effects of ethanol and cigarette smoke on hepatic and pulmonary xenobiotic metabolizing enzymes in rats

The combined effects of ethanol (EtOH) and cigarette smoke (CS) on hepatic and pulmonary monooxygenase (MO) activities (aniline 4-hydroxylase (AH), aminopyrine N-demethylase (AMND), 7-ethoxyresorufin O-deethylase (EROD), p-nitroanisole O-demethylase ( p-NAOD)), lipid peroxidation (LP) and reduced glutathione (GSH) levels and glutathione S-transferase (GST) activities toward several substrates (1-chloro-2,4-dinitrobenzene (CDNB), 1,2-dichloro-4-nitrobenzene (DCNB), ethacrynic acid (EAA), 1,2-epoxy-3-( p-nitrophenoxy)-propane (ENPP)) were determined and compared with those of EtOH or CS alone in rats. When the male adult rats (225–275 g) were treated with 10% EtOH (v/v) in their drinking for 21 days AH, AMND and EROD activities and LP and GSH levels increased significantly whereas GST activity for EAA decreased significantly in liver as compared to controls. EtOH did not change the hepatic p-NAOD and GST activities toward CDNB, DCNB and ENPP. In lung, EtOH increased GST activities toward CDNB and ENPP and LP level but decreased GST activity toward DCNB, significantly. No alterations were noted in pulmonary MO activities and GST activity toward EAA and GSH level by EtOH treatment. When the animals were exposed to CS five times a day, with 1 h intervals, for 3 days in a chamber where smoke and fresh air lead alternatively, AMND, EROD and p- NAOD activities, GST activity toward EAA and GSH level increased but LP level and GST activity for ENPP decreased significantly in liver. CS did not alter the hepatic AH and GST activities toward CDNB and DCNB. In lung, CS increased AH, EROD and p- NAOD activities and LP and GSH levels and decreased all the GST activities studied significantly. CS had no influence on pulmonary AMND activity. For the combined treatment, the animals were treated with 10% EtOH (v/v) in their drinking water for 21 days and during the last 3 days they were exposed to CS five times a day, with 1, h intervals, in a chamber where smoke and fresh air lead alternatively. In these animals, augmentation of elevations were noted in AH and p- NAOD activities and LP and GSH levels but not in EROD and AMND activities in liver. Combined treatment significantly decreased GST activity toward CDNB, ameliorated the alteration caused by either EtOH or CS treatment alone on GST activity toward EAA and potentiated the depression of GST activity toward ENPP to a greater degree. No change was observed in GST activity toward DCNB. In lung, combined treatment potentiated the elevations of AMND and p- NAOD activities and LP level and not those of AH and EROD activities. GST activities toward CDNB, DCNB and ENPP were highly elevated by the combined treatment. No changes were observed in pulmonary GSH level and GST activity for EAA by the combined treatment. These results reveal that the regulations of the hepatic and pulmonary MO and GST are differentially influenced by EtOH, CS and the combined treatment.

Early hepatic changes induced in rats by two hepatocarcinogenic organohalogen pesticides: bromopropylate and DDT.

The aim of the present studies was to describe the effect of two organohalogen pesticides: DDT and bromopropylate, on early changes in rat liver, proposed in the literature to be useful endpoints in screening of non-genotoxic hepatocarcinogens and/or liver tumor promoters. We investigated the effects on the following endpoints: hepatomegaly, mitogenesis (DNA synthesis, mitotic activity, percentage of binuclear cells) and cytochrome CYP2B1-dependent monooxygenase induction. The histological and cytochemical changes in the liver were also recorded. Male Wistar rats received bromopropylate in one, three or five daily oral doses of 125, 250, and 500 mg/kg body wt. day-1. DDT was applied as one, three, and five daily oral doses of 24 mg/kg body wt. day-1 (this dose is close to the mean hepatocarcinogenic dose in male Wistar rats: 34.1 mg/kg body wt. day-1). In the case of both pesticides the early effects observed consisted of hepatomegaly accompanied by an increase in the p-nitroanisole O-demethylase activity and hepatocyte proliferation. Hepatocyte proliferation was elevated during the total experimental period. Vacuolated cytoplasm and evident focal necrosis may suggest that the maximal increase in hepatocyte proliferation, preceding hepatomegaly, is at least partly related to a regenerative liver response to pesticides. In addition to the above-mentioned early changes, the present findings provide new evidence for the occurrence of dose-dependent abnormal mitoses (and c-mitoses) in the hepatocytes of the bromopropylate and DDT treated rats.

