Multiple Component Pharmacokinetics after Oral Administration of Gnaphalium affine Extract in Rats.

Gnaphalium affine is traditionally used to treat hyperuricemia and gout in China. Recently, the hypouricemic and renal protective effects of G. affine extract (GAD) have been deeply evaluated. However, little is known about the pharmacokinetics (PKs) of bioactive constituents in GAD. This study is aimed at investigating the individual and holistic pharmacokinetics of 10 bioactive components (including caffeic acid, caffeoylquinic acids, and flavonoids) in rats after single and multiple administrations of GAD. GAD is orally dosed to normal male rats at doses of 225, 450, or 900 mg/kg/day for 10 consecutive days and also orally administrated to uric acid nephropathy (UAN) rats at a dose of 900 mg/kg/day for 28 consecutive days. Integrated PKs of multiple components are calculated by area under the curve (AUC)-based weighting approach. All the components show a double-peak phenomenon in terms of their plasma concentration-time curves, suggesting that the components undergo enterohepatic circulation. The integrated AUC increases in a good dose-proportional manner with GAD dose. Compared with that in normal rats, the plasma exposure of caffeic acid and caffeoylquinic acids increases by 2.3- to 4.3-fold after 10-day chronic treatment of 900 mg/kg GAD in UAN rats. Modest drug accumulation is observed after 28-day chronic treatment.

The Derived Components of Gnaphalium hypoleucum DC. Reduce Quorum Sensing of Chromobacterium violaceum.

Gnaphalium hypoleucum DC. was first recorded in the Chinese National Pharmacopoeia "Yi Plant Medicine". There is no detailed report on its main components' activity in suppressing the quorum sensing activity (QS) of bacteria. Our study aimed to screen the main components in extracts of G. hypoleucum DC. in order to measure their effects on bacterial QS activity and to explore specific quorum sensing mechanisms that are affected by G. hypoleucum DC. extracts. Crude extracts of G. hypoleucum DC. contained significant amounts of two compounds shown to inhibit bacterial QS activity, namely apigenin and luteolin. Apigenin and luteolin in crude extracts of G. hypoleucum DC. showed substantial inhibition of pigment formation, biofilm production, and motility in Chromobacterium violaceum ATCC 12472 compared to the effects of other phytochemicals from G. hypoleucum DC. Apigenin and luteolin exhibited a strong QS inhibitory effect on C. violaceum, interfering with the violacein pigment biosynthesis by downregulating the vioB, vioC, and vioD genes. In the presence of signal molecules, the QS effect is prevented, and the selected compounds can still inhibit the production of the characteristic purple pigment in C. violaceum. Based on qualitative and quantitative research using genomics and bioinformatics, we concluded that apigenin and luteolin in crude extracts of G. hypoleucum DC can interfere with the generation of QS in C. violaceum by downregulating the vioB, vioC, and vioD genes. Indeed, G. hypoleucum DC. is used for the treatment of bacterial infections, and this research provides new ideas and potential alternative uses for medicinal plants.

The wild plant Gnaphalium lavandulifolium as a sentinel for biomonitoring the effects of environmental heavy metals in the metropolitan area of México Valley.

Biomonitoring is a valuable tool for assessing the presence and effects of air pollutants such as heavy metals (HM); due to their toxicity and stability, these compounds can affect human health and the balance of ecosystems. To assess its potential as a sentinel organism of HM pollution, the wild plant Gnaphalium lavandulifolium was exposed to four sites in the metropolitan area of México Valley (MAMV): Altzomoni (ALT) Coyoacán (COY), Ecatepec (ECA), and Tlalnepantla (TLA) during 2, 4, and 8 weeks, between October and November 2019. Control plants remained under controlled conditions. The chemical analysis determined twelve HM (Al, As, Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, V, and Zn) in the leaves. Macroscopic damage to the leaves, later determined in semi-thin sections under light microscopy, lead to a finer analysis. Transmission electron microscope (TEM) showed major structural changes: chromatin condensation, protoplast shrinkage, cytoplasm vacuolization, cell wall thinning, decreased number and size of starch grains, and plastoglobules in chloroplasts. All these characteristics of stress-induced programed cell death (sPCD) were related to the significant increase of toxic HM in the leaves of the exposed plants compared to the control (p < 0.05). Immunohistochemistry revealed a significant amount of proteases with caspase 3-like activity in ECA and TLA samples during long exposure times. Ultrastructural changes and sPCD features detected confirmed the usefulness of G. lavandulifolium as a good biomonitor of HM contamination. They supported the possibility of considering subcellular changes as markers of abiotic stress conditions in plants.

