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Showing papers on "Xanthine published in 1998"


Journal ArticleDOI
TL;DR: These phenolic compounds were shown to be effective to protect biological systems against various oxidative stresses and protected red cells against oxidative hemolysis.

233 citations


Journal ArticleDOI
TL;DR: It is suggested that oxygen free radicals contribute to ionic, high-osmolality contrast medium nephrotoxicity in hypomagnesemic patients with mild renal disease and that magnesium attenuates the neph rotoxicity mediated by oxygenFree radicals.

219 citations


Journal ArticleDOI
TL;DR: XO decomposes RSNO by O·̄2-dependent and -independent pathways, and in the presence of oxygen it leads to peroxynitrite formation, and it is found that CysNO is an electron acceptor substrate for XO with aK m of 0.7 mm.

195 citations


Journal ArticleDOI
TL;DR: Results indicate that the time window of ischemia capable to induce preconditioning in liver is defined by the relative tissue concentrations of adenosine and xanthine.

125 citations


Journal ArticleDOI
TL;DR: The ribonuclease sensitivities of some internal-loop residues changed upon the addition of xanthine, suggesting that the internal loop of the XBA RNA is involved in the ligand binding.
Abstract: RNAs that bind to xanthine (2,6-dioxypurine) were isolated from a population of 10(12) random sequences by in vitro selection. These xanthine-binding RNAs were found to have a 10 nt consensus sequence at an internal loop in the most probable secondary structure. By trimming one of the xanthine-binding RNAs, a representative xanthine-binding RNA (designated as XBA) of 32 nt residues was prepared. The dissociation constant of this RNA for xanthine was determined to be 3.3 microM by equilibrium filtration experiments. The XBA RNA can bind to guanine as well, whereas it hardly accommodates adenine, cytosine or uracil. The K d values for various xanthine/guanine analogues were determined, and revealed that the N1H, N7 and O6 moieties of the ligand are involved in the binding with the XBA RNA. The ribonuclease sensitivities of some internal-loop residues changed upon the addition of xanthine, suggesting that the internal loop of the XBA RNA is involved in the ligand binding. Interestingly, the consensus sequence of the xanthine/guanine-binding RNAs is the same as a sequence in one of the internal loops of the hairpin ribozyme, except for a substitution that is neutral with respect to xanthine/guanine binding.

107 citations


Journal ArticleDOI
TL;DR: It is concluded that both human xanthine dehydrogenase (XD) and XO can oxidise NADH to generate ROS, and the conversion of XD to XO is not necessary for post-ischaemic ROS generation.
Abstract: Xanthine oxidase (XO) is conventionally known as a generator of reactive oxygen species (ROS) which contribute to hypoxic-reperfusion injury in tissues. However, this role for human XO is disputed due to its distinctive lack of activity towards xanthine, and the failure of allopurinol to suppress reperfusion injury. In this paper, we have employed native gel electrophoresis together with activity staining to investigate the role human xanthine dehydrogenase (XD) and XO in hypoxic reperfusion injury. This approach has provided information which cannot be obtained by conventional spectrophotometric assays. We found that both XD and XO of human umbilical vein endothelial cells (HUVECs) and lymphoblastic leukaemic cells (CEMs) catalysed ROS generation by oxidising NADH, but not hypoxanthine. The conversion of XD to XO was observed in both HUVECs and CEMs in response to hypoxia, although the level of conversion varied. Purified human milk XD generated ROS more efficiently in the presence of NADH than in the presence of hypoxanthine. This NADH oxidising activity was blocked by the FAD site inhibitor, diphenyleneiodonium (DPI), but was not suppressible by the molybdenum site inhibitor, allopurinol. However, in the presence of both DPI and allopurinol the activities of XD/XO were completely blocked with either NADH or hypoxanthine as substrates. We conclude that both human XD and XO can oxidise NADH to generate ROS. Therefore, the conversion of XD to XO is not necessary for post-ischaemic ROS generation. The hypoxic-reperfusion injury hypothesis should be reappraised to take into account the important role played by XD and XO in oxidising NADH to yield ROS.

