Anti-Inflammatory Effects of Mitrephora sirikitiae Leaf Extract and Isolated Lignans in RAW 264.7 Cells

Mitrephora sirikitiae Weeras., Chalermglin & R.M.K. Saunders has been reported as a rich source of lignans that contribute to biological activities and health benefits. However, cellular anti-inflammatory effects of M. sirikitiae leaves and their lignan compounds have not been fully elucidated. Therefore, this study aimed to investigate the anti-inflammatory activities of methanol extract of M. sirikitiae leaves and their lignan constituents on lipopolysaccharide (LPS)-induced inflammation in RAW 264.7 mouse macrophage cells. Treatment of RAW 264.7 cells with the methanol extract of M. sirikitiae leaves and its isolated lignans, including (−)-phylligenin (2) and 3′,4-O-dimethylcedrusin (6) significantly decreased LPS-induced prostaglandin E₂ (PGE₂) and nitric oxide (NO) productions. These inhibitory effects of the extract and isolated lignans on LPS-induced upregulation of PGE₂ and NO productions were derived from the suppression of cyclooxygenase 2 (COX-2) and inducible nitric oxide synthase (iNOS) production, respectively. In addition, treatment with 2-(3,4-dimethoxyphenyl)-6-(3,5-dimethoxyphenyl)-3,7-dioxabicyclo[3.3.0]octane (3) and mitrephoran (5) was able to suppress LPS-induced tumor necrosis factor alpha (TNF-α) secretion and synthesis in RAW 264.7 cells. These results demonstrated that M. sirikitiae leaves and some isolated lignans exhibited potent anti-inflammatory activity through the inhibition of secretion and synthesis of PGE₂, NO, and TNF-α.     

Diterpenoids Isolated from Podocarpus macrophyllus Inhibited the Inflammatory Mediators in LPS-Induced HT-29 and RAW 264.7 Cells

Species of Podocarpus are used traditionally in their native areas for the treatment of fevers, asthma, coughs, cholera, chest pain, arthritis, rheumatism, and sexually transmitted diseases. To identify natural products having efficacy against inflammatory bowel disease (IBD), we identified a new, 16-hydroxy-4β-carboxy-O-β-D-glucopyranosyl-19-nor-totarol (4) together with three known diterpenoids from P. macrophyllus. Furthermore, all the extracts, fractions, and isolates 1–4 were investigated for their anti-inflammatory effects by assessing the expression on nitric oxide (NO) production and proinflammatory cytokines in lipopolysaccharide (LPS)-stimulated RAW 264.7 and HT-29 cells. Among them, nagilactone B (2) exhibited a potent anti-inflammatory effect against NO production on RAW 264.7 cells; therefore, nagilactone B was further assessed for anti-inflammatory activity. Western blot analysis revealed that nagilactone B significantly decreased the expression of LPS-stimulated protein, inducible nitric oxide synthase (iNOS), cyclooxygenase (COX)-2, and phosphorylated extracellular regulated kinase (pERK)1/2. In addition, nagilactone B downregulated tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-8 levels in LPS-induced macrophages and colonic epithelial cells. To our best knowledge, this is the first report on the inhibitory effect of nagilactone B (pure state) and rakanmakilactone G against NO production in LPS-stimulated RAW 264.7 cells. Thus, diterpenoids isolated from P. macrophyllus could be employed as potential therapeutic phytochemicals for IBD.     

