Antimalarial activities of medicinal plants traditionally used in the villages of Dharmapuri regions of South India

Ethnopharmacological relevance

An ethnopharmacological investigation of medicinal plants traditionally used to treat diseases associated with fevers in Dharmapuri region of South India was undertaken. Twenty four plants were identified and evaluated for their in vitro activity against Plasmodium falciparum and assessed for cytotoxicity against HeLa cell line.

Aim of the study

This antimalarial in vitro study was planned to correlate and validate the traditional usage of medicinal plants against malaria.

Materials and methods

An ethnobotanical survey was made in Dharmapuri region, Tamil Nadu, India to identify plants used in traditional medicine against fevers. Selected plants were extracted with ethyl acetate and methanol and evaluated for antimalarial activity against erythrocytic stages of chloroquine (CQ)-sensitive 3D7 and CQ-resistant INDO strains of Plasmodium falciparum in culture using the fluorescence-based SYBR Green I assay. Cytotoxicity was determined against HeLa cells using MTT assay.

Results

Promising antiplasmodial activity was found in Aegle marmelos [leaf methanol extract (ME) (IC₅₀ = 7 μg/mL] and good activities were found in Lantana camara [leaf ethyl acetate extract (EAE) IC₅₀ = 19 μg/mL], Leucas aspera (flower EAE IC₅₀ = 12.5 μg/mL), Momordica charantia (leaf EAE IC₅₀ = 17.5 μg/mL), Phyllanthus amarus (leaf ME IC₅₀ = 15 μg/mL) and Piper nigrum (seed EAE IC₅₀ = 12.5 μg/mL). The leaf ME of Aegle marmelos which showed the highest activity against Plasmodium falciparum elicited low cytotoxicity (therapeutic index > 13).

Conclusion

These results provide validation for the traditional usage of some medicinal plants against malaria in Dharmapuri region, Tamil Nadu, India.     

Antiplasmodial activity of extracts from seven medicinal plants used in malaria treatment in Cameroon

Aim of the study

In a search for new plant-derived biologically active compounds against malaria parasites, we have carried out an ethnopharmacological study to evaluate the susceptibility of cultured Plasmodium falciparum to extracts and fractions from seven Cameroonian medicinal plants used in malaria treatment. We have also explored the inhibition of the Plasmodium falciparum cysteine protease Falcipain-2.

Materials and methods

Plant materials were extracted by maceration in organic solvents, and subsequently partitioned or fractionated to afford test fractions. The susceptibility of erythrocytes and the W2 strain of Plasmodium falciparum to plant extracts was evaluated in culture. In addition, the ability of annonaceous extracts to inhibit recombinant cysteine protease Falcipain-2 was also assessed.

Results and discussion

The extracts showed no toxicity against erythrocytes. The majority of plant extracts were highly active against Plasmodium falciparumin vitro, with IC₅₀ values lower than 5 μg/ml. Annonaceous extracts (acetogenin-rich fractions and interface precipitates) exhibited the highest potency. Some of these extracts exhibited modest inhibition of Falcipain-2.

Conclusion

These results support continued investigation of components of traditional medicines as potential new antimalarial agents.     

Antimalarial compounds from the aerial parts of Flacourtia indica (Flacourtiaceae)

Aim of the study

In the Comoros Islands, the aerial parts of Flacourtia indica are used in traditional medicine to treat malaria. Because of the important use of this plant, the phytochemistry of the aerial parts was investigated.

Materials and methods

Three compounds were isolated from the decoction of this plant material, Pyrocatechol, Homaloside D and Poliothrysoside. The in vitro antiplasmodial activity on the chloroquine-resistant strain (W2) of Plasmodium falciparum and the cytotoxicity on two complementary human cell lines (THP1, HepG2), of AcOEt extract obtained after liquid/liquid extraction of the decoction and pure compounds, were evaluated.

Results

The Poliothrysoside isolated from the AcOEt extract presented a strong antiplasmodial activity (IC₅₀ = 7.4 μM) and a good selectivity index (>28) similar to chloroquine.

Conclusion

This study reports for the first time antiplasmodial activity for Flacourtia indica, for its AcOEt extract and the three major constituents and confirms its traditional use.     

Evaluation of 13 selected medicinal plants from Burkina Faso for their antiplasmodial properties

Aim of the study

The aim of this study was to evaluate the antiplasmodial properties of 13 plants used against malaria in traditional medicine in Burkina Faso.

Materials and methods

In vitro antiplasmodial activity of dichloromethane, methanol and aqueous crude extracts obtained from vegetal samples collected in Burkina Faso was first evaluated on the Plasmodium falciparum 3D7 chloroquine-sensitive strain using a colorimetric method.

