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Plants of the Strychnos genus, which include about 200 species, are used for multiple traditional purposes as hunting poison, for example, and have shown interesting pharmacological properties, especially curarizing and tetanizing, but also against malaria. Many monoterpene indole alkaloids have already been isolated and identified. Among them, there is strychnine, a famous alkaloid that can cause death by asphyxiation.Investigate alkaloidic molecular diversity from Strychnos genus using molecular networking technique and study the Strychnos genus from a chemotaxonomic point of view.Twenty-eight different species and different plant parts were ground into powder using a grinder. The methanolic extracts were carried out using a pressurized solvent extraction and the alkaloid extract was performed manually with a separating funnel. The extracts were analyzed by HPLC-ESI(+)-Q/TOF. The data were processed using MZmine 2 software and the molecular network was generated on the GNPS platform. The study of the generated molecular network allowed the detection of various alkaloids. Among these is the famous strychnine which has been detected in 7 new Strychnos species not yet described as strychnine producers. This identification was investigated using orthogonal approaches, namely TLC, NMR, HPLC-UV and UHPLC-ESI(+)-Q/TOF analyses. The LOD by HPLC-UV of strychnine was also determined.Further analyses allowed to confirm the presence of strychnine in S. densiflora trunk barks but also to show the presence of strychnine with high probability in the trunk barks of S. camptoneura, S. congolana, S. boonei, and S. tchibangensis, and in the leaves of S. usambarensis. About the trunk barks of S. tricalyisoides, the probability of a strychnine content remains low.This work exemplified the efficiency of molecular networking in identifying known metabolites (major and minor alkaloids) involved in the chemotaxonomic study of plants from Strychnos genus.

International audience; Annonaceous acetogenins are natural products held responsible for atypical parkinsonism due to chronic consumption in traditional medicine or as food. Due to their long-term neurotoxic effects in humans, the complete chemical characterization of acetogenins in complex mixtures has become crucial. Characterization by tandem mass spectrometry (MS/MS) of acetogenins using collisioninduced dissociation (CID) from lithium adducts provides additional structural information compared to protonated or sodiated species such as ketone location on the acetogenins backbone. However, diagnostic ions are present at a very low intensity limiting this approach to abundant species in the extracts. Copper adducts led to diagnostic fragment ions that allow to identify the position of oxygen rings and hydroxyl substituents. Fragmentation rules were established based on acetogenin standards allowing the identification of 45 over the 77 analogues observed in an extract of Annona muricata by LC-MS/MS using post-column infusion of copper sulfate (CuSO4) solution. Molecular networks that were generated thanks to specific fragmentations obtained with copper, led to the distinction of THF ring position or to the identification of hydroxylated lactone for instance, while networks generated from lithium adducts mainly distinguish analogues with a hydroxyl group at C4 from the other structures.

We herein report on the first chemical assessment of Erythrococca anomala (Juss. ex Poir.) Prain (Euphorbiaceae), a genus that was – to the best of our knowledge – not studied yet from a phytochemical perspective. A molecular networking strategy was implemented to rapidly identify the known specialized metabolites from untargeted MS/MS analyses of E. anomala leaves ethanolic extract. This strategy allowed for the identification of diverse C-glycosyl flavones and a cursory examination of MS/MS spectra could extend the GNPS-provided annotation to pinpoint the structural novelty of further derivatives. The isolation of the sought-after structures could be streamlined based on MS-guidance and their structures, determined through extensive NMR analyses, displayed structural features in line with MS²-based predictions. Anticipating sharp structural features at an early stage of the dereplication process through a critical assessment of the tandem mass spectrometric landmarks was essential to embark on the isolation of the newly reported structures owing to the elevated number of flavonoid glycosides isomers thereof formerly known, which would have deterred us from isolating them without the support of additional tandem mass spectrometric information. The isolation of the main components of the ethanolic extract completed the currently provided chemical report on E. anomala, also resulting in the description of a new phenylethanoid derivative (3) and of a new orcinol-based dimer (4). Anomaloflavone (1) exhibit significant activities with minimal inhibitory concentration values of 25 µg/mL against Staphylococcus aureus and Mycobacterium smegmatis while failing to exert an antibacterial activity against Pseudomonas aeruginosa, while being devoid of cytotoxicity against SiHa cells.