Prunetin 4′-O-Phosphate,a Novel Compound,in RAW 264.7 Macrophages Exerts Anti-Inflammatory Activity via Suppression of MAP Kinases and the NFκB Pathway

Biorenovation, a microbial enzyme-assisted degradation process of precursor compounds, is an effective approach to unraveling the potential bioactive properties of the derived compounds. In this study, we obtained a new compound, prunetin 4′-O-phosphate (P4P), through the biorenovation of prunetin (PRN), and investigated its anti-inflammatory effects in lipopolysaccharide (LPS)-treated RAW 264.7 macrophage cells. The anti-inflammatory effect of P4P was evaluated by measuring the production of prostaglandin-E₂ (PGE₂), nitric oxide (NO), which is an inflammation-inducing factor, and related cytokines such as tumor necrosis factor-α (TNFα), interleukin-1β (IL1β), and interleukin-6 (IL6). The findings demonstrated that P4P was non-toxic to cells, and its inhibition of the secretion of NO—as well as pro-inflammatory cytokines—was concentration-dependent. A simultaneous reduction in the protein expression level of pro-inflammatory proteins such as cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) was observed. Moreover, the phosphorylation of mitogen-activated protein kinases (MAPKs) such as extracellular signal-regulated kinases (ERKs), c-Jun N-terminal kinase (JNK), p38 MAPK (p38), and nuclear factor kappa B (NFκB) was downregulated. To conclude, we report that biorenovation-based phosphorylation of PRN improved its anti-inflammatory activity. Cell-based in vitro assays further confirmed that P4P could be applied in the development of anti-inflammatory therapeutics.     

Iridal-Type Triterpenoids Displaying Human Neutrophil Elastase Inhibition and Anti-Inflammatory Effects from Belamcanda chinensis

The aim of this study is to explore anti-inflammatory phytochemicals from B. chinensis based on the inhibition of pro-inflammatory enzyme, human neutrophil elastase (HNE) and anti-inflammatory activities in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophage. Three stereoisomers of iridal-type triterpenoids (1–3) were isolated from the roots of B. chinensis and their stereochemistries were completely identified by NOESY spectra. These compounds were confirmed as reversible noncompetitive inhibitors against HNE with IC₅₀ values of 6.8–27.0 µM. The binding affinity experiment proved that iridal-type triterpenoids had only a single binding site to the HNE enzyme. Among them, isoiridogermanal (1) and iridobelamal A (2) displayed significant anti-inflammatory effects by suppressing the expressions of pro-inflammatory cytokines, such as iNOS, IL-1β, and TNF-α through the NF-κB pathway in LPS-stimulated RAW264.7 cells. This is the first report that iridal-type triterpenoids are considered responsible phytochemicals for anti-inflammatory effects of B. chinensis.     

Anti-Inflammatory Activity and Mechanism of Isookanin,Isolated by Bioassay-Guided Fractionation from Bidens pilosa L.

Bidens pilosa L. (Asteraceae) has been used historically in traditional Asian medicine and is known to have a variety of biological effects. However, the specific active compounds responsible for the individual pharmacological effects of Bidens pilosa L. (B. pilosa) extract have not yet been made clear. This study aimed to investigate the anti-inflammatory phytochemicals obtained from B. pilosa. We isolated a flavonoids-type phytochemical, isookanin, from B. pilosa through bioassay-guided fractionation based on its capacity to inhibit inflammation. Some of isookanin’s biological properties have been reported; however, the anti-inflammatory mechanism of isookanin has not yet been studied. In the present study, we evaluated the anti-inflammatory activities of isookanin using lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. We have shown that isookanin reduces the production of proinflammatory mediators (nitric oxide, prostaglandin E₂) by inhibiting the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in LPS-stimulated macrophages. Isookanin also inhibited the expression of activator protein 1 (AP-1) and downregulated the LPS-induced phosphorylation of p38 mitogen-activated protein kinase (MAPK) and c-jun NH₂-terminal kinase (JNK) in the MAPK signaling pathway. Additionally, isookanin inhibited proinflammatory cytokines (tumor necrosis factor-a (TNF-α), interleukin-6 (IL-6), interleukin-8 (IL-8), and interleukin-1β (IL-1β)) in LPS-induced THP-1 cells. These results demonstrate that isookanin could be a potential therapeutic candidate for inflammatory disease.     

