Investigation of alternative two-phase solvent systems for purification of natural products by centrifugal partition chromatography.

INTRODUCTION: Centrifugal partition chromatography (CPC) is a liquid-liquid chromatography characterised by its solvent flexibility. The compounds undergoing separation are subjected to a continuous partition process between two immiscible phases in a column space free of solid support. In the context of green chemistry, it is important to substitute halogenated and petroleum-based solvents commonly used in purification processes. OBJECTIVES: The main goal of the current study was to replace classical solvents used in CPC (e.g., hexane and methanol) by green and renewable alternatives. METHODS: Solvents were first selected based on literature. Their commercial availability, price, recyclability, toxicity and ability to form two phases were particularly sought after. KEY FINDINGS: The new two-phase solvent systems were evaluated for the purification of two compounds of interest: piperine and cannabidiol. Using these alternative two-phase solvent systems allows us to isolate natural products with a high purity level (> 95%). CONCLUSION: Substituting petroleum-based solvents with bio-sourced, renewable alternatives reduces the environmental impact of CPC. Herein, new biphasic solvent systems were built using hexamethyldisiloxane, ethyl isobutyrate and 2-methyl tetrahydrofuran in combination with ethanol and water. Furthermore, this research provides a scientific basis for developing new and sustainable solvent systems in CPC.

Phytochemical study of Boswellia dalzielii oleo-gum resin and evaluation of its biological properties.

Boswellia dalzielii is a resin-producing tree endemic to West and Central Africa, used by local populations for various medicinal purposes. In this study, B. dalzielii gum resin was analyzed by GC-MS and UHPLC-MS to identify and quantify volatile and non-volatile compounds. Its main volatile constituents were α-pinene (54.9%), followed by α-thujene (4.4%) and α-phellandren-8-ol (4.0%). Pentacyclic triterpenoids such as ß-boswellic acids and their derivatives were quantified by UHPLC-MS and their content was shown to reach around 22% of the gum resin. Since some of the volatile and non-volatile compounds identified in this work are known to possess biological effects, the bioactivities of B. dalzielii ethanolic extract, essential oil, as well as fractions of the oil and extract were evaluated. Some of these samples exhibited interesting anti-inflammatory properties, and their antioxidant, anti-ageing and skin-bleaching activities were also tested.

Commiphora wildii Merxm. Essential Oil: Natural Heptane Source and Co-Product Valorization.

As an alternative to fossil volatile hydrocarbon solvents used nowadays in perfumery, investigation on essential oil of Commiphora wildii Merxm. oleo gum resin as a source of heptane is reported here. Heptane, representing up to 30 wt-% of this oleo gum resin, was successfully isolated from the C. wildii essential oil, using an innovative double distillation process. Isolated heptane was then used as a solvent in order to extract some noble plants of perfumery. It was found that extracts obtained with this solvent were more promising in terms of sensory analysis than those obtained from fossil-based heptane. In addition, in order to valorize the essential oil depleted from heptane, chemical composition of this oil was found to obtain, and potential biological activity properties were studied. A total of 172 different compounds were identified by GC-MS in the remaining oil. In vitro tests-including hyaluronidase, tyrosinase, antioxidant, elastase and lipoxygenase, as well as inhibitory tests against two yeasts and 21 bacterial strains commonly found on the skin-were carried out. Overall, bioassays results suggest this heptane-depleted essential oil is a promising active ingredient for cosmetic applications.

Anti-breast cancer potential of Anonidium mannii (Oliv.) Engl. & Diels barks ethanolic extract: UPLC-ESI-QTOF-MS detection of anticancer alkaloids.

