In vivo antimalarial activities of Enantia polycarpa stem bark against Plasmodium berghei berghei in mice

Ethnopharmacological relevance

Enantia polycarpa (PC) Engl. Et Diels (Annonaceae) is used in traditional medicine as an antimalarial remedy in Southern Nigeria.

Aim of the study

The antimalarial activities of ethanolic stem bark extracts of Enantia polycarpa was studied in vivo, in mice infected with Plasmodium berghei berghei.

Materials and methods

The ethanolic stem bark extract of Enantia polycarpa was administered at doses ranging from 200 to 600 mg/kg/day to Plasmodium berghei infected mice in both early and established models of antiplasmodial studies.

Results

The extract of Enantia polycarpa exhibited promising antimalarial activity against both early and established infections. At a dose of 600 mg/kg the extract achieved a 75.8% and 72% chemosuppression of parasitaemia in the study of acute and established infections, respectively. The extract also prolonged mean survival time of Plasmodium berghei infected mice during the study of established infection. The mean survival time of mice administered Enantia polycarpa extract at 600 mg/kg/day (27 days) was significantly longer than infected/untreated control (12 days). For the acute toxicity study the extract had an intraperitoneal LD₅₀ of 186 mg/kg but caused no mortality when administered orally at doses as high as 2,000 and 4,000 mg/kg.

Conclusions

Collectively, the results indicate that Enantia polycarpa is safe when administered orally and possesses promising antimalarial activity, thus supporting its use in traditional medicine for the treatment of malaria.     

Analgesic and antimalarial activities of crude leaf extract and fractions of Acalypha wilkensiana

Aim of the study

Antiplasmodial and analgesic activities of leaf extract and fractions of Acalypha wilkensiana were evaluated to ascertain the folkloric claim of its antimalarial and analgesic activities.

Materials and methods

The crude leaf extract (220–659 mg/kg) and fractions (chloroform and aqueous; 440 mg/kg) of Acalypha wilkensiana were investigated for antiplasmodial activity against chloroquine sensitive Plasmodium berghei infections in mice and for analgesic activity against chemical and heat-induced pains. The antiplasmodial activity during early and established infections as well as prophylactic activity were investigated. Chloroquine (5 mg/kg) and pyrimethamine (1.2 mg/kg) were used as positive controls. Thin films made from tail blood of each mouse were used to assess the level of parasitaemia of the mice. Analgesic activity of the crude extract was also evaluated against acetic acid, formalin and heat-induced pains.

Results

The extract and its fractions dose-dependently reduced parasitaemia induced by chloroquine sensitive Plasmodium berghei infection in prophylactic, suppressive and curative models in mice. These reductions were statistically significant (p < 0.001). They also improved the mean survival time (MST) from 16 to 22 days relative to control (p < 0.01–0.001). The activities of extract/fractions were incomparable to that of the standard drugs used (chloroquine and pyrimethamine). On chemically and thermally induced pains, the extract inhibited acetic acid and formalin-induced inflammation as well as hot plate-induced pain in mice. These inhibitions were statistically significant (p < 0.001) and in a dose-dependent fashion.

Conclusion

The antiparasitaemic and analgesic effects may in part be mediated through the chemical constituents of the plant.     

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.     

A new protoberberine alkaloid from Meconopsis simplicifolia (D. Don) Walpers with potent antimalarial activity against a multidrug resistant Plasmodium falciparum strain

Ethnopharmacological relevance

The aerial components of Meconopsis simplicifolia (D. Don) Walpers are indicated in Bhutanese traditional medicine for treating malaria, coughs and colds, and the infections of the liver, lung and blood. This study is to validate the ethnopharmacological uses of this plant and also identify potent antimalarial drug leads through bioassays of its crude extracts and phytochemical constituents.

Materials and methods

Meconopsis simplicifolia (D. Don) Walpers was collected from Bhutan and its crude MeOH extract was subjected to acid-base fractionation. Through repeated extractions, separations and spectroscopic analysis, the alkaloids obtained were identified and tested for their antimalarial and cytotoxicity activities.

