Inhibitory Potential of Quercetin Derivatives Isolated from the Aerial Parts of Siegesbeckia pubescens Makino against Bacterial Neuraminidase.

This study aimed to isolate bacterial neuraminidase (BNA) inhibitory O-methylated quercetin derivatives from the aerial parts of S. pubescens. All the isolated compounds were identified as O-methylated quercetin (1-4), which were exhibited to be noncompetitive inhibitors against BNA, with IC50 ranging from 14.0 to 84.1 µM. The responsible compounds (1-4) showed a significant correlation between BNA inhibitory effects and the number of O-methyl groups on quercetin; mono (1, IC50 = 14.0 µM) > di (2 and 3, IC50 = 24.3 and 25.8 µM) > tri (4, IC50 = 84.1 µM). In addition, the binding affinities between BNA and inhibitors (1-4) were also examined by fluorescence quenching effect with the related constants (KSV, KA, and n). The most active inhibitor 1 possessed a KSV with 0.0252 × 105 L mol-1. Furthermore, the relative distribution of BNA inhibitory O-methylated quercetins (1-4) in S. pubescens extract was evaluated using LC-Q-TOF/MS analysis.

Siegesbeckiae Herba Extract and Chlorogenic Acid Ameliorate the Death of HaCaT Keratinocytes Exposed to Airborne Particulate Matter by Mitigating Oxidative Stress.

Airborne particulate matter with a size of 10 µm or less (PM10) can cause oxidative damages and inflammatory reactions in the skin. This study was conducted to discover natural products that are potentially useful in protecting the skin from PM10. Among the hot water extracts of a total of 23 medicinal plants, Siegesbeckiae Herba extract (SHE), which showed the strongest protective effect against PM10 cytotoxicity, was selected, and its mechanism of action and active constituents were explored. SHE ameliorated PM10-induced cell death, lactate dehydrogenase (LDH) release, lipid peroxidation, and reactive oxygen species (ROS) production in HaCaT cells. SHE decreased the expression of KEAP1, a negative regulator of NRF2, and increased the expression of NRF2 target genes, such as HMOX1 and NQO1. SHE selectively induced the enzymes involved in the synthesis of GSH (GCL-c and GCL-m), the regeneration of GSH (GSR and G6PDH), and GSH conjugation of xenobiotics (GSTκ1), rather than the enzymes that directly scavenge ROS (SOD1, CAT, and GPX1). SHE increased the cellular content of GSH and mitigated the oxidation of GSH to GSSG caused by PM10 exposure. Of the solvent fractions of SHE, the n-butyl alcohol (BA) fraction ameliorated cell death in both the absence and presence of PM10. The BA fraction contained a high amount of chlorogenic acid. Chlorogenic acid reduced PM10-induced cell death, LDH release, and ROS production. This study suggests that SHE protects cells from PM10 toxicity by increasing the cellular antioxidant capacity and that chlorogenic acid may be an active phytochemical of SHE.

Kirenol, darutoside and hesperidin contribute to the anti-inflammatory and analgesic activities of Siegesbeckia pubescens makino by inhibiting COX-2 expression and inflammatory cell infiltration.

ETHNOPHARMACOLOGICAL RELEVANCE: The Chinese traditional medicine of Siegesbeckia pubescens Makino (SM), which has the effect of healing rheumatism and promoting joint health, is often used to treat rheumatoid arthritis and ischemic stroke. AIM OF THE STUDY: To clarify the mechanisms underlying the anti-inflammatory and analgesic influence of active components in the ethanol extract of Siegesbeckia pubescens Makino (ESM). MATERIALS AND METHODS: The active ingredients in the ESM were identified practicing high-performance liquid chromatography-diode array detection (HPLC-DAD). Four models including xylene-induced ear oedema, complete Freund's adjuvant (CFA)-induced hind paw oedema, acetic acid-induced pain writhing and lipopolysaccharide (LPS)-induced RAW264.7 cell migration, were used to clarify the anti-inflammatory and analgesic mechanisms of the active ingredients in the ESM. RESULTS: (1) Three active ingredients of kirenol, darutoside and hesperidin were identified in the ESM, with relative proportion of 0.6%, 0.2% and 0.01%, respectively; hesperidin was reported for the first time in the ESM. (2) Both the ESM and its active ingredients could effectively alleviate the degree of swelling of the auricle and toes, increase the threshold of heat pain, decrease the overexpression of inflammatory protein cyclooxygenase-2 (COX-2) in the skin tissue of the tested parts of the toes, and reduce the number of writhes induced by acetic acid in mice. (3) ESM and its active ingredients also dose-dependently inhibited the migration of RAW264.7 cells. CONCLUSIONS: ESM and its active ingredients can effectively attenuate the expression of inflammatory factors induced by chemical inflammation, prevent the infiltration of inflammatory cells, and exert good anti-inflammatory and antinociceptive activities.

A new monoterpenoid glycoside from aerial parts of Siegesbeckia pubescens / 中草药

Objective: To study the chemical constituents of the aerial parts of Siegesbeckia pubescens. Methods: A systematic separation of chemical constituents was conducted. Detailed chemical investigation of S. pubescens led to the isolation of six compounds by comprehensive chromatographic Methods: (Extraction, Silic gel C.C, Sephadex LH-20, ODS C. C). The structures of them were fully determined based on spectroscopic analysis including IR, HR-ESI-MS, ESI-MS, 1H-NMR, 13C-NMR, DEPT, HSQC, and HMBC spectrum. Results: A total of six compounds were isolated from the fraction of n-butanol extract of S. pubescens including one new monoterpenoid glycoside named (2Z,5E)-7-hydroxy-3,7-dimethyl-2,5-octadiene-1-O-[α-L-rhamnpyranosyl (1→6)]-β-D-glucopyranoside (1), four known diterpenoid glucosides named pubeside D (2), ent-15,16,19-trihydroxypimar- 8(14)-en-2-one-19-O-β-glucopyranoside (3), ent-15,16-dihydroxypimar-8(14)-en-2-one-19-oic-β-glucopyranoside (4), and neodarutoside (5), and eleutherazine B (6). Conclusion: Compound 1 is a new compound, named as siegeside F, and compound 6 is isolated from this species for the first time.

