Alembert T. Tchinda

Antiparasitic activities of two sesquiterpenic lactones isolated from Acanthospermum hispidum D.C.

ETHNOPHARMACOLOGICAL RELEVANCEnAerial parts of Acanthospermum hispidum D.C. are often used by traditional healers in Benin for various diseases and especially for malaria.nnnAIM OF THE STUDYnTo identify active compounds from extracts of Acanthospermum hispidum D.CV. leaves previously shown to possess antimalarial properties and analyse in vivo activity and toxicity of crude extracts.nnnMATERIALS AND METHODSnCompounds were isolated from aerial part of Acanthospermum hispidum D.C. and structurally elucidated using extensive spectroscopic analysis. Antiplasmodial activity was evaluated in vitro against a chloroquine-sensitive strain of Plasmodium falciparum (3D7) using the measurement of the plasmodial lactate dehydrogenase activity and in vivo against Plasmodium berghei berghei by the 4-day suppressive test. Selectivity of extract and purified compounds on Plasmodium parasites were evaluated by using MTT test on J774 macrophage like murine cells and WI38 human normal fibroblasts and also against two other parasites: Trypanosoma brucei brucei and Leishmania mexicana mexicana. Acute and sub-acute toxicities of a crude extract were evaluated on mice.nnnRESULTSnTwo known sesquiterpenic lactones were isolated: 1 (15-acetoxy-8β-[(2-methylbutyryloxy)]-14-oxo-4,5-cis-acanthospermolide) and 2 (9α-acetoxy-15-hydroxy-8β-(2-methylbutyryloxy)-14-oxo-4,5-trans-acanthospermolide). 1 and 2 showed in vitro antiplasmodial activity against the chloroquine-sensitive strain (3D7) with IC(50) of 2.9±0.5 and 2.23±0.09μM respectively. Only 2 showed a high selectivity index (SI: 18.4) on Plasmodium compared to cytotoxicity against human fibroblasts cell line (WI38). 1 and 2 also showed interesting antiparasitic activities in vitro against Trypanosoma brucei brucei (IC(50) of 2.45±0.49 and 6.36±1.42μM respectively) and Leishmania mexicana mexicana (IC(50) of 0.94±0.05 and 2.54±0.19μM respectively). Furthermore, crude acidic water extract and fractions containing one of the two isolated compounds displayed a weak in vivo antimalarial activity against Plasmodium berghei berghei with a long half-life causing a delayed effect. In vivo acute (2000mg/kg) and sub-acute (1000mg/kg) toxicity tests on the crude acidic water extract did not show toxicity.nnnCONCLUSIONnCrude acidic water extract, fractions and pure isolated compounds from Acanthospermum hispidum showed promising in vitro antiplasmodial activity. Despite our study did not show in vivo acute and subacute toxicities of the crude acidic water extract, its weak in vivo antimalarial activity and the in vitro cytotoxicity of pure compounds and enriched extracts containing 1 and 2 indicate that the aerial parts of Acanthospermum hispidum should be used with caution for malaria treatments.

Antiplasmodial Alkaloids from the Stem Bark of Strychnos malacoclados.

From the stem bark of Strychnos malacoclados, one new bisindole alkaloid, 3-hydroxylongicaudatine Y (1), was isolated along with the known alkaloids vomicine (2), bisnordihydrotoxiferine (3), divarine (4), longicaudatine (5), longicaudatine Y (6), and longicaudatine F (7). All the compounds were tested for their antimalarial activity against the chloroquine-sensitive 3D7 and -resistant W2 strains of Plasmodium falciparum. Longicaudatine was the most active compound with IC₅₀ values of 0.682 and 0.573 µM, respectively. The activity of compounds 1, 3, 4, 6, and 7 against the two strains ranged from 1.191 to 6.220 µM and 0.573 to 21.848 µM, respectively. Vomicine (2), the only monomer isolated, was inactive. The alkaloids of the longicaudatine-type ( 1, 5-7) were more active than those of the caracurine-type (3- 4). The presence of the ether bridge in the molecule seems to increase the antiplasmodial activity. Compounds 1, 5, and 7 were tested against the WI-38 human fibroblast cell line. Longicaudatine was the most cytotoxic compound with an IC₅₀ of 2.721 µM. Longicaudatine F was 40-46 times more active against the two strains of P. falciparum than against the human fibroblasts and was thus considered as the more selective alkaloid. The structures of the compounds were determined based on the analysis of their spectral data.

Strychnobaillonine, an unsymmetrical bisindole alkaloid with an unprecedented skeleton from Strychnos icaja roots.

