Liver Microsomal Drug Metabolism Research Articles

Liquid Biopsies or Therapeutic Drug Monitoring for CYP Activity Profile Determination.

During pharmacotherapy, knowledge about the actual drug and metabolite concentrations in plasma is often critical. Individual dose adjustments can be performed based on pre-emptive genotyping of certain absorption, distribution, metabolism, and excretion (ADME) genes but also using therapeutic drug monitoring (TDM). Analyses of liquid biopsies for tumor-derived components are well-established and have been found to be a good complement to biopsy examinations. Recently, liquid biopsy-based quantification of cell-free RNA (cfRNA) in plasma exosomes was proposed as a proxy measurement for the expression of different hepatic ADME genes and for the rate of drug metabolism, constituting an alternative to TDM. In this study, we validated these findings by examining the correlation between mRNA expression of eight different CYP genes in liver and the corresponding rate of enzyme-specific drug metabolism in 96 donor-matched liver samples. Analyses of CYP-dependent drug metabolism in liver microsomes in comparison to the level of mRNA expression for the different CYP genes revealed a mean Pearson correlation coefficient of 0.28. The highest correlations (0.33-0.34) were obtained for CYP2D6 and CYP3A4 and the weakest correlations were observed for CYP1A2 and CYP2B6 (0.18-0.21). In all cases, the correlations obtained were too weak to demonstrate a predictive relationship, likely due to different regulatory and post-translational events controlling the rate of enzyme activity. Our results reinforce the notion that, whilst liquid biopsy-based approaches might have utility for prediction of hepatic CYP protein expression, they are not currently an important substitute for TDM.

7‐Dimethylamino‐4‐trifluormethylcoumarin (Coumarin‐152) as a fluorogenic substrate of CYP3A4 and a tool for studying functional interrelationships among human cytochromes P450

In this study we probe the use of 7‐Dimethylamino‐4‐trifluromethylcoumarin (Coumarin 152, C152) as a fluorogenic substrate for in vitro studies of drug metabolism in human liver microsomes (HLM). Examining spectral properties of the formed fluorescent products and using LC‐MS/MS for monitoring C152 metabolism we demonstrate sequential formation of two N‐demethylated products – 7‐Methylamino‐4‐trifluoromethylcoumarin (desmethyl‐Coumarin 152, DC152) and amino‐4‐trifluoromethylcoumarin (Coumarin 151, C151). The predominant product formed by HLM is DC152, which accounts for >70% of C152 metabolized. Using insect cell microsomes containing recombinant P450 enzymes (Supersomes™) we found that C152 is metabolized by CYP2B6, CYP3A4, CYP2C19, CYP1A2 and CYP2C9 listed in the order of decreasing rates of turnover. The KM values exhibited by CYP2B6, CYP3A4, CYP1A2 and CYP2C19 were of the same order of magnitude (1.8 – 4.8 μM). Importantly, the major metabolizers of C152 – CYP2B6, CYP3A4, CYP2C19 and CYP1A2 – demonstrated different ability to catalyze the second demethylation step, with CYP1A2 being the most potent catalyst in this reaction. While over half of the substrate metabolized by CYP1A2 underwent the second demethylation, the fraction of C151 formed by other enzymes did not exceed 30%, similar to what is observed in HLM. Although the rate of turnover of C152 with CYP2B6 is higher than that observed with CYP3A4, the latter appears to be the major metabolizer of C152 due to its high abundance in HLM. This is confirmed by ketoconazole (KCZ) inhibition experiments. We demonstrate that in both CYP3A4 Supersomes™ and HLM over 80% of C152 turnover is suppressed by KCZ, whereas the activities of CYP1A2, CYP2B4 and CYP2C19 with C152 remain largely unaffected. Our results demonstrate that C152 can be used for probing functional interrelationships between multiple cytochrome P450 species in HLM as an example of a substrate that is almost exclusively metabolized by CYP3A4 though having similar affinities to most major human drug‐metabolizing cytochromes P450. In addition, we establish an easy and reliable fluorimetric assay for monitoring and analyzing the metabolism of C152 that allows discriminating between the two sequential steps of its N‐demethylation.Support or Funding InformationThis research was supported by the National Institute On Alcohol Abuse And Alcoholism of NlH under Award Number R21AA024548

Role of Cell Death in Toxicology: Does It Matter How Cells Die?