104 citations


Journal ArticleDOI
TL;DR: The possibility that T-4,5-D and perhaps other putative intraneuronal metabolites formed by the O2*-/H2O2/oxo-iron-mediated oxidations of 5-HT might be endotoxins that contribute to neurodegeneration in brain regions innervated by serotonergic neurons caused by MA, ischemia-reperfusion, and other neurodegnerative brain disorders is raised.
Abstract: Many new lines of evidence implicate both superoxide anion radical (O2*-) and biogenic amine neurotransmitters in the pathological mechanisms that underlie neuronal damage caused by methamphetamine (MA), glutamate-mediated oxidative toxicity, ischemia-reperfusion, and other neurodegenerative brain disorders In this investigation the oxidation of 5-hydroxytryptamine (5-HT, serotonin) by an O2*--generating system (xanthine/xanthine oxidase) in buffered aqueous solution at pH 74 has been studied The major product of the O2*--mediated oxidation of 5-HT is tryptamine-4,5-dione (T-4, 5-D) However, O2*- and H2O2, cogenerated by the xanthine oxidase-mediated oxidation of xanthine to uric acid, together react with trace levels of iron that contaminate buffer constituents to give a chemically ill-defined oxo-iron species This species mediates the oxidation of 5-HT to a C(4)-centered carbocation intermediate that reacts with 5-HT to give 5,5'-dihydroxy-4, 4'-bitryptamine (4,4'-D) and with uric acid to give 9-[3-(2-aminoethyl)-5-hydroxy-1H-indol-4-yl]-2,6,8-triketo-1H,3H, 7H-purine (7) as the major products These products differ from those formed in the HO*-mediated oxidation of 5-HT under similar conditions When the reaction is carried out in the presence of the intraneuronal nucleophile glutathione (GSH), T-4,5-D is scavenged to give 7-(S-glutathionyl)tryptamine-4,5-dione, whereas the putative carbocation intermediate is scavenged to give 4-(S-glutathionyl)-5-hydroxytryptamine T-4,5-D also reacts with the sulfhydryl residues of a model protein, alcohol dehydrogenase, and inhibits its activity Previous investigators have proposed that T-4, 5-D is a serotonergic neurotoxin This raises the possibility that T-4,5-D and perhaps other putative intraneuronal metabolites formed by the O2*-/H2O2/oxo-iron-mediated oxidations of 5-HT might be endotoxins that contribute to neurodegeneration in brain regions innervated by serotonergic neurons caused by MA, ischemia-reperfusion, and other neurodegenerative brain disorders

100 citations


Journal ArticleDOI
TL;DR: Results suggests that sufficient substrate exists in plasma to produce micromolar levels of hydrogen peroxide and xanthine oxidase may catalyze these reactions.

99 citations


Journal ArticleDOI
TL;DR: Fourteen Rhodobacter capsulatus mutants unable to grow with xanthine as sole nitrogen source were isolated by random Tn5 mutagenesis and revealed a higher degree of similarity to eukaryotic x anthine dehydrogenases than to the xanthin dehydrogenase‐related aldehyde oxidoreductase from Desulphovibrio gigas.
Abstract: Fourteen Rhodobacter capsulatus mutants unable to grow with xanthine as sole nitrogen source were isolated by random Tn5 mutagenesis. Five of these Tn5 insertions were mapped within two adjacent chromosomal EcoRI fragments hybridizing to oligonucleotides synthesized according to conserved amino acid sequences of eukaryotic xanthine dehydrogenases. DNA sequence analysis of this region revealed two open reading frames, designated xdhA and xdhB, encoding xanthine dehydrogenase. The deduced amino acid sequence of XDHA contains binding sites for two [2Fe-2S] clusters and FAD, whereas XDHB is predicted to contain the molybdopterin cofactor. In contrast to R. capsulatus, these three cofactor binding sites reside within a single polypeptide chain in eukaryotic xanthine dehydrogenases. The amino acid sequence of xanthine dehydrogenase from R. capsulatus showed a higher degree of similarity to eukaryotic xanthine dehydrogenases than to the xanthine dehydrogenase-related aldehyde oxidoreductase from Desulphovibrio gigas. The expression of an xdhA-lacZ fusion was induced when hypoxanthine or xanthine was added as sole nitrogen source. Mutations in nlfR1 (ntrC) and nifR4 (rpoN, encoding σ) had no influence on xdh gene expression. A putative activator sensing the availability of substrate seems to respond to xanthine but not to hypoxanthine. The transcriptional start site of xdhA was mapped by primer extension analysis. Comparison with known promoter elements revealed no significant homology. Xanthine dehydrogenase from R. capsulatus was purified to homogeneity. The enzyme consists of two subunits with molecular masses of 85 kDa and 50 kDa respectively. N-terminal amino acid sequencing of both subunits confirmed the predicted start codons. The molecular mass of the native enzyme was determined to be 275 kDa, indicating an αβ-subunit structure. Analysis of the molybdenum cofactor of xanthine dehydrogenase from R. capsulatus revealed that it contains the molybdopterin cofactor and not a molybdopterin dinucleotide derivative.

