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PubMed1988

Regional differences in lipid composition and incorporation of saturated and unsaturated fatty acids into microsomal membranes of rat small intestine.

Incorporation of [1-14C]palmitic (16:0) and [1-14C]linoleic (18:2 omega 6) acids into microsomal membranes of proximal (jejunum) and distal (ileum) regions of rat small intestine was investigated, and the lipid composition, including fatty acid profiles of membrane phospholipids, was determined. Jejunal microsomes contained significantly higher amounts of total phospholipids, phosphatidylcholine, and phosphatidylinositol, and lower amounts of cholesterol and sphingomyelin when compared with ileal microsomes. Jejunal microsomal phospholipids contained higher levels of stearic (18:0), 18:2 omega 6, and eicosapentaenoic (20:5 omega 3) acids followed by reduced levels of oleic (18:1 omega 9), arachidonic (20:4 omega 6), and docosahexaenoic (22:6 omega 3) acids when compared with those from the ileum, except for phosphatidylinositol where no significant difference between 20:4 omega 6 content of each site was observed. In both jejunal and ileal microsomes, incorporation of [1-14C]18:2 omega 6 was significantly higher than that of [1-14C]16:0. Incorporation of both [1-14C]16:0 and [1-14C]18:2 omega 6 was significantly higher in jejunal microsomal lipid fractions (phospholipids, diacylglycerols, triacylglycerols) when compared with the ileal microsomal fraction. These data suggest that (1) jejunal and ileal microsomal membranes differ from each other in terms of lipid composition and lipid synthesis, (2) site variations in the specificity of acyltransferases for different fatty acids exist, and (3) higher delta 9-, delta 6-, delta 5-, and delta 4-desaturase activities exist in ileal compared with jejunal enterocytes.

Animals↗
PubMed2004

Correlation between UDP-glucuronosyltransferase genotypes and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone glucuronidation phenotype in human liver microsomes.

The nicotine-derived tobacco-specific nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone, is one of the most potent and abundant procarcinogens found in tobacco and tobacco smoke, and glucuronidation of its major metabolite, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), is an important mechanism for 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone detoxification. Substantial interindividual variability in urinary NNAL glucuronide formation has been observed in smokers and tobacco chewers. To determine whether genetic variations may play a role in this interindividual variability, NNAL-glucuronidating activities were analyzed in 78 human liver microsomal specimens and compared with the prevalence of missense polymorphisms in the two major NNAL-glucuronidating enzymes UGT1A4 and UGT2B7. In vitro assays using liver microsomal specimens from individual subjects demonstrated a 70- and 50-fold variability in NNAL-N-Gluc and NNAL-O-Gluc formation, respectively, and a 20-fold variability in the ratio of NNAL-N-Gluc:NNAL-O-Gluc formation. Microsomes from subjects with a homozygous polymorphic UGT1A4(24Thr)/UGT1A4(24Thr) genotype exhibited a significantly higher (P < 0.05) level of NNAL-N-Gluc activity compared with microsomes from subjects with the wild-type UGT1A4(24Pro)/UGT1A4(24Pro) genotype, and a significantly higher (P < 0.05) number of subjects with liver microsomes having high NNAL-N-Gluc formation activity contained the UGT1A4(24Thr)/UGT1A4(24Thr) genotype. Microsomes from subjects with the homozygous polymorphic UGT2B7(268Tyr)/UGT2B7(268Tyr) genotype exhibited a significantly lower level (P < 0.025) of NNAL-O-Gluc activity when compared with microsomes from subjects with the wild-type UGT2B7(268His)/UGT2B7(268His) genotype, and a significantly (P < 0.05) higher number of subjects with liver microsomes having low NNAL-O-Gluc formation activity contained the UGT2B7(268Tyr)/UGT2B7(268Tyr) genotype. These data suggest that the UGT1A4 codon 24 and UGT2B7 codon 268 polymorphisms may be associated with altered rates glucuronidation and detoxification of NNAL in vivo.

Codon↗
PubMed1968

Effects of a short-term fast on albumin synthesis studied in vivo, in the perfused liver, and on amino acid incorporation by hepatic microsomes.