My research activity started with studies on drug metabolism in rat liver microsomes in the early 1960s. The CO-binding pigment (cytochrome P450) had been discovered a few years earlier and was subsequently found to be involved in steroid hydroxylation in adrenal cortex microsomes. Our early studies suggested that it also participated in the oxidative demethylation of drugs catalyzed by liver microsomes, and that prior treatment of the animals with phenobarbital caused increased levels of the hemoprotein in the liver, and similarly enhanced rates of drug metabolism. Subsequent studies of cytochrome P450-mediated metabolism of toxic drugs in freshly isolated rat hepatocytes characterized critical cellular defense systems and identified mechanisms by which accumulating toxic metabolites could damage and kill the cells. These studies revealed that multiple types of cell death could result from the toxic injury, and that it is important to know which type of cell death results from the toxic injury.

Predicted contributions of cytochrome P450s to drug metabolism in human liver microsomes using relative activity factor were dependent on probes

Contributions of cytochrome P450 (CYP450) isoforms to drug metabolism are often predicted using relative activity factor (RAF) method, assuming RAF values were independent of probe. We aimed to report probe-dependent characteristic of RAF values using CYP3A4 or CYP2C9 probes.Metabolism of four CYP3A4 probes (testosterone, midazolam, verapamil and atorvastatin) and three CYP2C9 probes (tolbutamide, diclofenac and S-warfarin) in human liver microsomes (HLM) and cDNA-expressed recombinant CYP450 (Rec-CYP450) systems were characterized and RAFCL value was estimated as ratio of probe intrinsic clearance in HLM to that in Rec-CYP450. CYP450i contributions to metabolic reaction of a probe were predicted using other probes and compared with data from specific inhibitions. Contributions of CYP3A4 and CYP2C9 to metabolism of deoxypodophyllotoxin and nateglinide were also predicted.RAF values were dependent on probes, leading to probe-dependently predicted contributions. Predicted contributions of CYP3A4 to formations of 6β-hydroxytestosterone, 1′-hydroxymidazolam, norverapamil, ortho-hydroxyatorvastatin and para-hydroxyatorvastatin using other probes were 47.46–219.46%, 21.62–98.87%, 186.49–462.44%, 21.87–101.15% and 53.62–247.97%, respectively. Predicted contributions of CYP3A4 and CYP2C9 to nateglinide metabolism were 8.18–37.84% and 36.08–94.04%, separately.In conclusion, CYP450i contribution to drug metabolism in HLM estimated using RAF approach were probe-dependent. Therefore, contribution of each isoform must be confirmed by multiple probes.

Metabolic inhibition and kinetics of raloxifene by pharmaceutical excipients in human liver microsomes

This study was originally undertaken to establish the in vitro metabolic conditions and then evaluate the effect of pharmaceutical excipients (PEs) on drug metabolism in uridine diphosphoglucuronic acid-supplemented human liver microsomes. Poorly bioavailable raloxifene was chosen as a model drug. Intact drug and its two glucuronide metabolites were successfully isolated using gradient HPLC analysis and LC/MS analysis. Formation of raloxifene metabolites was affected by buffer compositions, incubation time and initial raloxifene concentrations. Under optimized metabolic conditions, 41.0% of raloxifene was converted to its metabolites after 2h incubation. This metabolic inhibition of raloxifene by the PEs occurred in a dose-dependent manner and accordingly formed two glucuronide metabolites. In the metabolic kinetics using Lineweaver-Burk analyses, Cremophor® EL competitively inhibited formation of metabolites while sodium lauryl sulfate (SLS), polyvinylpyrrolidone K30 (PVP) and Tween® 80 significantly inhibited in a mixed competition. Although some PEs showed inhibition on glucuronidation of raloxifene in vitro, their effects on in vivo bioavailability of raloxifene need to be confirmed directly due to the dilution factors and other complicated situations influencing the bioavailability.

