OA20988A - Phytomedicine based on extracts of two medicinal plants for the treatment of uncomplicated malaria and its preparation process. - Google Patents

Abstract

The invention relates to a completely safe phytomedicine, composed of extracts of two medicinal plants: Alchornea cordifolia and Annikia polycarpa, formulated in granules packaged in capsules and intended to treat malaria. It acts by reducing Plasmodium berghéi parasitemia by at least 75%. The remedy according to the invention derives its antiplasmodial activity from metabolites which are flavonoids, alkaloids, polyphenols including ellagic acid whose combined actions block the development of strains of Plasmodium falciparum and destroy it.

Description

Phytomedicine based on extracts of two medicinal plants for the treatment of uncomplicated malaria and its preparation process

The invention which is the subject of this patent application relates to the pharmaceutical field. It describes an antimalarial medication based on a combination of aqueous extracts of two medicinal plants: Alchornea cordifolia and Annikia polycarpa.

The invention also relates to the process for preparing the phytomedicine.

Malaria is a potentially fatal human disease caused by parasites transmitted by the bites of infected female Anopheles mosquitoes. According to the World Health Organization (WHO), in 2020, there were an estimated 241 million cases of malaria worldwide. And the estimated number of deaths attributable to malaria stood at 627 000. The WHO African Region bears a large and disproportionate share of the global malaria burden. In 2020, 95% of malaria cases and 96% of deaths due to the disease were recorded in this Region. Children under 5 years old accounted for 80% of 15% of malaria deaths in the Region. The first symptoms: fever, headache and chills usually appear 10 to 15 days after the infectious bite and may be mild and difficult to recognize. If left untreated, P. falciparum malaria can progress to serious or even fatal illness within 24 hours.

Despite advances in many countries, including the recent declaration that China is now malaria-free, malaria remains entrenched in Africa, where more than 90% of malaria morbidity and mortality is seen. In the past, Africa has suffered greatly from the effects of resistance to chloroquine, the mainstay of malaria treatment for decades, with increasing mortality from Plasmodium falciparum in the last decades of the 20th century. These increases were 25 reversed earlier this century, with an estimated 57% decrease in the malaria death rate from 2000 to 2015.

Malaria still remains endemic in Côte d'Ivoire with the majority of the population (81%) living in regions with an estimated incidence and cases varying from 300% to more than 500% of inhabitants. Overall, there are five levels of endemicity classified into 30 levels: very low, low, moderate, high and very high, which respectively cover 2%, 17%,

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30%, 18% and 34% of the population. Furthermore, data from the parasitic prevalence survey for malaria and anemia in children aged 6 to 59 months (EPPA 2016) reveal an average parasitic prevalence of 37%. Malaria is transmitted by three vector species complexes in Côte d'Ivoire: An. gambiae s.l, An. funestus s.l. and 5 An. nili s.l.

The economic burden of malaria is considerable. In Ivory Coast for example, according to studies, the direct costs associated with malaria represent on average 12 to 14% of the income of households whose active members also lose 4 to 7 days of work (A.S. Kouadio et al). According to Tchicaya et al, the direct cost of malaria represented 213,223.85 euros (139,865 to 677 FCFA), absenteeism linked to malaria at 64,738.33 euros (4,249,507 FCFA) and the cost of productivity at 933,042 .73 euros (612,035,910 FCFA).

The fight against malaria revolves around two major axes: prevention and treatment. Prevention is done through tools including insecticide-treated mosquito nets, indoor insecticide spraying and chemoprevention of seasonal malaria for children under five years old. These means are usually deployed as part of large-scale campaigns. As for treatment, its main objective is to ensure the rapid and complete elimination of malaria to avoid the progression of a simple case to severe malaria which can lead to death. For the WHO, the best treatment available, particularly for P. falciparum malaria, is combination therapy 20 based on artemisinin (CTA). Other medications such as artemisinin and its derivatives are recommended: atovaquone-proguanil, chloroquine (or hydroxychloroquine), clindamycin, doxycycline, mefloquine, primaquine, quinine and quinidine. However, these medications have the disadvantage of producing numerous undesirable effects: headaches, anxiety, depression, insomnia, nightmares, abdominal pain, visual disturbances and itching, attention disorders, hallucinations, fatigue, fever and cardiovascular disorders, digestive disorders, photosensitivity of the skin.

Phytotherapy is not left out, thus, currently on the African market, particularly in the West African sub-region, we find numerous antimalarials in the form of raw powders resulting from the direct pulverization of plant parts and put in the form 30 capsules. The metabolites of these plant powders which have not been previously extracted will not be quickly made available and will have a delayed action.

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The majority of scientific work carried out on the antiplasmodial effectiveness of medicinal plants focuses preferentially on organic extracts. Very few studies focus on aqueous extracts. For example, only one study carried out on a 72-hour cold aqueous macerate of Alchornea cordifolia leaves refers to antiplasmodial efficacy tests (Ezeokeke et al2015). As for the antiplasmodial efficacy tests of extracts of bark from the trunk of Annickia polycarpa, they were evaluated using an organic solvent, in this case I ethanol (Anosa et al 2014). No study of antiplasmodial activity on an aqueous extract of this plant has been found in the literature.

