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This study aimed to investigate the effect of Gnaphalium affine extract (GAE) (0.04, 0.2 and 1 mg/g protein) on the gel properties of porcine myofibrillar proteins (MPs) in a simulated Fenton oxidation system, using tea polyphenols (TPs) at similar concentrations of 0.04, 0.2, and 1 mg/g protein, respectively, as a contrast. The findings revealed that as the TP concentration increased, the water retention of MP gels decreased significantly (p < 0.05). In contrast, MP gels containing medium and high concentrations of GAE exhibited significantly higher water retention than those with low concentrations of GAE (p < 0.05). When the concentration of GAE was increased to 1 mg/g protein, the strength of MP gels was significantly reduced (p < 0.05) by 33.32% compared with the oxidized control group, suggesting that low and medium GAE concentrations support MP gel formation. A texture profile analysis indicated that an appropriate GAE concentration improved gel structure and texture. Dynamic rheological characterization revealed that low concentrations of TP (0.04 mg/g protein) and low and medium concentrations of GAE (0.04 and 0.2 mg/g protein) strengthened the protein gel system. Conversely, high concentrations of TP and GAE (1.0 mg/g protein) damaged the protein gel system or even promoted the collapse of the gel system. Scanning electron microscopy revealed that higher TP concentrations disrupted the gel, whereas low and medium GAE concentrations maintained a more continuous and complete gel network structure compared with the oxidized control group. This indicates that an appropriate GAE concentration could effectively hinder the destruction of the gel network structure by oxidation. Therefore, based on the obtained results, 0.2 mg/g protein is recommended as the ideal concentration of GAE to be used in actual meat processing to regulate the oxidization and gel properties of meat products.
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OBJECTIVE To screen the quality biomarkers of Gnaphalium affine with anti-chronic obstructive pulmonary disease (COPD) effect and determine their contents. METHODS The effective components and targets of “G. affine” with anti- COPD effect were predicted by using network pharmacology as a search criterion. HPLC fingerprints for 10 batches of G. affine were established by using Similarity Evaluation System of TCM Chromatographic Fingerprint (2012 edition); common peak identification and similarity evaluation were conducted; cluster analysis (CA), principal component analysis (PCA), and orthogonal partial least squares-discriminant analysis (OPLS-DA) were performed to screen differential components as quality maker that affected the quality of G. affine using variable importance projection (VIP)>1 as the standard. The same HPLC method was adopted to determine the contents of the differential components in 10 batches of samples. RESULTS A total of 10 flavonoids (such as quercetin, luteolin, and chlorogenic acid) and organic acid components, were identified through network pharmacology search, with 91 targets closely related to anti-COPD. A total of 9 common peaks were identified in 10 batches of samples, with similarity greater than 0.90. Among them, the differential components included chlorogenic acid, caffeic acid, 1,3-O- dicaffeoylquinic acid and apigenin 7-O-β-D-glucopyranoside; S3, S4, S6, S7 and S10 were clustered into one category, S2, S5, S8 and S9 clustered into one category, and S1 clustered into one category. The contents of chlorogenic acid, caffeic acid, 1,3-O- dicaffeoylquinic acid, and apigenin 7-O-β-D-glucopyranoside in 10 batches of G. affine ranged 0.070-7.653, 0.010-0.097, 0.001- 0.036, 0.508-6.627 mg/g, respectively. CONCLUSIONS Chlorogenic acid, caffeic acid, 1,3-O-dicaffeoylquinic acid, apigenin 7- O-β-D-glucopyranoside can serve as the potential quality marker for the anti-COPD effect of G. affine, with the highest content of chlorogenic acid in G. affine produced in Ji’an, Jiangxi province, and the highest content of caffeic acid in G. affine produced in Ji’an, Jiangxi province and Sanming, Fujian province. The contents of the last two components are highest in G. affine produced in Chaoshan, Guangdong province.
