Isolation of compounds from the roots of Ambrosia artemisiifolia and their effects on human cancer cell lines.

Common ragweed (Ambrosia artemisiifolia L.) is an invasive plant in Europe with spreading use in the contemporary folk medicine. The chemical composition of the above-ground parts is extensively studied, however, the metabolites of the roots are less discovered. By multiple chromatographic purification of the root extracts, we isolated thiophene A (1), n-dodecene (2), taraxerol-3-O-acetate (3), α-linoleic acid (4), (+)-pinoresinol (5), and thiophene E (7,10-epithio-7,9-tridecadiene-3,5,11-triyne-1,2-diol) (6). The 1H NMR data published earlier for 1 were supplemented together with the assignment of 13C NMR data. Thiophene E (6), which is reported for the first time from this species, exerted cytotoxic and antiproliferative effects on A-431 epidermoid skin cancer cells, whereas taraxerol-3-O-acetate (3) and α-linoleic acid (4) had slight antiproliferative effect on gynecological cancer cell lines. Thiophene E (6) and taraxerol-3-O-acetate (3) displayed antiproliferative and cytotoxic effects on MRC-5 fibroblast cells. Thiophene E (6) exerted weak antibacterial activity (MIC 25 µg/mL) on MRSA ATCC 43300, on Staphylococcus aureus ATCC 25923, Escherichia coli AG100 and E. coli ATCC 25922 both thiophenes were inactive. Although the isolated compounds exerted no remarkable cytotoxic or antiproliferative activities, the effects on MRC-5 fibroblast cells highlight the necessity of further studies to support the safety of ragweed root.

Aconitum Alkaloid Songorine Exerts Potent Gamma-Aminobutyric Acid-A Receptor Agonist Action In Vivo and Effectively Decreases Anxiety without Adverse Sedative or Psychomotor Effects in the Rat.

Songorine (SON) is a diterpenoid alkaloid from Aconitum plants. Preparations of Aconitum roots have been employed in traditional oriental herbal medicine, however, their mechanisms of action are still unclear. Since GABA-receptors are possible brain targets of SON, we investigated which subtypes of GABA-receptors contribute to the effects of SON, and how SON affects anxiety-like trait behavior and psychomotor cognitive performance of rats. First, we investigated the effects of microiontophoretically applied SON alone and combined with GABA-receptor agents picrotoxin and saclofen on neuronal firing activity in various brain areas. Next, putative anxiolytic effects of SON (1.0-3.0 mg/kg) were tested against the GABA-receptor positive allosteric modulator reference compound diazepam (1.0-5.0 mg/kg) in the elevated zero maze (EOM). Furthermore, basic cognitive effects were assessed in a rodent version of the psychomotor vigilance task (PVT). Local application of SON predominantly inhibited the firing activity of neurons. This inhibitory effect of SON was successfully blocked by GABA(A)-receptor antagonist picrotoxin but not by GABA(B)-receptor antagonist saclofen. Similar to GABA(A)-receptor positive allosteric modulator diazepam, SON increased the time spent by animals in the open quadrants of the EOM without any signs of adverse psychomotor and cognitive effects observed in the PVT. We showed that, under in vivo conditions, SON acts as a potent GABA(A)-receptor agonist and effectively decreases anxiety without observable side effects. The present findings facilitate the deeper understanding of the mechanism of action and the widespread pharmacological use of diterpene alkaloids in various CNS indications.

Antiproliferative and cytotoxic effects of sesquiterpene lactones isolated from Ambrosia artemisiifolia on human adenocarcinoma and normal cell lines.