Carbonate-(14)C was used to label the hepatic intracellular arginine pool and direct measurement of albumin synthesis was made in six rabbits before and after an 18-36 hr fast. 18 perfusion studies were performed with livers derived from fed and fasted rabbits (18-24 hr). Microsomal amino acid-incorporating ability with leucine-(3)H and phenylalanine-(14)C was compared in 17 studies, using microsomes isolated from livers taken from fed and fasted rabbits and from isolated perfused livers whose donors were fed and fasted. Albumin synthesis is rapidly inhibited by fasting. Albumin synthesis decreased 33% in vivo and 53% in the perfused liver. The microsomes from perfused livers taken from fed animals did not demonstrate a significantly reduced capacity to incorporate leucine-(3)H or phenylalanine-(14)C into protein. Microsomes derived from perfused and nonperfused livers whose donors were fasted incorporated 32-54% less tracer than microsomes obtained from fed donor rabbits. Microsomes separated from perfused livers removed from fed and fasted rabbits responded to polyuridylic acid stimulation and phenylalanine-(14)C incorporation rose from 58 to 171%. An 18-36 hr fast inhibits albumin production in vivo and in the perfused liver. The microsomal system is less active in the fasted state and perfusion per se does not inhibit the microsomal response.

Albumins↗
PubMed1979

Effects of glucocorticoids on microsomal membrane synthesis in hepatocytes from adrenalectomized rats.

Biochemical and morphological studies were performed on livers from normal, adrenalectomized (ADX), and ADX and dexamethasone (DEX)-treated rats to investigate the effects of glucocorticoids on microsomal membrane synthesis. Overnight fasted normal, ADX and ADX rats treated 2 or 4 h with DEX received [3H]leucine and [14C]glycerol. Livers were removed, and tissue specimens were prepared for electron microscopy and tissue fractionation. Liver microsomal subfractions were prepared and subsequently washed to produce rough and smooth microsomal membranes. Radioactivity and membrane composition were determined, and glucose-6-phosphatase activity was measured in washed microsomal membranes. Adrenalectomy caused decreased microsomal membrane synthesis. Two and 4 h of DEX administration restored microsomal membrane synthesis to normal levels. ADX also caused an alteration in composition of the microsomal membranes (reflected in decreased phospholipid-protein ratios), which was restored to normal levels by 4 h after DEX administration. The earliest effects of the hormone on membrane synthesis were observed in smooth microsomes as part of a smooth endoplasmic reticulum (SER) proliferation. These findings were supported by observations made with the electron microscope. The proliferating SER was enriched in at least one component: glucose-6-phosphatase. Although the specific relationship of SER to glucocorticoid action remains unclear, the interpretation is offered that SER proliferation and alteration in glucose-6-phosphatase distribution are component parts of the total response of the hepatocyte to glucocorticoids.

Adrenalectomy↗
PubMed1985

Differences in microsomal steroid metabolism between the inner and outer zones of the guinea pig adrenal cortex.

Previous investigations established that cells isolated from the outer zone (zona fasciculata + zona glomerulosa) of the guinea pig adrenal cortex produced far more cortisol and androstenedione than those from the inner zone (zona reticularis). Studies were done to determine whether differences in microsomal metabolism might contribute to the zonal differences in steroid secretion. Cytochromes P-450 and b5 concentrations were greater in inner zone microsomes as were the magnitudes of the type I difference spectra produced by progesterone and 17 alpha-hydroxyprogesterone. Basal NADPH-cytochrome P-450 reductase activity was greater in the outer zone, but steroid substrates (progesterone, 17 alpha-hydroxyprogesterone) increased reductase activity in the inner zone and decreased activity in the outer zone. 21-Hydroxylase activity was far greater in inner than outer zone microsomes, but 17 alpha-hydroxylase activity was greater in the outer zone. As a result, progesterone was converted primarily to 17 alpha-hydroxyprogesterone by outer zone microsomes, but 11-deoxycorticosterone was the major metabolite produced by inner zone preparations. In addition, with 17 alpha-hydroxyprogesterone as substrate, the major product produced by outer zone microsomes was androstenedione, indicating relatively high C17-20-lyase activity. Inner zone microsomes by contrast, converted 17 alpha-hydroxyprogesterone primarily to the 21-hydroxylated metabolite, 11-deoxycortisol, with little production of androstenedione. The rate of conversion of pregnenolone to progesterone was also greater with outer than inner zone microsomes. The results suggest that differences in the patterns of microsomal steroid metabolism contribute to the greater secretion of cortisol and androstenedione by adrenocortical outer zone cells than by inner zone cells.