Results

Thirteen extracts obtained from 8 different species were found to exhibit antiplasmodial activity (IC₅₀ < 50 μg/ml). Five species demonstrated a moderate activity (15 μg/ml < IC₅₀ < 50 μg/ml): Boswellia dalzielii (leaves), Waltheria indica (roots and aerial parts), Bergia suffruticosa (whole plant), Vitellaria paradoxa (bark) and Jatropha gossypiifolia (leaves). The best results were obtained with extracts from the Dicoma tomentosa whole plant, from Psorospermum senegalense leaves and from Gardenia sokotensis leaves. These extracts found to display promising antiplasmodial activity, with IC₅₀ values ranging from 7.0 to 14.0 μg/ml.The most active plant extracts were then tested for in vitro activity on the Plasmodium falciparum W2 chloroquine-resistant strain and also for in vitro cytotoxicity on normal human fibroblasts (WI-38) in order to determine the selectivity index.

Conclusions

Dicoma tomentosa (Asteraceae) and Psorospermum senegalense (Clusiaceae) appeared to be the best candidates for further investigation of their antiplasmodial properties, reported for the first time by this study.     

In vitro antiplasmodial activity of plants used in Benin in traditional medicine to treat malaria

Aim of the study

The aim of the study was to evaluate the in vitro antiplasmodial activity of crude extracts of 12 plant species traditionally used in Benin for the treatment of malaria in order to validate their use.

Materials and methods

For each species, dichloromethane, methanol and total aqueous extracts were tested. The antiplasmodial activity of extracts was evaluated using the measurement of the plasmodial lactate dehydrogenase activity on chloroquine-sensitive (3D7) and resistant (W2) strains of Plasmodium falciparum. The selectivity of the different extracts was evaluated using the MTT test on J774 macrophage-like murine cells and WI38 human normal fibroblasts.

Results

The best growth inhibition of both strains of Plasmodium falciparum was observed with the dichloromethane extracts of Acanthospermum hispidum DC. (Asteraceae) (IC₅₀ = 7.5 μg/ml on 3D7 and 4.8 μg/ml on W2), Keetia leucantha (K. Krause) Bridson (syn. Plectronia leucantha Krause) (Rubiaceae) leaves and twigs (IC₅₀ = 13.8 and 11.3 μg/ml on 3D7 and IC₅₀ = 26.5 and 15.8 μg/ml on W2, respectively), Carpolobia lutea G.Don. (Polygalaceae) (IC₅₀ = 19.4 μg/ml on 3D7 and 8.1 μg/ml on W2) and Strychnos spinosa Lam. (Loganiaceae) leaves (IC₅₀ = 15.6 μg/ml on 3D7 and 8.9 μg/ml on W2). All these extracts had a low cytotoxicity.

Conclusion

Our study gives some justifications for the traditional uses of some investigated plants.     

Investigation of plant extracts in traditional medicine of the Brazilian Cerrado against protozoans and yeasts

Aim of the study

To investigate the activities of the 217 plant extracts in traditional medicine of the Brazilian Cerrado against protozoans and yeasts.

Materials and methods

Plant extracts were prepared by the method of maceration using solvents of different polarities. The growth inhibition of chloroquine-resistant Plasmodium falciparum strain (FcB1) was determined by measuring the radioactivity of the tritiated hypoxanthine incorporated. Activity against Leishmania (Leishmania) chagasi and Trypanosoma cruzi was measured by the MTT colorimetric assay. The antifungal tests were carried out by using the CLSI method. The active extracts were tested also by cytotoxicity assay using NIH-3T3 cells of mammalian fibroblasts.

Results

Two hundred and seventeen extracts of plants were tested against Plasmodium falciparum. The eleven active extracts, belonging to eight plant species were evaluated against L. (L.) chagasi, Trypanosoma cruzi, yeasts and in NIH-3T3 cells. The results found in these biological models are consistent with the ethnopharmacological data of these plants. The ethyl acetate extract of Diospyros hispida root showed IC₅₀ values of 1 μg/mL against Plasmodium falciparum. This extract demonstrated no toxicity against mammalian cells, resulting in a significant selectivity index (SI) of 435.8. The dichloromethane extract of Calophyllum brasiliense root wood was active against Cryptococcus gattii LMGO 01 with MIC of 1.95 μg/mL; and Candida albicans ATCC 10231 and Candida krusei LMGO 174, both with MIC of 7.81 μg/mL. The same extract was also active against Plasmodium falciparum and L. (L.) chagasi with IC₅₀ of 6.7 and 27.6 μg/mL respectively. The ethyl acetate extract of Spiranthera odoratissima leaves was active against Cryptococcus gattii LMGO 01 with MIC of 31.25 μg/mL, and against Plasmodium falciparum with IC₅₀ of 9.2 μg/mL and Trypanosoma cruzi with IC₅₀ of 56.3 μg/mL.