Melonine is a basic monoterpene indole alkaloid (MIA) skeleton from Melodinus philliraeoides that was reported in 1983. The scarcity of its spectroscopic data questioned the validity of its structure. This prompted us to reisolate this molecule and to revise its structure into an unprecedented MIA scaffold. DFT-validated biosynthetic paths to both this new core and the originally reported form are proposed. The pathway to the original structure of melonine seems to be thermodynamically feasible, and that compound may exist as a natural product.

Hemitheion (1), a new sulfur-containing vobasane-type indole alkaloid, was isolated, together with three known compounds, vobasine (2), gelsedine (3), and gelsemicine (4), from the alkaloid extract of the stems of Mostuea brunonis Didr. (Gelsemiaceae). Compound 1 could be straightforwardly isolated. Its structure was elucidated by a combination of spectroscopic methods. Besides corresponding to a formerly postulated biosynthetic intermediate toward theionbrunonines, hemitheion (1) stands among the few monomeric vobasanes lacking an oxygen at C-3. Hemitheion (1) showed moderate antiplasmodial activity in the micromolar range against the strain FcB1 of Plasmodium falciparum and no cytotoxic activity against the MRC-5 cell line at 20 μM.

In less than 10 years, molecular networking (MN) strategy has revolutionized the art of Natural Products (NP) isolation to enter a rational workflow greatly increasing the probabilities of isolating new chemical entities. To pinpoint and streamline the isolation of new Monoterpene Indole Alkaloids (MIAs) in producing plants, we rendered publicly available the MIA database (MIADB), comprising MS

Mareya micrantha (Euphorbiaceae) is an Ivorian medicinal plant widely used in gastrointestinal disorders. So as to understand and identify metabolites possibly implicated in the activity of the plant on intestinal muscles, this study has been undertaken. From the crude ethanolic extract EE of the leaves, hexanic (EH), chloromethylene (EC), ethyl acetatic (EA) and butanolic (EB) sub-extracts were prepared as well as the hydroethanolic residual phase dried (Ar) and their effects on the isolated duodenum of rabbits were determined. If the sub-extracts EH, EA and EB, but also Ar, confirmed the myostimulant activity of the crude extract EE, this was not the case of EC which showed myorelaxant activity. This sub-extract, subjected to purification, led to the isolation of three major triterpenoids type 29-norcucurbitacin (1-3) whose structures were confirmed using the combined analyses of their spectroscopic data (UV, MS, 1D NMR and 2D). All these molecules have shown myorelaxant activity, the most significant one being 2 with an IC50 value of 36.9 µM. These molecules would act by blocking both intracellular and extracellular calcium.

Bioassay guided fractionation of the stem barks of Isolona cooperi led to the isolation of a new lactone, apoprunellelactone (APL, 1), and two known compounds, 5-[1-hydroxyhexyl]-2H-furan-2-one (2) and oleic acid (3). Their structures were elucidated by spectral analysis including MS, UV, IR, 1D and 2D-NMR spectroscopy. Evaluated for its antiprotozoal activities, APL (1) was found to be the most active on Leishmania donovani and L. major promastigotes with EC50 values of 16.3 and 8.2 µM, respectively. Against Trypanosoma brucei brucei trypomastigote forms, the activity of APL was moderated (MEC = 38.0 µM). Its hemisynthetic ester acetic derivative (1c) was 2-42 times more active than that of the APL and reference drugs, justifying further in vivo evaluation of the two compounds (1 and 1c) on Leishmania sp and Trypanosoma brucei brucei/mice models.

The structure elucidation of theionbrunonine C, a thioether-bridged dimeric monoterpene indole alkaloid (MIA), and more generally, one of the very few sulfur-containing MIA, is reported after its isolation from Mostuea brunonis (Gelsemiaceae). This unstable structure had already been targeted for isolation in our former, molecular network-guided, investigation of this plant, but this compound had degraded before sufficient spectroscopic data could have been acquired for a complete structure assignment. With this constraint in mind, the rapid acquisition of nuclear magnetic resonance (NMR) data enabled retrieving sufficient spectroscopic information for full structure elucidation, although from a partial set of spectroscopic information (

Inaequalisines A and B (1 and 2), the first examples of hybrid alkylated phenylpropane monoterpene indole alkaloids, were isolated from the roots of Callichilia inaequalis, guided by the combined use of molecular networking and substructure annotation. Their structures, including absolute configuration, were elucidated by spectroscopic methods and ECD calculations. A possible biosynthetic pathway for 1 and 2 was postulated.