CYP2D-related Metabolism in Animals of the Canoidea Superfamily – Species Differences

CYP2D-related drug metabolism in liver microsomes from animals of the Canoidea super family, i.e. mink (Mustela vison), bears (Ursus arctos), foxes (Vulpes vulpes) and dogs, were investigated. Propranolol, bunitrolol and imipramine, which are typically substrates of CYP2D subfamilies, were used in the experiment. All the animals of the Canoidea superfamily that were tested lacked the ability to catalyse 7-hydroxylation of propranolol, which is one of the major metabolic pathways in rats. Stereoselectivity of propranolol metabolism was towards (S)-propranolol in all the reactions of the animals tested with the exception of mink, which showed a selective tendency towards (R)-propranolol in N-dealkylation. As far as metabolic patterns of (R)- and (S)-propranolol are concerned, bears, foxes and dogs are alike, but minks are somewhat different. Liver microsomes from mink showed, among the animals of the Canoidea superfamily, the lowest propranolol hydroxylase activity at 4- and 5-positions and imipramine 2-hydroxylation and {N-}demethylation activities. We could not detect bunitrolol 4-hydroxylation in mink liver microsomes at the low substrate concentration used. We conclude that mink have the lowest activity of CYP2D-related xenobiotic metabolism among the Canoidea superfamily.

Effects of Kampo Extracts on Drug Metabolism in Rat Liver Microsomes: Rhei Rhizoma Extract and Glycyrrhizae Radix Extract Inhibit Drug Oxidation

In the present study, the effects on drug oxidations in rat liver microsomes in vitro using 126 Kampo extracts were investigated. Although the effects of inhibition on drug oxidations were dependent on the Kampo extracts and probe reactions studied, most of the Kampo extracts showed inhibitory effects on both N-demethylations of aminopyrine and erythromycin in rat liver microsomes. Among the Kampo extracts studied herein, Daio-kanzo-to exhibited the most remarkable inhibitory effect on both reactions. The Rhei Rhizoma extract inhibited not only aminopyrine and erythromycin N-demethylations, but also phenacetin O-deethylation, 7-ethoxycoumarin O-deethylation, ethanol oxidation and tolbutamide 4-hydroxy-lation in rat liver microsomes. The Glycyrrhizae Radix extract also showed a remarkable inhibitory effect on phenacetin O-deethylation as well as aminopyrine and erythromycin N-demethylations. In contrast, the Glycyrrhizae Radix extract virtually showed no effect on ethanol oxidation.

Inhibition of drug metabolism in human liver microsomes by nizatidine, cimetidine and omeprazole

1. The inhibitory effects of cimetidine, nizatidine and omeprazole on the metabolic activity of CYP2C9, 2C19, 2D6 and 3A were investigated in human liver microsomes. Both cimetidine and omeprazole inhibited each of the CYP subfamily enzymes; in particular, omeprazole extensively inhibited the hydroxylation of S-mephenytoin (CYP2C19, K1 = 7.1 μM). Nizatidine exhibited no inhibition of any of the CYP isoforms examined.2. Cimetidine inhibited the hydroxylation of tolbutamide but not of diclofenac, whereas omeprazole inhibited the hydroxylation of diclofenac but not that of tolbutamide. The ability to inhibit CYP2C9 varied with incubation time, as measured by the metabolic rate constant for the substrates. Therefore, suitable substrates and incubation times must be selected in inhibition studies examining metabolic clearance and the mechanism of inhibition of these drugs.3. Nizatidine did not inhibit the metabolism of cisapride, glibenclamide, benidipine and simvastatin. Omeprazole inhibited the metabolism of cisapride (K1 = 0.4 μm), glibenclamide (11.7 μm) and benidipine (6.5 μm), whereas cimetidine inhibited the metabolism of glibenclamide (11.6 μm). To avoid drug-drug interactions, care needs to be taken to select suitable medicines for co-administration with anti-ulcer drugs.

