Rejoining of radiation-induced single-strand breaks in deoxyribonucleic acid of Escherichia coli: effect of phenethyl alcohol.
(17/635)
Single-strand breaks in deoxyribonucleic acid of Escherichia coli B/r cells exposed to 20 krads of gamma radiation could be rejoined by incubation of irradiated cells in growth medium. In the presence of 0.25% phenethyl alcohol, this repair was completely inhibited although deoxyribonucleic acid and protein syntheses were suppressed only partially. (+info)
Accumulation of the capacity of initiation of deoxyribonucleic acid replication in Escherichia coli.
(18/635)
Several temperature-sensitive initiation mutants of Escherichia coli were examined for the ability to initiate more than one round of replication after being held at nonpermissive temperature for approximately 1.5 generation equivalents. The capacity for initiation was measured by residual synthesis experiments and rate experiments under conditions where protein synthesis and ribonucleic acid synthesis were inhibited. Results of the rate and density transfer experiments suggest that the cells may initiate more than one round of replication in the absence of protein or ribonucleic acid synthesis. This contrasts with the results of the residual synthesis experiments which suggest that, under these conditions, only one round of synthesis is achieved. These findings suggest that the total amount of residual synthesis achieved in the presence of an inhibitor may be both a function of the number of initiation events which occur and the effect of the inhibitor of protein or ribonucleic acid synthesis on chain elongation. (+info)
Hydroperoxide specificity of plant and human tissue lipoxygenase: an in vitro evaluation using N-demethylation of phenothiazines.
(19/635)
Since hydroperoxide specificity of lipoxygenase (LO) is poorly understood at present, we investigated the ability of cumene hydroperoxide (CHP) and tert-butyl hydroperoxide (TBHP) to support cooxidase activity of the enzyme toward the selected xenobiotics. Considering the fact that in the past, studies of xenobiotic N-demethylation have focused on heme-proteins such as P450 and peroxidases, in this study, we investigated the ability of non-heme iron proteins, namely soybean LO (SLO) and human term placental LO (HTPLO) to mediate N-demethylation of phenothiazines. In addition to being dependent on peroxide concentration, the reaction was dependent on enzyme concentration, substrate concentration, incubation time, and pH of the medium. Using Nash reagent to estimate formaldehyde production, the specific activity under optimal assay conditions for the SLO mediated N-demethylation of chlorpromazine (CPZ), a prototypic phenothiazine, in the presence of TBHP, was determined to be 117+/-12 nmol HCHO/min/mg protein, while that of HTPLO was 3.9+/-0.40 nmol HCHO/min/mg protein. Similar experiments in the presence of CHP yielded specific activities of 106+/-11 nmol HCHO/min/mg SLO, and 3.2+/-0.35 nmol HCHO/min/mg HTPLO. As expected, nordihydroguaiaretic acid and gossypol, the classical inhibitors of LOs, as well as antioxidants and free radical reducing agents, caused a marked reduction in the rate of formaldehyde production from CPZ by SLO in the reaction media fortified with either CHP or TBHP. Besides chlorpromazine, both SLO and HTPLO also mediated the N-demethylation of other phenothiazines in the presence of these organic hydroperoxides. (+info)
Edge-to-face CH/pi interaction between ligand Phe-phenyl and receptor aromatic group in the thrombin receptor activation.
(20/635)
In the ligand/receptor interaction, the side chain phenyl group of phenylalanine (Phe) is involved in a so-called hydrophobic interaction, in which the Phe-phenyl group functions as a p element or merely as a hydrophobic element. The thrombin receptor-tethered ligand SFLLRNP consists of the Phe-2 residue essential for receptor activation. In order to explore the molecular characteristics of this Phe-2-phenyl group, a complete set of S/Phe/LLRNP peptides comprising six different difluorophenylalanine isomers [(F(2))Phe] was newly synthesized and assayed to evaluate their ability to induce the aggregation of human platelets. The assay results clarified several important structural elements to conclude that Phe-2-phenyl of S/Phe/LLRNP is in the edge-to-face CH/pi interaction with the receptor aromatic group, utilizing the Phe-phenyl edge along with adjacent benzene hydrogens at positions (2-3) or (5-6). It was also found that the fluorine atom at position 4 increases the acidity of the hydrogen mainly at its ortho position, resulting in a reinforcement of the CH/pi interaction and thus in an enhancement of biological activity. The H-->F replacement in the benzene ring was found to provide an effective structural examination to the Phe residue; i.e., to identify the hydrogens in the CH/pi interaction, and to strengthen the CH/pi interaction. (+info)
Red cells from glutathione peroxidase-1-deficient mice have nearly normal defenses against exogenous peroxides.
