Biomedical subjects

V Ravindranath

Publications and source records attributed to V Ravindranath.

66 records · Page 4Linked to original sources

PubMed1990

Preparation of brain microsomes with cytochrome P450 activity using calcium aggregation method.

Microsomes have been conventionally prepared by centrifugation of the postmitochondrial supernatant at 100,000g using an ultracentrifuge. Liver microsomes have been prepared by low speed centrifugation following sedimentation of the microsomal membranes in the presence of calcium ions. However, this method has not been suitable for the preparation of microsomes from extrahepatic tissues as it often results in the loss of cytochrome P450 activity. Brain microsomes prepared by the traditional calcium aggregation method results in the loss of cytochrome P450. We now describe a modification of the calcium aggregation method for the rapid preparation of rat and mouse brain microsomes. This involves the incorporation of glycerol, dithiothreitol, and EDTA in the preparation of microsomes. Such preparations do not differ in their cytochrome P450 content and associated monooxygenase activity from the traditionally prepared microsomes using ultracentrifugation. Electron microscopic analysis also does not reveal any differences between the microsomes prepared by the two methods. As brain microsomes are relatively unstable and are obtained in low yields, rapid isolation of large quantities of microsomes, possible using the present method, should be very useful.

Animals↗

PubMed1989

Xenobiotic metabolism in human brain--presence of cytochrome P-450 and associated mono-oxygenases.

The cytochromes P-450, a family of heme proteins, play an important role in the oxidation of drugs and carcinogens, as well as endogenous substrates. We report the presence of cytochrome P-450 and associated mono-oxygenase activity in human brain regions and their selective enrichment in the brainstem. Immunocytochemical studies on human medulla with antibodies raised to phenobarbital-inducible rat liver cytochrome P-450 indicate that the enzyme is primarily localized in the neuronal cell bodies and to a lesser extent in the axons. These observations indicate that the human brain could be involved in metabolism of xenobiotics and endogenous compounds, mediated through cytochrome P-450.

Adult↗

PubMed1989

High activity of cytochrome P-450-linked aminopyrine N-demethylase in mouse brain microsomes, and associated sex-related difference.

The presence of cytochrome P-450 and associated mono-oxygenase activities was examined in brain microsomes from male and female mice. Although the cytochrome P-450 level in male mouse brain was very low as compared with mouse liver, the aminopyrine N-demethylase and morphine N-demethylase specific activities in male mouse brain were much higher than those observed in mouse liver. Ethoxycoumarin O-de-ethylase and aniline hydroxylase activities were, however, not detected in mouse brain. Sex-related differences were observed in both the cytochrome P-450 levels and aminopyrine N-demethylase activity in mouse brain, the levels of both being higher in male mouse brain as compared with female mouse brain. Aminopyrine N-demethylase activity in mouse brain microsomes was dependent on the presence of oxygen and NADPH and could be inhibited by piperonyl butoxide, N-octyl imidazole and carbon monoxide. Antiserum raised to the phenobarbital-inducible form of rat liver cytochrome P-450 [P-450(b+e)] inhibited mouse brain aminopyrine N-demethylase activity by around 80+ mouse brain microsomal protein exhibited cross-reactivity against this antiserum when examined by Ouchterlony double diffusion and immunoblotting. The present results indicate the presence of a phenobarbital-inducible form of cytochrome P-450 (or a form of cytochrome P-450 that is similar immunologically) in mouse brain microsomes, which is associated with a sex-related difference.

Aminopyrine N-Demethylase↗

PubMed1989

Low glutathione levels in brain regions of aged rats.

Glutathione (GSH) was measured in 6 regions of brain and liver of young adult, middle-aged and aged rats. GSH levels were significantly lower in cortex, cerebellum, striatum, thalamus and hippocampus of aged rats, while no changes were observed in liver as compared to young adult rats. On the other hand, lipid peroxidation as measured by thiobarbituric acid-reactive products increased significantly in all the regions of brain examined and in the liver of aged rats. Since GSH plays an important role as a cellular protectant against oxygen radical-mediated injury, decreased levels of GSH in aged rat brain are indicative of the vulnerability of the aged cerebral tissue to oxidative injury.