ETHNOPHARMACOLOGICAL RELEVANCE: Breast cancer is a serious threat in low-income as well as developed countries. To face this, many herbal preparations are prescribed by traditional healers in Cameroon, among which is Anonidium mannii commonly called "wild soursop". AIM: This study was undertaken to assess the anti-tumor effect of A. mannii ethanolic extract on cancer cell growth and against DMBA-induced mammary tumors in rats. MATERIALS AND METHODS: The well characterized MTT bioassay was used to assess the cytotoxic potential of A. mannii ethanolic extract in liver (HepG2), prostate (DU145 & PC3) and breast (MCF-7) cancer cell lines. Considering the fact that breast cells were the most sensitive to the extract, a 7,12-dimethylbenz(a)anthracene (DMBA)-induced breast tumor rat model was used to assess the possible anticancer effect of A. mannii extract. Indeed, rats were treated with either tamoxifen (3.3 mg/kg BW) or A. mannii extract (16.5, 50 and 150 mg/kg BW) or vehicle (2% ethanol) for 20 weeks. Tumor incidence, tumor mass and volume, oxidative stress status in tumor as well as tumor histoarchitecture were evaluated. RESULTS: A 24 h incubation of tested cells with the A. mannii extract significantly slowed cell growth in a concentration-dependent manner with an interesting effect in breast cells (IC50 ~61.5 µg/mL). As compared to the DMBA rats, those treated with A. mannii extract (50 and 150 mg/kg) showed reduced breast tumor incidence (28%), tumor burden (95.34% at 50 mg/kg and 99.14% at 150 mg/kg) and tumor volume (~92%). A. mannii extract counteracted the high proliferation of terminal mammary ducts induced by DMBA, mainly at 50 mg/kg. Furthermore, the extract decreased MDA and nitrite levels but increased SOD activity in the mammary gland. High Performance Liquid Chromatography coupled with Mass Spectrometry (HPLC-MS) analysis detected potential anticancer and antioxidant alkaloids in A. manni extract, which are close to those found in Annona muricata. CONCLUSION: These results provide evidence on the in vitro and in vivo anticancer effects of A. mannii, and therefore support its use in traditional medicine system to fight against cancer.

New Isoflavones with Antioxidant Activity Isolated from Cornulaca monacantha.

In the course of phytochemical and chemotaxonomical investigations of Cornulaca monacantha (Amaranthaceae), two new isoflavones, 3-(2-hydroxyphenyl)-5,7-dimethoxy-6-(methoxymethyl)-4H-1-benzopyran-4-one (1) and 7-hydroxy-3-(4-hydroxyphenyl)-5-methoxy-6-(methoxymethyl)-4H-1-benzopyran-4-one (2) were isolated from the fresh aerial parts of C. monacantha among with three known compounds named vanillic acid (3), N-cis-feruloyltyramine (4) and N-trans-feruloyltyramine (5). Their structures were elucidated by means of spectroscopic methods including one- and two-dimensional NMR and HR-ESI-MS techniques. The isolated compounds exhibited interesting antioxidant activity determined by DPPH, ABTS and TAC tests.

Metabolomic study of volatile compounds emitted by lavender grown under open-field conditions: a potential approach to investigate the yellow decline disease.

INTRODUCTION: Fine lavender and lavandin are perfume and medicinal plants originate from the South of France and are widely cultivated for their essential oils. Recently, cultivated plants suffered from a severe decline in France, due to the propagation of the yellow decline disease. This disease is caused by the stolbur phytoplasma, a bacterium transmitted by a sap-sucking insect, the planthopper. OBJECTIVES: In order to understand the complex relationships between host plant, pest, pathogen and environment responsible for the yellow decline of lavender, we use a metabolomic approach to highlight changes in chemical emissions from asymptomatic ("healthy") and symptomatic ("infected") plants. METHODS: Volatile compounds produced by fine lavender and lavandin were collected in the field using a dynamic headspace extraction approach. Afterwards, compounds trapped on Tenax adsorbent were thermodesorbed and analysed using an automated thermal desorption-gas chromatography-mass spectrometry (ATD-GC-MS). Multivariate statistical analyses was performed using principal component analysis and partial least square discriminant analyses. RESULTS: The untargeted screening of volatiles allowed the separation of asymptomatic and symptomatic plants according to their emissions. The approach was sufficiently accurate so as to separate the emissions according to the different stages of infection. Twelve compounds were found to be deregulated metabolites of yellow disease infection, common to fine lavender (variety 7713) and lavandin (variety abrial). CONCLUSION: The metabolomic approach allowed for the effective identification of chemical variations between infected and healthy plants in a complex field environment.