Characterization of the independent and combined effects of two inhibitors on oxidative drug metabolism in rat liver microsomes

To evaluate how two inhibitors influence oxidative drug metabolism, this study investigated the inhibitory effects of mexiletine with cimetidine and mexiletine with lidocaine, both individually and in combination, on the oxidative metabolism of two probe substrates, aminopyrine and aniline in rat liver microsomes. Mexiletine was a competitive inhibitor of aminopyrine N-demethylation, whereas cimetidine was a mixed type of inhibitor ( K i = 2.00 ± 0.04 and 0.20 ± 0.02 mM, respectively). For aniline hydroxylation, mexiletine exhibited a mixed type of inhibition, whereas lidocaine was a noncompetitive inhibitor ( K i = 0.60 ± 0.07 and 8.50 ± 0.12 mM, respectively). The combined inhibition of either mexiletine with cimetidine or mexiletine with lidocaine on aminopyrine and aniline metabolism was close to the fully additive effects of the individual compounds when their individual concentrations were below a 2-fold K i concentration, regardless of the apparent kinetic inhibition type. The combined inhibition was less than fully additive when the individual concentrations were twice the K i or above. These results demonstrate that, when two inhibitors of oxidative drug metabolism are combined, both the k i values and the concentrations of inhibitors play important roles in determining the extent of additive inhibition of enzyme activity.

Ketamine as a probe for medetomidine stereoisomer inhibition of human liver microsomal drug metabolism.

Medetomidine (MED) is a novel, selective, alpha 2 adrenergic agonist with potent sedative, hypnotic, and analgesic properties, currently undergoing evaluation as an anesthetic adjuvant. The pharmacologic effects of MED are stereospecific, due entirely to the D-isomer (DMED), whereas the L-isomer (LMED) is essentially inactive. DMED, a 4(5)substituted imidazole, has been shown to inhibit adrenal steroidogenesis and human liver microsomal alfentanil metabolism, reactions mediated by cytochrome P-450. The mechanism of MED inhibition of cytochrome P-450 is unknown. The purpose of this investigation was to determine the mechanism of DMED inhibition of human cytochrome P-450-mediated microsomal metabolism, using ketamine as a probe. Ketamine undergoes extensive hepatic biotransformation and has been used previously to characterize the effects of imidazole anesthetics on human P-450-catalyzed drug metabolism. Ketamine N-demethylation by microsomes from three human livers was measured by gas chromatography-mass spectrometry with selected-ion monitoring. DMED was a potent, competitive inhibitor of S(+) ketamine N-demethylation, with a Ki of 0.11-0.18 microM for the high affinity ketamine demethylase. The IC50 for DMED inhibition of therapeutic concentrations of racemic ketamine (10 microM) was 0.15 +/- 0.02 microM. Preincubation of DMED with microsomes and an NADPH generating system prior to ketamine addition had no additional effect on the inhibition of ketamine demethylase activity, thereby implicating the parent compound rather than a DMED metabolite as the inhibitory species. LMED, although pharmacologically inactive, had a greater inhibitory effect than DMED on racemic ketamine and ketamine enantiomer demethylation at therapeutic concentrations. Spectral studies showed that DMED interacted with microsomal cytochrome P-450 to elicit a Type II binding spectrum.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of antipyrine metabolism by low-dose contraceptives with gestodene and desogestrel