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ObjectiveTo establish a rapid and stable liquid chromatography-mass spectrometry(LC-MS) for simultaneous analysis of 17 chemical components in Gnaphalium affine aboveground parts with flowers, so as to provide experimental basis for improving the quality standard of this herb. MethodUltra performance liquid chromatography-quadrupole/electrostatic field orbitrap mass spectrometry(UPLC-Q-Exactive Orbitrap MS) was used for the quantitative analysis of 17 constituents in 15 batches of G. affine from different origins, the separation was performed on an ACQUITY UPLC® BEH C18 column(2.1 mm×100 mm, 1.7 μm) with the mobile phase of methanol(A)-0.1% formic acid aqueous solution(B) for gradient elution(0-1.0 min, 8%A; 1.0-4.0 min, 8%-26%A; 4.0-9.0 min, 26%A; 9.0-14.0 min, 26%-34%A; 14.0-14.5 min, 34%-45%A; 14.5-15.0 min, 45%-60%A; 15.0-18.0 min, 60%-90%A; 18.0-19.0 min, 90%A; 19.0-19.01 min, 90%-8%A; 19.01-20.0 min, 8%A), the flow rate was 0.3 mL·min-1, the column temperature was 40 ℃ and the injection volume was 2 μL. And the electrospray ionization was used with full scanning in both positive and negative ion modes, and the scanning range was m/z 100-1 000. ResultThe established method has been verified by the methodology and could be used for the simultaneous quantification of 17 components in G. affine. The content ranges of the 17 components(quinic acid, gallic acid, protocatechuic acid, neochlorogenic acid, chlorogenic acid, cryptochlorogenic acid, caffeic acid, 1,3-O-dicaffeoylquinic acid, isochlorogenic acid A, isoquercitrin, 1,5-O-dicaffeoylquinic acid, apigenin-7-O-glucoside, astragalin, isochlorogenic acid C, luteolin, apigenin and hispidulin) in 15 batches of G. affine samples was 39.60-179.12, 0.17-0.84, 2.41-8.38, 4.33-31.50, 13.63-180.38, 2.43-14.75, 1.16-19.68, 0.49-5.63, 55.77-445.16, 0.23-10.26, 62.04-530.10, 1.11-18.01, 11.36-90.61, 12.22-65.98, 7.22-69.84, 3.37-45.65, 0.30-2.59 μg·g-1, respectively. The content of organic acids was higher than that of flavonoids in G. affine, and the contents of 1,5-O-dicaffeoylquinic acid, isochlorogenic acid A, quinic acid and chlorogenic acid were higher. Meanwhile, the content of flavonoids in the samples from Guizhou was higher than that from Jiangsu, while the content of organic acids in the samples from Jiangsu was higher than that from Guizhou. ConclusionThe established method can be used for the rapid and accurate determination of 17 components in G. affine, which clarifies the content range of the main components in this herb, and can provide a reference for the selection of quality control markers of G. affine.
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BACKGROUND: Gnaphalium affine D. Don extract (GAD) enhanced efficacy and reduced toxicity of benzbromarone (BBR) in combination use. However, little is known about effects of GAD on the pharmacokinetics (PKs) and metabolic enzymes of BBR. PURPOSE: To investigate the pharmacokinetic (PK) and pharmacodynamic (PD) mechanism of the herb-drug interactions (HDIs) between GAD and BBR. STUDY DESIGN AND METHODS: Intragastric single BBR (4.5 or 50 mg/kg), single BBR (4.5 or 50 mg/kg) + single GAD (450 mg/kg, 2 h after BBR-administration), or single BBR (4.5 or 50 mg/kg) + multiple GAD (450 mg/kg/day, once daily for 7 days) were administered to both sexes for BBR PK studies in normal rats. Intragastric multiple BBR (4.5 mg/kg/day), or multiple BBR (4.5 mg/kg/day) + multiple GAD (450 mg/kg/day, 2 h after BBR-administration) were administered for BBR PK and PD studies in male rats with hyperuricemic nephropathy (HN). The cumulative anti-hyperuricemic effects of BBR and BBR+GAD were determined by plasma uric acid (UA) concentration-time curve and area under curve (AUCUA). An in vivo cocktail approach was employed to determine the effects of GAD on cytochrome P450 (CYP) 2C11(9) and 1A2 - mediated drug metabolism. RESULTS: In normal rats, the repeated dose administration of GAD induced a significant increase of BBR AUC and prolonged the mean residence time (MRT) (p < 0.05). systemic exposure to BBR and metabolically derived hydroxybenzbromarones was significantly greater in female compared with male rats (p < 0.05). In HN rats, post-administration of GAD resulted in significantly higher bioavailability and enterohepatic recycling (ER) of BBR relative to the BBR alone administrated group from the prolongation of terminal elimination half-life (T1/2) and MRT of BBR (p < 0.05). Significantly higher urate-lowering effect of BBR+GAD compared with BBR alone was generally observed at post-dosing most time points with a maximal effect of 84.3% (acute treatment), 71.4% (7-day subchronic treatment) and 82.5% (14-day subchronic treatment) reduction in UA levels. Additionally, GAD showed a significant inhibitory effect on CYP2C11(9)-mediated tolbutamide (probe substrate) metabolism with ≥ 1.25 but < 2-fold increase in AUCtolbutamide. CONCLUSIONS: PD synergism demonstrated with the BBR+GAD combination could be explained by the PK interaction observed partially from CYP2C11(9)-mediation and enterohepatic recycling.