Aging↗

PubMed1989

Production of deoxynivalenol by Fusarium isolates from samples of wheat associated with a human mycotoxicosis outbreak and from sorghum cultivars.

Fusarium isolates from specific diseased sorghum plants and rain-soaked wheat and wheat flour associated with human mycotoxicosis in India have been screened for their toxigenic potential. Of the 322 isolates screened, 11 isolates were found to produce deoxynivalenol in concentrations ranging from 0.01 to 186 micrograms g-1. The occurrence of deoxynivalenol-producing fusaria in a nontemperate region and deoxynivalenol production in low concentrations by Fusarium moniliforme are reported for the first time.

Disease Outbreaks↗

PubMed1987

Hepatotoxicity of precocene I in rats. Role of metabolic activation in vivo.

The mechanism of the hepatotoxicity of precocene I has been investigated in male, Sprague-Dawley rats. Administration of a single dose of precocene I caused a large depletion of liver glutathione (GSH) levels that was both time and dose dependent. Concomitant with the decrease of liver GSH, there was an increase in serum glutamic pyruvic transaminase (GPT) levels which was also time and dose dependent. Administration of a single dose of [4-3H]precocene I resulted in extensive covalent binding of the radiolabel to liver proteins and DNA in the liver; the extent of binding increased with increasing dose. Treatment of the rats with the mixed-function oxidase inhibitor piperonyl butoxide, before administration of precocene I, significantly decreased the proportion of the radiolabel bound covalently to proteins and DNA, although the total radioactivity (bound and unbound) in the liver remained the same. Piperonyl butoxide pretreatment limited both the liver GSH depletion and the hepatic necrosis normally caused by precocene I. These results are consistent with the view that the hepatotoxicity of precocene I is due to reactive metabolites formed through cytochrome P-450 mediated metabolism of precocene I.

Alanine Transaminase↗

PubMed1986

2-Methylfuran toxicity in rats--role of metabolic activation in vivo.

Administration of a single ip dose of 2-methylfuran (2-MF) to male Sprague-Dawley rats at a dose of 100 mg/kg produced centrilobular necrosis of the liver and bronchial injury of the lung, the severity of the lesions increasing with increasing doses up to 400 mg/kg. Kidneys, however, showed no visible evidence of tissue damage even at the highest dose. Liver injury was also evidenced by an increase in serum glutamic pyruvic transaminase (SGPT) levels. Tissue distribution and covalent binding studies conducted over a dose of 50-200 mg/kg of [14C]2-MF indicated that the total radioactivity present per gram of wet tissue was in the order of liver greater than kidney greater than lung greater than blood. Covalent binding of the label to protein was greatest in the liver followed by kidney and the lung. Radioactivity bound covalently per milligram of DNA was also highest in the liver followed by kidney. Tissue distribution and covalent binding studies were conducted over a period of 0.5 to 24 hr after an ip dose of 100 mg/kg of [14C]2-MF. Maximal covalent binding was observed in the liver at 4 hr. At all time points binding of the label was greatest in liver, followed by kidney. Liver glutathione levels were depressed following 2-MF administration. Pretreatment of rats with phenobarbital markedly increased the covalent binding to protein and DNA and caused a twofold increase in SGPT compared to rats treated with 2-MF alone. Pretreatment with 3-methylcholanthrene had no effect on either parameter. Administration of N-octylimidazole, an inhibitor of cytochrome P-450, prior to administration of the radiolabeled 2-MF decreased the covalent binding of the label to protein and DNA. Moreover, the SGPT levels remained the same in the pretreated rats compared to the rats treated with vehicle alone. Thus, pretreatment with phenobarbital, an inducer of cytochrome P-450, enhanced both covalent binding and toxicity while prior treatment with N-octylimidazole, an inhibitor of cytochrome P-450 decreased covalent binding and prevented hepatotoxicity of 2-MF. These results support the view that at least some of the toxic effects of 2-MF are mediated by reactive metabolite(s) formed in vivo.