Postmortem changes in drug-metabolizing enzymes of rat liver microsome

Postmortem changes in the drug-metabolizing enzymes in rat liver microsome were studied. Parameters investigated were: microsomal protein, NADPH-cytochrome P-450 reductase activity, NADH-cytochrome b 5 reductase activity, cytochrome b 5 content, cytochrome P-450 content, aminopyrine N-demethylase activity, aniline p-hydroxylase activity, p-nitroanisole O-demethylase activity, uridine diphosphate (UDP)-glucuronyl transferase activity and glutathione S-transferase activity. Nearly all the parameters based on microsomal protein decreased during autolysis and the time-dependent decrement ratios of the parameters changed by various amounts. Cytochrome b 5 content decreased more rapidly than that of other components. By 36 h post mortem, levels of cytochrome b 5 were not detectable. By 48 h post mortem, NADPH-cytochrome P-450 reductase activity decreased to 91%, NADH-cytochrome b 5 reductase activity decreased to 94%, and cytochrome P-450 content decreased to 92% of relative activities. By 48 h post mortem, aminopyrine N-demethylase activity decreased to 87%, aniline p-hydroxylase activity decreased to 98% and p-nitroanisole O-demethylase activity decreased to 75% of relative activities. The activity of p-nitroanisole O-demethylase appeared to be more stable than that of aminopyrine N-demethylase or aniline p-hydroxylase. These results demonstrate that there are multiple forms of isozymes of the cytochrome P-450-linked monooxygenase system. Hepatic transferases showed decrease patterns different to those of monooxygenases, so UDP-glucuronyl transferase activity of ∼32% of relative activity was detected at 48 h post mortem. Thus, UDP-glucuronyl transferase activity appeared to be more stable than the cytochrome P-450-linked monooxygenases. These results show that these activities and components would be useful as markers of postmortem time. The causes of the variety of instability of these enzyme systems are discussed.

Stability of microsomal monooxygenases in murine liver S9 fractions derived from phenobarbital and beta-naphthoflavone induced animals under various long-term conditions of storage.

The aim of this study was to define the long-term stability of metabolizing enzymes in activating preparations for short-term genotoxicity bioassays under various storage conditions. Expressions of cytochrome P450 content, NADPH-cytochrome (P450) c-reductase activity, and of the several monooxygenases, such as aminopyrine N-demethylase (class IIIA P450), p-nitroanisole O-demethylase (mixed), dinemorphan N-demethylase (IIB1), ethoxyresorufin O-deethylase (IA1), ethoxycoumarin O-deethylase (mixed), and pentoxyresorufin O-dealkylase (IIB1), were examined in S9 fractions derived from Na-phenobarbital (PB) plus beta-naphthoflavone (beta-NF) induced male and female mice, stored at -80 degrees C, or lyophilized and stored at -20 degrees C. Lipid peroxidation was also determined. Cytochrome P450 and the associated activities were decreased by 30-82% within 9 months of storage. The pattern and degree of relative stabilities were different for the various isoforms. The IA1-like activity, for example, was much more stable (approximately 49% loss) than IIB1-like activities (up to 82% loss). In general, lyophilized enzymes were less stable than directly frozen preparations. In addition, immediately after freeze-drying (lyophilization), a marked decrease in activity of up to 35% was observed. On the contrary, demethylation of aminopyrine and p-nitroanisole remains almost constant over 6 months storage at -196 degrees C. The results obtained indicate that either fresh, daily made S9 fractions or, alternatively, fractions stored in liquid nitrogen (up to 6 months) are recommended for mutagenesis studies.

