Wound healing activity and phytochemical screening of purified fractions of Sempervivum tectorum L. leaves on HCT 116.

INTRODUCTION: Sempervivum tectorum L. (Crassulaceae), is a succulent perennial plant widespread in Mediterranean countries and commonly used in traditional medicine for ear inflammation, ulcers and skin rashes as a refrigerant and astringent. OBJECTIVE: To demonstrate the therapeutic effects of the plant, various fractions were purified and characterised. The potential wound healing activity, proliferation rate and intracellular signalling cascades were investigated by using human epithelial colorectal carcinoma (HCT 116) cells. METHODOLOGY: An extraction method without organic solvents was applied for the first time. The purification was carried out by droplet counter current chromatography (DCCC) coupled with high-performance liquid chromatography (HPLC) and electrospray ionisation mass spectrometry (ESI-MS) data. By nuclear magnetic resonance (NMR) [1 H, 13 C and two-dimensional (2D) experiments] pure components were identified. Wound healing and cell proliferation assays were utilised to determine the role of the isolated S. tectorum (SVT) fraction on cellular migration and proliferation. The signalling pathways elicited from the SVT fractions, were analysed by Western blot analysis. RESULTS: In this study two rare natural components were identified, namely monosaccharide sedoheptulose and polyalcohol 2-C-methyl-D-erythritol, along with known organic acids and flavonoids. The fractions with high level of sedoheptulose enhance the proliferation and the cellular migration of epithelial HCT 116 cells. The intracellular signalling cascades elicited from the purified fractions induce the c-Src-mediated transactivation of EGFR and the activation of the STAT3 pathway which, in turn, are crucially involved in the cellular proliferation and migration. CONCLUSIONS: Our study demonstrates the efficacy of purified fractions of S. tectorum L. in enhancing cellular proliferation and migration, suggesting their potential role as topical therapeutic treatments for wound healing.

Antioxidant activity and chemical components as potential anticancer agents in the olive leaf (Olea europaea L. cv Leccino.) decoction.

Epidemiological studies have shown that a reduced risk of chronic diseases such as cancer and cardiovascular diseases is correlated with a regular consumption of fruits and vegetable, many of which are rich in polyphenols. The additive and synergistic effect of phytochemicals in fruits and vegetables may reduce chronic diseases related to oxidative stress in human body. Olea europaea L. leaf are rich in phenolic components, which have been proposed to play a role in cancer prevention. The purpose of this study was to identify the main components in the Olea europaea L. leaf (cv. Leccino) preserved during the decoction preparation, in order to delineate the antioxidant activities of the crude extracts and its isolated compounds by using different in vitro assays including DPPH radicalscavenging capacity, total antioxidant capacity (TAC), xanthine oxidase (XO) inhibitory effect and the ability to delay the linoleic acid peroxidation process (ALP). The aqueous decoction was partitioned obtaining four extracts and the n-butanol extract showed the highest antioxidant activity and the highest total phenolic content. Phytochemical investigation leads to the isolation of thirteen secondary metabolites including simple phenolics, flavonoids, secoiridoids whose structures were elucidated by spectroscopic data (1D and 2D NMR) and spectrometric techniques. A significant free radical scavenging effect against DPPH has been evidenced in fraxamoside (1) (EC50 62.6 µM) and taxifolin (5) (EC50 50.0 µM), isolated for the first time from the water decoction. The most active compound in the TAC evaluation, was the 3,4 dihydro-phenyl glycol (8) (0.90 caffeic acid equiv.) while taxifolin and fraxamoside resulted as the most efficient inhibitors of XO activity (IC50 2.7 and 5.2 µM, respectively). Secoxyloganin (4), oleuropein (2) and tyrosol (6) showed the highest ALP activity. This study adds to the growing body of data supporting the bioactivities of phytochemicals and their potential impact on human health.

Novel steroidal components from the underground parts of Ruscus aculeatus L.

Two new furostanol saponins 1–2 and three new sulphated glycosides 3a,b and 4 were isolated from the underground parts of Ruscus aculeatus L., along with four known furostanol and one spirostanol saponins 5–9 and three free sterols. All of the structures have been elucidated on the basis of spectroscopic data 1D and 2D NMR experiments, MS spectra and GC analyses.

Imbricatolic acid from Juniperus communis L. prevents cell cycle progression in CaLu-6 cells.

