Metabolite profiling, enzyme inhibitory activity and antioxidant potential of different extracts from Glycyrrhiza foetida Desf. (Fabaceae, Galegeae, Glycyrrhizinae).

Obesity, a risk factor for the development of type 2 diabetes, represents a socio-economic burden for healthcare systems. Consequently multiple studies aiming to facilitate early diagnostic and prevent complications have demonstrated the promising role of phenolic derivatives on diabetes management. In this frame, the present study aimed to establish for the first time the chemical ingredients and to appraise the antioxidant and the enzyme inhibitory properties of Glycyrrhiza foetida aerial parts and roots extracts using methanol, ethyl acetate and chloroform solvents. The phytochemicals were analyzed via (HPLC-MS/MS). In addition, the extracts were tested for inhibitory activities against α-amylase, α-glucosidase, acetylcholinesterase, butyrylcholinesterase, and tyrosinase. The antioxidant capabilities were evaluated using various methods (phospho­molybdenum, DPPH, ABTS, FRAP, CUPRAC, and metal chelating assays). Notably, twenty-nine secondary metabolites, namely eleven phenolic acids, seventeen flavonoids and a non-phenolic acid have been identified. Delphinidin 3,5-diglucoside, rutin, isoquercitrin, hyperoside, catechin, phloridzin and hesperidin were the major compounds in the aerial parts extracts while trans-cinnamic acid was the dominant compound in roots. Among phenolic acids, vanillic acid was found to be the main constituent in all extracts. The tested extracts were found to yield TPC in amounts ranging from 34.08 to 49.36 mg GAE/g and TFC ranging from 7.01 to 45.74 mg RE/g. G. foetida extracts showed interesting in vitro antioxidant properties. Additionally, all extracts showed a significant anti-tyrosinase activity, with inhibition values ranging between 52.84 and 80.34 mg KAE/g. Thus, G. foetida remains an exceptional matrix for further investigations and application in cosmetics and food additives.

Chemical composition, antifungal and antibiofilm activities of essential oils from Glycyrrhiza foetida (Desf.) growing in Tunisia.

This study was designated to investigate the chemical composition, the antifungal activity and antibiofilm properties of Glycyrrhiza foetida (Desf.) growing in Tunisia and recognized for its pharmacological and therapeutic effects. The chemical analysis of essential oil samples prepared via hydrodistillation of the aerial parts was performed by gas chromatography-mass spectrometry (GC-MS). Moreover, the antifungal activity of G. foetida essential oil was developed against three dermatophyte strains, two molds and Candida spp. yeasts using the broth microdilution assay. According to the percentages, the main constituents are δ-cadinene (13.9%), (E)-caryophyllene (13.2%) and γ-cadinene (8.3%). The efficiency of the essential oil in inhibiting Candida albicans biofilms formation was also evaluated in terms of inhibitory percentages. The results showed that C. albicans and Microsporum canis were the most sensitive to G. foetida essential oil with a complete inhibition at 0.4 and 0.2 mg ml-1 , respectively. Candida albicans biofilm development was reduced by 80% by the volatile oil at a concentration of 0.8 mg ml-1 . The essential oil of G. foetida has a promising role in the control of fungal agents with medical interest and in inhibition of Candida biofilm development.

Re-establishment of Sileneneglecta Ten. (Caryophyllaceae) with taxonomic notes on some related taxa.

Sileneneglecta has been misunderstood and confused with S.nocturna, although several morphological characters (petal shape, calyx indumentum, hairiness of stamen filaments, seed size, seed-coat surface and shape) allow separation of these species. Moreover, S.mutabilis (which has been considered conspecific with S.neglecta) and S.martinolii (an alleged endemic species to south-western Sardinia) are considered here as taxonomic synonyms of S.nocturna and S.neglecta, respectively. These taxonomic conclusions are strongly supported by multivariate morphometric analyses of 21 characters.