A reinvestigation of the roots of Strychnos icaja resulted in the isolation of a new bisindole alkaloid named strychnobaillonine (1) with original C-17-N-1 and C-23-C-17 junctions, in addition to sungucine, bisnordihydrotoxiferine, and strychnohexamine (2). Compound 1 showed potent activity against the chloroquine-sensitive 3D7 strain of Plasmodium falciparum in vitro with an IC50 value of 1.1 μM. The structures of the compounds were defined by detailed spectroscopic analyses, especially 1H and 13C NMR, DEPT, HSQC, COSY, NOESY, HMBC, and HRESIMS. The proposed absolute configuration was based on biosynthetic considerations and spectroscopic data (CD, NMR) supported by molecular modeling.

Antiplasmodial activity of Mezoneuron benthamianum leaves and identification of its active constituents

ETHNOPHARMACOLOGICAL RELEVANCEnDecoctions of the leaves of M. benthamianum Baill. are used by traditional healers in Guinea to treat malaria and this use was validated by a preliminary clinical assay.nnnAIM OF THE STUDYnTo evaluate the in vitro antiplasmodial activity and to identify active compounds from extracts of M. benthamianum leaves.nnnMATERIAL AND METHODSnAntiplasmodial activity of extracts, fractions and pure compounds was evaluated in vitro against a chloroquine-sensitive strain of Plasmodium falciparum (3D7) using the measurement of the plasmodial lactate dehydrogenase activity. Selectivity of extracts and purified compounds for Plasmodium parasites was evaluated by using WST-1 test on HeLa human cells. Compounds were isolated using normal phase silica gel column chromatography and prepHPLC and their structures elucidated using extensive spectroscopic analysis.nnnRESULTSnHydroethanolic extracts (70% v/v) of M. benthamianum leaves showed a moderate in vitro activity against P. falciparum 3D7, with IC50 in the range 22.5 - 32.6µg/mL, depending on the batch; while a dark precipitate formed during ethanol evaporation showed higher activity (IC50 =6.5µg/mL). The fractionation was performed on this most active fraction and was followed by in vitro antiplasmodial assay. Active compounds (5, 7, 8) belong to several phytochemical classes, contributing together to the global antiplasmodial activity of the hydroethanolic extract against P. falciparum parasite. This study finally allowed the isolation of three diterpenes including two new compounds named Mezobenthamic acids A (1) and B (2) and neocaesalpin H (3), as well as quercetin (4), kaempferol (7), resveratrol (6), gallic acid (9) and its ethylester (5), β-sitosterol glucoside (10) and 13b-hydroxy-pheophorbide a (8).nnnCONCLUSIONnThis study gives some concrete evidence to support the ethnopharmacological use of Mezoneuron benthamianum leaves extract in the management of malaria. The active compounds can be further studied for their antiplasmodial potential, as well as their suitability to be used as quality markers for the standardization of this herbal drug from the Guinean traditional pharmacopeia.

Antiplasmodial activity of Heinsia crinita (Rubiaceae) and identification of new iridoids.

ETHNOPHARMACOLOGICAL RELEVANCEnHeinsia crinita is used in traditional medicine for the treatment of febrile illness and erectile dysfunction. Its stem bark powder is found in some peripheral markets in the Democratic Republic of the Congo (DRC) as a remedy against malaria. Investigations were conducted on crude extracts of leaves, fruits and stem barks in view to validate their use and to determine which plant part possesses the best antiplasmodial properties.nnnMATERIALS AND METHODSnDifferent plant parts were extracted with methanol, ethanol and dichloromethane. Based on the preliminary assays, the dichloromethane extract of the stem bark was subjected to fractionation using preparative HPLC system and column chromatography. This step led to the isolation of two new iridoids which had their structures elucidated by NMR, UV, MS and FT-IR spectroscopic techniques. Extracts and pure compounds were tested in vitro against the 3D7 strain of Plasmodium falciparum. The inhibition of the parasite growth was evaluated in vitro by colorimetric method (p-LDH assay) and their cytotoxicity evaluated in vitro against the human non-cancer fibroblast cell line (WI38) through WST1 assay. The in vivo antiplasmodial activity was assessed by the inhibition of Plasmodium berghei growth in infected mice treated with the ethanol extract of H. crinita stem bark at the concentrations of 200 and 300mg/Kg/day per os, using a protocol based on the 4-d suppressive test of Peters and compared to a non-treated negative control group of mice (growth =100%). Finally the antioxidant activity of the same extract was evaluated using ABTS, DPPH and cell-based assays.nnnRESULTSnA moderate in vitro antiplasmodial activity was observed for the dichloromethane extract of the stem bark of H. crinita (IC50 =29.2±1.39µg/mL) and for the two new iridoids, lamalbide 6, 7, 8- triacetate (IC50 =16.39±0.43µg/mL) as well as for its aglycone lamiridosin 6, 7, 8-triacetate (IC50 =0.44.56±1.12µg/mL). The ethanolic stem bark extract (200 and 300mg/kg/day, oral route) showed a moderate in vivo antimalarial activity in Plasmodium berghei-infected mice with 27.84±2.75% and 48.54±3.76% of inhibition of the parasite growth, respectively (p<0.01).). This extract displayed high cellular antioxidant activity using dichlorofluorescein-diacetate (DCFDA) on HL-60 monocytes. These crude extracts and pure compounds tested at the higher concentration of 100µg/mL did not show any cytotoxicity against WI38 cells.nnnCONCLUSIONSnThe results showed that H. crinita extracts possess antimalarial activity and contain some unusual iridoids with moderate antiplasmodial activity, therefore justifying to some extent its traditional use by the local population in DRC for this purpose. This is the first report of the isolation and antiplasmodial activity of these two new iridoids.