Alanine Transaminase↗

PubMed1985

Metabolic activation of 2-methylfuran by rat microsomal systems.

2-Methylfuran (2-MF), a constituent of cigarette smoke and coffee, causes necrosis of liver, lungs, and kidneys in rodents. 2-MF is metabolically activated by mixed-function oxidases to acetylacrolein, a reactive metabolite that binds covalently to microsomal protein. The hepatic microsomal metabolism of 2-MF to reactive metabolite required the presence of NADPH and oxygen and was dependent on incubation time and substrate concentration. The microsomal metabolism of 2-MF was inducible by pretreatment of rats with phenobarbital and was inhibited by piperonyl butoxide and N-octyl imidazole, which indicates that the metabolism of 2-MF may be mediated by cytochrome P-450. Acetylacrolein was a potent inhibitor of mixed-function oxidase and completely inhibited the microsomal metabolism of 2-MF, indicating that 2-MF is a suicide substrate for the enzyme. The sulfhydryl nucleophile cysteine was a better trapping agent of the reactive metabolite of 2-MF than N-acetylcysteine or glutathione. Lysine decreased the covalent binding of 2-MF metabolites, presumably by reacting with the aldehyde group of acetylacrolein. In addition, in the presence of NADPH, 2-MF was bioactivated by both pulmonary and renal cortical microsomes to reactive metabolites that were covalently bound to microsomal proteins.

Acrolein↗

PubMed1984

Reactive metabolites from the bioactivation of toxic methylfurans.

An important mechanism of toxicity of furans involves the cytochrome P-450 monooxygenase-catalyzed bioactivation of the compound in situ directly within the target tissues to highly reactive electrophilic products. The unsaturated aldehydes acetylacrolein and methylbutenedial have been identified as the principal reactive intermediates of 2- and 3-methylfuran, respectively, that are produced and bound covalently to tissue macromolecules in hepatic and pulmonary microsomal systems in vitro.

Animals↗

PubMed1981

In vitro studies on the intestinal absorption of curcumin in rats.

When everted sacs of rat intestines were incubated with 50-750- micrograms of curcumin in 10 ml incubation medium, 30-80% of the added curcumin disappeared from the mucosal side. No curcumin was however detectable in the serosal fluid. Less than 3% of the added curcumin was found in the tissue at the highest concentration. In experiments with [3H[ curcumin, 5-6% of added radioactivity was found in the serosal side. TLC examination of the mucosal extract showed the presence of 2 compounds, 1 corresponding to curcumin and the other to a less polar, colourless compound. The serosal fluid had no curcumin but a compound whose RF was identical with the colourless compound present in the mucosal side. These experiments indicated that curcumin undergoes transformation during absorption from the intestine.

Animals↗

PubMed1980

Absorption and tissue distribution of curcumin in rats.

After oral administration of 400 mg curcumin to rats, about 60% of the dose was absorbed. No curcumin was detectable in urine. The urinary excretion of conjugated glucuronides and sulfates significantly increased. No curcumin was present in heart blood. Only traces (less than 5 microgram/ml) in portal blood and negligible quantities in liver and kidney (< 20 micrograms/tissue) were observed from 15 min upto 24 h after administration of curcumin. At the end of 24 h the concentration of curcumin remaining in the lower part of the gut namely caecum and large intestine amounted to 38% of the quantity administered.

Animals↗

PubMedDate not supplied

Metabolism of curcumin--studies with [3H]curcumin.

Radioactivity was detectable in blood, liver and kidney following dosage with 400, 80 or 10 mg of [3H]curcumin. The major route of elimination of the label was the feces; the urinary excretion of the label was very low regardless of the dose. At the lower doses of 80 mg and 10 mg of [3H]-curcumin, most of the label was excreted within 72 h, while with 400 mg, considerable amounts of the label was present in the tissues 12 days after dosage. The percentage of curcumin absorbed (60-66% of the given dose) remained constant regardless of the dose administered.

Animals↗