Caffeoylquinic acid derivatives rich extract from Gnaphalium pensylvanicum willd. Ameliorates hyperuricemia and acute gouty arthritis in animal model.

BACKGROUND: The Gnaphalium pensylvanicum willd. is used in China as a folk medicine to treat anti-inflammatory, cough and rheumatism arthritis. The aim of this study was to evaluate the potential of the extract of G. pensylvanicum to treat hyperuricemia and acute gouty arthritis in animal model. METHODS: G. pensylvanicum 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. pensylvanicum extract showed activity in reducing serum uric acid (Sur) through effect renal glucose transporter 9 (GLUT9), organic anion transporter 1 (OAT1) and urate transporter 1 (URAT1) mainly and inhibited XO activity in vivo of mice with PO induced hyperuricemia. The extract of G. pensylvanicum also showed significant anti-inflammatory activity and reduced the paw swelling on MSU crystal-induced paw edema model. Meanwhile, 13 caffeoylquinic acid derivatives and 1 flavone were identified by UPLC-ESI-MS/MS as the main active component of G. pensylvanicum. CONCLUSIONS: The extract of G. pensylvanicum showed significant effect on evaluated models and therefore may be active agents for the treatment of hyperuricemia and acute gouty arthritis.

Screening of Potential Xanthine Oxidase Inhibitors in Gnaphalium hypoleucum DC. by Immobilized Metal Affinity Chromatography and Ultrafiltration-Ultra Performance Liquid Chromatography-Mass Spectrometry.

In this study, a new method based on immobilized metal affinity chromatography (IMAC) combined with ultrafiltration-ultra performance liquid chromatography-mass spectrometry (UF-UPLC-MS) was developed for discovering ligands for xanthine oxidase (XO) in Gnaphalium hypoleucum DC., a folk medicine used in China for the treatment of gout. By IMAC, the high flavonoid content of G. hypoleucum could be determined rapidly and efficiently. UF-UPLC-MS was used to select the bound xanthine oxidase ligands in the mixture and identify them. Finally, two flavonoids, luteolin-4'-O-glucoside and luteolin, were successfully screened and identified as the candidate XO inhibitors of G. hypoleucum. They were evaluated in vitro for XO inhibitory activity and their interaction mechanism was studied coupled with molecular simulations. The results were in favor of the hypothesis that the flavonoids of G. hypoleucum might be the active content for gout treatment by inhibiting XO.

Inhibition of xanthine oxidase activity by gnaphalium affine extract.

OBJECTIVE: To evaluate the inhibitory effect of Gnaphalium affine extracts on xanthine oxidase (XO) activity in vitro and to analyze the mechanism of this effect. METHODS: In 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. RESULTS: Four 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. CONCLUSIONS: These 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.

Cadmium accumulation characteristics of the winter farmland weeds Cardamine hirsuta Linn. and Gnaphalium affine D. Don.

