Compositional differences in flavonoids are varied in the big family of Compositae. By summarizing our previous analytical studies and other scientific evidences, new strategy will be possible to further analyze flavonoids and utilize them for human health. The HPLC analytical method has been established in terms of linearity, sensitivity, accuracy, and precision. Herbs of the family of Compositae have considerable amounts of peroxynitrite (ONOO-)-scavenging effects and their phenolic substances. These effects may contribute to the prevention of disease associated with excess production of ONOO-, depending on the high content of flavonoid substances.
Asteraceae* ; Chromatography, High Pressure Liquid ; Flavonoids ; Humans ; Peroxynitrous Acid ; Phenol

Transient forebrain ischemia results in the delayed neuronal death in the CA1 area of the hippocampus. The present study was performed to determine effects of aminoguanidine, a selective iNOS inhibitor, on the generation of peroxynitrite and delayed neuronal death occurring in the hippocampus following transient forebrain ischemia. Transient forebrain ischemia was produced in the conscious rats by four-vessel occlusion for 10 min. Treatment with aminoguanidine (100 mg/kg or 200 mg/kg, i.p.) or saline (0.4 ml/100 g, i.p.) was started 30 min following ischemia-reperfusion and the animals were then injected twice daily until 12 h before sacrifice. Immunohistochemical method was used to detect 3-nitrotyrosine, a marker of peroxynitrite production. Posttreatment of aminoguanidine (200 mg/kg) significantly attenuated the neuronal death in the hippocampal CA1 area 3 days, but not 7 days, after ischemia-reperfusion. 3-Nitrotyrosine immunoreactivity was enhanced in the hippocampal CA1 area 3 days after reperfusion, which was prevented by the treatment of aminoguanidine (100 mg/kg and 200 mg/kg). Our findings showed that (1) the generation of peroxynitrite in the hippocampal CA1 area 3 days after ischemia-reperfusion was dependent on the iNOS activity; (2) the postischemic delayed neuronal death was attenuated in the early phase through the prevention of peroxynitrite generation by an iNOS inhibitor.
Animals ; Hippocampus* ; Ischemia* ; Neurons ; Peroxynitrous Acid* ; Prosencephalon* ; Rats* ; Reperfusion

Lactuca raddeana (Compositae) is used to treat obesity and complications due to diabetes. The five phenolic compounds including chlorogenic acid, chicoric acid, luteolin 7-O-glucoside, luteolin 7-O-glucuronide, luteolin were qualitatively identified by LC-ESI-MS analysis. The contents were quantitatively determined by HPLC, under the condition of a Capcell Pak C18 column (5 microm, 250 mm x 4.6 mm i.d.) and a gradient elution of 0.05% trifluoroacetic acid (TFA) and 0.05% TFA in MeOH-H2O (60 : 40). The contents of chicoric acid (100.99 mg/g extract) and luteolin 7-O-glucoside (101. 69 mg/g extract) were high, while those of other three phenolic substances were very low. The 3T3-L1 adipocyte cells treated with chicoric acid and luteolin 7-O-glucuronide significantly suppressed the accumulation of fat, suggesting they are effective against obesity. Since high level of peroxynitrite (ONOO) causes cardiovascular disease in obese patients, its scavenging activity was also studied.
Adipocytes* ; Asteraceae ; Cardiovascular Diseases ; Chlorogenic Acid ; Chromatography, High Pressure Liquid ; Humans ; Luteolin ; Obesity ; Peroxynitrous Acid ; Phenol* ; Trifluoroacetic Acid

Many herbs have been used as therapeutics in Korean traditional medicine. In view of their clinical indications, anti-oxidant activity may contribute to their pharmacological effects. However, anti-oxidant information on these plants has not been available. In this study, seventy herbs which have been used in Korean traditional medicine were selected and screened for anti-oxidant activity using their water extracts. The anti-oxidant activity was assessed by their ability to inhibit three oxidation reactions; luminol/Fenton reagent, 2, 7-dichlorodihydrofluorescein (DCHF)/Fenton reagent and DCHF/peroxynitrite. In each assay, 70 herbs were divided into two groups; anti-oxidant group which inhibited the respective oxidation reaction and was majority (about 60 herbs), and pro-oxidant group which enhanced the oxidation reaction but was minority (more or less 10 herbs). When the herbs were listed in the order of their anti-oxidant strength, the orders obtained from each assay were found to be quite similar. The upper top rankers (more or less 10 herbs) in each assay showed strong activity compared to the others. The uppermost rankers in each assay were Rubus coreanus Miquel/ Rubus schizostylus, Schisandra chinensis Baillon/ Schizandra chinensis and Terminalia chebula Retzius/ Terminalia chebula. Of the pro-oxidant herbs, about 4-5 herbs were strongly pro-oxidant, which enhanced the control oxidation reactions to 150-300%. But the meaning of this observation is not known since few of them in one assay were also anti-oxidant in other assays. The results obtained in the present study may serve as information for understanding pharmacological effects of these herbs and developing new drugs from them.
Hydrogen Peroxide ; Iron ; Luminescence ; Medicine, Korean Traditional ; Peroxynitrous Acid ; Schisandra ; Terminalia ; Water

