Total Amount Of Cytochrome P-450 Research Articles

In vivo effects of Diosgenin, isolated from Asparagus officinalis, compared to Phenobarbital in rats

Event Abstract Back to Event In vivo effects of Diosgenin, isolated from Asparagus officinalis, compared to Phenobarbital in rats Magdalena Kondeva-Burdina1*, Mitka Mitcheva1, Ilina Krasteva2 and Stefan Nikolov2 1 Medical University-Sofia, Department of Pharmacology, Pharmacotherapy and Toxicology, Bulgaria 2 Medical University-Sofia, Department of Pharmacognosy and Botany, Bulgaria Diosgenin is a well-known antihypercholesterolemic agent. Biliary cholesterol secretion is mediated by different transporters and is stimulated by cholesterol and non-cholesterol steroids – Cholate and Diosgenin. It is known that nuclear receptors CAR and PXR take part in the regulation of hepatic drug metabolism. There are data that Diosgenin increased the expression of lipid-sensing nuclear receptor Pregnane X Receptor (PXR). Phenobarbital (PB) and Phenytoin are the most well known activators of PXR and CAR. PXR has been shown to take part in the regulation of hepatic metabolism – especially biochemically and genetically activation of CYP3A genes. According to these data, it is interesting to investigate in vivo the possible effect of Diosgenin, isolated from Asparagus officinalis, on the activity of some drug-metabolizing enzyme systems, compared to Phenobarbital in rats. Rats were treated with Diosgenin (50 mg/kg p.o./6 days) and Phenobarbital (75 mg/kg i.p./4 days). The total quantity of Cytochrome P450, enzyme activity of Ethylmorphine N-demethylase (EMND) (CYP3A) and Aniline Hydroxilase (AH) (CYP2E1) were measured spectrophotometrically in isolated rat microsomes. As marker of lipid peroxidation and cell protection, we measured the quantity of malondialdehyde (MDA) and reduced glutathione (GSH). Diosgenin and Phenobarbital, administered alone, increased statistically significant, compared to control, the total amount of Cytochrome P450 – with 36% and with 85%, respectively; the activity of EMND with 58% and with 184%, respectively. Diosgenin didn’t influenced the AH activity compared to Phenobarbital. Both Diosgenin and Phenobarbital have no statistically significant effect on GSH level. On the quantity of MDA, Diosgenin has no effect, while Phenobarbital increased it with 27%. Our results showed that Diosgenin, isolated from Astragalus officinalis, exhibited inducible effect similar to the effect of Phenobarbital on the activity of some drug-metabolizing enzyme systems. Keywords: Diosgenin, Phenobarbital, induction, drug-metabolizing enzymes Conference: 8th Southeast European Congress on Xenobiotic Metabolism and Toxicity - XEMET 2010, Thessaloniki, Greece, 1 Oct - 5 Oct, 2010. Presentation Type: Poster Topic: Xenobiotic metabolism Citation: Kondeva-Burdina M, Mitcheva M, Krasteva I and Nikolov S (2010). In vivo effects of Diosgenin, isolated from Asparagus officinalis, compared to Phenobarbital in rats. Front. Pharmacol. Conference Abstract: 8th Southeast European Congress on Xenobiotic Metabolism and Toxicity - XEMET 2010. doi: 10.3389/conf.fphar.2010.60.00145 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 28 Oct 2010; Published Online: 04 Nov 2010. * Correspondence: Dr. Magdalena Kondeva-Burdina, Medical University-Sofia, Department of Pharmacology, Pharmacotherapy and Toxicology, Sofia, Bulgaria, magdalenakondeva@hotmail.com Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Magdalena Kondeva-Burdina Mitka Mitcheva Ilina Krasteva Stefan Nikolov Google Magdalena Kondeva-Burdina Mitka Mitcheva Ilina Krasteva Stefan Nikolov Google Scholar Magdalena Kondeva-Burdina Mitka Mitcheva Ilina Krasteva Stefan Nikolov PubMed Magdalena Kondeva-Burdina Mitka Mitcheva Ilina Krasteva Stefan Nikolov Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Safety assessment of sanguiritrin, alkaloid fraction of Macleaya cordata, in rats