Effect of cadmium on hepatic microsomal monooxygenase activities in guinea pigs with low and high ascorbic acid intake

The influence of in vivo administration of cadmium on hepatic microsomal cytochrome P-450 and cytochrome b 5 content, aniline hydroxylase, p-nitroanisole O-demethylase, and NADPH-cytochrome c reductase activities was investigated in guinea pigs with low (2 mg/animal/day) and high (100 mg/animal/day) ascorbic acid intake. Male guinea pigs received 10 mg Cd/L drinking water for 5 and 12 weeks. Cadmium administration resulted in significant decreases in cytochrome P-450 content and NADPH-cytochrome c reductase activity in the group of guinea pigs with low ascorbic acid intake, while the decreases were less evident in guinea pigs with high ascorbic acid intake. The activities of aniline hydroxylase and O-demethylase were not apparently changed by Cd administration, except for the group of guinea pigs with high ascorbic acid intake in 12th week of cadmium treatment, when increased activities were observed. These findings suggest that hepatotoxic effect of cadmium manifested by inhibition of hepatic microsomal metabolism may be in part beneficially moderated by enhanced ascorbic acid intake.

Effect of E-5110, a novel non-steroidal anti-inflammatory drug, on trimethadione metabolism as an indicator of hepatic drug-oxidizing capacity in beagle dog.

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.

Correspondence of increased debrisoquine 4-monooxygenase activity with seizure-susceptibility in Mongolian gerbils

Four drug-metabolizing activities in Wistar rat and Mongolian gerbil liver microsomes were investigated to clarify the biochemical basis of convulsions. No significant associations were observed between the susceptibility to the tonic-clonic seizures in Mongolian gerbils and the drug-metabolizing enzymatic activities of p-nitroanisole O-demethylase, aminopyrine demethylase and benzo[a]pyrene-3-monooxygenase. However, a significant association was observed with the debrisoquine 4-monooxygenase (cytochrome P-450db1-linked monooxygenase system) activity, which is known to metabolize exogenous and endogenous parkinsonism-inducing neurotoxins such as MPTP and TIQ, and to activate carcinogens during the detoxication of xenobiotics. The mean activity of debrisoquine 4-monooxygenase in the seizure-sensitive group (male, 380; female, 350 pmol/h/mg protein) remained significantly higher (about 4–5 times) than that in the seizure-resistant group (male, 90; female, 80 pmol/h/mg protein). More Mongolian gerbils were extensive metabolizers of debrisoquine in the seizure-sensitive group than in the seizure-resistant group. These results suggest a possibility that the cytochrome P-450db1-linked monooxygenase system may activate the potential neurotoxins which are associated with tonic-clonic seizures in the Mongolian gerbil.

Blood cholinesterase in rats fed an insect resistance apple clone containing a natural cholinesterase inhibitor

A crab apple clone (Malus brevipes 1021), highly resistant to the apple maggot, is being used in breeding programs developing commercial apple cultivars. This study has discovered that this crab apple contains a natural cholinesterase (ChE) inhibitor that caused a 17.5% in vitro inhibition of rat blood ChE activity. This crab apple also showed a relatively high total (titratable) acidity of 1.28%. The commercial, nonresistant, apple cultivar McIntosh was capable of causing a 7.9% inhibition of blood ChE in vitro. The total acidity in McIntosh was 0.45%. A 4-wk feeding study compared 2 groups of 5-wk-old Fischer 344 male rats fed diets containing 45% of either M. brevipes or McIntosh freeze-dried apples to a third (control) group of rats fed a semipurified diet. In vivo blood ChE activities were similar in all groups of rats, as well as hemoglobin, hematocrit, and red blood cell counts. The liver mixed-function oxidase activity through aminopyrine N-demethylase in the rats fed the apple diets was higher than the controls, but p-nitroanisole O-demethylase activity was induced only in the animals fed the maggot-resistant crab apple. Lowered growth with concomitant lowered food intake, in the otherwise healthy rats fed the maggot-resistant crab apple diet, was attributed to the less palatable, highly acidic fruit. This study indicates that the natural ChE inhibitor in the insect-resistant apple M. brevipes is apparently detoxified upon ingestion.

Dual effects of a novel thienodiazepine platelet-activating factor antagonist, on drug-oxidizing enzymes in beagle dog.

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)

The responses of hepatic monooxygenases of guinea pig to cadmium and nickel.