Bioguided identification of daucosterol, a compound that contributes to the cytotoxicity effects of Crateva adansonii DC (capparaceae) to prostate cancer cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Crateva adansonii DC (Capparaceae) is a shrub used to treat tumors in Cameroon. In our previous reports, a Crateva adansonii dichloromethane-methanol (DCM/MeOH) extract was shown to prevent chemically induced tumors in Wistar rats. AIM OF STUDY: To determine the bioactive principle of Crateva adansonii extract and to elucidate its underlying mechanism. MATERIALS AND METHODS: An activity-guided fractionation was realized using MTT assay. To investigate if the bioactive compound daucosterol (CA2) accounted for the previously observed anticancer effects of the C. adansonii extract, it was tested on cell growth, cell proliferation, cell cycle, cell death mechanism and cell migration. In addition, cell cycle- and apoptosis-regulating proteins were assessed by Western blotting. RESULTS: Daucosterol (CA2), a steroid saponin, was identified as major anticancer principle of the C. adansonii extract. Daucosterol significantly inhibited LNCaP, DU145 and PC3 prostate carcinoma cell growth and proliferation at the optimal concentration of 1 µg/mL. It also significantly increased the number of late apoptotic (DU145) and apoptotic (PC3) cells. The number of cells in S phase increased in DU145, while the number of G0/G1 cells decreased. Cell cycle proteins (cdk1, pcdk1, cyclin A and B) were down-regulated in DU145 and PC3 cells, whereas only cdk2 was down-regulated in PC3 cells. Moreover, the anti-apoptotic Akt, pAKT and Bcl-2 proteins were down-regulated, while the pro-apoptotic protein Bax was up-regulated. CA2 induced anti-metastatic effects by decreasing chemotaxis and cell migration, while it increased cell adhesion to fibronectin and collagen matrix. CONCLUSION: These results suggest that daucosterol is the major active principle responsible at least in part for the anticancer effect of the extract of Crateva adansonii.

Development of a Headspace Solid-Phase Microextraction Gas Chromatography-Mass Spectrometry Method to Study Volatile Organic Compounds (VOCs) Emitted by Lavender Roots.

A headspace solid-phase microextraction (HS-SPME) method combined with gas chromatography-mass spectrometry (GC/MS) was developed and optimized for the extraction and the analysis of volatile organic compounds (VOCs) from lavandin and fine lavender roots. Optimal parameters to extract volatile molecules from ground and intact roots were determined using a divinylbenzene-carboxen-polydimethylsiloxane (DVB/CAR/PDMS) coating fiber at 70 °C for 60 min. A total of 99 VOCs, including 40 monoterpenoids, 15 sesquiterpenoids, 1 diterpenoid and 2 coumarins were detected. The main compounds detected in lavandin roots were fenchol, borneol, and coumarin. Performances of the optimized SPME GC/MS method were evaluated via the comparison of VOC emissions between roots from different cultivars of fine lavender (7713 and maillette) and lavandin (abrial and grosso). Chemometric analysis, using partial least squares-discriminant analysis (PLS-DA), suggests fifteen significant features as potential discriminatory compounds. Among them, ß-phellandrene allows discrimination between lavender and lavandin varieties.

Lupeol, the major compound of the dichloromethane extract of Millettia macrophylla Benth (Fabaceae), displays estrogenic effects in ovariectomized rats.

The present work deals with the assessment of the in vitro and in vivo estrogenic effects of the triterpenoids (lupenone, lupeol, and stigmastenone) isolated from Millettia macrophylla extract. The in vitro estrogenicity was performed by a reporter gene assay and estrogen receptor-α (ERα) target gene expression, whereas the in vivo estrogenicity was evaluated by a 3-day uterotrophic assay in ovariectomized rats. As results, lupenone and lupeol did not transactivate ERα as well as ERß of human embryonic kidney 293T (HEK293T) cells. However, lupeol seems to be antagonistic to estrogen (E2) only in HEK293T-ERα (10-9 and 10-8  µM). Furthermore, lupeol slightly upregulated GREB1 gene expression at the concentration of 1 µM, suggesting a weak activation of endogenous ERα. In vivo, only lupeol at a dose of 1 mg/kg significantly increased the uterine wet weight (p < 0.05), uterine (p < 0.05), and vaginal (p < 0.01) epithelial heights. The concomitant administration of lupeol (1 mg/kg) with a pure antiestrogen fulvestrant abrogated its effects only in the vagina, whereas in combination with E2, lupeol exhibited a significant antiestrogen-like effect in uterine wet weight and synergistic effects on endometrium. Lupeol has estrogenic effects that is partly through ERs transcriptional activity and does involve alternative mechanisms that are still to be uncovered.