To examine the effects of the two newest monophasic oral contraceptives on liver microsomal drug metabolism, plasma kinetics and urinary metabolite excretion of antipyrine, a model substrate for liver microsomes, were evaluated. Plasma lipid and lipoprotein levels, and in particular the high-density lipoprotein subfractions, were also monitored in view of their apparent regulation by a P450-dependent system. Ten healthy volunteers were treated with each contraceptive for a period of 3 months in a crossover trial. Both contraceptives significantly reduced antipyrine clearance by 34.6% (gestodene) and 43.3% (desogestrel) by impairing the oxidative metabolism, particularly to the 4-hydroxy and 3-hydroxymethyl metabolites, with little difference between the two associations. In addition, with both a comparable highly significant rise of plasma triglyceride levels, apolipoproteins A-I and A-II and the high-density lipoprotein-3 subfraction was observed. Treatment with these new monophasic contraceptives may reduce the metabolism of concomitantly given drugs undergoing oxidative transformations.

Hepatic metabolism of artemisinin drugs—I. Drug metabolism in rat liver microsomes

1. In this communication, metabolism of the semisynthetic antimalarial drugs of the artemisinin class (β-arteether, β-artelinic acid and dihydroartemisinin) in rat liver microsomes, is reported. 2. Dihydroartemisinin was the major early metabolite of arteether (57%) and artelinic acid (80%); in addition, arteether was hydroxylated in the positions 9α- and 2α- of the molecule. 3. Dihydroartemisinin was further metabolized by extensive hydroxylation of its molecule; we were able to identify four hydroxylated derivatives of DQHS, but not the exact positions of the hydroxyl groups. 4. The rates of NADPH-supported metabolism of arteether, artelinic acid and dihydroartemisinin in rat liver microsomes were: 4.0, 2.5 and 1.3 nmol/min/mg of microsomal protein, respectively. 5. The apparent affinity constants of arteether and artelinic acid for the microsomal metabolizing system, calculated from the rates of product formation, were 0.54 mM and 0.33 mM (for arteether) and 0.11 mM (for artelinic acid), respectively. The appearance of two affinity constants indicated that arteether was metabolized by two different isoenzymes of cytochrome P-450 in rat liver microsomes.

Inhibition of Hepatic Microsomal Drug Metabolism in Rats by Five Calcium Antagonists

The effects of five calcium antagonists, verapamil, diltiazem, nifedipine, darodipine and isradipine, on rat liver microsomal drug metabolism in vitro and in vivo were studied. All compounds prolonged hexobarbital-induced sleeping time in a dose-dependent manner (doses 3.0 and 30.0 mg/kg, except nifedipine 0.3 and 3.0 mg/kg) and inhibited cytochrome P450-dependent N-demethylation of aminopyrine in vitro in rat liver microsomes. The incubation of all compounds with microsomes resulted in the apparent formation of formaldehyde, suggesting either N- or O-demethylation. Diltiazem, isradipine and darodipine gave rise to a type I spectral change. Nifedipine seemed to produce a type II spectral change. A spectrum of verapamil changed from a type I to a type II as concentration increased. These results indicate that all calcium antagonists studied interact with P450 and are in vitro inhibitors of microsomal drug metabolism in the rat and the inhibition brings out pharmacokinetic drug-drug interactions in vivo.

