FR3143326A3 - Pharmaceutical or dietetic composition comprising an extract of Hyphaene thebaica and an extract of Olea europaea usable as an antihypertensive and hypolipidemic agent. - Google Patents

Abstract

The invention relates to a new combination of an aqueous extract of the stripped pericarp of the fruit of Hyphaene thebaica (doum palm) and an extract of young leaves of Olea europaea under the name IFANOSINE for the treatment of arterial hypertension or hyperlipidemia, said combination being in a form suitable for oral administration.

Description

Pharmaceutical or dietetic composition comprising an extract of Hyphaene thebaica and an extract of Olea europaea usable as an antihypertensive and hypolipidemic agent.

The present invention relates to a new pharmaceutical or dietary composition usable as a phytomedicine or antihypertensive or hypolipidemic food supplement. This composition formulated only with the dry aqueous extracts of Olive leaf and dry aqueous extracts of the pericarp of Hyphaene thebaica fruit.

Some definitions.

To better understand this text it is desirable to define some words and terms used in it. The majority of all the terms and words used are defined in the illustrated dictionary of medical terms by Garnier Delamare, ^28th edition or in the latest Larousse edition in French.

Antagonist: The potency of a reversible competitive antagonist is quantified by the pA2 defined as the negative logarithm of the antagonist concentration that requires doubling the agonist concentration to maintain the same effect. In other words, the pA2 is the concentration of the antagonist that reduces the effect of the agonist by half.

Antihypertensive: Which opposes hypertension. It is a medicine or food supplement based on hypertension plants.

Bioavailability: This is the biological availability of drugs. It is the possibility of an active ingredient of a drug to be absorbed in sufficient quantity and quickly enough to be effective. It is the quantity of active ingredient that reaches the receptors and the speed at which this phenomenon occurs. It depends on different factors: solubility, speed of dissolution of the active substance, composition and physical state of the excipients, pharmaceutical form of the drug.

Drug candidate: A substance or combination of substances still under development presented as potentially possessing curative or preventive properties with respect to human or animal diseases, as well as any product under development intended to be administered to humans or animals with a view to establishing a medical diagnosis, restoring, correcting or modifying their organic functions. In the context of this invention, the drug candidate designates the anticancer or antibiotic, or the antifungal, or the antiparasitic.

Median effective concentration (EC50): is a measure of the concentration of a drug, antibody or toxicant that induces a response halfway (median) between the baseline and the maximum effect after a certain time of exposure to it.

Median inhibitory concentration (IC ₅₀ ): is a measure of the effectiveness of a given compound in inhibiting a specific biological or biochemical function. _Often , the compound in question is a potential drug.

Screening: In the Larousse dictionary, screening refers to the operation of sorting materials, with a view to granulometric classification. In the context of this document, screening refers to the rapid sorting of anticancer drugs, antibiotics, and antifungals.

Plant drug: Plant raw material intended for the preparation of medicine.

Efficacy: Efficacy refers to the established pharmacological and clinical effects in relation to a defined treatment regimen and a specified dosage.

Excipient: It is a vehicle for a drug. It is a substance to which the active ingredients are incorporated to make them easily absorbable.

Aqueous extract: By aqueous extract we mean the extract using only water as solvent in liquid or vapor form.

Hypolipidemic: Which reduces the level of lipids in the blood.

Medicine: Substance or combination of substances presented as having curative or preventive properties with regard to human or animal diseases, as well as any product that can be administered to humans or animals in order to establish a medical diagnosis, restore, correct or modify their organic functions. In the context of this invention, the drug candidate designates the anticancer or the antibiotic, or the antifungal, or the antiparasitic.

Essential medicine: A medicine that meets the priority health care needs of the population. These medicines are generally used for priority pathologies and are included in the national list of essential medicines.

Phytomedicine: Medicine whose active substances are plant raw materials.

Phytotherapy: Set of medical practices using substances of plant origin for therapeutic purposes.

Active ingredient of a drug: or active substance, is the component of the drug responsible for the therapeutic effect.

