Isolation of chemical constituents from Filago vulgaris and antiproliferative activity of the plant extract and its flavonoid against human tumor cell lines.

Phytochemical investigation of the whole plant of Filago vulgaris Lam. (Asteraceae) resulted in the isolation and characterization of seven compounds, including a rare methoxylated flavonol (araneol), tetrahydrofurofuranolignans (pinoresinol and syringaresinol), p-hydroxybenzaldehyde, vanillin, vanillic acid and scopoletin. The structures of the compounds were determined by NMR and mass spectroscopy. All compounds were first obtained from this species and reported for the genus Filago. Our results demonstrate that highly methoxylated flavonols lacking substituents on ring B and lignans can be regarded as taxonomic markers for the tribe Inuleae. The lipophilic extract of F. vulgaris was found to have antiproliferative activity against HeLa cells (62.1±0.9% inhibition at 30 µ/ml), and araneol was highly effective against this tumour cell line (IC50 8.36 µ M).

Sesquiterpene Lactones and Flavonoids from Psephellus pyrrhoblepharus with Antiproliferative Activity on Human Gynecological Cancer Cell Lines.

Multistep chromatographic separations of the chloroform extract of the Turkish endemic plant Psephellus pyrrhoblepharus (Boiss.) Wagenitz (syn. Centaurea pyrrhoblephara Boiss.) resulted in the isolation of six guaianolid-type sesquiterpenes, chlorojanerin (1), 19-deoxychlorojanerin (2), 15-hydroxyjanerin (3), aguerin B (4), cynaropicrin (5), eleganin (6); three flavonoids, apigenin, 6-methoxyluteolin and jaceosidine; two glycosides, benzyl-1-O-ß-d-glucoside and 3(Z)-hexenyl-1-O-ß-d-glucoside; and the coumarin scopoletin. The structures were established by the interpretation of their ESI-MS and 1D and 2D NMR data including 1H-NMR, JMOD, 1H,1H-COSY, HSQC, HMBC, and NOESY experiments. All compounds were isolated for the first time from P. pyrrhoblepharus. Compounds 1-6, the isolated flavonoids and scopoletin were evaluated for their antiproliferative activities on human gynecological cancer cell lines (SiHa, HeLa, and MDA-MB-231 cells) using the MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay. Chlorojanerin (1), 19-deoxychlorojanerin (2), aguerin B (4), cynaropicrin (5), eleganin (6) were shown to have noteworthy effects on all of the tested cell lines, while apigenin, jaceosidine, and 6-methoxyluteolin were moderately active on HeLa cells. The highest activities were demonstrated by the chlorine-containing derivatives chlorojanerin (1) and 19-deoxychlorojanerin (2) with IC50 values of 2.21 and 2.88 µM, respectively, against the triple negative breast cancer model MDA-MB-231 cells.

Isolation of Chemical Constituents of Centaurea virgata Lam. and Xanthine Oxidase Inhibitory Activity of the Plant Extract and Compounds.

BACKGROUND: Centaurea virgata Lam. is a species widely used in the traditional medicine in Turkey for the treatment of diabetes, allergy and gastric ulcers. The rationale of its use in the therapy has not been studied previously, therefore the present work aimed at the chemicalpharmacological evaluation of the plant. OBJECTIVE: The xanthine oxidase (XO) inhibitory activity of the MeOH extract and its subextracts (n-hexane, CHCl3 and remaining MeOH-H2O) prepared from C. virgata was investigated in vitro. Moderate activity was exerted in case of the CHCl3 extract (98.9 ± 15.8 µg/mL), therefore constituents of this extract were analysed. METHOD: Different purification steps, such as VLC, CPC, PLC and crystallization were used for the isolation, and ESIMS, NMR, LC-MS and authentic standards were applied for identification of the compounds. XO inhibitory and DPPH assays were used for evaluation of the bioactivities. RESULTS: Sesquiterpenes [8α-hydroxysonchucarpolide, 8α-(3,4-dihydroxy-2-methylenebutanoyloxy)- dehydromelitensine, and cnicin], flavones (apigenin, hispidulin, salvigenin, eupatorin, 3'-methyleupatorin), and the flavonol isokaempferide were isolated from the active extract. The XO-inhibitory activity of these compounds was analyzed using allopurinol as a positive control (IC50 7.49 ± 0.29 µM). It was found that sesquiterpenes and flavonoids, containing 7- OMe group, are inactive. CONCLUSION: 7-Hydroxyflavones (apigenin and hispidulin) exerted significant XO inhibitory effect with IC50 values of 0.99 ± 0.33 µM and 4.88 ± 1.21 µM, respectively. Therefore, these compounds are responsible for the XO-inhibitory effect of the extract. The free radical scavenging activity of the isolated flavonoids was determined by DPPH assay, and it was stated that none of the compounds have substantial antioxidant activity, therefore the reduced generation of reactive oxygen species may be the consequence only of XO inhibition.

