Pharmaceutical formulations and processes for their manufacture

Abstract

The present disclosure relates to a pharmaceutical formulation and a process for its preparation. The pharmaceutical formulation comprises cinnamic acid and at least one excipient. The pharmaceutical formulation of the present disclosure has improved patient compliance and reduced side effects. The pharmaceutical formulation of the present disclosure can be used in the treatment of cyclophosphamide-induced neutropenia. The present disclosure further relates to a process for the preparation of cinnamic acid. The process is simple and economical.

Description

[Technical field]

[0001] Field The present disclosure relates to pharmaceutical formulations and processes for their preparation.

[0002] definition As used in this disclosure, the following terms are generally intended to have the meanings set forth below, except to the extent the context in which they are used dictates otherwise.

[0003] MMAD (mass median aerodynamic diameter) then refers to the diameter, by mass, of which 50% of the particles of an aerosol are larger and 50% are smaller. FPF (fine particle fraction) refers to the fine particle dose divided by the total emitted dose. First-pass metabolism refers to the phenomenon whereby a drug is metabolized at a specific location in the body, resulting in a reduced concentration of the active drug when it reaches the site of action or the systemic circulation.

[0004] D10 refers to the percentage of particles with a diameter below the specified value being 10%. D50 refers to the percentage of particles with a diameter smaller than the specified value and larger than the specified value that is 50%. Also known as the median diameter. D90 refers to the percentage of particles having a diameter below a specified value of 90%.

[0005] F-MELT® Type C is a proprietary formulation of carbohydrate, disintegrant, and inorganic ingredients designed as a premix for orally disintegrating tablets (ODT). GCSF, granulocyte colony-stimulating factor (G-CSF or GCSF), also known as colony-stimulating factor 3 (CSF3), is a glycoprotein that stimulates the bone marrow to produce and release granulocytes and stem cells into the bloodstream.

[0006] An FD&C color is a synthetic color additive permitted for use in food, drugs, and cosmetics in qualified batches by the Federal Food, Drug, and Cosmetic Act of 1938 and subsequent statutes. Pure Picrside 1 is a naturally occurring iridoid glycoside and is obtained from the plant, Picrorhiza kurroa.

[0007] Picroside 1 with salt is obtained by forced alkali hydroxide decomposition of naturally occurring Picroside 1. The salt remains intact in the decomposed form. Picroside 1 without salt is a cinnamic acid according to the present disclosure, obtained by forced alkaline hydroxide decomposition of naturally occurring picroside 1. The salt is removed in this decomposed form by using a second fluid medium (methanol). [Background technology]

[0008] background The following background information herein is relevant to the present disclosure but is not necessarily prior art.

[0009] 3-Phenyl-2-propenoic acid, commonly called cinnamic acid, is a white crystalline solid with a low density and a sweet, honey-like aroma. Cinnamic acid and its derivatives are widely distributed in various fruits, vegetables, and flowers. Cinnamic acid is found as both trans-cinnamic acid and cis-cinnamic acid. The more stable isomer is the trans-isomer, which occurs naturally and is a common commercial product. Cinnamic acid acts as an antioxidant, antibacterial agent, healing agent, antifungal agent, and anticancer agent, etc. It is used as a precursor for the synthesis of thermoplastics and flavoring agents, and is also widely used in cosmetics and health foods.

[0010] In the current scenario, cancer is the most lethal cause of death worldwide and a highly prominent disease worldwide, affecting millions of people mainly in the United States, Europe, and other regions. Currently, there are numerous available treatments for the prevention and / or treatment of malignant cancer cells, including chemotherapy, hormonal therapy, immunotherapy, radiation therapy, surgery, and targeted therapy. Among them, chemotherapy is more prevalent and is frequently used for the purpose of treating cancer cells. Cyclophosphamide, an alkylating chemotherapy agent, is a widely used drug in the clinic to treat malignant and non-malignant tumors. However, despite its wide spectrum of clinical applications, it exhibits severe cytotoxicity to normal cells in both humans and experimental animals. Its metabolites can interact with cellular macromolecules such as proteins, membrane lipids, RNA, and DNA, and induce apoptosis. One of its metabolites, i.e., acrolein, causes oxidative stress leading to DNA damage in normal cells and causes toxicity in various organs. One of the most severely affected sites is the hematopoietic compartment of the bone marrow. Indeed, neutropenia is the most common and frequent side effect of cytotoxic chemotherapy. Other treatments are more costly and have serious and / or life-threatening adverse events, such as anemia, anorexia, thrombocytopenia, alopecia, and peripheral neuropathy.