Subject(s)
Gnaphalium , Herb-Drug Interactions , Animals , Benzbromarone/pharmacology , Female , Male , Plant Extracts/pharmacology , Rats , Tolbutamide/pharmacokineticsABSTRACT
ETHNOPHARMACOLOGICAL RELEVANCE: Gnaphalium affine D. Don is an important Traditional Chinese herbal Medicine (TCM) used to treat hyperuricemia, asthma, rheumatic arthritis, antitussive, expectorant and cardiovascular in folk medicine because of anti-inflammatory and anti-oxidant activity. The aim of this study was to investigate the potential beneficial effect of G. affine extract (GAE) on hydrogen peroxide (H2O2)-induced apoptosis and explore the possible underlying mechanism in cardiomyocyte. MATERIALS AND METHODS: The ingredients of GAE were isolated and tentatively identified by HPLC-ESI-Q-Qribatrip-MS/MS. The cardioprotective and anti-oxidant effects of GAE were evaluated in the experimental model with H2O2 induced apoptosis in H9c2 cells. H9c2 cells were pretreated for 3 h with or without GAE or with GAE plus PX866 (PI3K inhibitor), then exposed to H2O2 for 6 h, H9c2 cells viability were detected by CCK8 kit, the content of intracellular reactive oxygen species (ROS) and malondialdehyde (MDA) and intracellular superoxide dismutase (SOD) activity were measured by the commercial biochemical kits, western blotting, immunohistochemical (IHC), immunofluorescence (IF) and reverse transcription-polymerase chain reaction (RT-PCR) assays were performed to evaluate the proteins and mRNA expression, propidium iodide (PI) staining was adopted to indicate H9c2 cells apoptosis. RESULTS: Firstly, seventeen polyphenols and flavonoids compounds with the characteristics of anti-inflammatory and anti-oxidant in GAE were tentatively identified by HPLC-ESI-Q-Qribatrip-MS/MS. In the experimental model, GAE not only significantly improved cells viability, but also showed anti-oxidant effects through improving SOD activity, up-regulating nuclear factor E2-related factor 2 (Nrf2), and decreasing intracellular concentration of ROS and MDA and the proteins expression of p47phox, p67phox and gp91phox. On the other hand, GAE revealed anti-apoptotic effect through up-regulating the expression of B-cell lymphoma-2 (Bcl-2), down-regulating Bcl2-associated X (BAX) and cleaved-caspase 3. Furthermore, GAE significantly facilitated phosphorylation of AKT and glycogen synthase kinase-3 beta (GSK-3ß) but not AMPK, while the effects were blocked by PX866 (PI3K inhibitor). CONCLUSIONS: Our data suggested that GAE showed strong anti-oxidant effect to ameliorate oxidative stress and attenuate apoptosis induced by H2O2 in H9c2 cells by targeting PI3K/AKT/GSK-3ß signaling pathway.
