Antioxidant activity of Smoke Shield in-vitro and in-vivo.

Smoke Shield is a proprietory formulation containing extract of turmeric (Curcuma longa), obtained by supercritical carbon dioxide gas extraction and post-supercritical hydroethanolic extraction, together with extracts of green tea and other spices whose presence synergistically increases the activity of turmeric. This study evaluates the antioxidant potentials of Smoke Shield in-vitro and in experimental animals, as well as in human models. Smoke Shield was found to scavenge superoxide radicals generated by photoreduction of riboflavin (50% inhibitory concentration = 91 microg mL(-1)) and hydroxyl radicals generated by Fenton reaction (50% inhibitory concentration = 95 microg mL(-1)) and reduced lipid peroxidation. Administration of Smoke Shield to mice was found to elevate antioxidant enzymes such as catalase and superoxide dismutase in blood as well as in liver and kidney. Glutathione-S-transferase activity was found to be significantly elevated in liver and kidney of animals treated with Smoke Shield. Glutathione levels were also significantly elevated in blood. Glutathione reductase was significantly elevated in kidney. Administration of Smoke Shield decreased the lipid peroxidation in serum, liver and kidney, as well as reduced the levels of conjugated dienes and hydroperoxides. Administration of Smoke Shield to smokers was found to increase the superoxide dismutase and glutathione in blood and decrease glutathione peroxidase. Smoke Shield inhibited phase I enzymes as represented by aniline-hydroxylase and aminopyrenedemethylase in-vitro. These results indicate that Smoke Shield has potent antioxidant activity, could inhibit phase I enzymes and increase detoxifying enzymes, which makes it an effective chemoprotective herbal formulation.

Antioxidant activity of anti-inflammatory plant extracts.

The antioxidant properties of twenty medical herbs used in the traditional Mediterranean and Chinese medicine were studied. Extracts from Forsythia suspensa, Helichrysum italicum, Scrophularia auriculata, Inula viscosa, Coptis chinensis, Poria cocos and Scutellaria baicalensis had previously shown anti-inflammatory activity in different experimental models. Using free radical-generating systems H. italicum. I. viscosa and F. suspensa protected against enzymatic and non-enzymatic lipid peroxidation in model membranes and also showed scavenging property on the superoxide radical. All extracts were assayed at a concentration of 100 microg/ml. Most of the extracts were weak scavengers of the hydroxyl radical and C. chinensis and P. cocos exhibited the highest scavenging activity. Although S. baicalensis inhibited the lipid peroxidation in rat liver microsomes and red blood cells, the extract showed inhibitory actions on aminopyrine N-demethylase and xanthine oxidase activities as well as an pro-oxidant effect observed in the Fe3+-EDTA-H2O2 system. The results of the present work suggest that the anti-inflammatory activities of the same extracts could be explained, at least in part, by their antioxidant properties.

Inhibition of human hepatic cytochrome P450s and steroidogenic CYP17 by nonylphenol.

Effect of nonylphenol on aminopyrine N-demethylase activity, a typical drug-metabolizing enzyme activity, by ten kinds of human hepatic cytochrome P450s (CYP) and on progesterone 17alpha-hydroxylase activity by steroidogenic CYP17 was investigated. When determined at 2 mM substrate concentration, nonylphenol (1 mM) most efficiently inhibited aminopyrine N-demethylation by CYP2C9 and CYP2C19, by 61% and 59%, respectively, followed by CYP2D6, CYP1A2, CYP2C18 and CYP2C8 (46-51%), whereas inhibition of the activities by other CYPs was less than 27%. Additionally, nonylphenol competitively inhibited diclofenac 4'-hydroxylation by CYP2C9 and S-mephenytoin 4'-hydroxylation by CYP2C19 with Ki values of 5.3 and 37 microM, respectively. Furthermore, nonylphenol exhibited a competitive inhibition of progesterone 17alpha-hydroxylase activity by CYP17 with Ki value of 62 microM. These results suggest that nonylphenol inhibits human hepatic CYPs, especially CYP2C9 and CYP2C19, and steroidogenic CYP17 activities.