96 citations


Journal ArticleDOI
TL;DR: Uric acid at concentrations similar to its physiological levels, and also related analogues are able to suppress oxidative degradation of LDL components, in view of the various mechanisms underlying atherogenesis in vivo.

91 citations


Journal ArticleDOI
TL;DR: Substantially slower reaction rates of superoxide with S-nitrosothiols relative to the reaction rate with NO are consistent with the contention that the transient formation ofS-nitrosonium equivalent in biological systems may protect NO from its rapid destruction by superoxide, thus enabling these compounds to serve as carriers or buffers of NO.

Journal ArticleDOI
TL;DR: The results show that ascorbic acid is a poor inhibitor of nitration or deamination under acidic conditions such as those of the stomach, and the ability of plant phenolics to scavenge reactive nitrogen species derived from acidic nitrite may contribute to the protective effects of tea polyphenols against gastric cancer.
Abstract: Exposure of tyrosine or DNA bases to acidic nitrite at low pH results in the nitration of tyrosine and the formation of base deamination products, respectively. At pH 1, hypoxanthine and xanthine are formed from the deamination of adenine and guanine, respectively, whereas under the same conditions, uracil is not detected. The yield of 3-nitrotyrosine derived from interaction of equimolar nitrite and tyrosine at pH 1 is approximately 50% of that obtained from equimolar peroxynitrite-tyrosine interactions at pH 7. 4. The ability of a range of plant phenolic constituents to prevent damage mediated by acidic nitrite was also examined in comparison with the activity of vitamin C. The epicatechin/gallate family of flavonols, constituents of green tea, red wine, etc., demonstrates the most extensive inhibitory properties against both tyrosine nitration and base deamination. The results also show that ascorbic acid is a poor inhibitor of nitration or deamination under acidic conditions such as those of the stomach. The ability of plant phenolics to scavenge reactive nitrogen species derived from acidic nitrite may contribute to the protective effects of tea polyphenols against gastric cancer.

Journal ArticleDOI
TL;DR: Recently, Sanofi Research isolated a recombinant urate oxidase (SR29142) as a cDNA clone from Aspergillus flavus, expressed in the yeast strain Saccharomyces cerevisiae, and is undergoing clinical studies to demonstrate its efficacy and safety in this population of patients at highest risk of developing ATLS.
Abstract: Acute tumour lysis syndrome (ATLS) is a metabolic derangement (hyperuricaemia, hyperphosphataemia, hyperkalaemia and hypocalcaemia) associated with lymphoproliferative malignancies. The nature and severity of the metabolic alterations are variable. Major complications are oliguric acute renal failure and delays in initiating chemotherapy. Current management of ATLS includes hydration, alkalinization, diuretics, when indicated, and the reduction of uric acid levels using allopurinol or urate oxidase. Allopurinol inhibits xanthine oxidase, an enzyme that catalyses the conversion of hypoxanthine and xanthine to uric acid. Urate oxidase (Uricozyme), a naturally occurring proteolytic enzyme in many mammals, degrades uric acid to allantoins, which are ten times more soluble than uric acid and easily eliminated by the kidneys. Recently, Sanofi Research isolated a recombinant urate oxidase (SR29142) as a cDNA clone from Aspergillus flavus, expressed in the yeast strain Saccharomyces cerevisiae. Preclinical studies have documented its biological effects as a urolytic enzyme. Twenty-eight healthy male volunteers received SR29142, and a rapid decline of uric acid below measurable levels was seen within 4 h in all patients receiving a dose of more than 0.10 mg kg(-1). Currently, SR29142 is undergoing clinical studies in both Europe and the USA in patients with acute leukaemias or B-cell non-Hodgkin's lymphoma to demonstrate its efficacy and safety in this population of patients at highest risk of developing ATLS or its life-threatening sequelae.