Anthocyanin Profile,Antioxidant, Anti-Inflammatory,and Antimicrobial against Foodborne Pathogens Activities of Purple Rice Cultivars in Northern Thailand

Five glutinous purple rice cultivars and non-glutinous purple rice cultivated in different altitudes in the north of Thailand were collected. The samples were extracted using ethanol and determined for anthocyanins using HPLC. The total phenolic content (TPC), total flavonoid content (TFC), and the antioxidant, anti-inflammatory, and antimicrobial activities against foodborne pathogens were investigated. The highland glutinous cultivar named Khao’ Gam Luem-Phua (KGLP) extract had significantly high levels of cyanidin 3-O-glucoside, peonidin 3-O-glucoside, delphinidin 3-O-glucoside, TPC, and TFC, as well as exerting a potent antioxidant activity through ABTS assay (524.26 ± 4.63 VCEAC, mg l-ascorbic-ascorbic/g extract), lipid peroxidation (IC₅₀ = 19.70 ± 0.31 µg/mL), superoxide anions (IC₅₀ = 11.20 ± 0.25 µg/mL), nitric oxide (IC₅₀ = 17.12 ± 0.56 µg/mL), a suppression effect on nitric oxide (IC₅₀ = 18.32 ± 0.82 µg/mL), and an inducible nitric oxide synthase production (IC₅₀ = 23.43 ± 1.21 µg/mL) in combined lipopolysaccharide-interferon-γ-activated RAW 264.7 murine macrophage cells. Additionally, KGLP also exhibited antimicrobial activity against foodborne pathogens, Staphylococcus aureus, Escherichia coli, Salmonella Enteritidis, and Vibrio parahaemolyticus. These results indicate that Thai glutinous purple rice cultivated on the highland could be a potent natural source of antioxidants, anti-inflammatories, and antimicrobial agents for use as a natural active pharmaceutical ingredient in functional food and nutraceutical products.     

Cordyceps militaris Fungus Extracts-Mediated Nanoemulsion for Improvement Antioxidant,Antimicrobial, and Anti-Inflammatory Activities

This study aimed to produce and optimize a Cordyceps militaris-based oil-in-water (O/W) nanoemulsion (NE) encapsulated in sea buckthorn oil (SBT) using an ultrasonication process. Herein, a nonionic surfactant (Tween 80) and chitosan cosurfactant were used as emulsifying agents. The Cordyceps nanoemulsion (COR-NE) was characterized using Fourier-transform infrared spectroscopy (FT-IR), dynamic light scattering (DLS), and field-emission transmission electron microscope (FE-TEM). The DLS analyses revealed that the NE droplets were 87.0 ± 2.1 nm in diameter, with a PDI value of 0.089 ± 0.023, and zeta potential of −26.20 ± 2. The small size, low PDI, and stable zeta potential highlighted the excellent stability of the NE. The NE was tested for stability under different temperature (4 °C, 25 °C, and 60 °C) and storage conditions for 3 months where 4 °C did not affect the stability. Finally, in vitro cytotoxicity and anti-inflammatory activity were assessed. The results suggested that the NE was not toxic to RAW 264.7 or HaCaT (human keratinocyte) cell lines at up to 100 µL/mL. Anti-inflammatory activity in liposaccharides (LPS)-induced RAW 264.7 cells was evident at 50 µg/mL and showed inhibition of NO production and downregulation of pro-inflammatory gene expression. Further, the NE exhibited good antioxidant (2.96 ± 0.10 mg/mL) activity and inhibited E. coli and S. aureus bacterial growth. Overall, the COR-NE had greater efficacy than the free extract and added significant value for future biomedical and cosmetics applications.     

Inflammatory and Cytotoxic Activities of Abietane Terpenoids from Nepeta bracteata Benth.