Polyoxygenated germacranes from Daucus carota and their antimalarial transmission blocking activity.

Chemical analysis of the aerial parts obtained from a Tunisian specimen of Daucus carota yielded to the isolation of six undescribed polyoxygenated germacranes and one elemanolide, along with one known metabolite. The stereostructures of the undescribed compounds were determined by extensive spectroscopic analysis including 1D and 2D NMR and HR-ESI-MS analysis. Due to their structural similarity with the Plasmodium transmission-blocking agent daucovirgolide G, the isolated metabolites were evaluated for their inhibitory activity on the development of Plasmodium early sporogonic stages. Three compounds proved to inhibit ookinete formation showing a good transmission blocking efficacy, but the low potency exhibited by these compounds when compared to daucovirgolide G further supports the observation that strict structural requirements do exist for the antimalarial activity of germacranolides.

Chemical Composition and Antimicrobial Activity of Teucrium Capitatum L. Subsp. Lusitanicum (Schreb.) T. Navarro & Rosua Essential Oil.

Chemical composition and antimicrobial activity of Teucrium capitatum L. subsp. lusitanicum essential oil was investigated for the first time in the present study. Qualitative and quantitative analyses of the chemical composition by gas chromatography and mass spectrometry (GC-FID and GC-MS) revealed the presence of 60 compounds representing 97.6% of the whole constituents. The main compounds were germacrene D (47.1%), spathulenol (5.8%), α-selinene (5.3%), germacrene A (2.9%), δ-cadinene (2.8%) and cubenol (2.7%). In vitro, the antimicrobial activity was investigated against five bacterial strains along with the yeast Candida albicans using broth microdilution assay. T. capitatum subsp. lusitanicum essential oil showed significant activity against the gram-positive bacteria Staphylococcus aureus (MIC = MBC = 78 µg mL-1), Bacillus subtilis (MIC = MBC = 156 µg mL-1) and the yeast C. albicans (MIC = MFC = 156 µg mL-1). The great potential of antimicrobial effects is most likely due to the very high percentage of sesquiterpene hydrocarbons particularly to germacrene D, for which the antimicrobial properties have been previously reported.

Chemical diversity and biological activities of essential oils from native populations of Clinopodium menthifolium subsp. ascendens (Jord.) Govaerts.

This study is focused on the analysis of regional variation of the chemical compositions of three Clinopodium menthifolium subsp. ascendens (Jord.) Govaerts Tunisian accession, as well as their inhibition toward fungi and insect pests. The diversity of the chemical constituents and biological activities in front of the aforementioned variations was found to be remarkable. Essential oils were obtained by hydrodistillation of the aerial parts and analyzed by gas chromatography-mass spectrometry (GC-MS). A total of 41, 42, and 30 compounds were identified respectively from Clinopodium menthifolium essential oils harvested from three Tunisian regions, namely Ain-Draham (ADEO), Babouch (BEO), and Tabarka (TEO). All analyzed oils were rich in oxygenated monoterpenes with different major constituents. Piperitenone (34.5%), cis-piperitone oxide (26.1%), and pulegone (47.9%) were the dominant compounds in the three volatile oils, respectively. The antifungal activity was investigated in vitro using six targeted fungal strains (Aspergillus flavus, Aspergillus terreus, Candida albicans, Microsporum canis, Microsporum gypseum, and Trichophyton mentagrophytes). The toxicity and repellency of essential oils were evaluated against the stored product pest Tribolium confusum. The tested samples were differently effective toward the target fungi and the pest depending on the variability of their chemical compositions. BEO exhibited the highest fungitoxic properties toward A. terreus mold, M. canis dermatophyte, and C. albicans yeast (the MIC values ranged from 40 to 400 µg mL-1). In addition, the data showed that TEO repelled T. confusum moderately (PR = 42.5% at 2 h after exposure). Concerning the contact treatment, both ADEO and BEO were proved to possess slightly toxic effects toward T. confusum pest (% of mortality 27.5-32.5% at 5% concentration). The results showed that the geographic origin greatly influenced the chemical composition and the associated bioactivities of Clinopodium menthifolium subsp. ascendens.

