Four pentacyclic triterpenoids with antifungal and antibacterial activity from Curtisia dentata (Burm.f) C.A. Sm. leaves

ETHNOPHARMACOLOGICAL RELEVANCE: Curtisia dentata is used in southern Africa to treat various diseases of bacterial and fungal origin in humans and animals to such a degree that the species is vulnerable and declining [Dold, A.R., Cocks, M.L., 2001. Traditional veterinary medicine in the Alice district of the Eastern Cape Province, South Africa. South African Journal of Science 97, 375-379]. Preliminary studies indicated good activity against Candida albicans. AIM OF THE STUDY: The phytochemistry and antimicrobial activity of this plant species has not been extensively investigated, and a closer examination of the compounds responsible for antimicrobial activity was warranted. MATERIALS AND METHODS: Broth microdilution assay and bioautography were used to evaluate antibacterial and antifungal activity in Curtisia dentata leaf extracts and fractions. Bioassay-directed fractionation using column chromatography yielded four compounds characterised by spectroscopic methods. RESULTS: Lupeol (1), betulinic acid (2), ursolic acid (3) and 2alpha-hydroxyursolic acid (4) were isolated from Curtisia dentata leaves. Betulinic acid, ursolic acid and 2alpha-hydroxyursolic acid appreciably inhibited fungal growth with MIC values ranging from 8 to 63mug/mL. CONCLUSIONS: The isolation of four antibacterial and antifungal triterpenoids is reported for the first time from Curtisia dentata. This study provides information on the antimicrobial compounds of this species, as well as a preliminary rationale for the use in traditional South African medicine.     

Phytochemical study guided by the myorelaxant activity of the crude extract,fractions and constituent from stem bark of Hymenaea courbaril L.

Ethnopharmacological relevance

Hymenaea courbaril L. (Caesalpinoideae) is used in Brazilian folk medicine to treat anemia, kidney problems, sore throat and other dysfunctions of the respiratory system, such as bronchitis and asthma, although such properties are yet to be scientifically validated.

Aim of the study

In order to give a scientific basis to support the traditional use of Hymenaea courbaril, this study was designed to evaluate antioxidant, myorelaxant and anti-inflammatory properties of the ethanol extract from stem bark and its fractions. The myorelaxant effect of astilbin, a flavonoid isolated from the bioactive ethyl acetate fraction (EAF), has also been evaluated.

Material and methods

In the present study ethanol extract from stem bark (EEHC) and fractions were analyzed using bioassay-guided fractionation. The following activities were investigated: antioxidant by 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay, myorelaxant on rat tracheal smooth muscle, and anti-inflammatory using ovalbumin-induced leukocytosis and airway hyperresponsiveness in rats.

Results

The results of the present investigation show that the whole extract of Hymenaea courbaril and some of its fractions strongly scavenged DPPH radical. The extract showed myorelaxant activity on rat trachea, being EAF its highest efficient fraction. Bio-guided study allowed the isolation of astilbin, a well-known flavonoid. The activity induced by this compound indicates that it may be partly responsible for the myorelaxant effect of EAF. EAF reduced contractions that depended on divalent cation inflow through voltage-operated Ca²⁺ channels (VOCCs) or receptor-operated Ca²⁺ channels (ROCCs), but it was more potent to inhibit VOCC- than ROCC-dependent contraction induced by Ca²⁺ addition in ACh-enriched Ca²⁺-free medium. Oral pretreatment of antigen-challenged animals with EAF prevented airway hyperresponsiveness on KCl-induced contraction and reduced the number of total white cells, particularly eosinophils and neutrophils in bronchoalveolar lavage.

Conclusions

This study provided scientific basis that Hymenaea courbaril presents potential antioxidant, myorelaxant and anti-inflammatory actions, which support its use in folk medicine to treat inflammatory airway diseases.     

Analysis of propolis from Baccharis dracunculifolia DC. (Compositae) and its effects on mouse fibroblasts

This paper confirms Baccharis dracunculifolia DC. (Compositae) as the main botanical source of the propolis from southeastern Brazil (state of São Paulo) investigated to ascertain specific biological activity in relation to mouse NIH-3T3 fibroblasts, skin cells directly involved in the cicatrization processes. Flavonoid and total phenolic compounds were determined by spectrophotometry, and chemical composition by HPLC; the chromatographic profile, characterized largely by flavonoids and aromatic acids, was found to be qualitatively similar to that of Baccharis dracunculifolia DC. The adsorption of phenolic compounds in the propolis to skin powder was also investigated, and 68% of these compounds adsorbed to the skin powder. At concentrations from 0.12 to 7.81 μg/ml, the propolis revealed no statistical significant differences from its control solutions; however, at concentrations of 31.25 μg/ml or more, the propolis was toxic to NIH-3T3 cells. Thus, the propolis from Baccharis dracunculifolia DC. (Compositae) presents an in vitro concentration-dependent toxicity on mouse NIH-3T3 fibroblasts.     