Results

Phytochemical studies resulted in the isolation of one new protoberberine type alkaloid which we named as simplicifolianine and five known alkaloids: protopine, norsanguinarine, dihydrosanguinarine, 6-methoxydihydrosanguinarine and oxysanguinarine. Among the five of the alkaloids tested, simplicifolianine showed the most potent antiplasmodial activities against the Plasmodium falciparum strains, TM4/8.2 (chloroquine–antifolate sensitive strain) and K1CB1 (multidrug resistant strain) with IC₅₀ values of 0.78 μg/mL and 1.29 μg/mL, respectively. The compounds tested did not show any significant cytotoxicity activities against human oral carcinoma KB cells and normal Vero cells of African kidney epithelial cells.

Conclusions

This study validated the traditional uses of the plant for the treatment of malaria and identified a new alkaloid, simplicifolianine as a potential antimalarial drug lead.     

Ethnopharmacology guided screening of traditional Indian herbs for selective inhibition of Plasmodium specific lactate dehydrogenase

Ethnopharmacological relevance

Medicinal plants traditionally used to treat malaria can provide quality leads towards identifying novel anti-malarial drugs. Here we combined this approach with target based drug discovery and explored Plasmodium specific lactate dehydrogenase (LDH) inhibitory activity of 8 Indian plants which are ethnically used to treat malaria.

Methods

LDH from Indian Plasmodium falciparum and Plasmodium vivax strains, were cloned and expressed in Escherichia coli, followed by purification of recombinant enzymes (rPfLDH and rPvLDH respectively). Extracts of 8 plants in different organic and aqueous solvents, were screened for their inhibitory activity on rPfLDH, rPvLDH and mammalian LDHs. Phyllanthus amarus aqueous extract was further tested for in vitro parasiticidal activity.

Results

Aqueous extract of Phyllanthus amarus Schum. and Thonn. and chloroform extract of Murraya koenigii (L.) Spreng. exhibited profound and exclusive inhibitory effect on Plasmodium falciparum LDH (IC₅₀=11.2 μg/ml±0.4) and Plasmodium vivax LDH (IC₅₀=6.0 μg/ml±0.6) respectively. Moreover, Phyllanthus amarus aqueous extract also demonstrated antiplasmodial activity in vitro, on Chloroquine sensitive and resistant strains of Plasmodium falciparum (IC₅₀=7.1 μg/ml±0.5 and 6.9 μg/ml±0.7 respectively).

Conclusion

Target specific screening of traditional herbs used in malaria treatment has proffered Phyllanthus amarus and Murraya koenigii extracts as hits which can optimistically provide novel antimalarial drugs.     

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.     

Substantiation of the ethnopharmacological use of Conyza sumatrensis (Retz.) E.H.Walker in the treatment of malaria through in-vivo evaluation in Plasmodium berghei infected mice

Ethnopharmacological relevance

Scientific validation of ethnopharmacologically used plants and their utilization for therapeutic interventions can be a source of affordable treatment especially for neglected diseases in endemic areas. Conyza sumatrensis is a plant which finds its use in treating malaria like fevers but lacks proper scientific validation. Our study has tried to address this gap by authenticating its traditional use for the treatment of malaria.

Aim of the study

Evaluate the antimalarial activity of extracts derived from Conyza sumatrensis for its ethnopharmacological validation.

Materials and methods

Shade dried leaves were extracted with varying concentrations of ethanol and concentrated for bio-evaluation. Swiss albino mice infected with 1×10⁶ parasitized red blood cells, were orally administered with test extracts for 7 days in two sets of experiments. The first set was used to evaluate alcoholic, hydroalcoholic and aqueous extracts while the second set was used to evaluate the dose response of alcoholic extract ranging from 500–1600 mg/kg. Mean survival time, parasitaemia and haemoglobin levels were considered to interpret the antimalarial potential. Phytochemical analysis for the presence of flavonoids, alkaloids tannins, total phenolics, riboflavin and thiamine was also carried out.

Results

Among the three extracts administered at 1000 mg/kg, chemo suppression was significantly (p<0.001) observed in the alcoholic extract (62.59±12.52%) followed by hydroalcoholic (41.81±19.04%, p<0.01) and aqueous (32.04±19.04%, P<0.05) indicating that the active constituents leach out in ethanol. The dose response study involving the ethanol extract concluded the optimum dose to be 1000 mg/kg, as also evidenced by the haemoglobin levels.

Conclusion

The plant exhibits moderate antimalarial activity which can be further prospected for active fractions or pure molecules for adjunctive therapy.     