Chemical composition, antifungal and antiproliferative activities of essential oils from Thymus numidicus L.

In this study, Thymus numidicus Poir. plant material was collected from two different locations in north-western Tunisia and the aerial parts essential oils (EOs) were extracted via hydro-distillation. Gas chromatography coupled to mass spectrometry (GC-MS) and flame ionisation detection (GC-FID) were used for the qualitative analysis and quantification of the volatile constituents. Thymol (50.1-52.8%) was identified as the main compound of both EOs. To evaluate the potential application of the EOs as antifungal agents, the in vitro inhibitory effects were tested against six fungal strains; a strong antifungal activity of one sample was observed (MIC = 40-400 µg/mL). The in vitro antiproliferative activity was investigated on two human cancer cell lines, i.e. the colonic (HCT116) and breast adenocarcinoma (MCF7) using the colourimetric MTT assay. Again, the same sample demonstrated to possess good antiproliferative activity against both cancer cell lines, with IC50 values of 26.9 and 11.7 µg/mL, respectively.

The effect of pressure on the characteristics of supercritical carbon dioxide extracts from Calamintha nepeta subsp. nepeta.

Chemists and industrialists are continuously attempting to develop greener and more environmentally benign chemical processes to extract essential oils and bioactive metabolites of high purity, finding various applications in cosmetics, detergents, nutraceuticals and pharmaceuticals. An increase preferenced for natural products over synthetic ones has made supercritical fluid technology a primary alternative for the generation of high-value bioactive ingredients. This effective technique requires only moderate temperatures, eliminates clean-up steps and avoids the use of harmful organic solvents. In this context, our study was focused on the chemical analysis of Calamintha nepeta subsp. nepeta aromatic extracts obtained with supercritical carbon dioxide. The effect of different operating conditions on the capacity of the lipophilic solvent to extract the targeted volatile components was also studied. The process was carried out at a fairly low constant temperature of 40°C, and with varying the pressure from 90 to 300 bar. The chemical composition of the extracts was analyzed by gas chromatography-mass spectroscopy. The results showed that the composition pattern, the concentrations of individual components and the quality of the extractable analytes were affected by pressure increase. The extraction yields varied from 0.73 to 1.21 wt% at 90 and 300 bar, respectively.

Differentiation of Nanoparticles Isolated from Distinct Plant Species Naturally Growing in a Heavy Metal Polluted Site.

Leaves harvested from the plants of two different species (Dittrichia viscosa and Cichorium intybus) grown in their autogenous environment near a steel manufacturing company were characterized for naturally accumulated nanoparticles. These plant species are known to accumulate heavy metals. It was, however, unknown if these species would also accumulate these heavy metals in the form of nanoparticles. The isolated solid fractions were analyzed using dynamic light scattering, X-ray fluorescence, and transmission electron microscopy. These analyses revealed the presence of nanoparticles within the plants. The composition of nanoparticles found in each plant species is distinct: (i) for Dittrichia viscosa, the nanoparticle composition matched the heavy metal pollution anticipated from the surrounding industries; (ii) for Cichorium intybus, the nanoparticle composition was similar to the most abundant elements in the soil. The different behavior is a reflection of the phytoaccumulator characteristics of both species. This study provides the first evidence of sequestration of heavy metals in the form of nanoparticles by plants grown autogenously in polluted areas and will have implications in waste management of phytoremediation systems and in understanding the heavy metal life-cycle in the environment.

Chemical Composition and Antioxidant Activity of Essential Oils and Hexane Extract of Onopordum arenarium from Tunisia.