Bare pericarp: the fruit of Hyphaene thebaica is a leathery, ovoid drupe with a single stone, exceptionally two, embedded in a dry pulp (this is what we call in this invention pericarp), floury, fibrous, pinkish or yellowish; brick-brown integument, granular to the touch (this is the part that is removed, hence the expression bare).

Screening: In this document, screening refers to the rapid sorting of anticancer drugs or antibiotics, or antifungals. Screening can be replaced by cribrage.

Xenobiotic: A xenobiotic is a product that is foreign to biological chemistry and can behave as toxic or allergenic to the body. These are mainly household and industrial chemicals. In the context of this invention, xenobiotics refer to drugs or drug candidates, chemicals, trace elements, total or partial extracts of plants for therapeutic purposes.

Technical field of the invention

The present invention relates to a pharmaceutical or dietary composition combining an association of the dry aqueous extracts of two plants, the Egyptian doum palm (Hyphaene thebaica L. Mart.), and the olive leaves (Olea europaea). The mixture of the extracts of two plants is made according to a well-determined formula obtained from the tests obtained on the in vitro work on the isolated organs. The extracts of the doum palm are obtained from the bare pericarp of the fruit, while only the young olive leaves are used in this composition.

State of the prior art;

The use of the two plants has been well known since antiquity for their therapeutic virtue in the fight against several pathologies including hypertension and lipidemia. However, the use of these two plants remains confined to the stage of plant-based food supplements since their effectiveness remains limited in treating hypertension or excess lipids in the blood. Several studies report their use in preclinical and clinical settings in humans and animals. The big difference between these studies and use is a new formulation made from the preclinical studies that the applicant has obtained. This new formulation which gives the composition according to the invention an efficacy and clinical benefits superior to the chemical drugs used in clinics.

Presentation of the invention

1. Être plus efficace de l’extrait d’olivier seule comme hypotenseur et hypolipidémiant,
2. Être plus efficace que l’extrait du palmier doum seul comme hypotenseur et hypolipidémiant,
3. Ce phytomédicament doit être plus efficace que la Prazosine,
4. La biodisponibilité intestinale doit être supérieur à l’extrait de l’Olivier et du palmier doum séparément.
5. Le coût de revient doit être accessible à tous pour permettre aux démunis à avoir accès au phytomédicament.

The aim of the invention is to create a new phytomedicine by combining the aqueous extracts of young leaves (young shoots) of Olive tree and the aqueous extract of the bare pericarp of Doum palm having antihypertensive and hypolipidemic properties. Above all, the new phytomedicine must have the following characteristics:

1. Being more effective than olive extract alone as a hypotensive and hypolipidemic agent,
2. Be more effective than doum palm extract alone as a hypotensive and hypolipidemic agent,
3. This herbal medicine should be more effective than Prazosin,
4. Intestinal bioavailability should be higher than the extract of Olive and Doum palm separately.
5. The cost price must be accessible to all to allow the poor to have access to phytomedicine.

In the context of the invention, the pharmaceutical and/or dietary composition called IFANOSINE (mixture of dry aqueous extracts of Hyphaene thebaica and Olea europaea as active ingredients) is a pharmaceutical or dietary composition which meets the following conditions:

1. The formula for combining the active ingredients of Ifanosine based on the pA2 obtained in preclinical studies.

For 1.6 grams of Ifanosine principle, it is necessary to combine 1 gram of dry aqueous extract of young olive leaves with 0.6 grams of dry aqueous extract of the bare pericarp of the doum palm.

2. The intestinal bioavailability of the active ingredient Ifanosine must be greater than that of both the olive extract and the doum palm.

3. Clinical benefit in humans must be proven

Dosage forms

According to the invention, the composition can be presented in the form of tablets, capsules, granules, syrup, oral solution or any other forms known to those skilled in the art comprising the extracts of the two plants.