Comparison of microsomal drug-metabolizing enzymes in 14 rat inbred strains

Drug metabolic capacity in liver microsomes of 14 rat inbred strains was investigated. Cytochrome P-450 content as well as the following enzyme activities were measured: NADPH cyt. c(P-450) reductase (Red.), aminopyrine N-demethylase (APDM), ethoxycoumarin O-deethylase (ECOD), 1-naphthol: UDP-glucuronosyltransferase (NGT) and hydrolysis of acetylsalicylic acid (ASA; measured at pH5.5 and pH7.4). All enzymes measured were found to exhibit statistically significant inter-strain differences. In males the enzyme activities varied over a 7.3-fold (ECOD) to 1.4-fold (cytochrome P-450) range. Other inter-strain differences were generally larger than 2-fold: ASA-hydrolysis at pH 5.5 and 7.4 (3.9- and 3.3-fold variation, respectively), NGT and Red. (2.1-fold variation) and APDM (1.8-fold variation). In females similar, but somewhat smaller inter-strain differences were observed. Correlations between different enzyme activities were generally poor (correlation coefficients r < 0.7). An exception was the correlation between ASA-hydrolysis at pH 5.5 and pH 7.4 ( r = 0.79). We conclude that ASA hydrolysis at pH 5.5 and 7.4 is mediated by the same enzyme or by coregulated enzymes and that all other activities are mediated by different or differentially regulated enzymes. Based on analysis of variance and subsequent inter-strain comparisons, all strains appear to express a unique profile of liver microsomal drug metabolism. No two strains are identical with respect to all activities measured. We suggest that differences between inbred rat strains and particularly the difference in balance between different enzymes in various strains can be used advantageously in pharmacological and toxicological experiments.

Liver microsomal drug metabolism in ethanol-treated hamsters.

Administration of ethanol in drinking water to Syrian golden hamsters for 1-3 weeks caused alterations of microsomal cytochrome P-450-dependent monooxygenase activities in the liver accompanied by a slight elevation in cytochrome P-450 content. Ethanol treatment resulted in an increase in the activities for ethanol oxidation, aniline p-hydroxylation and dimethylnitrosamine N-demethylation. In particular, when dimethylnitrosamine was used as a substrate, the rate of formaldehyde formation was enhanced by 2- to 2.7-fold, while ethanol oxidation and aniline p-hydroxylation were increased by 1.5- to 2- and 1.2- to 1.3-fold, respectively. On the other hand, the activities of 7-ethoxycoumarin O-deethylase, benzphetamine N-demethylase and benzo[a]pyrene 3-hydroxylase were apparently decreased after ethanol treatment. These results for hamsters were significantly different from those reported for rats.

The influence of in vitro procedures which diminish NADPH cytochrome c reductase activity on oxidative drug metabolism in lung and liver microsomes

Rabbit lung and liver microsomes were subjected to three procedures which decreased NADPH cytochrome c reductase activity; flavoprotein antibody, trypsin and subtilisin digestion. The effects on benzphetamine and p-nitroanisode demethylation and amine metabolic-intermediate complex formation were investigated. In general, the proteolytic digestion had a greater inhibitory effect on oxidation reactions for a given loss of NADPH cytochrome c reductase activity than did flavoprotein antibody; and of the two proteases, subtilisin, which also diminises the cytochrome b 5 reduction pathway, had a greater inhibitory effect than trypsin. Subtilisin digestion had similar effects in both liver and lung microsomes; a loss of flavoprotein without a loss of cytochrome P-450; but whereas all three oxidative reactions decreased in unison as the flavoprotein was lost in the liver, benzphetamine demethylation was less susceptible to flavoprotein depletion than the other two reactions in lung microsomes. With trypsin digestion flavoprotein was removed without loss of cytochrome P-450 only in lung microsomes; in liver microsomes the cytochrome P-450 was susceptible to tryptic degradation. In lung microsomes, benzphetamine and p-nitroanisole demethylations were less susceptible to flavoprotein loss than metabolic-intermediate complex formation.

Inhibition of drug metabolism by the antimalarial drugs chloroquine and primaquine in the rat

