Chemical constituents isolated from Polygala japonica leaves and their inhibitory effect on nitric oxide production in vitro

A methanolic extract of dried leaves of Polygala japonica Houtt (Polygalaceae) significantly attenuated nitric oxide production in lipopolysaccharide-simulated BV2 microglia. Five anthraquinones chrysophanol (1), emodin (2), aloe-emodin (3), emodin 8-O-β-D-glucopyranoside (4) and trihydroxy anthraquinone (5), and four flavonoids kaempferol (6), chrysoeriol (7), kaempferol 3-gentiobioside (8) and isorhamnetin (9) were isolated from the methanolic extract using bioactivity-guided fractionation. Among them, compounds 1–4, 6 and 7 showed significant inhibitory effect on lipopolysaccharide-induced nitric oxide production in BV2 microglia at the concentrations ranging from 1.0 to 100.0 μM.     

Chemical constituents of Polygala tenuifolia roots and their inhibitory activity on lipopolysaccharide-induced nitric oxide production in BV2 microglia

A methanolic extract of the roots of Polygala tenuifolia (Polygalaceae) significantly attenuated nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated BV2 microglia cells. Five xanthones, 1-hydroxy-7-methoxyxanthone (1), 3,6-dihydroxy-1,2,7-trimethoxyxanthone (2), 1,3,6-trihydroxy-2,7-dimethoxyxanthone (3), 1,7-dihydroxy-2,3-dimethoxyxanthone (4) and 1,7-dihydroxy-3-methoxyxanthone (5), and five phenylpropanoids, 4-hydroxy-3-methoxypropiophenone (6), methyl 4-hydroxy-3-methoxycinnamic acid (7), 3,4,5-trimethoxycinnamic acid (8), 4-methoxycinnamic acid (9) and β-d-(3-O-sinapoyl) fructofuranosyl-α-d-(6-O-sinapoyl)glucopyranoside (10), were isolated from CHCl(3) fraction using bioactivity-guided fractionation. Among these compounds, compounds 1, 2, 4, 5 and 7 showed significant inhibitory effects on LPS-induced NO production in BV2 microglia cells at the concentration ranging from 10.0 to 100.0 μM.     

Inhibitory alkaloids from Dictamnus dasycarpus root barks on lipopolysaccharide-induced nitric oxide production in BV2 cells

The methanolic extract of Dictamnus dasycarpus root barks afforded one new glycosidic quinoline alkaloid, 3-[1β-hydroxy-2-(β-D-glucopyranosyloxy)-ethyl)-4-methoxy-2(1H)-quinolinone (1), together with nine known compounds, preskimmianine (2), 8-methoxy-N-methylflindersine (3), dictamine (4), γ-fagarine (5), halopine (6), skimmianine (7), dictangustine-A (8), iso-γ-fagarine (9), isomaculosidine (10). The isolated alkaloids significantly inhibited nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated BV2 cells. Among them, compounds 3 and 7 showed the most potent inhibitory activities on LPS-induced NO production.     

Inhibition of nitric oxide synthase expression by a methanolic extract of Crescentia alata and its derived flavonols.

In order to validate the use of Crescentia alata (Bignoniaceae) in the traditional medicine of Guatemala as an antiinflammatory remedy, the methanolic (MeOH) extract has been evaluated in vivo for antiinflammatory activity on carrageenin paw edema in rats and in vitro on Escherichia coli lipopolysaccharide- (LPS)-induced nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) expression in J774.A1 macrophage cell line. This extract exerted in vivo a significant anti-inflammatory activity at the highest dose tested. The same extract showed in vitro an inhibitory activity on inducible nitric oxide synthase expression and on NO formation in LPS-primed J774.A1 cells. Subsequent fractionation and analysis of the extract has led to the isolation and characterization as major constituents of two flavonol glycosides: quercetin 3-O-alpha-L-rhamnopyranosyl-(1->6)-beta-D-glucopyranoside (rutin) 1, kaempferol 3-O-alpha-L-rhamnopyranosyl-(1->6)-beta-D-glucopyranoside (kaempferol 3-O-rutinoside) 2, and flavonol aglycone, kaempferol 3. Their structures were elucidated by spectral methods. The bioassay-directed analysis of flavonols 1-3 indicated that kaempferol (3) was the most active compound contained in the MeOH extract because it reduced in vitro both NO production and iNOS expression in LPS-primed J774.A1 cells, whereas rutin (1) and kaempferol 3-O-rutinoside (2) showed no significant activity. The MeOH extract and all of flavonoids tested did not show in vitro significant cytotoxic effect in J774.A1 macrophage cell line.     