Antibacterial and anti-inflammatory effects of extracts and fractions from Polygonum capitatum

Aim of the study

This study was designed to investigate the antibacterial and anti-inflammatory activities of the extracts and the structure-based fractions from P. capitatum so as to provide the evidence for the traditional use of this plant in the treatment of urinary tract infections and to clarify the structural types that were responsible for the clinical use of the plant.

Materials and methods

The dry whole plant of P. capitatum was extracted with water and 70% aqueous ethanol and then separated, respectively, into a fraction enriched in polysaccharides and proteins (PP) and four other fractions enriched in gallic acid and its analogues (GAA), flavonoids (FV), tannins (TN), and triterpenoids and steroids (TS). UV spectral or chemical methods were used for the confirmation of the five fractions. The in vitro antibacterial activities of the aqueous (AE) and 70% aqueous ethanol (70EE) extracts as well as the fractions against gram-positive and gram-negative bacteria were initially evaluated by a disc diffusion test. The anti-bacterial potencies of the active extracts or fractions were then assessed in vitro by determining the MICs and MBCs. The anti-inflammatory activity was evaluated employing the xylene-induced mouse ear edema model.

Results

Except for fraction PP, AE, 70EE, and the four fractions (GAA, FV, TN, and TS) exhibited varying degrees of antibacterial and anti-inflammatory activities. The results of the minimal inhibition concentration (MIC) and minimal bactericidal concentration (MBC) indicated that the crude extracts or fractions FV and TN all possess bacteriostatic and bactericidal properties. Fractions FV and TS showed significantly anti-inflammatory activity (P < 0.01) with the inhibition rates of 86.15 and 73.71% at 0.6 g/kg, respectively, as compared to 76.93% of the positive control dexamethasone.

Conclusions

The overall results suggested that the traditional use of this plant for the treatment of urinary tract infections were attributed to the presence of antibacterial and anti-inflammatory agents. The results also provided evidence that the studied plant extracts, as well as some of the fractions obtained from this plant might be potential sources for antimicrobial and anti-inflammatory drug development.     

Seselin from Plumbago zeylanica inhibits phytohemagglutinin (PHA)-stimulated cell proliferation in human peripheral blood mononuclear cells

Effects of seselin (C(14)H(12)O(3); MW 228) identified from Plumbago zeylanica on phytohemagglutinin (PHA)-stimulated cell proliferation were studied in human peripheral blood mononuclear cells (PBMC). The data demonstrated that seselin inhibited PBMC proliferation-activated with PHA with an IC(50) of 53.87+/-0.74 microM. Cell viability test indicated that inhibitory effects of seselin on PBMC proliferation were not through direct cytotoxicity. The action mechanisms of seselin may involve the regulation of cell cycle progression, interleukin-2 (IL-2) and interferon-gamma (IFN-gamma) production in PBMC. Since cell cycle analysis indicated that seselin arrested the cell cycle progression of activated PBMC from the G(1) transition to the S phase. Seselin suppressed IL-2 and IFN-gamma production in a concentration-dependent manner. Furthermore, seselin significantly decreased the IL-2 and IFN-gamma gene expression in PHA-activated PBMC. Therefore, results elucidated for the first time that seselin is likely an immunomodulatory agent for PBMC.     

Isolation and characterization of antimicrobial constituents of Searsia chirindensis L. (Anacardiaceae) leaf extracts

Ethnopharmacological relevance

Searsia chirindensis is used in South African traditional medicine for management of bacterial infections such as diarrhoea. Aim of the study was to examine the phytochemical composition from the leaves of Searsia chirindensis that is responsible for the ethnomedicinal use of this plant.

Materials and methods

The crude extract (80% methanol) was extracted sequentially with dichloromethane (DCM), ethyl acetate (EtOAc) and n-butanol. The extracts and isolated compounds were tested for their antibacterial activity against Gram-negative (Campylobacter jejuni, Escherichia coli and Shigella flexneri) and Gram-positive (Staphylococcus aureus) bacterial strains using the microdilution method. Bioguided fractionation of EtOAc fraction afforded five phenolic compounds. Structural elucidation was carried out using NMR (1D and 2D) spectroscopic analyses.

Results

Of the three fractions obtained from the crude extract, EtOAc was the most active and its fractionation afforded methyl gallate (1), and four flavonol glycosides: myricetin-3-O-arabinopyranoside (2), myricetrin-3-O-rhamnoside (3), kaempferol-3-O-rhamnoside (4) and quercetin-3-O-arabinofuranoside (5). These compounds are reported from Searsia chirindensis for the first time. All the compounds showed good antibacterial activity against all bacterial strains tested. Their minimum inhibitory concentrations ranged from 30 to 250 µg/mL.