[0011] Because there are no approved medications for the effective management and treatment of cyclophosphamide-induced neutropenia, there is room for research and development of formulations that are safe and have enhanced efficacy.

[0012] Therefore, a need is felt for pharmaceutical formulations that alleviate the above disadvantages herein. Summary of the Invention

[0013] assignment Some of the problems of the present disclosure that are met by at least one embodiment herein are as follows: It is an object of the present disclosure to ameliorate one or more of the problems in the prior art, or at least to provide a useful alternative. The object of the present disclosure is to provide a pharmaceutical formulation.

[0014] Another object of the present disclosure is to provide a pharmaceutical formulation of cinnamic acid. Another object of the present disclosure is to provide a pharmaceutical formulation of cinnamic acid that is cost-effective and can be used for all types of chemotherapy-induced neutropenia.

[0015] The problem of the present disclosure is to provide a pharmaceutical formulation in the form of a dry powder inhalation formulation of cinnamic acid which avoids first pass metabolism. Another object of the present disclosure is to provide a pharmaceutical formulation in the form of a dry powder inhalation formulation of cinnamic acid that exerts equivalent / enhanced efficacy at reduced doses.

[0016] Another object of the present disclosure is to provide a pharmaceutical formulation of cinnamic acid in the form of an injectable formulation. Another object of the present disclosure is to provide a pharmaceutical formulation of cinnamic acid in the form of an oral dosage form.

[0017] Another objective of the present disclosure is to provide a simple process for the preparation of cinnamic acid. Other objects and advantages of the present disclosure will become more apparent from the following description, but the following description is not intended to limit the scope of the present disclosure.

[0018] overview The present disclosure relates to pharmaceutical formulations and processes for the preparation of pharmaceutical formulations.

[0019] In one aspect, the pharmaceutical formulation comprises cinnamic acid in an amount ranging from 0.1% to 10% by weight, based on the total weight of the formulation, and at least one excipient in an amount ranging from 90% to 99.9% by weight, based on the total weight of the formulation. The cinnamic acid is selected from cis-cinnamic acid and trans-cinnamic acid.

[0020] In another aspect, the present disclosure relates to a process for the preparation of cinnamic acid. The process includes mixing picroside 1 with a first fluid medium, and subsequently adding an alkali hydroxide under stirring to obtain a mixture. The mixture is maintained at a temperature ranging from 25°C to 35°C for a predetermined period of time, and subsequently neutralizing the mixture with hydrochloric acid to obtain a neutralized mixture. From the neutralized mixture, the fluid medium is removed under vacuum at a temperature ranging from 40°C to 50°C to obtain a dry powder containing decomposed picroside 1 and salts. The dry powder is mixed with a second fluid medium to obtain a solution containing decomposed picroside 1 without salts. The solution is decanted, and the second fluid medium is evaporated from the solution at a temperature ranging from 40°C to 50°C to obtain cinnamic acid (decomposed picroside 1 without salts). [Brief description of the drawings]

[0021] BRIEF DESCRIPTION OF THE ACCOMPANYING DRAWINGS The present disclosure will now be described with the aid of the accompanying drawings, in which: [Figure 1A] FIG. 1A is a graph illustrating the pattern of drug distribution per release from stage 2 to stage 6 of an NGI (next generation impactor) according to the present disclosure; [Figure 1B] FIG. 1B illustrates a graph presenting the drug distribution pattern per discharge from the device to the MOC (micro-orifice collector, which captures in a collection cup the very small particles of drug that are usually collected in the final filter in other impactors) in accordance with the present disclosure;

[0022] [Diagram 2] FIG. 2 illustrates a graph showing cumulative (% undersize) particle size distribution and drug distribution at various stages of a dry powder inhalation (DPI) formulation of cinnamic acid according to the present disclosure;

