首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 31 毫秒
1.
Spin-trapping investigators are largely limited by the instability of the radical adducts. Spin trap 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline N-oxide (DEPMPO) forms very stable alkoxyl radical adducts. However, the presence of two chiral centers in the DEPMPO alkoxyl radical adduct results in two diastereomers with distinctive ESR spectra, which complicates the interpretation of the ESR spectra. We have analyzed the high resolution ESR spectra of the DEPMPO/?OCH3 radical adduct. DEPMPO/?OCH3 has been synthesized by the nucleophilic addition of alcohols to DEPMPO. The electron spin resonance (ESR) spectrum of DEPMPO/?OCH3 in oxygen-free methanol solution reveals superhyperfine structure with hyperfine coupling constants as small as 0.3?G. In order to simplify the analysis of the electron spin resonance (ESR) spectrum, we synthesized the DEPMPO/?OCD3 radical adduct. Computer simulation of the DEPMPO/?OCD3 ESR spectrum revealed two diastereomers. Hyperfine coupling constants of γ-protons and 17O from the –OCH3 group were also determined. ESR spectra of DEPMPO/?OCH3 in phosphate buffer have also been characterized. The presence of specific hyperfine couplings from the –OCH3 group can be used for the unambiguous identification of the DEPMPO/?OCH3 radical adducts. We suggest that the analysis of high resolution ESR spectra can be used for the unambiguous characterization of DEPMPO radical adducts.  相似文献   

2.
It has been previously reported that the spin trap 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline N-oxide (DEPMPO) can form stable radical adducts with superoxide radical. However, the presence of diastereomers of DEPMPO radical adducts and the appearance of superhyperfine structure complicates the interpretation of the ESR spectra. It has been suggested that the superhyperfine structure in the ESR spectrum of DEPMPO/OOH is a result of conformational exchange between conformers. The analysis of the temperature dependence of the ESR spectrum of DEPMPO/OOH and of its structural analog DMPO/OOH have demonstrated that both ESR spectra contain exchange effects resulting from conversion between two conformers. Computer simulation calculates a conformer lifetime on the order of 0.1 μs for DMPO/OOH at room temperature. However, temperature dependence of the ESR spectrum of DEPMPO/OOH suggests that superhyperfine structure does not depend on the conformational exchange. We have now found that the six-line ESR spectrum with superhyperfine structure should be assigned to a DEPMPO-superoxide-derived decomposition product. Therefore, ESR spectra previously assigned to DEPMPO/OOH contain not only the two diastereomers of DEPMPO/OOH but also the decomposition product, and these spectra should be simulated as a combination of four species: two conformers of the first diastereomer, one conformer of the second diastereomer and the superoxide-derived decomposition product. The presence of four species has been supported by the temperature dependence of the ESR spectra, nucleophilic synthesis of radical adducts, and isotopic substitution experiments. It is clear that to correctly interpret DEPMPO spin trapping of superoxide radicals, one must carefully consider formation of secondary radical adducts.  相似文献   

3.
Polyunsaturated fatty acid (PUFA) peroxyl radicals play a crucial role in lipid oxidation. ESR spectroscopy with the spin-trapping technique is one of the most direct methods for radical detection. There are many reports of the detection of PUFA peroxyl radical adducts; however, it has recently been reported that attempted spin trapping of organic peroxyl radicals at room temperature formed only alkoxyl radical adducts in detectable amounts. Therefore, we have reinvestigated spin trapping of the linoleic, arachidonic, and linolenic acid-derived PUFA peroxyl radicals. The slow-flow technique allowed us to obtain well-resolved ESR spectra of PUFA-derived radical adducts in a mixture of soybean lipoxygenase, PUFA, and the spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO). However, interpretation of the ESR spectra was complicated by the overlapping of the PUFA-derived alkoxyl radical adduct spectra. In order to understand these spectra, PUFA-derived alkoxyl radical adducts were modeled by various alkoxyl radical adducts. For the first time, we synthesized a wide range of DMPO adducts with primary and secondary alkoxyl radicals. It was found that many ESR spectra previously assigned as DMPO/peroxyl radical adducts based on their close similarity to the ESR spectrum of the DMPO/superoxide radical adduct, in conjunction with their insensitivity to superoxide dismutase, are indeed alkoxyl radical adducts. We have reassigned the PUFA alkylperoxyl radical adducts to their corresponding alkoxyl radical adducts. Using hyperfine coupling constants of model DMPO/alkoxyl radical adducts, the computer simulation of DMPO/PUFA alkoxyl radical adducts was performed. It was found that the trapped, oxygen-centered PUFA-derived radical is a secondary, chiral alkoxyl radical. The presence of a chiral carbon atom leads to the formation of two diastereomers of the DMPO/PUFA alkoxyl radical adduct. Therefore, attempted spin trapping of the PUFA peroxyl radical by DMPO at room temperature leads to the formation of the PUFA alkoxyl radical adduct.  相似文献   

