Field of the Invention

The present invention relates to an oral pharmaceutical composition and process for its preparation thereof. The composition is a combination of specific amino acid with plant extract(s) which is useful for treating dengue.

Background of the Invention
Dengue is emerging as a major health problem in tropical countries and is drawing the attention of government and all health agencies due to its fatality and limitations of treatment in conventional system.
Dengue is caused by the “aedes aegy mosquito” which breeds in clean water. The symptoms of dengue include high fever, joint pains, headache, nausea, vomiting, dip in blood pressure along with a characteristic skin rash. Though the fever does not last beyond a week, in some cases there is severe drop in the level of platelets which turn fatal.
The incidence of dengue has grown dramatically around the world in recent decades. A vast majority of cases are asymptomatic and hence the actual numbers of dengue cases are underreported and many cases are misclassified. One estimate indicates 390 million dengue infections per year (95% credible interval 284–528 million), of which 96 million (67–136 million) manifest clinically (with any severity of disease). Another study, of the prevalence of dengue, estimates that 3.9 billion people, in 128 countries, are at risk of infection with dengue viruses. India in 2017 has seen 15,7220 cases of dengue and 250 deaths due to the disease, as per the National Vector Borne Disease Control Programme (NVBDCP) data. In 2016, the country registered 129166 cases and 245 deaths.
Apart from eliminating the mosquitos, work is ongoing for medication targeted directly at the virus. It is classified as a neglected tropical disease. In the modern era of medicine, herbal materials and plants continue to play an important role in drug discovery and development. The demand for plant-based medicines is ever growing since crude or processed products obtained from plants are believed to have fewer or no adverse effects as compared to the drugs that are synthetic small molecules.

It is well known in the art that certain plant herbs exhibit anti-dengue or antiviral activity. The EP Patent No. EP2389184 relates to the anti-dengue activity of the Cissampelos pareira extracts.

CN110123921 patent discloses an ethanol extract of banana leaves have some inhibition of dengue virus action.

MY142050 discloses a plant derived compound; piperine, for treating flavivirus infection, in particular dengue viral infection. It further discloses that piperine can be applied as a non-peptide protease inhibitor which specifically acts on dengue virus serotype 2 NS2B/3 protease complexes and hence piperine can be used as the dengue protease inhibitors for treatment of dengue viral infection.

IN2011MU00840 claims a process for preparation of antiviral herbal composition which helps in treatment of hepatitis (A, B, C, D, E, F), viral fever (chicken guinea, dengue, swine flu), whooping cough, warts on skin as well as herpes zoaster.

IN2014MU00411 discloses novel formulation syrup consisting of extracts of Papaya Leaves (Carica papaya), Tulsi leaves (Ocimum sanctum), Nimba or Neem leaves (Azidarachta indica) and Kalmegh (Andrographis paniculata). The syrup is an adjuvant for the treatment of the Vishama Jwara (Dengue fever) with antithrombocytopenic and antiviral properties.

IN201811017566 relates to a formulation comprising Tinosporia Cordifolia; Syzygium Cumini; Psidium Guajava; Carica Papaya; Piper Nigrum; Ricinus Communis; Glycrrhiza Glabra; Calotropis Gigantea; Achyranthes Aspera; Jaggery; and Ocimum Sanctum. The formulation is used for increasing blood platelet counts in a body.

WO2006/067537 pertains to a synergistic antipyretic formulation comprising extracts of plants Berberis aristata, Tinospora cordifolia, Alstonia scholaris, Andrographis paniculata and Hedychium spicatum, optionally alongwith pharmaceutically acceptable additives; its use in treating fever, and lastly, a process for the preparation said synergistic antipyretic formulation.

Asian Pac J Trop Biomed 2011; 1(4): 330-333; Nisar Ahmad et. al., “Dengue fever treatment with Carica papaya leaves extracts” reported that Carica papaya leaves aqueous extract exhibited potential activity against Dengue fever.