Conclusions

Antibacterial activity demonstrated by the extracts and isolated compounds provides credence to the ethnomedicinal use of Searsia chirindensis against diarrhoea.     

Antipsychotic and sedative effects of the leaf extract of Crassocephalum bauchiense (Hutch.) Milne-Redh (Asteraceae) in rodents

ETHNOPHARMACOLOGICAL RELEVANCE: Crassocephalum bauchiense (Hutch.) Milne-Redh (Asteraceae) has been used as a medicine for the treatment of epilepsy, insomnia, dementia and psychotic disorders in Cameroonian traditional medicine. AIM OF THE STUDY: This study was designed to examine whether the aqueous extract and the alkaloid fraction prepared from the leaves of Crassocephalum bauchiense possess antipsychotic and sedative properties in rodents. MATERIALS AND METHODS: The rectal temperature of mice was recorded with a probe thermometer at a constant depth. Novelty-induced rearing behavior is used to evaluate a central excitatory locomotor behavior in mice. The antipsychotic effects of the extracts were assessed using the apomorphine animal model of psychosis. The catalepsy test was tested based on the ability of the leaves extracts of Crassocephalum bauchiense to alter the duration of akinesia by placing the naive mice with both forelegs over a horizontal bar. The extracts of Crassocephalum bauchiense effects were evaluated on sodium pentobarbital-induced sleeping time. In addition, gamma-aminobutyric acid concentrations in the brain treated mice were also estimated. RESULTS: The aqueous extract and the alkaloid fraction from Crassocephalum bauchiense caused dose-dependent inhibition of novelty-induced rearing behavior, decreased the apomorphine-induced stereotypy and fighting, and had significant fall of the body temperature. The aqueous extract prolonged the sodium pentobarbital sleeping time. This prolongation was not reversed by bicuculline, a light-sensitive competitive antagonist of GABA(A) receptors complex. However, the effect of the aqueous extract on sodium pentobarbital-induced sleeping time was blocked by N-methyl-β-carboline-3-carboxamide, a partial inverse agonist of the benzodiazepine site in the GABA(A) receptor complex and flumazenil, a specific antagonist of the benzodiazepine site in the GABAA receptor complex. In biochemical experiments, the concentration of the inhibitory amino acid, gamma-aminobutyric acid, was significantly increased in the brain of animals treated with the aqueous extract of Crassocephalum bauchiense and sodium valproate. CONCLUSIONS: The results show that the antipsychotic and sedative properties of Crassocephalum bauchiense are possibly mediated via the blockade of dopamine D-2 receptors and GABAergic activation, respectively. However, pharmacological and chemical studies are continuing in order to characterize the mechanism(s) responsible for these neuropharmacological actions and also to identify the active substances present in the extracts of Crassocephalum bauchiense.     

Hypotensive and vasorelaxant effects of the procyanidin fraction from Guazuma ulmifolia bark in normotensive and hypertensive rats

THE AIM OF THE STUDY: was to investigate the in vivo and in vitro cardiovascular activity of a procyanidin fraction (PCF) obtained from acetone extract of Guazuma ulmifolia bark which has traditionally been used as an antihypertensive agent. RESULTS: 10 mg/kg PCF doses orally administered to sugar-fed hypertensive rats decreased both the systolic arterial pressure and the heart rate, whereas the same doses intravenously administered induced arterial hypotension which was attenuated by NG-nitro-L-arginine methylester (L-NAME 31 mg/kg) pretreatment. In these experiments we employed carbachol as a positive control test. The PCF reduced the contraction induced by norepinephrine (1x10(-7) M) in isolated aortic rings of normotensive (IC50=35.3+/-12.4 ng/mL) and sugar-fed hypertensive (IC50=101.3+/-57.2 ng/mL) rats. This relaxant activity was inhibited by either vascular endothelium removal or L-NAME (30 microM) pretreatment, while indomethacin (10 microM) or atropine (10 microM) had no effect. Preliminary analysis of the PCF by HPLC-DAD-MS and FAB+ mass spectrometry allowed the detection of the main components such as the complex of procyanidin oligomers consisting mainly of tetramers and trimers. CONCLUSIONS: Guazuma ulmifolia bark possesses long-lasting antihypertensive and vasorelaxing properties linked to the endothelium related factors, where nitric oxide is involved.     

