RESUMO
The present study focused on the investigation of the inhibition of cyclooxygenase-2 and nuclear factor kappa B1 gene expression, nitric oxide production, leukotriene biosynthesis (5-lipoxygenase), and cyclooxygenase-1 and cyclooxygenase-2 enzymes of Onopordum acanthium, and the isolation and identification of its active compounds. From the chloroform soluble part of the MeOH extract prepared from aerial parts, lignans [pinoresinol (1), syringaresinol (2), and medioresinol (3)] and flavonoids [hispidulin (4), nepetin (5), apigenin (6), and luteolin (7)] were isolated by a combination of different chromatographic methods. The structures of the compounds were determined by means of mass spectrometry and 1D- and 2D-nuclear magnetic resonance spectroscopy, and by comparison of the spectral data with literature values. Extracts of different polarity and the isolated compounds obtained from the aerial parts, together with those previously isolated from the roots of the plant [4ß,15-dihydro-3-dehydrozaluzanin C (8), zaluzanin C (9), 4ß,15,11ß,13-tetrahydrozaluzanin C (10), nitidanin diisovalerianate (11), 24-methylenecholesterol (12), and 13-oxo-9Z,11E-octadecadienoic acid (13)], were evaluated for their inhibitory effects on cyclooxygenase-2 and nuclear factor kappa B1 gene expression, inducible nitric oxide synthase, 5-lipoxygenase, and cyclooxygenase-1 and cyclooxygenase-2 enzymes in in vitro assays. It was found that O. acanthium extracts exert strong inhibitory activities in vitro and some lignans, flavonoids, and sesquiterpenes may play a role in these activities. 4ß,15-Dihydro-3-dehydrozaluzanin C and zaluzanin C at 20 µM were the most active constituents tested against lipopolysaccharide/interferon-γ-induced nitric oxide production (100.4 ± 0.5â% and 99.4 ± 0.8â%) in the inhibition of cyclooxygenase-2 (98.6 ± 0.2â% and 97.0 ± 1.1â%) and nuclear factor kappa B1 gene expression (76.7 ± 7.3â% and 69.9 ± 3.4â%). Furthermore, it was shown that these inhibitory effects are not due to cytotoxicity of the compounds.
Assuntos
Inibidores de Ciclo-Oxigenase/farmacologia , Subunidade p50 de NF-kappa B/genética , Óxido Nítrico/metabolismo , Onopordum/química , Extratos Vegetais/farmacologia , Araquidonato 5-Lipoxigenase/genética , Araquidonato 5-Lipoxigenase/metabolismo , Linhagem Celular/efeitos dos fármacos , Ciclo-Oxigenase 1/genética , Ciclo-Oxigenase 1/metabolismo , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Inibidores de Ciclo-Oxigenase 2/farmacologia , Inibidores de Ciclo-Oxigenase/química , Avaliação Pré-Clínica de Medicamentos/métodos , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Inibidores de Lipoxigenase/química , Inibidores de Lipoxigenase/farmacologia , Estrutura Molecular , Subunidade p50 de NF-kappa B/antagonistas & inibidores , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Extratos Vegetais/química , Plantas Medicinais/químicaRESUMO
In order to identify active constituents and to gain some information regarding their mode of action, extracts from leaves of Epipremnum pinnatum were tested for their ability to inhibit inflammatory gene expression in endothelial- and monocyte-like cells (HUVECtert and THP-1, respectively). Bioactivity-guided fractionation using expression of PTGS2 (COX-2) mRNA as a readout resulted in the isolation of two C13 megastigmane glycosides, gusanlungionoside C (1) and citroside A (3), and the phenylalcohol glycoside phenylmethyl-2-O-(6-O-rhamnosyl)-ß-D-galactopyranoside (2). Further analysis identified six additional megastigmane glycosides and the aglycones ß-damascenone (10), megastigmatrienone (11), 3-hydroxy-ß-damascenone (12), and 3-oxo-7,8-dihydro-α-ionol (13). Pharmacological analysis demonstrated that 10 inhibits LPS-stimulated induction of mRNAs encoding for proinflammatory cytokines and leukocyte adhesion molecules, such as TNF-α, IL-1ß, IL-8, COX-2, E-selectin, ICAM-1, and VCAM-1 in HUVECtert and THP-1 cells. 10 inhibited induction of inflammatory genes in HUVECtert and THP-1 cells treated with different agonists, such as TNF-α, IL-1ß, and LPS. In addition to mRNA, also the upregulation of inflammatory proteins was inhibited by 10 as demonstrated by immune assays for cell surface E-selectin and secreted TNF-α. Finally, using a luciferase reporter construct, it was shown, that 10 inhibits NF-κB-dependent transcription. Therefore, we hypothesize that inhibition of NF-κB by ß-damascenone (10) may represent one of the mechanisms underlying the in vitro anti-inflammatory activity of Epipremnum pinnatum extracts.