Inhibitory alkaloids from Dictamnus dasycarpus root barks on lipopolysaccharide-induced nitric oxide production in BV2 cells

The methanolic extract of Dictamnus dasycarpus root barks afforded one new glycosidic quinoline alkaloid, 3-[1β-hydroxy-2-(β-D-glucopyranosyloxy)-ethyl)-4-methoxy-2(1H)-quinolinone (1), together with nine known compounds, preskimmianine (2), 8-methoxy-N-methylflindersine (3), dictamine (4), γ-fagarine (5), halopine (6), skimmianine (7), dictangustine-A (8), iso-γ-fagarine (9), isomaculosidine (10). The isolated alkaloids significantly inhibited nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated BV2 cells. Among them, compounds 3 and 7 showed the most potent inhibitory activities on LPS-induced NO production.     

Dalesconols B inhibits lipopolysaccharide induced inflammation and suppresses NF-κB and p38/JNK activation in microglial cells

Therapeutic strategies designed to inhibit the activation of microglia may lead to significant advancement in the treatment of most neurodegenerative diseases. Dalesconols B, also termed as TL2, is a newly found polyketide from a mantis-associated fungus and has been reported to exert potent immunosuppressive effects. In the present study, the anti-inflammatory effects of TL2 was investigated in lipopolysaccharide (LPS)-treated BV2 microglia and primary microglia cells. Our observations indicated that pretreatment with TL2 significantly inhibited the production of NO and PGE2 and suppressed the expression of pro-inflammatory mediators such as inducible nitric oxide synthase (iNOS), COX-2, TNF-α, IL-1β, IL-6, MCP-1 and MIP-1α in LPS-stimulated BV2 microglia. The nuclear translocation of NF-κB and the phosphorylation level of Akt, p38 and JNK MAP kinase pathways were also inhibited by TL2 in LPS-treated BV2 microglia. Moreover, TL2 also decreased Aβ-induced production of TNF-α, IL-1β and IL-6 in BV2 microglia. Additionally, TL2 protected primary cortical neurons against microglia-mediated neurotoxicity. Overall, our findings suggested that TL2 might be a promising therapeutic agent for alleviating the progress of neurodegenerative diseases associated with microglia activation.     

Covalent anthocyanin-flavonol complexes from flowers of chive, Allium schoenoprasum

The structures of eight anthocyanins have been determined in acidified methanolic extract of pale-purple flowers of chive, Allium schoenoprasum. Four of them have been identified as the anthocyanin-flavonol complexes (cyanidin 3-O-beta-glucosideAII) (kaempferol 3-O-(2-O-beta-glucosylFIII-beta-glucosideFII)-7-O-beta-gl ucosiduronic acidFIV) malonateAIII (AII-6-->AIII-1, FIV-2-->AIII-3), 1, (cyanidin 3-O-(3-O-acetyl-beta-glucosideAII) (kaempferol 3-O-(2-O-beta-glucosylFIII-beta-glucosideFII)-7-O-beta-gl ucosiduronic acidFIV) malonateAIII (AII-6-->AIII-1, FIV-2-->AIII-3), 2, and their 7-O-(methyl-O-beta-glucosiduronateFIV) analogous, 3 and 4. Pigments 1 and 2 are the first final identification of covalent complexes between an anthocyanin and a flavonol, while 3 and 4 are formed during the isolation process. The other four anthocyanins (5-8) were found to be the 3-acetylglucoside, 3-glucoside, 3-(6-malonylglucoside) and 3-(3,6-dimalonylglucoside) of cyanidin. The three latter pigments have earlier been identified as the major anthocyanins of the chive stem. The covalent anthocyanin-flavonol complexes show intramolecular association between the anthocyanidin (cyanidin) and flavonol (kaempferol) units, which influence the colour.     

Flavonol glycosides of Warburgia ugandensis leaves

Four new flavonol gycosides: kaempferide 3-O-beta-xylosyl (1-->2)-beta-glucoside, kaempferol 3-O-alpha-rhamnoside-7,4'-di-O-beta-galactoside, kaempferol 3,7,4'-tri-O-beta-glucoside and quercetin 3-O-[alpha-rhamnosyl (1-->6)] [beta-glucosyl (1-->2)]-beta-glucoside-7-O-alpha-rhamnoside, were characterized from a methanolic leaf extract of Warburgia ugandensis. The known flavonols: kaempferol, kaempferol 3-rhamnoside, kaempferol 3-rutinoside, myricetin, quercetin 3-rhamnoside, kaempferol 3-arabinoside, quercetin 3-glucoside, quercetin, kaempferol 3-rhamnoside-4'-galactoside, myricetin 3-galactoside and kaempferol 3-glucoside were also isolated. Structures were established by spectroscopic and chemical methods and by comparison with authentic samples.     