In a preliminary study, we found that the cadmium (Cd) concentrations in shoots of the winter farmland weeds Cardamine hirsuta Linn. and Gnaphalium affine D. Don exceeded the critical value of a Cd-hyperaccumulator (100 mg kg(-1)), indicating that these two farmland weeds might be Cd-hyperaccumulators. In this study, we grew these species in soil containing various concentrations of Cd to further evaluate their Cd accumulation characteristics. The biomasses of C. hirsuta and G. affine decreased with increasing Cd concentrations in the soil, while the root/shoot ratio and the Cd concentrations in shoot tissues increased. The Cd concentrations in shoots of C. hirsuta and G. affine reached 121.96 and 143.91 mg kg(-1), respectively, at the soil Cd concentration of 50 mg kg(-1). Both of these concentrations exceeded the critical value of a Cd-hyperaccumulator (100 mg kg(-1)). The shoot bioconcentration factors of C. hirsuta and G. affine were greater than 1. The translocation factor of C. hirsuta was less than 1 and that of G. affine was greater than 1. These findings indicated that C. hirsuta is a Cd-accumulator and G. affine is Cd-hyperaccumulator. Both plants are distributed widely in the field, and they could be used to remediate Cd-contaminated farmland soil in winter.

Chemical profiling and quantification of flavones in several Pseudognaphalium and Gnaphalium species of Mexican gordolobo using UHPLC/PDA/MS.

The inflorescences of the Mexican gordolobo are used as a folk medicine to treat various respiratory diseases. Currently, the botanical species that bear the name Mexican gordolobo belong to the genera Gnaphalium and Pseudognaphalium. Despite a long history of traditional use, most Mexican gordolobo species have never been fully chemically characterized, and the range of constituents in the species has not been comprehensively reported. To establish a quality control and chemical characterization method, a total of 49 samples belonging to 18 species of Pseudognaphalium and four species of Gnaphalium were studied. Nine flavones were quantified using a UPLC-PDA method. The method was validated in terms of linearity (R2 > 0.99), precision (intra- and inter-day: 0.1-3.9%), accuracy (96-103%), detection limit (10 ng/mL), limit of quantification (25 ng/mL) and robustness. 3-Methylquercetin, luteolin, quercetin, 3,5-dihydroxy-6,7,8-trimethoxyflavone, apigenin and gnaphaliin A were present at relatively high levels in most of the samples analyzed. The samples of P. oxyphyllum and P. liebmannii showed the highest content of the 9 compounds analyzed. Whereas the samples of the 5 species of Gnaphalium showed the lowest levels, including non-detectable, of the 9 compounds quantified. This marks an important difference with Pseudognaphalium species. Furthermore, using UHPLC-ESI-QToF data with targeted and non-targeted approaches, 57 compounds, were identified in Mexican gordolobo samples. Flavonoids were the main group of compounds found in Mexican gordolobo.

The genus Gnaphalium L. (Compositae): phytochemical and pharmacological characteristics.

The genus Gnaphalium, a herb distributed worldwide, comprises approximately 200 species of the Compositae (Asteraceae) family that belongs to the tribe Gnaphalieae. Some species are traditionally used as wild vegetables and in folk medicine. This review focuses on the phytochemical investigations and biological studies of plants from the genus Gnaphalium over the past few decades. More than 125 chemical constituents have been isolated from the genus Gnaphalium, including flavonoids, sesquiterpenes, diterpenes, triterpenes, phytosterols, anthraquinones, caffeoylquinic acid derivatives, and other compounds. The extracts of this genus, as well as compounds isolated from it, have been demonstrated to possess multiple pharmacological activities such as antioxidant, antibacterial and antifungal, anti-complement, antitussive and expectorant, insect antifeedant, cytotoxic, anti-inflammatory, antidiabetic and antihypouricemic properties. The present review compiles the information available on this genus because of its relevance to food and ethnopharmacology and the potential therapeutic uses of these species.

Two new phenolic glycosides from Gnaphalium affine D. Don and their anti-complementary activity.

Two new phenolic glycosides, named gnaphaffine A and B (compounds 1 and 2), were isolated from Gnaphalium affine. together with six known compounds, including caffeic acid (3), everlastoside L (4), isorhamnetin-7-O-ß-D-glucopyranoside (5), quercetin- 3-O-ß-D-glucopyranoside (6), scutellarein-7-O-ß-D-glucoside (7) and api-genin-7-O-ß-D- glucopyranoside (8). Their structures were elucidated by spectroscopic methods, including ESI-MS and 2D NMR spectroscopy (HMQC and HMBC). All compounds were evaluated for their anti-complementary activity on the classical pathway of the complement system in vitro.