Siegesbeckia pubescens Makino inhibits Pam3CSK4-induced inflammation in RAW 264.7 macrophages through suppressing TLR1/TLR2-mediated NF-κB activation.

BACKGROUND: Siegesbeckia pubescens Makino (SP) is one of the important plant origins for the anti-inflammatory Chinese herbal medicine of Siegesbeckiae Herba. The current investigations indicated that the anti-inflammatory effects of SP were associated with the toll-like receptors (TLRs)-mediated nuclear factor-κB (NF-κB) and the mitogen-activated protein kinase (MAPK) signaling pathways. METHODS: Raw 264.7 macrophages were pretreated with the 50% ethanol extract of SP (SPE, 50-200 µg/mL) and then co-treated with Pam3CSK4 (200 ng/mL) for another 12 h. The inhibitory effect of SPE on Pam3CSK4-stimulated NO release and post-inflammatory cytokines secretions were determined using Griess reagent and Elisa kits, respectively. The influence of SPE on NF-κB and MAPKs signaling relevant proteins was measured by Western blotting analysis, while the intracellular nitric oxide (NO) generation and NF-κB/p65 nuclear translocation were determined using Leica TCS SP8 laser scanning confocal microscope. Moreover, the effect of SPE on luciferase reporter gene in NF-κB-luc DNA transfected raw 264.7 cells was determined using the Dual-Glo luciferase assay system kit. RESULTS: SPE dose-dependently (50-200 µg/mL) attenuated Pam3CSK4-induced NO release, post-inflammatory cytokines (IL-6, TNF-α and MCP-1) secretions and intracellular NO generation in raw 264.7 cells. Biologically, SPE suppressed Pam3CSK4-induced expressions of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), phosphorylation of NF-κB/p65 and IκBα, but did not significantly show effect on the proteins involved in MAPKs signaling (p38, ERK and JNK). The results were further confirmed by NF-κB-luc reporter gene assay and p65 nuclear translocation assay. CONCLUSIONS: In conclusion, SPE ameliorated Pam3CSK4-induced inflammation in raw 264.7 cells through suppressing TLR 1/2-mediated NF-κB activation.

Secondary Metabolites of Endophytic Fungus Colletotrichum capsici Derived from Siegesbeckia pubescens Makino / 中国药学杂志

OBJECTIVE: To identify the secondary metabolites of endophytic fungus C. capsici. METHODS: Various column chromatographic methods were used to isolate and purify compounds, such as silica gel, Sephadex LH-20 and semi-prep HPLC. The structures were established on the basis of spectral analysis. RESULTS: Thirteen compounds were isolated and identified as GKK1032(1), altertoxin I(2), ergosta-7, 22-dien-3β, 5α, 6β-triol(3), ergosterol(4), 20-hydroxylergosta-4, 6, 8(14), 22-tetraen-3-one(5), ergosta-7, 22-diene-3, 6-dione(6), isocyathisterol(7), ganodermaside D(8), TCA 3a(9), TCA 3b(10), nicotinamide(11), tyrosol(12) and 4-hydroxybenzaldehyde(13). CONCLUSION: All these compounds are isolated from C. capsici for the first time.

Rapid extraction and analysis method for the simultaneous determination of 21 bioflavonoids in Siegesbeckia pubescens Makino.

A novel and rapid microwave extraction and ultra high performance liquid chromatography coupled with a triple quadrupole-linear ion trap mass spectrometry method was developed and validated for the determination of 21 bioflavonoids in Siegesbeckia pubescens Makino. The optimal conditions for the extraction of flavonoids from Siegesbeckia pubescens Makino involved the use of methanol as the extraction solvent, a microwave temperature of 70°C, an extraction time of 11 min, and a solvent-to-solid ratio of 40 mL/g. Chromatographic separation was achieved on a Waters ACQUITY UPLC BEH C18 column(100 × 2.1 mm, 1.7 µm) with a gradient mobile phase (A: 0.3% v/v aqueous formic acid and B: acetonitrile) at a flow rate of 0.25 mL/min. All calibration curves showed good linearity (r > 0.999) within the test ranges. The method developed was validated with acceptable sensitivity, intra- and interday precision, reproducibility, and extraction recoveries. The validated method was successfully applied to determine the contents of 21 bioflavonoids in Siegesbeckia pubescens Makino from different sources.

Two new oxylipins from Siegesbeckia pubescens / 中草药

Objective: To study the chemical constituents from the aerial parts of Siegesbeckia pubescens. Methods: The compounds were separated and purified by silica gel column chromatography and TLC, and their structures were determined by analyses on the physicochemical properties and spectral data. Results: Three oxylipins were obtained and identified as 3-(myristoyloxy)-2-(isobutyloxy)-4-methylpentanoic acid (1), 3-(myristoyloxy)-2-hydroxy-4-methylpentanoic acid (2), and γ-dodecyl-α-hydroxy-γ-lactone (3) from the ethanol extract of S. pubescens. Conclusion: The two new oxylipins 1 and 2, named siegesbeclipin A and siegesbeclipin B, and known compound 3 is obtained from this plant for the first time.