OBJECTIVETo evaluate the inhibitory effect of Gnaphalium affine extracts on xanthine oxidase (XO) activity in vitro and to analyze the mechanism of this effect.

METHODSIn this in vitro study, Kinetic measurements were performed in 4 different inhibitor concentrations and 5 different xanthine concentrations (60, 100, 200, 300, 400 Μmol/L). Dixon and Lineweaver-Burk plot analysis were used to determine Ki values and the inhibition mode for the compounds isolated from Gnaphalium affine extract.

RESULTSFour potent xanthine oxidase inhibitors were found in 95% ethanolic (v/v) Gnaphalium affine extract. Among them, the flavone Eupatilin exhibited the strongest inhibitory effect on XO with a inhibition constant (Ki) of 0.37 Μmol/L, lower than the Ki of allopurinol (4.56 mol/L), a known synthetic XO inhibitor. Apigenin (Ki of 0.56 Μmol/L, a proportion of 0.0053‰ in Gnaphalium affine), luteolin (Ki of 2.63 Μmol/L, 0.0032‰ in Gnaphalium affine) and 5-hydroxy-6,7,3',4'-tetramethoxyflavone (Ki of 3.15 Μmol/L, 0.0043‰ in Gnaphalium affine) also contributed to the inhibitory effect of Gnaphalium affine extract on XO activity.

CONCLUSIONSThese results suggest that the use of Gnaphalium affine in the treatment of gout could be attributed to its inhibitory effect on XO. This study provides a rational basis for the traditional use of Gnaphalium affine against gout.

Flavonoids ; pharmacology ; Gnaphalium ; chemistry ; Xanthine Oxidase ; antagonists & inhibitors

A double targets of high throughput screening model for xanthine oxidase inhibitors and superoxide anion scavengers was established. In the reaction system of xanthine oxidase, WST-1 works as the probe for the ultra oxygen anion generation, and product uric acid works as xanthine oxidase activity indicator. By using SpectraMax M5 continuous spectrum enzyme sign reflectoscope reflector, the changes of these indicators' concentration were observed and the influence factors of this reaction system to establish the high throughput screening model were studied. And the model is confirmed by positive drugs. In the reaction system, the final volume of reaction system is 50 μL and the concentrations of xanthine oxidase is 4 mU x mL(-1), xanthine 250 μmol x L(-1) and WST-1 100 μmol x L(-1), separately. The Z'-factor of model for xanthine oxidase inhibitors is 0.537 4, S/N is 47.519 9; the Z'-factor of model for superoxide anion scavengers is 0.507 4, S/N is 5.388 9. This model for xanthine oxidase inhibitors and superoxide anion scavengers has more common characteristics of the good stability, the fewer reagent types and quantity, the good repeatability, and so on. And it can be widely applied in high-throughput screening research.
Enzyme Inhibitors ; pharmacology ; Free Radical Scavengers ; pharmacology ; High-Throughput Screening Assays ; Superoxides ; Uric Acid ; Xanthine ; Xanthine Oxidase ; antagonists & inhibitors

OBJECTIVETo evaluate the antioxidant activities of different chemical constituents from Astragalus mongholicus Bunge and their protection against xanthine (XA)/xanthine oxidase (XO)-induced toxicity in PC12 cells.

METHODSThe compounds of Astragalus mongholicus Bunge were isolated by chromatography and the structures were elucidated on the basis of spectral data interpretation. Their antioxidant activities were detected by 1, 1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging activities in a cell-free system. Meanwhile, the effects against XA/XO-induced toxicity were assessed using MTT assay in PC12 cells.

RESULTSTen principal constituents were isolated and identified as formononetin (I), ononin (II), calycosin (III), calycosin-7-O-beta-D-glucoside (IV), 9,10-dimethoxypterocarpan-3-O-beta-D-glucoside (V), adenosine (VI), pinitol (VII), daucosterol (VIII), beta-sitoster (IX) and saccharose (X) from Astragalus mongholicus Bunge. The compounds I, III, and IV scavenged DPPH free radicals in vitro. Formononetin and calycosin were found to inhibit XA/XO-induced cell injury significantly, with an estimated EC50 of 50 ng/mL.

CONCLUSIONCompound II, VI, and VII are first reported in this plant. Calycosin exhibits the most potent antioxidant activity both in the cell-free system and in the cell system.