Nepeta bracteata Benth. is used clinically to treat tracheal inflammation, coughs, asthma, colds, fevers, adverse urination, and other symptoms, along with functions in clearing heat and removing dampness. However, there have been few studies characterizing the material basis of its efficacy. Therefore, the aim of this study was to screen for compounds with anti-inflammatory activities in N. bracteata Benth. Using silica gel, ODS C18, and Sephadex LH-20 column chromatography, as well as semipreparative HPLC, 10 compounds were separated from N. bracteata Benth. extract, including four new diterpenoids (1–4), one amide alkaloid (5), and five known diterpenoids (6–10). The structures of all the isolates were elucidated by HR-ESI-MS, NMR, and CD analyses. Using lipopolysaccharide (LPS)-stimulated RAW 264.7 cells, we investigated the anti-inflammatory activities of compounds 1–10. It is worth noting that all were able to inhibit nitric oxide (NO) production with IC₅₀ values < 50 μM and little effect on RAW 264.7 macrophage viability. Compounds 2 and 4 displayed remarkable inhibition with IC₅₀ values of 19.2 and 18.8 μM, respectively. Meanwhile, screening on HCT-8 cells demonstrated that compounds 2 and 4 also had moderate cytotoxic activities with IC₅₀ values of 36.3 and 41.4 μM, respectively, which is related to their anti-inflammatory effects.     

Study of the Antioxidant and Anti-Inflammatory Properties of the Biological Extracts of Psophocarpus tetragonolobus Using Two Extraction Methods

Psophocarpus tetragonolobus has long been used in traditional medicine and cuisine. In this study, Psophocarpus tetragonolobus extracts were isolated by maceration and ultrasound-assisted extraction and were evaluated for their antioxidant and anti-inflammatory effects in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. The obtained results show that both extracts (maceration and ultrasound) were rich in bioactive molecules and exerted substantial antioxidant and anti-inflammatory effects. The P. tetragonolobus extracts’ treatment in LPS-stimulated RAW264.7 macrophages resulted in a significant downregulation of the expressions of tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and IL-1β mRNA. In addition, the P. tetragonolobus extracts’ treatment attenuated inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) protein expression. Our observations indicate that there is no significant difference between the two studied extracts of P. tetragonolobus in terms of biological properties (specifically, antioxidant and anti-inflammatory effects. Regardless of the extraction method, P. tetragonolobus could be used for treating diseases related to oxidative stress and inflammatory reactions.     

Chemical Composition,Antibacterial, Anti-Inflammatory,and Enzyme Inhibitory Activities of Essential Oil from Rhynchanthus beesianus Rhizome

Rhynchanthus beesianus W. W. Smith, an edible, medicinal, and ornamental plant, is mainly cultivated in China and Myanmar. The essential oil (EO) from R. beesianus rhizome has been used as an aromatic stomachic in China. The chemical composition and biological activities of EO from R. beesianus rhizome were reported for the first time. Based on gas chromatography with flame ionization or mass selective detection (GC-FID/MS) results, the major constituents of EO were 1,8-cineole (47.6%), borneol (15.0%), methyleugenol (11.2%), and bornyl formate (7.6%). For bioactivities, EO showed a significant antibacterial activity against Staphylococcus aureus, Enterococcus faecalis, Bacillus subtilis, Escherichia coli, Pseudomonas aeruginosa, and Proteus vulgaris with the diameter of the inhibition zone (DIZ) (8.66–10.56 mm), minimal inhibitory concentration (MIC) (3.13–6.25 mg/mL), and minimal bactericidal concentration (MBC) (6.25–12.5 mg/mL). Moreover, EO (128 μg/mL) significantly inhibited the production of proinflammatory mediators nitric oxide (NO) (92.73 ± 1.50%) and cytokines tumor necrosis factor-α (TNF-α) (20.29 ± 0.17%) and interleukin-6 (IL-6) (61.08 ± 0.13%) in lipopolysaccharide (LPS)-induced RAW264.7 macrophages without any cytotoxic effect. Moreover, EO exhibited significant acetylcholinesterase (AChE) inhibitory activity (the concentration of the sample that affords a 50% inhibition in the assay (IC₅₀) = 1.03 ± 0.18 mg/mL) and moderate α-glucosidase inhibition effect (IC₅₀ = 11.60 ± 0.25 mg/mL). Thus, the EO could be regarded as a bioactive natural product and has a high exploitation potential in the cosmetics and pharmaceutical industries.     