[0023] [Figure 3A] FIG. 3A illustrates the HPLC chromatogram of pure picroside 1 according to the present disclosure; [Figure 3BC] FIG. 3B illustrates an HPLC chromatogram of degraded picroside 1 (cinnamic acid of the present disclosure) without salt, according to the present disclosure; FIG. 3C illustrates an HPLC chromatogram of degraded picroside 1 with salt, according to the present disclosure;

[0024] [Figure 3D] FIG. 3D illustrates the mass chromatograms of pure picroside 1 and methanol (blank solution) according to the present disclosure; FIG. 3E illustrates the mass spectrum of decomposed picroside 1 without salt according to the present disclosure; [Figure 3FG] FIG. 3D illustrates the FTIR spectrum of pure picroside 1 and methanol according to the present disclosure; FIG. 3F illustrates the FTIR spectrum of decomposed picroside 1 without salt according to the present disclosure;

[0025] [Figure 3HI] FIG. 3G illustrates a 1H-NMR spectrum of decomposed picroside 1 without salt according to the present disclosure; FIG. 3H illustrates a 13C-NMR spectrum of decomposed picroside 1 without salt according to the present disclosure; FIG. 3I illustrates a UV spectrum of decomposed picroside 1 without salt according to the present disclosure; [Figure 3J] FIG. 3J illustrates the UV spectrum of commercially available synthetic cinnamic acid in accordance with the present disclosure;

[0026] [Figure 4A] FIG. 4A illustrates the comparative effects of pure picroside 1 and picroside 1 degraded with salt on cyclophosphamide-induced hematological changes versus control and GCSF (granulocyte colony-stimulating factor); [Figure 4B] FIG. 4B illustrates the comparative effects of pure picroside 1 and degraded picroside 1 with salt on cyclophosphamide-induced hepatotoxicity versus control and GCSF (granulocyte colony-stimulating factor);

[0027] [Figure 5A]FIG. 5A illustrates the comparative effects of pure picroside 1, salt-free decomposed picroside 1 (cinnamic acid of the present disclosure), low-dose commercial synthetic cinnamic acid (0.6 mg / tablet), and high-dose commercial synthetic cinnamic acid (6 mg / tablet) treatment on cyclophosphamide-induced hematological changes versus control and GCSF (granulocyte colony-stimulating factor); and [Figure 5AB] Figure 5A illustrates the comparative effects of pure picroside 1, salt-free decomposed picroside 1 (cinnamic acid of the present disclosure), commercial synthetic cinnamic acid (0.6 mg / tablet) low-dose treatment, and commercial synthetic cinnamic acid (6 mg / tablet) high-dose treatment on cyclophosphamide-induced hematological changes on control and GCSF (granulocyte colony-stimulating factor). Figure 5B illustrates the comparative effects of pure picroside 1, salt-free decomposed picroside 1 (cinnamic acid of the present disclosure), commercial synthetic cinnamic acid (0.6 mg / tablet) low-dose treatment, and commercial synthetic cinnamic acid (6 mg / tablet) high-dose treatment on cyclophosphamide-induced hepatotoxicity and renal changes on control and GCSF (granulocyte colony-stimulating factor). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0028] Detailed Description Aspects of the present disclosure will now be described with reference to the accompanying drawings.

[0029] The embodiments are provided to thoroughly and completely convey the scope of the present disclosure to those skilled in the art. Numerous details are set forth with respect to specific components and methods to provide a thorough understanding of the embodiments of the present disclosure. It will be apparent to those skilled in the art that the details provided in the embodiments should not be construed as limiting the scope of the present disclosure. In some embodiments, well-known processes, well-known device structures, and well-known techniques are not described in detail.

[0030] The terminology used in this disclosure is only for describing specific embodiments, and such terminology shall not be considered to limit the scope of the disclosure. The forms "a", "an", and "the" used in this disclosure may be intended to include the plural form, unless the context clearly suggests otherwise. The terms "comprises", "comprising", "including", and "having" are open-ended transitional phrases and thus specify the presence of stated features, integers, steps, operations, elements, modules, units, and / or components, but do not prohibit the presence or addition of one or more other features, integers, steps, operations, elements, components, and / or groups thereof. The specific order of steps disclosed in the methods and processes of the present disclosure should not be construed as necessarily requiring their execution exactly as described or illustrated. It should also be understood that additional or alternative steps may be employed.