CONTEXT: Ambrosia artemisiifolia L. (Asteraceae) contains sesquiterpene lactones as characteristic secondary metabolites. Many of these compounds exert antiproliferative and cytotoxic effects. OBJECTIVE: To isolate the sesquiterpene lactones from the aerial part of A. artemisiifolia and to elucidate their cytotoxic, antiproliferative and antibacterial effects. MATERIALS AND METHODS: The compounds were identified by one-dimensional (1D) and 2D NMR, HR-MS spectroscopy from the methanol extract. Isolated compounds were investigated for their cytotoxic and antiproliferative effects on human colonic adenocarcinoma cell lines and human embryonal lung fibroblast cell line using MTT assay. The selectivity of the sesquiterpenes was calculated towards the normal cell line. To check the effect of drug interactions between compounds and doxorubicin, multidrug-resistant Colo 320 cells were used. RESULTS: A new seco-psilostachyinolide derivative, 1,10-dihydro-1'-noraltamisin, and seven known compounds were isolated from the methanol extract. Acetoxydihydrodamsin had the most potent cytotoxic effect on sensitive (Colo205) cell line (IC50 = 7.64 µM), also the strongest antiproliferative effect on Colo205 (IC50 = 5.14 µM) and Colo320 (IC50 = 3.67 µM) cell lines. 1'-Noraltamisin (IC50 = 8.78 µM) and psilostachyin (IC50 = 5.29 µM) showed significant antiproliferative effects on the multidrug-resistant Colo320 cell line and had moderate selectivity against human embryonal lung fibroblast cell line. Psilostachyin C exhibited cytotoxic effects on Colo205 cells (IC50 = 26.60 µM). None of the isolated compounds inhibited ABCB1 efflux pump (EP; P-glycoprotein) or the bacterial EPs. DISCUSSION AND CONCLUSIONS: Acetoxydihydrodamsin, 1'-noraltamisin, and psilostachyin showed the most remarkable cytotoxic and antiproliferative activity on tumour cell lines and exerted selectivity towards MRC-5 cell line.

Bearberry in the treatment of acute uncomplicated cystitis (BRUMI): protocol of a multicentre, randomised double-blind clinical trial.

BACKGROUND: Bearberry (Arctostaphylos uva-ursi) leaf is available as a treatment of uncomplicated cystitis in several European countries. The antimicrobial activity of its extracts and some of its individual constituents has been observed in vitro; however, the efficacy of bearberry compared with standard antimicrobial therapy has not been assessed yet. OBJECTIVE: The objective of the study is to assess the safety and non-inferiority of bearberry as an alternative therapy in the treatment of acute uncomplicated cystitis in comparison with standard antibiotic therapy (fosfomycin). METHODS AND ANALYSIS: This is a randomised controlled double-blinded multicentre trial. Eligible patients will be premenopausal women with a sum score of ≥6 for the typical acute uncomplicated cystitis symptoms (frequency, urgency, painful urination, incomplete emptying, suprapubic pain and visible haematuria) reported on the Acute Cystitis Symptom Score (ACSS) typical domain and pyuria. Patients will be randomly assigned to receive 3 g single dose of fosfomycin powder and two placebo tablets three times a day for 7 days or B a single dose of placebo powder and two tablets containing a dry extract of Uvae ursi folium. At least 504 patients (allocated as 1:1) will need to be enrolled to access non-inferiority with a non-inferiority limit of 14% for the primary endpoint.Improvement of symptoms of uncomplicated cystitis (based on the ACSS score) at day 7 is defined as the primary endpoint, whereas several secondary endpoints such as the number and ratio of patients with bacteriuria at day 7, frequency and severity of side effects; recurrence of urinary tract infection, concurrent use of other over the counter (OTC) medications and food supplements will be determined to elucidate more detailed differences between the groups. The number of recurrences and medications taken for treatment will be monitored for a follow-up period of 90 days (80-100 days). ETHICS AND DISSEMINATION: This study has been approved by the Scientific and Research Ethics Committee of the Hungarian Medical Research Council (IV/4225-1/2021/EKU). The results will be disseminated by publication of peer-reviewed manuscripts. TRIAL REGISTRATION NUMBER: NCT05055544.

Achillea spp.: A comprehensive review on its ethnobotany, phytochemistry, phytopharmacology and industrial applications.