Earth and Environmental Science 101 (2017) 012005, M Ichsyani et. al., “Antiviral effects of Curcuma longa L. against dengue virus in vitro and in vivo” revealed that C. longa could potentially be used as antiviral against DENV with low cytotoxicity and effective inhibition.

Pharmacognosy Journal, Volume: 9 Issue: 6 Pages: 873-879, Rahmani Arshad Husaini et al.; “Saffron (Crocus sativus) and its Active Ingredients: Role in the Prevention and Treatment of Disease” summarizes the study of saffron and its ingredients based on in vivo and in vitro and role in disease cure and prevention.

It has been indicated herein above that considering the growing prevalence of dengue, there exist a continuing need for new compositions and methods for the effective treatment of the said disease. In fact, efforts of the inventors of the present invention directed to find a solution to these problems have resulted in a pharmaceutical composition comprising specific amino acid with plant extracts, which has been found to be useful for the treatment of dengue.

Summary of the Invention
In one aspect of the present invention, there is provided an oral pharmaceutical composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acids and optionally one or more pharmaceutical excipient(s).

In yet another aspect there is provided a process for preparation of an oral pharmaceutical composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acids and optionally one or more pharmaceutical excipient(s).

In an aspect, there is provided use of the pharmaceutical composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s) for the treatment of dengue.

In another aspect, there is provided a method for treating or preventing of dengue, comprising administering to the subject, an oral pharmaceutical composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s).

In another further aspect, there is provided use of an oral pharmaceutical composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s), for the manufacture of a medicament for treating or preventing dengue.

In a still further aspect, there is provided a pharmaceutical kit comprising: (a) therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s); and (b) optionally a package insert comprising instructions for using the said pharmaceutical composition.

These and other aspects and advantages of the present invention will be apparent to those skilled in the art from the following description.

Brief Description of Drawings of the Invention
Figure 1 represents change in the platelet counts of the patient shown in Table 2.
Figure 2 represents change in the platelet counts of the patient shown in Table 3.

Detailed Description of the Invention
It should be understood that the detailed description and specific examples, while indicating embodiments of the invention, are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art. One skilled in the art, based upon the description herein, may utilize the present invention to its fullest extent. The following specific embodiments are to be construed as merely illustrative, and not limitative of the remainder of the disclosure in any way whatsoever.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention belongs.
Definitions
For the purpose of the disclosure, listed below are definitions of various terms used to describe the present invention. Unless otherwise indicated, these definitions apply to the terms as they are used throughout the specification and the appended claims, either individually or as part of a larger group. They should not be interpreted in the literal sense. They are not general definitions and are relevant only for this application.

It should be noted that, as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the content clearly dictates otherwise.

It should be noted that the term “or” is generally employed in its sense including “and/or” unless the content clearly dictates otherwise.

As used herein, the term "about" means approximately and in the context of numerical values the term “about” can be construed to estimate a value that is ±10% of the value or range recited.

The term "dengue" refers to a mosquito-borne viral disease occurring in tropical and subtropical areas. The virus responsible for causing dengue, is called dengue virus (DENV) and is spread by several species of female mosquitoes of the Aedes type, principally A. aegypti. There are four types of DENV serotypes (DENV1, DENV2, DENV3 and DENV4).

The term “excipient(s)” as used herein means a diluent, binder, disintegrant, glidant, lubricant, coating material or the like, which is non-toxic, and inert, which does not have undesirable effects on a subject to whom it is administered and is suitable for delivering a therapeutically active agent to the target site without affecting the therapeutic activity of the said agent.

The term “treating”, “treat” or “treatment” as used herein includes preventive (prophylactic) and palliative treatment.

