Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Detection of peroxyl and alkoxyl radicals produced by reaction of hydroperoxides with rat liver microsomal fractions.

ABSTRACT: E.s.r. spin trapping using the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) was used to detect peroxyl, alkoxyl and carbon-centred radicals produced by reaction of t-butyl hydroperoxide (tBuOOH) with rat liver microsomal fraction. The similarity of the hyperfine coupling constants of the peroxyl and alkoxyl radical adducts to those obtained previously with isolated enzymes suggests that these species are the tBuOO. and tBuO. adducts. The effects of metal-ion chelators, heat denaturation, enzyme inhibitors and reducing equivalents demonstrate that these species arise from reaction of tBuOOH with a haem enzyme such as cytochrome P-450 or cytochrome b5. In the absence of NADPH or NADH the previously undetected peroxyl radical adduct is the major species observed. In the presence of these reducing equivalents the alkoxyl and carbon-centred radical adducts predominate, which is in accord with product studies on similar systems. These results demonstrate that both reductive and oxidative decomposition of tBuOOH can occur in rat liver microsomal fraction with the reductive pathway favoured in the presence of NADH or NADPH.

SUBMITTER: Davies MJ

PROVIDER: S-EPMC1135621 | biostudies-other | 1989 Jan

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Interactions of some acceptors with superoxide anion radicals formed by the NADPH-specific flavoprotein in rat liver microsomal fractions.

Project description:In rat liver microsomal fractions oxidation of adrenaline was effected by superoxide anion radicals (O2-), whereas cytochrome c, 2,6-dichlorophenol-indophenol and ferricyanide accepted electrons from NADPH-specific flavoprotein only directly. Nitro Blue Tetrazolium was reduced both by O2- and by the direct acceptance of electrons. Elevation of pH and addition of menadione shift the Nitro Blue Tetrazolium reduction towards the O2--dependent pathway. From the values of the kinetic constants for interaction of adrenaline and Nitro Blue Tetrazolium with NADPH-specific flavoprotein, the rates of generation of O2- in rat liver microsomal fraction were determined.

| S-EPMC1172711 | biostudies-other

A Theoretical Kinetic Study on Concerted Elimination Reaction Class of Peroxyl-hydroperoxyl-alkyl Radicals (•OOQOOH) in Normal-alkyl Cyclohexanes.

Project description:The concerted elimination reaction class of peroxyl-hydroperoxyl alkyl radicals (•OOQOOH) plays a crucial role in the low-temperature combustion of normal-alkyl cyclohexanes. The generation of the relatively unreactive HO2 radicals in this reaction is one of the factors leading to the negative temperature coefficient (NTC) behavior, which hinders the low-temperature oxidation of normal-alkyl cyclohexanes. In this study, 44 reactions are selected and divided into 4 different subclasses according to the nature of the carbon atom where the H atom is eliminated and the reaction center position. Utilizing the CBS-QB3 method, we compute the energy barriers for the concerted elimination reactions of peroxyl-hydroperoxyl alkyl radicals. Following this, we assess both the high-pressure limit and pressure-dependent rate constants for all reactions by applying TST and RRKM/ME theory. These calculations allow for the development of rate rules, which come to fruition through an averaging process involving the rate constants of representative reactions within each subclass. Our work provides accurate rate constants and rate rules for this reaction class, which can aid in constructing more accurate combustion mechanisms for normal-alkyl cyclohexanes.

| S-EPMC10536632 | biostudies-literature

Purification of membrane-bound galactosyltransferase from rat liver microsomal fractions.

Project description:1. Rat liver microsomal preparations incubated in 1% Triton X-100 at 37 degrees C for 1h released about 60% of the membrane-bound UDP-galactose-glycoprotein galactosyltransferase (EC 2.4.1.22) into a high-speed supernatant. The supernatant galactosyltransferase which was solubilized but not purified by this treatment had a higher molecular weight than the serum enzyme as shown by Sephadex G-100 column chromatography. 2. The galactosyltransferase present in the high-speed supernatant was purified 680-fold by an affinity-column-chromatographic technique by using a column of activated Sepharose 4B coupled with alpha-lactalbumin. The galactosyltransferase ran as a single band on polyacrylamide gels and contained no sialyltransferase, N-acetylglucosaminyltransferase or UDP-galactose pyrophosphatase activities. 3. The purified membrane enzyme had properties similar to serum galactosyltransferase. It had an absolute requirement for Mn(2+) that could not be replaced by Ca(2+), Mg(2+), Zn(2+) or Co(2+), and was active over a wide pH range (6-8) with a pH optimum of 6.5. The apparent K(m) for UDP-galactose was 10.8mum. The protein alpha-lactalbumin modified the enzyme to a lactose synthetase by increasing substrate specificity for glucose in preference to N-acetylglucosamine and fetuin depleted of sialic acid and galactose. 4. The molecular weight of the membrane enzyme was 65000-70000, similar to that of the purified serum enzyme. Amino acid analyses of the two proteins were similar but not identical. 5. Sephadex G-100 column chromatography of the purified membrane enzyme showed a small peak (2-5%) of higher molecular weight than the purified serum enzyme. Inclusion of 1mm-epsilon-aminohexanoic acid in the isolation procedures increased this peak to as much as 30% of the total enzyme recovered. Increasing the epsilon-aminohexanoic acid concentration to 100mm resulted in no further increase in this high-molecular-weight fraction.

| S-EPMC1164830 | biostudies-other

Palmitate activation and esterification in microsomal fractions of rat liver.

