Effect of resveratrol analogue, DMU-212, on antioxidant status and apoptosis-related genes in rat model of hepatocarcinogenesis.

The aim of the study was to examine whether antioxidant properties of 3,4,4',5-tetramethoxystilbene (DMU-212) contribute to its anticarcinogenic activity and whether DMU-212 affects the expression of apoptosis-related genes. Two-stage model of hepatocarcinogenesis was used; male Wistar rats were challenged with N-nitrosodiethylamine (NDEA), 200 mg/kg body weight (b.w.), intraperitoneal, then phenobarbital (PB) in drinking water (0.05%) was administered. Simultaneously, DMU-212 was given per os at a dose 20 or 50 mg/kg b.w. two times a week for 16 weeks. DMU-212 caused a moderate decrease in hepatic thiobarbituric acid reactive substances and protein carbonyls concentration elevated in rats treated with NDEA/PB. The activity of antioxidant enzymes examined reduced by NDEA/PB treatment was not restored in rats coadministered with DMU-212. Effects of DMU-212 on messenger RNA (mRNA) expression of antioxidant enzymes in rats challenged with NDEA/PB were diversified; no changes in their protein expression were noted in any of the groups. The expression of 17,000 genes was analyzed by Affymetrix® Rat Gene 1.1 ST Array; 15 apoptosis-related genes were selected and validated by RT-q PCR. The combined treatment with NDEA/PB and DMU-212 increased the mRNA level of some genes driving mitochondria-mediated apoptosis, whereas the mRNA expression of some anti-apoptotic genes triggering receptor-mediated apoptosis was reduced. The expression of genes encoding caspases-4, -8, -9, and -12 was also increased in rats treated with DMU-212. Although antioxidant effect of DMU-212 in rats challenged with NDEA/PB was moderate, its potential anticarcinogenic properties were demonstrated as evidenced by modulation of apoptosis-related genes.

Hepatoprotective effect of the extract and isocytisoside from Aquilegia vulgaris.

The hepatoprotective effect of the ethanol extract (AvEE) and the main fl avonoid compound 4'-methoxy-5,7-dihydroxy fl avone 6-C-beta-glucopyranoside (isocytisoside, ISOC) from the leaves and stems of Aquilegia vulgaris L. were studied using the CCl(4)-induced hepatotoxicity test. The acute toxicity test in mice showed that AvEE can be classi fi ed as nontoxic since a dose of 3000 mg/ kg did not cause mortality. The barbiturate-induced sleeping time prolonged by CCl(4) administration to mice was signi fi cantly reduced after AvEE treatment proving the protective effect of the extract on microsomal drug-metabolizing enzymes.AvEE and ISOC administered to rats 48 h, 24 h and 2 h before, and 6 h after CCl(4) intoxication caused a signi fi cant decrease in the CCl(4)-induced elevation of hepatic enzymes activity in serum, i.e. sorbitol dehydrogenase (SDH), glutamate oxaloacetate and glutamate pyruvate transaminases (GOT, GPT). Both substances induced CCl(4)-diminished erythrocyte superoxide dismutase (SOD) and reduced the activities of glutathione peroxidase (GPx) and glutathione reductase (GR) preliminarily enhanced by CCl(4). The hepatoprotective properties of AvEE and ISOC were con fi rmed by pathomorphological examination of the liver.

Modulation of antioxidant defence system by dietary fat in rats intoxicated with o-toluidine.

o-Toluidine was administered to rats in the diet for four weeks at levels approximately 40, 80 and 160 mg/kg b.w. per day. Two types of diet have been used, standard (4% fat) and high fat (14% fat). Activity of antioxidant enzymes, level of glutathione and thiobarbituric acid reactive substances were measured in liver. Glutathione peroxidase was significantly increased in all treated groups while glutathione S-transferase and glutathione reductase were elevated in rats fed high-fat diet. o-Toluidine slightly enhanced catalase activity regardless of the kind of diet. Superoxide dismutase was the only enzyme whose activity was lowered in almost all treated groups. Enzymatic and nonenzymatic microsomal lipid peroxidation was enhanced 2- to 3-fold in both diet groups. Reduced glutathione level in liver was 2.3- to 4.0-fold increased in all treated groups. Our findings indicate that free radical processes can be involved in the toxic effects of o-toluidine and dietary fat can modify the response of some antioxidant enzymes to this compound.

Effect of sesquiterpene lactones on antioxidant enzymes and some drug-metabolizing enzymes in rat liver and kidney.

Previously we have reported that several sesquiterpene lactones isolated from Helenium aromaticum and Telekia speciosa showed pro-oxidative properties and caused glutathione level depletion in rat liver in vivo. In the present study we examined the in vivo effect of these lactones on antioxidant enzyme systems and some drug metabolizing enzymes in the liver and the kidney of rats. We found that the majority of the compounds increased the hepatic activity of glutathione peroxidase (GPx), glutathione reductase (GR), and catalase (CAT), but superoxide dismutase (SOD) activity was distinctly lowered by five lactones. A few of the compounds tested caused a decrease in the hepatic cytochrome P450 content and reduced the activity of NADPH-cytochrome P450 reductase, aminopyrine demethylase, aniline hydroxylase and glutathione-S-transferase. Results for the kidney showed fewer changes in activities of both classes of enzymes when compared to the liver. Not all lactones affected the enzymes under test, the most active were: linifolin, helenalin, mexicanin 1 and telekin. 6 alpha-Hydroxy-2,3-dihydroaromaticin behaved differently towards monooxygenases since it induced the activity of aminopyrine demethylase and aniline hydroxylase.

Gas chromatographic method for the determination of toluidines in spiked urine samples.

