NO Production In RAW Research Articles

Two new phenolic glucosides from marine-derived fungus Aspergillus sp.

Two new phenolic glucosides, including a new O-glycoside (1) and a new C-glycoside (2), were isolated from a marine-derived fungus Aspergillus sp. The structures of new compounds were elucidated through interpretations of spectroscopic evidence and high-resolution electrospray ionization mass spectrometry. The hexose unit of 1 was identified as β-D-glucose by comparison with an authentic sample via HPLC after acid hydrolysis and derivatization. All compounds were evaluated for their ability to inhibit LPS-induced NO production in RAW264.7 macrophages, but none of them displayed significant activity.

Two New Quinazoline Derivatives from the Moss Endophytic Fungus Aspergillus sp. and Their Anti-inflammatory Activity

Two new quinazoline derivatives versicomides E (1) and F (2), and 10 known compounds (3–12) were isolated from the moss endophytic fungus Aspergillus sp. Their structures were determined on the basis of extensive spectroscopic data analysis and ECD calculations. Among them, the compound 7 (6-hydroxy-3-methoxyviridicatin) was first reported as a natural product. Inhibition on LPS-induced NO production in RAW 264.7 murine macrophages found that compounds 5, 7 and 8 showed significant inhibitory effects on NO production, with IC50 values of 49.85, 22.14 and 46.02 μM respectively.Graphic

Tetrahydroprotoberberine N-oxides from Chelidonium majus and their inhibitory effects on NO production in RAW 264.7 cells

Abstract Two new tetrahydroprotoberberine N-oxides, 7R,14S-cis-tetrahydrocoptisine N-oxide (1) and 7R,14R-trans-tetrahydrocoptisine N-oxide (2) along with fourteen known compounds (3–16) were isolated from the aerial parts of Chelidonium majus. Their structures were elucidated by spectroscopic and spectrometric methods, such as 1D and 2D-NMR (HSQC, HMBC, NOESY) and HRESIMS. The absolute configurations of 1 and 2 were established by comparison of their experimental and calculated ECD data. All isolates were evaluated for their inhibitory effects on the nitric oxide production in LPS-induced RAW 264.7 macrophages, and compounds 2, 6–10, 13, and 15 showed significant inhibitory effects with the IC50 values ranging from 1.1–31.9 μM.

Chemical constituents from the branches and leaves of Alchornea annamica

Phytochemical investigation of the branches and leaves of Alchornea annamica led to isolation of ten secondary metabolites, including two new megastigmane glucosides alnamicosides A (1) and B (2). The structure elucidation was confirmed by 1 D and 2 D NMR, ECD as well as HR-QTOF-MS experiments. The megastigmane derivatives 1 − 3 exhibited inhibitory effects on LPS-induced NO production in RAW264.7 cells with IC50 values of 78.72 ± 1.90, 77.40 ± 9.40 and 82.16 ± 4.56 µM, respectively. This is the first report on chemical constituents and biological activity of the plant A. annamica.

Polyhydroxylated steroid derivatives from the starfish Pentaceraster regulus

A new polyhydroxysteroid glycoside, regulusoside D (1), and a new polyhydroxylated steroid, (24S)-cholestane-3β,5,6β,8,15α,24-hexol (2), together with seven known compounds were purified from the starfish Pentaceraster regulus collected near Con Co Islands, Vietnam. The structure elucidation was confirmed by extensive analysis of their NMR and HR-QTOF mass experiments. Among isolated compounds, regulusoside D (1), (24S)-cholestane-3β,5,6β,8,15α,24-hexol (2), granulatoside A (4), 5α-cholestane-3β,6β,7α,15α,16β,26-hexol (6), 5α-cholestane-3β,6β,7α,8β,15α,16β,26-heptol (7) and 5α-cholestane-3β,6α,8,15α,16β,26-hexol (8) exhibited strong inhibitory effects on LPS-induced NO production in RAW264.7 cells with IC50 values of 9.13 ± 1.17, 9.19 ± 0.77, 8.63 ± 3.31, 12.61 ± 0.51, 14.83 ± 3.02 and 10.98 ± 1.22 µM, respectively.