The subchronic safety of sanguiritrin, a mixture of sanguinarine (SA) and chelerythrine (CHE) quaternary benzo[c]phenanthridine alkaloids (QBA), obtained from Macleaya cordata was assessed. Rats were fed a diet containing 120 ppm sanguiritrin (100 ppm QBA) for 109 days. The feed consumption and the animal weight were monitored. The content of QBA in selected tissues and plasma was determined using HPLC. It was evidenced that 2% of QBA were absorbed through the GIT while 98% were excreted in the feces. In plasma, bilirubin, urea, creatinine, glomerular filtration, AST, ALT, GMT, ALP and total antioxidant capacity were determined. In liver, GSH level, lipoperoxidation products, SOD and GPx activities and total amount of cytochrome P450 were evaluated. Damage to nuclear DNA was assessed; a <sup>32</sup>P-postlabeling assay proved that no DNA-adducts were detected in nuclear and mitochrondrial DNA in liver. No adverse effects were observed on rat organism. QBA had no influence on the gut mucosal epithelium, liver tissue and any biochemical parameters tested. Oxidative stress was not manifested during the experiment.

Studies on interactions between functional foods or dietary supplements and medicines. I. Effects of Ginkgo biloba leaf extract on the pharmacokinetics of diltiazem in rats.

The effects of Ginkgo biloba leaf extract (GBE), one of the most widely used herbal dietary supplements in Japan, on the pharmacokinetics of diltiazem (DTZ), a typical probe of cytochrome P450 (CYP) 3A, were examined in rats. The simultaneous addition of GBE to small intestine and liver microsomes inhibited the formation of N-demethyl DTZ (MA), an active metabolite of DTZ produced by CYP3A, in a concentration-dependent manner, with an IC(50) of about 50 and 182 microg/ml, respectively. This inhibition appeared to be caused, at least in part, by a mechanism-based inhibition. Both the rate of formation of MA and total amount of CYP in intestinal or hepatic microsomes after a single oral pretreatment with GBE (20 mg/kg) decreased transiently. The pretreatment significantly decreased the terminal elimination rate constant and increased the mean residence time, after intravenous administration of DTZ (3 mg/kg). Furthermore, it significantly increased the area under the concentration-time curve and absolute bioavailability after oral administration of DTZ (30 mg/kg). These results indicated that the concomitant use of GBE in rats increased the bioavailability of DTZ by inhibiting both intestinal and hepatic metabolism, at least in part, via a mechanism-based inhibition for CYP3A.

Effect of glycerol-induced acute renal failure and di-2-ethylhexyl phthalate on the enzymes involved in biotransformation of xenobiotixs.

The effects of di-(2-ethylhexyl)-phthalate (DEPH) on the levels of cytochrome P-450 and b5 monooxygenases were studied in the rat kidney and liver in acute renal failure induced by glycerol. Intramuscular injection of glycerol (50%,10 ml x kg(-1)) to rats produced proximal tubular damage and acute renal failure. The indicators of renal function, serum urea and creatinine significantly increased (480 and 350 percent, respectively). In control and glycerol-treated animals DEPH had no significant effect on the concentrations of serum urea and creatinine. Twenty-four hours after glycerol injection the total amount of cytochrome P-450 and b5 significantly decreased in renal but increased in liver microsomal fractions. Moreover, 48 and 72 hours after glycerol injection the level of cytochrome P-450 and b5 significantly increased in both organs. A single dose of DEPH (2 ml x kg(-1), i.p.) also elevated the total cytochrome P-450 and b5 in control animals. This enhancing effect of DEPH was additive to that of glycerol in glycerol-induced acute renal failure. These results indicate that DEPH and glycerol evoked pathological changes may affect the metabolism of xenobiotics plus endogenous hormones in the liver and in kidney.