Journal ArticleDOI
TL;DR: The xanthine content of seven Ilex spp. was studied by HPLC using UV and a photodiode-array detector as discussed by the authors, and the results showed that I. paraguariensis has the highest amounts of caffeine and theobromine.
Abstract: The xanthine (caffeine, theobromine and theophylline) content of decoctions of seven Ilex spp. was studied by HPLC using UV and a photodiode-array detector. Ilex paraguariensis (‘Mate’ or ‘Yerba Mate’) is widely used in South America as a tealike beverage and the other six species are used as substitutes or adulterants of it. The results showed that I. paraguariensis has the highest amounts of caffeine and theobromine. Traces of caffeine were detected in I. theezans, I. dumosa, I. microdonta and I. pseudobuxus. Traces of theobromine were detected in I. argentina and I. microdonta. From the seven Ilex species investigated, theophylline was detected in I. theezans, I. dumosa and I. pseudobuxus, but only I. pseudobuxus has quantifiable amounts: 0.6 mg% (6 ppm). © 1998 John Wiley & Sons, Ltd.

Journal ArticleDOI
TL;DR: A novel function for xanthine compounds is delineated and the molecular features that enablexanthine activation of CFTR are identified, which may be useful in the development of new molecules for studying the pharmacology of chloride channels.
Abstract: On the basis of their structure, we compared the ability of 35 xanthine derivatives to activate the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel stably expressed in chinese hamster ovary (CHO) cells using the cell-attached patch clamp and iodide efflux techniques. Activation of CFTR channels was obtained with 3-mono, 1,3-di or 1,3,7-tri-substituted alkyl xanthine derivatives (enprofylline, theophylline, aminophylline, IBMX, DPMX and pentoxifylline). By contrast, xanthine derivatives substituted at the C8- or N9-position failed to open CFTR channels. The CFTR chloride channel activity was blocked by glibenclamide (100 μM) but not by DIDS (100 μM). Activation of CFTR by xanthines was not mimicked by the calcium ionophore A23187, adenosine, UTP, ATP or the specific phosphodiesterase inhibitors rolipram, Ro 20-1724 and milrinone. In addition, we found no correlation between the effect of xanthines on CFTR and on the cellular cyclic AMP or ATP levels. We then synthesized a series of 3,7-dimethyl-1-alkyl xanthine derivatives; among them, 3,7-dimethyl-1-propyl xanthine and 3,7-dimethyl-1-isobutyl xanthine both activated CFTR channels without increasing the intracellular cyclic AMP level, while the structurally related 3,7-dimethyl-1-(2-propenyl) xanthine and 3,7-dimethyl-1-(oxiranyl methyl) xanthine were inactive. Our findings delineate a novel function for xanthine compounds and identify the molecular features that enable xanthine activation of CFTR. These results may be useful in the development of new molecules for studying the pharmacology of chloride channels. British Journal of Pharmacology (1998) 123, 683–693; doi:10.1038/sj.bjp.0701648

Journal ArticleDOI
TL;DR: Observations reveal that .NO will not serve as an indirect antioxidant by inhibiting XO-derived production of reactive species and that the Xo-derived products O*-2 and uric acid readily modify the reactivities of .NO and ONOO-.

Journal ArticleDOI
TL;DR: It is concluded that oxygen free radicals generated by xanthine andxanthine oxidase released to the bloodstream are involved in the systemic organ failure associated with acute pancreatitis.
Abstract: In the present study we evaluate the possibility that xanthine oxidase released by damaged pancreas could act as a source of oxidative damage in systemic tissues during the early stages of acute pancreatitis. This was accomplished by evaluating the effects of xanthine oxidase inhibition with oxypurinol infused into the portal vein. Under these conditions, we inhibited the enzyme before it reached the liver and other distant organs, without inducing changes in the severity of pancreatic damage. Results indicate that pancreatitis parallels increases in xanthine oxidase activity in plasma. Superoxide radicals generated by this enzyme appears to be involved in the decrease of reduced glutathione levels in the plasma and liver. In addition, xanthine oxidase inhibition prevents the infiltration of neutrophils into the lungs. We conclude that oxygen free radicals generated by xanthine and xanthine oxidase released to the bloodstream are involved in the systemic organ failure associated with acute pancreatitis.

