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Biomedical subjects

A Gescher

Publications and source records attributed to A Gescher.

137 records · Page 8Linked to original sources
PubMedDate not supplied

Determination of quercetin in human plasma by HPLC with spectrophotometric or electrochemical detection.

A reversed-phase high-performance liquid chromatographic method for the determination of quercetin in human plasma following intravenous infusion is described. Quercetin in plasma was extracted with methanol-dimethyl sulphoxide (4:1 v/v) and separated on a C18 Hypersil-BDS column with 44% (v/v) methanol in 0.1 M ammonium acetate (pH 5.15) containing 0.27 mM EDTA as the mobile phase. The drug was detected specifically and sensitively at its absorption maximum of 375 nm, or electrochemically, with a detection limit of 80 ng/mL and 2 ng/mL, respectively.

Chromatography, High Pressure Liquid↗
PubMedDate not supplied

The reaction of alkylnitronates with glutathione.

Nitroalkanes are agents of occupational importance, and 2-nitropropane has been shown to be mutagenic and hepatotoxic. Here the reaction of alkylnitronates, such as propyl-2-nitronate, with glutathione and other thiol nucleophiles has been studied by using TLC, HPLC, and spectroscopic methods. Propyl-2-nitronate, but not 2-nitropropane, reacted with glutathione in acidic media. The reaction yielded an inseparable mixture of oxidized glutathione and a product that was identified as S-nitrosoglutathione. S-Nitrosoglutathione is known to be generated by reaction of nitrous acid with glutathione and furnishes a characteristic UV spectrum, but its NMR and mass spectral properties are described here for the first time. The 1H NMR spectrum of S-nitrosoglutathione showed in principle the resonance signals of glutathione except that the cysteine beta-protons gave two broad signals shifted downfield by 1 ppm as compared to the resonance frequencies of the glutathione cysteine beta-protons. The interpretation of the spectrum was aided by investigation of the properties of S-nitroso-N-acetylcysteine, the product of the reaction of N-acetylcysteine with propyl-2-nitronate. The nitronates of primary nitroalkanes, such as nitromethane, nitroethane, or 1-nitropropane, did not react with glutathione. The reaction between propyl-2-nitronate and glutathione did not occur at pH values greater than 5; therefore, the relevance of these findings to the disposition of propyl-2-nitronate in vivo is unclear.

Chromatography, High Pressure Liquid↗
PubMedDate not supplied

Chemical synthesis and cytotoxic properties of N-alkylcarbamic acid thioesters, metabolites of hepatotoxic formamides.

The S-linked cysteine and glutathione conjugates of N-methylformamide and N-ethylformamide, together with a series of methyl ester derivatives thereof, have been synthesized and characterized by 1H NMR, 13C NMR, FAB-MS, and FAB tandem mass spectrometry. In vitro cytotoxicity assays showed that all of the title conjugates were toxic to isolated mouse hepatocytes when incubated at concentrations in excess of 1 mM and that they served as potent growth inhibitors of murine TLX5 lymphoma cells when present at levels of 10-100 microM. Both of these effects were reversed by the addition to incubation media of glutathione (10 mM). The possibility is raised that N-alkylcarbamic acid thioester conjugates, which are formed during the metabolism of N-alkylformamides in mammalian systems, may act as important mediators of the antineoplastic and/or hepatotoxic activity of the parent formamides, possibly through their ability to liberate methyl isocyanate at cell membranes.

Animals↗
PubMedDate not supplied

N-alkylformamides are metabolized to N-alkylcarbamoylating species by hepatic microsomes from rodents and humans.