Improvement in liver fibrosis, functionality and hemodynamics in CCl 4-cirrhotic rats after injection of the Liver Growth Factor

Background/Aims: Most substances used in experimental models of cirrhosis are chosen either as protectors of lipid peroxidation, as antifibrogenic agents or as vitamins, among others. In this report, we analyze the improvement produced, in established cirrhosis (CCl 4 plus phenobarbital) in rats, by intraperitoneal injection of Liver Growth Factor, a hepatic mitogen with activity both in vivo and in vitro. Methods: Following confirmation of CCl 4-induced cirrhosis, Liver Growth Factor (4.5 μg per rat×2 injections/week for 3 weeks) was administered to one group of rats (Cirr+LGF). The remaining rats (Cirr) received saline. The groups were compared in terms of serum enzymes, tissue damage, total liver collagen, collagenase activity, microsomal enzyme activities, splanchnic and systemic hemodynamics and portosystemic shunting. Results: Treatment of rats presenting CCl 4-induced cirrhosis with Liver Growth Factor decreased serum aminotransferase levels and increased levels of serum albumin and total protein. The Liver collagen content was lower in rats treated with Liver Growth Factor (2.96 vs 4.32 mg/g liver, p< 0.01). Microscopic studies revealed that the livers of rats receiving Liver Growth Factor showed decreases in fibrosis, necrosis and inflammatory infiltration, as well as a recovery of architectural integrity. Liver function was improved after treatment with Liver Growth Factor, as indicated by the rate constant for elimination of aminopyrine, which increased from 0.0063 to 0.0170 ( p<0.05). This increase was accompanied by a higher total amount of cytochrome P-450 as well as of certain P-450 isoenzymes, especially those that are hormone-dependent, such as P-450 3A. The improved liver histology and function observed in Cirr+LGF rats was associated with decreases in portal pressure (14.4 vs 9.4 mmHg, p<0.01) and portosystemic shunting (55.8 vs 11.5%, p<0.01), as well as increases in mean arterial pressure and systemic vascular resistance, and a reduction in ascites. Conclusions: Administration of the hepatic mitogen, Liver Growth Factor, to CCl 4-cirrhotic rats decreased liver collagen and reorganized the hepatic extracellular matrix, resulting in an improvement in liver function, reduced portal pressure and amelioration of ascites.

Comparative 32P-postlabeling analysis of benzo[a]pyrene--DNA adducts formed in vitro upon activation of benzo[a]pyrene by human, rabbit and rodent liver microsomes.

An interspecies comparison was made of the DNA-adducts formed in vitro upon incubation of rat liver DNA (RL-DNA) with benzo[a]pyrene (BP) in the presence of liver microsomes. Incubations were carried out with RL-DNA, BP (100 microM) and liver microsomes from hamsters, mice, rabbits, rats, 3-methylcholanthrene (3MC) pretreated rats and from humans. To analyse the adduct profiles, the 32P-postlabeling technique with the nuclease P1-enhancement procedure was used. The total amount of adduct formed varied greatly with the species; also the number of adduct spots detected was different, ranging from 1 to 5. In all incubations the BP-N2-deoxyguanosine adduct was formed. Relative to the total adduct level, the level of this adduct varied from 26% with rat, 54% with hamster, 56% with 3MC-pretreated rat, 58% with mouse and 75% with rabbit, to 100% with human liver microsomes. In human liver microsomes both the total amount of cytochrome P-450 per mg microsomal protein and the ethoxyresorufin O-deethylation (EROD) activity were low compared to that in animal liver microsomes. In microsomes from 3MC-pretreated rats the EROD activity was strongly induced. There was no correlation between EROD activity in non-induced microsomes and total adduct level. To compare BP-DNA adduct formation in human white blood cells (WBC) with that in RL-DNA, WBC were incubated with BP and 3MC-pretreated rat microsomes. The adduct profile in WBC-DNA differed from that observed after incubation of RL-DNA: the BP-N2-deoxyguanosine adduct in WBC-DNA accounted for 97% of the total adduct level. It is concluded that the 32P-postlabeling method is a suitable technique to investigate both qualitative and quantitative differences in BP-DNA adduct formation between species. Furthermore, the incubation of microsomes from the liver (or other sources) with a genotoxic agent and isolated DNA or cells can be a useful approach to study the formation and stability of reactive intermediates that are able to bind to DNA, also with respect to differences between species or tissue.

