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WO2011077455A2 - Mise au point d'une formulation médicamenteuse efficace à base d'herbes pour le traitement de la drépanocytose et procédé correspondant - Google Patents

Mise au point d'une formulation médicamenteuse efficace à base d'herbes pour le traitement de la drépanocytose et procédé correspondant Download PDF

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Publication number
WO2011077455A2
WO2011077455A2 PCT/IN2010/000840 IN2010000840W WO2011077455A2 WO 2011077455 A2 WO2011077455 A2 WO 2011077455A2 IN 2010000840 W IN2010000840 W IN 2010000840W WO 2011077455 A2 WO2011077455 A2 WO 2011077455A2
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WIPO (PCT)
Prior art keywords
extract
alcohol
carica papaya
sickle cell
wrightia
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Ceased
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PCT/IN2010/000840
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English (en)
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WO2011077455A3 (fr
Inventor
Vinod Domaji Rangari
Virendra Sukhadeo Borkar
Anwar Siraj Daud
Showkat Ahmad Patni
Jayant Janrao Thakre
Milind Pandurang Mane
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UNIJULES LIFE SCIENCES Ltd
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UNIJULES LIFE SCIENCES Ltd
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Publication of WO2011077455A3 publication Critical patent/WO2011077455A3/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18—Magnoliophyta (angiosperms)
    • A61K36/185—Magnoliopsida (dicotyledons)
    • A61K36/24—Apocynaceae (Dogbane family), e.g. plumeria or periwinkle
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • A61P7/06—Antianaemics