In vivo antimalarial evaluation of Gomphostenins

Aim of the study

The present study aims to evaluate the in vivo antimalarial potential of the leaf extract of Gomphostemma niveum and two new compounds; named as Gomphostenin (GN–6) and acetyl Gomphostenin (GN–10) isolated and purified from this plant against Plasmodium berghei in mice.

Materials and methods

The blood schizontocidal activity was performed in early infection of Plasmodium berghei and also in established infection of Plasmodium berghei. The mice were orally administrated with various doses of water and chloroform extracts of leaves of Gomphostemma niveum (GN–W and GN–C, 300, 400 and 500 mg/kg/day), GN–6 and GN–10 (50, 100, 150 and 200 mg/kg/day). Chloroquine (8 mg/kg) used as a positive control, while an equal volume of saline plus 0.5% tween-80 was used as a negative control.

Results

GN–W, GN–C and pure compounds (GN–6 and GN–10) produced a dose dependent chemosuppression effect at various dose levels. GN–10 exhibited highest percent of chemosuppression i.e. 92.65% at a dose level of 200 mg/kg/day. In case of curative test, the survival period of the parasitized infected mice was significantly prolonged at 200 mg/kg dose of GN–10.

Conclusions

The studies have indicated that clerodane class of diterpenes GN–6 and GN–10 certainly holds great promise for malaria control and will be useful in antimalarial chemotherapy.     

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 and in vivo antimalarial activity and cytotoxicity of extracts and fractions from the leaves,root-bark and stem-bark of Triclisia gilletii

Ethnopharmacological Relevance

To evaluate the in vitro antiplasmodial activity and cytotoxicity, and the in vivo activity of extracts and fractions from the leaves, root-bark and stem-bark of Triclisia gilletii (De Wild) Staner (Menispermaceae), used in traditional medicine against malaria.

Materials and Methods

The aqueous and 80% MeOH extracts, and a series of fractions and subfractions from the leaves, stem and root-bark of Triclisia gilletii were tested in vitro for their antiplasmodial activity against a Congolese-sensitive strain of Plasmodium falciparum, against the chloroquine and pyrimethamine-resistant K1 strain of Plasmodium falciparum, for cytotoxicity against MRC-5 cells, and in vivo in mice infected with Plasmodium berghei berghei.

Results

Many samples from the three plant parts exhibited pronounced activity against the Congolese chloroquine-sensitive strain of Plasmodium falciparum with some IC₅₀ values <0.02 µg/ml, and against the K1 strain, with some IC₅₀ <0.25; the selectivity was higher against the Congolese strain. At oral doses of 200 and 400 mg/kg body weight in infected mice, the aqueous, 80% methanol and total alkaloid extracts from the three plant parts produced more than 65% and 75% chemosuppression, respectively. The antiplasmodial activity of these three plant parts of Triclisia gilletii can at least in part be attributed to bisbenzylisoquinoline alkaloids, and supports its use for the treatment of uncomplicated malaria in traditional medicine.     

Antimalarial and safety evaluation of Pluchea lanceolata (DC.) Oliv. & Hiern: In-vitro and in-vivo study

Ethnopharmacological relevance

Many of the effective therapeutic strategies have been derived from ethnopharmacologically used natural products. Pluchea lanceolata is an herb employed in Indian folk medicine for malaria like fever but it lacks proper pharmacological intervention.

Aim of the study

To evaluate antimalarial and safety profile of Pluchea lanceolata: an in-vitro, in-vivo for its ethnopharmacological validation.

Materials and methods

Methanol, butanol, ethyl acetate, chloroform, hexane extracts and its isolate, taraxasterol acetate (TxAc) were obtained from air dried aerial part of Pluchea lanceolata. These were tested in-vitro against chloroquine-sensitive strain of Plasmodium falciparum NF54 by measuring the parasite specific lactate dehydrogenase activity. The most potent hexane extract and TxAc were further validated for in-vivo antimalarial and safety evaluation. TxAc, a pentacyclic-triterpene isolated from the most active fraction was further evaluated with special emphasis on inflammatory mediators involved in malaria pathogenesis. Murine malaria was induced by intra-peritoneal injection of Plasmodium berghei infected red blood cells to the male Swiss inbred mice. Mice were orally treated following Peters 4-Day suppression test. In-vivo antimalarial efficacy was examined by evaluating the parasitaemia, percent survival, mean survival time, blood glucose, haemoglobin and pro-inflammatory mediators involved in malaria pathogenesis.