Role of C-5 chiral center in R-(+)-pulegone-mediated hepatotoxicity: metabolic disposition and toxicity of 5, 5-dimethyl-2-(1-Methylethylidene)-cyclohexanone in rats.

Metabolic disposition of 5, 5-dimethyl-2-(1-methylethylidene)-cyclohexanone (I) was examined in rats. Compound (I) was administered orally (250 mg/kg of body weight/day) to rats for 5 days. The following urinary metabolites were isolated and identified: 4,5,6,7-tetrahydro-3,6, 6-trimethylbenzofuran (III), 3,3-dimethylcyclohexanone (VI), 5, 5-dimethyl-3-hydroxy-2-(1-methylethylidene)-cyclohexanone (X), 5, 5-dimethyl-2-(1-hydroxymethylethyl)-cyclohexanone (IX), 3-hydroxy-5-hydroxymethyl-5-methyl-2-(1-methylethylidene)-cyclo hexano ne (XI), 5,6-dihydro-3,6,6-trimethyl-2(4H)-benzofuranone (VIII), and 5,5-dimethyl-3-hydroxy-2-(1-carboxy ethylidene)-cyclohexanone (XIII). Incubation of compound (I) with phenobarbital (PB)-induced rat liver microsomes in the presence of NADPH resulted in the formation of a metabolite, tentatively identified as a furanoterpene (III) based on proton magnetic resonance, gas chromatography, and gas chromatography-mass spectroscopy analyses. The formation of III was inhibited to a significant extent by carbon monoxide, metyrapone, SKF 525-A, and cytochrome c, suggesting the participation of PB-induced microsomal cytochrome P-450 system in the conversion of I to III. Compound I gave type I spectral change in the PB-induced liver microsomes and the dissociation constant (Ks) for I was 38.5 microM. Intraperitoneal administration of a single dose (250 mg/kg) of I to rats resulted in 26, 23, and 41% decreases in the levels of cytochrome P-450, glucose-6-phosphatase, and aminopyrine N-demethylase, respectively, at the end of 24 h. During this period, a 11-fold increase in serum glutamate pyruvate transaminase level was also observed. However, a decrease in the level of cytochrome P-450 and glucose-6-phosphatase, and an increase in serum glutamate pyruvate transaminase values were comparatively more pronounced when R-(+)-pulegone (250 mg/kg) or CCl(4) (0.6 ml/kg) was administered to rats. Pretreatment of rats with PB potentiated the hepatotoxicity caused by I, whereas pretreatment with 3-methylcholanthrene protected from it. This suggests that PB-induced cytochrome P-450-catalyzed reactive metabolites may be responsible for the toxic effects caused by I.

Effect of vitamin E on lipid peroxidation and liver monooxigenase activity in experimental influenza virus infection.

Influenza virus infection was associated with development of oxidative stress in liver of mice, viz. increase in amount of lipid peroxidation products, decrease in cytochrome P-450 and NADP. H-cytochrome c-reductase activity, and inhibition of liver monooxygenases (aniline hydroxylase, ethylmorphine-N-demethylase, amidopyrine-N-demethylase and analgin-N-demethylase). These effects were most pronounced on the 7th day after virus inoculation as compared to the 5th one. Supplementation of mice with vitamin E before virus inoculation leads to liver protection against oxidative stress and toxicosis. A marked decrease of lipid peroxidation products and an increase of cytochrome P-450 and activities of monooxygenases was established. The stabilizing effect of vitamin E was dose-dependent and was most pronounced on the 5th day after virus inoculation as compared to the 7th one.

Triphenyltin acetate-mediated in vitro inactivation of rat liver cytochrome P-450.

