Flore Nardella

Antiplasmodial and anti-inflammatory effects of an antimalarial remedy from the Wayana Amerindians, French Guiana: Takamalaimë (Psidium acutangulum Mart. ex DC., Myrtaceae)

ETHNOPHARMACOLOGICAL RELEVANCE Field investigations highlighted the use of Psidium acutangulum Mart. ex DC (syn. P. persoonii McVaugh), a small tree used by the Wayana Amerindians in Twenke-Taluhwen and Antecume-Pata, French Guiana, for the treatment of malaria, and administered either orally in the form of a decoction or applied externally over the whole body. This use appears limited to the Wayana cultural group in French Guiana and has never been reported anywhere else. Our goal was to evaluate the antimalarial and anti-inflammatory activities of a P. acutangulum decoction to explain the good reputation of this remedy. MATERIALS AND METHODS Interviews with the Wayana inhabitants of Twenke-Taluhwen and Antecume-Pata were conducted within the TRAMAZ project according to the TRAMIL methodology, which is based on a quantitative and qualitative analysis of medicinal plant uses. A decoction of dried aerial parts of P. acutangulum was prepared in consistency with the Wayana recipe. In vitro antiplasmodial assays were performed on chloroquine-resistant FcB1 ([(3)H]-hypoxanthine bioassay) and 7G8 (pLDH bioassay) P. falciparum strains and on chloroquine sensitive NF54 ([(3)H]-hypoxanthine bioassay) P. falciparum strain. In vitro anti-inflammatory activity (IL-1β, IL-6, IL-8, TNFα) was evaluated on LPS-stimulated human PBMC and NO secretion inhibition was measured on LPS stimulated RAW murine macrophages. Cytotoxicity of the decoction was measured on L6 mammalian cells, PBMCs, and RAW cells. A preliminary evaluation of the in vivo antimalarial activity of the decoction, administered orally twice daily, was assessed by the classical four-day suppressive test against P. berghei NK65 in mice. RESULTS The decoction displayed a good antiplasmodial activity in vitro against the three tested strains, regardless to the bioassay used, with IC50 values of 3.3µg/mL and 10.3µg/mL against P. falciparum FcB1 and NF54, respectively and 19.0µg/mL against P. falciparum 7G8. It also exhibited significant anti-inflammatory activity in vitro in a dose dependent manner. At a concentration of 50µg/mL, the decoction inhibited the secretion of the following pro-inflammatory cytokines: TNFα (-18%), IL-1β (-58%), IL-6 (-32%), IL-8 (-21%). It also exhibited a mild NO secretion inhibition (-13%) at the same concentration. The decoction was non-cytotoxic against L6 cells (IC50>100µg/mL), RAW cells and PBMC. In vivo, 150µL of the decoction given orally twice a day (equivalent to 350mg/kg/day of dried extract) inhibited 39.7% average parasite growth, with more than 50% of inhibition in three mice over five. The absence of response for the two remaining mice, however, induced a strong standard deviation. CONCLUSIONS This study highlighted the in vitro antiplasmodial activity of the decoction of P. acutangulum aerial parts, used by Wayana Amerindians from the Upper-Maroni in French Guiana in case of malaria. Its antioxidant and anti-inflammatory potential, which may help to explain its use against this disease, was demonstrated using models of artificially stimulated cells.

Purpureone, an antileishmanial ergochrome from the endophytic fungus Purpureocillium lilacinum

Abstract The ethyl acetate extracts prepared from the mycelia of three endophytic fungi Purpureocillium lilacinum, Aspergillus sp., and Fusarium sp., isolated from the roots of Rauvolfia macrophylla (Apocynaceae) were screened for their antiprotozoal activity in vitro against Plasmodium falciparum (NF54), Leishmania donovani, Trypanosoma brucei rhodesiense, and Trypanosoma cruzi. Amongst these extracts, the one from P. lilacinum showed potent antileishmanial activity against L. donovani (IC50 value of 0.174 μg mL−1) with good selectivity (SI=94.9) toward the L6 cell line, whereas the other extracts were inactive and not selective. The fractionation and purification of the active extract from P. lilacinum by column chromatography over silica gel yielded a new ergochromone derivative (1), together with six known compounds: (22E,24R)-stigmasta-5,7,22-trien-3-β-ol (2), (22E,24R)-stigmasta-4,6,8(14),22-tetraen-3-one (3), emodin (4), chrysophanol (5), aloe-emodin (6), and palmitic acid, whose structures were elucidated spectroscopically. Compound 1 was tested in vitro for its antiparasitic activities against the above listed parasites and for its antimicrobial activity against Staphylococcus aureus, Bacillus cereus, Listeria monocytogenes, Escherichia coli, Providencia stuartii, Klebsiella pneumoniae, and Pseudomonas aeruginosa. The compound displayed potent antileishmanial activity against L. donovani with an IC50 value of 0.63 μg mL−1 (0.87 μm) with good selectivity (SI=49.5) toward the L6 cell line. It also exhibited good antibacterial activity against three of the tested microbial strains B. cereus, E. coli ATCC879, and P. stuartii ATCC29916 with minimum inhibitory concentrations below 62.6 μg mL−1. Compound 1 is thus a promising active compound that could be investigated for antileishmanial and antimicrobial drug development.