Animals ; Astragalus Plant ; chemistry ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Free Radical Scavengers ; chemistry ; pharmacology ; Free Radicals ; metabolism ; Isoflavones ; chemistry ; pharmacology ; PC12 Cells ; Rats ; Xanthine ; toxicity ; Xanthine Oxidase ; toxicity

To determine whether oxygen free radicals are responsible for the pathogenesis of the cholestasis induced by ligation of common bile duct (CBD) variables which reflect the hepatic function in the serum, the amount of superoxide radical production, and xanthine oxidase(XO) activity were studied. The activity of serum alanine aminotransferase, bilirubin level in the serum and the amount of superoxide radical production were lower in a CBD ligation with allopurinol treated group than in a CBD ligation without allopurinol treated group. Abnormalities of the microscopic structures were reduced in a CBD ligation with allopurinol treated group than in a CBD ligation without allopurinol treated group. Allopurinol, an inhibitor of XO, prevented the hepatic damage induced by CBD ligation through the inhibition of XO. These experiments demonstrate that oxygen free radicals are responsible for the pathogenesis of the cholestatic liver.
Allopurinol/*pharmacology ; Animal ; Bile Ducts ; Cholestasis/*pathology ; Enzyme Inhibitors/*pharmacology ; Free Radicals ; Ligation ; Liver/*pathology ; Male ; Rats ; Rats, Sprague-Dawley ; Superoxides/metabolism ; Xanthine Oxidase/analysis/*antagonists & inhibitors

In the present study, two new acetylene conjugate compounds, dibutyl (2Z, 6Z)-octa-2, 6-dien-4-yne dioate (1), and dibutyl (2E, 6E)- octa-2, 6-dien-4-yne dioate (2), were isolated from the dry stem leaves of Viscum album, along with nine known compounds (3 - 11). Their structures were confirmed on the basis of spectroscopic data. Compounds 1 and 8 showed antioxidant activity against xanthine oxidase (XOD) and 1,1-diphenyl-2-picrylhydrazyl radical 2,2-diphenyl-1-(2,4,6-trinitrophenyl) hydroxyl (DPPH), with the IC of 1.22 and 1.33 μmol·L, and the SC of 4.34 and 8.22 μmol·L, respectively.
Acetylene ; chemistry ; Antioxidants ; chemistry ; pharmacology ; Biphenyl Compounds ; chemistry ; Molecular Structure ; Picrates ; chemistry ; Plant Extracts ; chemistry ; pharmacology ; Plant Leaves ; chemistry ; Viscum album ; chemistry ; Xanthine Oxidase ; chemistry

We evaluated DNA protection effect of heat shock protein (HSP) against cytotoxic effects of exogenous nitric oxide (NO) and reactive oxygen intermediate (ROI). Cultured human corneal fibroblasts were divided into 4 groups. Control (Group I) was not exposed to a sub-lethal heat treatment. Other 3 groups were exposed to 43 degrees C for 1 hr, then incubated at 37 degrees C during different duration (1, 6, 24 hr, Group II, III, IV, respectively). Expression pattern of HSP 70 was analyzed by Western blot. Cell viability was measured by MTT assay and the relationship between HSP 70 expression and DNA damage was examined by terminal deoxyribonucleotidyl transferase mediated dUTP-digoxigenin nick and labeling (TUNEL) stain and single cell gel electrophoresis. Expression pattern of HSP 70 was dependent on recovery times. Cell viability following heat treatment was significantly increased and the TUNEL positive cell number was decreased at 6 hr. In single cell gel electrophoresis, tail moments were increased in a dose-dependent manner by SNAP and X/XO. Following heat treatment, tail moments showed decreased significantly at 6 hr. These results suggest that induction of HSP 70 by sub-lethal heat treatment is closely related with cytoprotective effects against oxidative stresses in human corneal fibroblasts.
Cell Survival ; Cells, Cultured ; Cornea/*cytology ; DNA Damage ; Dose-Response Relationship, Drug ; Fibroblasts/cytology/drug effects/*metabolism ; Heat ; Heat-Shock Proteins 70/genetics/*metabolism ; Humans ; In Situ Nick-End Labeling ; Nitric Oxide/metabolism ; Nitric Oxide Donors/pharmacology ; *Oxidative Stress ; Reactive Oxygen Species/metabolism ; Research Support, Non-U.S. Gov't ; S-Nitroso-N-Acetylpenicillamine/pharmacology ; Xanthine/pharmacology ; Xanthine Oxidase/pharmacology