Conclusion

The active extracts for protozoans and human pathogenic yeasts are considered promising to continue the search for the identification and development of leading compounds.     

Effects of Phyllanthus urinaria extract on HepG2 cell viability and oxidative phosphorylation by isolated rat liver mitochondria

Ethnopharmacological relevance

Phyllanthus urinaria is widely used as anti-inflammatory, anti-diarrheal and hepatoprotective medicines in Asian countries such as India, China and Thailand. In Thailand, Phyllanthus urinaria is traditionally used as an adjuvant or alternative medicine for cancer patients, including liver cancer. However, there is limited scientific evidence supporting its use in cancer particularly hepatocellular carcinoma.

Aim of the study

To investigate the cytotoxic effect of Phyllanthus urinaria extract on human hepatocellular carcinoma HepG2 cells and the effect on oxidative phosphorylation by isolated rat liver mitochondria.

Materials and methods

HepG2 cells and isolated rat liver mitochondria were treated with the 50% methanolic extract of Phyllanthus urinaria. Cytotoxicity of the extract was assessed by trypan blue exclusion and MTT assay. Rates of oxygen consumption of isolated mitochondria were determined with a Clark oxygen electrode.

Results

It was found that the hydromethanolic extract induced cell death of HepG2 cells in a dose-dependent fashion. The IC₅₀ of Phyllanthus urinaria extract measured by trypan blue exclusion and MTT assay were 431 ± 65 μg/ml and 445 ± 62 μg/ml, respectively. Morphological changes of the cells were also observed. With isolated rat liver mitochondria, the extract slightly stimulated mitochondrial state 4 respiration but profoundly depressed state 3 respiration and respiratory control ratio.

Conclusions

The extract impairs energy metabolism by acting as inhibitor of oxidative phosphorylation and weak mitochondrial uncoupler. These mitochondrial effects may play a role in the cytotoxic action of Phyllanthus urinaria extract on HepG2 cells. These results provide preliminary experimental evidence supporting the use of Phyllanthus urinaria against hepatocellular carcinoma and open the possibility of considering this plant an adjunctive medicine for the treatment of this deadly disease.     

Prophylactic and curative activities of extracts from Warburgia ugandensis Sprague (Canellaceae) and Zanthoxylum usambarense (Engl.) Kokwaro (Rutaceae) against Plasmodium knowlesi and Plasmodium berghei

Ethnopharmacological relevance

This work reports the anti-plasmodial activities of Warburgia ugandensis and Zanthoxylum usambarense commonly used as phytomedicines against malaria by some Kenyan communities.

Aim of study

To determine the anti-plasmodial activities of extracts from Warburgia ugandensis and Zanthoxylum usambarense against Plasmodium knowlesi and Plasmodium berghei.

Materials and methods

Eight plant extracts were screened for in vitro anti-plasmodial activity against Plasmodium knowlesi, in a 96-well plate incubated at 37 °C on a RPMI culture medium supplemented with baboon serum. Of the eight, three were investigated for prophylactic and curative activities in BALB/c mice against drug-sensitive Plasmodium berghei in a 4-day test at a dose rate of 200 mg/kg/day.

Results

Inhibitory concentrations (IC₅₀) values of between 3.14 and 75 μg/ml, up to 69% chemosuppression of parasites growth and over 80% survivorship of treated mice were observed.

Conclusion

The two medicinal plants, Warburgia ugandensis and Zanthoxylum usambarense possess bioactive compounds against malaria parasites and could be exploited for further development into malaria therapy.     

In vitro antiplasmodial activity and cytotoxicity of nine plants traditionally used in Gabon

Aim of the study

As part of a project to identify new compounds active on malarial parasites, we tested the in vitro antiplasmodial activity of nine plants traditionally used to treat malaria symptoms in Haut-Ogooué Province, South-East Gabon.

Materials and methods

Dichloromethane and methanolic extracts of each plant were tested for their antiplasmodial activity on two chloroquine-resistant strains of Plasmodium falciparum (FCB and W2), based on lactate dehydrogenase activity. Cytotoxicity was assessed with the MTT test on MRC-5 human diploid embryonic lung cells.