4.
It has been previously reported that the spin trap 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline N-oxide (DEPMPO) can form stable radical adducts with superoxide radical. However, the presence of diastereomers of DEPMPO radical adducts and the appearance of superhyperfine structure complicates the interpretation of the ESR spectra. It has been suggested that the superhyperfine structure in the ESR spectrum of DEPMPO/?OOH is a result of conformational exchange between conformers. The analysis of the temperature dependence of the ESR spectrum of DEPMPO/?OOH and of its structural analog DMPO/?OOH have demonstrated that both ESR spectra contain exchange effects resulting from conversion between two conformers. Computer simulation calculates a conformer lifetime on the order of 0.1?μs for DMPO/?OOH at room temperature. However, temperature dependence of the ESR spectrum of DEPMPO/?OOH suggests that superhyperfine structure does not depend on the conformational exchange. We have now found that the six-line ESR spectrum with superhyperfine structure should be assigned to a DEPMPO-superoxide-derived decomposition product. Therefore, ESR spectra previously assigned to DEPMPO/?OOH contain not only the two diastereomers of DEPMPO/?OOH but also the decomposition product, and these spectra should be simulated as a combination of four species: two conformers of the first diastereomer, one conformer of the second diastereomer and the superoxide-derived decomposition product. The presence of four species has been supported by the temperature dependence of the ESR spectra, nucleophilic synthesis of radical adducts, and isotopic substitution experiments. It is clear that to correctly interpret DEPMPO spin trapping of superoxide radicals, one must carefully consider formation of secondary radical adducts.  相似文献   

5.
It has been proposed that the C-phenyl-N-tert-butylnitrone/trichloromethyl radical adduct (PBN/CCl3) is metabolized to either the C-phenyl-N-tert-butylnitrone/carbon dioxide anion radical adduct (PBN/CO2) or the glutathione (GSH) and CCl4-dependent PBN radical adduct (PBN/[GSH-CCl3]). Inclusion of PBN/CCl3 in microsomal incubations containing GSH, nicotinamide adenine dinucleotide phosphate (NADPH), or GSH plus NADPH produced no electron spin resonance (ESR) spectral data indicative of the formation of either the PBN/[GSH-CCl3] or PBN/CO2 radical adducts. Microsomes alone or with GSH had no effect on the PBN/CCl3 radical adduct. Addition of NADPH to a microsomal system containing PBN/CCl3 presumably reduced the radical adduct to its ESR-silent hydroxylamine because no ESR signal was observed. The Folch extract of this system produced an ESR spectrum that was a composite of two radicals, one of which had hyperfine coupling constants identical to those of PBN/CCl3. We conclude that PBN/CCl3 is not metabolized into either PBN/[GSH-CCl3] or PBN/CO2 in microsomal systems.  相似文献   