When Cd (3.58 mg CdCl2.H2O/kg, ip) was administered to male guinea pigs 72 h prior to sacrifice, the metal significantly inhibited the aniline 4-hydroxylase (AH) (16%), ethylmorphone N-demethylase (EMND) (26%), and aminopyrine N-demethylase (AMND) (18%) activities and cytochrome P-450 (12%) and cytochrome b5 (10%) levels. Cd did not alter the hepatic microsomal heme level. Cd, however, significantly increased the hepatic microsomal p-nitroanisole O-demethylase (p-NAOD) (53%) activity. When Ni (59.5 mg NiCl2 x 6H2O/kg, sc) was administered to the guinea pigs 16 h prior to sacrifice, the metal significantly depressed AH (49%), p-NAOD (66%), EMND (47%), and AMND (37%) activities, and cytochrome P-450 (15%), cytochrome b5 (24%), and microsomal heme (28%) levels. For the combined treatment, animals received the single dose of Ni 56 h after the single dose of Cd and then were killed 16 h later. In these animals, significant inhibitions were noted in AH (51%), EMND (47%), and AMND (30%) activities, and cytochrome P-450 (15%), cytochrome b5 (26%), and microsomal heme (30%) compared to those of controls. In the case of p-NAOD activity, the influence was in favor of Ni, i.e., the inhibition was about 61% by the combined treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Suppression of liver microsomal drug-metabolizing enzyme activities in adult female rats pretreated with cannabidiol.

The suppression by cannabidiol (CBD) of the liver microsomal drug-metabolizing enzyme activities in female rats was demonstrated and its mechanism was examined. Pretreatment of rats with CBD (10 mg/kg, i.p.) caused temporary decreases in contents of cytochrome P450 (P450) and b5 and NADPH-cytochrome c (P450) reductase activity compared with values from the vehicle control group. p-Nitroanisole O-demethylase, aniline hydroxylase, d-benzphetamine N-demethylase and delta 9-tetrahydrocannabinol 11-hydroxylase were also decreased by the CBD pretreatment. The latter two activities took a longer time to return to control levels than the former two. However, the CBD pretreatment, which reduced the protein level of P450 UT-2 (CYP2C11) in adult male rats, did not decrease the protein level of P450 F-1 (CYP2C6) or F-2 (CYP2C12) in liver microsomes from female rats. These results suggest that the mechanisms by which CBD suppresses liver microsomal drug-metabolizing enzyme activities are different in male and female rats.

Effect of single interferon and combined interferon-methyl-cholanthrene treatment on various hepatic microsomal enzyme activities in mice

The effect of temperature on enzymatic activity and stability was studied with respect to the monooxygenase activities of aminopyrine-N-demethylase (APD) and p-nitroanisole O-demethylase (pNAD) under incubation conditions for the liver microsomal assay. The activities of S9 liver fractions of mice induced with sodium phenobarbital and β-naphthoflavone were determined during a period of preincubation in a range of temperatures from 30 to 44°C.The greatest value of the mean specific activity was found at 40–42°C for both APD and pNAD. The rapid increase of lipid peroxidation after 1 h of incubation at temperatures higher than 42°C can provide an explanation of the enhancement of the rate of inactivation.In order to determine whether biological response is affected by the modifications induced by temperature in the metabolic activating system, tester strain D7 of Saccharomyces cerevisiae was used to assay the genetic activity of the well known premutagenic agent cyclophosphamide by incubating the mixtures both at the traditional temperature of 37°C and at 42°C. We suggest that the use of more favourable conditions for LMA with respect to enzymatic activity, than the traditional ones could improve the reliability and the sensitivity of such tests.

Differential effects of swimming and running on microsomal metabolism in middle-aged and aged Fischer 344 rats