The genus Achillea genus houses more than 100 species, a number of them are popularly used in traditional medicine for spasmodic gastrointestinal, gynecological and hepatobiliary disorders, hemorrhages, pneumonia, rheumatic pain, inflammation, wounds healing etc. Members of the genus contain a wide variety of volatile and non-volatile secondary metabolites, including terpenes, polyphenols, flavonoids and others. Multiple studies have assessed the biological effects and other aspects of Achillea spp. In a number of preclinical studies, Achillea plants and their essential oils have demonstrated promising antibacterial properties against a number of human and plant pathogens. Besides, the plants have displayed strong antioxidative and potent anti-proliferative and anticancer properties in various cellular and animal models. Achillea plants have widely been used as food preservative in food industry. Clinical studies have indicated its potential against multiple sclerosis (MS), irritable bowel syndrome (IBS), ulcerative colitis, episiotomy wound, primary dysmenorrhea, oral mucositis etc. The present work focuses to provide a brief overview on folk knowledge, phytochemistry, biological activity and applications of Achillea plants. There is a close relationship between the traditional ethnobotanical usage and pharmacological and clinical data from different Achillea spp. The application of Achillea plants and their extracts seems to be a promising alternative for antimicrobial and antioxidant purposes in food, pharmaceutical and cosmetic industries.

Phenanthrenes from Juncus Compressus Jacq. with Promising Antiproliferative and Anti-HSV-2 Activities.

Juncaceae species are rich sources of phenanthrenes. The present study has focused on the isolation and structure determination of biologically active components from Juncus compressus. Eleven compounds (nine phenanthrenes and two flavonoids) have been isolated from the plant by the combination of different chromatographic methods. Two compounds (compressins A (Compound 1) and B (Compound 2)) are novel natural products, while seven phenanthrenes (effusol (Compound 3), effususol (Compound 4), juncusol (Compound 5), 2-hydroxy-1-methyl-4-oxymethylene-5-vinyl-9,10-dihydrophenanthrene (Compound 6), 7-hydroxy-1-methyl-2-methoxy-5-vinyl-9,10-dihydrophenanthrene (Compound 7), effususin A (Compound 8), and dehydroeffusol (Compound 9)), and two flavonoids (apigenin (Compound 10) and luteolin (Compound 11) were isolated for the first time from the plant. Compressin B (Compound 2) is a dimeric phenanthrene, in which two juncusol monomers (Compound 5) are connecting through their C-3 atoms. The structure elucidation of the isolated compounds was carried out using 1D, 2D NMR spectroscopic methods and HR-MS measurements. In vitro investigation of the antiproliferative effect of the phenanthrenes on two cervical (HeLa and SiHa) and an ovarian human tumor cell line (A2780) revealed that compounds have remarkable antiproliferative activity, mainly on the HeLa cell line. Moreover, juncusol (Compound 5) proved to possess significant antiviral activity against the herpes simplex 2 virus (HSV-2).

Antiproliferative and Antimicrobial Activities of Selected Bryophytes.

One-hundred and sixty-eight aqueous and organic extracts of 42 selected bryophyte species were screened in vitro for antiproliferative activity on a panel of human gynecological cancer cell lines containing HeLa (cervix epithelial adenocarcinoma), A2780 (ovarian carcinoma), and T47D (invasive ductal breast carcinoma) cells using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and for antibacterial activity on 11 strains using the disc-diffusion method. A total of 99 extracts derived from 41 species exerted ≥25% inhibition of proliferation of at least one of the cancer cell lines at 10 µg/mL. In the cases of Brachythecium rutabulum, Encalypta streptocarpa, Climacium dendroides, Neckera besseri, Pleurozium schreberi, and Pseudoleskeella nervosa, more than one extract was active in the antiproliferative assay, whereas the highest activity was observed in the case of Paraleucobryum longifolium. From the tested families, Brachytheciaceae and Amblystegiaceae provided the highest number of antiproliferative extracts. Only 19 samples of 15 taxa showed moderate antibacterial activity, including the most active Plagiomnium cuspidatum, being active on 8 tested strains. Methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus aureus were the most susceptible to the assayed species. This is the first report on the bioactivities of these 14 species.

Investigation of Antimicrobial, Antioxidant, and Xanthine Oxidase--Inhibitory Activities of Phellinus (Agaricomycetes) Mushroom Species Native to Central Europe.