Results

The methanolic extract of Staudtia gabonensis and the dichloromethane extract of Adhatoda latibracteata showed high antiplasmodial activity (IC₅₀ < 1 μg/ml) and low cytotoxicity, with selectivity indexes of about 58.25 and 16.43, respectively. The methanolic extract of Monodora myristica and the dichloromethane extract of Afromomum giganteum also showed promising activity (1 < IC₅₀ < 10 μg/ml) and low cytotoxicity, with selectivity indexes about 15.70 and 12.48, respectively. Dichloromethane extracts of Monodora myristica and Leonotis Africana showed moderate activity (10 < IC₅₀ < 40 μg/ml), with selectivity indexes about 6.07 and 28.89, respectively. Both extracts of Culcasia lancifolia had IC₅₀ values of 10-40 μg/ml but high cytotoxicity (selectivity indexes <2.77). The methanolic extract of Dorstenia klaineana had moderate antiplasmodial activity (IC₅₀ around 17 μg/ml) but strong cytotoxicity (0.43 μg/ml), giving a selectivity index of about 0.03.

Conclusions

Most extracts of nine selected plants traditionally used to treat malaria in Gabon had interesting antiplasmodial activity in vitro. This supports continued investigations of traditional medicines in the search for new antimalarial agents. The compounds responsible for the observed antiplasmodial effects are under investigation.     

In vivo antimalarial activity of Keetia leucantha twigs extracts and in vitro antiplasmodial effect of their constituents

Ethnopharmacological relevance

The West African tree Keetia leucantha (Rubiaceae) is used in traditional medicine in Benin to treat malaria. The twigs dichloromethane extract was previously shown to inhibit in vitro Plasmodium falciparum growth with no cytotoxicity (>100 µg/ml on human normal fibroblasts).

Materials and methods

The dichloromethane and aqueous extracts of twigs of K. leucantha were evaluated in vivo against Plasmodium berghei NK 173 by the 4-day suppressive test and in vitro against a chloroquine-sensitive strain of Plasmodium falciparum (3D7) using the measurement of the plasmodial lactate dehydrogenase activity. Bioguided fractionations were realized and compounds were structurally elucidated using extensive spectroscopic analysis.

Results

The in vivo antimalarial activity of K. leucantha dichloromethane and aqueous twigs extracts were assessed in mice at the dose of 200 mg/kg/day. Both extracts exhibited significant effect in inhibiting parasite growth by 56.8% and 53.0% (p<0.0001) on day 7-postinfection. An LC–MS analysis and bioguided fractionations on the twigs dichloromethane extract led to the isolation and structural determination of scopoletin (1), stigmasterol (2), three phenolic compounds: vanillin (3), hydroxybenzaldehyde (4) and ferulaldehyde (5), eight triterpenic esters (6–13), oleanolic acid and ursolic acid. The antiplasmodial activity of the mixture of the eight triterpenic esters showed an antiplasmodial activity of 1.66±0.54 µg/ml on the 3D7 strain, and the same range of activity was observed for isolated isomers mixtures.

Conclusions

This is the first report on the in vivo activity of K. leucantha extracts, the isolation of thirteen compounds and analysis of their antiplasmodial activity. The results obtained may partially justify the traditional use of K. leucantha to treat malaria in Benin.     

Antimalarial and safety evaluation of extracts from Toddalia asiatica (L) Lam. (Rutaceae)

Ethnopharmacological relevance

Toddalia asiatica (L) Lam. (Rutaceae) is a medicinal plant traditionally used in Kenya by many communities for the treatment of malaria and other ailments. All parts of the plant are claimed to have medicinal value, but the root bark in particular is believed to be more potent. Decoctions or infusions of the roots are taken orally to treat malaria, fever and stomach ache.

Aim of the study

To evaluate antimalarial activity of aqueous and organic extracts prepared from Toddalia asiatica and determine in vitro and in vivo safety of the extracts.

Materials and methods

Aqueous, ethyl acetate, hexane and methanol extracts were obtained from Toddalia asiatica root bark, fruits and leaves. In vitro antiplasmodial activity was done using chloroquine-sensitive (D6) and chloroquine-resistant (W2) Plasmodium falciparum strains and the concentration causing 50% inhibition of radioisotope incorporation (IC₅₀) was determined. In vivo assay was done by administering mice infected with Plasmodium berghei four consecutive daily doses of the extracts through oral route following Peters 4-Day suppressive test. The percentage suppression of parasitaemia was calculated for each dose level by comparing the parasitaemia in untreated control with those of treated mice. Quinine hydrochloride was used as positive control while double distilled water or 20% Tween-80 was used as a negative control. In vivo acute toxicity was determined in mice using standard procedures. In vitro cytotoxicity assay was carried out using actively dividing sub-confluent Vero cells.