Those skilled in the art know and use various excipients as binders or lubricants for manufacturing powders, granules or other preparations. Examples of excipients which can thus be used according to the invention are: lactose, starch, dextrin, calcium phosphate, calcium carbonate, natural or synthetic aluminum silicate, magnesium oxide, hydrated aluminum hydroxide, magnesium stearate, sodium bicarbonate, rice starch or flour. Other excipients not mentioned above but which are also suitable are described in Remington's Pharmaceutical Sciences by E.W. Martin, and can be used according to the invention.

In these different formulations, and for example those mentioned above, the composition can also be presented in powder for dilution or syrup. This powder may not contain the active ingredients and the excipients or bulking agents.

Description of embodiments;

The implementation of the invention is done by preclinical studies on the relaxation of the rat aorta and by clinical trials in humans.

Materials and methods

Animals

To conduct this study, the target organs, the aorta and intestine, will be collected respectively from male Wistar strain albino rats aged 3 to 4 months and weighing an average of 350 g. These animals are raised in-house in a conventional and approved animal facility.

All animals are housed in a suitable environment where temperature, light and ventilation are controlled. They are fed with standard diet for laboratory animals and water ad libitum and identified by a sheet containing various information: laboratory name, name of the experiment supervisor, species, strain, sex, date of birth and weaning of the experimental animal. They are euthanized by intraperitoneal injection of phenobarbital.

Extraction of plant material

About thirty Doum palm fruits were purchased in southern Chad and formally authenticated. A maceration with 125 g of stripped mesocarp powder was carried out in 1 L of distilled water for 24 hours under a magnetic stirrer. The filtrate was concentrated at a temperature of 50 ° C using an oven. This operation made it possible to obtain 27.82 g of aqueous extract, i.e. a yield of 22.25%.

Physiological solution and pharmacological tools

Ringer's solution with the following composition: 115mM NaCl, 25mM _NaHCO3 , _{1.3mM MgCl2 ,} 1.2mM CaCl2, 2.4mM _K2HPO4 , 0.4mM _{KH2PO4 ,} was used as survival solution.

Adrenaline (Sigma France): Adrenaline is a neurotransmitter released following stimulation of the sympathetic nerves. It acts on the heart and blood vessels by activating different subtypes of α 1 & 2 and β 1 & 2 adrenergic receptors. Phenylephrine (Sigma, France): a specific agonist of α1 adrenergic receptors. Prazosin: (purchased in a pharmacy, TEVA brand) antagonist of α1 adrenergic receptors. All dilutions are made in Ringer.

In vitro study of the effects of extracts on contraction of smooth muscles of isolated rat aorta

Micro-dynamometer technique: Principle

The principle is to measure the contractile activity caused by a drug or a contracting substance. This technique consists of provoking with an agonist (adrenaline, phenylephrine) at known concentrations, a sustained contraction of the aorta. Then, determine the concentration range of the aqueous extract which would cause a partial or total relaxation of the contraction induced by the agonist.

Aortic harvesting and assembly

After anesthetizing the rat with pentobarbital (60 mg/kg, i.p.), it was placed in the dorsal decubitus position on a dissection board and a thoracotomy was performed. This allowed the aorta to be exposed. A fragment of the aorta was carefully removed as close as possible to the heart. It was freed from surrounding tissues, cleaned, then cut into rings of 3 mm maximum length and immediately immersed in Ringer's solution. After this, the aortic rings were each mounted between two hooks and a bracket connected to a sensor. The assembly thus produced is maintained in survival by placing it in a 3 ml isolated organ tank containing physiological solution thermostated at 37 ° C and continuously oxygenated (O2: 95%; CO2: 5%). Then, an equilibration period of 60 minutes with rinsing every 20 minutes was carried out. The organs were subjected to a basic voltage of 1.5g. Voltage variations resulting from variations in vessel diameter were captured and converted into electrical signals using an isometric "Transducer" (TBC Laboratories, Paris, France). These electrical signals were visualized on a graphic recorder using data acquisition software (BiodacqSoft, TBC Laboratories, Paris, France).