The in vitro effects of the organotin (OT) compound triphenyltin acetate (TPTA) on cytochrome P-450 content and functions were investigated in liver microsomes from untreated, phenobarbital (PB)- or beta-naphthoflavone- (betaNAF) pretreated rats. At a concentration of 0.5 mM, TPTA caused a marked loss in the spectrally detectable content of cytochrome P-450 up to 27% of its original value, along with an increase in the inactive form cytochrome P-420. Both effects were most pronounced in betaNAF-treated microsomes, which showed a shift in the hemoprotein absorption maximum from 448 nm to 451 nm, but in all cases TPTA failed to affect either cytochrome b5 or total heme content, or to increase the production of malondialdehyde. These results suggest that lipid peroxidation of microsomal membranes or damage to the heme moiety should be excluded as contributing factors in the hemoprotein loss. TPTA also produced a concentration-related functional inactivation of cytochrome P-450 that was most pronounced in betaNAF-exposed microsomal preparations, as denoted by a striking reduction in the ethoxyresorufin O-deethylase (EROD) activity (IC50 = 0.088 mM). In contrast, the activities of cytochrome P-450-independent microsomal enzymes such as NADPH cytochrome c reductase and indophenyl acetate esterase (IPA-EST) were not markedly affected even by 0.5 mM TPTA (-30%). As assessed by Lineweaver-Burk plots, the mechanism of inhibition appeared to be noncompetitive for IPA-EST and of mixed type (competitive-noncompetitive) for EROD. Among sulfhydryl-containing compounds, dithiothreitol was considerably more effective than albumin and reduced glutathione in preventing cytochrome P-450 inactivation and even was able to partially reverse the hemoprotein damage when added after TPTA; glycerol, which is known to protect the hydrophobic environment of cytochrome P-450, was as effective as albumin. This study indicates that TPTA behaves as an almost specific and powerful in vitro inhibitor of cytochrome P-450-dependent monooxygenases, apparently through the interaction with critical sulfhydryl groups of the hemoprotein.

Antimutagenic and anticarcinogenic potentials of some Thai vegetables.

Fifteen kinds of commonly consumed Thai vegetables were sequentially extracted with hexane, chloroform and methanol, and then tested for antimutagenic activities against direct-acting (AF-2 and NaN3) and indirect-acting (AFB1 and B(a)P) mutagens using Ames' Salmonella mutagenicity test with Salmonella typhimurium TA100 as tester strain. It was found that only the methanol extract of neem leaves contain weak antimutagen inhibiting the mutagenicities of both direct-acting mutagens. Interestingly, all vegetables studied were found to contain chemical compounds, mainly nonpolar ones, capable of inhibiting the mutagenicity of AFB1, while only some vegetables contain chemical compounds capable of inhibiting the mutagenicity of B(a)P, which is also an indirect-acting mutagen. Studies on anticarcinogenic potentials demonstrated that Thai bitter gourd fruits, but not sweet basil leaves, at the concentration of 6.25% and 12.5% in the diet, partially inhibited DMBA-induced mammary gland carcinogenesis in female Sprague-Dawley rats when fed to the animals 2 weeks prior to DMBA. Results in the present study therefore demonstrated that most Thai vegetables contain antimutagens inhibiting the mutagenicity of some indirect-acting mutagen, particularly AFB1. The mechanism of their antimutagenicity may probably be the inhibition of the activity of metabolic-activating enzymes in rat liver homogenates. Very interestingly, our results clearly reveal that Thai bitter gourd fruits, which possess Phase II enzymes inducing property, as well as the ability to reduce Phase I enzyme activities in rat liver, contain some anticarcinogens or chemopreventive agents. However, sweet basil leaves that possess both Phase I and Phase II enzyme-inducing properties may not contain any anticarcinogen, at least against DMBA-induced mammary gland carcinogenesis.

Zinc ethylene-bis-dithiocarbamate (Zineb)-mediated inhibition of monooxygenases and lipid peroxidation in bovine liver microsomes.