Results

Inhibitory concentrations of ethyl acetate extract of Toddalia asiatica fruits showed high activity against chloroquine resistant (W2) strains of Plasmodium falciparum (IC₅₀=1.87 μg/ml), followed by root bark aqueous extract (IC₅₀=2.43 μg/ml). Tested in vivo against Plasmodium berghei, the fruit ethyl acetate extract (500 mg/kg) and root bark aqueous extract (250 mg/kg) reduced malaria parasitaemia by 81.34% and 56.8% respectively. Higher doses were found to be less effective in vivo. Acute toxicity and cytotoxictiy of the tested extracts, with the exception of hexane extract from the roots, showed LD₅₀>1000 mg/kg and CC₅₀>100 μg/ml respectively.

Conclusions

The results obtained contribute to the validation of traditional use of Toddalia asiatica and provides in vivo and safety data of the plant extracts tested for the first time. Ethyl acetate extract of the fruits was active against chloroquine resistant Plasmodium falciparum as well as against Plasmodium berghei. These findings confirm the suitability of Toddalia asiatica as a good candidate for further tests to obtain a prototype for antimalarial medicine.     

In vitro and in vivo antimalarial and cytotoxic activity of five plants used in congolese traditional medicine

Aim of the study

The in vitro antiplasmodial activity and cytotoxicity of methanolic and dichloromethane extracts from five Congolese plants were evaluated. The plants were selected following an ethnobotanical survey conducted in D.R. Congo and focusing on plants used traditionally to treat malaria. The in vivo antimalarial activity of aqueous and methanolic extracts active in vitro was also determined in mice infected by Plasmodium berghei berghei.

Materials and methods

The growth inhibition of Plasmodium falciparum strains was evaluated using the measurement of lactate dehydrogenase activity. The extracts (aqueous, CH₃OH, EtOH and CH₂Cl₂) were prepared by maceration and tested in vitro against the 3D7 (chloroquine sensitive) and W2 (chloroquine resistant) strains of Plasmodium falciparum and against the human normal fetal lung fibroblasts WI-38 to determine the selectivity index. Some extracts were also used at the dose of 300 mg/kg to evaluate their activity in mice infected since 4 days by Plasmodium berghei.

Results

Two plants presented a very high activity (IC₅₀ < 3 μg/ml). These plants were Strychnos icaja roots bark (MeOH and CH₂Cl₂) and Physalis angulata leaves (MeOH and CH₂Cl₂). One plant (Anisopappus chinensis whole plant, MeOH and CH₂Cl₂) presented a high activity (IC50 < 15 μg/ml). The extracts of Anisopappus chinensis and Physalis angulata showed also a good inhibition of parasitemia in vivo. Flavonoids, phenolic acids and terpenes were identified in these plants by a general phytochemical screening method.

Conclusion

Three plants showed a very interesting antiplasmodial activity (Anisopappus chinensis, Physalis angulata and Strychnos icaja) and one of them showed a good selectivity index (>10, Anisopappus chinensis). Anisopappus chinensis and Physalis angulata were also active in vivo.     

In vitro xanthine oxidase inhibitory activity of the fractions of Erythrina stricta Roxb.

Aim of the study

To assay the in vitro xanthine oxidase inhibitory activity of the various fractions of the hydromethanolic extract of the leaves of Erythrina stricta and to determine its enzyme inhibition mechanism.

Materials and methods

Xanthine oxidase inhibitory activity was assayed spectrophotometrically under aerobic conditions and the degree of enzyme inhibition was determined by measuring the increase in absorbance at 295 nm associated with uric acid formation. Enzyme kinetics was carried out using Lineweaver-Burk plots using xanthine as the substrate.

Results

Among the fractions tested, the chloroform fraction exhibited highest potency (IC₅₀ 21.2 ± 1.6 μg/ml) followed by the pet–ether (IC₅₀ 30.2 ± 2.2 μg/ml), ethyl acetate (IC₅₀ 44.9 ± 1.4 μg/ml) and residual (IC₅₀ 100 ± 3.3 μg/ml) fractions. The IC₅₀ value of allopurinol used, as the standard was 6.1 ± 0.3 μg/ml. Enzyme inhibition mechanism indicated that the mode of inhibition was of a mixed type.

Conclusion

These results suggest that the use of Erythrina stricta for the treatment of gout could be attributed to its xanthine oxidase inhibitory activity.     