Subject(s)
Glycogen Synthase Kinase 3 beta/metabolism , Gnaphalium , Hydrogen Peroxide/toxicity , Myocytes, Cardiac/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Plant Extracts/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Animals , Apoptosis/drug effects , Apoptosis/physiology , Cell Line , Cell Survival/drug effects , Cell Survival/physiology , Drug Delivery Systems/methods , Myocytes, Cardiac/drug effects , Oxidative Stress/drug effects , Oxidative Stress/physiology , Plant Extracts/administration & dosage , Plant Extracts/isolation & purification , Rats , Signal Transduction/drug effects , Signal Transduction/physiologyABSTRACT
Objective To provide the experimental basis for the subsequent genetic diversity research through establishing and optimizing the inter-simple sequence repeat PCR (ISSR-PCR) reaction system of Gnaphalium affine. Methods The single-factor experimental method and full experimental method were used to optimize the ISSR-PCR reaction system of Gnaphalium affine. Under the optimal system, after screening primers and corresponding annealing temperatures, the systematic feasibility was verified. Results The optimal ISSR-PCR reaction system was consisted of 10 μl Premix Taq DNA polymerase, 0.3 μmol/L primer, 10 ng DNA template, and sterilized water added to 20 μl. Finally, 10 primers were screened from 100 universal primers, and verification results indicated the system had high stability, good reproducibility, and the selected primers had good polymorphism. Conclusion The ISSR-PCR amplification system of Gnaphalium affine was established for the first time and the primers with appropriate annealing temperatures were filtered out, which provided a reference for the subsequent genetic diversity research of Gnaphalium affine.
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Gnaphalium affine D. Don, a medicinal and edible plant, has been used to treat gout in traditional Chinese medicine and popularly consumed in China for a long time. A detailed phytochemical investigation on the aerial part of G. affine led to the isolation of two new esters of caffeoylquinic acid named (-) ethyl 1, 4-di-O-caffeoylquinate (1) and (-) methyl 1, 4-di-O-caffeoylquinate (2), together with 35 known compounds (3-37). Their structures were elucidated by spectroscopic data and first-order multiplet analysis. All the isolated compounds were tested for their xanthine oxidase inhibitory activity with an in vitro enzyme inhibitory screening assay. Among the tested compounds, 1 (IC50 11.94 µmol·L-1) and 2 (IC50 15.04 µmol·L-1) showed a good inhibitory activity. The current results supported the medical use of the plant.
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Drugs, Chinese Herbal/chemistry , Drugs, Chinese Herbal/pharmacology , Gnaphalium/chemistry , Phytochemicals/chemistry , Plant Extracts/pharmacology , Quinic Acid/analogs & derivatives , Xanthine Oxidase/antagonists & inhibitors , Adenine/analogs & derivatives , Adenine/chemistry , Adenine/isolation & purification , Enzyme Activation/drug effects , Flavonoids/chemistry , Flavonoids/isolation & purification , Gout Suppressants/chemistry , Gout Suppressants/isolation & purification , Gout Suppressants/pharmacology , Hydroxybenzoates/chemistry , Hydroxybenzoates/isolation & purification , Molecular Structure , Nuclear Magnetic Resonance, Biomolecular , Phytochemicals/isolation & purification , Phytochemicals/pharmacology , Plant Components, Aerial/chemistry , Plant Extracts/chemistry , Plant Extracts/isolation & purification , Quinic Acid/chemistry , Quinic Acid/isolation & purificationABSTRACT
AIM To establish an HPLC method for the simultaneous content determination of seven constituents in Gnaphalium affine D.Don.METHODS The analysis of 80% methanol of G.affine was performed on a 30 ℃ Atlantis (C) T3 column (4.6 mm× 250 mm,5 μm),with the mobile phase comprising of acetonitrile-formic acid flowing at 1.0 mL/min in a gradient elution manner,and the detection wavelength was set at 288 nm.RESULTS Seven constituents showed good linear relationships within their own ranges (R2 ≥0.999 8),whose average recoveries were 98.58%-103.8% with the RSDs of 0.88%-1.74%.CONCLUSION This accurate,stable and reproducible method can be used for the quality control of G.affine.
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Gnaphalium affine D. Don, a medicinal and edible plant, has been used to treat gout in traditional Chinese medicine and popularly consumed in China for a long time. A detailed phytochemical investigation on the aerial part of G. affine led to the isolation of two new esters of caffeoylquinic acid named (-) ethyl 1, 4-di-O-caffeoylquinate (1) and (-) methyl 1, 4-di-O-caffeoylquinate (2), together with 35 known compounds (3-37). Their structures were elucidated by spectroscopic data and first-order multiplet analysis. All the isolated compounds were tested for their xanthine oxidase inhibitory activity with an in vitro enzyme inhibitory screening assay. Among the tested compounds, 1 (IC 11.94 μmol·L) and 2 (IC 15.04 μmol·L) showed a good inhibitory activity. The current results supported the medical use of the plant.