6.
Lipid radicals: properties and detection by spin trapping   总被引:1,自引:0,他引:1  
Unsaturated lipids are rapidly oxidized to toxic products such as lipid hydroperoxides, especially when transition metals such as iron or copper are present. In a Fenton-type reaction Fe2+ converts lipid hydroperoxides to the very short-lived lipid alkoxyl radicals. The reaction was started upon the addition of Fe2+ to an aqueous linoleic acid hydroperoxide (LOOH) emulsion and the spin trap in the absence of oxygen. Even when high concentrations of spin traps were added to the incubation mixture, only secondary radical adducts were detected, probably due to the rapid re-arrangement of the primary alkoxyl radicals. With the commercially available nitroso spin trap MNP we observed a slightly immobilized ESR spectrum with only one hydrogen splitting, indicating the trapping of a methinyl fragment of a lipid radical. With DMPO or 5-diethoxyphosphoryl-5-methyl-1-pyrroline N-oxide (DEPMPO) adducts were detected with carbon-centered lipid radical, with acyl radical, and with the hydroxyl radical. We also synthesized lipophilic derivatives of the spin trap DEPMPO in order to detect lipid radical species generated in the lipid phase. With all spin traps studied a lipid-derived carbon-centered radical was obtained in the anaerobic incubation system Fe2+/LOOH indicating the trapping of a lipid radical, possibly generated as a secondary reaction product of the primary lipid alkoxyl radical formed. Under aerobic conditions an SOD-insensitive oxygen-centered radical adduct was formed with DEPMPO and its lipophilic derivatives. The observed ESR parameters were similar to those of alkoxyl radical adducts, which were independently synthesized in model experiments using Fe3+-catalyzed nucleophilic addition of methanol or t-butanol to the respective spin trap.  相似文献   

7.
The study of the important role of peroxyl radicals in biological systems is limited by their difficult detection with direct electron spin resonance (ESR). Many ESR spectra were assigned to 5,5-dimethyl-1-pyrroline N-oxide (DMPO)/peroxyl radical adducts based only on the close similarity of their ESR spectra to that of DMPO/superoxide radical adduct in conjunction with their insensitivity to superoxide dismutase, which distinguishes the radical adduct from DMPO/superoxide radical adduct. Later, the spin-trapping literature reported that DMPO/peroxyl radical adducts have virtually the same hyperfine coupling constants as synthesized alkoxyl radical adducts, raising the issue of the correct assignment of peroxyl radical adducts. However, using 17O-isotope labelling, the methylperoxyl and methoxyl radical adducts should be distinguishable. We have reinvestigated the spin trapping of the methylperoxyl radical. The methylperoxyl radical was generated in aerobic solution with 17O-molecular oxygen either in a Fenton system with dimethylsulfoxide or in a chloroperoxidase system with tert-butyl hydroperoxide. Two different spin traps, DMPO and 2,2,4-trimethyl-2H-imidazole-1-oxide (TMIO), were used to trap methylperoxyl radical. 17O-labelled methanol was used to synthesize methoxyl radical adducts by nucleophylic addition. It was shown that the 17O hyperfine coupling constants of radical adducts formed in methylperoxyl radical-generating systems are identical to that of the methoxyl radical adduct. Therefore, methylperoxyl radical-producing systems form detectable methoxyl radical adduct, but not detectable methylperoxyl radical adducts at room temperature. One of the possible mechanisms is the decomposition of peroxyl radical adduct with the formation of secondary alkoxyl radical adduct. These results allow us to reinterpret previously published data reporting detection of peroxyl radical adducts. We suggest that detection of 17O-alkoxyl radical adduct from 17O-labelled molecular oxygen can be used as indirect evidence for peroxyl radical generation.  相似文献   

8.
The ability of horseradish peroxidase (E.C. 1.11.1.7. Donor: H2O2 oxidoreductase) to catalytically oxidize 2-deoxyribose sugars to a free radical species was investigated. The ESR spin-trapping technique was used to denionstrate that free radical species were formed. Results with the spin trap 3.5-dibronio-4-nitrosoben-zene sulphonic acid showed that horseradish peroxidase can catalyse the oxidation of 2-deoxyribose to produce an ESR spectrum characteristic of a nitroxide radical spectrum. This spectrum was shown to be a composite of spin adducts resulting from two carbon-centered species, one spin adduct being characterized by the hyperfine coupling constants aN = 13.6GandaHβ = 11.0G, and the other by aN = 13.4G and aH = 5.8 G. When 2-deoxyribose-5-phosphate was used as the substrate, the spectrum produced was found to be primarily one species characterized by the hyperfine coupling constants aN = 13.4G and aH= 5.2. All the radical species produced were carbon-centered spin adducts with a β hydrogen, suggesting that oxidation occurred at the C(2) or C(5) moiety of the sugar. Interestingly, it was found that under the same experimental conditions, horseradish peroxidase apparently did not catalyze the oxidation of either 3-deoxyribose or D-ribose to a free radical since no spin adducts were found in these cases.