Results

Hexane extract and TxAc showed promising antimalarial activity in-vitro and in-vivo condition. TxAc attributed in inhibition of the pro-inflammatory cytokines as well as afford to significant increase in the blood glucose and haemoglobin level when compared with vehicle treated infected mice. We have not observed the synergistic action of combinations of chloroquine and TxAc from our experimental results. In-vitro and in-vivo safety evaluation study revealed that hexane extract is non toxic at higher concentration.

Conclusion

Present study further validates the ancient Indian traditional knowledge and use of Pluchea lanceolata as an antimalarial agent. Study confirms the suitability of Pluchea lanceolata as a candidate for further studies to obtain a prototype for antimalarial medicine.     

In vivo antimalarial activity,toxicity and phytochemical screening of selected antimalarial plants

Ethnopharmacological relevance

Malaria continues to kill over a million people each year and in many populations affected by malaria, conventional drugs are often unaffordable or inaccessible. Historically, plants have been a prominent source of antimalarial drugs. Those plants currently used by indigenous people to treat malaria should be documented and investigated as potential sources of new antimalarial drugs.

Aim of the study

To investigate in vivo antimalarial activity, toxicity and carry out phytochemical screening of selected plants which have been used in traditional medicine for the treatment of malaria.

Materials and methods

Organic and water extracts of four medicinal plants used for the treatment of malaria in traditional health systems of Msambweni people in Kenya were tested for antimalarial activity against Plasmodium berghei and brine shrimp lethality. They were also screened for their major phytochemical constituents.

Results

Aqueous extract of the stem bark of Adansonia digitata exhibited highest chemosuppression of parasitaemia, >60% in a murine model of Plasmodium berghei infected mice. Aqueous and organic extracts of Launaea cornuta and Zanthoxylum chalybeum were toxic to the brine shrimp (LD₅₀<1000 μg/ml) while aqueous and organic extracts of Adansonia digitata and aqueous extracts of Canthium glaucum were not toxic to brine shrimp (LD₅₀>1000 μg/ml). Phytochemical screening revealed the presence of alkaloids and flavonoids in all the crude extracts of the selected plant species studied. Sesquiterpene lactones and saponis were present in organic extracts but absent in the aqueous extracts of Adansonia digitata, Canthium glaucum, Launaea cornuta and Zanthoxylum chalybeum.

Conclusion

The results showed that the crude extracts of Adansonia digitata and Canthium glaucum demonstrated promising antimalarial activity and there is potential for isolation of lead compounds from their extracts.     

Zanthoxylum capense constituents with antimycobacterial activity against Mycobacterium tuberculosis in vitro and ex vivo within human macrophages

Ethnopharmacological relevance

Zanthoxylum capense Thunb. (Rutaceae) is a medicinal plant traditionally used in Mozambique to treat tuberculosis.

Aims of the study

The main aim of the study was to find antimycobacterial lead compounds from Zanthoxylum capense. Another goal was to provide scientific validation for the use of this plant in traditional medicine.

Methods and materials

By bioassay-guided fractionation, 16 compounds were isolated and screened for their in vitro antimycobacterial activity against two different strains of Mycobacterium tuberculosis. Their in vitro cytotoxicity to human THP-1 macrophages was also assessed. The compounds with favourable selectivity index values (SI>10) were further investigated for their ability to inhibit the growth of Mycobacterium tuberculosis H37Rv in an intracellular macrophage model of infection.

Results

The best results were obtained for a benzophenanthridine alkaloid, decarine (1), and an N-isobutylamide, N-isobutyl-(2E,4E)-2,4-tetradecadienamide (15), which showed high activity against Mycobacterium tuberculosis H37Rv (MIC of 1.6 μg/ml), and a low macrophage cytotoxicity (IC₅₀>60 μg/ml), indicating considerable selective activity. The benzophenanthridine alkaloid 6-acetonyldihydronitidine (6) revealed cytotoxicity (IC₅₀ 1.7 μg/ml), despite the determined MIC of 6.2–12.5 μg/ml. In infected macrophages, decarine (1) was able to reduce bacterial survival by almost two log units at a concentration of 6.2 μg/ml 5 days post-drug exposure. Compound 15 exhibited an intermediate activity at drug concentrations ranging from 6.2 to 25 μg/ml.