[0031] Cyclophosphamide, an alkylating chemotherapy agent, is a drug that has been widely applied in clinical practice for the treatment of malignant and non-malignant tumors. However, despite its wide spectrum of clinical applications, it exhibits severe cytotoxicity to normal cells in both humans and experimental animals. Its metabolites can interact with cellular macromolecules such as proteins, membrane lipids, RNA, and DNA, and induce apoptosis. One of its metabolites, namely acrolein, causes oxidative stress leading to DNA damage in normal cells and causes toxicity in various organs. One of the most severely affected sites is the hematopoietic compartment of the bone marrow. Indeed, neutropenia is the most common and frequent side effect of cytotoxic chemotherapy. Other treatments are more costly and have severe and / or life-threatening adverse events such as anemia, anorexia, thrombocytopenia, alopecia, and peripheral neuropathy.

[0032] Given the current situation, and the absence of approved drugs for the effective management and treatment of cyclophosphamide-induced neutropenia, there is room for the development of pharmaceutical compositions that are safe and effective. The present disclosure provides pharmaceutical formulations and processes for their preparation.

[0033] In one aspect of the present disclosure, a pharmaceutical formulation comprises cinnamic acid as a pharma- ceutical active agent in an amount ranging from 0.1% to 10% by weight, based on the total weight of the formulation, and at least one excipient in an amount ranging from 90% to 99.9% by weight, based on the total weight of the formulation.

[0034] The cinnamic acid is selected from trans-cinnamic acid and cis-cinnamic acid. The excipient is at least one selected from the group consisting of a solvent, a colorant, a lubricant, a diluent and a disintegrant.

[0035] According to the present disclosure, the solvent is at least one selected from water, ethanol and isopropyl alcohol. The coloring agent is selected from FD and C color. The lubricant is at least one selected from magnesium stearate, talc, silica, stearic acid. The disintegrant is at least one selected from carboxymethylcellulose and hydroxypropylmethylcellulose. The diluent is selected from polyethylene glycol, dimethyl sulfoxide, ethyl lactate, and a combination mixture of D-mannitol-xylitol-microcrystalline cellulose-crospovidone-anhydrous dibasic calcium phosphate.

[0036] In one embodiment, the pharmaceutical formulation is in the form selected from oral formulations, injection formulations, and inhalation formulations, metered dose inhalation formulations, ointments, gels, patches, ophthalmic formulations, and sprays.In an exemplary embodiment, the pharmaceutical formulation is in the form of an oral formulation.In another exemplary embodiment, the pharmaceutical formulation is in the form of an injection formulation.In yet another embodiment, the pharmaceutical formulation is in the form of a dry powder inhalation formulation.

[0037] In one embodiment, the pharmaceutical formulation is in the form of an oral formulation. The oral formulation is in a dosage form selected from the group consisting of tablets, capsules, ointments, gels, mouthwashes, and suspensions. In an exemplary embodiment, the dosage form of the oral pharmaceutical formulation is a tablet. The oral formulation comprises cinnamic acid in an amount ranging from 0.1% to 5% by weight based on the total weight of the formulation, and at least one excipient in an amount ranging from 95% to 99.9% by weight based on the total weight of the formulation.

[0038] In one embodiment, the excipient is at least one selected from the group consisting of a solvent, a colorant, a lubricant, a diluent, and a disintegrant. In one embodiment, the solvent is at least one selected from the group consisting of water, ethanol, and isopropyl alcohol. In an exemplary embodiment, the solvent is water.

[0039] In one embodiment, the solvent is in an amount ranging from 10% to 50% by weight based on the total weight of the excipient. In an exemplary embodiment, the amount of solvent is a sufficient amount (weight %) based on the total weight of the excipient.

[0040] In one embodiment, the colorant is at least one selected from an FD&C color. In an exemplary embodiment, the colorant is selected from Iron Oxide yellow NF, brilliant Blue supra, FD&C Green 3, and sunset yellow.

[0041] In one embodiment, the colorant is in an amount ranging from 0.5% to 2% by weight based on the total weight of the excipient. In an exemplary embodiment, the amount of colorant is 0.7% by weight based on the total weight of the excipient.