Definitions

  • the invention pertains to extraction of active ingredients from seeds of Wrightia tinctoria R. Br. and unripe fruits of Carica papaya L. and making- a nutraceutical formulation for treatment and management of Sickle cell disease.
  • Sickle cell disease is an inherited blood disorder that affects red blood cells.
  • People with sickle cell disease have red blood cells that contain mostly hemoglobin S, an abnormal type of hemoglobin. Hemoglobin is the main substance of the red blood cell. It helps red blood cells carry oxygen from the air in our lungs to all parts of the body.
  • Normal red blood cells contain hemoglobin A. Hemoglobin S and hemoglobin C are abnormal types of hemoglobin. Normal red blood cells are soft and round and can squeeze through tiny blood vessels. Normally, red blood cells live for about 120 days before new ones replace them. People with sickle cell conditions make a different form of hemoglobin called hemoglobin S (S stands for sickle). Red blood cells containing mostly hemoglobin S do not live as long as normal red blood cells (normally about 16 days). They also become stiff, distorted in shape and have difficulty in passing through the body's small blood vessels. When sickle-shaped cells
  • Sickle Cell Anemia SS
  • SC Sickle-Hemoglobin C Disease
  • AS Sickle Beta-Plus Thalassemia
  • Sickle Beta-Zero Thalassemia Sickle Cell trait
  • Health maintenance for patients with sickle cell disease starts with early diagnosis, preferably in the newborn period and includes penicillin prophylaxis, vaccination against Pneumococcus bacteria and folic acid supplementation. Treatment of complications often includes antibiotics, pain management, intravenous fluids, blood transfusion and surgery, all backed by psychosocial support. Like all patients with chronic disease, patients are best managed in a comprehensive multi-disciplinary program of care.
  • Blood transfusions help benefit sickle cell disease patients by reducing recurrent pain crises, risk of stroke and other complications. Because red blood cells contain iron, and there is no natural way for the body to eliminate it, patients who receive repeated blood transfusions can accumulate iron in the body until it reaches toxic levels. It is important to remove excess iron from the body, because it can gather in the heart, liver, and other organs and may lead to organ damage. Treatments are available to eliminate iron overload. Patients frequently experience a vicious circle of events called a "Sickle cell disease crisis", in which low oxygen tension in tissue causes sickling, which leads to ruptured RBC and further leads to serious decrease in RBC count and more sickling which often causes death.
  • Hydroxyurea was found to reduce the frequency of severe pain, acute chest syndrome and the need for blood transfusions in adult patients with sickle cell disease. Droxia, the prescription form of hydroxyurea, was approved by the FDA in 1998 and is now available for adult patients with sickle cell anemia.
  • the invention comprises a pharmaceutical dosage composition comprising anti-sickling effective amount of extract of seeds of Wrightia tinctoria R. Br. with or without combination with extract of raw fruits of Carica papaya L; and optional addition of pharmaceutically compatible excipients or/and diluents, and extracts or fractions from other plants.
  • the said extracts comprises a hydro-alcoholic extract.
  • the alcohol used in the hydroalcoholic extract may comprise, without limitation, methyl alcohol, ethyl alcohol, isopropyl alcohol, OF n-butyl alcohol, amyl alcohol, isoamyl alcohol or any other alcoholic solvent or a mixtures of alcohols or mixture of alcohols along with other polar solvents such as acetone or mixture of alcohol with less polar solvents such as diethyl ether, ethyl acetate, dichloromethane or toluene.
  • the pharmaceutical dosage composition of this invention comprises drying of the said hydroalcoholic extract. The said drying may be done in vaccum or by any other method of drying that preserves the antisickling activity of the extract.
  • the pharmaceutical dosage composition of this invention may comprise at least about 500 mg each of the extract of Wrightia tinctoria R. Br. or / and Carica papaya L.
  • this invention comprises a process of preparation of extract of this invention wherein: powdered seeds of Wrightia tinctoria R. Br. are macerated and extracted with aqueous alcohol, the extract separated from the residue.
  • the said alcohol may comprise, without limitation, methyl alcohol, ethyl alcohol, isopropyl alcohol, n-butyl alcohol, amyl alcohol, isoamyl alcohol or any other alcoholic solvent or a mixtures of alcohols or mixture of alcohols along with other polar solvents such as acetone or mixture of alcohol with less polar solvents such as diethyl ether, ethyl acetate, dichloromethane or toluene , the extract is separated from the residue by percolation or filtration, residual plant material optionally re-extracted, the filtrates of the plant extract combined, subjected to further concentration and dried by vaccum drying or other methods of drying.
  • Extract of Carica papaya L used in this invention is prepared from its raw fruit, which is peeled, cream coloured seeds inside the fruit discarded, the fruit cut into pieces, soaked in water and/or 0.9% sodium chloride solution and incubated at room temperature for 12 to 72 hrs. to get the extract.
  • Illustrative pharmaceutical dosage composition of this invention may comprise, per dosage form, 0.5 g extract of seeds of Wrightia tinctoria R. Br. with or without combination with 0.5 g extract of raw fruits of Carica papaya L.
  • the said dosage form may comprise, without limitation, a capsule, a tablet, a pill, an oral thin films, soft gel capsules, an elixir or a syrup suspension.
  • the said excipients may comprise one or more of a binder; an additive; a disintegration; a lubricant; a fliudifier; a sweetening agent and a flavoring agent.
  • the said binder may comprise microcrystalline cellulose, tragacanth or gelatin; the said additive may comprise starch or lactose; the said disintegration agent may comprise alginic acid, primogel, maize starch; lubricants such as magnesium stearate; the said fliudifier may comprise colloidal silicon oxide, the said sweetening agent comprises sucrose or saccharine; the said flavouring agent comprises a mint flavor, an orange flavor of methyl salicylate.