Conclusions

The high antimycobacterial activity of decarine found, both in vitro and ex vivo against mycobacteria, and the low cytotoxicity towards human macrophages indicate that it may be valuable as a lead scaffold for the development of anti-TB drugs.     

Toxicity studies of Tithonia diversifolia A. Gray (Asteraceae) in rats

Objective

To investigate the toxicity of an ethanolic extract of the aerial parts of Tithonia diversifolia, used in Nigeria to treat malaria, in rats.

Materials and methods

A 70% ethanol extract was administered orally to adult Wistar rats at various dosages (400–1600 mg/kg) and the animals sacrificed and various organs examined at a range of times from 30 min up to 24 h after administration.

Results

The studies showed a dose- and time-dependent toxic effect, which was reversible on the kidney and liver while there was no noticeable adverse effect on the morphology of the heart, spleen and brain.

Conclusion

A 70% ethanol extract of the aerial parts of Tithonia diversifolia, which had previously been shown to reduce parasitemia in mice infected with Plasmodium, displayed kidney and liver toxicity at the lowest dose tested. The use of this plant extract against malaria therefore raises concerns over its safety.     

Screening of antiplasmodial properties among some traditionally used Iranian plants

Ethnopharmacological relevance

An investigation of plants was undertaken through interviews and literature surveys on plants used to treat malaria or cancer or microbial diseases in Iran.

Aim of study

In vitro and in vivo antiplasmodial tests were carried out on selected plants traditionally used in Iran.

Materials and methods

Thirty-two plants were extracted with methanol and tested for their in vitro (pLDH assay) activity against Plasmodium falciparum, in vivo activity against Plasmodium berghei and assessed for any cytotoxicity against the human cancer cell line MCF7 and the normal cell MDBK.

Results

Extracts from four plants, Buxus hyrcana Pojark. (Buxaceae), Erodium oxyrrhnchum M. Bieb. (Geraniaceae), Glycyrrhiza glabra L. (Fabaceae) and Ferula oopoda (Boiss and Bushe) Boiss. (Apiaceae) were found to have significant antiplasmodial activity (IC₅₀ ranging from 4.7 to 26.6 μg/ml). These findings lend support to the use of Buxus hyrcana and Glycyrrhiza glabra in traditional medicine. The chloroformic fraction also was active against K1 and 3D7 strains. The chloroformic fraction was studied at 10 mg per kg body weight mouse per day. This fraction reduced parasitaemia by 86.1% compared to untreated control mice.

Conclusion

Glycyrrhiza glabra showed antiplasmodial activity and has selectivity for Plasmodium falciparum and Plasmodium berghei when tested on mammalian cells. This is the first report that mentioned in vitro and in vivo antiplasmodial activity of Glycyrrhiza glabra.     

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.     

Antiplasmodial activity of sesquiterpene lactones and a sucrose ester from Vernonia guineensis Benth. (Asteraceae)

Ethnopharmacological relevance

Aqueous preparations of Vernonia guineensis Benth. (Asteraceae) are used in Cameroonian folk medicine as a general stimulant and to treat various illnesses and conditions including malaria, bacterial infections and helminthic infestations.

Materials and methods

Ten gram samples of the leaf and tuber powders of Vernonia guineensis were extracted separately using dichloromethane, methanol and distilled water. The extracts were dried in vacuo and used in bioassays. These extracts and three compounds previously isolated from Vernonia guineensis [vernopicrin (1), vernomelitensin (2) and pentaisovalerylsucrose (3)] were screened for antiplasmodial activity against chloroquine (CQ)-sensitive (Hb3) and CQ-resistant (Dd2) Plasmodium falciparum lines.

Results

Crude extracts and pure compounds from Vernonia guineensis showed antiplasmodial activity against both Hb3 and Dd2. The IC₅₀ values of extracts ranged from 1.64 to 27.2 µg/ml for Hb3 and 1.82–30.0 µg/ml for Dd2; those for compounds 1, 2 and 3 ranged from 0.47 to 1.62 µg/ml (1364–1774 nM) for Hb3 and 0.57–1.50 µg/ml (1644–2332 nM) for Dd2. None of the crude extracts or pure compounds was observed to exert toxic effects on the erythrocytes used to cultivate the Plasmodium falciparum lines.