Project description:Palmitoyl-CoA synthetase activity in the microsomal fraction of rat liver was measured directly by palmitoyl-CoA production, and indirectly by converting the palmitoyl-CoA into palmitoylcarnitine under optimum conditions. Even in the latter system, palmitoyl-CoA accumulated. The rate of palmitoyl-CoA hydrolysis and the inhibition of palmitoyl-CoA synthetase by palmitoyl-CoA were each estimated to be less than 10% of the maximum rate of palmitoyl-CoA production. The concentration of palmitoyl-CoA present in the assay systems used for measuring palmitate esterification to glycerol phosphate and the activity of palmitoyl-CoA synthetase by using the carnitine-linked determination were measured. These concentrations were not altered by the addition of glycerol phosphate, or of carnitine plus carnitine palmitoyltransferase. The relationship between the activity of palmitoyl-CoA synthetase and the rate of glycerolipid synthesis was investigated. The latter activity was measured by using palmitoyl-CoA generated from palmitate, palmitoyl--AMP or palmitoylcarnitine. It is concluded that, at optimum substrate concentrations, the activity of glycerol phosphate acyltransferase is rate-limiting in the synthesis of phosphatidate by rat liver microsomal fractions. The implications of these results in the measurement of palmitoyl-CoA synthetase and in the control of glycerolipid synthesis are discussed.

| S-EPMC1172437 | biostudies-other

Hydroxycinnamyl Derived BODIPY as a Lipophilic Fluorescence Probe for Peroxyl Radicals.

Project description:Herein, we describe the synthesis of a fluorescent probe NB-2 and its use for the detection of peroxyl radicals. This probe is composed of two receptor segments (4-hydroxycinnamyl moieties) sensitive towards peroxyl radicals that are conjugated with a fluorescent reporter, dipyrrometheneboron difluoride (BODIPY), whose emission changes depend on the oxidation state of the receptors. The measurement of the rate of peroxidation of methyl linoleate in a micellar system in the presence of 1.0 µM NB-2 confirmed its ability to trap lipid peroxyl radicals with the rate constant kinh = 1000 M-1·s-1, which is ten-fold smaller than for pentamethylchromanol (an analog of ?-tocopherol). The reaction of NB-2 with peroxyl radicals was further studied via fluorescence measurements in methanol, with ?,?'-azobisisobutyronitrile (AIBN) used as a source of radicals generated by photolysis or thermolysis, and in the micellar system at pH 7.4, with 2,2'-azobis(2-amidinopropane) (ABAP) used as a thermal source of the radicals. The reaction of NB-2 receptors with peroxyl radicals manifests itself by the strong increase of a fluorescence with a maximum at 612-616 nm, with a 14-fold enhancement of emission in methanol and a 4-fold enhancement in the micelles, as compared to the unoxidized probe. Our preliminary results indicate that NB-2 behaves as a "switch on" fluorescent probe that is suitable for sensing peroxyl radicals in an organic lipid environment and in bi-phasic dispersed lipid systems.

| S-EPMC7022944 | biostudies-literature

Detection of free radicals produced from the reaction of cytochrome P-450 with linoleic acid hydroperoxide.

Project description:The ESR spin-trapping technique was employed to investigate the reaction of rabbit cytochrome P-450 1A2 (P450) with linoleic acid hydroperoxide. This system was compared with chemical systems where FeSO4 or FeCl3 was used in place of P450. The spin trap 5, 5'-dimethyl-1-pyrroline N-oxide (DMPO) was employed to detect and identify radical species. The DMPO adducts of hydroxyl, O2-., peroxyl, methyl and acyl radicals were detected in the P450 system. The reaction did not require NADPH-cytochrome P-450 reductase or NADPH. The same DMPO-radical adducts were detected in the FeSO4 system. Only DMPO-.OH radical adduct and carbon-centred radical adducts were detected in the FeCl3 system. Peroxyl radical production was completely O2-dependent. We propose that polyunsaturated fatty acids are initially reduced to form alkoxyl radicals, which then undergo intramolecular rearrangement to form epoxyalkyl radicals. Each epoxyalkyl radical reacts with O2, forming a peroxyl radical. Subsequent unimolecular decomposition of this peroxyl radical eliminates O2-. radical.

| S-EPMC1218956 | biostudies-other

Benoxacor is enantioselectively metabolized by rat liver subcellular fractions.