It can be readily seen that 2-deoxyribose and 2-deoxyribose-5-phosphate can be oxidized by HRP/H2O2 to form a free radical species that can be detected with the ESR spin-trapping technique. There are two probable sites for the formation of a CH type radical on the 2-deoxyribose sugar, these being the C(2) and the C(5) carbons. The fact that there is a species produced from 2-deoxy-ribose, but not 2-deoxy-ribose-5-phosphate, suggests that there is an involvement of the C(5) carbon in the species with the 1 1.0G β hydrogen. In the spectra formed from 2-deoxy-ribose, there is a big difference in the hyperfine splitting of the β hydrogens, suggesting that the radicals are formed at different carbon centers, while the addition of a phosphate group to the C(5) carbon seems to inhibit radical formation at one site. In related work, the chemiluminescence of monosaccharides in the presence of horseradish peroxidase was proposed to be the consequence of carbon-centered free radical formation (10).  相似文献   

9.
Photosensitizers newly developed for photodynamic therapy of cancer need to be assessed using accurate methods of measuring reactive oxygen species (ROS). Little is known about the characteristics of the reaction of singlet oxygen (1O2) with spin traps, although this knowledge is necessary in electron spin resonance (ESR)/spin trapping. In the present study, we examined the effect of various reductants usually present in biological samples on the reaction of 1O2 with 5,5-dimethyl-1-pyrroline-N-oxide (DMPO). The ESR signal of the hydroxyl radical (•OH) adduct of DMPO (DMPO-OH) resulting from 1O2-dependent generation of •OH strengthened remarkably in the presence of reduced glutathione (GSH), 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox), ascorbic acid, NADPH, etc. A similar increase was observed in the photosensitization of uroporphyrin (UP), rose bengal (RB) or methylene blue (MB). Use of 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide (DEPMPO) as a spin trap significantly lessened the production of its •OH adduct (DEPMPO-OH) in the presence of the reductants. The addition of DMPO to the DEPMPO-spin trapping system remarkably increased the signal intensity of DEPMPO-OH. DMPO-mediated generation of •OH was also confirmed utilizing the hydroxylation of salicylic acid (SA). These results suggest that biological reductants enhance the ESR signal of DMPO-OH produced by DMPO-mediated generation of •OH from 1O2, and that spin trap-mediated •OH generation hardly occurs with DEPMPO.  相似文献   

10.
We applied a spin trap, 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline-N-oxide (DEPMPO), to detect O2·- generation during phagocytosis in human polymorphonuclear leukocytes (PMNs). PMNs were activated with serum-opsonized zymosan (sOZ) in the presence of DEPMPO. The ESR spectra mainly consisted of Cu,Zn-SOD-sensitive DEPMPO-OOH spin adducts. To clarify where these spin-adducts were present, cells after stimulation were separated from extracellular fluid by brief centrifugation and resuspended in Hanks' balanced salt solution. ESR examination showed that DEPMPO-OOH adducts were present in both fractions. When cells were stimulated by phorbol myristate acetate (PMA), the DEPMPO-OOH was detected in extracellular fluid but not in the cell fraction. Furthermore, DEPMPO-OOH adducts were quickly converted into ESR-silent compounds by addition of cell lysate of PMNs. These results indicate that DEPMPO is useful to detect O2·- of extracellular space including the intraphagosome but not that of intracellular space in sOZ-stimulated phagocytes.  相似文献   