Purification of two valepotriates from Centranthus ruber by centrifugal partition chromatography: From analytical to preparative scale.

Considering chemical complexity of plant crude extracts, purification of natural products is a rate limiting process to identify new compounds as well as to obtain standard references for quantitative or qualitative purposes. In the present work, a centrifugal partition chromatography (CPC) method was developed to isolate and produce high quality reference standards of valtrate and 7-homovaltrate from Centranthus ruber L. roots. These two compounds are controversial aglycon iridioids regulated by the legislation on plant-based dietary supplements. A new biphasic solvent system suitable for CPC separation of valepotriates was developed. It was composed of methanol/hexane/water (5/5/0.8, v/v/v). It yielded a partition coefficient near 1 and a theoretical selectivity of 1.3 between both targeted compounds. Optimization of CPC experimental parameters at the analytical scale (50 mL- and 100 mL-column capacity) enabled compounds' separation with a flow rate of 8 mL/min at 2500 rpm. Then a scale up from a 100 mL-column capacity to a 1000 mL-column capacity has been studied using the "free-space between peaks" concept. It allowed an injected quantity 16 times higher in comparison to the maximal loading capacity of the 100 mL-column. Both valtrate and 7-homovaltrate were recovered in one single step with a purity over 97%. Further MS and NMR characterization allowed to confirm unambiguously the compounds' structures. The highly efficient CPC separation developed in this work provides valepotriates in amounts suitable for further study and strong bases for future industrial development.

The hydro-ethanolic extract of Acacia seyal (Mimosaceae) stem barks induced death in an ER-negative breast cancer cell line by the intrinsic pathway of apoptosis and inhibited cell migration.

BACKGROUND: Despite the significant developments occurring in the treatment of cancer, it still remains the second deadly disease, responsible for 8.2 million deaths every year. Various natural substances have been studied for active molecules of tumor suppression in the past and the tropical flora, by its diversity, continues to provide new antitumor drugs. Acacia seyal is a plant used in Cameroonian traditional system to treat cancer. It exhibited cytotoxic effects towards human breast adenocarcinoma cells. The present work was therefore designed to elucidate the underlying mechanisms by which A. seyal extract induced its cytotoxic effect. METHODS: The cell death mechanism (apoptosis or necrosis) and cell cycle analyses were assessed using flow cytometry. The levels of reactive oxygen species (ROS), mitochondrial membrane potential (ΔΨm), caspases activities as well as Bcl-2 and Bcl-xL protein contents were assessed in MDA-MB-231 cells. Afterwards, cell migration/invasion was also assessed. RESULTS: The A. seyal extract induced apoptosis in MDA-MB-231 cells, while it failed to do so in MCF-7 cells. It induced cell cycle arrest in G2/M phase. Further it induced a decrease in ΔΨm, an increase in ROS levels and caspases activities as well as a down regulation in Bcl-2 and Bcl-xL protein contents in MDA-MB-231 cells. Moreover, A. seyal extract exhibited anti-migration, anti-invasion activities in MDA-MB-231 cells. CONCLUSION: These results demonstrate that A. seyal extract induced its antitumor effects mainly by interference in metastasis related events, by triggering apoptosis through a ROS-mediated mitochondrial pathway.

UHPLC-HRMS/MS Based Profiling of Algerian Lichens and Their Antimicrobial Activities.