A capillary gas-chromatographic method was developed for the analysis of a mixture of toluidines in urine. The method is based on the extraction of toluidines with toluene and derivatisation with heptafluorobutyric anhydride to form a product for electron capture detection. The procedure gave a linear response at concentrations of 0.02-0.20 microg/ml with sufficient reproducibility. The method is simple, requires little sample pretreatment and is being considered for biomonitoring workers exposed to toluidines.

Effect of several sesquiterpene lactones on lipid peroxidation and glutathione level.

Seven sesquiterpene lactones isolated from Helenium aromaticum: helenalin, mexicanin I, linifolin A, geigerinin, and from Telekia speciosa: 6 alpha-hydroxy-2,3-dihydroaromaticin, asperilin, telekin have been tested for their hydroxyl radical scavenging activity and effect on lipid peroxidation. All compounds were found to be potent hydroxyl radical scavengers but did not affect lipid peroxidation in vitro. In vivo they exerted pro-oxidative properties and caused glutathione level depletion and elevation in glutathione peroxidase activity.

Effect of toluidines and dinitrotoluenes on caffeine metabolic ratio in rat.

Caffeine (1,3,7-trimethylxanthine, CA) is metabolised by N-demethylation to three primary metabolites: theophylline (TP), paraxanthine (PX) and theobromine (TB). This process is mediated in 95% by CYP1A2. Thus the measurement of CA demethylated metabolites can be used as a marker of CYP1A2 activity in vivo. In the present study, caffeine and its primary metabolites were determined simultaneously in plasma of rats pretreated with three isomers of toluidine at doses: 1, 10, 60 mg/kg b.w., p.o. and four isomers of dinitrotoluene (DNT) at doses: 100 and 200 mg/kg b.w., p.o. Caffeine metabolite ratios in plasma: TB/CA, PX/CA, TP/CA, TB + PX + TP/CA were calculated and compared to those of control rats. Administration of toluidines resulted in a 2-20 fold increase of the concentration ratios of metabolites to caffeine. All toluidines seem to be inducers of CYP1A2. To the best of our knowledge this is the first information concerning the effect of toluidines on caffeine metabolism. Two out of the four tested dinitrotoluenes slightly affected CYP1A2 activity; 2,3- and 3,4-DNT increased estimated parameters 2-6 fold. Two others, 2,4- and 2,6-DNT can be considered as moderate hepatotoxic agents decreasing CA metabolic ratios to 4-70% of the control values.

Metabolism of 1-(4-hydroxy-3-methoxyphenyl)-2-propanone, a smoke flavour ketone, in rat.

1. Metabolites of 1-(4-hydroxy-3-methoxyphenyl)-2-propanone (HMP-one), a smoke flavour compound, were isolated from rat urine using hydrolysis, ether extraction, t.l.c. and g.l.c. 2. Three metabolites were identified by mass spectrometry and independent synthesis, namely: 1-(3, 4-dihydroxyphenyl)-2-propanone (Met I), 1-(3, 4-dihydroxyphenyl)-2-propanol (Met II), and 1-(4-hydroxy-3-methoxyphenyl)-2-propanol (Met III). 3. A g.l.c. method for the quantitative determination of the parent compound and metabolites in urine was devised. Unchanged HMP-one accounted for about 74% dose, with Met I 11%, Met II 5%, and Met III 9%. All compounds were excreted both as sulphate and glucuronide conjugates.

[Emission of volatile components in carpenter's kumylotox into air]. / Emisja lotnych skladników Kumylotoxu stolarskiego do powietrza.

Study was made of Carpenter's Kumylotox, a fungicidal preparation containing: p-cumylphenol, dibutyl phthalate, machine oil, chloroparaffin, a 15% solution of ker-1500 rubber in painter's naphta, and petrol for pastas. The preparation was applied onto boards placed in an experimental chamber at 1-week intervals. In air of the chamber, dibutyl phthalate and p-cumylphenol were determined quantitatively by gas chromatography. The presence of hydrocarbons was recorded by the same method, without quantitative determination. Analyses were continued until the disappearance of the investigated from air. It was found that already after 2 weeks the p-cumylphenol level dropped below the allowable concentration amounting to 0.015 mg/dm3. The dibutyl phthalate level decreased to the allowable concentration (0.05 mg/m3) only after 9 weeks of board ageing. According to analysis by the GC-MS method, aromatic hydrocarbons disappeared from the chamber's air already after 5 weeks, and the remaining hydrocarbons--after 9 weeks.

Metabolism of 5-methyl-2-furaldehyde in rat. III. Identification and determination of 5-methyl-2-furylmethylketone.

1. Continuous extraction, column chromatography and t.l.c. were employed to isolate a minor metabolite of 5-methyl-2-furaldehyde from rat urine. 2. The metabolite was identified by mass spectrometry and independent synthesis as 5-methyl-2-furylmethylketone. 3. A method for quantitative determination of the metabolite in urine was devised. About 7% of the parent compound was metabolized to 5-methyl-2-furylmethylketone.

Studies on kinetics of anturan excretion in man.

The kinetics of anturan excretion was studied. Experiments were carried out on ten healthy volunteers. The drug was given orally once applying three different doses: 100, 200, 400 mg. The contents of the drug in urine were determined by means of the modified method worked out by Wallace. It was found about 42 per cent of the dose was excreted with urine in an unchanged form. The process of anturan excretion may be described according to the one-compartment open kinetic model. The half-life of excretion is 3.5 hours, the excretion constant is 0.20. The formula showing the course of anturan excretion in time has been given. Five examples of the quantitative exposure test have been proposed; they allow the calculation of the absorbed drug dose and thus the degree of poisoning. The test can be also helpful in controlled therapy.