Carbon Multiplicity Editing in Long-Range Heteronuclear Correlation NMR Experiments: A Valuable Tool for the Structure Elucidation of Natural Products

A recently developed NMR method to simultaneously obtain both long-range heteronuclear correlations and carbon multiplicity information in a single experiment, ME-selHSQMBC, is demonstrated as a potentially useful technique for chemical shift assignment and structure elucidation of natural products presenting complicated NMR spectra. Carbon multiplicities, even for C/CH2 and odd for CH/CH3 resonances, can be distinguished directly from the relative positive/negative phase of cross-peaks. In addition, connectivity networks can be further extended by incorporating a TOCSY propagation step. Staurosporine (1) and sungucine (2) are utilized as model compounds to demonstrate these techniques.

Vitellaroside, A New Cerebroside from Vitellaria paradoxa (Sapotaceae) and its Bioactivities

A new cerebroside (2R)-2-hydroxy-N-[(Z,2S,3S,4R)-1-O-β-D-glucopyranosyl-3,4-dihydroxynonadec-8-en-2-yl] nonacosanamide (1) was isolated from the wood of roots of V. paradoxa along with six known compounds including catechin (2), quercetin (3), spinasterol 3-O-β-D -glucopyranoside (6), gallic acid (7) and a mixture of β-sitosterol (4) and stigmasterol (5). The structure of the new compound was established by 1D (1H and 13C NMR) and 2D NMR (COSY and HSQC) spectroscopy, extensive mass spectrometry and by comparison with published data. The antibacterial, α-glucosidase and alkaline phosphatase (AP) inhibitory activities of all the pure compounds were evaluated. The antibacterial activities were evaluated against three gram negative bacteria (Escherichia coli; Salmonella typhimurium and Pseudomonas aeruginosa) while APs inhibitory activities were evaluated on h-TNAP and h-IAP. Significant antibacterial activity was recorded for quercetin (3) against P. aeruginosa. Most of the compounds except 1 and 6 were found to be inhibitors of α-glucosidase. The highest inhibitory potential being recorded for quercetin (3) with IC50 value of 4.30 ± 0.01 μM, 55 fold higher than the standard drug acarbose (IC50=234.6 ± 2.01 μM). All tested compounds exhibited moderate inhibitory activities against APs. h-TNAP inhibitory values were ranged between 41.24 ± 1.33 μM and 312.54 ± 6.44 μM while h-IAP inhibitory values were in the range of 47.95 ± 0.35 μM and 777.47 ± 18.55 μM. Quercetin (3) was found to be the most active h-IAP inhibitor (IC50=47.95 ± 0.35 mM), whereas, spinasterol 3-O-β-D-glucopyranoside (6) was found to be the most active h-TNAP inhibitor (IC50=41.24 ± 1.33 mM). The new compound (1) showed moderate inhibition on h-IAP (78.11 ± 3.70 μM) and on h-TNAP (88.84 ± 2.70 μM).

Lipids constituents from Gardenia aqualla Stapf & Hutch

Abstract Gardenia aqualla a plant of the Rubiaceae family is being used extensively in Africa, particularly in Cameroon as an herbal medicine. Therefore it is necessary to have knowledge of the constituents of the plant of our native species. Thus, the aim of the present study was to investigate chemical constituents of this herbal medicine. Nine compounds, including one alkane, n-Nonacosane (1), two aliphatic alcohols n-heptatriacontanol (2) and n-Docosanol(3), one fatty ester, Heptadecyl heptacosanoate (4), two sugars, D-mannitol (5) and D-mannitol acetate (6), and a mixture of three phytosterols, β-sitosterol (7), stigmasterol (8) and fucosterol (9), have been isolated and purified from the stem barks of Gardenia aqualla Stapf & Hutch. Their structures were elucidated using spectroscopic analyses, including 1D and 2D NMR and ESI-MS. The fatty acid ester, heptadecyl heptacosanoate (4) is reported here for the first time. All the isolated compounds were tested for their antimicrobial activities against four Gram negative bacteria (Salmonella Typhimurium ATCC6539, Pseudomonas aeruginosa ATCC9721, Escherichia coli, and S. Typhi) and four yeasts (Candida albicans ATCC9002, Candida parapsilosis ATCC22019, Candida krusei and C. albicans).