Conclusion

In Cameroonian folk medicine, Vernonia guineensis may be used to treat malaria in part due to the antiplasmodial activity of sesquiterpene lactones (1, 2), a sucrose ester (3) and perhaps other compounds present in crude plant extracts. Exploring the safety and antiplasmodial efficacy of these compounds in vivo requires further study.     

Anthelmintic activity of Ziziphus nummularia (bark) and Acacia nilotica (fruit) against Trichostrongylid nematodes of sheep

Ethnopharmacological relevance

Ziziphus nummularia (Rhamnaceae) and Acacia nilotica (Fabaceae) are being used as anthelmintics in ethnoveterinary medicinal system of Pakistan.

Aim of the study

Present study was conducted to determine the anthelmintic activity of Ziziphus nummularia (bark) and Acacia nilotica (fruit) in order to justify their traditional use in veterinary medicine.

Materials and methods

In vitro anthelmintic activity of crude methanolic extract (CME) of both the plants was determined against Haemonchus contortus by the adult motility assay, the egg hatch test and the larval development assay. In vivo anthelmintic activity was evaluated in sheep naturally infected with gastrointestinal nematodes by administering increasing doses of crude powder (CP) and CME (1.0–3.0 g/kg).

Results

Both the plants exhibited dose- and time-dependent anthelmintic effects by causing mortality of worms, and inhibiting egg hatching and larval development. Acacia nilotica (LC₅₀ = 512.86 and 194.98 μg/ml) was found to be more potent than Ziziphus nummularia (LC₅₀ = 676.08 and 398.11 μg/ml) in egg hatch test and larval development assay, respectively. In vivo, maximum fecal egg count reduction (84.7%) was recorded on day 13 post-treatment in sheep treated with Ziziphus nummularia CME (3.0 g/kg) followed by 78.5% on same day with Acacia nilotica CME (3.0 g/kg).

Conclusions

These data show that both Ziziphus nummularia and Acacia nilotica possess anthelmintic activity in vitro and in vivo, justifying their use in traditional veterinary medicine in Pakistan.     

Anthelmintic activity of Arisaema franchetianum and Arisaema lobatum essential oils against Haemonchus contortus

Ethnopharmacological relevance

Arisaema franchetianum and Arisaema lobatum are two perennial plants native to China. Arisaema franchetianum is universally used to promote the subsidence of induration and swelling, quicken blood and relieve pains, and kill intestinal parasites in humans and animals. Arisaema lobatum is used to treat malaria, intestinal parasites, and snake and insect bites in humans and animals. The aim of this study was to determine the composition of the essential oils from Arisaema franchetianum and Arisaema lobatum and evaluate the anthelmintic effect against Haemonchus contortus.

Materials and methods

Two oils were investigated by GC and GC–MS. The anthelmintic bioassay tests of Arisaema franchetianum and Arisaema lobatum essential oil, linalool and carvacrol were performed using egg hatch assay (EHA), larval development assay (LDA) and larval migration inhibition assay (LMIA).

Results

Fifty six components representing 96.88% of the Arisaema franchetianum oil and 64 components representing 96.88% of the Arisaema lobatum oil were identified. Carvacrol and linalool were found to be the major constituents of two oils. In the EHA, greater than 99% inhibition were observed with Arisaema franchetianum oil at 10 mg/mL (CE₅₀ 1.63 mg/mL) and Arisaema lobatum oil at 5 and 10 mg/mL (CE₅₀ 0.48 mg/mL). In the LDA, both oils induced complete inhibition at 10 mg/mL, with the CE₅₀ being 1.10 mg/mL for Arisaema franchetianum oil and 0.73 mg/mL for Arisaema lobatum oil. In the LMIA, the Arisaema franchetianum oil and Arisaema lobatum oil at best inhibited 74.1% and 95.6% of larval migration at 10 mg/mL, respectively. Carvacrol exhibited similar activity to Arisaema lobatum essential oil and linalool did not show high activity in every assay.

Conclusions

These data show for the first time that the essential oils obtained from Arisaema franchetianum or Arisaema lobatum had promising anthelmintic activity against Haemonchus contortus. Arisaema plant may offer an alternative source for the control of gastrointestinal nematodes of sheep and goats.