During our research to contribute to the elucidation of the chemical composition of the root bark of E. suaveolens, six non-nitrogenous cassane diterpenoids (1–6) were isolated and identified. Of these secondary metabolites, three have never been previously described: Cassan-13,15-dien-3-oxo-17-oic acid (2), Cassan-15-en-[7,17]-γ-lactone (3) and 6α-hydroxy-cassamic acid (5). The other are known but, never isolated from the root barks of E. suaveolens (Fabaceae): Cassan-13,15-dien-17-oic acid (1), 6α-hydroxy-cassamic acid methyl esther (4) and cassamic acid (6). Their structures were established according to the spectroscopic data (NMR 1D and 2D, HR-ESI-MS and IR), in comparison with those of literature. The originality of this study lies in the fact that three new natural molecules were isolated and identified. In addition, all the isolated compounds (1–6) were reported for the first time from the root barks of E. suaveolens.

Traditional natural products discovery workflows implying a combination of different targeting strategies including structure- and/or bioactivity-based approaches, afford no information about new compound structure until late in the discovery pipeline. By integrating a MS/MS prediction module and a collaborative library of (bio)chemical transformations, we have developed a new platform, coined MetWork, that is able of anticipating the structural identity of metabolites starting from any identified compound. In our quest to discover new monoterpene indole alkaloids, we demonstrate the utility of the MetWork platform by anticipating the structures of five previously undescribed sarpagine-like N-oxide alkaloids that have been targeted and isolated from the leaves of Alstonia balansae using a molecular networking-based dereplication strategy fueled by computer-generated annotations. This study constitutes the first example of non peptidic molecular networking-based natural product discovery workflow, in which the targeted structures were initially generated, and therefore anticipated by a computer prior to their isolation.

Hemitheion (

International audience; Four undescribed alkaloids have been isolated from the bulbs of the previously unstudied Crinum scillifolium. These compounds were targeted following a state-of-the-art molecular networking strategy comprising a dereplication against in silico databases and re-ranking of the candidate structures based on taxonomically informed scoring. The unreported structures span across a variety of Amaryllidaceae alkaloids appendages. Their structures were unambiguously elucidated by thorough interpretation of their HRESIMS and 1D and 2D NMR data, and comparison to literature data. DFT-NMR calculations were performed to support the determined relative configurations of scillitazettine and scilli-N-desmethylpretazettine and their absolute configurations were mitigated by comparison between experimental and theoretically calculated ECD spectra. The lack of a methyl group on the nitrogen atom in the structure of scilli-N-desmethylpretazettine series is highly unusual in the pretazettine/tazettine series but the most original structural feature in it lies in its 11α disposed hydrogen, which is new to pretazettines. The antiplasmodial as well as the cytotoxic activities against the human colon cancer cell line HCT116 were evaluated, revealing mild to null activities.

Five new cassane-type diterpenoid heterosides, i. e. two cassane-type amides (1-2), two erythrophlamine-type amine esters (3-4) and a non‑nitrogenous erythrophlamine analogue (5) were isolated from the root barks (1-2) and the seeds (3-5) of Erythrophleum suaveolens. Their structures were unambiguously established by interpretation of their HRESIMS, 1D and 2D NMR data, and chemical degradation for sugar determination. Compounds 3-5 were evaluated for their cytotoxicity against a panel of three cell lines, revealing modest to strong activities.

Bioassay guided fractionation of the stem barks of

Covering: up to 2019Landmark advances in bioinformatics tools have recently enhanced the field of natural products research, putting today's natural product chemists in the enviable position of being able to perform the efficient targeting/discovery of previously undescribed molecules by expediting the prioritization of the isolation workflow. Among these advances, MS/MS molecular networking has appeared as a promising approach to dereplicate complex natural product mixtures, leading to a real revolution in the "art of natural product isolation" by accelerating the pace of research of this field. This review illustrates through selected cornerstone studies the new thinking in natural product isolation by drawing a parallel between the different underlying philosophies behind the use of molecular networking in targeting natural products.