The effect of the antimalarial drugs chloroquine (CQ) and primaquine (PQ) on rat liver microsomal drug metabolism has been studied in vitro and in vivo. After acute administration, PQ increased pentobarbitone sleeping time in a dose-related manner [control, 94.0 ± 9.4 min; 10mg/kg, 137.0 ± 2.4 min; 20mg/kg, 197.0 ± 7.5 min; 50 mg/kg, 269.0 ± 2.9 min (means ± S.E.M.)], prolonged zoxazolamine paralysis time (control, 140.0 ± 10.0 min; 50 mg/kg, 341.5 ± 25.6 min) and decreased antipyrine blood clearance from 2.17 ± 0.19 to 0.86 ± 0.12 ml/min. CQ showed no effect on pentobarbitone sleeping time or zoxazolamine paralysis time, but decreased antipyrine clearance from 2.17 ± 0.19 to 1.11 ± 0.18 ml/min. Both drugs inhibited aminopyrine N-demethylase activity, although the concentration required to produce 50% inhibition was much greater for CQ (10 mM) than for PQ (approximately 0.1 mM). Lineweaver-Burk plots showed that CQ inhibited competitively whereas PQ inhibition was apparently non-competitive. Ethoxyresorufin O-deethylase activity decreased by about 40 and 50% in the presence of CQ and PQ respectively (250 nM, equimolar with substrate). There was no evidence of induction following chronic administration of CQ and PQ (50 mg/kg/day for 4 days). There was an apparent decrease in cytochrome P-450 content and aminopyrine N-demethylase activity was decreased. These results demonstrate that PQ and CQ inhibit hepatic drug metabolism both in vitro and in vivo and that PQ appears to be the more potent inhibitor.

The effect of cimetidine on in vitro and in vivo microsomal drug metabolism in the rat

The effect of cimetidine on rat liver microsomal drug metabolism in vitro and in vivo was studied. Cimetidine inhibits aminopyrine N-demethylation and benzo[ a]pyrene hydroxylation in a noncompetitive manner with inhibition constants between 1 and 10 mM. Benzo[ a]pyrene hydroxylation in liver microsomes from 3-methylcholanthrene-pretreated rats is not appreciably inhibited by cimetidine indicating some specificity in terms of different cytochrome P-450 forms. Cimetidine gives rise to a type II spectral change with a spectral dissociation constant of about 0.1 mM. The prolonged administration of cimetidine does not result in the induction of hepatic drug metabolism. Pretreatment of rats with cimetidine prolongs aminopyrine half-life and hexobarbital sleeping time. These results demonstrate that cimetidine is an in vitro inhibitor of microsomal drug metabolism in the rat and this inhibition leads to pharmacokinetic drug-drug interactions in vivo.

Selective induction of rat liver microsomal cytochrome P-448 by 3,4,5,3',4'-pentachlorobiphenyl and its effect on liver microsomal drug metabolism.

Pretreatment of rats with 3, 4, 5, 3', 4'-pentachlorobiphenyl (PenCB) at a single i.p. dose of 10 mg/kg caused a marked increase of cytochrome P-448 (P448) in liver microsomes. The liver microsomes of PenCB-treated rats (PenCB-microsomes) showed similar but somewhat different properties from those of 3-methylcholanthrene (MC)-treated rats (MC-microsomes) with respect to its CO difference spectrum, electrophoretic pattern on sodium dodecyl sulfate (SDS)-polyacrylamide gel and immunochemical reactivity with the antibodies prepared against the hemoproteins from phenobarbital (PB)- or β-naphthoflavone (BNF)-treated rat liver microsomes. Furthermore, PenCB pretreatment decreased not only demethylation of aminopyrine and codeine, but also hydroxylation of aniline, to which PenCB-microsomes showed lower affinity than MC-microsomes. Trace amounts of PenCB inhibited the metabolism of aniline and aminopyrine by MC-microsomes but not by untreated rat liver microsomes. On SDS-polyacrylamide gel electrophoretograms the ratio of cytochrome P450 (P450) to P448 in PenCB-microsomes was much smaller than that in MC-microsomes. These results may suggest that a part of the differences between the catalytic properties of PenCB- and MC-microsomes is due to the inhibitory effect of residual PenCB in PenCB-microsomes or/and the marked change of a relative ratio of P450 to P448 in PenCB-microsomes. A possibility that PenCB may induce a different P448 from that induced by MC is also discussed.

56 - Developmental change of activities of drug metabolism in fetal liver microsomes with advancing gestation

56 - Developmental change of activities of drug metabolism in fetal liver microsomes with advancing gestation