Imbricatolic acid was isolated from the methanolic extract of the fresh ripe berries of Juniperus communis (Cupressaceae) together with sixteen known compounds and a new dihydrobenzofuran lignan glycoside named juniperoside A. Their structures were determined by spectroscopic methods and by comparison with the spectral data reported in literature. Imbricatolic acid was evaluated for its ability to prevent cell cycle progression in p53-null CaLu-6 cells. This compound induces the upregulation of cyclin-dependent kinase inhibitors and their accumulation in the G1 phase of the cell cycle, as well as the degradation of cyclins A, D1, and E1. Furthermore, no significant imbricatolic acid-induced apoptosis was observed. Therefore, this plant-derived compound may play a role in the control of cell cycle.

Lauroside B, a megastigmane glycoside from Laurus nobilis (bay laurel) leaves, induces apoptosis in human melanoma cell lines by inhibiting NF-κB activation.

Malignant melanoma is a highly aggressive tumor that frequently resists chemotherapy, so the search for new agents for its treatment is of great importance. In the present study, the antiproliferative propensity against human melanoma cell lines of lauroside B (1), a megastigmane glycoside isolated from Laurus nobilis (bay laurel) leaves, was investigated. This compound suppressed the proliferation of three human melanoma cell lines, namely, A375, WM115, and SK-Mel-28. The 1-induced inhibition of human melanoma cell proliferation was due to the induction of apoptosis, as demonstrated by FACS analysis with annexin V/PI staining and confirmed by activation of caspase-3 and by the cleavage of poly(ADP-ribose) polymerase (PARP). Growing evidence implicates NF-κB as an important contributor to metastasis and increased chemoresistance of melanoma. Thus, it was hypothesized that 1-induced apoptosis could be associated with suppression of NF-κB activation. The results showed that exposure of human melanoma cells to 1 inhibited IκB-α degradation and constitutive NF-κB DNA-binding activity as well as the expression, regulated by NF-κB, of two antiapoptotic genes, XIAP and c-FLIP. Induction of apoptosis by 1 in human aggressive melanoma cell lines has a potential high biological value.

Identification of minor secondary metabolites from the latex of Croton lechleri (Muell-Arg) and evaluation of their antioxidant activity.

Dragon's blood (Sangre de drago), a viscous red sap derived from Croton lechleri Muell-Arg (Euphorbiaceae), is extensively used by indigenous cultures of the Amazonian basin for its wound healing properties. The aim of this study was to identify the minor secondary metabolites and test the antioxidant activity of this sustance. A bioguided fractionation of the n-hexane, chloroform, n-butanol, and aqueous extracts led to the isolation of 15 compounds: three megastigmanes, four flavan-3-ols, three phenylpropanoids, three lignans, a clerodane, and the alkaloid taspine. In addition to these known molecules, six compounds were isolated and identified for the first time in the latex: blumenol B, blumenol C, 4,5-dihydroblumenol A, erythro-guaiacyl-glyceryl-beta-O-4'- dihydroconiferyl ether, 2-[4-(3-hydroxypropyl)-2-methoxyphenoxy]-propane-1,3-diol and floribundic acid glucoside. Combinations of spectroscopic methods ((1)H-, (13)C- NMR and 2D-NMR experiments), ESI-MS, and literature comparisons were used for compound identification. In vitro antioxidant activities were assessed by DPPH, total antioxidant capacity and lipid peroxidation assays. Flavan-3-ols derivatives (as major phenolic compounds in the latex) exhibited the highest antioxidant activity.

Identification of a new sesquiterpene polyol ester from Celastrus paniculatus.

A new sesquiterpene polyol ester characterized as 1alpha,6beta,8beta-triacetoxy-9beta-benzoyloxydihydro-beta-agarofuran (1), along with the three known compounds: 1alpha,6beta,8alpha-triacetoxy-9alpha-benzoyloxydihydro-beta-agarofuran (2), angulatueoid C (3), and 1alpha,6beta,8beta,14-tetraacetoxy-9alpha-benzoyloxydihydro-beta-agarofuran (4), was isolated from the CCl (4)-soluble fraction of Celastrus paniculatus methanolic seed extract. All four compounds produce at a concentration of 1 microg/mL a relaxant effect on the isolated rat ileum of 31.4 +/- 7.5, 24.3 +/- 9.6, 24.4 +/- 4.9, and 16.2 +/- 7.2 %, respectively.