Quercetin-Rich Ethanolic Extract of Polygonum odoratum var Pakphai Leaves Decreased Gene Expression and Secretion of Pro-Inflammatory Mediators in Lipopolysaccharide-Induced Murine RAW264.7 Macrophages

Polygonum odoratum var. Pakphai has been used in traditional Thai medicine for the treatment of flatulence and constipation and to relieve the inflammation caused by insect bites. Quercetin (Q), which is abundant in plant-based foods, has been found to exert anti-inflammatory properties. This study evaluated the anti-inflammatory activity of P. odoratum ethanolic extract in RAW264.7 macrophage cells. Leaves were extracted with 50% ethanol, phenolics and flavonoids were then analyzed using UHPLC-QTOF-MS and HPLC-DAD. RAW264.7 cells were induced with lipopolysaccharides (LPSs). They were then treated with the extract and prostaglandin E₂ (PGE₂), and interleukin-6 (IL-6) and tumor necrotic factor-alpha (TNF-α) concentrations were determined. Levels of cyclooxygenase-2 (COX-2), inducible nitric oxide synthase (iNOS), IL-6 and TNF-α mRNAs were analyzed using qRT-PCR. Chemical analysis demonstrated that the extract was abundant with Q while also containing catechin, gallic acid, epicatechin gallate and coumarin. The extract increased the viability of RAW264.7 cells and dose-dependently decreased nitric oxide production, PGE₂, IL-6 and TNF-α levels in the medium from the LPS-induced RAW264.7 cell culture. Consistently, COX-2, iNOS, IL-6 and TNF-α mRNA levels were decreased in a concentration-dependent manner (p < 0.05). Thus, the quercetin-rich ethanolic extract derived from P. odoratum var Pakphai leaves can exert anti-inflammatory activity in LPS-induced RAW264.7 cells through a reduction of the pro-inflammatory mediator response.     

Inhibition of LPS-induced no production in RAW264.7 cell lines: synthesis and molecular modeling of α-ketophosphonates of febuxostat

In an effort to develop novel anti-inflammatory agents, we synthesized phosphorylated derivatives of febuxostat using various phosphates/phosphonites by Michaelis-Arbuzov reaction. Their inhibitory activity on nitric oxide (NO) production, reactive oxygen species (ROS) production and cytotoxicity were evaluated using lipopolysaccharide (LPS) activated macrophages RAW264.7 assay, flow-cytometry analysis and MTT method respectively. Some of these compounds showed potent inhibition of NO and ROS production without obvious cytotoxicity. Furthermore, docking simulations were performed to positional compounds 3g, 3h, 3i and 3j into the COX-2 active site to determine the probable binding model. Physicochemical parameters revealed that most of the compounds possessed drug-like properties.     

Synthesis and Investigation of Flavanone Derivatives as Potential New Anti-Inflammatory Agents