Cytochrome P-450 ( P-450) content as well as p-nitroanisole (pNA) O-demethylase and UDP-glucuronyltransferase (UDPGT) activities were determined in livers of middle-aged (MA; 12 or 18 months) and aged (24–26 months) rats exercised by either treadmill running or swimming. In addition, aniline hydroxylase activity was measured in MA runners and aged swimmers and compared to respective sham and non-handled controls. Treadmill exercise consisted of running aged and MA rats on a motorized treadmill for 16 and 20 m/min respectively, 60 min/day and 4 times per week, for 8 weeks. Sham rats were placed on the treadmill twice per week for 5 mins at 8 m/min. No differences were found in any parameter comparing sham rats to non-handled controls. Running did not affect body weight or hepatic microsomal protein during the 8-week study. A 33–35% decline in microsomal P-450 content in treadmill exercised MA and aged rats was found. PNA O-demethylase activity was decreased 30% in MA and 45% in aged runners and aniline metabolism was inhibited 21% in MA rats. UDPGT activity was not affected by running in MA or aged rats. Swimming exercise was accomplished by placing the rats in a tank of water (32–33°C) filled to a depth of 2 ft. Swim time was 60 min twice daily, 5 times per week. The aged and MA rats were trained for 6 months and 1 year, respectively. Two control groups, non-swimming sedentary (dry control) and 1 min swim/day sham (wet control), were utilized. MA and aged wet controls and swimmers weighed 8% and 15% less respectively, than MA and aged sedentary rats. Microsomal protein was significantly increased in MA swimmers compared to sedentary (20%) and wet control (35%) bu no change was found with swimming in the aged rats. The results of the enzymatic studies were variable in the MA rats. Increases in P-450 content were found in wet controls (16%) and swimmers (27%) of the MA group, but only the swimming change was significant. No significant change was determined for pNA metabolism between swimmers and wet (22%) or dry (17%) controls. Aged swimmers and wet controls were more consistent, with no change in any of the parameters except aniline metabolism which was significantly increased in wet controls (25%) and swimmers (28%) as compared to dry controls. No significant change in UDPGT activity was measured in either age group of swimmers. Thus, while runing resulted in consistent results including an inhibition of P-450 content and activity within and between age groups, swimming results were variable and most changes included the wet control. Therefore, the results of the swimming study appear to be influenced by some components associated with exposuer to the water which may or may not be related to the exercise itself.

Insecticide resistance in the fall armyworm, Spodoptera frugiperda (J. E. Smith)

A strain of the fall armyworm, Spodoptera frugiperda (J. E. Smith), collected from corn in North Florida showed resistance to commonly used insecticides. Resistance to pyrethroids (permethrin, cypermethrin, cyhalothrin, fenvalerate, tralomethrin, bifenthrin, tetramethrin, and fluvalinate) ranged from 2- to 216-fold; the highest resistance level observed was to fluvalinate. Resistance to organophosphorus insecticides (chlorpyrifos, methyl parathion, diazinon, sulprofos, dichlorvos, and malathion) ranged from 12- to 271-fold; the highest resistance level observed was to methyl parathion. Resistance to carbamates (methomyl, carbaryl, and thiodicarb) ranged from 14- to >192-fold with the highest resistance level being observed with carbaryl. Detoxication enzyme assays revealed that activities of microsomal oxidases (aldrin epoxidase, heptachlor epoxidase, biphenyl hydroxylase, p-nitroanisole O-demethylase, and phorate sulfoxidase) and hydrolases (helicin β-glucosidase and acetylcholinesterase) were 1.4- to 6.6-fold higher in the field strain than in the susceptible strain. Levels of cytochrome P450 and cytochrome b 5 were 1.7- and 1.3-fold higher, respectively, in the field strain than in the susceptible strain. In addition, the bimolecular rate constants for inhibition of acetylcholinesterase by dichlorvos and carbaryl were 2.8- and 3.5-fold higher, respectively, in the susceptible strain than in the field strain. There was no difference in the rate of cuticular penetration of carbaryl in both strains. The results indicated that the broad spectrum of insecticide resistance observed in the field strain was due to multiple resistance mechanisms, including increased detoxication of these insecticides by microsomal oxidases and target site insensitivity such as insensitive acetylcholinesterase.

Inhibitory Effect of Cannabidiol Hydroxy-quinone, an Oxidative Product of Cannabidiol, on the Hepatic Microsomal Drug-Metabolizing Enzymes of Mice.

Cannabidiol hydroxy-quinone (CBDHQ) was identified as an air oxidation product of cannabidiol (CBD). The in vitro incubation of mouse hepatic microsomes with CBDHQ resulted in a decrease of cytochrome P-450 content. CBDHQ inhibited the hepatic microsomal drug-metabolizing enzymes of mice. This inhibitory effect was stronger than that of CBD. CBDHQ (150 microM) inhibited aniline hydroxylase, p-nitroanisole O-demethylase and aminopyrine N-demethylase in the microsomes by 70, 52 and 77%, respectively, whereas the same concentration of CBD caused the inhibition by 39, 30 and 26%, respectively. CBDHQ (91.5 microM) significantly decreased total heme content by 21% and free SH groups by 11% in the microsomes. The results indicate that CBDHQ, which is an oxidation product of CBD, inhibits the hepatic microsomal drug-metabolizing enzymes through the decrease of cytochrome P-450 content.