PURPOSE: To assess the role of nitric oxide in the development of cataract by detection of nitrotyrosine, which is a reaction product of peroxynitrite with tyrosine. METHODS: We have collected aqueous humor and anterior capsule of lens after cataract surgery from 14 eyes. We classified lens opacities into three types such as nuclear sclerosis, cortical and posterior subcapsular opacity. Nitrite, stable oxidative product of nitric oxide, was measured in the aqueous humor and immunohistochemical staining for nitrotyrosine was performed to examine anterior capsules. RESULTS: The concentrations of nitric oxide in aqueous humor were 0.0698+/-0.0036, 0.0652+/-0.0116, and 0.0700+/-0.0022 mM in nuclear sclerosis, cortical and posterior subcapsular opacity, respectively and no significant differences existed among lens opacity types. In lenses with cortical and posterior subcapsular opacity, the expression of nitrotyrosine was observed around epithelial cell and intercellular space. There was no expression of nitrotyrosine in lenses with nuclear sclerosis. CONCLUSIONS: This results suggest that nitration of tyrosine by peroxynitrite is possibly related to the cataractogenesis of lenses with cortical and posterior subcapsular opacity.
Aqueous Humor ; Capsules ; Cataract* ; Epithelial Cells ; Extracellular Space ; Nitric Oxide* ; Peroxynitrous Acid ; Sclerosis ; Tyrosine

Previously, we reported that glucose-deprived astrocytes are more vulnerable to the cytotoxicity of peroxynitrite, the reaction product of nitric oxide and superoxide anion. The augmented vulnerability of glucose-deprived astrocytes to peroxynitrite cytotoxicity was dependent on their proliferation rate. Inhibition of cell cycle progression has been shown to inhibit the apoptotic cell death occurring in cerebral ischemia-reperfusion. In the present study, we demonstrate that the increased death of glucose-deprived astrocytes by peroxynitrte was largely blocked by the cell cycle phase G2/M transition blocker etoposide. However, the cytoprotective effect of etoposide was not associated with its inhibition of cell cycle progression. Instead, etoposide effectively scavenged peroxynitrite. However, etoposide did not scavenge individual nitric oxide and superoxide anion and it did not prevent the hydrogen peroxide-induced cytotoxicity. The present results indicate that etoposide prevents the toxicity of peroxynitrite in astrocytes by directly scavenging peroxynitrite, not by inhibiting cell cycle progression.
Astrocytes ; Cell Cycle ; Cell Death ; Etoposide ; Hydrogen ; Nitric Oxide ; Peroxynitrous Acid ; Superoxides

BACKGROUND AND OBJECTIVES: Recent evidence has shown that nitric oxide (NO) levels are increased in allergic rhinitis. However, the role of this molecule in the pathophysiology of allergic rhinitis is still poorly understood. Peroxynitrite (OONO-), the reactive metabolites of NO, causes nitrotyrosine formation by the nitration of tyrosine residues, and promotes deleterious effects on protein function. We attempted in this study to clarify whether NO and nitrotyrosine in the nasal secretion could be increased in the early or the late phase reaction after allergen challenge. MATERIALS AND METHODS: Samples were obtained from thirteen patients with allergic rhinits to Dermatophagoides Pteronyssinus with the filter paper absorption method. The samples were collected right before, 30 minutes after, and 8 hours after the allergen challenge. Then we estimated the concentrations of nitrite (NO2-) and nitrate (NO3-). Nitrotyrosine in nasal secretions was determined by Western blot analysis in three patients. RESULTS: The nitrite/nitrate concentration in nasal secretions did not show significant changes between the baseline, the early, and the late phase (p>0.05). In the Western blot analysis, the concentration of nitrotyrosine was increased in the late phase. CONCLUSIONS: Although the NO was not increased after the allergen challenge, nitrotyrosine, the evidence of the peroxynitrite effect to tyrosine residues of the protein, was increased in the late phase of the reaction rather than the early phase. Because the peroxynitrite is an metabolite of NO, we can estimate that the overall NO effect has an influence on the late phase of the allergic reaction, and it can be presumed that NO has an influence on the long-term deterioration on the nasal mucosa by cytotoxic effect of peroxynitrite, rather than on the immediate reaction of allergic rhinitis.
Absorption ; Blotting, Western ; Dermatophagoides pteronyssinus ; Humans ; Hypersensitivity ; Nasal Mucosa ; Nitric Oxide* ; Peroxynitrous Acid* ; Rhinitis* ; Tyrosine