Lichens are complex symbiotic organisms able to produce a vast array of compounds. The Algerian lichen diversity has only prompted little interest even given the 1085 species listed. Herein, the chemodiversity of four Algerian lichens including Cladonia rangiformis, Ramalina farinaceae, R. fastigiata, and Roccella phycopsis was investigated. A dereplication strategy, using ultra high performance liquid chromatography-high resolution-electrospray ionization-mass spectrometry (UHPLC-HRMS/MS), was carried out for a comprehensive characterization of their substances including phenolics, depsides, depsidones, depsones, dibenzofurans, and aliphatic acids. Some known compounds were identified for the first time in some species. Additionally, the lichenic extracts were evaluated for their antifungal and antimicrobial activities on human pathogenic strains (Candida albicans, C. glabrata, Aspergillus fumigatus, Staphylococcus aureus, and Escherichia coli). Cyclohexane extracts were found particularly active against human pathogenic fungi with MIC80 values ranging from 8 to 62.5 µg/mL, without cytotoxicity. This study highlights the therapeutic and prophylactic potential of lichenic extracts as antibacterial and antifungal agents.

Cosmetic potentials of Prunus domestica L. leaves.

BACKGROUND: The current study presents new insight on the phytochemical content and biological activities of five Prunus domestica L. varieties ('Quetsche blanche de Létricourt', 'Mirabelle de Nancy', 'Perdrigon violet', 'Mirabelle de Provence', 'Reine-claude dorée'). RESULTS: The plum leaves were found to possess promising anti-aging activities by their capacity to inhibit 1,1-diphenyl-2-picrylhydrazyl (DPPH), elastase, hyaluronidase and lipoxygenase. After solid phase extraction (SPE), chlorogenic acid, rutin, quercetin and their derivatives were putatively identified by high-performance liquid chromatography coupled with tandem mass spectrometry using an electrospray ionization source (HPLC/ESI-MS/MS). CONCLUSION: The plum leaf, a by-product, provides an interesting valuable resource for use as a natural cosmetic product or as a food supplement. © 2017 Society of Chemical Industry.

Estrogen-like and tissue-selective effects of 7-methoxycoumarin from Ficus umbellata (Moraceae): an in vitro and in vivo study.

BACKGROUND: Ficus umbellata is a medicinal plant previously shown to endow estrogenic properties. Its major component was isolated and characterized as 7-methoxycoumarin (MC). Noteworthy, coumarins and the respective active metabolite 7-hydroxycoumarin analogs have shown aromatase inhibitory activity, which is of particular interest in the treatment of estrogen-dependent cancers. The present work aimed at evaluating the estrogenic/antiestrogenic effects of MC in vitro and in vivo. METHODS: To do so, in vitro assays using E-screen and reporter gene were done. In vivo, a 3-day uterotrophic assay followed by a postmenopausal-like rat model to characterize MC as well as F. umbellata aqueous extract in ovariectomized Wistar rats was performed. The investigations focused on histological (vaginal and uterine epithelial height) and morphological (uterine wet weight, vagina stratification and cornification) endpoints, bone mass, biochemical parameters and lipid profile. RESULTS: MC induced a significant (p < 0.05) MCF-7 cell proliferation at a concentration of 0.1 µM, but did not inhibit the effect induced by estradiol in both E-screen and reporter gene assays. In vivo, MC treatment did not show an uterotrophic effect in both rat models used. However, MC (1 mg/kg) induced a significant increase (p < 0.01) of vaginal epithelial height. No significant change was observed with MC in abdominal fat weight, serum lipid levels and bone weight. CONCLUSION: These results suggest that MC has a weak estrogenic activity in vitro and in vivo that accounts only in part to the estrogenicity of the whole plant extract. MC could be beneficial with regard to vagina dryness as it showed a tissue specific effect without exposing the uterus to a potential tumorigenic growth.

Ficus umbellata Vahl. (Moraceae) Stem Bark Extracts Exert Antitumor Activities In Vitro and In Vivo.