Adrenal Cortex↗
PubMed1988

Effects of tocopherol depletion on the regional differences in adrenal microsomal lipid peroxidation and steroid metabolism.

Studies were done to assess the contribution of alpha-tocopherol to the regional differences in microsomal lipid peroxidation (LP) and steroid metabolism in the guinea pig adrenal cortex. In normal guinea pigs, ferrous ion (Fe2+)- and ascorbic acid-induced LP are far greater in microsomal preparations from the inner adrenal zone (zona reticularis) than in those from the outer zones (zona fasciculata plus zona glomerulosa). The amounts of unsaturated fatty acids, substrates for LP, are similar in the two zones, but alpha-tocopherol concentrations are 4-5 times greater in outer than inner zone microsomes. Tocopherol depletion by dietary deprivation had little effect on LP in vitro in inner zone microsomes, but substantially increased LP in outer zone preparations. As a result, tocopherol deficiency eliminated the zonal differences in microsomal LP. Unsaturated FFA concentrations were lower in tocopherol-deficient microsomal preparations than in those from tocopherol-sufficient animals, suggesting peroxidative losses in vivo. Tocopherol deficiency decreased steroid C-17,20 lyase activity in outer zone microsomes, but had no effect on activity in inner zone preparations, eliminating the normal zonal difference in activity (outer greater than inner). The results indicate that alpha-tocopherol is a major determinant of adrenal LP and is responsible for the regional differences in microsomal LP in guinea pig adrenal cortex; the effects of ascorbic acid on LP in each zone are also affected by alpha-tocopherol. alpha-Tocopherol may influence the functional zonation of the adrenal cortex by selectively protecting outer zone steroidogenic enzymes from oxidative degradation.

Adrenal Glands↗
PubMed1989

In vitro metabolism of 25-hydroxyvitamin D3 by human trophoblastic homogenates, mitochondria, and microsomes: lack of evidence for the presence of 25-hydroxyvitamin D3-1 alpha- and 24R-hydroxylases.

In vitro studies were performed to assess the ability of term human trophoblastic tissue to metabolize 25-hydroxyvitamin D3 (25OHD3) and to compare this metabolism to that occurring in porcine renal mitochondria and microsomes. Human trophoblastic homogenates, containing a NADPH-generating system, were able to produce 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] at a rate of 225 pg/mg protein.h, but did not produce detectable quantities (less than 20 pg/mg protein.h) of 24,25-dihydroxyvitamin D3 [24,25-(OH)2D3]. Similarly, mitochondria and microsomes isolated from term human trophoblastic tissue produced 1,25-(OH)2D3 [249 +/- 156 and 199 +/- 82 (mean +/- SD) pg/mg protein.h, respectively] in the presence of an NADPH-generating system, but failed to produce detectable quantities (less than 200 pg/mg protein.h) of 24,25-(OH)2D3. The production of 1,25-(OH)2D3 from the trophoblastic mitochondria and microsomes could be increased by adding 140,000 x g trophoblastic cytosol to the subcellular incubation tubes. This treatment had no effect on the production of 24,25-(OH)2D3. The component(s) present in trophoblastic cytosol responsible for the increased 1,25-(OH)2D3 production by trophoblastic mitochondria and microsomes was shown to be heat labile, trypsin resistant, and less than 1000 mol wt in size. Comparing characteristics of the porcine renal 1 alpha- and 24R-hydroxylase systems with those of the human trophoblastic system revealed that 1) 1,2-dianilinoethane and EDTA totally blocked synthesis of 1,25-(OH)2D3 in trophoblastic mitochondria and microsomes, but had no effect on the synthesis of 1,25-(OH)2D3 by renal mitochondria; and 2) ketoconazole greatly inhibited the synthesis of 1,25-(OH)2D3 and 24,25-(OH)2D3 by renal mitochondria, but had no effect on the production of 1,25-(OH)2D3 by trophoblastic mitochondria or microsomes. Finally, production of 24,25-(OH)2D3 could not be demonstrated in trophoblastic homogenates, mitochondria, or microsomes, while the production of this compound was readily evident in renal mitochondria, but not microsomes. The results of this study question the existence of the 25-hydroxyvitamin D3-1 alpha- and 24R-hydroxylase systems in the trophoblastic portion of the human placenta. This study also suggests that 1,25-(OH)2D3 can be produced in vitro by a mechanism other than enzymatic 1 alpha-hydroxylation. The possibility exists that the mechanism involves the insertion of oxygen at the 1 position of 25-(OH)D3 by free radical chemistry.