[0042] In one embodiment, the lubricant is at least one selected from the group consisting of magnesium stearate, talc, silica, and stearic acid. In an exemplary embodiment, the lubricant is magnesium stearate.

[0043] In one embodiment, the lubricant is in an amount ranging from 0.5% to 2% by weight, based on the total weight of the excipient. In an exemplary embodiment, the amount of lubricant is 0.7% by weight, based on the total weight of the excipient.

[0044] In one embodiment, the disintegrant is at least one selected from the group consisting of carboxymethylcellulose, hydroxypropylmethylcellulose, and F melt type C. In an exemplary embodiment, the disintegrant is F melt type C.

[0045] In one embodiment, the disintegrant is in an amount ranging from 95% to 98% by weight, based on the total weight of the excipient. In an exemplary embodiment, the amount of disintegrant is 96% by weight, based on the total weight of the excipient.

[0046] In one embodiment, the diluent is selected from polyethylene glycol, dimethylsulfoxide, ethyl lactate, and a combination mixture of D-mannitol-xylitol-microcrystalline cellulose-crospovidone-dicalcium phosphate anhydrous.

[0047] In one embodiment, the total amount of excipients is in the range of 95% to 99.9% by weight based on the total weight of the formulation.

[0048] In one embodiment, the pharmaceutical formulation is an injectable formulation. The injectable formulation comprises cinnamic acid in an amount ranging from 1% to 10% by weight based on the total weight of the formulation, and at least one excipient in an amount ranging from 90% to 99% by weight based on the total weight of the formulation.

[0049] In one embodiment, the at least one excipient is selected from the group consisting of polyethylene glycol, water, ethanol, ethyl lactate, propylene glycol, and dimethyl sulfoxide. In an exemplary embodiment, the excipient is a combination of alcohol and polyethylene glycol, alcohol and water, alcohol and dimethyl sulfoxide, alcohol and ethyl lactate, alcohol and propylene glycol.

[0050] In one embodiment, the pharmaceutical formulation is a dry powder inhalation formulation comprising micronized cinnamic acid, a first lactose, a second lactose and at least one excipient.

[0051] In one embodiment, the micronized cinnamic acid has a particle size ranging from 0.1 μm to 10 μm. In an exemplary embodiment, the particle size of the micronized cinnamic acid is 1 to 5 μm.

[0052] In one embodiment, the micronized cinnamic acid is in an amount ranging from 1% to 5% by weight relative to the total weight of the formulation. In an exemplary embodiment, the amount of micronized cinnamic acid is 2.4% by weight.

[0053] In one embodiment, the first lactose has a particle size in the range of 20 μm to 300 μm. In an exemplary embodiment, the particle size of the first lactose is 35 to 190 μm. In one embodiment, the first lactose is in an amount ranging from 10% to 30% by weight, based on the total weight of the formulation. In an exemplary embodiment, the amount of the first lactose is 19% by weight.

[0054] In one embodiment, the first lactose is respitose SV010. In one embodiment, the average particle size (d10) of the first lactose is in the range of 35 μm to 65 μm. In one embodiment, the average particle size (d50) of the first lactose is in the range of 95 μm to 125 μm.

[0055] In one embodiment, the average particle size (d90) of the first lactose is in the range of 160 μm to 190 μm. In one embodiment, the second lactose has a particle size in the range of 0.1 μm to 10 μm. As an example, the second lactose has a particle size of 0.8 to 7.5 μm.

[0056] In one embodiment, the second lactose is in an amount ranging from 75% to 85% by weight, based on the total weight of the formulation. In an exemplary embodiment, the amount of the second lactose is 78% by weight. In one embodiment the second lactose is Pharmatose-450M.

[0057] In one embodiment, the average particle size (d10) of the second lactose is in the range of 0.01 μm to 0.1 μm. In one embodiment, the average particle size (d50) of the second lactose is in the range of 1 μm to 5 μm. In one embodiment, the average particle size (d90) of the second lactose is in the range of 1 μm to 10 μm.

[0058] In one embodiment, the mass ratio of the first lactose to the second lactose is in the range of 1:2 to 1:5. In an exemplary embodiment, the mass ratio of the first lactose to the second lactose is 1:4. In one embodiment, the excipient is selected from magnesium stearate, calcium stearate and zinc stearate. In an exemplary embodiment, the excipient is magnesium stearate.