Hepatic bovine microsomes were incubated with Zineb concentrations ranging from 2.5 mM to 2.5 microM. Only the higher concentrations of the fungicide (2.5 and 0.25 mM) elicited a sharp decline in cytochrome P450, cytochrome b5 and total sulphydryl groups content as well as in the activities of NADPH cytochrome c reductase, aminopyrine N-demethylase and aniline 4-hydroxylase. The loss of cytochrome P450 was matched by a concomitant increase in the amount of cytochrome P420, which represents a catalytically inactive form of cytochrome P450. The same concentrations of the fungicide, either alone or in the presence of NADPH 1 mM, failed to increase the amount of thiobarbituric reactive substances with respect to control incubations, thereby excluding the possibility of lipid peroxidation as a contributing factor in the loss of cytochrome P450 and in the inhibition of cytochrome P450-mediated metabolism. It is concluded that Zineb can depress monooxygenase activity in bovine hepatic microsomes mainly through the denaturation of cytochrome P450 and the impaired transfer of reducing equivalents to the complex cytochrome P450-substrate. These mechanisms might also account for the inhibition in lipid peroxidation brought about by the fungicide.

Role of 3,4-dichlorophenyl methyl sulfone, a metabolite of o-dichlorobenzene, in the changes in hepatic microsomal drug-metabolizing enzymes caused by o-dichlorobenzene administration in rats.

2,3- and 3,4-Dichlorophenyl methyl sulfoxides and 2,3- and 3,4-dichlorophenyl methyl sulfones (2,3- and 3,4-DCPSO2Mes) were detected in the urine of rats administered o-dichlorobenzene (o-DCB). After administration of o-DCB to rats, swift decreases were observed in the concentrations of o-DCB in blood, liver, and kidneys, whereas 3,4-DCPSO2Me appeared in blood, liver, kidneys, and adipose tissue. The concentrations of 3,4-DCPSO2Me in the blood and three tissues reached maxima at 24 hr. Both aminopyrine N-demethylase and aniline hydroxylase activities and cytochrome P450 content of hepatic microsomes decreased 24 hr after administration of o-DCB. In contrast, 3,4-DCPSO2Me increased the activities of these enzymes and cytochrome P450 and b5 contents in rat liver microsomes. In both antibiotic-pretreated and bile duct-cannulated rats dosed with o-DCB, the concentrations of 2,3- and 3,4-DCPSO2Mes in blood, liver, kidneys, and adipose tissue were dramatically reduced. These findings suggest that the process of formation of methylsulfonyl metabolites of o-DCB involves biliary secretion of DCPSO2Mes and/or their precursors which will be subjected to metabolism by intestinal microflora. In antibiotic-pretreated rats, the inhibitory effects of o-DCB administration on the activities of aminopyrine- and aniline-metabolizing enzymes and the contents of cytochromes P450 and b5 in hepatic microsomes were greater than those observed in the intact rats. In bile duct-cannulated rats, the decrease in aminopyrine N-demethylase activity after administration of o-DCB was greater than that observed in the intact rats. These findings suggest that the apparent inhibition of drug-metabolizing enzymes by o-DCB is the result of simultaneous contrary effects, namely, the inductive effect of 3,4-DCPSO2Me and the stronger inhibitory effect of an unknown factor(s) on drug-metabolizing enzymes.

A protein inhibitor of hepatic drug metabolizing enzymes from Ehrlich ascites cells.

Hepatic drug metabolizing enzymes were significantly decreased in Ehrlich ascites tumour-bearing mice. A protein inhibitor of hepatic drug metabolizing enzymes was isolated from Ehrlich ascites cells and purified. This involved ammonium sulphate fractionation (60-80%), DEAE, phosphocellulose, Sephadex G-100 and hydroxyapatite column chromatography. Purification attained was 800-fold. The inhibitory protein was effective in decreasing all the components of hepatic mixed-function-oxidase system and drug metabolizing enzymes both in vivo and in vitro. This novel inhibitor may have potential applications in chemical carcinogenesis.

Induction of hepatic microsomal P450 by 4-phenylalkylpyridines in rat: chain length-dependent and sex-related differential induction of P450s.