Microsomal ethanol-oxidizing system in Euglena gracilis. Similarities between Euglena and mammalian cell systems

1. 1. ADH activity of Euglena grown with 50 mM ethanol decreased, but MEOS activity increased with a corresponding increase in the total amount of cytochrome P-450. 2. 2. Phenobarbital treatment increased the total amount of cytochrome P-450. 3. 3. CO and KCN, cytochrome P-450 ligands, diminished acetaldehyde formed from ethanol oxidation by MEOS. 4. 4. The amounts of NAD(P)H cytochrome c reductases and cytochrome b 5 type, components of microsomal monooxygenase reaction, have been spectrophotometrically measured. 5. 5. NAD(P)H cytochrome c reductases activities were induced by phenobarbital. 6. 6. DMSO, an inhibitor of rabbit MEOS, inhibited O 2 consumption (11–20%) by Euglena grown with an ethanol, but not a lactate medium. 7. 7. These studies indicate the presence of cytochrome P-450-dependent MEOS in Euglena similar to that in the mammalian hepatic cell.

Induction of cytochrome P-450 isozymes upon repeated administration of nifedipine

In experiments on male Wistar rats it has been found that nifedipine administration at a dose of 10 mg/kg body weight i.p. daily for 20 days did not significantly increase the total amount of cytochrome P-450 but markedly increased the 7 alpha-, 16 beta- and 6 beta-hydroxylation of androstenedione in liver microsomes, suggesting the induction of cytochromes P-450IIA1, P-450IIB1, and P-450IIIA1, respectively. The induction of cytochrome P-450IIIB1 was also confirmed immunochemically with polyclonal antibodies against cytochrome P-450IIB1/B2.

Role of heme in phenobarbital induction of cytochromes P450 and 5-aminolevulinate synthase in cultured rat hepatocytes maintained on an extracellular matrix

When hepatocytes are cultured on matrigel, a reconstituted basement membrane matrix, mRNAs for cytochrome P450 class IIB1/2 and class III genes can be induced by treatment with phenobarbital. We took advantage of this new system to critically evaluate the role of heme as a regulator of these cytochromes P450 and of 5-aminolevulinate synthase (ALA-S), the rate-limiting enzyme in heme biosynthesis. Phenobarbital treatment of rat cultures increased the total amount of cytochrome P450, activities catalyzed by IIB1/2 (benzyloxy- and pentoxyresorufin O-dealkylases) and ALA-S activity, and ALA-S mRNA. Treatments with phenobarbital combined with succinyl acetone, an inhibitor of heme biosynthesis at the step of 5-aminolevulinate dehydrase, blocked the induction of the proteins for cytochrome P450IIB1/2 and cytochrome P450IIIAI, as indicated by spectral, immunological, and enzymatic assays. However, at the same time, succinyl acetone cotreatment failed to inhibit the induction of the mRNAs for cytochrome P450IIB1/2 and cytochrome P450IIIA. Lack of effect on the cytochrome P450 mRNAs was selective inasmuch as treatment with phenobarbital combined with succinyl acetone synergistically increased both ALA-S activity and ALA-S mRNA, presumably by blocking formation of heme, the feedback repressor of ALA-S. Indeed, the increase in ALA-S mRNA caused by the combined treatment was abolished by adding heme itself to the cultures. In contrast to earlier concepts, we conclude that in the intact hepatocyte, phenobarbital-induced cytochrome P450 induction is independent of changes in heme synthesis.