Animals↗
PubMed2005

Effect of nonspecific binding to microsomes and metabolic elimination of buprenorphine on the inhibition of cytochrome P4502D6.

Recently, the pharmaceutical industry has employed the high-throughput method for the evaluation of cytochrome P450 (CYP) inhibition, using a combination of the heterologously expressed enzyme and a fluoregenic substrate. When buprenorphine (BN), a potent mixed agonist-antagonist analgesic, was evaluated by this method, it exhibited potent inhibition of CYP2D6 with an IC50 value of 0.25 microM in recombinant CYP2D6-expressing insect cell microsomes (rCYP2D6 microsomes). In contrast, the IC50 value was 22.7 microM in human liver microsomes (HLM) using a classical method. Although the substrate concentrations in each study were set to near the Km values, there was a large discrepancy in IC50 values. When we investigated the effect of nonspecific binding to microsomes on the inhibitory potency, with a view to clarifying this discrepancy, the unbound fraction in microsomes (fu,mic) was 0.06-0.21 and 0.99 in HLM and rCYP2D6 microsomes, respectively. The corrected IC50 value (1.74 microM) using free BN concentrations was much smaller than the uncorrected value. On the other hand, it was observed that the concentration of BN in HLM decreased rapidly due to metabolism by CYP3A4 while that in rCYP2D6 microsomes decreased only slightly. We then investigated the effect of incubation time on the inhibitory potency, since the rapid elimination of BN in HLM could have been a cause of the discrepancy. The IC50 value for BN was observed to decrease slightly from 22.7 to 17.1 microM, following the shortening of the incubation time from 10 to 2 min in HLM. We conclude that nonspecific binding to microsomes of the inhibitor could affect the inhibitory potency against CYP2D6. If this factor is considered, a more precise estimate of the risk of adverse drug interaction could be achieved.

Animals↗
PubMed1989

7-ethenyloxycoumarin as a new substrate for fluorophotometric assay of hepatic microsomal epoxidizing activities.

7-Ethenyloxycoumarin (7-vinyloxycoumarin, VOC) was metabolized by rat liver microsomes in the presence of a reduced nicotinamide adenine dinucleotide phosphate-generating system to 7-hydroxycoumarin (HOC) and glycolaldehyde via the unstable epoxide, 7-(epoxyethoxy)coumarin, as an obligatory intermediate which had a half life of 5.4 min in 0.1 M phosphate buffer, pH 7.4, at 37 degrees C. The epoxide of VOC accumulated in the microsomal incubation mixture in the presence of the epoxide hydrolase inhibitor, 3,3,3-trichloropropene 1,2-oxide, was isolated and identified. HOC and glycolaldehyde were auto-decomposition products of the putative highly unstable intermediate, 7-(1',2'-dihydroxyethoxy)coumarin, mostly formed by microsomal epoxide hydrolase from the epoxide. Direct fluorophotometry of HOC made it possible to determine epoxidizing activities of very small quantities of the microsomes from untreated rat liver (greater than or equal to 5 micrograms protein). VOC was epoxidizied by rat liver microsomal cytochrome P-450, inducible by 3-methylcholanthrene (3-MC) and phenobarbital (PB), and the microsomal epoxidation reactions were inhibited by IgG preparations raised against the major cytochrome P-450 components isolated from 3-MC- and PB-pretreated rat liver microsomes. In the untreated, 3-MC- and PB-pretreated rat liver microsomes, at least two monooxygenase components with different affinity were strongly suggested by a kinetic study, carried out using the antibodies, to be involved in the epoxidation of VOC.

Animals↗
PubMed1989

Activation of liver microsomal glutathione S-transferase activity by heating.

The effect of heating on rat liver microsomal glutathione S-transferase activity was investigated. The microsomal glutathione S-transferase activity increased with an elevation of temperature and reached a maximal level at 50 degrees C for 30 min, at which point the enhancement was 2.6-fold. The microsomal glutathione S-transferase activity had one low Km value for 1-chloro-2,4-dinitrobenzene at room temperature. However, two Km values were observed in heated microsomes. N-Ethylmaleimide increased microsomal glutathione S-transferase activity 7.7-fold and a 2.5-fold increase in the activity was observed even after heating of microsomes at 55 degrees C for 10 min. On the other hand, there was no additional activation in the heated-microsomal glutathione S-transferase by glutathione disulfide or diamide with glutathione. The increased activity of the microsomal glutathione S-transferase activity is activated by heating through a mechanism different from the activation caused by sulfhydryl reagents.