The effects of 4-phenylalkylpyridines (chain length of 0-5, 7, 9 and 11 carbon atoms) on the induction of hepatic microsomal P450 and its multiple forms (1A1/2 (1A1, 1A2), 3A2, 2B1/2, 2C6 and 2E1) in the male and female rat have been investigated. P450 induction gradually declined with increasing chain length of the 4-phenylalkylpyridines. Immunoblot analysis revealed that three pyridine compounds having methylene units of 0, 1 and 2 only induced P4501A2, whereas those having 4, 5 or 7 methylene units only induced 1A1 in the male rat. In the female rat, however, we could not observe such a chain length-dependent differential induction of the P4501A subfamily. Induced levels of P4503A2 and 2E1 were dependent on total P450 specific content, but 2C6 was increased in a chain length-dependent manner in both sexes. These results provide new information on the differential effects of pyridine-containing compounds on P450 induction in the rat.

Brain mitochondrial cytochromes P450: xenobiotic metabolism, presence of multiple forms and their selective inducibility.

The capability of rat brain mitochondria to metabolize a variety of xenobiotics was examined. The presence of cytochrome P450 (P450) and associated monooxygenase activities were estimated in isolated rat brain mitochondria and compared with the corresponding activities in microsomes. Total P450 content in brain mitochondria from naive rats was twice that of the corresponding microsomal level. The ability of brain mitochondria to metabolize the potent carcinogen N-nitrosodimethylamine was more than twofold that of the corresponding microsomal activity, while the 7-ethoxycoumarin-O-deethylase activity was significantly lower in mitochondria. Immunoblot experiments using antisera to purified rat liver microsomal P450s, namely P450 (2B1/2B2), P4501A1, and P4502E1, and purified phenobarbital-inducible rat brain P450, revealed the presence of immunoreactive bands in isolated brain mitochondria. These various antibodies to P450 inhibited the brain mitochondrial monooxygenase activities to significant, though varying extent. The addition of antiserum to microsomal NADPH cytochrome P450 reductase did not affect the mitochondrial P450 associated monooxygenase activities, although it completely inhibited the corresponding microsomal activities. Chronic ethanol administration resulted in twofold induction of total P450 content and the monooxygenase activities known to be mediated by P4502E1, such as N-nitrosodimethylamine-N-demethylase and p-nitrophenol hydroxylase in brain mitochondria. Pretreatment of animals with phenobarbital resulted in the induction of aminopyrine N-demethylase activity in brain mitochondria. The study demonstrates the presence of multiple forms of P450 in the rat brain mitochondria, their inducibility, and their capability to metabolize xenobiotics.

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 (Ki = 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 (Ki = 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 Ki concentration, regardless of the apparent kinetic inhibition type. The combined inhibition was less than fully additive when the individual concentrations were twice the Ki or above. These results demonstrate that, when two inhibitors of oxidative drug metabolism are combined, both the Ki values and the concentrations of inhibitors play important roles in determining the extent of additive inhibition of enzyme activity.

Inhibition of microsomal lipid peroxidation and monooxygenase activities by eugenol.