Characterization and immunological activity of polysaccharides from Potentilla chinensis

Three novel polysaccharides (PCPW, PCPS1 and PCPS2) were isolated from Potentilla chinensis and subjected to structural analysis by using spectral and physicochemical methods. The molecular weights of PCPW, PCPS1 and PCPS2 were calculated to be 4.45 × 103 Da, 1.18 × 104 Da and 4.23 × 104 Da, respectively. Analysis of monosaccharides composition confirmed that PCPW was composed of mannose, rhamnose, glucose, galactose and arabinose, while the two acidic polysaccharides PCPS1 and PCPS2 were consisted of six monosaccharides, including mannose, rhamnose, galacturonic acid, glucose, galactose and arabinose respectively. In addition, the main linkages of glycosidic bonds of PCPS2 were 1, 4-linked-rha, 1, 4-linked-man,1, 4-linked-galA and 1, 6-linked-man. Immunological tests indicated that both PCPW and PCPS2 could increase NO production of RAW264.7 cells, and promote splenocyte proliferation. All three polysaccharides proved to be activators of NF-κB. Overall, three polysaccharides showed a good immunological activity and pose great potential as a novel food or drug additive.

Secondary Metabolites with Nitric Oxide Inhibition from Marine-Derived Fungus Alternaria sp. 5102.

Two new benzofurans, alternabenzofurans A and B (1 and 2) and two new sesquiterpenoids, alternaterpenoids A and B (3 and 4), along with 18 known polyketides (5−22), were isolated from the marine-derived fungus Alternaria sp. 5102. Their structures were elucidated on the basis of extensive spectroscopic analyses (1D and 2D NMR, HR-ESIMS, and ECD) and X-ray crystallography, as well as the modified Mosher’s method. Compounds 2, 3, 5, 7, 9–18, and 20–22 exhibited potent anti-inflammatory activity by inhibiting the production of NO in RAW264.7 cells activated by lipopolysaccharide with IC50 values in the range from 1.3 to 41.1 μM. Structure-activity relationships of the secondary metabolites were discussed.

The chemical constituents of Gentiana macrophylla pall. under acidic condition using gentiopicroside-rich secoiridoids extract

The chemical constituents of Gentiana macrophylla Pall. under acidic condition were explored using gentiopicroside-rich secoiridoids extract treated with hydrochloric acid. 2 new secoiridoid-like structures gentiagenin A (1) and B (2), together with 4 known compounds, swerimilegenin G (3), 4-hydroxyindan-1-one (4), erythrocentaurin (5) and swerimilegenin E (6) were identified using 1D, 2D NMR and HR-ESI-MS spectral technology. In the activity evaluation, the 2 new compounds, such as gentiagenin A (1) and B (2), exhibited stronger inhibitory effect on NO production in RAW264.7 macrophage cells than that of gentiopicroside.

Enhanced antioxidant, anti-inflammatory and α-glucosidase inhibitory activities of citrus hesperidin by acid-catalyzed hydrolysis

Hesperidin hydrolysates (HHS) was produced by the hydrolysis of hesperidin (HDN) in previous studies. The potential components in HHS were identified by LC-MS, and minor components (MCS) in HHS were isolated. Antioxidant activities by radical-scavenging capacities, reducing capacity and β-carotene-linoleate assay, anti-inflammatory effects by inhibiting NO production of RAW 264.7 cells, and α-glucosidase inhibitory effects of HDN, HHS, MCS and henperetin (HTN) were investigated in present study. HHS showed higher radical scavenging activities, higher reducing capacity, and higher inhibitory activity in the β-carotene-linoleate assay than HDN. HHS inhibited the production of NO and pro-inflammatory cytokines of RAW 264.7 cells more strongly than HDN. HHS also intensively inhibited α-glucosidase activity whereas HDN showed little activity. In addition, the effects of MCS on above activities showed it play a synergistic part with HTN. This work suggested that hydrolyzation of HDN enhance the activities, and provided valuable information on effective utilization of HDN.