Ten representative Central European phellinoid Hymenochaetaceae species (Phellinus sensu lato) were selected to examine their potential pharmacological activity. In this study 40 organic (n-hexane, chloroform, 50% methanol) and aqueous extracts with different polarities were analyzed for their antimicrobial, antioxidant, and xanthine oxidase (XO)--inhibitory properties. Fomitiporia robusta, Fuscoporia torulosa, Phellopilus nigrolimitatus, and Porodaedalea chrysoloma showed moderate antibacterial activity; Bacillus subtilis ATCC 6633, methicillin-resistant Staphylococcus aureus ATCC 43300, and Moraxella catarrhalis ATCC 43617 were the strains most susceptible to the examined fungal species. The in vitro antioxidant and XO assays demonstrated that most of the selected species possess remarkable antioxidant and XO-inhibitory activities. The water extracts in general proved to be more active antioxidants than organic extracts. In the case of F. torulosa, Ph. Nigrolimitatus, and P. chrysoloma, the results of DPPH tests correlate well with those obtained by oxygen radical absorbance capacity tests; these mushrooms presented high antioxidant activities in both assays. Future studies involving phellinoid Hymenochaetaceae species are planned, which may furnish novel results in terms of the species' pharmacological activity and the specific compounds responsible for the observed activity.

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.

Inhibition of COX-2 and NF-κB1 Gene Expression, NO Production, 5-LOX, and COX-1 and COX-2 Enzymes by Extracts and Constituents of Onopordum acanthium.

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 ± 0.5 % and 99.4 ± 0.8 %) in the inhibition of cyclooxygenase-2 (98.6 ± 0.2 % and 97.0 ± 1.1 %) and nuclear factor kappa B1 gene expression (76.7 ± 7.3 % and 69.9 ± 3.4 %). Furthermore, it was shown that these inhibitory effects are not due to cytotoxicity of the compounds.

Bioactivity-guided isolation of antiproliferative compounds from the roots of Onopordum acanthium.

Onopordum acanthium has been considered in traditional medicine to be effective against different cancers. The chloroform extracts of the root, which displayed antiproliferative effect against human tumor cell lines, was subjected to bioactivity-guided multistep chromatographic separation. This experiment resulted in the isolation of the sesquiterpene lactones 4beta,14-dihydro-3-dehydrozaluzanin C (1), zaluzanin C (2) and 4beta,15,11beta,13-tetrahydrozaluzanin C (3), the neolignan nitidanin-diisovalerianate (4), besides 13-oxo-9Z,11 E-octadecadienoic acid (5), 24-methylenecholesterol (6), alpha-linolenic acid, linoleic acid, stigmasterol and beta-sitosterol. The structures of the isolated compounds were established through analytical data (NMR, MS), and by comparison of these with those reported in the literature. All the aforementioned compounds were detected for the first time from this plant. The antiproliferative activities of compounds 1-6 were assessed on cervix adenocarcinoma HeLa, breast adenocarcinoma MCF7 and skin epidermoid carcinoma A431 cells by using the MTT assay. It was found that, 4beta,14-dihydro-3-dehydrozaluzanin C (1), the most active antiproliferative compound of the extract, exerted remarkable tumor cell growth inhibitory activity (IC50 2.7-15.1 microM).

Antifungal activity and composition of essential oils of Conyza canadensis herbs and roots.

Essential oils from herbs and roots of Conyza canadensis (horseweed), collected in Hungary, were obtained by hydrodistillation. The chemical compositions of the oils were analysed by combination of GC and GC/MS. The major constituent of the oil obtained from the aerial parts of horseweed was limonene (78%), while the main component of root oil was 2Z,8Z-matricaria ester. The antimicrobial activities of the oils were tested on Gram-positive (Enterococcus faecalis, Staphylococcus aureus, and Streptococcus pyogenes), Gram-negative (Escherichia coli, Pseudomonas aeruginosa) bacteria, reference fungal strains, and fungal strains isolated from patients (Candida, Cryptococcus, Trichophyton, Rhodotorula, and Aspergillus) by agar disc diffusion and broth dilution methods. None of the oils showed any activity against the tested bacterial strains, but exhibited moderate-to-strong activity against all fungi with the only exception of A. fumigatus. The highest zone of inhibition was observed in case of Cryptococcus neoformans and Trichophyton interdigitalis.

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.

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.