胡桃枝的化学成分及抑制一氧化氮生成的作用

以体外测定各化合物对抑制脂多糖(LPS)和γ干扰素(IFNγ)诱导的RAW264.7大鼠巨噬细胞NO的生成量为活性指标,从核桃中分离得到了5个化合物,分别为2乙氧基胡桃醌(1),3乙氧基胡桃醌(2),regiolone(3),(4S)4hydroxyαtetralone(4)和大黄素(5)。化合物1和2为首次从该植物中分离得到,化合物1具有较强的抑制大鼠巨噬细胞NO生成的作用。化合物1和2均为首次作为天然产物得到。     

Triterpene saponins from Randia formosa

Seven new triterpenoid saponins, randiasaponins I (1), II (2), III (3), IV (4), V (5), VI (6) and VII (7) as well as two known ones, ilexoside XXVII (8) and ilexoside XXXVII (9), were isolated from the methanolic extract of the leaves of Randia formosa. The structures of the new saponins were established as 3-O-alpha-L-arabinopyranosyl-3 beta,19 alpha,23-trihydroxyursa-12,20(30)-dien-28-oic acid 28-beta-D-glucopyranosyl ester (1), 3-O-beta-D-glucopyranosyl-(1-->3)-alpha-L-arabinopyranosyl rotundic acid (2), 3-O-beta-D-glucopyranosyl-(1-->3)-alpha-L-arabinopyranosyl pomolic acid 28-beta-D-glucopyranosyl ester (3), 3-O-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl pomolic acid 28-beta-D-glucopyranosyl ester (4), 3-O-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl siaresinolic acid 28-beta-D-glucopyranosyl ester (5), 3-O-alpha-L-arabinopyranosyl ilexosapogenin A 28-beta-D-glucopyranosyl ester (6), and 3-O-beta-D-glucopyranosyl ilexosapogenin A 28-beta-D-glucopyranosyl ester (7), based on spectral and chemical evidence. Besides the saponins, two common flavonoids kaempferol 3-O-rutinoside and rutin were also isolated.     

Inhibitory effects of flavonol glycosides from Cinnamomum osmophloeum on inflammatory mediators in LPS/IFN-gamma-activated murine macrophages

Four kaempferol glycosides were isolated from the leaves of Cinnamomum osmophloeum Kaneh, a Taiwan endemic tree. These compounds namely, kaempferitrin (1), kaempferol 3-O-beta-D-glucopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-7-O-alpha-L-rhamnopyranoside (2), kaempferol 3-O-beta-D-apiofuranosyl-(1-->2)-alpha-L-arabinofuranosyl-7-O-alpha-L-rhamnopyranoside (3), and kaempferol 3-O-beta-D-apiofuranosy-(1-->4)-alpha-L-rhamnopyranosyl-7-O-alpha-L-rhamnopyranoside (4). The structure of compound 2 was determined by spectroscopic analyses and acid hydrolysis. The isolates 1-4 were evaluated as inhibitors of some macrophage functions involved in the inflammatory process. These four compounds inhibited lipopolysaccharide (LPS) and interferon (IFN)-gamma-induced nitric oxide (NO), and cytokines [tumor necrosis factor (TNF)-alpha and interleukin (IL)-12] in a dose-dependent manner. The concentration of 50% inhibition (IC(50)) of NO by compounds 1, 3, 4 were 40, 15, 20microM, respectively. In parallel, these concentrations were approximately in a similar manner to that observed for TNF-alpha and IL-12 production. However, compound 2 inhibited NO and cytokines production by 30% at 100microM concentration. On the other hand, compounds 3 and 4 showed no inhibitory effect on the production of NO from macrophages, when inducible NO synthase was already expressed by the stimulation with LPS and IFN-gamma. Taken together, our results provide evidence that isolates of C. osmophloeum possess an anti-inflammatory potential which constitutes a previously unrecognized biological activity.     