The terms “pharmaceutical composition” or “composition” are used interchangeably and may refer to a composition comprising therapeutically effective amount of the plant extracts selected from the group consisting of Curcumin longa (Haldi), Thinopyrum intermedium (Wheatgrass) with more chlorophyll content, Carica papaya (Papaya), Crocus sativus L. (Saffron), Picrorhiza kurroa (Katuka) and Andrographis paniculata (Kalmegh); therapeutically effective amount of amino acids, i.e. L-Arginine, L-Tyrosine and one or more pharmaceutical excipient(s). It should be noted that the term "composition" should be construed in a broad sense and includes any composition which is intended for the purpose of achieving a therapeutic effect whether sold as a pharmaceutical product, for example carrying a label as to the intended indication, whether sold over the counter, or whether sold as a phyto-pharmaceutical.

The term “therapeutically effective amount” as used herein means an amount of the plant extracts and amino acid or the composition containing the extract, which is sufficient to significantly induce a positive modification in the condition to be regulated or treated, but low enough to avoid side effects, if any (at a reasonable benefit/risk ratio), within the scope of sound medical judgment. The therapeutically effective amount of the extract or composition will vary with the particular condition being treated e.g. dengue, the age and physical condition of the end user, the severity of the condition being treated/prevented, the duration of the treatment, the nature of concurrent therapy, the particular pharmaceutically acceptable carrier utilized, and like factors. As used herein, all percentages are by weight unless otherwise specified.

The term "subject" as used herein refers to an animal, particularly a mammal, and more particularly, a human. The term “mammal” used herein refers to warm-blooded vertebrate animals of the class Mammalian, including humans, characterized by a covering of hair on the skin and, in the female, milk-producing mammary glands for nourishing the young. The term mammal includes animals such as cat, dog, rabbit, bear, fox, wolf, monkey, deer, mouse, pig and the human.

In an embodiment, there is provided an oral pharmaceutical composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s).

In another embodiment, there is provided an oral pharmaceutical composition comprising of therapeutically effective amount of plant extracts selected from the group consisting of Curcumin longa (Haldi), Thinopyrum intermedium (Wheatgrass) with more chlorophyll content, Carica papaya (Papaya), Crocus sativus L. (Saffron), Picrorhiza kurroa (Katuka) and Andrographis paniculata (Kalmegh); therapeutically effective amount of amino acids, i.e. L-Arginine, L-Tyrosine and one or more pharmaceutical excipient(s).

In an embodiment, the pharmaceutical composition of the present invention optionally contains probiotics.

In an embodiment, there is provided a process for preparation of an oral pharmaceutical composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s).

Process for preparation of pharmaceutical composition:

In an embodiment, there is provided a process for preparation of an oral pharmaceutical composition comprising the steps of:
(i) placing amino acid and/or plant extracts (about 2000-3000 g of mixture) in a high energy planetary ball mill;
(ii) grinding the mixture formed in step (i) for about 1 hour until the particle size is reduced to about 0.1 to 1 µm using a disc speed upto 200 rpm (rotations per minute);
(iii) heating the resulting mixture obtained in step (ii) at a temperature raising from 100oC to 600oC to yield a fine homogeneous powder in a quantity of about 140-200 g;
(iv) optionally combining the homogeneous powder with one or more pharmaceutical excipient(s) to obtain the pharmaceutical composition; and
(v) optionally filling the pharmaceutical composition of step (v) in a capsule or optionally
compressing the pharmaceutical composition of step (v) into a tablet.

In an embodiment, the amino acid used in step (i) above is L-Arginine and L-Tyrosine.

In an embodiment, the plant extracts used in step (i) above are selected from the group consisting of Curcumin longa (Haldi), Thinopyrum intermedium (Wheatgrass), Carica papaya (Papaya), Crocus sativus L. (Saffron), Picrorhiza kurroa (Katuka) and Andrographis paniculata (Kalmegh).

The ingredients used in pharmaceutical composition are tabulated in Table 1 below:

Table 1

Ingredients Quantity (gm)
Curcumin longa/Haldi 300-600
Thinopyrum intermedium (Wheatgrass) extract
With higher chlorophyll content 50-200
Carica papaya (Papaya) extract 50-150
Crocus sativus L. (Saffron) extract 5-50
Picrorhiza kurroa (Katuka) extract 50-100
Andrographis paniculata (Kalmegh) extract 100-200
L-Arginine 100-300
L-Tyrosine 50-100

In an embodiment, the high energy planetary ball mill used in the process of preparing the compounds results in fusion and homogenization of the particles without any contamination.