RESUMO
Filipendula vulgaris Moench (dropwort) is used in traditional medicine for relieving various inflammation-related diseases. In the present study, the phytochemical profile of F. vulgaris aerial part (FVA) and root (FVR) methanolic extracts was evaluated by LC-DAD-HRMS analysis. Furthermore, their in vitro and in vivo anti-inflammatory effects, as well as their potential cytotoxicity, were assessed. Results showed that the extracts mainly contain phenolics like flavonoids, hydrolyzable tannins, procyanidins, and phenolic acid derivatives, including gaultherin. No in vitro cytotoxicity was found at the highest concentration (50⯵g/mL). FVA extract (50⯵g/mL) significantly inhibited cyclooxygenase-1 and -2 (COX-1 and COX-2) activities in vitro (>50% inhibition), and FVR extract considerably inhibited COX-2 activity (52.5⯱â¯2.7%) without affecting COX-2 gene expression in LPS-stimulated THP-1â¯cells. The extracts demonstrated prominent in vivo anti-inflammatory potential upon oral administration in rats. Especially FVA extract at 100 and 200â¯mg/kg significantly inhibited carrageenan-induced edema formation. From these results, it can be concluded that F. vulgaris extracts possess interesting anti-inflammatory properties.
Assuntos
Anti-Inflamatórios/farmacologia , Filipendula/química , Compostos Fitoquímicos/química , Extratos Vegetais/farmacologia , Animais , Anti-Inflamatórios/administração & dosagem , Linhagem Celular , Linhagem Celular Tumoral , Cromatografia Líquida/métodos , Ciclo-Oxigenase 1/efeitos dos fármacos , Ciclo-Oxigenase 1/genética , Ciclo-Oxigenase 1/metabolismo , Ciclo-Oxigenase 2/efeitos dos fármacos , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Inibidores de Ciclo-Oxigenase/farmacologia , Relação Dose-Resposta a Droga , Sequestradores de Radicais Livres/farmacologia , Expressão Gênica , Humanos , Concentração Inibidora 50 , Masculino , Espectrometria de Massas/métodos , Metanol/química , Óxido Nítrico/metabolismo , Compostos Fitoquímicos/farmacologia , Componentes Aéreos da Planta/química , Extratos Vegetais/química , Raízes de Plantas/química , Ratos WistarRESUMO
The rue (Ruta graveolens) copiousness in rural areas of the Campania Region based a thorough chemical and biological investigation aimed at exploring the seasonal variability of phenol constituents in rue leaves and its influence on their antioxidant, cytotoxic and anti-inflammatory capabilities. To this purpose, hydroalcoholic extracts were prepared from plant samples seasonally collected. LC-ESI-MS/MS techniques were employed to analyze qualitatively and quantitatively the seasonal rue phenol content, whereas different chemical antioxidant assays (by DPPH, ABTS, Fe(3+) RP, ORAC, and FCR methods) and XTT redox metabolic activity assay were performed to screen the seasonal phenol complex-related antioxidant and cytotoxic power. The ability of the rue leaf extracts to counteract cyclooxygenase-2 (COX-2) expression was also evaluated. Data obtained highlighted that the adopted extraction procedure markedly pauperized the furanocoumarin content in all the prepared rue extracts. Flavonol glycosides, along with the flavone acacetin and two sinapic acid derivatives were the main constituents of the spring harvest-derived extract, which exerted the highest antioxidant capability in cell-free systems and was capable to inhibit COX-2 synthesis by 44% comparably to dexamethasone, used as positive control. Data provide new insights for developing a proper management of rue plants for new safe industrial purposes in herbal medicine field.