Previously we reported that eugenol (4-allyl-2-methoxyphenol) inhibits non-enzymatic peroxidation in liver mitochondria (E. Nagababu and N. Lakshmaiah, 1992, Biochemical Pharmacology. 43, 2393-2400). In the present study, we examined the effect of eugenol on microsomal mixed function oxidase mediated peroxidation using Fe+3-ADP-NADPH, carbon tetrachloride (CCL4)-NADPH and cumene hydroperoxide (CumOOH) systems. In the presence of eugenol the formation of thiobarbituric acid reactive substances (TBARS) was decreased in all the systems (IC50 values: 14 microM for Fe+3-ADP-NADPH, 4.0 microM for CCl4-NADPH and 15 microM for CumOOH). Oxygen uptake was also inhibited to a similar extent with Fe+3-ADP-NADPH and CumOOH systems. A comparative evaluation with other antioxidants showed that in Fe+3-ADP-NADPH and CumOOH systems, the antioxidant efficacy was in the order: butylated hydroxytoluene (BHT) > eugenol > alpha-tocopherol, while in CCl4-NADPH system the order was alpha-tocopherol > BHT > eugenol. Time course of inhibition by eugenol indicated interference in initiation as well as propagation of peroxidation. Eugenol did not inhibit cytochrome P-450 reductase activity but it inhibited P-450 - linked monooxygenase activities such as aminopyrine-N-demethylase, N-nitrosodimethylamine demethylase, benzo(a)pyrene hydroxylase and ethoxyresorufin-O-deethylase to different extents. However, CumOOH supported monooxygenases (aminopyrine-N-demethylase and benzo(a)pyrene hydroxylase) required much higher concentrations of eugenol for inhibition. The concentration of eugenol required to inhibit monooxygenase activities was more than that required to inhibit peroxidation in all the systems. Eugenol elicited type 1 changes in the spectrum of microsomal cytochrome P-450. These results suggest that the inhibitory effect of eugenol on lipid peroxidation is predominantly due to its free radical quenching ability. Eugenol significantly protected against the degradation of cytochrome P-450 during lipid peroxidation with all the systems tested. These findings suggest that eugenol has the potential to be used as a therapeutic antioxidant. Further evaluation may throw more light on this aspect.

Antimutagenic and mutagenic potentials of Chinese radish.

The edible part of fresh Chinese radish was chopped into small pieces, lyophilized, and then extracted sequentially with hexane, chloroform, and methanol. The solvent in each fraction was removed by evaporation under reduced pressure at 50-55 degrees C, and the residue was dissolved in dimethylsufoxide just before being tested for antimutagenicity as well as mutagenicity using the Salmonella/mammalian microsome mutagenicity test. We found that none of the three fractions exhibited any mutagenicity toward S. typhimurium strains TA98 and TA100 when tested either in the presence or absence of S-9 mix. Interestingly, however, hexane and chloroform extracts could strongly inhibit the mutagenicities of both direct mutagens (e.g., 2-(2-furyl)-3-(5-nitro-2-furyl)acrylamide and sodium azide) and indirect mutagens (e.g., aflatoxin B1). In contrast, however, these two fractions did not inhibit the mutagenicity of benzo[a]pyrene, which is also an indirect mutagen. Both hexane and chloroform extracts could also markedly inhibit the activities of rat liver aniline hydroxylase and aminopyrine demethylase. The methanol fraction could inhibit neither the mutagenicities of direct or indirect mutagens tested nor the activities of those two rat liver enzymes. Results of the present study demonstrate that Chinese radish may not contain any mutagenic compound but does contain some nonpolar compounds with antimutagenic activity toward both direct and indirect mutagens. In addition, the antimutagenic activity toward aflatoxin B1 may be partly due to the inhibition of enzymes necessary for activation of this mutagen.

Inhibition of hepatic microsomal drug metabolism by atracurium administration in the rat.

The muscle relaxant atracurium is known to undergo extrahepatic degradation via Hofmann elimination and ester hydrolysis. The purpose of the present study was to evaluate the effects of atracurium on hepatic P450-dependent enzyme activities. Thirty-two male Sprague-Dawley rats were anaesthetized, mechanically ventilated, and randomly allocated to one of four study groups: group 1 received saline, group 2 atracurium, group 3 vecuronium, and group 4 pancuronium intravenously for a period of 3 hr. Equipotent doses of the muscle relaxants were applied; the doses had been obtained in a pilot study using evoked electromyography. At the end of the study period, the livers were removed and analyzed. All three muscle relaxants may lead to inhibition of hepatic drug metabolism. Atracurium influences hepatic P450, although it is predominantly degraded in extrahepatic tissues. Further studies are needed to evaluate the contribution of the major metabolite laudanosine to this inhibitory action.

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

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

Inhibition of dexamethasone-induced cytochrome P450-mediated mutagenicity and metabolism of aflatoxin B1 by Chinese medicinal herbs.