Investigation of the Antiproliferative Properties of Natural Sesquiterpenes from Artemisia asiatica and Onopordum acanthium on HL-60 Cells in Vitro.

Plants and plant extracts play a crucial role in the research into novel antineoplastic agents. Four sesquiterpene lactones, artecanin (1), 3ß-chloro-4α,10α-dihydroxy-1α,2α-epoxy-5α,7αH-guaia-11(13)-en-12,6α-olide (2), iso-seco-tanapartholide 3-O-methyl ether (3) and 4ß,15-dihydro-3-dehydrozaluzanin C (4), were isolated from two traditionally used Asteraceae species (Onopordum acanthium and Artemisia asiatica). When tested for antiproliferative action on HL-60 leukemia cells, these compounds exhibited reasonable IC50 values in the range 3.6-13.5 µM. Treatment with the tested compounds resulted in a cell cycle disturbance characterized by increases in the G1 and G2/M populations, while there was a decrease in the S phase. Additionally, 1-3 elicited increases in the hypodiploid (subG1) population. The compounds elicited concentration-dependent chromatin condensation and disruption of the membrane integrity, as revealed by Hoechst 33258-propidium staining. Treatment for 24 h resulted in significant increases in activity of caspases-3 and -9, indicating that the tested sesquiterpenes induced the mitochondrial pathway of apoptosis. The proapoptotic properties of the sesquiterpene lactones were additionally demonstrated withannexin V staining. Compounds 1 and 2 increased the Bax/Bcl-2 expression and decreased the expressions of CDK1 and cyclin B2, as determined at the mRNA level by means of RT-PCR. These experimental results indicate that sesquiterpene lactones may be regarded as potential starting structures for the development of novel anticancer agents.

Effects of Chelidonium majus extracts and major alkaloids on hERG potassium channels and on dog cardiac action potential - a safety approach.

Chelidonium majus or greater celandine is spread throughout the world, and it is a very common and frequent component of modern phytotherapy. Although C. majus contains alkaloids with remarkable physiological effect, moreover, safety pharmacology properties of this plant are not widely clarified, medications prepared from this plant are often used internally. In our study the inhibitory effects of C. majus herb extracts and alkaloids on hERG potassium current as well as on cardiac action potential were investigated. Our data show that hydroalcoholic extracts of greater celandine and its alkaloids, especially berberine, chelidonine and sanguinarine have a significant hERG potassium channel blocking effect. These extracts and alkaloids also prolong the cardiac action potential in dog ventricular muscle. Therefore these compounds may consequently delay cardiac repolarization, which may result in the prolongation of the QT interval and increase the risk of potentially fatal ventricular arrhythmias.

Antiproliferative activity of Artemisia asiatica extract and its constituents on human tumor cell lines.