Lipid peroxidation-induced loss of hepatic drug metabolizing capacity in liver fluke infection: Restoring effects of GSH

Indole-3-carbinol (IC), a compound present in cabbage and other cruciferous vegetables, was fed for 3 wk to young male Sprague-Dawley rats at dietary levels of 50, 500, 5000 and 7500 ppm. Because lower body weights were recorded in the animals fed the two highest levels of IC, pair-fed control groups of rats were used in a second experiment. Increased relative liver weights with enhanced activities of hepatic aminopyrine-N-demethylase, p-nitroanisole O-demethylase and aryl hydrocarbon hydroxylase (AHH) accompanied by higher cytochrome P-450 levels were observed only in the rats fed the two highest levels of IC either ad lib. or in the paired-feeding study. AHH activity increased also in the kidney, with enhanced glucuronide excretion, in the rats fed either of the two highest levels of IC. Intestinal AHH exhibited a significant induction only as a result of the 7500-ppm treatment. The dose-response relationships differed among the tissues. The induction of tissue mixed-function oxidases by only extremely high dietary levels of IC supports the conclusion that this indole is not the specific vegetable constituent that induces these enzymes in rats fed Cruciferae.

Enhanced Metabolism and Knockdown Resistance in a Field Versus a Laboratory Strain of the Soybean Looper (Lepidoptera: Noctuidae)

Soybean looper, Pseudoplusia includens (Walker), larvae collected from a field where permethrin failed to provide adequate control and for which a reduction in susceptibility to permethrin had been demonstrated (3-fold at LC50) were compared with an established laboratory colony with respect to knockdown resistance ( kdr ) and in vitro metabolic capacity for a variety of substrates. The time necessary to achieve 50% knockdown of the field population (22.9 ± 1.3 min) after the topical application of 1 μg permethrin was significantly greater than that required for the laboratory colony (18.4 ± 1.0 min). Rates of metabolism for first-generation larvae from the field population were significantly greater than for larvae from the laboratory culture for substrates of glutathione transferase (l-chloro- 2,4-dinitrobenzene, 2.7-fold), monooxygenases (p-nitroanisole O-demethylase, l.8-fold), and hydrolases ( alpha -naphthyl acetate [1.5-fold], p-nitrophenyl acetate [1.5-fold], and permethrin [1.5-fold]). Significant differences between populations were not observed for NADPH cytochrome c reductase nor acephate hydrolysis. Results of the study indicate that a combination of target site insensitivity and increased activity of several enzymes involved in insecticide metabolism including a trans -permethrin hydrolase may be contributing to the reduced susceptibility of the field population relative to the laboratory colony.

Cadmium deposition and hepatic microsomal activity in mice fed swiss chard grown on municipal incinerator refuse ash

Incineration of municipal solid waste results in the production of millions of tons of ash that may be typically high in heavy metals such as cadmium. Disposal of such ash in landfills capped with soil could lead to absorption of such metals by plants and deposition in foraging animal tissues. In this study, weanling, male mice were fed swiss chard that was grown on soil amended with 10% w/w municipal incinerator refuse ash. Cadmium was taken up by the swiss chard (8.15 ppm, dry wt). The mice fed diets containing 25% of ash-grown chard showed mean kidney and liver concentrations of cadmium (ppm, dry wt), respectively, of 2.80 ± 0.30 and 0.45 ± 0.03. Control mice fed soil-grown chard showed significantly lower kidney and liver concentrations of cadmium, i.e. 0.39 ± 0.02 and 0.05 ± 0.00 ppm. Since refuse incinerator ashes may contain various organic toxicants that can be hepatic microsomal inducers, the relative liver weights and hepatic microsomal aminopyrine N-demethylase and p-nitroanisole O-demethylase activities of mice fed control or ash-grown chard were measured. No consistent increases in these latter parameters were found in the ash-grown chard fed mice as compared with the control animals.