Animals↗
PubMed1989

Effect of the calcium-binding protein regucalcin on the Ca2+ transport system in rat liver microsomes: the protein stimulates Ca2+ release.

The effect of the calcium-binding protein regucalcin on the Ca2+ transport system in the liver microsomes from fed rats was investigated. Ca2+ transport was assayed by the method of Millipore filtration to estimate microsomal 45Ca2+ accumulation following addition of 10 mM adenosine triphosphate (ATP). 45Ca2+ uptake was retarded by the presence of regucalcin (1.0-4.0 microM). This retardation was remarkable at 1 min after regucalcin addition, while appreciable retardation was no longer seen at 5 min. Regucalcin (2.0 microM)-induced retardation of 45Ca2+ uptake was prevented by the presence of calmodulin (5 micrograms/ml). Calmodulin alone (1 and 5 micrograms/ml) caused a significant increase in 45Ca2+ uptake at 3 min after the start of incubation. Also, regucalcin (2.0 microM)-induced retardation of 45Ca2+ uptake was completely blocked by the presence of a Ca2(+)-trapping agent, oxalate (3 mM). On the other hand, 45Ca2+, which accumulated in microsomes during 5 min after ATP addition, was markedly released by the addition of regucalcin. This release was dose-dependent (0.5-4.0 microM). Guanosine triphosphate (GTP; 10-100 microM) caused a significant release of 45Ca2+ from the microsomes. The presence of regucalcin (2.0 microM) further enhanced the GTP effect. Regucalcin (2.0 microM)-induced release of 45Ca2+ was not blocked by the presence of the protein thiol-protecting agent dithiothreitol (0.1 mM). The presence of oxalate (3 mM) completely blocked the effect of regucalcin on 45Ca2+ release from the microsomes. These results indicate that regucalcin stimulates Ca2+ release from liver microsomes, and that the protein retards the microsomal Ca2+ uptake. The present study suggests that regucalcin can regulate the Ca2+ transport system in rat liver microsomes.

Animals↗
PubMed1994

Activation of microsomal glutathione S-transferase in tert-butyl hydroperoxide-induced oxidative stress of isolated rat liver.

The activation of microsomal glutathione S-transferase in oxidative stress was investigated by perfusing isolated rat liver with 1 mM tert-butyl hydroperoxide (t-BuOOH). When the isolated liver was perfused with t-BuOOH for 7 min and 10 min, microsomal, but not cytosolic, glutathione S-transferase activity was increased 1.3-fold and 1.7-fold, respectively, with a concomitant decrease in glutathione content. A dimer protein of microsomal glutathione S-transferase was also detected in the t-BuOOH-perfused liver. The increased microsomal glutathione S-transferase activity after perfusion with t-BuOOH was reversed by dithiothreitol, and the dimer protein of the transferase was also abolished. When the rats were pretreated with the antioxidant alpha-tocopherol or the iron chelator deferoxamine, the increases in microsomal glutathione S-transferase activity and lipid peroxidation caused by t-BuOOH perfusion of the isolated liver was prevented. Furthermore, the activation of microsomal GSH S-transferase by t-BuOOH in vitro was also inhibited by incubation of microsomes with alpha-tocopherol or deferoxamine. Thus it was confirmed that liver microsomal glutathione S-transferase is activated in the oxidative stress caused by t-BuOOH via thiol oxidation of the enzyme.

Animals↗
PubMed1990

Differential association of endogenous proenkephalin-derived peptides with membranes of microsomes from rat striatum, adrenal medulla and heart ventricle.