Oldenlandia diffusa (OD) and Scutellaria barbata (SB) have been used in traditional Chinese medicine for treating liver, lung and rectal tumours. We previously showed that they inhibited mutagenesis, DNA binding and metabolism of aflatoxin B1 (AFB1) and benzo(a)pyrene (BaP) bioactivated by Aroclor 1254-induced rat S9. The purpose of this study was to investigate the effects of OD and SB on the mutagenicity of AFB1 in Salmonella typhimurium TA100 using dexamethasone (DXM)-induced rat hepatic S9, on cytochrome P450-linked aminopyrine N-demethylase (APND) activity in DXM-induced hepatic microsomes and on the metabolism of AFB1 by DXM-induced S9 using high-performance liquid chromatography (HPLC). The experimental results showed that OD and SB consistently inhibited the mutagenicity of AFB1 bioactivated by either non-induced or DXM-induced S9. These effects correlated with the inhibition of cytochrome P450-linked APND activity in DXM-induced microsomes and with an inhibition of DXM-induced S9 mediated metabolism of [3H]AFB1 as determined by HPLC. Since DXM treatment has been associated with an induction of the CYP3 enzyme family, these results suggest that OD and SB may possess antimutagenic and antitumorigenic activity towards AFB1 through an inhibition of CYP3-mediated metabolism of AFB1.

General anesthetics inhibit cytochrome P450 monooxygenases and arachidonic acid metabolism.

Identification of a specific biomolecular target appropriately sensitive to a wide array of anesthetics has been elusive. At concentrations close to their respective ED50's for anesthesia in man or other species, 18 compounds, differing in potencies up to 66,000 fold, inhibited cytochrome P450 mediated metabolism of aminopyrine, a synthetic substrate, and arachidonic acid (AA), an endogenous substrate, in isolated liver microsomes. There was a highly significant correlation for both substrates between the absolute concentrations required for anesthesia (EC50) and for inhibition of P450 activity (Ki or IC50). The mean Ki/EC50 ratio was 0.97 for inhibition of aminopyrine demethylase. The mean IC50/EC50 ratios were 0.42 and 0.64 for inhibition of two AA-derived products and 2.8 for a third; a mean ratio of 1.4 for inhibition of overall AA metabolism suggests interaction of general anesthetics with a composite of P450 isozymes. The universal cytochrome P450 monooxygenases, in conjunction with other lipid oxygenases (cyclooxygenases and lipoxygenases) participate in the second messenger AA cascade. In nerve cells the sensitivity of these enzymes to hydrophobic neurodepressant drugs may underlie the state of general anesthesia: reversible disruption of intracellular and intercellular signalling without impairment of enzymes vital to cell respiration.

Effects of interleukin 1 alpha on the activities and gene expressions of the cytochrome P450IID subfamily.

The mechanism by which recombinant human interleukin 1 alpha (rhIL-1 alpha) inhibits the activities of drug-metabolizing enzymes of rat liver microsomes, especially debrisoquine monooxygenase and bufuralol monooxygenase (both cytochrome P450IID supported reactions), as well as other enzymes, was investigated by injecting IL-1 alpha into rats. rhIL-1 alpha suppressed the activities of various P450-linked monooxygenase systems such as aminopyrine N-demethylase, benzphetamine N-demethylase, and 7-ethoxycoumarin O-deethylase. It also suppressed the activities of debrisoquine monooxygenase and bufuralol monooxygenase. On the other hand, IL-1 alpha had little effect on the activity of p-nitroanisole N-demethylase. The suppression of debrisoquine monooxygenase and bufuralol monooxygenase activities was caused by a decrease in the amounts of immunoreactive P450IID protein and its mRNA. The reduction rates in the level of immunoreactive P450IID protein and its mRNA were comparable. These results suggest that at the mRNA level, the enzymatic activities of debrisoquine monooxygenase and bufuralol monooxygenase are down-regulated by IL-1 alpha.