11.
Rebamipide, an antiulcer agent, is known as a potent hydroxyl radical (OH) scavenger. In the present study, we further characterized the scavenging effect of rebamipide against OH generated by ultraviolet (UV) irradiation of hydrogen peroxide (H2O2), and identified the reaction products to elucidate the mechanism of the reaction. Scavenging effect of rebamipide was accessed by ESR using DMPO as a OH-trapping agent after UVB exposure (305 nm) to H2O2 for 1 min in the presence of rebamipide. The signal intensity of OH adduct of DMPO (DMPO-OH) was markedly reduced by rebamipide in a concentration-dependent fashion as well as by dimethyl sulfoxide and glutathione as reference radical scavengers. Their second order rate constant values were 5.62 × 1010, 8.16 × 109 and 1.65 × 1010 M-1 s-1, respectively. As the rebamipide absorption spectrum disappeared during the reaction, a new spectrum grew due to generation of rather specific reaction product. The reaction product was characterized by LC-MS/MS and NMR measurements. Finally, a hydroxylated rebamipide at the 3-position of the 2(1H)-quinolinone nucleus was newly identified as the major product exclusively formed in the reaction between rebamipide and the OH generated by UVB/H2O2. Specific formation of this product explained the molecular characteristics of rebamipide as a potential OH scavenger.  相似文献   

12.
The spin trap 5-(diethoxyphosphoryl)-5-methyl-1-pyrroline N-oxide (DEPMPO) forms a superoxide adduct with a half-life of almost 15 min. DEPMPO is very hydrophilic and its use for the detection of radicals in the lipid phase (lipid-derived radicals and superoxide generated in the lipid phase) is therefore limited due to its very low concentration in the lipid phase. For the detection of lipid-derived radicals, three derivatives of DEPMPO with increasing degree of lipid solubility have been investigated: 5-(di-n-propoxyphosphoryl)-5-methyl-1-pyrroline N-oxide (DPPMPO), 5-(di-n-butoxyphosphoryl)-5-methyl-1-pyrroline N-oxide (DBPMPO), and 5-(bis-(2-ethylhexyloxy)phosphoryl)-5-methyl-1-pyrroline N-oxide (DEHPMPO). As compared with the spin trap DMPO, the half-lives of the respective superoxide adducts were clearly higher in aqueous solutions of the spin traps, which facilitates qualitative ESR measurements. The stability of the superoxide spin adducts formed with the various lipophilic spin traps in aqueous buffer were similar to those observed with DEPMPO (half-life: 7-11 min.). In model experiments using Fe(3+)-catalyzed nucleophilic addition of methanol or tert-butanol to the respective spin trap the respective alkoxyl radical adducts were formed in aqueous solution as transient species in the presence of high concentrations of the alcohol. Upon dilution with water the alkoxyl group was substituted by water, giving the respective hydroxyl adduct of the spin trap. Care must therefore be taken when Fenton-type reactions are used for the generation of radicals such as the use of Fe(2+) complexes with phosphate or DTPA or inactivation of iron by addition of "Desferal" (Novarti's Pharma GmbH, Vienna, Austria) after a short incubation time. Addition of Fe(2+) under anaerobic conditions to an aqueous suspension of linoleic acid hydroperoxide and the spin trap resulted in the detection of three different species: a carbon-centered radical adduct, an acyl radical adduct, and the hydroxyl adduct. In the presence of oxygen a different species was observed with DEPMPO, DPPMPO, and DBPMPO, which was only slightly suppressed upon the addition of SOD, possibly the respective spin adduct of either the alkylperoxyl radical or, in analogy to DMPO, a secondary alkoxyl radical.  相似文献   