The extract of Artemisia asiatica herb with antiproliferative activity against four human tumor cell lines (A2780, A431, HeLa, and MCF7) was analyzed by the MTT assay, and bioassay-directed fractionation was carried out in order to identify the compounds responsible for the cytotoxic activity. Guaianolide (1-4), seco-guianolide (5), germacranolide (6) and eudesmanolide sesquiterpenes (7), monoterpenes (8, 9), including the new compound artemisia alcohol glucoside (8), and flavonoids (10-16) were isolated as a result of a multistep chromatographic procedure (CC, CPC, PLC, and gel filtration). The compounds were identified by means of UV, MS, and NMR spectroscopy, including (1)H-and (13)C-NMR, (1)H-(1)H COSY, NOESY, HSQC, and HMBC experiments. The isolated compounds 1-16 were evaluated for their tumor cell growth-inhibitory activities on a panel of four adherent cancer cell lines, and different types of secondary metabolites were found to be responsible for the cytotoxic effects of the extract. Especially cirsilineol (13), 3ß-chloro-4α,10α-dihydroxy-1α,2α-epoxy-5α,7αH-guai-11(13)-en-12,6α-olide (3), and iso-seco-tanapartholide 3-O-methyl ester (5) exerted marked cytotoxic effects against the investigated cell lines, while jaceosidin (12), 6-methoxytricin (15), artecanin (2), and 5,7,4',5'-tetrahydroxy-6,3'-dimethoxyflavone (14) were moderately active. All the sesquiterpenes and monoterpenes are reported here for the first time from this species, and in the case of artecanin (2), 3α-chloro-4ß,10α-dihydroxy-1ß,2ß-epoxy-5α,7αH-guai-11(13)-en-12,6α-olide (4), ridentin (6), and ridentin B (7), previously unreported NMR spectroscopic data were determined.

Pest management systems affect composition but not abundance of phytoseiid mites (Acari: Phytoseiidae) in apple orchards.

We examined the faunal composition and abundance of phytoseiid mites (Acari: Phytoseiidae) in apple orchards under different pest management systems in Hungary. A total of 30 apple orchards were surveyed, including abandoned and organic orchards and orchards where integrated pest management (IPM) or broad spectrum insecticides (conventional pest management) were applied. A total of 18 phytoseiid species were found in the canopy of apple trees. Species richness was greatest in the organic orchards (mean: 3.3 species/400 leaves) and the least in the conventional orchards (1.4), with IPM (2.1) and abandoned (2.7) orchards showing intermediate values. The phytoseiid community's Rényi diversity displayed a similar pattern. However, the total phytoseiid abundance in the orchards with different pest management systems did not differ, with abundance varying between 1.8 and 2.6 phytoseiids/10 leaves. Amblyseius andersoni, Euseius finlandicus, and Typhlodromus pyri were the three most common species. The relative abundance of A. andersoni increased with the pesticide load of the orchards whereas the relative abundance of E. finlandicus decreased. The abundance of T. pyri did not change in the apple orchards under different pest management strategies; regardless of the type of applied treatment, they only displayed greater abundance in five of the orchards. The remaining 15 phytoseiid species only occurred in small numbers, mostly from the abandoned and organic orchards. We identified a negative correlation between the abundance of T. pyri and the other phytoseiids in the abandoned and organic orchards. However, we did not find any similar link between the abundance of A. andersoni and E. finlandicus.

Antiproliferative constituents of the roots of Conyza canadensis.

Bioassay-guided fractionation of the N-hexane and CHCl3 phases of the methanol extract of the roots of Conyza canadensis (L.) Cronquist led to the isolation of two new dihydropyranones named conyzapyranone A (1) and B (2), and the known 4 Z,8 Z-matricaria- γ-lactone (3), 4 E,8 Z-matricaria- γ-lactone (4), 9,12,13-trihydroxy-10(E)-octadecenoic acid (5), epifriedelanol (6), friedeline (7), taraxerol (8), simiarenol (9), spinasterol (10), stigmasterol, ß-sitosterol, and apigenin. The structures were determined by means of ESIMS and 1D and 2D NMR spectroscopy, including ¹H-¹H COSY, NOESY, HSQC, and HMBC experiments. The isolated compounds were evaluated for their antiproliferative activities and were demonstrated to exert considerable cell growth-inhibitory activity against human cervix adenocarcinoma (HeLa), skin carcinoma (A431), and breast adenocarcinoma (MCF-7) cells. Some of the active components, including 2, 4, and 10, proved to be substantially more potent against these cell lines than against noncancerous human foetal fibroblasts (MRC-5) and can therefore be considered selective antiproliferative natural products.

Bioactivity-guided isolation of antiproliferative compounds from Centaurea arenaria.