A Ficus umbellata is used to treat cancer. The present work was therefore designed to assess antitumor potentials of F. umbellata extracts in nine different cell lines. Cell cycle, apoptosis, cell migration/invasion, levels of reactive oxygen species (ROS), mitochondrial membrane potential (MMP), caspases activities as well as Bcl-2 and Bcl-xL protein content were assessed in MDA-MB-231 cells. The 7,12-dimethylbenz(a)anthracene (DMBA)-induced carcinogenesis in rats were also used to investigate antitumor potential of F. umbellata extracts. The F. umbellata methanol extract exhibited a CC50 of 180 µg/mL in MDA-MB-231 cells after 24 h. It induced apoptosis in MCF-7 and MDA-MB-231 cells, while it did not alter their cell cycle phases. Further, it induced a decrease in MMP, an increase in ROS levels and caspases activities as well as a downregulation in Bcl-2 and Bcl-xL protein contents in MDA-MB-231 cells. In vivo, F. umbellata aqueous (200 mg/kg) and methanol (50 mg/kg) extracts significantly (p < 0.001) reduced ovarian tumor incidence (10%), total tumor burden (58% and 46%, respectively), average tumor weight (57.8% and 45.6%, respectively) as compared to DMBA control group. These results suggest antitumor potential of F. umbellata constituents possibly due to apoptosis induction mediated through ROS-dependent mitochondrial pathway.

Ethanol-extracted Cameroonian propolis exerts estrogenic effects and alleviates hot flushes in ovariectomized Wistar rats.

BACKGROUND: Since the biological properties of propolis depend to the plants that can be found in a specific region, propolis from unexplored regions attracts the attention of scientists. Ethanolic extract of Cameroonian propolis (EEP) is used to treat various ailments including gynecological problems and amenorrhea. Since there were no scientific data to support the above claims, the present study was therefore undertaken to assess estrogenic properties of Cameroonian propolis. METHODS: To achieve our goal, the ability of EEP to induce MCF-7 cells proliferation in E-screen assay as well as to activate estrogen receptors α (ERα) and ß (ERß) in cell-based reporter gene assays using human embryonic kidney cells (HEK293T) transfected with ERs was tested. Further, a 3-day uterotrophic assay was performed and the ability of EEP to alleviate hot flushes in ovariectomized adult rats was evaluated. RESULTS: In vitro, EEP showed an antiestrogenic activity in both HEK293T ER-α and ER-ß cells. In vivo, EEP induced a significant increase in a bell shape dose response manner of the uterine wet weight, the total protein levels in the uterus, the uterine and vaginal epithelium height and acini border cells of mammary gland with the presence of abundant eosinophil secretions. Moreover, EEP induced a significant decrease in the total number, average duration as well as frequency of hot flushes after 3 days of treatment in rat (equivalent to a month in woman). The dose of 150 mg/kg exhibited the most potent estrogenic effects among all the tested doses. The UPLC-HRMS analysis showed the presence of caffeic acid derivatives and trirtepernoids in EEP, which are well known endowed with estrogenic properties. CONCLUSION: These results suggest that Ethanolic extract of Cameroonian propolis has estrogen-like effects in vivo and may alleviate some menopausal problems such as vaginal dryness and hot flushes. Ethanol-extracted Cameroobian propolis exhibited in vitro and in vivo estrogen-like effects. This extract may contain promising phytoestrogens.

Chemical Composition, Antimicrobial and Insecticidal Activities of Flowers Essential Oils of Alpinia zerumbet (Pers.) B.L.Burtt & R.M.Sm. from Martinique Island.

Alpinia species, used as ornamental plants and flavoring agents, are renowned for their therapeutic properties and their subsequent use in traditional medicines throughout the world. Alpinia zerumbet (Pers.) B.L.Burtt & R.M.Sm. is the most common Alpinia species encountered in Martinique. Several essential oils (EOs) obtained by hydrodistillation of A. zerumbet flowers collected in various locations on the island at different seasons were analysed to evaluate the influence of the collection period and area on the EOs' chemical compositions and to assess their bioactivity. A combination of GC-FID and GC/MS techniques was used to examine the volatile constituents, leading to the identification of a total of 71 components accounting for 97.8 - 99.3% of the respective total GC-FID areas: among them, nineteen compounds were characterized for the first time in A. zerumbet EOs. The antimicrobial activity of these EOs was assessed against eight bacterial, yeast and fungal strains and two multi-resistant strains: some significant bacteriostatic and fungistatic activities of A. zerumbet flower oils were evidenced. Finally, an interesting insecticidal activity of the flower EO was highlighted for the first time against Aedes aegypti.