13.
Studies documenting spin trapping of lipid radicals in defined model systems have shown some surprising solvent effects with the spin trap DMPO. In aqueous reactions comparing the reduction of H2O2 and methyl linoleate hydroperoxide (MLOOH) by Fez+, hydroxyl (HO·) and lipid alkoxyl (LO·) radicals produce identical four-line spectra with line intensities 1:2:2:1. Both types of radicals react with commonly-used HO· scavengers, e.g. with ethanol to produce ·C(CH3)HOH and with dirnethylsulfoxide (DMSO)togive ·CH3. However, DMSO radicals (either ·CH3or ·OOCH3) react further with lipids, and when radicals are trapped in these MLOOH systems, multiple adducts are evident. When acetonitrile is added to the aqueous reaction systems in increasing concentrations, ·CH2CN radicals resulting from HO· attack on acetonitrile are evident, even with trace quantities of that solvent. In contrast, little, if any, reaction of LO· with acetonitrile occurs, even in 100% acetonitrile. A single four-line signal persists in the lipid systems as long as any water is present, although the relative intensity of the two center lines decreases as solvent-induced changes gradually dissociate the nitrogen and β-hydrogen splitting constants. Extraction of the aqueous-phase adducts into ethyl acetate shows clearly that the identical four-line spectra in the H202 and MLOOH systems arise from different radical species in this study, but the lack of stability of the adducts to phase transfer may limit the use of this technique for routine adduct identification in more complex systems. These results indicate that the four-line 1:2:2:1. aN = aH = 14.9G spectrum from DMPO cannot automatically be assigned to the HO· adduct in reaction systems where lipid is present, even when the expected spin adducts from ethanol or DMSO appear confirmatory for HO-. Conclusive distinction between HO· and LO· ultimately will require use of 13C-labelled DMPO or HPLC-MS separation and specific identification of adducts when DMPO is used as the spin trap.  相似文献   

14.
Short-lived free radicals formed in the reaction of 11 substrates and radiolytically produced hydroxyl radicals were trapped successfully with 5, 5-dimethyl-1-pyrroline-N-oxide (DMPO) in dilute aqueous solution. The in situ radiolysis steady-state ESR spectra of the spin adducts were analyzed to determine accurate ESR parameters for these spin adducts in a uniform environment. Parent alkyl radicals include methyl, ethyl, 1-propyl and 2-propyl (1-methylethyl). Hydroxyalkyl parent radicals were hydroxymethyl, hydroxyethyl, 2-hydroxy-2-propyl (1-methyl-1-hydroxyethyl), 1-hydroxypropyl and 2-hydroxy-2-methylpropyl. Carboxyl radical (carbon dioxide anion, formate radical) and sulfite anion radical were the sigma radicals studied. The DMPO spin adduct of 1-propyl was identified for the first time. For most spin adducts, g factors were also determined for the first time. In DMPO spin adducts of hydroxyalkyl radicals, nitrogen and C(2)-proton hyperfine coupling constants are smaller than those of alkyl radical adducts; the hydroxyalkyl spin adducts possess larger g values than their unsubstituted counterparts. These changes are ascribed to the spread of pi conjugation to include the hydroxyl group. Strong evidence of spin addend-aminoxyl group interaction can be seen in the asymmetrical line shapes in the hydroxyethyl and the hydroxypropyl spin adducts.  相似文献   

15.
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.  相似文献   

16.
The spin traps substituted with some groups at the 4-position of dimethyl-1-pyrroline N-oxide(DMPO) were compared with DMPO itself regarding their abilities as spin traps and their physical properties. 4,5,5-Trimethyl-l-pyroHine N-oxide (4MDMPO) and 5,5-dimethyl- 4-phenyl-l-pyrolline N-oxide (4PDMPO) were synthesized by the Bonnett method, and 5,5-dimethyl-4-hydroxymethyl-l-pyrolline N-oxide (4HMDMPO) was made by a unique method from 2(5H)-furanone. The melting points of 4MDMPO, 4PDMPO and 4HMDMPO were higher than that of DMPO. The magnitude of hydrophilicity was in the order of 4HMDMPO, DMPO, 4MDMPO, and 4PDMPO based on the partition coefficient experiments in a 1-octanol-water system. Several radicals, O2, HO-, -CH3, -CH2OH, -CH(CH3)OH, (CH3)3 CO and H radicals, were trapped with these DMPO derivatives for comparison with the trapping by DMPO itself. Spin adducts of O J with the three DMPO derivatives showed ESR spectra similar to that of DMPO. In spite of the formation of diastereomers arising from spin trapping, the line-width enlargement was very small. The intensities and the decay rates of the spectra of 4MDMPO-O2-, 4PDMPO-O2- 4HMDMPO-O2- and DMPO-O2- were almost equal. In the trapping of the OH radical by 4MDMPO, 4PDMPO and 4HMDMPO, the eight-line ESR spectra observed were different from the well-known four-line spectrum of DMPO-OH.  相似文献   