Assuntos
Antineoplásicos/farmacologia , Extratos Vegetais/farmacologia , Ruta/química , Estações do Ano , Antineoplásicos/química , Linhagem Celular Tumoral , Sobrevivência Celular , Humanos , Fitoterapia , Extratos Vegetais/química , Folhas de Planta/químicaRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Meadowsweet (Filipendula ulmaria (L.) Maxim, Rosaceae) has been traditionally used in most European countries for the treatment of inflammatory diseases due to its antipyretic, analgesic, astringent, and anti-rheumatic properties. However, there is little scientific evidence on F. ulmaria anti-inflammatory effects regarding its impact on cyclooxygenases enzymatic activity and in vivo assessment of anti-inflammatory potential. This study aims to reveal the anti-inflammatory activity of methanolic extracts from the aerial parts (FUA) and roots (FUR) of F. ulmaria, both in in vitro and in vivo conditions. MATERIALS AND METHODS: The characteristic phenolic compounds in F. ulmaria extracts were monitored via high performance thin layer chromatography (HPTLC). The in vitro anti-inflammatory activity of F. ulmaria extracts was evaluated using cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) enzyme assays, and an assay for determining COX-2 gene expression. The in vivo anti-inflammatory effect of F. ulmaria extracts was determined in two doses (100 and 200 mg/kg b.w.) with hot plate test and carrageenan-induced paw edema test in rats. Inflammation was also evaluated by histopathological and immunohistochemical analysis. RESULTS: FUA extract showed the presence of rutoside, spiraeoside, and isoquercitrin. Both F. ulmaria extracts at a concentration of 50µg/mL were able to inhibit COX-1 and -2 enzyme activities, whereby FUA extract (62.84% and 46.43% inhibition, respectively) was double as effective as the root extract (32.11% and 20.20%, respectively). Extracts hardly inhibited the level of COX-2 gene expression in THP-1 cells at a concentration of 25µg/mL (10.19% inhibition by FUA and 8.54% by FUR). In the hot plate test, both extracts in two doses (100 and 200mg/kg b.w.), exhibited an increase in latency time when compared with the control group (p<0.05). In the carrageenan-induced acute inflammation test, FUA at doses of 100 and 200mg/kg b.w., and FUR at 200mg/kg, were able to significantly reduce the mean maximal swelling of rat paw until 6h of treatment. Indomethacin, FUA, and FUR extracts significantly decreased inflammation score and this effect was more pronounced after 24h, compared to the control group (p<0.05). CONCLUSIONS: The observed results of in vitro and, for the first time, in vivo anti-inflammatory activity of meadowsweet extracts, provide support of the traditional use of this plant in the treatment of different inflammatory conditions. Further investigation of the anti-inflammatory compounds could reveal the mechanism of anti-inflammatory action of these extracts.
Assuntos
Anti-Inflamatórios/farmacologia , Etnofarmacologia , Filipendula/química , Extratos Vegetais/farmacologia , Animais , Anti-Inflamatórios/isolamento & purificação , Anti-Inflamatórios/uso terapêutico , Linhagem Celular Tumoral , Ciclo-Oxigenase 1/genética , Ciclo-Oxigenase 1/metabolismo , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Inibidores de Ciclo-Oxigenase/isolamento & purificação , Inibidores de Ciclo-Oxigenase/farmacologia , Inibidores de Ciclo-Oxigenase/uso terapêutico , Relação Dose-Resposta a Droga , Edema/tratamento farmacológico , Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Dor Nociceptiva/tratamento farmacológico , Componentes Aéreos da Planta/química , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/uso terapêutico , Raízes de Plantas/química , Ratos WistarRESUMO
Thymus longicaulis C. Presl. (Lamiaceae) is a small aromatic perennial herb typical of the Illyric-Mediterranean flora, traditionally used as remedy for cold, flu, cough, nephritis and abdominal pain. In order to carry out a thorough chemical and biological screening of the plant and to explore phenophases influence on its polyphenol content, samples of the plant were collected at different phases during its life cycle (July/October 2012 and January/April 2013). Each sample, previously extracted using a hydroalcoholic solution, was phytochemically analyzed for its metabolic constitution applying LC-DAD-ESI-MS/MS techniques. Although identified metabolites were differently concentrated at the various collection times, T. longicaulis leaf extracts were mainly constituted by low molecular weight phenols, and flavonoids. Rosmarinic acid was found as the main metabolite in Oct12 sample. Chemopreventive efficacy of the investigated extracts, by means of their anti-inflammatory, cytotoxic and antioxidant activities, was assessed. To this purpose, each extract underwent an extensive screening towards five human cell lines: CCRF-CEM (leukemia); U251 (glioblastoma); MDA-MB-231 (breast cancer); HCT-116 (colon cancer) and MRC-5 (lung fibroblasts) through XTT [2,3bis(2-metoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)carbonyl]-2H tetrazolium hydroxide] test. The ability of the extracts to counteract cyclooxygenase-2 (COX-2) expression was also evaluated by COX-2 expression assay in human THP-1 monocyte-derived macrophages. COX-2 inhibition could represent a valuable anticancer strategy as it is associated with carcinogenesis and over-expressed in a variety of human malignancies. Oct12 extract, which was particularly rich in rosmarinic acid and methylapigenin, exhibited a strong antioxidant and anti-inflammatory effectiveness.