Antipsychotic and sedative effects of the leaf extract of Crassocephalum bauchiense (Hutch.) Milne-Redh (Asteraceae) in rodents

Ethnopharmacological relevance

Crassocephalum bauchiense (Hutch.) Milne-Redh (Asteraceae) has been used as a medicine for the treatment of epilepsy, insomnia, dementia and psychotic disorders in Cameroonian traditional medicine.

Aim of the study

This study was designed to examine whether the aqueous extract and the alkaloid fraction prepared from the leaves of Crassocephalum bauchiense possess antipsychotic and sedative properties in rodents.

Materials and methods

The rectal temperature of mice was recorded with a probe thermometer at a constant depth. Novelty-induced rearing behavior is used to evaluate a central excitatory locomotor behavior in mice. The antipsychotic effects of the extracts were assessed using the apomorphine animal model of psychosis. The catalepsy test was tested based on the ability of the leaves extracts of Crassocephalum bauchiense to alter the duration of akinesia by placing the naive mice with both forelegs over a horizontal bar. The extracts of Crassocephalum bauchiense effects were evaluated on sodium pentobarbital-induced sleeping time. In addition, gamma-aminobutyric acid concentrations in the brain treated mice were also estimated.

Results

The aqueous extract and the alkaloid fraction from Crassocephalum bauchiense caused dose-dependent inhibition of novelty-induced rearing behavior, decreased the apomorphine-induced stereotypy and fighting, and had significant fall of the body temperature. The aqueous extract prolonged the sodium pentobarbital sleeping time. This prolongation was not reversed by bicuculline, a light-sensitive competitive antagonist of GABA_A receptors complex. However, the effect of the aqueous extract on sodium pentobarbital-induced sleeping time was blocked by N-methyl-β-carboline-3-carboxamide, a partial inverse agonist of the benzodiazepine site in the GABA_A receptor complex and flumazenil, a specific antagonist of the benzodiazepine site in the GABAA receptor complex. In biochemical experiments, the concentration of the inhibitory amino acid, gamma-aminobutyric acid, was significantly increased in the brain of animals treated with the aqueous extract of Crassocephalum bauchiense and sodium valproate.

Conclusions

The results show that the antipsychotic and sedative properties of Crassocephalum bauchiense are possibly mediated via the blockade of dopamine D-2 receptors and GABAergic activation, respectively. However, pharmacological and chemical studies are continuing in order to characterize the mechanism(s) responsible for these neuropharmacological actions and also to identify the active substances present in the extracts of Crassocephalum bauchiense.     

Biological activities of Pseudevernia furfuracea (L.) Zopf extracts and isolation of the active compounds

Ethnopharmacological relevance

Pseudevernia furfuracea (L.) Zopf (Parmeliaceae) is a common epiphytic lichen in the conifer-hardwood forest of Anatolia. This species is used in traditional medicine in Turkey as a treatment for wounds, eczema and hemorrhoids.

Aim of the study

The present study was designed to investigate the active compounds from Pseudevernia. furfuracea, and the isolation studies yielded atraric acid ( Aslan et al., 2006) as the major compound and a mixture of methyl hematommate ( Baumann, 1960) and methyl chlorohematommate ( Bay?r et al., 2006). Furthermore, methanolic extract from thalli of Pseudevernia. furfuracea and its fractions and isolates ( 1, 2 and 3) were investigated for in vitro antimicrobial and antioxidant activities, and in vivo antinociceptive, anti-inflammatory and wound healing activities.

Material and methods

Antimicrobial activities of the samples were determined by using the disc diffusion technique. 2,2-diphenyl-1-picrylhydrazyl (DPPH) assay was used as a rapid TLC screening method to evaluate the antioxidant activity of Pseudevernia. furfuracea. The thiobarbituric acid (TBA) test was used to assess the efficacy of the extracts in protecting liposomes from lipid peroxidation. In vivo inhibitory effect of the extracts on the carrageenan-induced hind paw edema model in mice was studied for the assessment of anti-inflammatory activity. p-Benzoquinone-induced abdominal constriction test was used to explore the antinociceptive effects of the extracts. Moreover, the wound healing potential of the plant extracts that were evaluated by using in vivo incision and excision wound models on rats and mice, were comparatively assessed with a reference ointment Madecassol^®.

Results

Significant antimicrobial activities were observed against Gram (+) microorganisms and Candida krusei and Candida. dubliniensis in dichloromethane (DCM) and ethyl acetate (EtOAc) extracts and isolates. The methanol (MeOH), DCM and EtOAc extracts of the lichen were found to possess moderate inhibitory activity on lipid peroxidation. Methanolic extract of the lichen was found to possess significant inhibitory activity on the carrageenan-induced hind paw edema model in mice whereas the other fractions did not show any activity. While DCM and EtOAc extracts and fractions showed notable anti-inflammatory activity on carrageenan-induced hind paw edema model without inducing any apparent acute toxicity or gastric damage. Moreover, topical application of the ointment prepared with MeOH extract and EtOAc fraction onto the incised wounds exerted remarkable wound healing activity.