Subject(s)
Adenine , Chemistry , Drugs, Chinese Herbal , Chemistry , Pharmacology , Enzyme Activation , Flavonoids , Chemistry , Gnaphalium , Chemistry , Gout Suppressants , Chemistry , Pharmacology , Hydroxybenzoates , Chemistry , Molecular Structure , Nuclear Magnetic Resonance, Biomolecular , Phytochemicals , Chemistry , Pharmacology , Plant Components, Aerial , Chemistry , Plant Extracts , Chemistry , Pharmacology , Quinic Acid , Chemistry , Xanthine OxidaseABSTRACT
Gnaphalium affine D. Don, a medicinal and edible plant, has been used to treat gout in traditional Chinese medicine and popularly consumed in China for a long time. A detailed phytochemical investigation on the aerial part of G. affine led to the isolation of two new esters of caffeoylquinic acid named (-) ethyl 1, 4-di-O-caffeoylquinate (1) and (-) methyl 1, 4-di-O-caffeoylquinate (2), together with 35 known compounds (3-37). Their structures were elucidated by spectroscopic data and first-order multiplet analysis. All the isolated compounds were tested for their xanthine oxidase inhibitory activity with an in vitro enzyme inhibitory screening assay. Among the tested compounds, 1 (IC 11.94 μmol·L) and 2 (IC 15.04 μmol·L) showed a good inhibitory activity. The current results supported the medical use of the plant.
Subject(s)
Adenine , Chemistry , Drugs, Chinese Herbal , Chemistry , Pharmacology , Enzyme Activation , Flavonoids , Chemistry , Gnaphalium , Chemistry , Gout Suppressants , Chemistry , Pharmacology , Hydroxybenzoates , Chemistry , Molecular Structure , Nuclear Magnetic Resonance, Biomolecular , Phytochemicals , Chemistry , Pharmacology , Plant Components, Aerial , Chemistry , Plant Extracts , Chemistry , Pharmacology , Quinic Acid , Chemistry , Xanthine OxidaseABSTRACT
Objective To establish a HPLC method for the simultaneous determination of quercetin and mignonette in Gnaphalium affine. Methods The determination was performed on Diamonsil C18 (2) column (4.6 mm×250 mm, 5 μm), mobile phase was methanol-0.4% phosphoric acid solution (42:58), detection wavelength was set at 360 nm, column temperature was 30℃, flow rate was 1 mL · min-1 and the injection volume was 20 μL. Results The linear rages of quercetin and mignonette were 3.588-35.880 μg · mL-1 (R2=0.999 6) and 1.294-12.940 μg · mL-1 (R2=0.999 4) respectively. Their average recoveries (n=6) were 99.51% (RSD=0.31%) and 99.74% (RSD=2.28%) respectively.Quercetin Content was 0.39-0.58 mg·g-1, mignonette content was 0.17-0.27 mg·g-1.Conclusion The method is simple, accurate, reliable and repeatable, thus it can be used for the quality control of Gnaphalium affine.
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ETHNOPHARMACOLOGICAL RELEVANCE: The Gnaphalium affine D. Don is used in China as a folk medicine to treat gout, anti-inflammatory, antitussive and expectorant activities. The aim of this study was to evaluate the potential of the extract of G. affine to treat hyperuricemia and acute gouty arthritis in animal model. MATERIALS AND METHODS: G. affine extract was evaluated in an experimental model with potassium oxonate (PO) induced hyperuricemia in mice which was used to evaluate anti-hyperuricemia activity and xanthine oxidase (XO) inhibition. Therapies for acute gouty arthritis was also investigated on monosodium urate (MSU) crystal induced paw edema model. RESULTS: G. affine extract showed expressive results on active in reducing serum uric acid (Sur) through effect renal mGLUT9 and mURAT1 mainly and inhibit XO activity in vivo. The extract of G. affine also showed significant anti-inflammatory activity and reduced the paw swelling on MSU crystal-induced paw edema model. Meanwhile, eight major compounds were identified by HPLC-ESI-QTOF-MS/MS. CONCLUSIONS: The extract of G. affine showed significant effect on evaluated models and therefore may be active agents for the treatment of hyperuricemia and acute gouty arthritis.