(7<Emphasis Type="Italic">R</Emphasis>,8<Emphasis Type="Italic">S</Emphasis>)-Dehydrodiconiferyl Alcohol Suppresses Lipopolysaccharide-Induced Inflammatory Responses in BV2 Microglia by Inhibiting MAPK Signaling

(7R,8S)-Dehydrodiconiferyl alcohol (DDA), a lignan isolated from the dried stems of Clematis armandii, has been found to exert potential anti-inflammatory activity in vitro. In the present study, we investigated the effects and possible mechanisms of DDA on lipopolysaccharide (LPS)-mediated inflammatory response in murine BV2 microglia. Our results revealed that non-toxic concentrations (6.25–25 μM) of DDA markedly suppressed LPS-induced production of nitric oxide, expression of inducible nitric oxide synthase and cyclooxygenase-2, and release of inflammatory factors, including tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-6 in a concentration dependent manner. In addition, DDA time- and concentration-dependently attenuated LPS-induced phosphorylation of c-Jun N-terminal kinase 1/2 (JNK), but not protein kinase B, p38, or extracellular signal-regulated kinase 1/2. Moreover, DDA significantly suppress LPS-mediated nuclear factor-κB (NF-κB) activation by inhibiting phosphorylation and nuclear translocation of NF-κB p65. Collectively, our results demonstrated that DDA inhibited LPS-stimulated inflammatory response in BV2 cell, at least in part, through inhibition of NF-κB activation and modulation of JNK signaling.     

Flavonol and drimane-type sesquiterpene glycosides of Warburgia stuhlmannii leaves

An investigation of methanolic extract of Warburgia stuhlmannii leaves has led to the isolation of two new drimane-type sesquiterpene glycosides characterized as mukaadial 6-O-beta-D-glucopyranoside, mukaadial 6-O-alpha-L-rhamnopyranoside together with two other novel flavonol glycosides identified as 3',5'-O-dimethylmyricetin 3-O-beta-D-2",3"-diacetylglucopyranoside and 3'-O-methylquercetin 3-O-beta-D-2",3",4"-triacetylglucopyranoside. The known compounds; mukaadial, deacetylugandensolide, quercetin, kaempferol, kaempferol 3-O-alpha-L-rhamnopyranoside, quercetin 3-O-beta-D-glucopyranoside, kaempferol 7-O-beta-D-glucopyranoside, myricetin 3-O-alpha-L-rhamnopyranoside, quercetin 3-O-alpha-L-rhamnopyranoside, quercetin 3-O-sophoroside and isorhamnetin 3-O-beta-D-glucopyranoside were also isolated from the same extract.     

Anti-neuroinflammatory Effect of Emodin in LPS-Stimulated Microglia: Involvement of AMPK/Nrf2 Activation

AMPK/Nrf2 signaling regulates multiple antioxidative factors and exerts neuroprotective effects. Emodin is one of the main bioactive components extracted from Polygonum multiflorum, a plant possessing important activities for human health and for treating a variety of diseases. This study examined whether emodin can activate AMPK/Nrf2 signaling and induce the expression of genes targeted by this pathway. In addition, the anti-neuroinflammatory properties of emodin in lipopolysaccharide (LPS)-stimulated microglia were examined. In microglia, the emodin treatment increased the levels of LKB1, CaMKII, and AMPK phosphorylation. Emodin increased the translocation and transactivity of Nrf2 and enhanced the levels of HO-1 and NQO1. In addition, the emodin-mediated expression of HO-1 and NQO1 was attenuated completely by an AMPK inhibitor (compound C). Moreover, emodin decreased dramatically the LPS-induced production of NO and PGE₂ as well as the protein expression and promoter activity of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). In addition, emodin effectively inhibited the production of pro-inflammatory cytokines, TNF-α and IL-6, and reduced the level of IκBα phosphorylation, leading to the suppression of the nuclear translocation, phosphorylation, and transactivity of NF-κB. Emodin also suppressed the LPS-stimulated activation of STATs, JNK, and p38 MAPK. The anti-inflammatory effects of emodin were reversed by transfection with Nrf-2 and HO-1 siRNA and by a co-treatment with an AMPK inhibitor. These results suggest that emodin isolated from P. multiflorum can be used as a natural anti-neuroinflammatory agent that exerts its effects by inducing HO-1 and NQO1 via AMPK/Nrf2 signaling in microglia.     

Neolignans from Piper kadsura and their anti-neuroinflammatory activity

Bioassay-guided column chromatographic separation of the methanolic extract of dried aerial parts of Piper kadsura (Piperaceae) led to the isolation of a new neolignan, piperkadsin C (1), together with eight known neolignans (2–9). The structures of the isolated compounds were elucidated by combined spectroscopic methods. The anti-neuroinflammatory activities of these compounds were evaluated by assessing nitric oxide (NO) production in LPS-activated BV-2 cells, a microglia cell line. Piperkadsin C (1) and futoquinol (2) potently inhibited NO production with an IC₅₀ value of 14.6 and 16.8 μM in microglia cells, respectively. Compounds 3, 4, 5, 8, and 9 also exhibited moderate inhibition of NO production in BV-2 cells.     