Introduction Acetogenins are plant polyketides known to be cytotoxic and proposed as antitumor candidates. They are also suspected to be alimentary neurotoxins. Their occurrence as complex mixtures renders their dereplication and structural identification difficult using liquid chromatography coupled to tandem mass spectrometry and efforts are required to improve the methodology. Objective To develop a supercritical fluid chromatography (SFC) high-resolution tandem mass spectrometry method, involving lithium post-column cationisation, for the structural characterisation of Annonaceous acetogenins in crude extracts. Methodology The seeds of Annona muricata L. were extracted with methanol. Supercritical fluid chromatography of the extract, using a 2-ethylpyridine stationary phase column, was monitored using a high-resolution quadrupole time-of-flight mass spectrometer. Lithium iodide was added post-column in the make-up solvent. For comparison, the same extract was analysed using high-pressure liquid chromatography coupled to the same mass spectrometer, with a column based on solid core particles. Results Sensitivity was similar for both HPLC and SFC approaches. Retention behaviour and fragmentation pathways of three different isomer groups are described. A previously unknown group of acetogenins was also evidenced for the first time. Conclusion The use of SFC-MS/MS allows the reduction of the time of analysis, of environmental impact and an increase in the chromatographic resolution, compared to liquid chromatography. This new methodology enlightened a new group of acetogenins, isomers of montanacin-D. Copyright © 2017 John Wiley & Sons, Ltd.

The data are related to the research article entitled ‘‘New cassane diterpenoids from the root bark of Erythrophleum suaveolens’’ (Jacques et al., 2019). The article provides method of purification and data to determine structure of two novel cassane diterpenoid amines: 3β-hydroxy-3-methylbutanoyloxy-6α-hydroxy-nor-cassamine (1) and 3β-hydroxy-3-methylbutanoyloxy-erythrosuamine (2). This data in brief provides IR, NMR and Q-TOF-MS spectra, along with fragmentation pathways that allowed to the identification of both new compounds. Keywords: Data, NMR, IR, Cassane diterpenoid

This Data Descriptor announces the submission to public repositories of the monoterpene indole alkaloid database (MIADB), a cumulative collection of 172 tandem mass spectrometry (MS/MS) spectra from multiple research projects conducted in eight natural product chemistry laboratories since the 1960s. All data have been annotated and organized to promote reuse by the community. Being a unique collection of these complex natural products, these data can be used to guide the dereplication and targeting of new related monoterpene indole alkaloids within complex mixtures when applying computer-based approaches, such as molecular networking. Each spectrum has its own accession number from CCMSLIB00004679916 to CCMSLIB00004680087 on the GNPS. The MIADB is available for download from MetaboLights under the identifier: MTBLS142 (https://www.ebi.ac.uk/metabolights/MTBLS142)., Design Type(s)mass spectrometry data transformation objective • mass spectrometry data analysis objective • data integration objectiveMeasurement Type(s)mass spectrumTechnology Type(s)liquid chromatography-tandem mass spectrometryFactor Type(s)Sample Characteristic(s)Strychnos usambarensis • Picralima nitida • Geissospermum laeve • Pleiocarpa mutica • Alstonia • Callichilia inaequalis • Chimarris cymosa • Mostuea brunonis • Gonioma • Cinchona • Catharanthus roseus • Voacanga grandifolia Machine-accessible metadata file describing the reported data (ISA-Tab format)

Introduction Annonaceous acetogenins (AAGs) constitute a group of environmental neurotoxins, possibly implicated in sporadic atypical Parkinsonism/dementia complexes. The recent evidencing of complex mixtures of AAGs in edible fruits and derived food products requires efficient and practical analytical tools for an estimation of human exposure. Objective To develop a simple method for the direct quantitation of the majority of AAGs (sub-types 1a and 1b) within crude extracts, using commonly available 1H–NMR spectrometers, for food control. Methodology Method development was carried out on 400 MHz and 300 MHz spectrometers, for routine application on fruits crude extracts of Annona muricata L. The method was validated with annonacin and squamocin as reference compounds. Two internal standards (ISs), fumaric acid and dimethyl fumarate, were successfully used, in deuterated methanol (CD3OD) and deuterated chloroform (CDCl3), respectively. Results Quantitation was carried out using signals corresponding to the deshielded ethylenic protons characterising most AAGs, at δ 7.18 or δ 6.98 ppm in CDCl3. The limit of quantification (LOQ) was 2.5 mM, with acceptable accuracy, and the limit of detection (LOD) was 0.5 mM. The AAGs contents measured in seven distinct fruit samples of Annona muricata ranged from 14 μmol to 226 μmol of AAGs per 100 g fresh pulp (i.e. 0.14 mmol to 1.3 mmol of AAGs per fruit). Conclusion A simple, accurate and specific method for quantification of AAGs content was developed and validated for routine application to fruit pulp crude extracts. Copyright © 2017 John Wiley & Sons, Ltd.