Animals ; Chronic Disease ; Febuxostat ; Heart Failure ; drug therapy ; metabolism ; Humans ; Reactive Oxygen Species ; Thiazoles ; pharmacology ; therapeutic use ; Xanthine Oxidase ; antagonists & inhibitors

AIMTo explore the role of ligustrazin on dynamic changes of lipid peroxidation in hepatic ischemia/reperfusion injury (HIRI) and its mechanism.

METHODSThe HIRI model was used. Twenty rabbits were randomly divided into control group (n = 10) and ligustrazin group (n = 10). The xanthine oxidase (XO) activity, superoxide dismutase (SOD) activity,malondialdehyde (MDA) content and glutamic pyruvic transaminase (GPT) activity in plasma were observed before ischemia and at ischemia 25 min, reperfusion 25 min, reperfusion 60 min and reperfusion 120 min.

RESULTSThe XO activity, SOD activity, MDA content and GPT activity of ligustrazin group, as compared with control group, showed significant differences (P < 0.05 or P < 0.01) at total time points of reperfusion.

CONCLUSIONLigustrazin has notable anti-lipid peroxidation effect on HIRI, which is due to its inhibiting the generation of oxygen free radicals and its strengthening scavenging of oxygen free radicals.

Alanine Transaminase ; metabolism ; Animals ; Female ; Lipid Peroxidation ; drug effects ; Liver ; drug effects ; metabolism ; Male ; Malondialdehyde ; blood ; Pyrazines ; pharmacology ; Rabbits ; Reperfusion Injury ; metabolism ; Superoxide Dismutase ; metabolism ; Xanthine Oxidase ; metabolism

The attenuating effect of daidzein (DAI) on oxidative toxicity induced by Aroclor 1254 (A1254) was investigated in mouse testicular cells. Cells were exposed to A1254 alone or with DAI. The oxidative damage was estimated by measuring malondialdehyde (MDA) formation, superoxide dismutase (SOD) activity and glutathione (GSH) content. Results show that A1254 induced a decrease of germ cell number, an elevation in thiobarbituric acid reactive substances (TBARS) but a decrease in SOD activity and GSH content. However, simultaneous supplementation with DAI decreased TBARS level and increased SOD activity and GSH content. Consequently, dietary DAI may restore the intracellular antioxidant system to attenuate the oxidative toxicity of A1254 in testicular cells.
Animals ; Chlorodiphenyl (54% Chlorine) ; toxicity ; Hypoxanthine ; toxicity ; Isoflavones ; pharmacology ; Male ; Malondialdehyde ; metabolism ; Mice ; Mice, Inbred ICR ; Oxidation-Reduction ; Testis ; drug effects ; metabolism ; Xanthine Oxidase ; toxicity

Six compounds have been isolated from the leaves of Pyrenacantha staudtii, two of which are new compounds. The new compounds have been characterized as kaempherol 3-O-beta-rhamnopyranosyl (1-->6)-beta-D-glucopyranoside (1) and 4-beta-glucopyranosyl-(2-furyl)-5-methy-1,2-glucopyranoside phenylmethanone (2). The known compounds are 3-pyridinecarboxylic acid (3), beta-sitosterol (4), sitosterol 3-O-beta-glucopyranoside (5) and taraxerol (6). Their structures were determined by spectroscopic and chemical evidences. The two new compounds together with 3-pyridinecarboxylic acid showed significant in vitro xanthine oxidase inhibitory activity. To the best of our knowledge, this is the first report of these compounds from this plant.
Enzyme Inhibitors ; chemistry ; isolation & purification ; pharmacology ; Glucosides ; chemistry ; isolation & purification ; pharmacology ; Kaempferols ; chemistry ; isolation & purification ; pharmacology ; Magnoliopsida ; chemistry ; Molecular Structure ; Niacin ; chemistry ; isolation & purification ; pharmacology ; Plant Leaves ; chemistry ; Plants, Medicinal ; chemistry ; Xanthine Oxidase ; metabolism