Phenolic glycosides from Foeniculum vulgare fruit and evaluation of antioxidative activity.

Two diglucoside stilbene trimers and a benzoisofuranone derivative were isolated from Foeniculum vulgare fruit together with nine known compounds. Their structures were elucidated by spectral methods including 1D, 2D NMR and MS and chemical methods. Antioxidant activity was tested using three methods: DPPH(), total antioxidant capacity and assay of lipid peroxidation.

New constituents of sweet Capsicum annuum L. fruits and evaluation of their biological activity.

Four new acyclic diterpene glycosides named capsianosides (1-4), together with 12 known compounds, were isolated from the fresh sweet pepper fruits of Capsicum annuum L., a plant used as a vegetable food, spice, and external medicine. The chemical structures of new natural compounds, as well as their absolute configurations, were established by means of spectroscopic data including infrared, high-resolution mass spectrometry, and one- and two-dimensional nuclear magnetic resonance and by chemical derivatization. The known capsidiol (11) showed bacteriostatic properties in vitro against Helicobacter pylori with a minimum inhibitory concentration (MIC) of 200 microg/mL when compared with the commercial drug metronidazole (MIC, 250 microg/mL). Some purified components were also tested for their antioxidant activities.

New sesquiterpene lactones from Laurus nobilis leaves as inhibitors of nitric oxide production.

Two new metabolites 5alphaH,7alphaH-eudesman-4alpha,6alpha,11,12-tetraol (1) and 1beta,15-dihydroxy-5alphaH,7alphaH-eudesma-3,11(13)-dien-12,6alpha-olide ( 2) have been isolated from the methanolic extract of Laurus nobilis L. leaves. Their structures were determined through analysis of their one- and two-dimensional NMR spectral data ((1)H- and (13)C-NMR, DEPT, COSY, HMQC, HMBC and ROESY). The relative stereochemistry is proposed on the basis of combined J-based configuration analysis and ROESY data. In addition, three known sesquiterpene lactones santamarine (3), reynosin (4) and costunolide (5) along with blumenol C (6) were isolated and identified by spectral means. The isolated compounds 1 - 6 were found to inhibit nitric oxide (NO) production in lipopolysaccharide (LPS)-activated murine macrophages. The most active compound 2 potently inhibited NO (2)(-) release with an IC (50) value of 0.8 microM.

New sesquiterpenes with intestinal relaxant effect from Celastrus paniculatus.

The seed oil obtained from Celastrus paniculatus has been used in Ayurvedic medicine for the treatment of several complaints including intestinal disorders. The muscle relaxing activity of C. paniculatus has been studied in vitro on isolated preparations of rat intestine. C. paniculatus seeds extract (0.001 - 100 microg/mL) produced a concentration-related relaxation of the rat ileum [IC (50) (95 % c. l.) 0.24 (0.13 - 0.45) microg/mL]. Among the four fractions ( n-hexane, CCl (4), CHCl (3), n-BuOH) obtained from the methanolic extract of C. paniculatus, only the carbon tetrachloride fraction (0.01 - 10 000 ng/mL) induced a significant relaxation of the intestinal muscle [IC (50) (95 % c. l.): 1.9 (1.38 - 2.87) ng/mL]. Three new sesquiterpene polyol esters have been isolated from the carbon tetrachloride fraction. These new metabolites, characterised as 1alpha,8beta,14-triacetoxy-9beta-furoyloxydihydro-beta-agarofuran ( 1), 1alpha, 6beta,8beta,14-tetraacetoxy-9beta-benzoyloxydihydro-beta-agarofuran ( 2) and 1alpha,8beta-diacetoxy-9beta-benzoyloxydihydro-beta-agarofuran ( 3), at the concentration of 1 microg/mL produced a relaxant effect of 30.6 +/- 12.2 %, 26.9 +/- 4.7 % and 7.27 +/- 1.7 %, respectively.

Megastigmane and phenolic components from Laurus nobilis L. leaves and their inhibitory effects on nitric oxide production.

Laurus nobilis L. leaves are widely used in cooking and in folk medicine. Five new megastigmane glucosides (2-4, 7, and 9) named laurosides A-E and a new phenolic glucoside 12 were isolated from the methanolic extract of L. nobilis L. leaves, along with 10 known components: megastigmane (5), megastigmane glucosides (1, 6, 8, 10, and 11), aromatic compounds (13 and 14), and flavonoids (15 and 16). The structures and relative stereochemistry have been elucidated by one- and two-dimensional nuclear magnetic resonance experiments ((1)H and (13)C NMR, DEPT, correlation spectroscopy, heteronuclear multiple quantum correlation, heteronuclear multiple bond correlation, and nuclear Overhauser enhancement spectroscopy) and by chemical derivatization. The effect of isolated compounds on nitric oxide production in lipopolysaccharide-activated murine macrophages were examined.