Conclusion

The results of these experimental studies exhibited that nonpolar fractions of Pseudevernia. furfuracea have significant antimicrobial activity against especially Candida species and polar fractions (especially MeOH) display antioxidant, anti-inflammatory, antinociceptive and wound healing activities.     

Isolation of narciprimine from Cyrtanthus contractus (Amaryllidaceae) and evaluation of its acetylcholinesterase inhibitory activity

Ethnopharmacological relevance

Plants of the family Amaryllidaceae are used widely in traditional medicine in South Africa. Several of these, including representatives of the genus Cyrtanthus find use in the treatment of mental illness and age-related dementia.

Aim of the study

Based on the distribution of central nervous system-activating alkaloidal constituents within the genus Cyrtanthus, Cyrtanthus contractus was here explored for such compounds which could interact with acetylcholinesterase (AChE), of significance in the progression of neurodegeneration associated with Alzheimer's disease.

Materials and methods

The known phenanthridone alkaloid narciprimine was isolated via column chromatography of the ethanolic extract of bulbs of Cyrtanthus contractus. The structure of the compound was determined by high field 2D NMR and mass spectroscopic techniques. The classical method of Ellman et al. was used in the determination of AChE inhibitory activity.

Results

The isolation of narciprimine from Cyrtanthus contractus is a landmark find since it has previously only been identified in Zephyranthes, Narcissus and Lycoris, genera endemic to the Americas, Europe and Asia, respectively. Narciprimine exhibited micromolar inhibitory activity (IC₅₀ 78.9) against the enzyme acetylcholinesterase.

Conclusion

This work represents the first isolation of narciprimine from an African Amaryllidaceae species, which may be of chemotaxonomic significance. The AChE inhibitory activity of narciprimine, taken together with activities of other structurally similar inhibitors within the series affords further insight to the structural details of the lycorine alkaloid acetylcholinesterase inhibitory pharmacophore.     

Antimicrobial activity of southern African medicinal plants with dermatological relevance: From an ethnopharmacological screening approach,to combination studies and the isolation of a bioactive compound

Ethnopharmacological relevance

Ethnobotanical reports on more than 100 southern African medicinal plants with dermatological relevance have been highlighted, yet there is still limited scientific data to support claims for their antimicrobial effectiveness against skin pathogens. Guided by ethnobotanical data, this paper explores the antimicrobial efficacies of southern African medicinal plants used to treat skin ailments.

Aim of the study

To investigate the antimicrobial properties of southern African medicinal plants against dermatologically relevant pathogens. The study also aimed at providing a scientific rationale for the traditional use of plant combinations to treat skin diseases and the isolation of the bio-active compound from the most active species, Aristea ecklonii (Iridaceae).

Materials and methods

Organic and aqueous extracts (132) were prepared from 47 plant species and screened for antimicrobial properties against dermatologically relevant pathogens using the micro-titre plate dilution method. Four different plant combinations were investigated for interactive properties and the sum of the fractional inhibitory concentration (?FIC) calculated. Isobolograms were used to further investigate the antimicrobial interactive properties of Pentanisia prunelloides combined with Elephantorrhiza elephantina at varied ratios. A bioactivity-guided fractionation process was adopted to fractionate the organic leaf extract of Aristea ecklonii.

Results

Plants demonstrating notable broad-spectrum activities (MIC values ≤1.00 mg/ml) against the tested pathogens included extracts from Aristea ecklonii, Chenopodium ambrosioides, Diospyros mespiliformis, Elephantorrhiza elephantina, Eucalyptus camaldulensis, Gunnera perpensa, Harpephyllum caffrum, Hypericum perforatum, Melianthus comosus, Terminalia sericea and Warburgia salutaris. The organic extract of Elephantorrhiza elephantina, a plant reportedly used to treat acne vulgaris, demonstrated noteworthy antimicrobial activity (MIC value of 0.05 mg/ml) against Propionibacterium acnes. Similarly, Diospyros mespiliformis reported for its traditional use to treat ringworm, also displayed noteworthy antimicrobial activity against Trichophyton mentagrophytes (MIC 0.10 mg/ml) and Microsporum canis (MIC 0.50 mg/ml). The aqueous root extracts of Pentanisia prunelloides combined (1:1) with Elephantorrhiza elephantina displayed synergistic interactions (?FIC values 0.31–0.38) against Staphylococcus aureus, gentamycin–methicillin resistant Staphylococcus aureus, Staphylococcus epidermidis and Candida albicans. Fractionation of Aristea ecklonii resulted in the isolation of the known bio-active compound, plumbagin, displaying noteworthy antimicrobial activity (MIC range between 2.00 μg/ml and 16.00 μg/ml).