(21/635)
The role of glutathione peroxidase in red cell anti-oxidant defense was examined using erythrocytes from mice with a genetically engineered disruption of the glutathione peroxidase-1 (GSHPx-1) gene. Because GSHPx-1 is the sole glutathione peroxidase in the erythrocyte, all red cell GSH peroxidase activity was eliminated. Oxidation of hemoglobin and membrane lipids, using the cis-parinaric acid assay, was determined during oxidant challenge from cumene hydroperoxide and H(2)O(2). No difference was detected between wild-type red cells and GSHPx-1-deficient cells, even at high H(2)O(2) exposures. Thus, GSHPx-1 appears to play little or no role in the defense of the erythrocyte against exposure to peroxide. Simultaneous exposure to an H(2)O(2) flux and the catalase inhibitor 3-amino-1,2,4-triazole supported this conclusion. Hemoglobin oxidation occurred only when catalase was depleted. Circulating erythrocytes from the GSHPx-1-deficient mice exhibited a slight reduction in membrane thiols, indicating that high exposure to peroxides might occur naturally in the circulation. (Blood. 2000;96:1985-1988) (+info)
The antioxidant drug dipyridamole spares the vitamin E and thiols in red blood cells after oxidative stress.
(22/635)
OBJECTIVE: To test the antioxidant effect of therapeutic doses of dipyridamole on cellular membranes, human erythrocytes were chosen as an appropriate model to study oxidative stress induced by cumene hydroperoxide because of their high content in heme-Fe(2+). METHODS: The oxidative stress was induced by incubation with 160 micromoll(-1) cumene hydroperoxide and expressed by three main factors: lipid peroxidation by means of kinetics of decrease in fluorescence emission of the probe incorporated in the cell membranes, vitamin E oxidation and intracellular thiol content. The concentrations of dipyridamole tested (2-20 micromoll(-1)) did not exceed pharmacological doses. RESULTS: After 7 min of incubation at 25 degrees C with the oxidant and 20 micromoll(-1) dipyridamole thiol content was 50.1%+/-2.6 compared with 31.5%+/-2.4 in the absence of the drug. After 12 min vitamin E content was 88.3%+/-2.3 compared with 64.7%+/-3.4 of untreated cells in the absence of dipyridamole. Dipyridamole added 5 min after the oxidation reaction suppressed the fluorescence decrease for a time proportional to the drug concentration. CONCLUSIONS: Thus, at clinically realistic doses dipyridamole shows a concentration-dependent antioxidant effect. It protects membranes from oxidation and spares the antioxidant power of erythrocytes. (+info)
A GC-MS method for the detection of toluene and ethylbenzene in volatile substance abuse.
(23/635)
The interference of some substances with the gas chromatography-flame ionization detection and gas chromatography-Fourier transform infrared detection of toluene and ethylbenzene in volatile substance abuse poses problems. A gas chromatography-mass spectrometry (GC-MS) method that will overcome such interference has been developed for the detection of toluene and/or ethylbenzene in the headspace of preparations and products containing these substances and in the headspace of blood samples in the cases of volatile substance abuse. The method is based on converting toluene to benzoic acid via the formation of benzotrichloride. The latter compound was obtained upon the reaction of toluene with chlorine gas under direct sunlight conditions. In the presence of water, benzotrichloride was converted to benzoic acid. Ethylbenzene was converted to benzoic acid and two phenylethanols via the formation of side chain chloro-substituted phenylethanes followed by reaction with water. The chloro-substituted phenylethanes were obtained by the reaction of ethylbenzene with chlorine under direct sunlight conditions. The benzoic acid resulting from toluene and/or ethylbenzene and the two phenylethanols resulting from ethylbenzene were detected by GC-MS as their trimethylsilyl (TMS) derivatives. For the method to be viable for the detection of volatile substance abuse, the chlorination reactions were effected in the gaseous state. (+info)
Effects of volatile solvents on recombinant N-methyl-D-aspartate receptors expressed in Xenopus oocytes.
(24/635)
1. We have previously shown that toluene dose-dependently inhibits recombinant N-methyl-D-aspartate (NMDA) receptors at micromolar concentrations. This inhibition was rapid, almost complete and reversible. The NR1/2B combination was the most sensitive receptor subtype tested with an IC(50) value for toluene of 0.17 mM. 2. We now report on the effects of other commonly abused solvents (benzene, m-xylene, ethylbenzene, propylbenzene, 1,1,1-trichlorethane (TCE) and those of a convulsive solvent, 2,2,2-trifluoroethyl ether (flurothyl), on NMDA-induced currents measured in XENOPUS oocytes expressing NR1/2A or NR1/2B receptor subtypes. 3. All of the alkylbenzenes and TCE produced a reversible inhibition of NMDA-induced currents that was dose- and subunit-dependent. The NR1/2B receptor subtype was several times more sensitive to these compounds than the NR1/2A subtype. 4. The convulsant solvent flurothyl had no effect on NMDA responses in oocytes but potently inhibited ion flux through recombinant GABA receptors expressed in oocytes. 5. Overall, these results suggest that abused solvents display pharmacological selectivity and that NR1/2B NMDA receptors may be an important target for the actions of these compounds on the brain. (+info)