Hepatotoxic formamides such as N-methylformamide (NMF) and N,N-dimethylformamide (DMF) are metabolized in vivo to N-acetyl-S-(N-methylcarbamoyl)cysteine via oxidation at the formyl carbon, which yields a reactive intermediate. The hypothesis was tested that this biotransformation route can be studied in vitro with hepatic fractions. NMF was incubated with microsomes or cytosol obtained from BALB/c mice, and metabolically generated N-methyl-carbamoylating species were analyzed after derivatization with ethanol in base to furnish ethyl N-methylcarbamate. Generation of metabolite was catalyzed by microsomes, but not by cytosol. Detection of the N-methylcarbamoylating species was dependent on the presence in the incubation mixture of NMF, viable microsomes, NADPH, and a thiol-containing agent such as glutathione. Metabolite formation was inhibited by SKF 525-A (3 mM) and abolished when the incubation atmosphere consisted of an air/carbon monoxide mixture (1:1) instead of air. Metabolism was not induced by pretreatment of mice with phenobarbital or beta-naphthoflavone. N-Ethylformamide and the DMF metabolite N-(hydroxymethyl)-N-methylformamide, but not DMF, were metabolized by microsomes to the N-alkylcarbamoylating metabolite at a measurable rate. NMF metabolism was also observed with liver microsomes from Sprague-Dawley rats or from humans. In the case of rat microsomes the rate of metabolism was half of that measured with murine microsomes. The results suggest that (i) the metabolic toxification of NMF can be studied in hepatic microsomes and (ii) the oxidation of the formyl moiety in N-alkylformamides is catalyzed by cytochrome P-450.

Animals↗
PubMedDate not supplied

Investigation of the mechanistic basis of N,N-dimethylformamide toxicity. Metabolism of N,N-dimethylformamide and its deuterated isotopomers by cytochrome P450 2E1.

Dimethylformamide (DMF) is an industrial solvent with hepatotoxic properties. The toxicity of DMF has been associated with its metabolism to S-(N-methylcarbamoyl)glutathione (SMG). The major urinary metabolite of DMF is N-(hydroxymethyl)-N-methylformamide (HMMF). HMMF undergoes oxidation in the formyl moiety, possibly via the intermediacy of its hydrolysis product N-methylformamide (NMF), and the reactive intermediate thus generated reacts with glutathione to yield SMG. Experiments were conducted to elucidate enzymatic details of the metabolism of DMF. Generation of HMMF from DMF in microsomes from rats which had received acetone, an inducer of cytochrome P450 2E1, was increased by 175% over that observed in control microsomes. In liver microsomes from 4 humans the metabolism of DMF to HMMF was inhibited by a monospecific antibody against rat liver P450 2E1, and the metabolic rates were correlated with those of NMF to SMG, a process known to be mediated via P450 2E1. DMF was also metabolized by purified rat liver P450 2E1. The kinetic parameters which characterize the metabolism of DMF or its deuterated isotopomers to the respective HMMF isotopomers, of HMMF to SMG and of NMF to SMG in liver microsomes, were computed from Eadie-Hofstee plots. The affinity of DMF for the metabolizing enzyme in rat liver microsomes is considerably higher (apparent Km = 0.20 mM) than that of NMF (Km = 4.28 mM) or of HMMF (Km = 2.52 mM). The respective values observed with human microsomes are very similar. The apparent Km values for the N-methyl oxidation of N,N-dimethyldeuterioformamide ([2H1]DMF) and N,N-bis(trideuteriomethyl)formamide ([2H6]DMF) in rat microsomes are 0.14 and 0.21 mM, respectively. The apparent Vmax for the oxidation of [2H1]DMF is similar to that computed for DMF, and the Vmax for [2H6]DMF is less than half of that computed for DMF. The kinetic deuterium isotope effect (KDIE) on DMF metabolism was determined in incubations with rat microsomes in three ways: (i) the noncompetitive intermolecular KDIE by the ratio of Vmax/Km for DMF to Vmax/Km for [2H6]DMF, (ii) the competitive intermolecular KDIE as the quotient of metabolic products HMMF to N-(hydroxydideuteriomethyl)-N-(trideuteriomethyl)formamide in incubations of DMF together with [2H6]DMF, and (iii) the intramolecular KDIE as the quotient of the ratio of N-(hydroxymethyl)-N-(trideuteriomethyl)formamide to N-(hydroxydideuteriomethyl)-N-methylformamide generated from N-(trideuteriomethyl)-N-methylformamide ([2H3]DMF). The respective values were found to be (i) 2.4, (ii) 5.0, and (iii) 5.2. DMF inhibited the oxidation of NMF or HMMF to SMG.(ABSTRACT TRUNCATED AT 400 WORDS)

Amides↗
PubMedDate not supplied

The fate of N-methylformamide in mice. Routes of elimination and characterization of metabolites.