Journal ArticleDOI
TL;DR: Reperfusion during liver transplantation is associated with liberation of xanthine oxidase, hypoxanthine, andxanthine from the liver into the circulation and intravascular neutrophil activation takes place in the hepatic circulation.

Journal ArticleDOI
TL;DR: The detection of xanthine oxidase activity products by electron paramagnetic resonance (EPR) in presence of the spin-trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) and reversed-phase high-performance liquid chromatography (RP-HPLC) in COPD-BAL and BAL support the involvement ofxanthine oxidation in the mechanisms of superoxide production by BAL supernatant, which increases oxidative stress in chronic obstruct

Journal ArticleDOI
TL;DR: Results suggest that morphine increases striatal DA release and 5-hydroxytryptamine oxidative metabolism by a micro-opioid receptor-mediated mechanism mainly at extranigrostriatal sites and affects glutamate and AA release by amicro-opIOid receptor mediated mechanism acting also at nigral sites.

Journal ArticleDOI
Feng N. Ko1, Chen C Chu1, Chun N. Lin, Chia C Chang1, Che-M Teng1 
TL;DR: Since isoorientin-6"-O-glucoside did not possess pro-oxidant activity, it may be an effective water-soluble antioxidant that can prevent LDL against oxidation.

Journal ArticleDOI
TL;DR: Propofol had a direct scavenging activity against HOCl, superoxide-hydrogen peroxide and hydroxyl radical in the concentrations used, which may contribute to the effect of propofol on human leucocyte chemiluminescence.
Abstract: We have studied the ability of propofol and Intralipid to inhibit reactive oxygen species generated either by stimulated human leucocytes or cell-free systems using luminol chemiluminescence. Human leucocytes were stimulated by a chemotactic peptide, FMLP 1 mumol litre-1, or by a phorbol ester, PMA (protein kinase C activator) 0.1 mumol litre-1. In cell-free experiments, superoxide-hydrogen peroxide, hypochlorous acid or hydroxyl radical-induced chemiluminescence responses were initiated by xanthine 0.1 mmol litre-1 with xanthine oxidase 10 mu. ml-1, NaOCl 70 mumol litre-1 and FeSO4 3 mumol litre-1, respectively. Propofol with Intralipid, and to a lesser degree Intralipid alone, produced a concentration-dependent reduction in chemiluminescence from stimulated leucocytes. Similar attenuations were also observed using propofol with Intralipid on xanthine with xanthine oxidase-, HOCl- and ferrous iron-induced chemiluminescence. However, Intralipid produced a reduction only at high concentrations. Intralipid produced marked decreases in ferrous iron-induced chemiluminescence. This study suggests that propofol had a direct scavenging activity against HOCl, superoxide-hydrogen peroxide and hydroxyl radical in the concentrations used. These direct scavenging effects may contribute to the effect of propofol on human leucocyte chemiluminescence.