Immunochemical quantification of cytochrome P450IA and IIB subfamilies in the livers of metyrapone-treated rats. Relevance to the ability of metyrapone to prevent the loss of cytochrome P-450 in rat hepatocyte culture

Polyclonal antibodies to the major beta-naphthoflavone (BNF)-inducible form of cytochrome P-450 (P450IA) and to the major phenobarbitone (PB)-inducible form (P450IIB) have been used to quantify the contribution of these subfamilies to the total amount of cytochrome P-450 in rat livers and rat hepatocyte cultures treated with PB, BNF and metyrapone for 24 and 72 h. The P450IA and IIB subfamilies were not detectable (less than 5 pmol/mg of microsomal protein) in the livers of control rats, but administration of BNF resulted in the P450IA subfamily comprising more than 80% of the total hepatic cytochrome P-450. Administration of PB and metyrapone to rats did not elevate the level of this subfamily but elevated the levels of the P450IIB subfamily to 60% and 30% respectively of the total. Thus metyrapone is a 'PB-like' inducer. However, in contrast with their effects in vivo, treatment with PB and metyrapone of rat hepatocytes did not elevate the proportion of the P450IIB subfamily relative to that in untreated cells but rather, like BNF, increased the P450IA subfamily. This would account for the ability of metyrapone to produce in hepatocyte culture, like BNF, a pronounced induction of ethoxyresorufin O-de-ethylase activity, but it does not account for why of all inducers studied only metyrapone can maintain the total cytochrome P-450 content of cultured hepatocytes, or the activity of ethylmorphine N-demethylase. This activity is generally considered to be associated with the P450IIB subfamily, but the lack of effect of metyrapone on this subfamily in hepatocyte culture must suggest that metyrapone is able to prevent the loss of the total amount of the cytochrome by increasing the expression of other cytochromes P-450.

Sex differences in hepatic cytochrome P-450 monooxygenase activities in rainbow trout during an annual reproductive cycle

Hepatic microsomal cytochrome P-450 monooxygenase activities were investigated in rainbow trout during an annual reproductive cycle. The fish were kept in tanks supplied with fresh water at a constant temperature of 10 degrees C. The daily light and darkness cycle was adjusted to follow the natural photoperiod. Sampling was performed once every month for 1 year. Higher benzo(a)pyrene-hydroxylase (or aryl hydrocarbon hydroxylase; AHH), ethoxycoumarin-O-deethylase (ECOD) and ethylmorphine-N-demethylase (END) activities and cytochrome P-450 content were found during the late stage of sexual development in rainbow trout. When monooxygenase activities were expressed on a per cytochrome P-450 basis, sex-dependent differences were observed only for AHH and ECOD activities. It was thus found that sex-dependent variations of END were closely correlated with the total amount of cytochrome P-450. The results indicate that differences exist in hepatic cytochrome P-450 isoenzyme patterns between the sexes in rainbow trout. The similarity of the annual pattern of plasma levels of oestradiol and testosterone to that of sex-dependent differences in the cytochrome P-450 monooxygenases support the contention that sex steroids play a role in regulating the cytochrome P-450 system.

Temperature compensation of hepatic microsomal cytochrome P-450 activity in rainbow trout. I. Thermodynamic regulation during water cooling in autumn

1. 1. The molecular mechanism of temperature compensation in the hepatic polysubstrate monooxygenase system during the water cooling period in autumn was studied in rainbow trout ( Salmo gairdneri Richardson). 2. 2. When 7-ethoxycoumarin O-deethylase and benzo(a)pyrene hydroxylase activities were measured at ambient temperatures, practically no changes could be observed between August (20°C) and November (5°C). When measured at a constant temperature of 18°C over the same period, a three-fold increase occurred. 3. 3. The total amount of cytochrome P-450 remained constant. 4. 4. Cytochrome c reductase activity and the rate of the first reduction step of cytochrome P-450 increased two-fold without a change in the activation energy. 5. 5. Only minor changes in the microsomal fatty acid composition were seen during the adaptation. No phase shifts (breaks) in the temperature dependence of cytochrome c and cytochrome P-450 reduction, were observed either. 6. 6. The temperature compensation can mostly be explained by the reduction of cytochrome P-450 facilitated by an increased reductase/cytochrome P-450 ratio.