Proenkephalin-derived peptides, in common with other prohormones, are associated with membranes of microsomes and secretory granules in the bovine adrenal medulla. Post-translational processing of the precursor molecule varies depending upon the tissue. The relationship between post-translational events in different tissues was examined by studying the membrane association of endogenous proenkephalin-derived peptides in the crude microsomal fraction of rat adrenal medulla, brain striatum and heart ventricle. [Met]-Enkephalin and synenkephalin (proenkephalin(1-70)) immunoreactivities were quantified by radioimmunoassay after sequential enzymatic digestion with trypsin and carboxypeptidase B. Between 60 and 75% of total immunoreactive peptides present in intact microsomes of the three tissues were associated with membranes and specifically released with 2 M KSCN (pH 7.4). Analysis of the chromatographic profile of materials present in the soluble and associated fractions produced the following results. In the three tissues the materials associated with microsomal membranes corresponded to peptides larger than 3-5 kDa and displayed synenkephalin and [Met]-enkephalin immunoreactivity. Adrenal and heart microsomes showed a continuous pattern of membrane-associated proenkephalin-derived peptides of high, intermediate and low molecular weights containing the synenkephalin and [Met]-enkephalin sequences. These tissues, however, presented quantitative differences, as the highest concentrations belonged to materials larger and smaller than 12.5 kDa in adrenal and heart microsomes respectively. On the other hand, brain striatal microsomes displayed a discontinuous pattern of associated materials, with the absence of some products of high and intermediate molecular weight. Only in the soluble fraction of striatal microsomes were peptides detected of high and intermediate molecular weight containing the [Met]-enkephalin but not the synenkephalin sequence.(ABSTRACT TRUNCATED AT 250 WORDS)

Adrenal Medulla↗
PubMed1981

Activation of benzo(a)pyrene and 2-acetamidofluorene to mutagens by microsomal preparations from different animal species: role of cytochrome P-450 and P-448.

1. The metabolic activation of benzo(a)pyrene and 2-acetamidofluorene to mutagens was studied with liver microsomal preparations from rat, guinea-pig, hamster and mouse, untreated or pretreated with phenobarbitone or 3-methylcholanthrene. 2. Liver microsomal preparations from all animal species activated benzo(a)pyrene, that from mouse being the most efficient. Similarly, microsomal preparations from guinea-pig, hamster and mouse could activate 2-acetamidofluorene, but that from rat exhibited very weak activity. 3. Activation of benzo(a)pyrene into mutagenic intermediates by liver microsomal preparations was increased for all animals except mouse by pretreatment with 3-methylcholanthrene. In contrast, pretreatment with phenobarbitone decreased the activation by microsomal preparations from all species. 4. Activation of 2-acetamidofluorene by liver microsomal preparations from rat and guinea-pig, but not mouse and hamster, was increased by pretreatment of the animals with phenobarbitone. Pretreatment with 3-methylcholanthrene decreased the activation of this carcinogen by microsomal preparations from all species. 5. The metabolic activation of benzo(a)pyrene is catalysed by cytochrome P-448 but not cytochrome P-450. 6. The activation of 2-acetamidofluorene to mutagens may involve, in addition to the mixed-function oxidases, other microsomal enzyme systems.

2-Acetylaminofluorene↗
PubMed1987

Interaction of miconazole, ketoconazole and itraconazole with rat-liver microsomes.

The interaction of the antimycotics miconazole, ketoconazole and itraconazole with liver microsomes from untreated rats or from rats pretreated with phenobarbital or 3-methylcholanthrene, gave rise to type II difference spectra. The interactions of the antimycotics with control, phenobarbital-induced or 3-methylcholanthrene-induced microsomes were biphasic, except for the monophasic binding of ketoconazole to phenobarbital-induced microsomes. The N-demethylation of N,N-dimethylaniline, the O-demethylation of p-nitroanisole and the hydroxylation of aniline in microsomes from untreated and inducer-treated rats were lowered by miconazole and ketoconazole, the former being the more potent inhibitor. Control microsomes were less sensitive than induced microsomes. Itraconazole was almost devoid of inhibitory properties. The three antimycotics were non-competitive (mixed) inhibitors of enzyme activities in phenobarbital-induced microsomes. The Ki values were of the same order of magnitude as the Ks values, except for itraconazole. For the latter drug, Ki values were much greater than could be expected from the spectral studies. It is concluded that the antimycotics affect microsomal enzyme activities via a direct interaction of an azole-nitrogen with the haem group of cytochrome P-450. The interaction with mammalian cytochrome P-450 decreases from miconazole greater than ketoconazole much greater than itraconazole and is much weaker than the interaction of the antimycotics with yeast cytochrome P-450.

Aniline Hydroxylase↗
PubMed1991

Comparison of the effects of inducers of cytochrome P450 on Mongolian gerbil and rat hepatic microsomal monooxygenase activities.