International audience; The Annonaceous acetogenin annonacin is an environmental neurotoxin identified in the pulp of several fruits of the Annonaceae family, whose consumption was linked to the occurrence of sporadic atypical Parkinsonism with dementia. A method for its quantification in Rat brain homogenates by UPLC-MS/MS in selected reaction monitoring (SRM) mode was developed and validated. This method was applied to the quantitation of annonacin in Rat brain after intravenous (0.5 mg/kg) and oral (10 mg/kg, 100 mg/kg) administration.

The use of medicinal plants for the treatment of diseases including malaria is commonplace in Ghanaian traditional medicine, though the therapeutic claims for most plants remain unvalidated. Antiplasmodial activity of the aqueous extracts and successively obtained petroleum ether, ethyl acetate and methanol fractions of the whole Phyllanthus fraternus plant, the leaves of Tectona grandis, Terminalia ivorensis and Bambusa vulgaris, and roots of Senna siamea were studied against Plasmodium falciparum chloroquine-sensitive 3D7 and chloroquine-resistant W2 strains. The aqueous extracts were assessed against human umbilical vein endothelial cells (HUVECs) for cytotoxicity, and the organic solvent fractions against human O(+) erythrocytes for haemolytic effect. Both extracts and fractions demonstrated antiplasmodial activity to varied extents. The aqueous extract of T. ivorensis was the most active (3D7, IC50 0.64 ± 0.14; and W2, IC50 10.52 ± 3.55 μg/mL), and together with P. fraternus displayed cytotoxicity (CC50 6.25 ± 0.40 and 31.11 ± 3.31 μg/mL, respectively). The aqueous extracts were generally selective for 3D7 strain of P. falciparum (selectivity indexes (SIs) ≥3.48) but only that of S. siamea was selective for the W2 strain (SI > 2.1). The organic solvent fractions also displayed antiplasmodial activity with the methanol fractions of P. fraternus and T. grandis, and the fractions of B. vulgaris showing activity with IC50 below 1 μg/mL against P. falciparum 3D7 strain; some fractions showed haemolytic effect but with low to high selectivity indexes (SI ≥ 4). The results while justifying the traditional use of the plant materials in the treatment of malaria, however, suggest their cautious use.

The chemical investigation of the antimalarial plant Phyllanthus fraternus G. L. Webster (Phyllanthaceae) resulted in the discovery of the Securinega alkaloid (+)-allonorsecurinine ( 1 ), previously reported as a synthetic compound, together with the known ent -norsecurinine ( 2 ), nirurine ( 3 ), bubbialine ( 4 ), epibubbialine ( 5 ) and the lignan phyllanthin ( 6 ). The structure and absolute configuration of the new compound were elucidated on the basis of extensive spectroscopic analysis, optical rotation, and GIAO NMR shift calculation followed by CP3 analysis. The antiplasmodial activity of these compounds was evaluated against chloroquine-resistant (W2) and -sensitive (3D7) strains of Plasmodium falciparum . Among them, ent -norsecurinine ( 2 ) and (+)-allonorsecurinine ( 1 ) showed the strongest activity (IC 50 : 1.14 ± 0.32 and 2.57 ± 0.53 µM) respectively, against W2 but one of the weakest against 3D7.

Theionbrunonines A and B (1 and 2), the first examples of monoterpene bisindole alkaloids linked by a thioether bridge, were isolated from the stems of Mostuea brunonis, guided by a molecular networking-based dereplication strategy. Their structures were elucidated by a combination of spectroscopic data and ECD calculations. A plausible biosynthetic pathway for 1 and 2 was postulated. Theionbrunonines A and B (1 and 2) showed moderate antiplasmodial activities in the micromolar range against the strain FcB1 of Plasmodium falciparum and no cytotoxic activity against the MRC-5 cell line at 20 μM.

The two new acylphloroglucinol derivatives, methylene-bis-aspidinol AB (1) and mallopposinol (2), together with the nine known compounds, aspidinol B (3), methylene-bis-aspidinol (4), (+)-α-tocopherol (5), lupeol (6), stigmasterol (7), phytol (8), bergenin (9), squalene (11) and methyl gallate (10) were isolated from the leaves of Mallotus oppositifolius. Their structures were elucidated by spectral analysis including MS, 1D and 2D-NMR spectroscopy. In vitro trypanocidal and antileishmanial activities of compounds 1-9 were evaluated. Mallopposinol (2) and aspidinol B (3) displayed weak antileishmanial activities against Leishmania donovani promastigotes, with EC50 values of 21.3 and 38.8μM, respectively. Only the methylene-bis-aspidinol (4) exhibited trypanocidal activity against Trypanosoma brucei brucei trypomastigotes (LC100=0.8μM) similar to the reference drug pentamidine (LC100=0.4μM).