Variation in the composition of the heartwood flavonoids of Prunus avium by on-column capillary gas chromatography.

A fast and simple extraction procedure, coupled with on-column gas chromatography, has been developed in order to investigate the relative amounts of flavonoids in individual trees within several homogeneous populations of Prunus avium. Eight known flavonoid aglycones were isolated from the heartwood of P. avium and multivariate analysis was employed in order to characterise population groups.

Bioactive asterosaponins from the starfish Luidia quinaria and Psilaster cassiope. Isolation and structure characterization by two-dimensional NMR spectroscopy.

An investigation of the polar extracts from two starfish, Luidia quinaria and Psilaster cassiope, led to the isolation of five sulfated "asterosaponins". Two of them, named luidiaquinoside (1) and psilasteroside (2), are new compounds. Luidiaquinoside (1) contains a novel pentasaccharide chain composed of d-glucose, d-quinovose, and d-fucose, with the d-glucose unit being the branching point. Psilasteroside (2) contains a hexasaccharide chain in which an arabinose residue is detectable in the furanose form. Both of these compounds possess a Delta(9(11)),3beta,6alpha-dihydroxysteroidal nucleus with a 20-hydroxy, 23-oxo functionality. The structures of the new asterosaponins were elucidated by a combination of NMR techniques including (1)H-(1)H (COSY, TOCSY, and ROESY) and (1)H-(13)C (HMQC and HMBC) spectroscopy, ESIMS and HRFABMS spectrometry, and GC analyses. The new asterosaponins show marginal in vitro cytotoxicity against RBL-2H3 (rat basophilic leukemia) cell lines.

Minor steroidal alkaloids from the marine sponge Corticium sp.

Four new steroidal alkaloids, plakinamine G (1), plakinamine H (2), 4alpha-hydroxydemethylplakinamine B (3), and tetrahydroplakinamine A (4), along with three known compounds, were isolated from the marine sponge Corticium sp. The structures of these metabolites were elucidated largely by 1D and 2D NMR methods and accurate mass measurements (HR-EIMS). Compounds 1, 2, and 4 show significant in vitro cytotoxicity.

Antimicrobial furostanol saponins from the seeds of Capsicum annuum L. var. acuminatum.

Three new furostanol saponins named capsicoside E (1), capsicoside F (2), and capsicoside G (5) were obtained from the seeds of Capsicum annuum L. var. acuminatum along with known oligoglycosides (3, 4, and 6-10). On the basis of chemical and spectroscopic analyses, the structures of these new furostanol oligoglycosides were elucidated as 26-O-beta-D-glucopyranosyl-22-O-methyl-5alpha-furost-25(27)-en-2alpha,3beta,22xi,26-tetraol-3-O-beta-D-glucopyranosyl(1-->3)-beta-D-glucopyranosyl(1-->2)-[beta-D-glucopyranosyl(1-->3)]-beta-D-glucopyranosyl(1-->4)-beta-D-galactopyranoside (1), 26-O-beta-D-glucopyranosyl-(25R)-5alpha-furost-20(22)-en-2alpha,3beta,26-triol-3-O-beta-D-glucopyranosyl (1-->3)-beta-D-glucopyranosyl(1-->2)-[beta-D-glucopyranosyl(1-->3)]-beta-D-glucopyranosyl(1-->4)-beta-D-galactopyranoside (2), and 26-O-beta-D-gluco-pyranosyl-(25R)-5alpha-furosta-3beta,22xi,26-triol-3-O-beta-D-glucopyranosyl(1-->3)-beta-D-glucopyranosyl(1-->2)-[beta-D-glucopyranosyl(1-->3)]-beta-D-glucopyranosyl(1-->4)-beta-D-galactopyranoside (5). The isolated saponins showed higher antimicrobial activity against yeasts than against common fungi. Data indicated that the antiyeast activity was related to the combination of the oligosaccharide chain (S1, S2, or S3) with an O-methyl group at R(3) and the presence of a hydroxyl group at the C-2 position.