Methylobacterium gnaphalii sp. nov., isolated from leaves of Gnaphalium spicatum.

A pink-pigmented, facultatively methylotrophic bacterium, strain 23e(T), was isolated from the leaves of Gnaphalium spicatum (cudweed). The cells of strain 23e(T) were Gram-reaction negative, motile and non-spore-forming rods. On the basis of 16S rRNA gene sequence similarities, strain 23e(T) was related to Methylobacterium organophilum ATCC 27886(T) (97.1%) and Methylobacterium marchantiae JT1(T) (97%), and the phylogenetic similarities to all other Methylobacterium species with validly published names were less than 97%. Major cellular fatty acids were C(18:1)ω7c, C(16:00) and C(18:0). The results of DNA-DNA hybridization, phylogenetic analyses based on 16S rRNA and cpn60 gene sequences, fatty acid profiles, whole-cell matrix-assisted laser desorption/ionization time of flight/MS analysis, physiological and biochemical tests allowed genotypic and phenotypic differentiation of strain 23e(T) from the phylogenetically closest relatives. We propose that strain 23e(T) represents a novel species within the genus Methylobacterium, for which the name Methylobacterium gnaphalii sp. nov. is proposed. The type strain is 23e(T) (=DSM 24027(T)=NBRC 107716(T)).

[Study on the chemical constituents from Gnaphalium hypoleucum].

OBJECTIVE: To research the chemical constituents from Gnaphalium hypoleucum. METHODS: Compounds were separated by using silica gel, ODS, Sephadex LH-20 chromatography and their structures were elucidated by means of spectral data analysis. RESULTS: Ten compounds were isolated and identified as tetracosanoic acid (1), 5-hydroxy-3,6,7,8-tetramethoxyflavone (2), beta-sitosterol (3), 5-hydroxy-3,6,7,8,4'-pentamethoxyflavone (4), 5-hydroxy-3,6, 7,8,3',4'-hexamethoxyflavone (5), apigenin (6), luteolin (7), quercetin (8), luteolin4'-O-beta-D-glucoside (9), quercetin-4'-O-beta-D-glucoside (10). CONCLUSION: All of these compounds are isolated from this plant for the first time. Compound 5 is isolated from the genus for the first time.

Beneficial herb-drug interaction of Gnaphalium affine extract on benzbromarone: A pharmacokinetic and pharmacodynamic study in rats.

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.

Isolation and characterisation of antibacterial and anti-inflammatory compounds from Gnaphalium polycaulon.

ETHNO-PHARMACOLOGICAL RELEVANCE: Gnaphalium polycaulon commonly known as "cudweed" has been used throughout South America as an infusion to treat colds, bronchitis, fever or pneumonia. AIM OF THE STUDY: This study aimed to determine the antibacterial and anti-inflammatory activities of the aqueous extract of Gnaphalium polycaulon and identify the related compounds. MATERIALS AND METHODS: A bio-guided isolation of the active compounds of Gnaphalium polycaulon was carried out, selecting the fractions depending on their antibacterial, anti-inflammatory and cytotoxic activities. The antibacterial effect was studied against Haemophilus influenzae, Moraxella catarrhalis, Staphylococcus aureus and Streptococcus pneumoniae; and the anti-inflammatory study was performed by measuring the inhibition of NF-κB in BEAS-2B and IMR-90 cell cultures. RESULTS: Three compounds were obtained and characterised by nuclear magnetic resonance and mass spectrometry. These compounds are 2-(4-(1-H-tetrazol-1-yl) phenyl)-2-aminopropanoic acid (1), N-phenyl-4-(3-phenyl-1,2,4-thiadiazol-5-yl) piperazine-1-carboxamide (2) and N-(4-ethoxyphenyl)-4-(2-methylimidazo-[1,2-α] pyridine-3-yl) thiazol-2-amine (3). All compounds showed antibacterial activity with MIC values of 44.80-44.85, 0.017-0.021 and 0.0077-0.0079 µM, respectively, in the Haemophilus influenzae, Moraxella catarrhalis, Staphylococcus aureus and Streptococcus pneumoniae strains, while the positive control, Ofloxacin, had a MIC value of 27.64-27.67 µM. This was corroborated through a zone inhibition assay, where compound 3 (11.36-11.67 mm) was much more active than the positive control (Ofloxacin, 23.41-24.12 mm), while compounds 2 (26.47-27.64 mm) and 1 (28.39-29.76 mm) displayed similar antibacterial potential to the positive control. Finally, all the compounds presented NF-κB inhibitory activity, compounds 3 (IC50 = 0.0071-0.0073 µM) and 2 (IC50 = 0.016-0.019 µM) being the most promising. Compound 1 (IC50 = 44.24-44.26 µM) had less anti-inflammatory potential, being also the closest to the values displayed by the positive control (Celastrol, IC50 = 7.41 µM). CONCLUSION: In the present study, three compounds were isolated for the first time from the aqueous extract of Gnaphalium polycaulon. Their antibacterial and anti-inflammatory potential was tested and showcased.