New antiplasmodial alkaloids from Stephania rotunda

Ethnopharmacological relevance

Stephania rotunda Lour. (Menispermaceae) is a creeper growing in many countries of Asia and commonly found in the mountainous areas of Cambodia. As a folk medicine, it has been mainly used for the treatment of fever and malaria. The pharmacological activity is mostly due to alkaloids. Thus the aim of this study is to isolate new bioactive alkaloids from Stephania rotunda and to evaluate their in vitro antiplasmodial activity.

Materials and methods

Alkaloids were isolated and identified from dichloromethane and aqueous extracts using a combination of flash chromatography, high performance liquid chromatography, mass spectrometry and nuclear magnetic resonance. The purified compounds were tested for in vitro antiplasmodial activity on chloroquine-resistant W2 strain of Plasmodium falciparum.

Results

A new aporphine alkaloid named vireakine (2) along with two known alkaloids stephanine (1) and pseudopalmatine (8), described for the first time in Stephania rotunda, and together five known alkaloids tetrahydropalmatine (3), xylopinine (4), roemerine (5), cepharanthine (6) and palmatine (7) were isolated and identified. The structure of the new alkaloid was established on the basis of 1D and 2D NMR experiments and mass spectrometry. The compounds were evaluated for their in vitro antiplasmodial and cytotoxic activities. All tested compounds showed significant antiplasmodial activities with IC₅₀ ranged from 1.2 μM to 52.3 μM with a good selectivity index for pseudopalmatine with IC₅₀ of 2.8 μM against W2 strain of Plasmodium falciparum and IC₅₀>25 μM on K562S cells.

Conclusions

This study provides evidence to support the use of Stephania rotunda for the treatment of malaria and/or fever by the healers. Alkaloids of the tuber exhibited antiplasmodial activity and particularly cepharanthine and pseudopalmatine.     

Medicinal plants from the Yanesha (Peru): Evaluation of the leishmanicidal and antimalarial activity of selected extracts

Aim of the study

Ninety-four ethanolic extracts of plants used medicinally by the Yanesha, an Amazonian Peruvian ethnic group, for affections related to leishmaniasis and malaria were screened in vitro against Leishmania amazonensis amastigotes and against a Plasmodium falciparum chloroquine resistant strain.

Materials and methods

The viability of Leishmania amazonensis amastigote stages was assessed by the reduction of tetrazolium salt (MTT) while the impact on Plasmodium falciparum was determined by measuring the incorporation of radio-labelled hypoxanthine.

Results and conclusions

Six plant species displayed good activity against Plasmodium falciparum chloroquine resistant strain (IC₅₀ < 10 μg/ml): a Monimiaceae, Siparuna aspera (Ruiz & Pavon), A. DC., two Zingiberaceae, Renealmia thyrsoidea (Ruiz & Pavon) Poepp. & Endl. and Renealmia alpinia (Rottb.), two Piperaceae (Piper aduncum L. and Piper sp.) and the leaves of Jacaranda copaia (Aubl.) D. Don (Bignoniaceae).Eight species displayed interesting leishmanicidal activities (IC₅₀ < 10 μg/ml): Carica papaya L. (Caricaceae), Piper dennisii Trel (Piperaceae), Hedychium coronarium J. König (Zingiberaceae), Cestrum racemosum Ruiz & Pav. (Solanaceae), Renealmia alpinia (Rottb.) Zingiberaceae, Lantana sp. (Verbenaceae), Hyptis lacustris A. St.-Hil. ex Benth. (Lamiaceae) and Calea montana Klat. (Asteraceae). Most of them are used against skin affections by Yanesha people.Results are discussed herein, according to the traditional use of the plants and compared with data obtained from the literature.     

The bioactivity of novel furanoterpenoids isolated from Siphonochilus aethiopicus

Aim of the study

This study investigated the medicinal plant Siphonochilus aethiopicus (Zingiberaceae) for antiplasmodial activity.

Materials and methods

The ethyl acetate extract of Siphonochilus aethiopicus rhizomes was fractionated using solid phase extraction (SPE) and purified by high performance liquid chromatography. Structure elucidation was performed with nuclear magnetic resonance and mass spectrometry. The in vitro cytotoxicity and antiplasmodial activity was determined. In vivo schizontocidal activity was performed in a malaria mouse-model. Additional in vitro testing was done against Staphylococcus aureus, Klebsiella pneumoniae and Mycobacterium tuberculosis.