Flavonoids are polyphenols with broad known pharmacological properties. A series of 2,3-dihydroflavanone derivatives were thus synthesized and investigated for their anti-inflammatory activities. The target flavanones were prepared through cyclization of 2′-hydroxychalcone derivatives, the later obtained by Claisen–Schmidt condensation. Since nitric oxide (NO) represents an important inflammatory mediator, the effects of various flavanones on the NO production in the LPS-induced RAW 264.7 macrophage were assessed in vitro using the Griess test. The most active compounds were flavanone (4G), 2′-carboxy-5,7-dimethoxy-flavanone (4F), 4′-bromo-5,7-dimethoxy-flavanone (4D), and 2′-carboxyflavanone (4J), with IC50 values of 0.603, 0.906, 1.030, and 1.830 µg/mL, respectively. In comparison, pinocembrin achieved an IC₅₀ value of 203.60 µg/mL. Thus, the derivatives synthesized in this work had a higher NO inhibition capacity compared to pinocembrin, demonstrating the importance of pharmacomodulation to improve the biological potential of natural molecules. SARs suggested that the use of a carboxyl-group in the meta-position of the B-ring increases biological activity, whereas compounds carrying halogen substituents in the para-position were less active. The addition of methoxy-groups in the meta-position of the A-ring somewhat decreased the activity. This study successfully identified new bioactive flavanones as promising candidates for the development of new anti-inflammatory agents.     

Structural Characterization and Assessment of Anti-Inflammatory Activities of Polyphenols and Depsidone Derivatives from Melastoma malabathricum subsp. normale

The roots of Melastoma malabathricum subsp. normale (D. Don) Karst. Mey have been used in traditional ethnic medicine systems in China to treat inflammation-triggered ailments, such as trauma, toothache, and fever. Therefore, the aim of this study is to screen for compounds with anti-inflammatory activity in the title plant. The extract of M. malabathricum subsp. normale roots was separated using various chromatographic methods, such as silica gel, ODS C18, MCI gel, and Sephadex LH-20 column chromatography, as well as semi-preparative HPLC. One new complex tannin, named whiskey tannin D (1), and an undescribed tetracyclic depsidone derivative, named guanxidone B (2), along with nine known polyphenols (2–10) and three known depsidone derivatives (12–14) were obtained from this plant. The structures of all compounds were elucidated by extensive NMR and CD experiments in conjunction with HR-ESI-MS data. All these compounds were isolated from this plant for the first time. Moreover, compounds 1–4, 8, and 10–14 were obtained for the first time from the genus Melastoma, and compounds 1, 2, and 11–14 have not been reported from the family Melastomataceae. This is the first report of complex tannin and depsidone derivatives from M. malabathricum subsp. normale, indicating their chemotaxonomic significance to this plant. Compounds 1–12 were investigated for their anti-inflammatory activities on the production of the nitric oxide (NO) in lipopolysaccharide (LPS)-stimulated RAW264.7 cells, and compounds 1, 11, and 12 showed anti-inflammatory activities with IC₅₀ values of 6.46 ± 0.23 µM, 8.02 ± 0.35 µM, and 9.82 ± 0.43 µM, respectively. The structure–activity relationship showed that the catechin at glucose C-1 in ellagitannin was the key to its anti-inflammatory activity, while CH₃O- at C-16 of aromatic ring A in depsidone derivatives had little effect on its anti-inflammatory activity. The study of structure–activity relationships is helpful to quickly discover new anti-inflammatory drugs. The successful isolation and structure identification of these compounds, especially complex tannin 1, not only provide materials for the screening of anti-inflammatory compounds, but also provide a basis for the study of chemical taxonomy of the genus Melastoma.     

In vitro anti-inflammatory activities of naucleoffieine H as a natural alkaloid from Nauclea officinalis Pierrc ex Pitard,through inhibition of the iNOS pathway in LPS-activated RAW 264.7 macrophages

Abstract

Naucleoffieine H, a natural indole alkaloid, was isolated and identified from Nauclea officinalis Pierrc ex Pitard which is a traditional Chinese medicine used for the treatment of various diseases, such as cold, fever, bronchitis, pneumonia, acute tonsillitis, etc. In the present study, the effect of naucleoffieine H on the anti-inflammatory activities was investigated in LPS-induced RAW 264.7 macrophages. The results showed that naucleoffieine H significantly inhibited the release of nitric oxide (the level of nitrite as a stable biomarker of NO production) and tumor necrosis factor-α (TNF-α). Interesting, naucleoffieine H down-regulated the overexpression of inflammatory protein induced nitric oxide synthase (iNOS), but no effect on the expression cyclooxygenase-2 (COX-2) protein. In addition, this bioactive alkaloid suppressed enzymatic activity of iNOS activated by LPS. The above results indicated that naucleoffieine H suppress NO and TNF-α overproduction via block the iNOS pathway in LPS-induced RAW 264.7 macrophages.     