Journal ArticleDOI
TL;DR: Results indicate that leaves of Camellia ptilophylla exhibit unusual purine alkaloid metabolism as they have the capacity to synthesize theobromine from adenine nucleotides, but they lack adequate methyltransferase activity to convert of theobomine to caffeine in detectable quantities, and the leaves have a capacity to convert xanthine to theobroma, probably via 3-methylxanthine.
Abstract: The biosynthesis and metabolism of purine alkaloids in leaves ofCamellia ptilophylla (cocoa tea), a new tea resource in China, have been investigated. The major purine alkaloid was theobromine, with theophylline also being present as a minor component. Caffeine was not accumulated in detectable quantities. Theobromine was synthesized from [8-14C] adenine and the rate of its biosynthesis in the segments from young and mature leaves from flush shoots was approximately 10 times higher than that from aged leaves from 1-year old shoots. Neither cellfree extracts nor segments fromC. ptilophylla leaves could convert theobromine to caffeine. A large quantity of [2-14C] xanthine taken up by the leaf segments was degraded to14CO2 via the conventional purine catabolic pathway that includes allantoin as an intermediate. However, small amounts of [2-14C] xanthine were also converted to theobromine. Considerable amounts of [8-14C] caffeine exogenously supplied to the leaf segments ofC. ptilophylla was changed to theobromine. These results indicate that leaves ofC. ptilophylla exhibit unusual purine alkaloid metabolism as i) they have the capacity to synthesize theobromine from adenine nucleotides, but they lack adequate methyltransferase activity to convert of theobromine to caffeine in detectable quantities, ii) the leaves have a capacity to convert xanthine to theobromine, probably via 3-methylxanthine.

Patent
24 Feb 1998
TL;DR: In this paper, the use of 8-phenylxanthines, 8-cyloalkyl xanthines or 8- substituted xanthine derivatives to specifically modulate the physiologic role of the A2B adenosine receptor was discussed.
Abstract: The invention concerns the use of 8-phenylxanthines, 8-cyloalkylxanthines or 8- substituted xanthine derivatives to specifically modulate the physiologic role of the A2B adenosine receptor.

Journal ArticleDOI
TL;DR: Evidence is provided for antioxidant properties of phytic acid under in vitro conditions and neither phytics acid nor iron had any significant effect on liver oxidant or antioxidant status in vivo in growing rats.
Abstract: The objective of this study was to determine the effects of phytic acid on free radical generation in vitro and in growing rats. Electron spin resonance spectroscopy studies using 5, 5-dimethyl-1-pyrroline-N-oxide (DMPO) as a spin trap indicate a complete inhibition of hydroxyl radical formation via the iron-catalyzed Fenton reaction at molar phytic acid/iron ratios >5. However, phytic acid had no scavenging effect on superoxide radicals generated in the xanthine/xanthine oxidase reaction. For the in vivo study, male growing albino rats were fed purified diets based on casein, cornstarch and vitamin E-stripped corn oil differing in the concentration of iron (30 or 300 mg/kg), phytic acid (0 or 10 g/kg) and dl-alpha-tocopheryl acetate (0 or 50 mg/kg). At marginal dietary iron supply, phytic acid supplementation reduced apparent Fe absorption, thereby decreasing liver Fe concentration. Dietary iron and phytate had no effect on the level of hepatic alpha-tocopherol, reduced glutathione, thiobarbituric acid-reactive substances and protein carbonyls. The concentration of thiobarbituric acid-reactive substances and protein carbonyls in the liver decreased as dietary vitamin E was increased from 0 to 50 mg/kg diet. The results obtained provide evidence for antioxidant properties of phytic acid under in vitro conditions. However, neither phytic acid nor iron had any significant effect on liver oxidant or antioxidant status in vivo in growing rats.

Journal ArticleDOI
TL;DR: In this paper, sulfostyryl-xanthine derivatives were synthesized as water-soluble A2A-selective adenosine receptor (AR) antagonists.

Journal ArticleDOI
TL;DR: Lucigenin-amplified chemiluminescence experiments have shown that at low pH, the presence of hemoglobin reduces the level of superoxide anion generated by the xanthine/xanthine oxidase system (met-Hb is more efficient in reducing thelevel of O2- than oxy-hemoglobin).