Effect of two inducers of cellular differentiation on the glutathione status of human HL-60 promyelocytic leukaemia and A549 lung carcinoma cells

those of Hultmark et al. [2]: PB increases DPNA metabolism 9 times, when calculated per mg protein, and 3 times, when calculated per nmol P-450. But in mice, proteinrelated increase after PB is only 2 times (C57BL/6J) and 3 times (NMRI), and there is no P-450-related increase at all. That means, although PB did augment the total amount of cytochrome P-450, no isoenzyme(s) specific for DPNAdemethylation have been formed. Or: as there is no increase of DPNA-specific activity, DPNA is not a specific substrate for PB in mice. TCPOBOP even decreases DPNA specific activity of cytochrome P-450 (Table lb) in guinea-pigs. PCN decreases it in mice and rats. BNF decreases it in C57BL/6J/6Jbut not in NMRf-mice, and seems to decrease it in rats and guinea-pigs also. From these results may be drawn the following conclusions. (1) In mice, DPNA is not the prominent marker of PB induction as it is in rats. (2) Although TCPOBOP has been described as PB-like inducer [4] its effect on DPNA metabolism is clearly different from that of PB in C57BL/6J mice and rats. (3) DPNA may be called an atypical substrate, because, in C57BL/6J mice, DPNAspecific activity per cytochrome P-450 is decreased by as

Polycyclic aromatic hydrocarbon metabolism in mullets, Chelon labrosus, treated by polychlorinated biphenyls.

Contamination of the ocean by hydrocarbons has become a major environmental problem. Consequently, it is not surprising that residues of potentially toxic xenobiotics, such as benzo(a)pyrene (B(a)P), are present in marine species used as human food. The oxidative metabolism of aromatic hydrocarbons proceeds via intermediate arene oxides. Some of these metabolites are very reactive electrophiles and may interact with cellular macromolecules such as proteins, DNA and RNA. Thus, the ability of an organism to further metabolize arene oxides can be an important protective mechanism against the possible toxic effect of these molecules. The existence of hepatic cytochrome P-450 dependent monooxygenase activities in fish is now well established. The exposure of fish to polychlorinated biphenyls (PCB) increases both the monooxygenase activities and the total amount of cytochrome P-450 in the microsomal fractions of fish liver. The purpose of this study was to compare the toxication-detoxication balance in both control and PCB induced estuarine fish (Grey mullets).

Effects of phenobarbital, 3-methylcholanthrene and polychlorinated biphenyls on sex-specific forms of cytochrome P-450 in liver microsomes of rats.

The effects of treatment with phenobarbital, 3-methylcholanthrene or polychlorinated biphenyls (PCB) on the amounts of sex-specific forms of cytochrome P-450, namely P-450-male and P-450-female, in male and female rats were studied. Although treatment with phenobarbital, 3-methylcholanthrene or PCB markedly increased the total amount of hepatic cytochrome P-450, P-450-male and P-450-female were rather decreased or not significantly changed. Thus, the percentages of P-450-male and P-450-female in the total cytochrome P-450 were decreased in liver microsomes from the treated rats. The increases in specific cytochrome P-450, such as P-448-H, P-448-L, and P-450I-c accounted for the increase in the total amount of cytochrome P-450 in the treated rats. The treatment with phenobarbital or PCB increased the activities of testosterone 16 alpha-hydroxylase, benzo(a)pyrene hydroxylase and aminopyrine N-demethylase more markedly in female rats than in male rats. Similarly, the treatment with 3-methylcholanthrene increased benzo(a)pyrene hydroxylase more markedly in female rats. Therefore, the sex-differences in testosterone 16 alpha-hydroxylase, benzo(a)pyrene hydroxylase, and aminopyrine N-demethylase activities became smaller after the drug treatment. These results indicate that sex-specific P-450-male and P-450-female were unaffected, or even depressed by the agents in some cases.