Conclusion

Most of the plant extracts demonstrated pathogen specific antimicrobial effects with a few exhibiting broad-spectrum activities. Positive antimicrobial effects noted for plants such as Elephantorrhiza elephantina and Diospyros mespiliformis used for acne vulgaris and ringworm infections, respectively, give some validation to their reported traditiona l uses. Synergistic interactions noted for Pentanisia prunelloides combined with Elephantorrhiza elephantina validate an enhanced antimicrobial effect when used in combination. Noteworthy antimicrobial activities (MIC range between 2.00 μg/ml and 16.00 μg/ml) were observed for plumbagin isolated from Aristea ecklonii.     

Chemical variability along the value chains of turmeric (Curcuma longa): A comparison of nuclear magnetic resonance spectroscopy and high performance thin layer chromatography

Ethnopharmacological relevance

Herbal medicine value chains have generally been overlooked compared with food commodities. Not surprisingly, revenue generation tends to be weighted towards the end of the chain and consequently the farmers and producers are the lowest paid beneficiaries. Value chains have an impact both on the livelihood of producers and on the composition and quality of products commonly sold locally and globally and consequently on the consumers. In order to understand the impact of value chains on the composition of products, we studied the production conditions for turmeric (Curcuma longa) and the metabolomic composition of products derived from it. We aimed at integrating these two components in order to gain a better understanding of the effect of different value chains on the livelihoods of some producers.

Materials and methods

This interdisciplinary project uses a mixed methods approach. Case studies were undertaken on two separate sites in India. Data was initially gathered on herbal medicine value chains by means of semi-structured interviews and non-participant observations. Samples were collected from locations in India, Europe and the USA and analysed using ¹H NMR spectroscopy coupled with multivariate analysis software and with high performance thin layer chromatography (HPTLC).

Results

We investigate medicinal plant value chains and interpret the impact different value chains have on some aspects of the livelihoods of producers in India and, for the first time, analytically assess the chemical variability and quality implications that different value chains may have on the products available to end users in Europe. There are benefits to farmers that belonged to an integrated chain and the resulting products were subject to a higher standard of processing and storage.By using analytical methods, including HPTLC and ¹H NMR spectroscopy, it has been possible to correlate some variations in product composition for selected producers and identify strengths and weaknesses of some types of value chains. The two analytical techniques provide different and complementary data and together they can be used to effectively differentiate between a wide variety of crude drug powders and herbal medicinal products.

Conclusions

This project demonstrates that there is a need to study the links between producers and consumers of commodities produced in so-called ‘provider countries’ and that metabolomics offer a novel way of assessing the chemical variability along a value chain. This also has implications for understanding the impact this has on the livelihood of those along the value chain.     

Endothelium-dependent and independent vasorelaxation induced by an n-butanolic fraction of bark of Scutia buxifolia Reiss (Rhamanaceae)

Ethnopharmacological relevance

Scutia buxifolia has been widely used in Brazilian folk medicine as an anti-hypertensive agent.We evaluated the vascular effects and mechanism involved in the relaxation of aorta induced by an n-butanolic fraction (BuOH) from Scutia buxifolia.

Materials and Methods

Rat aortic rings precontracted by phenylephrine (1 μM) were exposed to cumulative concentrations (3–3000 μg/ml) of crude extracts or fractions obtained from bark or leaves of Scutia buxifolia. Classical receptor antagonists, channel and enzymatic inhibitors were used to check the mechanisms involved.

Results

The crude extracts of both leaves and bark of Scutia buxifolia, as well as several fractions, were able to induce partial or total relaxation of rat aortic rings. The BuOH fraction of bark of Scutia buxifolia was the most potent in endothelium-intact (E+) preparations, and also induced a partial, but very significant relaxation in endothelium-denuded (E−) vessels. The non-selective nitric oxide synthase inhibitor L-NAME, as well as the soluble guanylate cyclase inhibitor ODQ, vanished the relaxation in E+. In E− preparations, K⁺ channel blockers, such as tetraethylammonium, glibenclamide, 4-aminopyridine, and the large-conductance calcium-activated K⁺ channel blocker iberiotoxin, were able to significantly reduce the maximum relaxation elicited by BuOH fraction.

Conclusion

Our results demonstrated that BuOH fraction obtained from barks of Scutia buxifolia induced both endothelium-dependent and -independent relaxation in rat aortic rings. The endothelium-dependent relaxation is fully dependent on NO/cGMP system, while direct activation of K⁺ channels may explain, at least in part, the endothelium-independent relaxation induced by BuOH fraction of Scutia buxifolia.     