  • This invention also comprises a process/method of treatment of Sickle cell disease for treatment of Sickle cell anemia comprising administering a person in need thereof of two dosage compositions per day comprising, per dosage form, 0.5 g extract of seeds of Wrightia tinctoria R. Br. with or without combination with 0.5 g extract of raw fruits of Carica papaya L.
  • the invention also comprises a process of making a medicament that comprises per dosage form, 0.5 g extract of seeds of Wrightia tinctoria R. Br. with or without combination with 0.5 g extract of raw fruits of Carica papaya L.
  • the said sickle cell disease may comprise, without limitation, sickle cell anemia (SS), Sickle-hemoglobin C Disease (SC), Sickle beta-plus thalassemia, or sickle beta-zero thalassemia.
  • SS sickle cell anemia
  • SC Sickle-hemoglobin C Disease
  • Sickle beta-plus thalassemia Sickle beta-zero thalassemia.
  • the illustration of the pharmaceutical dosage compositon of this invention is done using extracts of Wrightia tinctoria R. Br. or / and Carica papaya L
  • the extracts or fractions from other plants including one or more of other species of the genus Wrightia having antisickling or complementary activity or an extract or fraction of other plant species that may have an antisickling activity or other medicinal activity may also be included as a combi-preparation.
  • the objective of the research is to perform extraction, formulation and antisickling activity studies with reference to treatment and management of Sickle cell disease.
  • Another object of the present invention is to provide a method for the preparation of the nutraceutical formulation.
  • Yet another object of the invention includes determination of in vitro antisickling potential of extracts from seeds of Wrightia tinctoria R. Br., and unripe fruits of Carica papaya L. Taking into consideration their therapeutic activity against Sickle Cell Anemia (SCA), it was envisaged to develop a nutraceutical supplement from the extracts of Wrightia tinctoria R. Br. and Carica papaya L. for the treatment and management of SCA and conduct in vivo studies of clinical trial on volunteers with the help of pathological laboratory test.
  • SCA Sickle Cell Anemia
  • Further object of the present invention is to use the extracts of Wrightia tinctoria R. Br. and Carica papaya L. preparation for the treatment and management of SCA as well as the pharmaceutical composition containing effective dosages of one or more medicinal plants or their extracts /fractions, in the formulation with suitable excipients and/or diluents.
  • Antisickling Activity is expressed in terms of inhibitory activity of the extract on sodium metabisulphite induced sickling activity as determined by method described by Thomas and Ajani, 1987 (Thomas, K.D.; Ajani, B.; Antisickling agent in an extract of unripe pawpaw fruit (Carica papaya); Transactions of the Royal Society of Tropical Medicine and Hygiene; 81 : 510-11 , 1987).
  • the process of extraction of the antisickling activity of this invention in hydroalcoholic extract comprised extraction of seeds of Wrightia tinctoria R. Br. by maceration, percolation and other suitable extraction process.
  • the powdered seeds of Wrightia. tinctoria R. Br. were extracted by hydroalcoholic (10% - 95% alcohol and balance is water.) maceration in stainless steel vessels, provided with a stirrer.
  • the said alcohol used was methyl alcohol, ethyl alcohol, isopropyl alcohol, n-butyl alcohol, amyl alcohol, isoamyl alcohol or any other alcoholic solvent or a mixtures of alcohols or mixture of alcohols along with other polar solvents such as acetone or mixture of alcohol with less polar solvents such as diethyl ether, ethyl acetate, dichloromethane or toluene.
  • the extract was filtered and residual plant material re-extracted in the same way. The residual plant material was pressed using fine muslin cloth for the recovery of absorbed solvent and the extract was filtered. The filtrates of the plant extract were combined and subjected to further concentration and drying.
  • the concentrated extract of Wrightia tinctoria R. Br. (WTE) was collected, vacuum dried and stored in air tight containers.
  • the extract of unripe fruit of Carica papaya L was used as a standard.
  • the fresh unripe papaya was plucked, peeled and the cream colour seeds inside the fruit were discarded.
  • the papaya fruits were cut into pieces and soaked in water and/or physiological saline (0.9% sodium chloride solution) and incubated at room temperature for 12 to 72 hrs.
  • the extract was sieved, collected and stored into bottles. Extract of unripe fruits of Carica papaya L. (CPE) was concentrated, vacuum dried and stored in air tight containers.
  • CPE Carica papaya L. extract
  • CD Crohn's Disease
  • the pharmaceutical formulation of the present invention may contain therapeutically antisickling effective amounts of the Wrightia tinctoria R. Br. (WTE) extract and Carica papaya L. extract (CPE) or a combination thereof, optionally with pharmaceutically compatible excipients, and may also contain additional extracts or fractions from other medicinal plants.
  • the oral formulation will generally include inert diluents. It may be filled in gelatin capsules or compressed into tablets.
  • the oral administration dosage forms may be capsules, tablets, pills, oral thin films, soft gel capsules, an elixir or a syrup suspension.
  • Capsules, tablets, pills, and similar compositions may, without limitation, contain the following ingredients in addition to the active extracts: a binder such as microcrystalline cellulose, tragacanth or gelatin; an excipient such as starch or lactose; a disintegration agent such as alginic acid, primogel, maize starch and the like; lubricants such as magnesium stearate; a fliudifier such as colloidal silicon oxide; sweetening agent such as sucrose or saccharine or a flavouring agent such as mint flavour, orange flavour of methyl salicylate.
  • a binder such as microcrystalline cellulose, tragacanth or gelatin
  • an excipient such as starch or lactose
  • a disintegration agent such as alginic acid, primogel, maize starch and the like
  • lubricants such as magnesium stearate
  • a fliudifier such as colloidal silicon oxide
  • sweetening agent such as sucrose
  • the Wrightia tinctoria R. Br. has shown remarkable potential as evident from the result of its in vitro antisickling activity studies and case studies on Human volunteers.
  • the formulation prepared form the extracts of Wrightia tinctoria R. Br. and/or unripe fruit of Carica papaya L. was promising for the treatment and management of sickle cell disease especially in cases of sickle cell anemia (SS), Sickle-hemoglobin C Disease (SC), Sickle beta-plus thalassemia and sickle beta-zero thalassemia.