Wayanin and guaijaverin, two active metabolites found in a Psidium acutangulum Mart. ex DC (syn. P. persoonii McVaugh) (Myrtaceae) antimalarial decoction from the Wayana Amerindians

ETHNOPHARMACOLOGICAL RELEVANCE Psidium acutangulum Mart. ex DC is a small tree used by the Wayana Amerindians from the Upper-Maroni in French Guiana for the treatment of malaria. AIM OF THE STUDY In a previous study, we highlighted the in vitro antiplasmodial, antioxidant and anti-inflammatory potential of the traditional decoction of P. acutangulum aerial parts. Our goal was then to investigate on the origin of the biological activity of the traditional remedy, and eventually characterize active constituents. MATERIALS AND METHODS Liquid-liquid extractions were performed on the decoction, and the antiplasmodial activity evaluated against chloroquine-resistant FcB1 ([(3)H]-hypoxanthine bioassay) and 7G8 (pLDH bioassay) P. falciparum strains, and on a chloroquine sensitive NF54 ([(3)H]-hypoxanthine bioassay) P. falciparum strain. The ethyl acetate fraction (D) was active and underwent bioguided fractionation. All the isolated compounds were tested on P. falciparum FcB1 strain. In vitro anti-inflammatory activity (IL-1β, IL-6, IL-8, TNFα) of the ethyl acetate fraction and of an anti-Plasmodium active compound, was concurrently assessed on LPS-stimulated human PBMC and NO secretion inhibition was measured on LPS stimulated RAW murine macrophages. Cytotoxicity of the fractions and pure compounds was measured on VERO cells, L6 mammalian cells, PBMCs, and RAW cells. RESULTS Fractionation of the ethyl acetate soluble fraction (IC50 ranging from 3.4 to <1µg/mL depending on the parasite strain) led to the isolation of six pure compounds: catechin and five glycosylated quercetin derivatives. These compounds have never been isolated from this plant species. Two of these compounds (wayanin and guaijaverin) were found to be moderately active against P. falciparum FcB1 in vitro (IC50 5.5 and 6.9µM respectively). We proposed the name wayanin during public meetings organized in June 2015 in the Upper-Maroni villages, in homage to the medicinal knowledge of the Wayana population. At 50µg/mL, the ethyl acetate fraction (D) significantly inhibited IL-1β secretion (-46%) and NO production (-21%), as previously observed for the decoction. The effects of D and guiajaverin (4) on the secretion of other cytokines or NO production were not significant. CONCLUSIONS The confirmed antiplasmodial activity of the ethyl acetate soluble fraction of the decoction and of the isolated compounds support the previous results obtained on the P. acutangulum decoction. The antiplasmodial activity might be due to a mixture of moderately active non-toxic flavonoids. The anti-inflammatory activities were less marked for ethyl acetate fraction (D) than for the decoction.

Antileishmanial and Antitrypanosomal Activities of Flavonoids

This chapter focuses on the ability of flavonoid aglycones and glycosides to inhibit the growth of protozoan parasites of the genera Leishmania and Trypanosoma in different in vitro and in vivo models, namely pathogenic species for humans implicated in visceral (L. donovani, L. infantum) and cutaneous leishmaniasis (L. braziliensis, L. major, L. enriettii, L. mexicana), African sleeping sickness (T. brucei), and Chagas disease (T. cruzi). Several hundred naturally occurring flavonoids and their synthetic analogues were selected from a literature survey combining flavonoids exhibiting IC50 values smaller than 50 µM in one or more of the commonly used assays for antileishmanial or antitrypanosomal activity assessment. The trend of this survey indicates that the most active compounds against Leishmania spp. belong to chalcone, biflavone, and aurone classes, regardless of the performed assay. Few compounds of these classes exhibit submicromolar antileishmanial activity against axenic or intracellular amastigotes with no in vitro cytotoxicity. Flavones and flavonols are globally less active than chalcones, biflavones, and aurones. Flavones, flavan-3-ols, and isoflavones are less studied and less active. The most promising compounds against Trypanosoma brucei belong to synthetic chalcone, isoflavan, and isoflavanol classes, displaying submicromolar activities on trypomastigotes and selectivity indexes hundred times higher. On T. cruzi, flavan-3-ols are the most potent compounds, followed by flavones and flavonols. Globally, fewer compounds have been evaluated on T. cruzi than on T. brucei, and their activities and selectivity are often lower.

Substituted flavones: a promising scaffold in the fight against malaria

Background Ever since 2008, when evidence of artemisinin resistant malaria was highlighted in Western Cambodia [1], the need for new drugs with an original structure and novel mechanisms of action is even more pressing. The study of traditional remedies such as Cinchona bark or Artemisia aerial parts led to the discovery of the most potent antimalarials, bearing out that nature is still an incredible source of inspiration. Based on this approach, we are developing new synthetic antimalarial agents with an original structure inspired by nature.