Performing the test

Effects of extracts on phenylephrine or adrenaline-induced contraction

The objective of this protocol was to evaluate the relaxing effect of aqueous extracts on aortic ring contraction. For this purpose, at the end of the equilibration period, the aortic rings were pre-contracted with a solution of adrenaline (1µg/ml) or phenylephrine (10 nM). All dilutions of the extract were carried out in Ringer.

First, after the maximum contraction was reached (resulting in a plateau on the curve), 1 mg/ml of extract was added to the tank using a micropipette. Second, the relaxant effect of the extract before contraction was studied. To do this, increasing concentrations ranging from 1 ng to 1 mg/ml were introduced in a non-cumulative manner into the isolated organ reservoir 3 minutes before the addition of adrenaline or phenylephrine. The total volume of the drug added did not exceed 10% of the total volume of the tank. Between each addition of the extract of a given concentration followed by either adrenaline or phenylephrine, the aortic rings were left to rest for 20 minutes during which two 10-minute rinses were performed. The experiment was repeated to ensure the reproducibility of the result obtained. The relaxation percentages at each concentration were calculated to follow the induced contraction.

Comparative study of the effect of extracts on aortic contraction with a clinically used antihypertensive drug: Prazosin

The objective of this test was to make a comparative study of the effect of the extract on the contraction of the aorta with an antagonist of the α2 receptors, which is an antihypertensive used in clinics. To do this, increasing doses of the plant extract (doum palm or Olivier) or Prazosin were added in a non-cumulative manner before the induction of the contraction by phenylephrine. Each of the doses of either the plant extract studied or Prazosin was tested with an increasing concentration range of phenylephrine. Between each addition of the aqueous extract or Prazosin followed by phenylephrine, the aortic rings were left to rest for 20 minutes during which two rinses of 10-min intervals were carried out.

Clinical trials

Clinical trials were conducted on patients by professionals after taking care to ensure patient safety and in accordance with current ethical standards. Clinical examinations and analyses of biological parameters were carried out. The following biological parameters were subject to statistical analyses to compare the effect of Ifanosine on the people treated to assess the clinical benefit. These are: diastolic and systolic blood pressure (DBP, PAS), total cholesterol (TC), lipoproteins (HDL, LDL), triglycerides and Apoproteins (apoA, apoB).

Statistical tests

All data for statistical analysis are expressed as mean ± standard deviation of the mean (SEM, n = number of experiments), except for _IC50 (drug concentration causing half the maximal response), or pA2 (antagonist concentration that reduces the agonist effect by half) which are presented as geometric means with their respective 95% confidence intervals. Statistical analyses were performed by one-way analysis of variance (ANOVA), followed by Dunnett's test when necessary. P < 0.05 was considered significant. Dose-response curves were analyzed by nonlinear regression (Graphpad program for Windows version 5.01. Graphpad, San Diego, CA, USA).

Brief description of the figures

, Effect of different plant extracts Olea europaea and aqueous leaf extract (OEL), Hyphaene thebaica (HT) and prazosin (PRZ) on phenylephrine-induced contraction of rat aortic rings.

, Typical recording of contraction on isolated aortic segment rings. Aqueous extract of Hyphaene thebaica at different concentrations (5-100 ng/ml) diluted in Ringer solution was added to organ baths 3 min before contraction induction with phenylephrine (10 nM). The reduction in contraction amplitude reflects the inhibitory effect of HT extract at different concentrations. Maximum contraction was measured at the plateau.

, The concentration-response curve of phenylephrine (PHE)-induced contraction on rat aortic rings in the presence of different concentrations of aqueous extract of Olea europaea leaf (OEL), as antagonist. Figure A, shows that there is a progressive shift in the concentration-response curve of phenylephrine in the presence of increasing concentration of OEL.

, The Schild plot of the concentration-response curve of PHE in the presence of different OEL concentrations indicates that the pA ₂ value was 1. The solid circles and vertical bars represent the means and standard deviations of 8 tissues from 3 rats, respectively.