[0059] In one embodiment, the excipient is present in an amount ranging from 0.1% to 1% by weight, based on the total weight of the formulation. In an exemplary embodiment, the amount of the excipient is 0.32% by weight.

[0060] In one embodiment, the mean mass aerodynamic diameter (MMAD) of the dry powder inhalation formulation ranges from 2 μm to 5 μm. In an exemplary embodiment, the mean mass aerodynamic diameter (MMAD) of the dry powder inhalation formulation is 4.26 μm.

[0061] The dry powder inhalation (DPI) formulation thus obtained is filled into size 3 HPMC (hydroxypropyl methylcellulose) capsules using a capsule filling machine. The present disclosure provides dry powder inhalation formulations that avoid first-pass metabolism. The formulations of the present disclosure are therapeutically effective at reduced doses, and improved patient compliance has also been observed.

[0062] In one embodiment, the pharmaceutical formulation of cinnamic acid is administered at a dose ranging from 1.5 mg / kg body weight to 30 mg / kg body weight. In one embodiment, the pharmaceutical preparation has antineutropenic activity.

[0063] In another embodiment, the pharmaceutical formulation has antitubercular activity, antimalarial activity and cardiovascular activity. In one embodiment, the pharmaceutical formulation is further combined with a known anti-tuberculosis drug.

[0064] In one embodiment, the pharmaceutical formulation has antiviral activity against COVID-19 and antibacterial activity against bacteria such as Bacillus subtilis and E. coli, present in the herbal extract powder.

[0065] In one embodiment, a pharmaceutical formulation of cis-cinnamic acid is 120 times more effective than the trans form of cinnamic acid. In one embodiment, the pharmaceutical formulations of cis-cinnamic acid are in the form of oral, injectable, inhaled, topical, and ophthalmic formulations.

[0066] In one embodiment, pharmaceutical formulations of cis-cinnamic acid are used to treat neutropenia, tuberculosis, malaria, and cardiovascular disease. The present disclosure further provides a method of treating neutropenia, tuberculosis, malaria, viral infection, bacterial infection and cardiovascular disease in a mammal, the method comprising administering to the mammal a therapeutically effective amount of cinnamic acid in an amount ranging from 1.5 mg / kg body weight to 30 mg / kg body weight.

[0067] In one aspect of the disclosure, the mammal is a human. In one embodiment of the present disclosure, the therapeutically effective amount of cinnamic acid is in the range of 15 mg / kg body weight to 25 mg / kg body weight.

[0068] In an embodiment of the disclosure, the pharmaceutical formulation is used to treat neutropenia, tuberculosis, malaria, viral infections, bacterial infections, and cardiovascular disease.

[0069] In an embodiment of the present disclosure, cinnamic acid is used to treat neutropenia, tuberculosis, malaria, viral infections, bacterial infections, and cardiovascular disease, and a therapeutically effective amount of cinnamic acid is in the range of 1.5 mg / kg body weight to 30 mg / kg body weight.

[0070] A therapeutically effective amount refers to the amount of a compound that, when administered to a patient, is effective in treating such disease or condition and is sufficient to produce the desired effect. In another aspect of the present disclosure, a process for the preparation of cinnamic acid is provided.

[0071] The process is explained in detail. In the first step, pure picroside 1 is mixed with a first fluid medium and simultaneously an alkali hydroxide is added under stirring to obtain a mixture. In one embodiment, the first fluid medium is water.

[0072] In one embodiment, the alkali is sodium hydroxide, potassium hydroxide, and calcium hydroxide. In one embodiment, the ratio of picroside 1 to alkali hydroxide is 0.01:50. In the second step, the mixture is maintained at a temperature ranging from 25 to 35° C. for a predetermined period of time, followed by neutralizing the mixture with hydrochloric acid to obtain a neutralized mixture.

[0073] In one embodiment, the predetermined period of time is in the range of 30 minutes to 2 hours. In an exemplary embodiment, the predetermined period of time is 1 hour. In the third step, the fluid medium is removed from the neutralized mixture under vacuum at a temperature ranging from 40° C. to 50° C. to obtain a dry powder containing decomposed picroside 1 along with salts.