Apomixis is not prevalent in subnival to nival plants of the European Alps.

BACKGROUND AND AIMS: High alpine environments are characterized by short growing seasons, stochastic climatic conditions and fluctuating pollinator visits. These conditions are rather unfavourable for sexual reproduction of flowering plants. Apomixis, asexual reproduction via seed, provides reproductive assurance without the need of pollinators and potentially accelerates seed development. Therefore, apomixis is expected to provide selective advantages in high-alpine biota. Indeed, apomictic species occur frequently in the subalpine to alpine grassland zone of the European Alps, but the mode of reproduction of the subnival to nival flora was largely unknown. METHODS: The mode of reproduction in 14 species belonging to seven families was investigated via flow cytometric seed screen. The sampling comprised 12 species typical for nival to subnival plant communities of the European Alps without any previous information on apomixis (Achillea atrata, Androsace alpina, Arabis caerulea, Erigeron uniflorus, Gnaphalium hoppeanum, Leucanthemopsis alpina, Oxyria digyna, Potentilla frigida, Ranunculus alpestris, R. glacialis, R. pygmaeus and Saxifraga bryoides), and two high-alpine species with apomixis reported from other geographical areas (Leontopodium alpinum and Potentilla crantzii). KEY RESULTS: Flow cytometric data were clearly interpretable for all 46 population samples, confirming the utility of the method for broad screenings on non-model organisms. Formation of endosperm in all species of Asteraceae was documented. Ratios of endosperm : embryo showed pseudogamous apomixis for Potentilla crantzii (ratio approx. 3), but sexual reproduction for all other species (ratios approx. 1·5). CONCLUSIONS: The occurrence of apomixis is not correlated to high altitudes, and cannot be readily explained by selective forces due to environmental conditions. The investigated species have probably other adaptations to high altitudes to maintain reproductive assurance via sexuality. We hypothesize that shifts to apomixis are rather connected to frequencies of polyploidization than to ecological conditions.

3-Hydroxydihydrobenzofuran glucosides from Gnaphalium polycaulon.

A new 3-hydroxydihydrobenzofuran glucoside, gnaphaliol 9-O-ß-D-glucopyranoside (2), was isolated from the aerial parts of Gnaphalium polycaulon together with 1-{(2R*,3S*-3-(ß-D-glucopyranosyloxy)-2,3-dihydro-2-[1-(hydroxyl methyl)vinyl]-1-benzofuran-5-yl}-ethanone or gnaphaliol 3-O-ß-D-glucopyranoside (1), (Z)-3-hexenyl O-ß-D-glucopyranoside (3) and adenosine (4). The absolute configurations at C-2 and C-3 positions of compound 1 were determined to be 2R and 3R. The structures of these compounds were elucidated on the basis of their physical and spectroscopic data.

Development and validation of a column high-performance liquid chromatography method for quantification of ganaphaliin A and B in inflorescences of Gnaphalium liebmannii Sch. Bp ex Klatt.