The fate of N-methylformamide has been investigated in male CBA/CA mice following the administration of this compound labeled with 14C either in the methyl or in the formyl group. The major route of elimination was found to be via the kidneys although a substantial quantity (39% of the dose) was eliminated via the lungs as CO2 in the case of [14C]formyl-labeled N-methylformamide. In addition to the unchanged compound three metabolites were found in the urine by TLC autoradiography. One of these metabolites was identified as methylamine after conversion to its 2,4-dinitrophenyl derivative. The derivative was isolated and shown to be N-methyl-2,4-dinitroaniline by mass spectrometry. Further evidence that methylamine was a metabolite of N-methylformamide was provided by ion pair HPLC analysis of urine from mice dosed with [14C]methyl-labeled N-methylformamide. The second metabolite was tentatively identified as N-hydroxymethylformamide which was present in the urine of mice dosed with either [14C]methyl- or [14C]formyl-labeled N-methylformamide. Formate was not a urinary metabolite of N-methylformamide. The identity of the third urinary metabolite remains unknown.

Animals↗
PubMedDate not supplied

Pharmacokinetics of hexamethylmelamine and pentamethylmelamine in mice.

Plasma pharmacokinetics of hexamethylmelamine (HMM) and pentamethylmelamine (PMM) were investigated after ip administration of doses of 50 or 100 mg/kg to M5076/73A ovarian tumor-bearing C57BL/6J female mice. The half-life of HMM was 44.5-49 mins; the half-life of PMM was 7.6-8.7 mins. The area under the concentration-versus-time curve (AUC) was also much higher for HMM than for PMM. The AUC for the metabolite N2,N2,N4,N6-tetramethylmelamine (TMM) was seven to ten times higher than that of the parent compounds. After incubation with mouse liver microsomes and cofactors, the rates of metabolism of HMM and PMM were essentially the same. Plasma protein binding was 94%, 71%, and 50% for HMM, PMM, and TMM, respectively.

Altretamine↗
PubMedDate not supplied

Inhibition of p-nitrophenol hydroxylase in rat liver microsomes by small aromatic and heterocyclic molecules.

The cytochrome P450 isoenzyme P4502E1 is constitutively expressed in human liver and catalyzes the oxidation of many known or suspected carcinogens of low molecular weight. In this structure-metabolism study, the role that heteroatoms in heterocyclic compounds play in determining their affinity for P4502E1 was investigated. The ability of 16 six-membered and 10 five-membered compounds to inhibit the hydroxylation of p-nitrophenol, which is specifically catalyzed by P4502E1, was studied in suspensions of microsomes from rat livers in which P4502E1 had been induced by inclusion of acetone in the drinking water. Apparent Ki values were extrapolated from kinetic models of Dixon or Cornish-Bowden plots for enzyme inhibition. Enzyme inhibition was generally of the non-or uncompetitive type. Pyridine was the most potent and benzene one of the least potent inhibitors, with Ki values of 0.4 microM and 8,400 microM, respectively. Pyridazine was less inhibitory than 1,3,5-triazine, which inhibited P4502E1 to a lesser degree than pyrazine and pyrimidine. Among the unsubstituted unsaturated five-membered ring molecules, pyrrole was a better inhibitor than furan or thiophene. 4-Methylimidazole was a much stronger inhibitor than imidazole or 1-and 2-methylimidazole. The ability of compounds to inhibit P4502E1 seems to depend in the main on the presence of a nitrogen atom in the molecule and on the ability of the nitrogen lone pair of electrons to ligand to the heme.

Animals↗