Peroxynitrite (ONOO-)-scavenging activities of nine Compositae herbs consisting of three Ixeris, two Youngia, two Cirsium and one of each Lactuca and Taraxacum species were evaluated. The contents of their ONOO- scavengers in the extracts were also determined on a HPLC using seven standard compounds, chlorogenic acid (CGA), chicoric acid (CA), luteolin 7-glucoside (luteolin-7-glc), luteolin 7-glucuronide (luteolin-7-glcU), luteolin, linarin and pectolinarin. Five of those compounds exhibited potent ONOO--scavenging activities: IC50, CA (0.76 microM), CGA (1.34 microM), luteolin (0.81 microM), luteolin-7-glc (0.86 microM) and luteolin-7-glcU (3.13 microM). Both CA and luteolin-7-glc were highly contained in I. dentata (19.71 mg/g and 13.58 mg/g, respectively), I. dentata var. albiflora (17.58 mg/g and 23.83 mg/g, respectively) and I. sonchifolia (65.71 mg/g and 6.99 mg/g, respectively). Among the nine herbs, those three Ixeris species had very low IC50 values over the range of 0.48 - 1.74 microg/mL, suggesting that they could be potential therapeutic vegetables, particularly for preventing diabetic complications or obesity, which can be caused by an excess production of ONOO-.
Asteraceae* ; Chlorogenic Acid ; Chromatography, High Pressure Liquid ; Cirsium ; Diabetes Complications ; Inhibitory Concentration 50 ; Luteolin ; Obesity ; Peroxynitrous Acid ; Phenol* ; Taraxacum ; Vegetables

Various proteomics and immunological methods including mass spectrometry combined with both liquid and 2-D PAGE, and immunodetection have been employed to identify and characterize nitrated proteins from pathological samples. Nitrosative modifications regulate cellular signal transduction and pathogenesis of inflammatory responses and neurodegenerative diseases. Nitric oxide generates reactive nitrosative species, such as peroxynitrite (ONOO-) that may be involved in a number of diseases. ONOO- can mediate protein tyrosine nitration which causes structural changes of affected proteins and leads to their inactivation. Protein tyrosine nitration is a biomarker of oxidative stress and also influences protein structure and function. Recent advances in mass spectrometry have made it possible to identify modified proteins and specific modified amino acid residues. This review focuses on the significance of protein tyrosine nitration and the progress achieved in analytical methods. Although mass spectrometry of nitrated peptides has become a powerful tool for the analysis of nitrated peptides, the low stoichiometry of protein tyrosine nitration clearly demands the use of affinity chromatography to enrich modified proteins (or peptides).
Chromatography, Affinity ; Mass Spectrometry ; Neurodegenerative Diseases ; Nitric Oxide ; Oxidative Stress ; Peptides ; Peroxynitrous Acid ; Proteins ; Proteomics ; Signal Transduction ; Tyrosine

The purpose of this study is to evaluate the extent of peroxynitrite generation in the pool of radicals/oxidants from intraocular inflammation using luminol-dependent chemiluminescence(LDCL)method. S-antigen induced uveitis was produced in Lewis rats. The rats were killed at the peak of inflammation, and the retinas and choroids were collected for the LDCL. Sodium bicarbonate was used to confirm the peroxynitrite signal. Superoxide dismutase(SOD), N-nitro-L-arginine methyl ester(LNAME)and aminoguanidine(AG)were tried to evaluate the inhibitory effect of superoxide and nitric oxide. LDCL counts for 6 inflamed and 6 control retina/choroid preparations were 66, 429+/-413 cpm and 13, 941+/-105 cpm, respectively(p<0.01). In the presence of bicarbonate, emission was increased by 125.3+/-6.6%(n=6)and the signal was sustained for 2 hours. SOD, L-NAME and AG suppressed the LDCL by 39.8+/-6.1%(n=3), 20.4+/-4.4%(n=3) and 35.9+/-4.0 %(n=3), These observations suggest that peroxynitrite contributes considerably to the generation of the total pool of reactive species by the cellular infiltrate. The presence of a high level of peroxynitrite, a potent oxidizing and nitrating agent, in inflamed retina may cause irreversible tissue damage to the photoreceptors.
Animals ; Choroid ; Inflammation ; Luminescence* ; NG-Nitroarginine Methyl Ester ; Nitric Oxide ; Peroxynitrous Acid ; Rats ; Retina ; Sodium Bicarbonate ; Superoxides ; Uveitis*