Crateva adansonii DC, an African ethnomedicinal plant, exerts cytotoxicity in vitro and prevents experimental mammary tumorigenesis in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: Crateva adansonii DC is a plant traditionally used in Cameroon to treat constipation, asthma, snakebites, postmenopausal complaints and cancers. AIM: The anticancer potential of the dichloromethane/methanol extract of C. adansonii stem barks was investigated using human breast cancer cell and 7,12 dimethylbenz(a)anththracene (DMBA)-induced mammary tumorigenesis model in rats. MATERIAL AND METHODS: The cytotoxicity of C. adansonii extract was assessed in vitro towards breast carcinoma (MCF-7 and MDA-MB-231) and non-tumoral cell lines (NIH/3T3 and HUVEC) by Alamar Blue assay. Furthermore, in vivo studies were performed on female Wistar rats treated either with C. adansonii extract at a dose of 75 or 300mg/kg body weight or with tamoxifen (3.3mg/kg body weight), starting 1 week prior DMBA treatment and lasted 12 weeks. The investigation focused on tumour burden, tumour DNA fingerprint, morphological, histological, hematological, and biochemical parameters. RESULTS: CC50 values for the in vitro assays were 289µg/mL against MCF-7 cells and >500µg/mL in others cells, leading to a selectivity index ≥1.73. C. adansonii extract significantly (p<0.001) revealed in vivo the reduction of the cumulative tumour yield (87.23%), total tumour burden (88.64%), average tumour weight (71.11%) and tumour volume (78.07%) at the dose of 75mg/kg as compared to DMBA control group. A weak effect was also observed at 300mg/kg. This extract showed a moderate hyperplasia at the dose of 75mg/kg while at 300mg/kg no significant change was noted as compared to DMBA group. It protected rats from the DNA alteration induced by DMBA and increased antioxydant enzymes activities in mammary gland tissue homogenates. In addition, Ultra-High Performance Liquid Chromatography/ESI-QTOF-Mass Spectrometry analysis of C. adansonii extract detected structure-related of many well-known anticancer agents such as flavane gallate, flavonol, phenylpropanoïds, sesquiterpene derivatives, gallotannins and lignans. The LD50 of C. adansonii was estimated to be greater than 5000mg/kg. CONCLUSIONS: These aforementioned results suggest that the C. adansonii extract may possess antitumor constituents, which could combat breast cancer and prevent chemically-induced breast cancer in rats.

Estrogenic effects of Ficus umbellata Vahl. (Moraceae) extracts and their ability to alleviate some menopausal symptoms induced by ovariectomy in Wistar rats.

ETHNOPHARMACOLOGICAL RELEVANCE: Ficus umbellata Vahl. (Moraceae) is a medicinal plant used in Cameroon to treat amenorrhea as well as other physiological disorders related to menopause. AIM OF STUDY: In order to justify scientifically its traditional use, the estrogen-like properties of the aqueous (AE) and methanol (MeOH) extracts of F. umbellata were investigated. MATERIAL AND METHODS: In vitro, the ability of different extracts of F. umbellata to activate estrogen receptors α (ERα) and ß (ERß) in cell-based reporter gene assays using human embryonic kidney (HEK293T) cells transfected with ERs was tested. In vivo, a 3-day uterotrophic assay and the capacity of the extracts to alleviate hot flushes in ovariectomized adult rats were tested. Using a bioassay-guided fractionation the major compound of F. umbellata was isolated and tested in vitro on HEK293T-ERα and ERß cells. RESULTS: AE and MeOH extracts significantly altered ERα as well as ERß activities. In vivo, both extracts significantly increase the uterine and vaginal epithelium thickness, and uterine total protein levels in a dose dependent manner. Interestingly, both extracts of F. umbellata at the dose of 100 mg/kg BW significantly decreased the total number, average duration as well as frequency of hot flushes in experimental rats compared to age-matched OVX controls. Finally, 7-methylumbelliferone, a coumarin was characterized as the major compound of F. umbellata; however this compound did not transactivate ERα as well ERß in vitro. CONCLUSION: These aforementioned results suggest that F. umbellata extracts as used by the traditional practitioner have estrogen-like effects and may alleviate some menopausal problems such as vaginal dryness and hot flushes.