Chemical constituents from Xanthii Fructus

This study was carried out to investigate the chemical constituents from Xanthii Fructus(the fruits of Xanthium sibiricum). The compounds were separated and purified by silica gel column chromatography, Sephadex LH-20 and ODS chromatography and semi-preparative HPLC. Base on HR-ESI-MS, NMR and other spectral data, their structures were identified. The anti-inflammatory activity of the isolated compounds was evaluated by lipopolysaccharide(LPS)-induced macrophage RAW264.7 as a screening model. A total of twenty-one compounds were isolated from the EtOAc fraction of 95% ethanol extract and identified as uracil(1), thymine(2), uridine(3), indole-3-carbaldehyde(4), indole-3-carboxylic acid(5), 2'-O-methyluridine(6), guanosine(7), 2,4(1H,3H)-quinazolinedione(8), 3-hydroxy-3-(2-hydroxyethyl)indolin-2-one(9), nicotinamide(10), N-acetyl-L-phenylalaninol(11), heliolactam(12), terresoxazine(13), caudatin(14), qingyangshengenin(15), caudatin-3-O-β-D-oleandropyranosyl-(1→4)-β-D-cymaropyranosyl-(1→4)-β-D-cymaropyranoside(16), caudatin-3-O-β-D-cymaropyranosyl-(1→4)-β-D-oleandropyranosyl-(1→4)-β-D-cymaropyranosyl-(1→4)-β-D-cymaropyranoside(17), caudatin-3-O-α-L-cymaropyranosyl-(1→4)-β-D-cymaropyranosyl-(1→4)-β-D-cymaropyranosyl-(1→4)-α-L-cymaropyranosyl-(1→4)-β-D-oleandropyranosyl-(1→4)-β-D-cymaropyranoside(18), qinyangshengenin-3-O-β-D-oleandropyranosyl-(1→4)-β-D-oleandropyranosyl-(1→4)-β-D-cymaropyranoside(19), qinyangshengenin-3-O-β-D-oleandropyranosyl-(1→4)-β-D-cymaropyranosyl-(1→4)-β-D-digitoxopyranoside(20), rostratamine-3-O-β-D-oleandropyranosyl-(1→4)-β-D-cymaropyranosyl-(1→4)-β-D-cymaropyranoside(21). Compounds 5-21 are obtained from genus Xanthium for the first time. Compounds 12 and 13 indirectly exhibited anti-inflammatory activity by suppressing LPS-induced NO production in RAW264.7 cells with IC_(50) values of(15.45±0.56) and(20.14±0.78) μmol·L~(-1), respectively.

콩 추출물의 라디칼 소거 및 항염증 활성

This study compared the antioxidant and anti-inflammatory capacities of acetone extracts from commercially available soybeans with reddish (Gangnang-kong, Ultari-kong, and Lentils), black (Heuk-tae, Seori-tae, and Seomok-tae), and yellow (Baek-tae and Chickpea) seed coats. Reddish soybeans had the highest 2,2- diphenyl-1-picrylhydrazyl (DPPH) radical-, superoxide-, and peroxynitrite-scavenging activities as well as the highest cupric reducing antioxidant capacities, followed by black soybeans; yellow soybeans showed the lowest value. The oxygen radical absorbance capacities and total phenolic contents of reddish and black soybeans were similar and higher than those of yellow soybeans were. Supercoiled DNA strand breakage induced by the hydroxyl radical was inhibited more by reddish soybeans compared to black or yellow soybeans. Lipopolysaccharide-induced NO production in RAW 264.7 cells was inhibited more by reddish or black soybeans than yellow soybeans. The antioxidant activities, including DPPH radical scavenging, superoxide scavenging, and cupric reducing antioxidant capacity, were strongly correlated with the total phenolic and total flavonoid contents. The anti-inflammatory capacity was strongly correlated with the total flavonoid content. Therefore, these findings show that reddish or black soybeans had more potent antioxidant and anti-inflammatory effects than yellow soybeans, possibly due to their high phenolic contents.