An HPLC method was developed for the simultaneous determination of gnaphaliin A and B, active compounds of Gnaphalium liebmannii Sch. Bp ex Klatt. The HPLC separation was performed on an Inertsil ODS-3 (150 x 4.6 mm id, 5 microm) RP C18 column operated at 40 degrees C; the isocratic mobile phase was 0.02% aqueous orthophosphoric acid-methanol-acetonitrile (50 + 30 + 20, v/v/v), with a run time of 20 min and flow rate of 1.5 mL/min. Detection with a photodiode array detector (PDAD) was at 270 nm. The method was validated for linearity, repeatability, LOD, and LOQ. The LOD and LOQ for gnaphaliin A and B were found to be in the range of 0.4-0.5 and 1.0-1.4 microg/mL, respectively. This is the first report of an analytical method developed for the quantitative analysis of flavones from Gnaphalium species by HPLC-PDAD with applications for raw material and commercial products.

The extract of Gnaphalium affine D. Don protects against H2O2-induced apoptosis by targeting PI3K/AKT/GSK-3ß signaling pathway in cardiomyocytes.

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.

Mycorrhizal densities decline in association with nonnative plants and contribute to plant invasion.

Belowground interactions between herbaceous native species and nonnative species is a poorly understood but emerging area of interest to invasive-species researchers. Positive feedback dynamics are commonly observed in many invaded systems and have been suspected in California grasslands, where native plants associate strongly with soil mutualists such as arbuscular mycorrhizal fungi. In response to disturbance, invading nonnative plants proliferate, and to the degree these species associate weakly with soil mutualists, we would expect mutualist efficacy to degrade over time. Degraded mutualist efficacy would negatively impact mutualist-dependent native species or their recruitment following a disturbance. We investigated the feedback dynamics of soil conditioned both with native and nonnative herbaceous communities of southern California grasslands to test this degraded mutualist hypothesis. Using a mesocosm approach, we inoculated each community with live soil originating from a remnant native grassland and varied the plant communities (i.e., native or nonnative) along a plant-species-richness gradient. After one year, we then used this conditioned soil for reciprocal feedback tests on a native and nonnative indicator species. We show that a native herbaceous forb (Gnaphalium californicum) grows best in soil conditioned by a diverse mix of other native species that includes G. californicum but is inhibited by soil conditioned by a diverse mix of nonnative species. We also show that an invasive, nonnative herbaceous forb (Carduus pycnocephalus) exhibits strong growth in soil lacking arbuscular mycorrhizal fungi and in soil conditioned by a diverse mix of nonnative species that include C. pycnocephalus, and that it is inhibited by the same soil that best promotes the native, G. californicum. Separate bioassays for mycorrhizal density show a reduction of arbuscular mycorrhizal fungi in the nonnative-conditioned soil relative to the native-conditioned soil, which suggests that nonnative species do not promote the growth of mycorrhizal fungi in the same way that native species do. The growth patterns resulting from the vegetative history of these distinct soil communities provide evidence of a biotic feedback mechanism that may account for the maintenance of persistent communities of nonnative (and often invasive) plants ubiquitous throughout California grasslands.

Solving the confusion of gnaphaliin structure: gnaphaliin A and gnaphaliin B identified as active principles of Gnaphalium liebmannii with tracheal smooth muscle relaxant properties.

Inflorescences of Gnaphalium liebmannii, commonly known as "Gordolobo", is the most important remedy in Mexican traditional medicine to treat respiratory diseases, including asthma. By a bioguided fractionation of the n-hexane extract of this plant, following the relaxant effect on guinea pig tracheal smooth muscle, the flavones 5,7-dihydroxy-3,8-dimethoxyflavone (1) and 3,5-dihydroxy-7,8-dimethoxyflavone (2) were identified as the active relaxant compounds. Compounds 1 and 2 showed more potent relaxant properties than aminophylline in this model. Both 1 and 2 have been described as gnaphaliin in the past; here EIMS data, NMR experiments for both compounds, and X-ray diffraction analysis for 1 provided structural information to suggest that 1 and 2 should be named gnaphaliins A and B, respectively.