  • the antisickling activity may be common to all species of genus Wrightia and all of them may be useful for antisickling activity, although species specific differences in quantum of activity may exist.
  • extract of Wrightia tinctoria R. Br. may be substituted by any other species of the genus Wrightia.
  • Determining antisickling activity was based on measuring the effect of the known concentration of an extract of a plant under test on sodium metabisulphite induced sickling of HbSS red blood cells, collected from confirmed non-crises sickle cell patients, in terms of number of Sickle cells with help of a Heamocytometer slide, wherein Carica papaya L. extract (CPE) is used as a standard and normal saline as controls.
  • CPE Carica papaya L. extract
  • the vital organs viz. kidney, spleen, heart, and liver were separated (weighed) and studied for the histopathology parameters. Rest 50% of the animals in each groups have been sacrificed on 30 th day for the blood hematology (Hb, TG, DC, WBC, etc) and blood biochemistry for investigation of the toxicity of the drugs, if any.
  • the vital organs viz. kidney, spleen, heart, and liver were separated, weighed and kept for the histopathology in the section. Histopathological studies of the separated organs viz. kidney, spleen, heart, and liver has revealed no abnormal feature in tissue study.
  • the method employed for antisickling activity involves inhibition of sodium metabisulphite induced sickling of red blood cells collected from confirmed non-crises sickle cell patients.
  • a drop of blood from a sickle cell patient (SS) was mixed with a drop of freshly prepared 2% sodium metabisulphite on a clean slide, mixed well and cover slipped.
  • the cover slip was gently pressed to remove excess mixture.
  • the excess mixture was removed with cotton wool and the edges of the cover slip sealed with vaseline to prevent air from going in.
  • the slide was incubated at 37°C for 30 min and then viewed under microscope. This sample was termed as 'Control'.
  • a drop of blood from a sickle cell anemia patients (SS) and a drop of freshly prepared 2% sodium metabisulphite were mixed together on a clean glass slide and then a drop of Carica papaya L. Extract (CPE) was added to the mixture on the slide and cover slipped. The edges were sealed with vaseline after gentle pressing to remove excess mixture. The slide was incubated at 37°C for another 30 min. The slide was viewed under microscope. These samples were termed as 'Standard'. The above process was repeated for the test samples.
  • a drop of blood from a sickle cell anemia patients (SS) and a drop of freshly prepared 2% sodium metabisulphite were mixed together on a clean glass slide and then a drop of Wrightia tinctoria R. Br.
  • Extract WTE
  • CPE Carica papaya L. Extract
  • the Neubers slide consists of two identical ruled areas separated by empty space and having two elevated ridges on both the sides. Either of the ruled areas is used for counting cells. Each has a square of 3 mm x 3 mm. 3 mm which is ruled into equal 9 square of 1 mm 2 each.
  • the 4 corners square are for WBC and a central square for RBC, which was used for counting of sickled RBCs,
  • the central square is divided into 25 larger squares of 0.2 mm which are further subdivided into 16 smaller square with an area of 1 /400 th of 1 mm 2 and depth of each small square is 0.1 mm or 1/10 mm.
  • CPE unripe Carica papaya L.
  • the slides of the two extracts namely Wrightia tinctoria R. Br.
  • hydro-alcoholic extract WTE
  • Carica papaya L. extract CPE
  • WTE hydro-alcoholic extract
  • CPE Carica papaya L. extract
  • the extracts were used in combination and their effects were observed in the sickling test.
  • the extracts were made in combinations coded as CPE, WTE and WTE + CPE. All these extract were taken in concentration of 10mg/10ml and the sickling test was performed and compared with control.
  • the procedure used for preparation of control slide is same as above in sickling test. All the RBCs in a slide with 2% sodium metabisuphite with no extracts sickled within 30 min. In the slides containing mixture of 2% sodium metabisuphite with standard Carica papaya L. extract (CPE) in saline has shown maximum antisickling activity. Similar slides as described above were prepared and sickling test of different extracts were performed.
  • CPE Carica papaya L. extract
  • the number of sickled cell in control slide was considered as 100%.
  • percentage of synergistic antisickling activity of different combinations of Wrightia tinctoria R. Br. extract (WTE) and Carica papaya l. extract (CPE) were calculated. All these different concentrations of the two extracts exhibited significant antisickling activity in a dose dependent manner; that is, the activity was at lowest detectable level in a dose of 1mg/10ml but the maximum antisickling activity was achieved in the 10 mg/ml or 0.1 ml/100ml of stock solution (Table-1).
  • the common nutrient materials such as Wrightia tinctoria R. Br. seeds and Carica papaya L. fruits, are the potential herbal drugs which can be used for the treatment of Sickle cell disease.
  • a nutraceutical formulation of these drugs in combination is expected to give good result in Sickle cell anemia patients.
  • Complete Blood Count (CBC) studies showed that the treatment with the nutraceutical formulation comprising of Wrightia tinctoria R. Br. and/or Carica papaya L. extracts might help treat Sickle cell anemia.
  • Case studies were performed on the Sickle cell anemia patients to find out the efficacy of the nutraceutical drug formulation.
  • CBC Complete Blood Count
  • CBC Complete Blood Count
  • CD Crohn's Disease
  • Tables 4 shows the results of case studies/case reports of the administration of the nutraceutical formulation of the invention on Sickle Cell anemia patients.
  • Table 4 Complete Blood Count (CBC) Study Report of the nutraceutical formulation on patients in case studies before and 5 months after treatment .
  • Patient 1 and Patient 3 received 2 capsules daily, one of Wrightia tinctoria R. Br. extract (500 mg per capsule) and one Carica papaya L. extract (1 g per capsule) and Patient 2 ** received only Wrightia tinctoria R. Br. extract (500 mg per capsule) twice a day.
  • Table 5 Complete Blood Count (CBC) Study Report of the nutraceutical formulation on patients in case studies before treatment and at one month interval thereafter upto 5 th month.
  • BT Complete blood count before treatment
  • AT Complete blood count after treatment.
  • CD Crohn's Disease