Structural Characterization and Assessment of Anti-Inflammatory and Anti-Tyrosinase Activities of Polyphenols from Melastoma normale

Polyphenols, widely distributed in the genus Melastoma plants, possess extensive cellular protective effects such as anti-inflammatory, anti-tyrosinase, and anti-obesity, which makes it a potential anti-inflammatory drug or enzyme inhibitor. Therefore, the aim of this study is to screen for the anti-inflammatory and enzyme inhibitory activities of compounds from title plant. Using silica gel, MCI, ODS C18, and Sephadex LH-20 column chromatography, as well as semipreparative HPLC, the extract of Melastoma normale roots was separated. Four new ellagitannins, Whiskey tannin C (1), 1-O-(4-methoxygalloyl)-6-O-galloyl-2,3-O-(S)-hexahydroxydiphenoyl-β-d-glucose (2), 1-O-galloyl-6-O-(3-methoxygalloyl)-2,3-O-(S)-hexahydroxydiphenoyl-β-d-glucose (3), and 1-O-galloyl-6-O-vanilloyl-2,3-O-(S)-hexahydroxydiphenoyl-β-d-glucose (4), along with eight known polyphenols were firstly obtained from this plant. The structures of all isolates were elucidated by HRMS, NMR, and CD analyses. Using lipopolysaccharide (LPS)-stimulated RAW2 64.7 cells, we investigated the anti-inflammatory activities of compounds 1–4, unfortunately, none of them exhibit inhibit nitric oxide (NO) production, their IC₅₀ values are all > 50 μM. Anti-tyrosinase activity assays was done by tyrosinase inhibition activity screening model. Compound 1 showed weak tyrosinase inhibitory activity with IC₅₀ values of 426.02 ± 11.31 μM. Compounds 2–4 displayed moderate tyrosinase inhibitory activities with IC₅₀ values in the range of 124.74 ± 3.12–241.41 ± 6.23 μM. The structure–activity relationships indicate that hydroxylation at C-3′, C-4′, and C-3 in the flavones were key to their anti-tyrosinase activities. The successful isolation and structure identification of ellagitannin provide materials for the screening of anti-inflammatory drugs and enzyme inhibitors, and also contribute to the development and utilization of M. normale.     

In Vitro Anti-Inflammatory Activity of Essential Oil and β-Bisabolol Derived from Cotton Gin Trash

Natural α-bisabolol has been widely used in cosmetics and is sourced mainly from the stems of Candeia trees that have become endangered due to over exploitation. The in vitro anti-inflammatory activity of cotton gin trash (CGT) essential oil and the major terpenoid (β-bisabolol) purified from the oil were investigated against lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages as well as the 3t3 and HS27 fibroblast cell lines. Nitric oxide (NO), prostaglandin E₂ (PGE₂), tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), and interleukin 8 (IL-8) were measured using Greiss reagent, enzyme-linked immunosorbent assay (ELISA), and cytokine bead array (CBA)-flow cytometry. Non-toxic concentrations of CGT oil and β-bisabolol (1.6–50.0 µg/mL) significantly inhibited the production of the inflammatory mediators in a dose-dependent manner. Maximal inhibition by β-bisabolol was 55.5% for NO, 62.3% for PGE₂, and 45.3% for TNF-α production in RAW cells. β-Bisabolol induced a level of inhibition similar to an equal concentration of α-bisabolol (50.0 µg/mL), a known anti-inflammatory agent. These results suggest β-bisabolol exerts similar in vitro effects to known topical anti-inflammatory agents and could therefore be exploited for cosmetic and therapeutic uses. This is the first study to report the in vitro anti-inflammatory activity of β-bisabolol in CGT essential oil.     