1. Basal cytochrome P450 content (nmol/mg protein) was higher in gerbil (1.10 +/- 0.01) than in rat (0.81 +/- 0.05) hepatic microsomes. Pretreatment of gerbils with phenobarbitone and beta-naphthoflavone increased P450 contents by 200% and 60% respectively. 2. 7-Ethoxycoumarin O-deethylase, coumarin 7-hydroxylase and 4-nitrophenol hydroxylase activities were generally higher in gerbil liver microsomes, whereas erythromycin N-demethylase, and 7-ethoxyresorufin and 7-pentoxyresorufin O-dealkylase activities were higher in rat microsomes. Microsomal benzphetamine N-demethylase activities were similar in both species. 3. Induction of specific cytochrome P450 isozymes increased similar monooxygenase activities of rat and gerbil microsomes. Phenobarbitone, beta-naphthoflavone, isoniazid and pregnenolone 16 alpha-carbonitrile principally increased benzphetamine N-demethylase, 7-ethoxyresorufin O-deethylase, 4-nitrophenol hydroxylase and erythromycin N-demethylase activities respectively. 4. Constitutive 7-ethoxyresorufin and 7-pentoxyresorufin O-dealkylase activities were markedly lower in gerbil microsomes compared with rat microsomes, and pretreatment of gerbils with cytochrome P450 inducers did not significantly increase these activities. 5. Hepatic microsomal coumarin 7-hydroxylase activities were approximately 30-200 times greater (depending on the inducer) in the gerbil than in rat. The gerbil, due to is high coumarin 7-hydroxylase activity, would appear to be a more appropriate species than rat for investigations of coumarin metabolism and toxicity relevant to humans.

Animals↗
PubMed1992

Metabolism of coumarin by rat, gerbil and human liver microsomes.

1. o-Hydroxyphenylacetaldehyde was the major metabolite of coumarin (1 mM) in rat, gerbil and human liver microsomes. 2. Treatment of rats with phenobarbitone (PB) or beta-naphthoflavone increased the o-hydroxyphenylacetaldehyde formed. 3-Hydroxycoumarin was the other main metabolite produced by rat liver microsomes. 3. Liver microsomal metabolism of coumarin in gerbil was extensive with 3-, 5-, 6-, 7- and 8-hydroxycoumarins, and 3,7- and 6,7-dihydroxycoumarins produced, in addition to o-hydroxyphenylacetaldehyde. The profile of the hydroxy metabolites was altered by in vivo treatment of gerbils with cytochrome P-450 inducers, but there was no increase of coumarin metabolism. 4. Coumarin was metabolized by human liver microsomes to o-hydroxyphenylacetaldehyde, 7-hydroxycoumarin, 3-hydroxycoumarin, and trace amounts of 5-, 6- and 8-hydroxycoumarins. 5. At low substrate concentrations (0-10 microM) hepatic microsomal metabolism of coumarin in gerbil resembled that in man, with 7-hydroxycoumarin being a major metabolite. However, the production of o-hydroxyphenylacetaldehyde was greater in gerbil than human liver microsomes. 6. At higher substrate concentrations (1 mM) metabolism of coumarin by liver microsomes from PB-treated gerbils most closely resembled that by human liver microsomes. 7. The gerbil would appear to be a more appropriate animal model than rat for studies to assess the toxicological hazard of coumarin for man.

Adolescent↗
PubMed1993

Comparative sulphoxidation of albendazole by sheep and cattle liver microsomes and the inhibitory effect of methimazole.

1. The comparative rates of oxidation of the benzimidazole anthelmintic, albendazole (ABZ), by sheep and cattle liver microsomes, and inhibition by the antithyroid compound methimazole (MTZ) were investigated. 2. ABZ was oxidized to its sulphoxide metabolite (ABZSO) in an NADPH concentration-dependent reaction. Heat inactivation of the microsomal flavin-containing mono-oxygenase system significantly decreased the NADPH consumption of microsomes in the presence of ABZ, MTZ and thiourea. 3. Oxidation of ABZ, MTZ and thiourea by sheep liver microsomes consumed significantly more NADPH than oxidation by cattle microsomes. 4. Neither the pro-ABZ drug, netobimin, nor the ABZ sulphone metabolite (ABZSO2) was modified by incubation with either sheep or cattle liver microsomes. 5. ABZSO was oxidized into ABZSO2 at a very slow rate and only when a high microsomal protein concentration was used. 6. MTZ was a potent inhibitor of ABZ sulphoxidation and the inhibition was significantly lower in cattle than in sheep microsomes.

Albendazole↗