Benzopyran Derivatives and an Aliphatic Compound from a Mangrove Endophytic Fungus Penicillium citrinum QJF-22.

Two new benzopyran derivatives, (2R,4S)-5-methoxy-2-methyl-3,4-dihydro-2H-1-benzopyran-4-ol and (2S,4R,2'S,4'R)-4,4'-oxybis(5-methoxy-2-methyl-3,4-dihydro-2H-1-benzopyran), and a new aliphatic compound, (3E,5Z,8S,10E)-8-hydroxytrideca-3,5,10,12-tetraen-2-one, together with three known benzopyran derivatives, were obtained from a mangrove endophytic fungus Penicillium citrinum QJF-22 collected in Hainan island. Their structures were determined by analysis of spectroscopic data and the relative configuration of (2R,4S)-5-methoxy-2-methyl-3,4-dihydro-2H-1-benzopyran-4-ol was also confirmed by single-crystal X-ray diffraction. The absolute configurations of four compounds were established by comparison of ECD spectra to calculations. The configuration of (3E,5Z,8S,10E)-8-hydroxytrideca-3,5,10,12-tetraen-2-one was confirmed by comparison of optical value to the similar compound. The configurations of the compounds (2S,4S)-5-methoxy-2-methyl-3,4-dihydro-2H-1-benzopyran-4-ol and (2R,4R)-5-methoxy-2-methyl-3,4-dihydro-2H-1-benzopyran-4-ol were first determined. (3R,4S)-3,4,8-Trihydroxy-3,4-dihydronaphthalen-1(2H)-one exhibited moderate inhibitory effects on LPS-induced NO production in RAW264.7 cells with IC50 of 44.7 μM, and without cytotoxicity to RAW264.7 cells within 50 μM.

Alkaloids including two rare variecolortides from the fungus Aspergillus ruber

Three new alkaloids, namely, (−)-(S)-variecolortide D (1a), (+)-(R)-variecolortide D (1b) and brevicompanine I (3), along with three known alkaloids (2, 4 and 5) were isolated from Aspergillus ruber WH4-1. Variecolortide D (1) is a racemic natural product composed of anthraquinone and isoechinulin alkaloid, separation of 1 by chiral HPLC, provided two optically pure enantioners (1a and 1b). The structures of all compounds were determined by extensive analysis of their spectroscopic data as well as by comparison with literature. 2 showed weak inhibitory activity against the production of NO in RAW264.7 cells stimulated by lipopolysaccharide (LPS), with inhibitory rate of 53.2 ± 3.8% at 30 μM.

Sesquiterpenoids from the roots of Daphne genkwa Siebold et Zucc. With potential anti-inflammatory activity

Six undescribed sesquiterpenoids, including three acorane-type sesquiterpenoids (daphneaines A-C), three guaiane-type sesquiterpenoids (daphneaines E-G) and three known analogues were isolated from the roots of Daphne genkwa Siebold et Zucc. Their gross structures were elucidated by comprehensive spectroscopic analyses. The relative configurations of daphneaines A-C were determined by NOESY experiments. In addition, the relative configuration of daphneaine G was elucidated by performing a quantum chemical calculation of the NMR chemical shifts coupled with an advanced statistical procedure DP4+. The comparison of experimental and calculated electronic circular dichroism (ECD) data led to the establishment of the absolute configurations of daphneaines A-G. All isolates were tested for their inhibitory activity towards the LPS-induced NO production in RAW 264.7 macrophages, and daphneaine F showed inhibitory effect on NO production with an IC50 value of 35.68 ± 3.18 μM.

Guaiane type of sesquiterpene with NO inhibitory activity from the root of Wikstroemia indica.