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Abstract

L'invention concerne une composition de formes posologiques comprenant une quantité efficace antidrépanocytique d'extrait de graines de Wrightia tinctoria R. Br. combinée ou non avec un extrait de fruits verts de Carica papaya L, un procédé de fabrication de cette composition et son utilisation dans le traitement de la drépanocytose.
PCT/IN2010/000840 2009-12-22 2010-12-22 Mise au point d'une formulation médicamenteuse efficace à base d'herbes pour le traitement de la drépanocytose et procédé correspondant Ceased WO2011077455A2 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
IN2948/MUM/2009 2009-12-22
IN2948MU2009 2009-12-22

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WO2011077455A2 true WO2011077455A2 (fr) 2011-06-30
WO2011077455A3 WO2011077455A3 (fr) 2011-08-11

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PCT/IN2010/000840 Ceased WO2011077455A2 (fr) 2009-12-22 2010-12-22 Mise au point d'une formulation médicamenteuse efficace à base d'herbes pour le traitement de la drépanocytose et procédé correspondant

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108686196A (zh) * 2018-08-21 2018-10-23 叶文文 小儿复气汤、其配方及配制方法

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US8383166B2 (en) * 2007-12-03 2013-02-26 Sequent Scientific Limited Stable hydrophobic topical herbal formulationn

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108686196A (zh) * 2018-08-21 2018-10-23 叶文文 小儿复气汤、其配方及配制方法

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