Project description:This study investigated the enantioselective metabolism of benoxacor, an ingredient of herbicide formulations, in microsomes or cytosol prepared from female or male rat livers. Benoxacor was incubated for ≤30 min with microsomes or cytosol, and its enantioselective depletion was measured using gas chromatographic methods. Benoxacor was depleted in incubations with active microsomes in the presence and absence of NADPH, suggesting its metabolism by hepatic cytochrome P450 enzymes (CYPs) and microsomal carboxylesterases (CESs). Benoxacor was depleted in cytosolic incubations in the presence of glutathione, consistent with its metabolism by glutathione S-transferases (GSTs). The depletion of benoxacor was faster in incubations with cytosol from male than female rats, whereas no statistically significant sex differences were observed in microsomal incubations. The consumption of benoxacor was inhibited by the CYP inhibitor 1-aminobenzotriazole, the CES inhibitor benzil, and the GST inhibitor ethacrynic acid. Estimates of the intrinsic clearance of benoxacor suggest that CYPs are the primary metabolic enzyme responsible for benoxacor metabolism in rats. Microsomal incubations showed an enrichment of the first eluting benoxacor enantiomer (E1-benoxacor). A greater enrichment occurred in incubations with microsomes from female (EF = 0.67 ± 0.01) than male rats (EF = 0.60 ± 0.01). Cytosolic incubations from female rats resulted in enrichment of E1-benoxacor (EF = 0.54 ± 0.01), while cytosolic incubations from male rats displayed enrichment of the second eluting enantiomer (E2-benoxacor; EF = 0.43 ± 0.01). Sex-dependent differences in the metabolism of benoxacor in rats could significantly impact ecological risks and mammalian toxicity. Moreover, changes in the enantiomeric enrichment of benoxacor may be a powerful tool for environmental fate and transport studies.

| S-EPMC7556730 | biostudies-literature

Cyclic metabolic pathway of a butylated hydroxytoluene by rat liver microsomal fractions.

Project description:A cyclic metabolic pathway was obtained when 3,5-di-t-butyl-4-hydroxytoluene (BHT) was incubated with a rat liver microsomal preparation. The pathway is as follows: BHT --> 4-hydroperoxy-4-methyl-2,6-di-t-butylcyclohexa-2,5-dienone (BHT-OOH) --> 4-hydroxy-4-methyl-2,6-di-t-butylcyclohexa-2,5-dienone (BHT-3(0)OH) --> BHT. This metabolic pathway suggests that antioxidants such as BHT owe their high efficacy, at least in part, to their metabolic regeneration.

| S-EPMC1168527 | biostudies-other

Biochemical and cytochemical localization of inosine-5'-diphosphatase in rat liver microsomal fractions.

Project description:A procedure has been developed for the biochemical and cytochemical localization of inosine-5'-diphosphatase (IDPase) in rat liver microsomal fractions. The concentration of Pb2+ ions used in the cytochemical incubation medium inhibited IDPase by 32%. IDP formed during the incubation was not hydrolysed further to inosine, and no significant non-enzymic hydrolysis of substrate occurred. When microsomal fractions were incubated cytochemically for IDPase and separated from unchanged substrate under conditions such that the microsomal membrane was not permeable to EDTA, approx. 80% of the reaction product was solubilized by EDTA. In the electron microscope, lead precipitate was observed on the cytoplasmic side (outside) of a major fraction of the vesicles and on the cisternal side (inside) of others. After extraction with EDTA, the lead precipitate was observed only on the cisternal side. Since it was shown that phosphate is trapped on the same side of the membrane as it is released, it is concluded that IDPase can release phosphate on either the cisternal or the cytoplasmic side of the microsomal membrane.

| S-EPMC1153366 | biostudies-other

Retrospective assessment of rat liver microsomal stability at NCATS: data and QSAR models.

Project description:Hepatic metabolic stability is a key pharmacokinetic parameter in drug discovery. Metabolic stability is usually assessed in microsomal fractions and only the best compounds progress in the drug discovery process. A high-throughput single time point substrate depletion assay in rat liver microsomes (RLM) is employed at the National Center for Advancing Translational Sciences. Between 2012 and 2020, RLM stability data was generated for ~ 24,000 compounds from more than 250 projects that cover a wide range of pharmacological targets and cellular pathways. Although a crucial endpoint, little or no data exists in the public domain. In this study, computational models were developed for predicting RLM stability using different machine learning methods. In addition, a retrospective time-split validation was performed, and local models were built for projects that performed poorly with global models. Further analysis revealed inherent medicinal chemistry knowledge potentially useful to chemists in the pursuit of synthesizing metabolically stable compounds. In addition, we deposited experimental data for ~ 2500 compounds in the PubChem bioassay database (AID: 1508591). The global prediction models are made publicly accessible ( https://opendata.ncats.nih.gov/adme ). This is to the best of our knowledge, the first publicly available RLM prediction model built using high-quality data generated at a single laboratory.

| S-EPMC7693334 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data