In the present study, volatile oils from Onopordum arenarium fresh flowers and stems were obtained by hydrodistillation and the non-polar aerial part hexane extract was prepared using a Soxhlet apparatus. The constituents of different organs were identified for the first time by gas chromatography equipped with flame ionization detector and gas chromatography coupled to mass spectrometry. A total of 29 and 25 compounds were identified constituting over 91.6 and 89.2% of the whole constituents from flower and stem volatile oils, respectively. Both organs were constituted mainly of long-chain hydrocarbons (23.3-36.4%) followed by oxygenated long-chain hydrocarbons (31.5-33.8%) and oxygenated monoterpenes (14.4-6.6%). The major identified compound was palmitic acid [25.5% in O. arenarium flower essential oil (EO) and 28.7% in the stem EO]. Eighteen compounds representing 80.7% of the whole constituents were identified in the n-hexane extract, which was characterized by high amounts of triterpenoids (39.6%) and dominated by lupeol acetate (19.2%) and ß-amyrin acetate (10.1%). Moreover, all extracts were evaluated for antioxidant potential using 1,1-diphenyl-2-picrylhydrazyl radical assay. The obtained results demonstrated that the EOs and the hexane extract could be a new source of natural potentially bioactive molecules.

Effect of pressure variation on the efficiency of supercritical fluid extraction of wild carrot (Daucus carota subsp. maritimus) extracts.

The present study was conducted to check the comparative qualities of essential oils prepared by hydrodistillation (HD) and supercritical fluid techniques. It constitutes the first attempt to investigate the chemical composition of Daucus carota subsp. maritimus extracts using supercritical fluid technology (SFE) as an environmentally clean innovative method of extraction. The effect of pressure on the nature of extractable substances from wild carrot has been performed at a constant temperature of 50 °C and two different pressures (100 and 300 bar). The experimental results showed that pressure had a significant enhancing effect on the fluid transport properties and therefore on yield values. The extraction yield increases from 1.167 to 2.986% while increasing pressure. The chemical compositions of the essential oils prepared by HD and SFE were analyzed on the basis of gas chromatography-mass spectroscopy (GC-MS). Thus, we noticed that all analyzed samples were enriched in geranyl acetate and ß­bisabolene, and that the quantities of different identified substances were extremely sensitive to the extraction method and to the pressure variation in case of SFE.

Daucovirgolides I-L, four congeners of the antimalarial daucovirgolide G from Daucus virgatus.

Repeated chromatographic purifications of aerial parts of the Tunisian plant Daucus virgatus led to the isolation of four new germacranolides, named daucovirgolides I-L (2-5), along with the Plasmodium transmission-blocking agent daucovirgolide G. The chemical structures of the new compounds were defined as mono- or di-angeloylated germacrane-type sesquiterpenoids by spectroscopic (mainly 1D and 2D NMR) and spectrometric methods (ESIMS). The low potency exhibited by daucovirgolides I-L further supports the observation that strict structural requirements do exist for the Plasmodium transmission blocking activity in the daucovirgolide series. In particular, the endocyclic double bond system seems to be crucial for bioactivity.

Chromatographic analysis, antimicrobial and insecticidal activities of the essential oil of Phlomis floccosa D. Don.

The present study was conducted to investigate for the first time the chemical composition and insecticidal and antimicrobial properties of Phlomis floccosa D. Don. essential oil isolated by hydro-distillation of its aerial parts. Qualitative and quantitative analyses of the essential oil on the basis of gas chromatography and mass spectrometry (GC/FID and GC/MS) revealed the presence of 59 organic volatiles representing 86.9% of the total constituents. The major compounds were germacrene-D (19.7%) followed by ß-caryophyllene (15.5%), caryophyllene oxide (8.3%), hexadecenoic acid (7.9%) and carvacrol (6.1%). The broth microdilution method was conducted to test the antimicrobial activity of the essential oil against five pathogens: Escherichia coli, Pseudomonas aeruginosa, Salmonella enterica, Staphylococcus aureus, Bacillus subtilis and the yeast Candida albicans. The oil of P. floccosa D. Don. shows promising activity against the Gram-positive bacteria S. aureus and B. subtilis and the yeast C. albicans (minimum inhibitory concentration = 625 µg.mL-1 ). The effectiveness of the essential oil was assessed on both Tribolium confusum and Sitophilus zeamais pest insects and it was more pronounced against T. confusum, showing a moderate mortality of 47.5%.