The antiproliferative effects of n-hexane, chloroform and aqueous methanol extracts prepared from the whole plant of Centaurea arenaria M.B. ex Willd. were investigated against cervix adenocarcinoma (HeLa), breast adenocarcinoma (MCF7) and skin epidermoid carcinoma (A431) cells, using the MTT assay. The chloroform extract displayed high tumour cell proliferation inhibitory activity (higher than 85% at 10 µg/mL concentration), and was therefore subjected to a bioassay-guided multistep separation procedure. Flavonoids (eupatilin, eupatorin, 3'-methyleupatorin, apigenin and isokaempferid), lignans (arctigenin, arctiin and matairesinol), the sesquiterpene cnicin, serotonin conjugates (moschamine and cis-moschamine), ß-amyrin and ß-sitosterin-ß-D-glycopyranoside, identified by means of UV, MS and NMR spectroscopy, were obtained for the first time from this species. The isolated compounds were also evaluated for their tumour cell growth inhibitory activities on HeLa, MCF7 and A431 cells, and different types of secondary metabolites were found to be responsible for the antitumour effects of the extracts; in addition to moderately active compounds (isokaempferid and moschamine), especially apigenin, eupatorin, arctigenin, arctiin, matairesinol and cnicin exert marked antitumour effects against these cell lines.

Bioactivity-guided isolation of cytotoxic sesquiterpenes and flavonoids from Anthemis ruthenica.

A new eudesmanolide sesquiterpene, sivasinolide 6-O-angelate (1), was isolated from the aerial parts of Anthemis ruthenica together with the known compounds chrysanin (2), tanacin (3), 3 beta-hydroxycostunolide (4), centauridin (5), and centaureidin (6). The compounds were obtained by means of bioactivity-guided fractionation from the CHCl (3) extract of the herb, which displayed high cytotoxic activity. The structures were determined by UV, HR-ESI-MS, and high-field 1D and 2D NMR spectral analyses, affording complete (1)H- and (13)C-NMR assignments for all compounds. The cytotoxic activities of the isolated sesquiterpenes and flavonoids were assessed against cervical adenocarcinoma HeLa, breast adenocarcinoma MCF7, and skin epidermoid carcinoma A431 cells using the MTT assay. It was found that, apart from centaureidin (6), which is extremely active (IC(50) 0.082, 0.13, and 0.35 microM on the HeLa, MCF7, and A431 cell lines, respectively), all these compounds exert high or moderate tumor cell-growth inhibitory activity (IC(50) 3.42-58.15 microM).

Antiproliferative activity of Hungarian Asteraceae species against human cancer cell lines. Part II.

The antiproliferative activities of aqueous and organic extracts prepared from 26 Hungarian species of the tribes Cynereae and Lactuceae (Asteraceae) were tested in vitro against HeLa (cervix epithelial adenocarcinoma), A431 (skin epidermoid carcinoma) and MCF7 (breast epithelial adenocarcinoma) cells by using the MTT assay. Of the tested 200 extracts of different plant parts obtained with n-hexane, chloroform, 50% methanol and water, 16 extracts displayed noteworthy cell growth inhibitory activity (>50% inhibition at a concentration of 10 microg/mL). The IC50 values of these extracts were determined, and their direct cytotoxic effects were measured. High differences between the antiproliferative and cytotoxic activities, demonstrating a real cell proliferation inhibitory activity rather than direct killing effects, were found for some Centaurea, Cirsium, Cichorium, Lactuca, Onopordum and Scorsonera extracts.

Antiproliferative effect of flavonoids and sesquiterpenoids from Achillea millefolium s.l. on cultured human tumour cell lines.