Isoflavonoid glycosides and rotenoids from Pongamia pinnata leaves

Chromatographic separation of a 70% aqueous methanol extract (AME) of Pongamia pinnata (Linn.) Pierre (Leguminosae) leaves has led to the isolation of two new isoflavonoid diglycosides, 4'-O-methyl-genistein 7-O-beta-D-rutinoside (2) and 2',5'-dimethoxy-genistein 7-O-beta-D-apiofuranosyl-(1"'-->6")-O-beta-D-glucopyranoside (6), and a new rotenoid, 12a-hydroxy-alpha-toxicarol (5), together with nine known metabolites, vecinin-2 (1), kaempferol 3-O-beta-D-rutinoside (3), rutin (4), vitexin (7), isoquercitrin (8), kaempferol 3-O-beta-D-glucopyranoside (9), 11,12a-dihydroxy-munduserone (10), kaempferol (11), and quercetin (12). Their structures were elucidated on the basis of chemical and spectroscopic analyses.     

Anti-inflammatory effects of spermidine in lipopolysaccharide-stimulated BV2 microglial cells

Background

Spermidine, a naturally occurring polyamine, displays a wide variety of internal biological activities including cell growth and proliferation. However, the molecular mechanisms responsible for its anti-inflammatory activity have not yet been elucidated.

Methods

The anti-inflammatory properties of spermidine were studied using lipopolysaccharide (LPS)-stimulated murine BV2 microglia model. As inflammatory parameters, the production of nitric oxide (NO), prostaglandin E₂ (PGE₂), interleukin (IL)-6 and tumor necrosis factor (TNF)-α were evaluated. We also examined the spermidine''s effect on the activity of nuclear factor-kappaB (NF-κB), and the phosphoinositide 3-kinase (PI3K)/Akt and mitogen-activated protein kinases (MAPKs) pathways.

Results

Pretreatment with spermidine prior to LPS treatment significantly inhibited excessive production of NO and PGE₂ in a dose-dependent manner, and was associated with down-regulation of expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Spermidine treatment also attenuated the production of pro-inflammatory cytokines, including IL-6 and TNF-α, by suppressing their mRNA expressions. The mechanism underlying spermidine-mediated attenuation of inflammation in BV2 cells appeared to involve the suppression of translocation of NF-κB p65 subunit into the nucleus, and the phosphorylation of Akt and MAPKs.

Conclusions

The results indicate that spermidine appears to inhibit inflammation stimulated by LPS by blocking the NF-κB, PI3K/Akt and MAPKs signaling pathways in microglia.     

A new kaempferol triglycoside from Fagonia taeckholmiana: cytotoxic activity of its extracts

In addition to apigenin, apigenin 7-O-glucoside, kaempferol 3-O-glucoside, kaempferol 3,7-di-O-rhamnoside, quercetin, and quercetin 3-O-glucoside, the methanolic extract of Fagonia taeckholmiana afforded a new compound identified as kaempferol 3-O-beta-l-arabinopyranosyl-(1-->4)-alpha-l-rhamnopyranoside-7-O-alpha-l-rhamnopyranoside. Identification of the isolated compounds was based on chemical and spectroscopic analyses including UV, FABMS, (1)H, (13)C and 2D NMR, and DEPT. The cytotoxic activities of the compounds against several cancer cell lines were determined.     

Chemical constituents isolated from Disporum viridescens leaves and their inhibitory effect on nitric oxide production in BV2 microglial cells

Excessive NO (nitric oxide) has been associated with the pathogenesis of various neurodegenerative diseases including Alzheimer’s disease (AD). In our screening system using LPS-activated BV2 microglial cells, the methanolic extract of Disporum viridescens leaves was found to have significant NO inhibitory activity. Bioactivity-guided isolation yielded a new phenylpropanoid characterized as 4-ally-2,6-dimethoxyphenyl 1-O-β-d-apiofuranosyl (1 → 6)-β-d-glucopyranoside (12) with 21 known compounds from the leaves of D. viridescens. Among them, compounds 2 and 4 significantly inhibited NO production. Thus, we further elucidated the anti-inflammatory mechanism of these lignans. Especially, compound 4 inhibited the expression of both inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) through the suppression of the MAPK signaling pathway. Taken together, the anti-inflammatory activities of the active constituents isolated from D. viridescens leaves could have therapeutic potential against neurodegenerative diseases.