Induction and catalytic activities of cytochromes P-450 b/ e and P-450 c in the liver microsomes of neonatal rats

1. 1. The activities of cytochrome P-450-dependent monooxygenases has been investigated in the liver microsomes of newborn rats (3–16 days after birth) induced with PB or 3-MC. 2. 2. It has been shown that the induction by PB and 3-MC results in the increase of both the total amount of cytochrome P-450 as determined by the CO-reduced spectrum and the amount of induced forms P-450 b/ e † † In designations of the major forms of cytochrome P-450 induced with PB and 3-MC we have used those of Thomas et al. (1981). Our cytochrome P-450 b/ e possessed the following properties: λ max of the CO-reduced spectrum is 450 nm, M r is 52,000, BPh- N-demethylation rate in the reconstituted system is 70 nmol formaldehyde min −1 nmol P-450 b/ e −1, its contents in PB-microsomes is 40–50% of the general one measured by the CO-reduced state. It is evident that anti- P-450 b used by us reacts with both P-450 b and P-450 e . P-450 c has the following characteristics: λ max of the CO-reduced spectrum is 447.5 nm, M r is 56,000, the rates of BP-hydroxylation and 7-ER-deethylation in reconstituted system are 17 and 7 nmol of product min −1 nmol P-450 c −1, respectively, the contents in 3-MC-microsomes is 60–70%. and P-450 c † respectively. 3. 3. In the course of induction of the specific forms of cytochrome P-450 BP-hydroxylase and 7-ER- O-deethylase activities increased at 3-MC-induction, while BPh- N-demethylase and BP-hydroxylase increased at PB-induction. 4. 4. Analysis of inhibition of monooxygenase reactions with antibodies has showed that only P-450 c was involved in metabolism of BP and 7-ER. Participation of P-450 b/ e in BPh N-demethylation was notably lower in the neonates in comparison to the adult rats. 5. 5. In the one-week-old rats induced with 3-MC a considerable rate of BP hydroxylation and 7-ER- O-deethylation (2–4.5 nmol of product min −1 mg −1) has been observed despite a small amount of P-450 (0.02–0.1 nmol/mg of protein). This fact shows the higher catalytic activity of this cytochrome P-450 in the neonates compared to similar characteristics of P-450 c in the 3-MC-induced microsomes. 6. 6. Metabolism of BP in the PB-microsomes of the neonatal rats was inhibited neither by anti- P-450 b/ e nor anti- P-450 c in contrast to the adults, where this reaction was inhibited by antibodies against P-450 b/ e .

Drug metabolism in diabetic subjects with fatty livers.

The effect of fatty degeneration of liver parenchyma on drug metabolism was investigated in 21 obese non-insulin-dependent diabetic subjects by measuring plasma antipyrine kinetics, hepatic cytochrome P-450, liver size and the extent of fatty infiltration. The hepatic drug metabolising capacity, as measured by total antipyrine clearance and the estimated total amount of cytochrome P-450, was at the same level as in non-diabetic control subjects with normal livers. Relative antipyrine clearance (per unit weight of liver) and cytochrome P-450 concentrations were significantly lower in the diabetics than in controls. The extent of fatty infiltration correlated poorly with the indices of drug metabolism. In non-insulin-dependent diabetics, slight to moderate hepatic fatty infiltration, without more serious structural distortion interfering with hepatic blood flow or hepatocellular function, seems to have only a minor influence on drug metabolism.

Effect of multiple administration of orphenadrine or mono-N-desmethylorphenadrine on cytochrome P-450 catalyzed reactions in the rat.