In an embodiment, the present invention relates to use of the pharmaceutical composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s) for the treatment of dengue; wherein the said composition is as described in one or more embodiments of the present invention as described herein above.

In an embodiment, the present invention relates to a method for treating or preventing of dengue, comprising administering to the subject, an oral pharmaceutical composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s); wherein the said composition is as described in one or more embodiments of the present invention as described herein above.

In an embodiment, the present invention relates to use of an oral pharmaceutical composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s), for the manufacture of a medicament for treating or preventing dengue; wherein the said composition is as described in one or more embodiments of the present invention as described herein above.

In an embodiment, the composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s), may be packaged in a suitable container depending upon the formulation and the method of administration of the composition. Suitable containers known to a person skilled in the art include blister pack or bottle pack.

In another embodiment, the present invention provides a pharmaceutical kit comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s). The kit may further comprise a package insert, including information about the indication, usage, doses, direction for administration, contraindications, precautions and warnings.

The pharmaceutical composition of the present invention may be formulated for oral administration by compounding the active ingredient i.e. the plant extract(s) and amino acid with the usual non-toxic pharmaceutically acceptable excipient(s) for powders, pills, tablets, coated tablets, pellets, granules, capsules, and any other form suitable for use. Formulations of the present invention encompass those which include talc, water, glucose, lactose, sucrose, gum acacia, gelatin, mannitol, starch paste, magnesium trisilicate, corn starch, keratin, colloidal silica, potato starch, urea, and cellulose and its derivatives such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; malt; gelatin; as well as other non-toxic compatible lubricants such as sodium lauryl sulfate and magnesium stearate, releasing agents, coating agents and other excipients suitable for use in manufacturing preparations, in solid, semisolid or liquid form and in addition auxiliary, stabilizing, thickening and coloring agents may be used. For preparing solid compositions such as tablets or capsules, the extract is mixed with a pharmaceutical carrier (e.g., conventional tableting ingredients such as corn starch, lactose, sucrose, sorbitol, talc, stearic acid, magnesium stearate, dicalcium phosphate or gums) and other pharmaceutical diluents (e.g., water) to form a solid composition. This solid composition is then subdivided into unit dosage forms containing an effective amount of the composition of the present invention. The tablets or pills containing the extract can be coated or otherwise compounded to provide a dosage form affording the advantage of prolonged action.

In an embodiment, the pharmaceutical composition is filled in capsules according to the desired dosage, depending upon the severity of the condition of the patient.

The selected dosage level will depend upon a variety of factors including the activity of the particular extract of the present invention employed, the route of administration, the time of administration, the rate of excretion of the particular composition being employed, the duration of the treatment, used in combination with the other extracts, the age, sex, weight, condition, general health and prior medical history of the patient being treated, and like factors well known in the medical arts. In general, however, doses employed for human treatment will typically be in the range of 1-1500 mg per day. In any case the required dose may be increased or decreased depending on the severity of the disease and the other parameters by the medical practitioner. For example, the doses in which the composition can be used may be 10 – 1000 mg/day or 5 -500 mg/day or any other suitable dose. The desired dose may conveniently be presented in a single dose or as divided doses administered at appropriate intervals, for example as one, two or three sub-doses as required per day.

The present invention will be more readily understood by referring to the following examples which are given to illustrate the invention but do not limit its scope.