Roots of Wikstroemia indica is widely used in China as a folk medicine in treatment of many diseases. However, active compounds of guaiane type of sesquiterpene remain largely unknown. In the present work, five new guaiane of type sesquiterpene compounds wikstronone A-E, along with one known guaiane type of sesquiterpene compound were isolated from the petroleum and CH2Cl2 fraction of roots of Wikstroemia indica. Structures of these compounds including absolute configuration were determined by extensive NMR and CD spectroscopic analysis. The inhibitory activity of all isolated compounds were assayed against LPS-induced NO production in RAW 264.7 macrophages by Griess reagent. Among all compounds, wikstronone A (1) showed remarkable NO inhibitory activity comparable to that of positive control Indometacin. Wikstronone D and E (4 and 5) showed weak NO inhibitory activity. The isolated guaiane type of sesquiterpene may be potent NO synthetase inhibitors.

Inhibitory Effect of 1,5-Dimethyl Citrate from Sea Buckthorn (Hippophae rhamnoides) on Lipopolysaccharide-Induced Inflammatory Response in RAW 264.7 Mouse Macrophages.

Hippophae rhamnoides L. (Elaeagnaceae; commonly known as “sea buckthorn” and “vitamin tree”), is a spiny deciduous shrub whose fruit is used in foods and traditional medicines. The H. rhamnoides fruit (berry) is rich in vitamin C, with a level exceeding that found in lemons and oranges. H. rhamnoides berries are usually washed and pressed to create pomace and juice. Today, the powder of the aqueous extract of H. rhamnoides berries are sold as a functional food in many countries. As part of our ongoing effort to identify bioactive constituents from natural resources, we aimed to isolate and identify those from the fruits of H. rhamnoides. Phytochemical analysis of the extract of H. rhamnoides fruits led to the isolation and identification of six compounds, namely, a citric acid derivative (1), a phenolic (2), flavonoids (3 and 4), and megastigmane compounds (5 and 6). Treatment with compounds 1–6 did not have any impact on the cell viability of RAW 264.7 mouse macrophages. However, pretreatment with these compounds suppressed lipopolysaccharide (LPS)-induced NO production in RAW 264.7 mouse macrophages in a concentration-dependent manner. Among the isolated compounds, compound 1 was identified as the most active, with an IC50 of 39.76 ± 0.16 μM. This value was comparable to that of the NG-methyl-L-arginine acetate salt, a nitric oxide synthase inhibitor with an IC50 of 28.48 ± 0.05 μM. Western blot analysis demonstrated that compound 1 inhibited the LPS-induced expression of IKKα/β (IκB kinase alpha/beta), I-κBα (inhibitor of kappa B alpha), nuclear factor kappa-B (NF-κB) p65, iNOS (inducible nitric oxide synthase), and COX-2 (cyclooxygenase-2) in RAW 264.7 cells. Furthermore, LPS-stimulated cytokine production was detected using a sandwich enzyme-linked immunosorbent assay. Compound 1 decreased interleukin 6 (IL-6) and tumor necrosis factor alpha (TNF-α) production in LPS-stimulated RAW 264.7 cells. In summary, the mechanism of action of 1 included the suppression of LPS-induced NO production in RAW 264.7 cells by inhibiting IKKα/β, I-κBα, NF-κB p65, iNOS, and COX-2, and the activities of IL-6 and TNF-α.

Rheological properties of a neutral polysaccharide extracted from maca (Lepidium meyenii Walp.) roots with prebiotic and anti-inflammatory activities

The rheological, prebiotic, and anti-inflammatory properties of neutral polysaccharide extracted from maca roots (MP) were investigated. MP was composed of arabinose, galactose, rhamnose, and glucose. In steady shear rheological properties, an increasing concentration of MP solution exhibited higher apparent viscosity (ηa,100) and consistency index (K) under acidic condition (pH 4). In dynamic rheological properties, the dynamic moduli (G′ and G″) in the frequency sweep test from the MP solution were increased with increasing concentration and decreasing pH. The changes in dynamic moduli of the MP solution with various concentrations and pH values were stable during 1 h storage at 4 °C due to the enhancement of hydrogen bonding. According to the results of the temperature sweep test, an increasing concentration of MP solution increased dynamic moduli under acidic conditions. The prebiotic properties of MP induced a higher growth of Bifidobacterium longum ATCC15707 and Lactobacillus rhamnosus ATCC 7469 than inulin and increased acetic acid, propionic acid, and butyric acid more than inulin in vitro. Furthermore, MP inhibited lipopolysaccharide-induced NO production in RAW 264.7 cells, which indicated that an increasing concentration of MP enhanced anti-inflammatory activities. Therefore, MP is a potential functional material for the food and pharmaceutical industries.