Multiple administration (i.p.) of orphenadrine or its mono-N-demethylated metabolite, tofenacine (day 1, 20 mg/kg; day 2-5, 30 mg/kg) results in a considerable induction (50%) of the total cytochrome P-450 content. In addition, approximately 6% of the total amount of cytochrome P-450 was found to be blocked by a metabolic intermediate, formed from orphenadrine or tofenacine. Induction is apparent in enhancing the in vitro N-demethylation of aminopyrine and ethylmorphine and the p-hydroxylation of aniline. Pretreatment induced orphenadrine metabolism in vitro. The metabolism of tofenacine, however, was reduced. Probably this is due to a specific inhibition caused by the irreversible interaction of the metabolic intermediate with cytochrome P-450. In vivo, no induction of the aminopyrine metabolism (30 mg/kg, i.v.) is apparent, i.e., no change in the clearance was observed after pretreatment. This is probably due to the presence of relatively high, inhibitory concentrations of tofenacine (in the vicinity of cytochrome P-450). These results show that during chronic administration of orphenadrine or tofenacine, the in vivo disposition of concomitantly ingested compounds is determined by the influence of induction, high substrate and/or metabolite levels and complexation of cytochrome P-450. Moreover, based on these results an hypothesis is put forward in order to explain the phenomenon of product inhibition, which has been suggested to occur in man under chronic orphenadrine dosing conditions.

Effect of cannabidiol on cytochrome P-450 and hexobarbital sleep time

The influences of acute and subacute cannabidiol (CBD) treatment and of subsequent drug withdrawal were investigated on hexobarbital-induced sleep time, on hepatic cytochrome P-450 concentration, on the in vitro formation of carbon monoxide (CO) associated with CBD metabolism, and on the kinetics of aminopyrine N-demethylase metabolism. In acutely treated mice, CBD prolonged sleep time, decreased cytochrome P-450 concentration, decreased the endogenous formation of CO, and increased an apparent K m for aminopyrine N-demethylase activity. In subacutely treated animals, tolerance developed to the effect on sleep time but not to that on cytochrome P-450 concentration nor on the endogenous formation of CO in vitro nor on the K m for the N-demethylase activity. Upon withdrawal from subacute treatment, tolerance to the sleep-time effect was still evident on day 14, but, by day 28, the sensitivity to CBD had returned to normal. In contrast, the cytochrome P-450 concentration returned to normal on day 14 of withdrawal, as did the K m for the N-demethylase activity and the ability of CBD to induce CO synthesis in vitro. The comparative results lead us to conclude that the CBD effect on sleep time does not correlate with either the total amount of cytochrome P-450 or with the CBD depressant effect on the cytochrome.

Effects of clophen A50, 3-methylcholanthrene, pregnenolone-16α-carbonitril and phenobarbital on the hepatic microsomal cytochrome P-450-dependen monooxygenase system in rainbow trout, Salmo gairdneri, of different age and sex

Four groups of rainbow trout (mature male and female, 1 2 - year-old juvenile, and 1 1 2 - year-old juvenile) were injected ip at Days 0 and 3 with 20 mg/kg 3-methylcholanthrene (3-MC), 500 mg/kg Clophen A50 (Cl A50), 40 mg/kg pregnenolone-16α-carbonitrile (PCN), respectively, and examined at Day 14. Further, one group of 1 1 2 - year-old juvenile was injected ip daily with 80 mg/kg phenobarbital (PB) and examined at Day 14. Only 3-MC and Cl A50 markedly elevated the total amount of cytochrome P-450 and the cytochrome P-450-dependent metabolism of paranitroanisole (PNA) and benzo( a)pyrene in the fish liver, whereas no such elevation were observed by PCN or PB. In order to characterize the hepatic cytochrome P-450-dependent metabolism, the Michaelis-Menten kinetic constants of PNA- O-demethylase and the responses of the reactions to α-naphtoflavone, metyrapone, and SKF 525A, in vitro, were studied. These studies (A) showed PCN to significantly elevate apparent K m of PNA metabolism, whereas no alterations of apparent K m were seen by 3-MC or Cl A50, (B) indicated the hepatic reactions to be mediated by several species of cytochrome P-450, (C) indicated the formation of alternative form(s) of cytochrome P-450 species in 3-MC- or Cl A50-induced fish when compared to control fish, (D) showed 3-MC- and Cl A50-induced reactions to be similar in character. The presence of differences due to age and sex in the response to the inducers and in the response to the inhibitors, in vitro, is indicated.