, The concentration-response curve of phenylephrine (PHE)-induced contraction of rat aortic rings in the presence of different concentrations of aqueous extract of Hyphaene thebaica (HT), as antagonist. , shows that there is a progressive shift in the concentration-response curve of phenylephrine in the presence of increasing concentration of HT.

, The Schild plot of the PHE concentration-response curve in the presence of different concentrations of HT indicates that the pA ₂ value was 0.6 (B). The solid circles and vertical bars represent the means and standard deviations of 8 tissues from 3 rats, respectively.

, The concentration-response curve of phenylephrine (PHE)-induced contraction of rat aortic rings in the presence of different concentrations of prazosin (PRZ), as antagonist. , shows that there is a progressive shift in the concentration-response curve of phenylephrine in the presence of increasing concentration of PRZ.

, The Schild plot of the concentration-response curve of PHE in the presence of different concentrations of PRZ indicates that the pA ₂ value was 4 nM (i.e., 1.53 µg) (series of 7). The solid circles and vertical bars represent the means and standard deviations, respectively, of 8 tissues from 3 rats.

, The concentration-response curve of phenylephrine (PHE)-induced contraction of rat aortic rings in the presence of different concentrations of Ifanosine extract (IFN), as antagonist. , shows that there is a progressive shift in the concentration-response curve of phenylephrine in the presence of increasing concentration of IFN.

, The Schild plot of the PHE concentration-response curve in the presence of different IFN concentrations indicates that the pA ₂ value was 0.4 pg. The solid circles and vertical bars represent the means and standard deviations of 8 tissues from 3 rats, respectively.

, These figures show the results obtained from clinical trials in humans. Variations in biological parameters: diastolic and systolic blood pressure (DBP, PAS).

, These figures show the results obtained from clinical trials in humans on total cholesterol (TC), as well as Apoproteins (apoA, apoB), showing that Ifanosine (IFN) shows a clinical benefit superior to that of Prazosin.

, These figures show the results obtained by clinical trials in humans on total cholesterol (TC), lipoproteins (HDL, LDL), triglycerides as well as Apoproteins (apoA, apoB), showing that Ifanosine (IFN) shows a clinical benefit superior to that of Prazosin.

[Table 1]: Determination of apparent permeability (bioavailability, P _app ). Intestinal bioavailability or apparent permeability (Papp = J/Co) was determined during the steady state of transport (at 60-150 min). The table shows that the intestinal bioavailability (Papp) of Ifanosine (IFN) is higher not than Hyphaene thebaica extract (HT), but also than Olea europaea (OEL). But the natural products are less permeable than the purified chemical Prazosin (PRZ).

Advantages of the invention and applications

The invention presented here has several advantages and possible applications:

Association of an aqueous extract of the bare pericarp of the Hyphaene thebaica fruit and an extract of young leaves of Olea europaea for the treatment of high blood pressure or hyperlipidemia, said association being in a form suitable for oral administration, characterized in that:

1) for 1.6 grams of active ingredient of the formulated product we find 1 gram of Olea europaea extract and 0.6 grams of Hyphaene thebaica;

2) The intestinal bioavailability of the formulated product is superior to Olea europaea extract alone or to Hyphaene thebaica extract alone.

2) the clinical benefit of the formulated finished product is greater than that of Prazosin.

The formulated finished product (Ifanosine) is characterized in that the forms of use and presentation are in the form of: capsules, tablets, syrup, powder for syrup.

The formulated finished product (Ifanosine) may have non-therapeutic use as a dietary product to prevent hypertension and hyperlipidemia.

Claims (2)

Association of an aqueous extract of the bare pericarp of the Hyphaene thebaica fruit and an extract of young leaves of Olea europaea for the treatment of high blood pressure, said association being in a form suitable for oral administration, characterized in that: for 1.6 grams of active ingredient of the formulated product obtained, there is 1 gram of Olea europaea extract and 0.6 grams of Hyphaene thebaica. Association according to claim 1, characterized in that the forms of use and presentation of the invention are in the form of: capsules, tablets, syrup, powder for syrup.