New Constituents from the Low Polar Fraction of the Fruits of Crataegus dahurica and Their Anti-Inflammatory Activity in RAW264.7 Cells.

The fruit of Crataegus dahurica Koehne was used to treat the disease of infantile indigestion and dyspepsia as an ethnic medicine and food. As a continuous work on finding the active constituents from the edible herbs, four new biphenyl derivatives (1-4), together with two known compounds (5 and 6), were obtained from the petroleum ether fraction of the fruits of C. dahurica. Their structures were determined by the extensive 1D and 2D NMR spectra and HR-MS spectrometry. Furthermore, the anti-inflammatory activities of all the isolated compounds were investigated, in which compound 4 showed moderately inhibitory effects on NO production in RAW264.7 cells without inducing cytotoxicity.

Synthesis of New Lathyrane Diterpenoid Derivatives from Euphorbia lathyris and Evaluation of Their Anti-Inflammatory Activities.

Euphorbia factor L3 , a lathyrane diterpenoid extracted from Euphorbia lathyris, was found to display good anti-inflammatory activity with very low cytotoxicity. To find more potent anti-inflammatory drugs, two series of Euphorbia factor L3 derivatives with fatty and aromatic acids were designed and synthesized. Among them, lathyrane derivative 5n exhibited most potent inhibition on LPS-induced NO production in RAW264.7 cells with no obvious cytotoxicity. To determine the key characteristics of Euphorbia factor L3 derivatives that contribute to anti-inflammatory activity, we conducted a structure-activity relationship study of these compounds.

Anti-inflammatory activity of the extracts from Rodgersia podophylla leaves through activation of Nrf2/HO-1 pathway, and inhibition of NF-κB and MAPKs pathway in mouse macrophage cells.

Recently, Rodgersia podophylla has been reported to exhibit anti-inflammatory activity. However, little is known about the potential mechanisms about its anti-inflammatory activity. We elucidated the anti-inflammatory mechanisms of leaves extracts from Rodgersia podophylla (RP-L) in RAW264.7 cells. LPS-induced NO was measured by Griess and mRNA of pro-inflammatory mediators was analyzed by RT-PCR. Cell viability was measured using MTT assay. The protein level was analyzed by Western blot. RP-L significantly inhibited the production of the pro-inflammatory mediators such as NO, iNOS, IL-1β and IL-6 in LPS-stimulated RAW264.7 cells. RP-L increased HO-1 expression in RAW264.7 cells, and the inhibition of HO-1 by ZnPP reduced the inhibitory effect of RP-L against LPS-induced NO production in RAW264.7 cells. Inhibition of p38, ROS and GSK3β attenuated RP-L-mediated HO-1 expression. Inhibition of ROS inhibited p38 phosphorylation and GSK3β expression induced by RP-L. In addition, inhibition of GSK3β blocked RP-L-mediated p38 phosphorylation. RP-L induced nuclear accumulation of Nrf2, and inhibition of p38, ROS and GSK3β abolished RP-L-mediated nuclear accumulation of Nrf2. Furthermore, RP-L blocked LPS-induced degradation of IκB-α and nuclear accumulation of p65. RP-L also attenuated LPS-induced phosphorylation of ERK1/2 and p38. In GC/MS analysis of RP-L, pyrogallol was detected as bioactive compound for anti-inflammatory activity of RP-L. Pyrogallol was observed to activate HO-1 expression through ROS/GSK3β/p38/Nrf2/HO-1 signaling. Our results suggest that RP-L exerts potential anti-inflammatory activity by activating ROS/GSK3β/p38/Nrf2/HO-1 signaling and inhibiting NF-κB and MAPK signaling in RAW264.7 cells. These findings suggest that RP-L may have great potential for the development of anti-inflammatory drug.