Phytochemical investigation of the stem bark of Erythrophleum suaveolens yielded four new cassane diterpenoids; 6α-hydroxy-cassamic acid, methyl ester, 4β-carbomethoxy-14-methyltotarol, 6α-hydroxy- nor -cassamine, and 8,9-dehydro- nor -cassamine, along with four known cassane diterpenoids. All structures were elucidated on the basis of one- and two-dimensional NMR, HRMS and ESIMS analysis.

International audience; Three new monoterpene indole alkaloids (13) have been isolated from the bark of Geissospermum laeve-, together with the known alkaloids (-)-leuconolam (4), geissolosimine (5), and geissospermine (6). The structures of 1-3 were elucidated by analysis of their HRMS and NMR spectroscopic data. The absolute configuration of geissolaevine (I). was deduced from the comparison of experimental and theoretically calculated ECD spectra. The isolation workflow was guided by a molecular networking-based dereplication strategy using an in-house database of monoterpene indole alkaloids. In addition, five known compounds previously undescribed in the Geissospermum genus were dereplicated from the G. laeve alkaloid extract network and were assigned with various levels of identification confidence. The antiparasitic activities against Plasmodium falciparum and Leishmania donovani as well as the Ortotoxic activity against the MRC-5 cell line were determined for compounds 1-5.

Four new cassane diterpenoid amides, 1–4 and six known cassane diterpenoid amides have been isolated from the stem bark of Erythrophleum suaveolens. The isolated compounds were subjected to phytochemical investigation. The structures of those compounds were elucidated on the basis of one- and two-dimensional NMR, HRMS and ESIMS analyses. Nor-cassamide was obtained as crystal and characterized by X-ray method. This was the first crystallographic description of cassane diterpenoid and it allowed to confirm the absolute configuration of the stereogenic centers and the stereochemistry of the double exocyclic bond.

International audience; Epidemiological and toxicological studies have suggested Annonaceaeous acetogenins to be environmental neurotoxins responsible for sporadic atypical parkinsonism/dementia in tropical areas. These compounds are present in the tropical genus Annona (Annonaceae), known for its fruit-yielding cultivated species such as Annona cherimolia. This species is widely cultivated in South America, Spain, and Portugal and yields acetogenins in its seeds, stems, and roots. The presence of these compounds in the pulp of its fruit and in derived food products is unclear. An innovative and sensitive methodology by HPLC-ESI-LTQ-Orbitrap with postcolumn infusion of lithium iodide was used to identify the presence of low levels of acetogenins in an A. cherimolia Mill. fruit-based commercial alcoholic beverage. More than 80 representatives were detected, and the 31 most intense acetogenins were identified. All together these findings indicate that this species should be considered as a risk factor within the framework of a worldwide problem of food toxicity.

Cladodes and fruits of Opuntia ficus-indica are used in traditional medicine for the treatment of abscess and skin inflammation. It was therefore interesting to assess whether an antileishmanial activity could be associated to skin healing. This study reports on the antileishmanial activity of Opuntia ficus-indica extracts from cladodes and fruits. Ethyl acetate extract from cladodes only exhibited an activity against Leishmania major with an IC50 value of 53.9 μg/mL, but ethyl acetate fruit extract, ethyl acetate cladode extract and methanol cladode extract were active also against Leishmania donovani with IC50 values at 70.3, 70.5 and 45.2 μg/mL, respectively. A poor activity of the fractions was monitored against Trypanosoma brucei brucei. Finally, a bioguided fractionation of fruits of Opuntia ficus-indica led to a pre-purified fraction that exhibited an IC50 of 9.3 μg/mL against Leishmania donovani intramacrophage amastigotes. The selectivity index defined as CC50/IC50 was higher than 10. In conclusion, the bioguided fractionation allowed to enhance the antileishmanial activity about ten-fold comparatively to those of the ethyl acetate fruit extract. Such an activity is worth of further investigations to identify the compounds responsible for the antileishmanial effect.