Examples
The plant extracts described herein above have been procured from the vendor. The details of the vendor are provided in the table below:
Name: LAVANYA AGRO INDUSTRIES
Address: Off.: Flat No. 105, First Floor, “Gupta Plaza”, Plot No. 13, Site No. 37 & 38, Local Shopping Centre, Kalkaji, New Delhi – 110019
Sf9 cells

Example 1
Preparation of pharmaceutical composition:
Ingredients Quantity (gm)
Curcumin longa/Haldi 492
Thinopyrum intermedium(Wheatgrass) extract (10:1) with higher chlorophyll content 123
Carica papaya (Papaya) extract (10:1) 92
Crocus sativus L. (Saffron) 38.5
Picrorhiza kurroa (Katuka) extract (10:1) 80
Andrographis paniculata (Kalmegh) extract (10:1) 150
L-Arginine 192
L-Tyrosine 60

The ingredients of Example 1 are mixed in a mill to obtain a homogeneous mixture. This mixture is then converted to the desired pharmaceutical composition which may be in the form of a capsule or a tablet by combining optionally with one or more conventional pharmaceutical excipient(s).

Example 2
Effect of pharmaceutical composition on patients diagnosed with dengue.

Study 1:
The composition of Example 1 was administered to a patient at a dose of 50 mg, thrice daily.
The treatment was continued for a period of four days. The platelet count was monitored and the increase in the platelets is demonstrated in Table 2.

Table 2: Effect of the composition on platelet count of the patient
Test
Values at different time points
(Unit: 103/µL)
Initial (prior to medication) After 1 day (on medication) After 2 days (on medication)
Platelet count (PLT)
129 166 225

Reference range:
Platelet count (PLT)

Average Count
(Unit: 103/µL)
Normal value 150-400

Conclusion: The treatment of patient with the composition of Example 1 caused significant increase in platelet count within a period of four days.

Study 2:
The composition of Example 1 was administered to a patient at a dose of 50 mg, thrice daily.
The treatment was continued for a period of four days. The platelet count was monitored and the increase in the platelets is demonstrated in Table 3.

Table 3: Effect of the composition on platelet count of the patient
Test
Values at different time points
(Unit: 103/µL)
Initial (prior to medication) After 1 day (on medication) After 3 days (on medication) After 4 days (on medication)
PLT
30 50 120 209

Conclusion: The treatment of patient with the composition of Example 1 caused significant increase in platelet count within a period of four days.

CLAIMS:We Claim:
1. An oral pharmaceutical composition comprising of therapeutically effective amount of plant extract(s), therapeutically effective amount of amino acid and optionally one or more pharmaceutical excipient(s).

2. The composition as claimed in claim 1, wherein the plant extracts are selected from the group consisting of Curcumin longa, Thinopyrum intermedium with more chlorophyll content, Carica papaya, Crocus sativus L., Picrorhiza kurroa and Andrographis paniculata.

3. The composition as claimed in claim 1, wherein the amino acids are selected from the group consisting of L-Arginine and L-Tyrosine.

4. A process for preparation of an oral pharmaceutical composition of claim 1 comprising the steps of:
(i) placing amino acid and/or plant extracts (about 2000-3000 g of mixture) in a high energy planetary ball mill;
(ii) grinding the mixture formed in step (i) for about 1 hour until the particle size is reduced to about 0.1 to 1 µm using a disc speed upto 200 rpm (rotations per minute);
(iii) heating the resulting mixture obtained in step (ii) at a temperature raising from 100oC to 600oC to yield a fine homogeneous powder in a quantity of about 140-200 g.
(iv) optionally combining the homogeneous powder with one or more pharmaceutical excipient(s) to obtain the pharmaceutical composition; and
(v) optionally filling the pharmaceutical composition of step (v) in a capsule or optionally
compressing the pharmaceutical composition of step (v) into a tablet.

5. Use of an oral pharmaceutical composition comprising of therapeutically effective amount of plant extracts, therapeutically effective amount of amino acids and optionally one or more pharmaceutical excipient(s) for the treatment of dengue.

6. A method of treating dengue, comprising administering to a subject in need thereof a therapeutically effective amount of plant extracts, therapeutically effective amount of amino acids and optionally one or more pharmaceutical excipient(s).

7. Use of an oral pharmaceutical composition comprising of therapeutically effective amount of plant extracts, therapeutically effective amount of amino acids and optionally one or more pharmaceutical excipient(s), for the manufacture of a medicament for treating dengue.