The anti-inflammatory activities of an extract and compounds isolated from Platycladus orientalis (Linnaeus) Franco in vitro and ex vivo

Ethnopharmacological relevance

As a Chinese traditional herbal medicine, leaves of Platycladus orientalis (Linnaeus) Franco (LPO) are used to treat coughs, excessive mucus secretion, chronic bronchitis, bronchiectasis, and asthma, etc. The experiments were carried out to investigate their anti-inflammatory properties and mechanisms, which could support the Chinese traditional uses of treating inflammatory airway diseases.

Materials and methods

The anti-inflammatory activities of the chloroform fraction (CHL) and pure compounds of LPO were evaluated for their abilities to inhibit pro-inflammatory enzymes in vitro, and production of tumor necrosis factor-α (TNF-α) and nitric oxide in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. Furthermore, the arachidonic acid metabolites, stimulated by calcium ionophore A23187, were also determined by HPLC.

Results

For the first time, the assays of eicosanoids in intact cells showed that the CHL, hinokiol, and acacetin had significant inhibitory effects on 5-hydroxy-eicosa-tetra-enoic acid (5-HETE) and leukotriene B₄ (LTB4) formations. And cell-free enzyme assays (5-lipoxygenase, leukotriene A₄-hydrolase, cyclooxgenase-2) demonstrated the potent inhibitory effects of the CHL, hinokiol and acacetin on 5-lipoxygenase (5-LOX). Then, the inhibitions of the CHL, hinokiol on NO biosynthesis and the inhibitions of the CHL, 8(14),15-pimaradien-3β,18-diol, and hinokiol on TNF-α release were also confirmed in the RAW264.7 murine macrophages.

Conclusion

The data indicate that the inhibitory effects of the CHL and its components (hinokiol and acacetin) on 5-LOX contribute to the anti-inflammatory activity of LPO. Moreover, the CHL and its components also show beneficial effects on NO and TNF-α production. Consequently, these results provide a rationale for LPO's traditional applications in the treatment of inflammatory airway diseases.     

In vivo anti-inflammatory and antinociceptive activity of the crude extract and fractions from Rosa canina L. fruits

The aqueous and ethanol extracts of Rosa canina L. (Rosaceae) fruits and the fractions prepared from the latter were investigated for their anti-inflammatory and antinociceptive activities in several in vivo experimental models. The ethanolic extract was shown to possess significant inhibitory activity against inflammatory models (i.e., carrageenan-induced and PGE(1)-induced hind paw edema models, as well as on acetic acid-induced increase in a capillary permeability model) and on a pain model based on the inhibition of p-benzoquinone-induced writhing in mice. Hexane, chloroform, ethylacetate, n-butanol and the remaining water fractions were obtained through bioassay-guided fractionation. Ethylacetate and n-butanol fractions displayed potent anti-inflammatory and antinociceptive activities at a dose of 919 mg/kg without inducing acute toxicity. Further attempts to isolate and define the active constituent(s) were inconclusive, possibly due to the synergistic interaction of components in the extract.     

Inhibitory effects of adlay bran (Coix lachryma-jobi L. var. ma-yuen Stapf) on chemical mediator release and cytokine production in rat basophilic leukemia cells

Ethnopharmacological relevance

Adlay (Job's tears, Coix lachryma-jobi L. var. ma-yuen Stapf) has long been used in China to treat rheumatism.

Aim of the study

We investigated the anti-allergic effects of adlay bran on rat basophilic leukemia (RBL)-2H3 cells.

Materials and methods

To evaluate the anti-allergic effects of adlay bran, the release of histamines and cytokines were measured using ELISA. To explore the mechanism of these effects, the protein expression levels were determined using western blotting.

Results

A 40.8 μg/mL concentration of the ethyl acetate fraction of the ethanolic extracts of adlay bran (ABE-EtOAc) effectively inhibited mast cell degranulation. The 40–100% EtOAc/Hex subfractions of ABE-EtOAc inhibited histamine release with an IC₅₀ of 71–87 μg/mL. Moreover, the ABE-EtOAc subfractions suppressed the secretion of interleukin (IL)-4, IL-6 and tumor necrosis factor-α in the RBL-2H3 cells, indicating that adlay bran can inhibit cytokine secretion in the late phase of the allergic reaction. In addition, adlay bran reduced the intracellular production of reactive oxygen species, inhibited the phosphorylation of Akt and decreased the expression of protein kinase C. Furthermore, six phenolic acids and one flavone were isolated. Of these compounds, luteolin showed the most potent inhibitory activity (IC₅₀ = 1.5 μg/mL).