Journal ArticleDOI
TL;DR: The catabolism of purines, the superoxide-generating PMPs, the ascorbate-glutathione cycle, and the dehydrogenase-mediated recycling of NADPH, are activated oxygen roles of leaf peroxisomes that add to other functions previously known for perxisomes from eukaryotic cells.
Abstract: Peroxisomes are subcellular organelles with an essentially oxidative type of metabolism. The presence in these organelles of superoxide dismutases and the generation of superoxide radicals (O2•−) was first demonstrated in plant tissues and in recent years different experimental evidence has suggested the existence of cellular functions related to activated oxygen species. Some of these functions are analyzed in this work. In purified intact peroxisomes from pea (Pisum sativum L.) leaves, xanthine oxidase and urate oxidase were found to be present. The occurrence and the level of the metabolites xanthine, hypoxanthine, uric acid, and allantoin were studied in extracts of pea leaf peroxisomes by HPLC. Xanthine, uric acid, and allantoin were detected in peroxisomes. These results suggest a cellular role for leaf peroxisomes in the catabolism of purines. In peroxisomal membranes, 3 polypeptides (PMPs) with molecular masses of 18, 29 and 32 kDa, respectively, have been shown to generate superoxide radicals. These PMPs were purified from pea leaf peroxisomal membranes and characterized. While the 18- and 32-kDa PMPs use NADH as electron donor for O2•− production, the 29-kDa PMP was clearly dependent on NADPH. Very recently, the occurrence in pea leaf peroxisomes of all the enzymes of the ascorbate-glutathione cycle has been demonstrated. NADPH is required for the glutathione reductase activity of the cycle and this implies the reduction of NADP+ to NADPH. This recycling function could be carried out by the NADP-dependent glucose-6-phosphate dehydrogenase (G6PDH), 6-phosphogluconate dehydrogenase (6PGDH), and isocitrate dehydrogenase (ICDH). These 3 dehydrogenases have been demonstrated to be present in the matrix of pea leaf peroxisomes. The catabolism of purines, the superoxide-generating PMPs, the ascorbate-glutathione cycle, and the dehydrogenase-mediated recycling of NADPH, are activated oxygen roles of leaf peroxisomes that add to other functions previously known for peroxisomes from eukaryotic cells.

Journal ArticleDOI
TL;DR: The decrease in Cyt-ox and increase in XO suggest that ROS are produced during DFP induced muscle fasciculations initiating lipid peroxidation and subsequent myopathy, and this may suggest a conversion from XD into XO.
Abstract: A possible role of radical oxygen species (ROS) initiated lipid peroxidation in diisopropylphosphorofluoridate (DFP)-induced muscle necrosis was investigated by quantifying muscle changes in F2-isoprostanes, novel and extremely accurate markers of lipid peroxidation in vivo. A significant increase in F2-isoprostanes of 56% was found in the diaphragm of rats 60 min after DFP-induced fasciculations. As possible source of ROS initiating lipid peroxidation, the cytocrome-c oxidase (Cyt-ox) and xanthine dehydrogenase-xanthine oxidase (XD-XO) systems were investigated. Within 30 min of onset of fasciculations Cyt-ox activity was reduced by 50% from 0.526 to 0.263 mumol/mg prot/min and XO activity increased from 0.242 to 0.541 mumol/mg prot/min. Total XD-XO activity was unchanged, indicating a conversion from XD into XO. In rats pretreatment with the neuromuscular blocking agent d-tubocurarine, prevented DFP-induced fasciculations, increases in F2-isoprostanes and changes in Cyt-ox or XD-XO. The decrease in Cyt-ox and increase in XO suggest that ROS are produced during DFP induced muscle fasciculations initiating lipid peroxidation and subsequent myopathy.

Journal ArticleDOI
TL;DR: The lack of ionic interactions between Thr-47 and PPi and an increased distance between the loop and the active site as compared to the human HGPRTase are proposed to be responsible for the high Km for PPi in the T. foetus HGXPRT enzyme-catalyzed reaction.
Abstract: Tritrichomonas foetus, an anaerobic flagellated protozoan, causes urogenital trichomoniasis in cattle. Hypoxanthine-guanine-xanthine phosphoribosyl transferase (HGXPRTase), an essential enzyme in T. foetus required for salvaging exogenous purine bases, has been regarded as a promising target for anti-tritrichomonial chemotherapy. The steady-state kinetic analyses of synthesis and pyrophosphorolysis of IMP, GMP, and XMP and product inhibition studies have been used to elucidate the reaction mechanisms. Double-reciprocal plots of initial velocities versus the varying concentrations of one substrate at a fixed concentration of the other show intersecting lines indicating a sequential mechanism for both the forward and the reverse reactions. In terms of the kcat/Km ratios, hypoxanthine is the most effective substrate whereas guanine and xanthine are converted equally well into their corresponding nucleotides. The minimum kinetic model from the data in product inhibition studies is an ordered bi−bi mechanism, ...