Sophorolipid Suppresses LPS-Induced Inflammation in RAW264.7 Cells through the NF-κB Signaling Pathway

Objectives: Biosurfactants with anti-inflammatory activity may alleviate skin irritation caused by synthetic surfactants in cleaning products. Sophorolipid (SL) is a promising alternative to synthetic surfactants. However, there are few reports on the anti-inflammatory activity of SL and the underlying mechanism. The purpose of this work is to verify that lipopolysaccharide (LPS)-induced inflammation could be inhibited through targeting the pathway of nuclear factor-κB (NF-κB) in RAW264.7 cells. Methods: The influence of SL on cytokine release was investigated by LPS-induced RAW264.7 cells using ELISA. The quantification of the protein expression of corresponding molecular markers was realized by Western blot analysis. Flow cytometry was employed to determine the levels of Ca²⁺ and reactive oxygen species (ROS). The relative expression of inducible nitric oxide synthase (INOS) and cyclooxygenase-2 (COX-2) was determined by RT-PCR. An immunofluorescence assay and confocal microscope were used to observe the NF-κB/p65 translocation from the cytoplasm into the nucleus. The likely targets of SL were predicted by molecular docking analysis. Results: SL showed anti-inflammatory activity and reduced the release of inflammatory cytokines including interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and nitric oxide (NO). The experimental results show that SL suppressed the Ca²⁺ and ROS levels influx in the LPS-induced RAW264.7 cells and alleviated the LPS-induced expression of iNOS and COX-2, the LPS-induced translocation of NF-κB (p65) from the cytoplasm into the nucleus, and the expression of phosphorylated proteins such as p65 and IκBα. Furthermore, molecular docking analysis showed that SL may inhibit inflammatory signaling by competing with LPS to bind TLR4/MD-2 through hydrophobic interactions and by inhibiting IKKβ activation through the hydrogen bonding and hydrophobic interactions. Conclusion: This study demonstrated that SL exerted anti-inflammatory activity via the pathway of NF-κB in RAW264.7 cells.     

The Anti-Inflammatory Properties of Chaga Extracts Obtained by Different Extraction Methods against LPS-Induced RAW 264.7

Chaga, a sclerotia formed by the Inonotus obliquus fungus, has been widely recognized for a number of medicinal properties. Although numerous scientific investigations have been published describing various biological activities of chaga from different geographical locations, little work has focused on chaga harvested in the USA or extraction techniques to maximize anti-inflammatory properties. The aim of this study was to investigate the anti-inflammatory properties of chaga collected in Maine (USA) extracted using traditional aqueous (hot water steeping) methods against lipopolysaccharide (LPS)-induced RAW 264.7 macrophages. Chaga extracts obtained from both conventional (ethanol/water) extraction methods and an accelerated solvent extraction method (ASE) at optimized conditions were compared to aqueous extracts (tea) obtained from chaga in the powder form (P) and powder form in tea bags (B) based on their effect on both nitric oxide (NO) production and pro-inflammatory cytokine expression, in particular, the expression of TNF-α, interleukin-6 (IL-6), and interleukin-β (IL-1β). Phenolic acid extracts from chaga and individual phenolic acid standards were also investigated for their effect on the same parameters. Results indicated that various chaga extracts have significant anti-inflammatory activity on LPS-stimulated RAW 264.7 cells. The inhibitory effect was through a decrease in the production of NO and the downregulation of TNF-α, IL-6, and IL-1β in RAW 264.7 macrophages. ASE1 (novel, optimized ethanol/water extraction) and P6 (six-minute steeping of powder in 100 °C water) extracts showed the highest inhibitory activity on NO production and on the expression of the inflammatory cytokines, compared to extracts obtained by conventional extraction methods.