Alchornea cordifolia, commonly called Djeka, in Ivory Coast, is a plant often found in multi-species recipes. The literature notably reports combinations of plants with antimalarial purposes containing Alchornea cordifolia. These are Alchornea cordifolia and Alstonia boonei; Alchornea cordifolia and Anthocleista djanolensis; Alchornea cordifolia and Vernonia colorata (Sylla et al, 2018). However, the antimalarial activities of these herbal combinations based on Alchornea cordifolia have not been corroborated by evaluation trials.

The antimalarial phytomedicines found in pharmacies and/or traditional medicine centers are, for example, “Die kouadio” from Laboratoires STerre which contains neem, kinkeliba, and Khaya senegalensis. There is also “Palutime” from Laboratoires Gamet: a combination of neem, false kinkeliba, Lippia knighti, golden chamomile and Tamarind. Also found on the market are 20 phytomedicines, Malarial, a combination of Cassia occidentalis, Lippia chevalieri, and Spilanthes oleraceae, another phytomedicine “Saye” containing a combination of Cochlospermum planchonii, Cassia alata and Phyllantus sp, and also the phytomedicine “Api-palu”, a combination of several medicinal plants.

To date, no combination of antimalarial plants containing Annikia 25 polycarpa has been found in the literature.

The present invention aims to develop a natural medicine against malaria, based on plants, non-toxic and having a rapid effect by incorporating a new plant in the formulation of phytomedicines indicated for the treatment of malaria.

In accordance with the invention, this goal is achieved with a phytomedicine composed of extracts of two medicinal plants: Alchornea cordifolia and Annikia polycarpa. The medicine according to

[4] the invention is an antimalarial combining aqueous extracts of the leaves of Alchornea cordifolia and the bark of the trunk of Annikia polycarpa. These parts of both plants possess potentiated antiplasmodial activity in vitro and in vivo for the effective treatment of uncomplicated malaria. These two plants have been the subject of numerous scientific studies to demonstrate their effectiveness and safety.

Indeed, the combined phytochemical composition of the two plants considerably reduces parasitemia. The remedy according to the invention derives its antiplasmodial activity from metabolites such as flavonoids, alkaloids including berberine and dihydroquinidine, polyphenols including ellagic acid identified as carrying antiplasmodial activity. And also has a reduction rate in Plasmodium berghéi parasitemia of at least 75%, with a dose of 300 mg.

The phytomedicine according to the invention contains metabolites already extracted from plant parts and are quickly released for rapid therapeutic action, suitable for antimalarial therapeutic activity. The phytomedicine acts by blocking protein synthesis, and by chelating the nucleic acids of the Plasmodium nucleic bases using alkaloids and flavonoids, which also inhibit the intra-erythrocyte growth of certain strains of Plasmodium falciparum. As for tannins, they destroy Plasmodium through an antioxidant effect by chelating metal ions, and reducing ferric iron (Fe ³⁺ ) to ferrous iron (Fe ²⁺ ).

In one embodiment, the phytomedicine according to the invention is formulated in granules and packaged in capsules in order to facilitate its ambulatory use. It is obtained from the following process:

- Harvesting and cleaning: the leaves of Alchornea cordifolia and the bark of the trunk of Annikia polycarpa are collected and washed carefully in drinking water;

- Decoction: the leaves and cut bark are heated separately until boiling in clean water to extract the metabolites;

- Concentration: each aqueous extract is concentrated by evaporation under vacuum until a dry matter content is at least ten times higher.

- Mixing, granulation and encapsulation: the concentrated aqueous extracts of leaves of Alchornea cordifolia and bark of the trunk of Annikia polycarpa are then mixed in identical proportion.

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-Granulation and drying: the homogeneous mixture is diluted in corn starch in an almost identical ratio and kneaded, to give a pasty mass which is transformed into grains, before being dried at around 50°C to a maximum humidity level of 10% to produce dry grains.

- Encapsulation: the dry grains are put into capsules for approximately 400 mg.

The entire process for preparing the phytomedicine according to the invention can be transposed to an industrial scale.

To ensure safety of use, safety tests of the remedy according to the invention at different doses were carried out in animals free from any infection. Administered at doses of 600mg and 800mg over seven (7) consecutive days twice a day, the phytomedicine shows an absence of hematological, biochemical and clinical toxicity upon repeated administration. Indeed, the biological parameters studied, CBC, uremia, creatinine, transaminases, blood sugar, cholesterol showed no significant difference with the untreated batch. As well as the clinical parameters studied which are the evolution of body weight and the study of the animal's behavior.

The phytomedicine according to the invention has numerous advantages which are as follows:

- it presents no toxicity for the patient suffering from malaria;

- the plants which compose it are easily accessible and available,

- its production is simple and inexpensive;

- it is very effective in treating simple malaria at a lower cost.