Angeloylated Germacranolides from Daucus virgatus and Their Plasmodium Transmission Blocking Activity.

Phytochemical investigation of the aerial parts of the Tunisian plant Daucus virgatus led to the isolation of eight new germacranolides named daucovirgolides A-H (1-8). The stereostructures of these sesquiterpene lactones, decorated by either one or two angeloyl groups, have been determined by a combination of MS, NMR spectroscopy, chemical derivatization, and comparison of experimental electronic circular dichroism curves with TDDFT-predicted data. Daucovirgolide G (7) proved to be the single member of this family to possess a marked inhibitory activity (92% at 50 µg/mL) on the development of Plasmodium early sporogonic stages, the nonpathogenic transmissible stages of malaria parasites, devoid of general cytotoxicity. The selective activity of daucovirgolide G points to the existence of strict structural requirements for this transmission-blocking activity and therefore of a well-defined, although yet unidentified, biological target.

Antiproliferative metabolites from the Northern African endemic plant Daucus virgatus (Apiaceae).

Chemical analysis of the dichloromethane fraction obtained from aerial parts of the Northern African endemic plant Daucus virgatus led to the isolation of three previously undescribed sesquiterpenoids, namely the daucane vaginatin B, a eudesmane and the elemane elemavirgolide, along with five known metabolites. The structures of these compounds were determined by a detailed MS and NMR analysis and they were evaluated for antiproliferative activity against three human cell lines, A375 (melanoma), MCF-7 (breast adenocarcinoma), and HACAT (keratinocyte). The phytoalexin 6-methoxymellein revealed a previously unreported antiproliferative activity, while the eudesmane and the elemane derivatives exhibited a selective activity (SI = 11.1 and 3.3, respectively) against melanoma tumor cell lines.

Chemical Composition, Antioxidant and Cytotoxic Activities of Essential Oil of the Inflorescence of Anacamptis coriophora subsp. fragrans (Orchidaceae) from Tunisia.

The chemical composition of the essential oil produced by steam distillation of the inflorescences of naturally growing Anacamptis coriophora (L.) R. M. Bateman, Pridgeon & M. W. Chase subsp.fragrans (Pollini) R. M. Bateman, Pridgeon & M. W. Chase (Orchidaceae) from Kroumiria, north-west Tunisia was studied by GC-MS, which led to the identification of 19 volatile components, representing 97% of the oil. The main constituents were methyl-(E)-p-methoxycinnamate (29.3%), 13-heptadecyn-1-ol (18.6%), 2,5-dimethoxybenzyl alcohol (14.1%) and 4-(1,1,3,3-tetramethylbutyl)-phenol (9.0%). DPPH radical scavenging revealed a weak antioxidant activity. In addition, the antiproliferative effects were evaluated on BxPC3 human pancreatic carcinoma cells and on 2008 human ovarian cancer cells showing significant effect. This is the first report of the chemical composition of essential oils obtained from A. coriophora subsp. fragrans inflorescences for North Africa. Further studies are needed to understand fully the possible mechanism of action behind the cytotoxic activity of the essential oil.

Essential Oil Constituents and Antioxidant Activity of Asplenium Ferns.