17.
The generation of free radicals in a superoxide (O2-)driven Fe+3 catalysed reactions with isolated myocytic sarcolemma using electron spin resonance was investigated. Incubation of highly purified canine myocytic sarcolemma in the presence of the spin trap, 2-methyl-2-nitrosopropane (MNP). followed by the addition of dihydroxyfurmarate (DHF) and Fe+3-ADP resulted in the generation and detection of radical adducts of this spin trap. Spin trapping of the alkyl radicals with 2-methyl-2-nitrosopropane led to the identification of methyl radical adduct following exposure to DHF/Fe+3-ADP. With sarcolemma and the alkyl nitroso compound, the only radical product trapped was the methyl radical formed by β-scission of alkoxyl radical. The participation of hydroperoxide-derived radicals in this system verified that the decomposition of unsaturated hydroperoxy fatty acid does proceed via a free radical mechanism.  相似文献   

18.
Electron paramagnetic resonance (EPR) spin trapping was used to detect lipid-derived free radicals generated by iron-induced oxidative stress in intact cells. Using the spin trap alpha-(4-pyridyl 1-oxide)-N-tert-butylnitrone (POBN), carbon-centered radical adducts were detected. These lipid-derived free radicals were formed during incubation of ferrous iron with U937 cells that were enriched with docosahexaenoic acid (22:6n-3). The EPR spectra exhibited apparent hyperfine splittings characteristic of a POBN/alkyl radical, aN = 15.63 +/- 0.06 G and aH = 2.66 +/- 0.03 G, generated as a result of beta-scission of alkoxyl radicals. Spin adduct formation depended on the FeSO4 content of the incubation medium and the number of 22:6-enriched cells present; when the cells were enriched with oleic acid (18:1n-9), spin adducts were not detected. This is the first direct demonstration, using EPR, of a lipid-derived radical formed in intact cells in response to oxidant stress.  相似文献   

19.
5-(2,2-Dimethyl-1,3-propoxy cyclophosphoryl)-5-methyl-1-pyrroline N-oxide (CYPMPO), a new cyclic DEPMPO-type nitrone was evaluated for spin-trapping capabilities toward hydroxyl and superoxide radicals. CYPMPO is colorless crystalline and freely soluble in water. Both the solid and diluted aqueous solution did not develop electron spin resonance (ESR) signal for at least 1 month at ambient conditions. CYPMPO can spin-trap superoxide and hydroxyl radicals in both chemical and biological systems, and the ESR spectra are readily assignable. Half life for the superoxide adduct of CYPMPO produced in UV-illuminated hydrogen peroxide solution was approximately 15 min, and in biological systems such as hypoxanthine (HX)/xanthine oxidase (XOD) the half-life of the superoxide adduct was approximately 50 min. In UV-illuminated hydrogen peroxide solution, there was no conversion from the superoxide adduct to the hydroxyl adduct. Although overall spin-trapping capabilities of CYPMPO are similar to DEPMPO, its high melting point, low hygroscopic property, and the long shelf-life would be highly advantageous for the practical use.  相似文献   

20.
To spin trap hydroxyl radical (HO*) with in vivo detection of the resultant radical adducts, the use of two spin traps, 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) and 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide (DEPMPO) (10 mmol/kg) has been compared. In mice treatment with 5-aminolevulinic acid and Fe3+ resulted in detection of adducts of hydroxyl radicals (HO*), but only with use of DEPMPO. Similarly, 'HO* adducts' generated via nucleophilic substitution of SO4*- adducts formed in vivo could be observed only when using DEPMPO as the spin trap. The reasons for the differences observed between DEPMPO and DMPO are likely due to different in vivo lifetimes of their hydroxyl radical adducts. These results seem to be the first direct in vivo EPR detection of hydroxyl radical adducts.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号