Results and discussion

The ethyl acetate extract showed in vitro activity against the chloroquine-sensitive (CQS) and chloroquine-resistant (CQR) strains of Plasmodium falciparum with IC₅₀-values of 2.9 μg/ml and 1.4 μg/ml, respectively. Bioassay-guided fractionation led to the isolation of three novel furanoterpenoids with moderate in vitro antiplasmodial activity. The crude extract showed very good in vivo activity. The compounds and crude extract were more active against the CQR strain than the CQS strain of Plasmodium falciparum. The SPE fractions were more active than the isolated compounds. The compounds did not show good activity against the micro-organisms tested. No in vitro cytotoxicity was observed.

Conclusion

This study provides evidence of antiplasmodial compounds present in Siphonochilus aethiopicus.     

Antiprotozoal screening and cytotoxicity of extracts and fractions from the leaves,stem bark and root bark of Alstonia congensis

Ethnopharmacological relevance

To evaluate the antiprotozoal activity and cytotoxicity of extracts and fractions from the leaves, root bark and stem bark of Alstonia congensis (Apocynaceae), used in traditional medicine against parasitic diseases.

Materials and methods

The aqueous and 80% MeOH extracts, and a series of fractions and subfractions from the leaves, stem and root bark of Alstonia congensis were tested in vitro for their antiprotozoal activity against Trypanosoma brucei brucei, Trypanosoma cruzi, Lesihamania infantum and the chloroquine and pyrimethamine-resistant K1 strain of Plasmodium falciparum. Their cytotoxicity on MRC-5 cells (human lung fibroblasts) was evaluated as well.

Results

The aqueous and 80% MeOH extracts and a series of subfractions of each plant part exhibited pronounced antiprotozoal activity against the K1 strain of Plasmodium falciparum with IC₅₀ values ranging from 2 to 5 µg/ml, and good activity against Trypanosoma brucei brucei and Trypanosoma cruzi with IC₅₀ values ranging between 5 and 10 µg/ml. The residual 80% MeOH extract from the leaves, and the total alkaloid extract from stem and root bark were the only subfractions active against Leishmania infantum with IC₅₀ values <10 µg/ml. None of the samples from the root bark was cytotoxic against MRC-cell lines (CC₅₀>64 µg/ml). In general, the aqueous extract (traditional decoction) showed the highest selectivity, especially against Plasmodium falciparum.

Conclusion

These results can partly support and justify the traditional use of these plant parts of Alstonina congensis as raw materials for the preparation of traditional remedies to treat parasitic diseases such as malaria and trypanosomiasis.     

The antiprotozoal activity of methylated flavonoids from Ageratum conyzoides L.

Aim of the study

The dichloromethane extract prepared from aerial parts of Ageratum conyzoides L. (Asteraceae), a plant commonly used in folk medicine for a number of illnesses including sleeping sickness, was recently found to exhibit a prominent activity (IC₅₀ = 0.78 μg/mL) against bloodstream forms of Trypanosoma brucei rhodesiense, the etiologic agent of East African Human Trypanosomiasis (East African Sleeping Sickness). This extract also exhibited noticeable activities against Leishmania donovani (Kala-Azar, IC₅₀ = 3.4 μg/mL) as well as Plasmodium falciparum (Malaria tropica, IC₅₀ = 8.0 μg/mL). In the current study, we sought for potentially active constituents of Ageratum conyzoides.

Materials and methods

Extracts prepared with solvents of different polarity were tested for activity against the above mentioned parasites as well as against Trypanosoma cruzi (Chagas’ disease) and for cytotoxicity using established protocols. The dicholoromethane extract showed the highest level of activity and was chosen for phytochemical studies aimed at the isolation of potential active constituents.

Results and conclusion

Five highly methoxylated flavonoids along with the chromene derivative encecalol methyl ether were isolated. All isolated compounds were previously reported from Ageratum conyzoides. While the chromene turned out to be inactive against the tested parasites, the flavonoids showed activity against the protozoan pathogens, some in the lower micromolar range. However, none of these isolated compounds was as active as the crude extract. This is the first report on antiprotozoal activity of this plant species and some of its constituents. The chemical principle accounting for the high activity of the crude extract, however, remains to be identified.     

In vitro antitrypanosomal and antiplasmodial activities of crude extracts and essential oils of Ocimum gratissimum Linn from Benin and influence of vegetative stage

Ethnopharmacological relevance

Different parts of Ocimum gratissimum Linn are largely used in folk medicine for the treatment of many diseases, some of which related to parasitical infections as fevers and headaches. In order to validate their use and to clarify the plant part which possesses the best antiparasitic properties, we decided to evaluate the in vitro antiplasmodial and antitrypanosomal activities of essential oils and crude extracts from leaves, stems and seeds of Ocimum gratissimum as well as their cytotoxicity.