The antiproliferative activities of n-hexane, chloroform, aqueous-methanol and aqueous extracts of the aerial parts of the Achillea millefolium aggregate on three human tumour cell lines were investigated by means of MTT assays. The chloroform-soluble extract exerted high tumour cell proliferation inhibitory activities on HeLa and MCF-7 cells, and a moderate effect on A431 cells; accordingly, it was subjected to detailed bioactivity-guided fractionation. As a result of the multistep chromatographic purifications (VLC, CPC, PLC, gel filtration), five flavonoids (apigenin, luteolin, centaureidin, casticin and artemetin) and five sesquiterpenoids (paulitin, isopaulitin, psilostachyin C, desacetylmatricarin and sintenin) were isolated and identified by spectroscopic methods. The antiproliferative assay demonstrated that centaureidin is the most effective constituent of the aerial parts of yarrow: high cell growth inhibitory activities were observed especially on HeLa (IC(50) 0.0819 microm) and MCF-7 (IC(50) 0.1250 microm) cells. Casticin and paulitin were also highly effective against all three tumour cell lines (IC(50) 1.286-4.76 microm), while apigenin, luteolin and isopaulitin proved to be moderately active (IC(50) 6.95-32.88 microm). Artemetin, psilostachyin C, desacetylmatricarin and sintenin did not display antiproliferative effects against these cell lines. This is the first report on the occurrence of seco-pseudoguaianolides (paulitin, isopaulitin and psilostachyin C) in the Achillea genus.

Antiproliferative activity of Hungarian Asteraceae species against human cancer cell lines. Part I.

Aqueous and organic extracts of 25 selected species from four tribes of Hungarian Asteraceae were screened in vitro for antiproliferative activity against HeLa (cervix epithelial adenocarcinoma), A431 (skin epidermoid carcinoma) and MCF7 (breast epithelial adenocarcinoma) cells, using the MTT assay. Twenty five of the 228 tested extracts from different parts of the species of Astereae (6), Inuleae (3), Heliantheae (5) and Anthemideae (11) demonstrated a substantial antiproliferative effect (at least 50% inhibition of cell proliferation) at 10 microg/mL against one or more of the cell lines. Complete dose-response curves were generated and IC(50) values were calculated for these active extracts, and their direct cytotoxic effects were determined. In summary, 11 of the tested 25 plants were found to be active and 4 of them (Anthemis ruthenica, Erigeron canadensis, Erigeron annuus and Inula ensifolia) had not been studied previously for either active compounds or anticancer properties.

Diterpenoids and flavonoids from the fruits of Vitex agnus-castus and antioxidant activity of the fruit extracts and their constituents.

From the n-hexane fraction of the fruits of Vitex agnus-castus, two labdane-type diterpenes, vitetrifolin B and C, were isolated by means of multiple chromatographic separations, together with the previously identified rotundifuran, vitexilactone and the sesquiterpene spathulenol. From the EtOAc fraction, eupatorin was identified for the first time, besides the known casticin, penduletin, vitexin and orientin. The n-hexane, EtOAc and MeOH-H(2)O fractions of the MeOH extract of Agni-casti fructus were subjected to in vitro antioxidant assays. The EtOAc extract displayed a significant concentration-dependent effect when tested by 1,1-diphenyl-2-picrylhydrasyl (DPPH) free radical assay (IC(50) = 68 microg/mL) and against the autooxidation of a standard rat brain homogenate (IC(50) = 14 microg/mL). The MeOH-H(2)O fraction was less active with 3643 microg/mL (DPPH test) and IC(50) = 125 microg/mL (rat brain homogenate), while the n-hexane phase proved to be inactive. The main flavonoid constituents of the EtOAc extract, casticin, vitexin and orientin were assayed for antioxidant activity and found that only casticin possesses a marked lipid peroxidation inhibitory effect (IC(50) = 0.049 mm) compared with that of the positive control ascorbic acid (IC(50) = 0.703 mm).

Norditerpenoid alkaloids from Consolida orientalis and complete 1H and 13C NMR signal assignments of some lycoctonine-type alkaloids.

A new norditerpene alkaloid was isolated, 18-demethylpubescenine (1), in addition to four known compounds, 14-demethyltuguaconitine (2), takaosamine (3), gigactonine (4), and delcosine (5), from fresh, whole plants of Consolida orientalis. The structure of 1 was established by spectroscopic methods, including various 2D NMR techniques and HRESIMS. As a result of a detailed NMR study, complete 1H NMR chemical shift assignments for alkaloids 1-5 are presented herein, and some 13C NMR signal assignments for 2-4 have been revised.