Subject(s)
Arthritis, Gouty/drug therapy , Gnaphalium/chemistry , Hyperuricemia/drug therapy , Plant Extracts/pharmacology , Animals , Anti-Inflammatory Agents/isolation & purification , Anti-Inflammatory Agents/pharmacology , Chromatography, High Pressure Liquid , Disease Models, Animal , Edema/drug therapy , Male , Mice , Mice, Inbred ICR , Oxonic Acid/toxicity , Tandem Mass Spectrometry , Uric Acid/blood , Xanthine Oxidase/antagonists & inhibitorsABSTRACT
Gnaphalium affine is an annual herbaceous plant that is used as a traditional medicine in some Latin American and Asian countries. However, systematic studies on its anti-inflammatory activity and signaling pathways have not yet been reported. In this study, we investigated the anti-inflammatory effect of G. affine methanol extract in lipopolysaccharide (LPS)-stimulated RAW 264.7 murine macrophage cells and fractioned the methanol extract into hexane, chloroform, ethyl acetate (EtOAc), butyl alcohol (BuOH), and distilled water (DW) by measuring the generation of nitric oxide (NO). G. affine inhibited the generation of NO and prostaglandin E2. The chloroform-soluble fraction most effectively inhibited LPS-stimulated NO production. We also examined the cytotoxicity of G. affine in three normal cell lines: RAW264.7, HEK293, and HaCaT. Cell viability assays showed that the methanol extract and chloroform-soluble fraction of G. affine had no cytotoxic effect on normal cell lines. The expression of pro-inflammatory mediators was also investigated. Western blotting and immunofluorescence showed that G. affine reduces the expression of iNOS, COX-2, and MAPKs, as well as activation of NF-κB in LPS-stimulated RAW264.7 cells. RT-PCR showed that G. affine also negatively regulates inflammatory cytokines at the gene expression level. Taken together, G. affine exerts its anti-inflammatory activity through inhibition of NO generation as a result of inhibiting NF-κB and MAPKs-related inflammatory signaling pathways. In addition, the result of GC-MS analysis revealed the presence of nineteen different types of constituents including guaiacol in the chloroform-soluble fraction of G. affine.
Subject(s)
Anti-Inflammatory Agents/pharmacology , Gnaphalium/chemistry , Mitogen-Activated Protein Kinases/metabolism , NF-kappa B/metabolism , Phytochemicals/pharmacology , Signal Transduction/drug effects , Animals , Anti-Inflammatory Agents/chemistry , Blotting, Western , Cell Line , Cell Survival/drug effects , Cyclooxygenase 2/genetics , Cyclooxygenase 2/metabolism , Dinoprostone/analysis , Enzyme-Linked Immunosorbent Assay , Gas Chromatography-Mass Spectrometry , Gnaphalium/metabolism , HEK293 Cells , Humans , Lipopolysaccharides/toxicity , Macrophages/cytology , Macrophages/drug effects , Macrophages/metabolism , Mice , Microscopy, Fluorescence , Mitogen-Activated Protein Kinases/antagonists & inhibitors , NF-kappa B/antagonists & inhibitors , Nitric Oxide/metabolism , Nitric Oxide Synthase Type II/genetics , Nitric Oxide Synthase Type II/metabolism , Phosphorylation/drug effects , Phytochemicals/chemistry , RAW 264.7 Cells , Real-Time Polymerase Chain ReactionABSTRACT
Objective: To study the chemical constituents from Gnaphalium affine and their anti-oxidative activities. Methods: Chemical constituents were isolated by column chromatography and semi-prepared HPLC, the structures were elucidated by spectral data and physicochemical properties. DPPH free radical scavenging activities of compounds 1,3-11, and 16 were determined. Results: Seventeen compounds were isolated and respectively identified as apigenin (1), dihydroapigenin (2), luteolin (3), chrysin (4), wogonin (5), stimasterol (6), β-sitosterol (7), ursolic acid (8), oleanolic acid (9), 19α-hydroxyl-oleanolic acid (10), 2α, 3α, 19α-trihydroxy-28-norurs-12ene (11), α-amyrin acetate (12), β-amyrin acetate (13), patriscabratine (14), aurantiamide acetate (15), 4'-hydroxydehydrokawain (16), and isovanillin (17). Compounds 1 and 3-5 had significant anti-oxidative activities. Conclusion: Compounds 2,4,5,11-15, and 17 are isolated from G. affine for the frist time.