Conclusion

Adlay bran extract reduced the release of histamines and cytokines and suppressed the production of Akt. These combined effects influenced the signal transduction in RBL-2H3 cells, thereby revealing the mechanisms of the anti-allergic effects of adlay.     

Bioactivity guided isolation of analgesic and anti-inflammatory constituents of Cnestis ferruginea Vahl ex DC (Connaraceae) root

Ethnopharmacological relevance

Cnestis ferruginea (CF) Vahl ex DC (Connaraceae) is a shrub widely used in Traditional African Medicine (TAM) for the treatment of various painful and inflammatory conditions.

Aim of the study

To isolate the active pharmacological constituents responsible for the anti-inflammatory and antinociceptive properties of the methanolic root extract of C. ferruginea.

Materials and methods

The crude methanolic root extract of CF was sequentially fractionated into four sub extracts (chloroform, ethylacetate, n-butanol and the remaining aqueous fraction). The aqueous–butanol fractions, having showed significant inhibition of inflammation and pain, were subjected to fractionation through successive column chromatography on silica gel 60–120 mesh, eluted with a gradient of CHCl₃–MeOH.Sixty five fractions were collected; fractions with similar TLC profiles were grouped into seven major fractions (1–7). Fraction 4 being the most active in bioassay was rechromatographed to obtain CF-2. Analgesic activity was evaluated using the acetic acid-induced writhing and hot plate tests in mice while carrageenan induced paw oedema test was used to investigate the anti-inflammatory actions of the fractions obtained.

Result

Amentoflavone (CF-2) was isolated from the aqueous/n-butanol fraction. CF-2 (12.5, 25 and 100 mg/kg; p.o) produced significant (P<0.05) dose dependent inhibition of pain response elicited by acetic acid and increased nociceptive reaction latency in hot plate test. In addition it produced significant (P<0.05) dose-dependent inhibition of oedema in the carrageenan-induced inflammation.

Conclusion

This study showed that amentoflavone is responsible for the analgesic and anti-inflammatory activity of Cnestis ferruginea.     

Analgesic activity of affinin, an alkamide from Heliopsis longipes (Compositae)

Heliopsis longipes (Compositae) is a Mexican plant used as analgesic in pain toothache. A solution of 10mug/ml of dichloromethane extract from this plant showed analgesic activity determined by means of GABA release in mice brain slices. Through a bioassay-directed separation, fractions G-1, G-2, G-4 and G-6 at the same concentration were active. Affinin was the unique and common active compound, and evoke the GABA release 0.5min after administration at 1x10(-4)M concentration. Inactive compound were undeca-2E-en-8,10-dyinoic acid isobutylamide, hinokinin, 2'-hydroxyhinokinin, 3beta-sn-glyceroyl-(1'-palmitoxy)urs-12-ene, 13(18)-ursen-3beta-ol, 13(18)-ursen-3beta-acetate, beta-sitosterol and stigmasterol. The analgesic activity of Heliopsis longipes could be associated to affinin.     

Antimicrobial activity of the crude extracts and five flavonoids from the twigs of Dorstenia barteri (Moraceae)

The aim of this study was to evaluate the antimicrobial activity of the crude extract of the twigs of Dorstenia barteri (DBT) as well as that of four of the five flavonoids isolated from this extract. Gram-positive bacteria (six species), Gram-negative bacteria (12 species) and fungi (four species) were used. The agar disc diffusion test was used to determine the sensitivity of the tested samples while the well micro-dilution was used to determine the minimal inhibition concentrations (MIC) and the minimal microbicidal concentration (MMC) of the active samples. The results of the disc diffusion assay showed that DBT, isobavachalcone (1), and kanzonol C (4) prevented the growth of all the 22 tested microbial species. Other compounds showed selective activity. The inhibitory activity of the most active compounds namely compounds 1 and 4 was noted on 86.4% of the tested microorganisms and that of 4-hydroxylonchocarpin (3) was observed on 72.7%. This lowest MIC value of 19.06microg/ml was observed with the crude extract on seven microorganisms namely Citrobacter freundii, Enterobacter aerogens, Proteus mirabilis, Proteus vulgaris, Bacillus megaterium, Bacillus stearothermophilus and Candida albicans. For the tested compounds, the lowest MIC value of 0.3microg/ml (on six of the 22 organisms tested) was obtained only with compound 1, which appeared as the most active compound. This lowest MIC value (0.3microg/ml) is about 4-fold lower than that of the RA, indicating the powerful and very interesting antimicrobial potential of isobavachalcone (1). The antimicrobial activities of DBT, as well as that of compounds 1, 3, 4, amentoflavone (5) are being reported for the first time. The overall results provide promising baseline information for the potential use of the crude extracts from DBT as well as some of the isolated compounds in the treatment of bacterial and fungal infections.