The other characteristics and advantages will appear more clearly on reading the detailed description which is based on a non-limiting embodiment of the phytomedicine according to the invention, based on Figure 1 of the single plate.

The invention is a phytomedicine composed of extracts of two medicinal plants: Alchomea cordifolia and Annikia polycarpa. In one embodiment, it is formulated in granules packaged in capsules in order to facilitate its ambulatory use. It is obtained following the following steps:

- Harvesting and cleaning: the leaves of Alchornea cordifolia and the bark of the trunk of

Annikia polycarpa are collected and washed carefully with distilled water.

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- Decoction: the leaves and cut bark are heated separately in clean water to extract the metabolites. In a preferred embodiment, 100 grams of Alchornea cordifolia leaves are introduced into 1 liter of distilled water and heated to approximately 70°C, with stirring at 500 rpm for a period of one hour. The collected aqueous extract then undergoes tangential filtration to collect 800 mL of substrate. And 300g of Annikia polycarpa bark are also treated as before to also give 800 mL of substrate.

- Concentration: each aqueous extract is concentrated by evaporation under vacuum, in an evaporator, at a temperature of 40°C until a concentrate of approximately 80 mL is obtained.

io - Mixture: the two concentrated aqueous extracts of leaves of Alchornea cordifolia and bark of the trunk of Annikia polycarpa are put together, the mixture is then stirred for 5 minutes until total homogenization is observed by visual observation.

- Granulation and drying: to obtain the grains, a mixture containing 46.5% concentrated aqueous extract and 53.5% corn starch is produced with stirring and kneading for 30 min to give a pasty mass. The pasty mass is granulated in a rotary granulator to give wet grains of size between 50 and 300 μm. The damp grains are then put in an electric oven at 50°C to obtain dry grains with a humidity level of less than 10% and sizes between 40 and 100pm.

Encapsulation. the dry grains are put into capsules using a semi-automatic capsule maker and by the leveling technique, to give capsules of size No. 2 containing 300 mg of plant extract for 75 mg of corn starch. This mixture conforming to the phytomedicine according to the invention can be stored for at least five years from the date of preparation.

For treatment, the phytomedicine according to the invention is administered at a rate of 2 to 25 capsules 2 to 3 times per day for a period of 7 days.

To ensure the safety of use of the phytomedicine, safety tests of the remedy according to the invention at different doses were carried out in animals free from any infection. Administered at doses of 600mg and 800mg over seven (7) consecutive days twice a day, the phytomedicine shows an absence of hematological, biochemical and clinical toxicity in repeated administration. Indeed, the biological parameters studied, CBC, uremia, creatinine, transaminases, blood sugar, cholesterol showed no difference.

[7] significant with the untreated batch. As well as the clinical parameters studied which are the evolution of body weight and the study of the animal's behavior.

Indeed, the combined phytochemical composition of the two reduced plants gives the phytomedicine according to the invention an effective antiplasmodial activity for simple malaria which considerably reduces parasitemia. The remedy according to the invention derives its antiplasmodial activity from metabolites such as flavonoids, alkaloids including berberine and dihydroquinidine, polyphenols including ellagic acid identified as carrying antiplasmodial activity. And has a reduction rate in Plasmodium berghéi parasitemia of at least 75%, with a dose of 300 mg. Its content of metabolites already extracted from plant parts allows rapid therapeutic action.

In its mode of action, the phytomedicine acts by blocking protein synthesis, and by chelating the nucleic acids of the plasmodium nucleic bases thanks to alkaloids and flavonoids, which also inhibit the intra-erythrocytic growth of certain strains of Plasmodium falciparum. As for tannins, they destroy the plasmodium through an antioxidant effect 15 by chelating metal ions, reducing ferric iron (Fe3+) to ferrous iron (Fe^ ⁴ ·).

Claims (3)

1- Phytomedicine characterized in that said phytomedicine is composed of aqueous extracts of the leaves of Alchornea cordifolia and the bark of the trunk of Annikia polycarpa having antiplasmodial activity through their composition of metabolites such as flavonoids, alkaloids including berberine and dihydroquinidine , polyphenols including ellagic acid, which can induce a reduction rate in Plasmodium berghéi parasitemia of at least 75% in simple malaria patients by blocking the development and destruction of Plasmodium falciparum strains. 2- Obtaining phytomedicine according to claim 1, characterized in that the process comprises the following steps: - harvesting and cleaning: the leaves of Alchornea cordifolia and the bark of the trunk of Annikia polycarpa are collected and washed carefully in drinking water; - decoction: the leaves and cut bark are heated separately until boiling in clean water to extract the metabolites; - concentration: each aqueous extract is concentrated by evaporation under vacuum until a dry matter content is at least ten times higher; mixing, granulation and encapsulation: the concentrated aqueous extracts are then mixed in a ratio of approximately 1:1, and diluted in corn starch to give a wet paste which is processed into grains and dried at approximately 40°C to reach a maximum humidity level of 10%, then to give dry grains which are put into capsules for 400 mg. 3- Phytomedicine according to claims 1 and 2, characterized in that the safety of said phytomedicine is hematological, biochemical and clinical.