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ETHNOPHARMACOLOGICAL RELEVANCE: Gnaphalium affine D. Don (GA) has been traditionally used as a medicinal herb in China for the treatment of many ailments including rheumatoid arthritis. However, the anti-arthritic mechanism of GA has still not been demonstrated. This study aims to reveal the anti-inflammatory activity and anti-arthritic mechanism of ethanol extract of G. affine D. Don. MATERIALS AND METHODS: Anti-inflammatory potential of GA was analyzed in vivo in carrageenan induced mice paw edema (acute study). Also, in vivo study was applied in collagen-induced arthritis (CIA) rats. In vitro experiments for analyzing the anti-inflammatory potential of GA were performed on rat alveolar macrophages cell line (NR8383). Analysis of nitric oxide release in NR8383 cells was done by Griess reaction. RT-PCR and western blotting experiment was performed to analyze the expression of phosphorylated p65 and IκBα/ß-actin in NF-κB pathway. The production of TNF-α, IL-1ß, and COX-2 in NR8383 cells were measured by enzyme-linked immunosorbent assay. The chemical profile of GA was analyzed by HPLC-VWD. RESULTS: GA significantly reduced the paw volume in carrageenan induced rat paw edema rat at different doses (300 and 600 mg/kg), compared with the standard indomethacin treatment. In CIA, GA can obviously ameliorate the inflammatory symptom, including cytokine, histological symptom and paw swelling. In the vitro study, GA was able to reduce the nitric oxide (NO) levels in NR8383 cells that had been stimulated with lipopolysaccharide (LPS). The level of TNF-α, IL-1ß, and COX-2 was also decreased with GA treatment in NR8383 cells that had been stimulated with lipopolysaccharide (LPS). Interestingly, GA was found to decrease the level of phosphorylated p65 and IκBα in NR8383 cells. Fifteen compounds were identified by HPLC-VWD with the reference substances and verified by LC-MS. CONCLUSIONS: The results of the experiment scientifically validated its traditional use in inflammatory conditions.
Subject(s)
Anti-Inflammatory Agents/pharmacology , Anti-Inflammatory Agents/therapeutic use , Gnaphalium , Plant Extracts/pharmacology , Plant Extracts/therapeutic use , Animals , Arthritis, Experimental/blood , Arthritis, Experimental/drug therapy , Arthritis, Experimental/pathology , Carrageenan , Cell Line , Edema/drug therapy , Edema/pathology , Female , Foot/pathology , Interleukin-1beta/blood , Interleukin-1beta/metabolism , Knee Joint/drug effects , Knee Joint/pathology , Male , Mice, Inbred ICR , NF-kappa B/metabolism , Nitric Oxide/metabolism , Phytotherapy , Rats, Wistar , Tumor Necrosis Factor-alpha/blood , Tumor Necrosis Factor-alpha/metabolismABSTRACT
The antioxidant activity of Gnaphalium affine extract (GAE) against H2O2-induced oxidative injury in Caco-2 cells was evaluated, and the main antioxidant component was isolated and identified by column chromatography, high performance liquid chromatography, time-of-flight mass spectrometer and nuclear magnetic resonance. In vitro assays, GAE showed remarkable antioxidant activity to scavenge free radicals (ABTS, DPPH, superoxide and hydroxyl radicals), inhibit lipid peroxidation and show reducing power. In food system, GAE exhibited the obvious capacity to inhibit the oxidation of peanut oil and lard, which may be attributed to its high content of phenolic compounds. Moreover, GAE could effectively protect Caco-2 cell against H2O2-induced oxidative injury. With the isolation and purification by chromatography, quercetin was identified as the main antioxidant component of GAE, which was capable of scavenging ABTS, DPPH, superoxide and hydroxyl radicals. These results suggest that G. affine is a potential source for preparing functional foods and nutraceuticals in food industry.