Materials and methods

The essential oils and ethanol crude extracts of leaves and stems of Ocimum gratissimum from Benin, were obtained in pre and full flowering stages. Seeds obtained only in full flowering stage, were also extracted. The oils were isolated by hydrodistillation and analyzed by GC/MS and GC/FID. Extracts and essential oils were tested in vitro against Trypanosoma brucei brucei and Plasmodium falciparum. Cytotoxicity was evaluated in vitro against Chinese Hamster Ovary (CHO) cells and the human non cancer fibroblast cell line (WI38) through MTT assay to evaluate the selectivity and toxicity was assessed against Artemia salina Leach.

Results

The essential oils and non-volatile crude extracts of Ocimum gratissimum were more active on Trypanosoma brucei brucei than on Plasmodium falciparum (3D7). This activity varies according to the vegetative stage (pre and full flowering) and the plant part (seeds, stems and leaves) extracted. The best growth inhibition of Trypanosoma brucei brucei was observed with ethanol crude extracts of leaves (IC₅₀=1.66±0.48 μg/mL) and seeds (IC₅₀=1.29±0.42 μg/mL) in full flowering stage with good selectivity (SI>10). The chemical composition of the essential oil from aerial parts (47 compounds), characterized by the presence as main constituents of p-cymene, thymol, γ-terpinene, β-myrcene and α-thujene, depends on the vegetative stage. The oil contained some minor compounds such as myrcene (IC₅₀=2.24±0.27 μg/mL), citronellal (IC₅₀=2.76±1.55 μg/mL), limonene (IC₅₀=4.24±2.27 μg/mL), with good antitrypanosomal activities. These oils and crude extracts were not toxic against Artemia salina Leach and had a low cytotoxicity except leaves and seeds ethanol extracts obtained in full flowering which showed toxicity against CHO and WI38 cells.

Conclusions

Our study shows that ethanol crude extracts of leaves and seeds of Ocimum gratissimum in full flowering stage can be a good source of antitrypanosomal agents. This is the first report about the relation between the plant part extracted, the vegetative stage of the plant, the antitrypanosomal and antiplasmodial activities and the cytotoxicity of essential oils and non-volatile extracts of Ocimum gratissimum from Benin.     

Antiprotozoal activity of Khaya anthotheca, (Welv.) C.D.C. a plant used by chimpanzees for self-medication

Ethnopharmacological relevance

Khaya species, endemic to Africa and Madagascar, continues to be valuable in indigenous traditional medicine. Their bitter tasting barks are decocted to treat fevers, several febrile conditions, microbial infections and worm infestations. In the Budongo rain forest of Western Uganda, non-human primates, especially chimpanzees and baboons, have been observed to eat the bitter non-nutritious bark and occasionally the seed.

Materials and methods

Extracts were prepared by sequential fractionation with solvents of increasing polarities and assayed using standard procedures. Bioassay guided purification of the petroleum ether extract by column chromatography yielded three pure limonoids, Grandifolione (1), 7-deacetylkhivorin (2) and 1,3-deacetyldeoxyhavenensin (3). The antitrypanosomal, antileishmanial and antiplasmodial activities of pure compounds (1) and (2) were evaluated in vitro against Plasmodium falciparum K1, Trypanosoma brucei rhodesiense STIB 900, Trypanosoma cruzi trypomastigotes (Tulahuen C4), and axenic Leishmania donovani MHOMET-67/L82 and for cytotoxicity against L6 rat skeletal myoblast cells, in parallel with standard drugs.

Results

Of the four extracts tested, the petroleum ether extract showed activity against Plasmodium falciparum (IC₅₀ 0.955 μg/ml) and Trypanosoma brucei rhodesiense (IC₅₀ 5.72 μg/ml). The pure compounds (1) and (2) demonstrated activity against Plasmodium falciparum (KI strain) and marginal activities against Trypanosoma brucei brucei, Trypanosoma brucei rhodesiense, Trypanosoma cruzi and Leishmania donovani.

Conclusion

The present study provides evidence justifying the use of Khaya preparations in traditional medicine to treat fevers and microbial infections. The observed antiprotozoal activity of grandifolione and 7-deacetylkhivorin from the seed of Khaya anthotheca further confirms the ethnomedicinal potential of this plant and supports the hypothesis that non-human hominids (chimpanzees and baboons) too, eat the bitter bark and seeds for self-medication and in general, the use of Khaya plant material for medication by humans in disease endemic tropical areas. The antiprotozoal activity of gradifolione, and, the antitrypanosomal and antileishmanial activities of 7-deacetylkhivorin are reported here for the first time.