Two fern species Asplenium adiantum-nigrum L. and Asplenium trichomanes L. collected from the Kroumiria region (Northwest of Tunisia) were individually submitted to hydrodistillation in a Clevenger type apparatus. Volatile organic compounds were identified by GC-MS and GC-FID. Thus, 35 compounds were identified in A. adiantum-nigrum essential oil accounting for 77.5% of the whole constituents dominated by palmitic acid (34.5%); however, only 29 volatiles were identified in A. trichomanes showing a high amount of phytol, an odorous diterpene alcohol, representing 14.4% of the total oil contents. The total phenolic content and the antioxidant effects of crude extracts from both pteridophytes were determined using Folin-Ciocalteu and 2,2'-diphenyl-1-picrylhydrazyl free radical-scavenging assays, respectively. A. adiantum-nigrum ethyl acetate extract is shown to be lower in total phenolic contents (49.3 mg gallic acid equivalent/g) than similar extract from A. trichomanes (55.4 mg GAE/g).

New bioactive labdane diterpenoids from Marrubium aschersonii.

A phytochemical investigation of the ethanol extract of Marrubium aschersonii Magnus (Lamiaceae) collected from Tunisia led to the isolation and identification of two new labdane diterpenoids, marrubaschs A (1) and B (2), along with two known compounds (3 and 4). Their structures were elucidated by spectroscopic methods including HRESIMS and NMR techniques. All compounds were evaluated for their inhibitory effects on the nitric oxide (NO) production induced by lipopolysaccharide in RAW 264.7 macrophage cells. Compound 2 exhibited weak inhibition of NO production with an IC50 value of 35 ± 1.0 µM.

Volatile Profile of Herniaria fontanesii Growing Spontaneously in Tunisia.

The essential oil extracted from Desfontaine's rupturewort, Herniaria fontanesii J. Gay subsp. fontanesii growing wildly in Tunisia, was analyzed using GC and GC-MS techniques. The free radical scavenging capacity and total phenol contents of three crude extracts having different polarities (n-hexane, ethyl acetate and methanol) were examined. Thus, a total of 35 constituents were identified in the Desfontaine's rupturewort essential oil representing 89.8% of the whole constituents. The oil was dominated by hexadecanoic acid, caryophyllene oxide, terpin-4-ol, khusimone and trans-sabinene hydrate. The total phenolic contents ranged from 16.91 to 92.27 mg of gallic acid/g of dry weight and they were found to be significantly higher in methanol than in polar ethyl acetate and hexane extracts. Correlations were observed between the phenolic contents and the antioxidant properties. Thus, the antioxidant activity of the methanol extract was superior to that of all samples tested (IC50 = 0.21 ± 0.04 mg/mL).

Essential Oil Composition, Antioxidant, Cytotoxic and Antiviral Activities of Teucrium pseudochamaepitys Growing Spontaneously in Tunisia.

The chemical composition, antioxidant, cytotoxic and antiviral activities of the essential oil obtained by hydrodistillation from the aerial parts of Teucrium pseudochamaepitys (Lamiaceae) collected from Zaghouan province of Tunisia are reported. The essential oil was analyzed by gas chromatography equipped with a flame ionization detector (GC-FID) and gas chromatography coupled with mass spectrometry (GC/MS). Thirty-one compounds were identified representing 88.6% of the total essential oil. Hexadecanoic acid was found to be the most abundant component (26.1%) followed by caryophyllene oxide (6.3%), myristicin (4.9%) and α-cubebene (3.9%). The antioxidant capacity of the oil was measured on the basis of the scavenging activity to the stable 2,2-diphenyl-1-picrylhydrazyl (DPPH). The IC50 value of the oil was evaluated as 0.77 mg·mL(-1). In addition, the essential oil was found to possess moderate cytotoxic effects on the HEp-2 cell line (50% cytotoxic concentration (CC50)=653.6 µg·mL(-1)). The potential antiviral effect was tested against Coxsackievirus B (CV-B), a significant human and mouse pathogen that causes pediatric central nervous system disease, commonly with acute syndromes. The reduction of viral infectivity by the essential oil was measured using a cytopathic (CPE) reduction assay.