[0074] In a fourth step, the dry powder is mixed with a second fluid medium to obtain a solution containing salt-free decomposed picroside 1. In one embodiment, the salt thus obtained (NaCl) is allowed to settle to the bottom (settling salt). In one embodiment, the second fluid medium is methanol, ethanol, propanol and butanol.

[0075] In the fifth step, the solution is decanted, followed by evaporation of the second fluid medium from the solution at a temperature of 40° C.-50° C. to obtain cinnamic acid (decomposed picroside 1 without salt). In one embodiment, after decantation, the settling salt is further washed with a second fluid medium, and the washed second fluid medium is evaporated to obtain cinnamic acid.

[0076] Cinnamic acid prepared according to the present disclosure in an amount ranging from 0.1% to 10% by weight is formulated with at least one excipient in an amount ranging from 90% to 99.9% by weight for the treatment of neutropenia, tuberculosis, malaria, viral infections, bacterial infections and cardiovascular disease, wherein the weight percentage of each component is based on the total weight of cinnamic acid and excipients.

[0077] The foregoing description of the embodiments is provided for illustrative purposes and is not intended to limit the scope of the present disclosure. Individual components of a particular embodiment are generally not limited to that particular embodiment, but are interchangeable. Such variations are not considered a departure from the present disclosure, and all such modifications are considered to be within the scope of the present disclosure.

[0078] The present disclosure is further illustrated in light of the following experiments, which are presented for illustrative purposes only and should not be construed as limiting the scope of the present disclosure. The following experiments can be scaled up to industrial / commercial scale, and the results obtained can be extrapolated to industrial scale.

[0079] Experimental details Example 1: Preparation of a pharmaceutical formulation of cinnamic acid in the form of tablets by using a wet granulation method: A pharmaceutical formulation of cinnamic acid in the form of tablets was prepared using wet granulation. The process includes the following steps; 2mg / tablet of cinnamic acid was dissolved in 5gm of water to obtain a solution of the drug. 67.5mg / tablet of F-melt type C and 0.5mg / tablet of blue pigment were mixed in a poly bag for 5 minutes through 40 and 80 sieves to obtain a mixture. The blend of F-melt type C, blue pigment, and drug solution was transferred to a wet granulator to obtain granules of cinnamic acid. The granules were dried in an oven at 60°C for 15 minutes to obtain dry granules. The dry granules were passed through a 40 sieve and charged into a blender to obtain a powder. 0.5mg / tablet of magnesium stearate was added to the powder and mixed in a poly bag for 5 minutes and transferred to a compression machine to obtain uncoated tablets of cinnamic acid. The uncoated tablets of cinnamic acid were characterized by evaluating various tablet parameters, as given in Table 2.

[0080] Table 1 [Table 1] F-Melt® Type C is a combination mixture of D-mannitol-xylitol-microcrystalline cellulose-crospovidone-dicalcium phosphate anhydrous.

[0081] Table 2 [Table 2]

[0082] Example 2: Comparative formulation Comparative formulations were prepared in a similar manner to that disclosed in Example 1 by varying the concentrations of ingredients depending on the formulation, as illustrated in Table 3 below. [Table 3]

[0083] Example 3A: Preparation of a pharmaceutical formulation of cinnamic acid (6 mg / ml) in an injectable form by using commercially available synthetic cinnamic acid: 6 mg of cinnamic acid was dissolved in 300 μl of 100% ethanol, sonicated for 5 minutes, and then diluted to 1 ml with 5% PEG400 / 5% DMSO / 5% ethyl lactate to obtain a clear solution of cinnamic acid in injectable form (injectable formulation). The formulation was evaluated for stability according to ICH guidelines. The stability parameters of the injectable formulation of cinnamic acid (6 mg / ml) are shown in Tables 4a and 4b below.

[0084] Table 4a [Table 4a]

[0085] Table 4b [Table 4b]

[0086] Inference: From Tables 4a and 4b, it is evident that the 6 mg / ml formulation is stable in all dilutions tested for stability. The presence of 30% ethanol ensures that the cinnamic acid of the present disclosure (degraded picroside 1 without salt) remains in a soluble form.

[0087] Example 3B: Preparation of a pharmaceutical formulation of commercially available synthetic cinnamic acid (2 mg / ml) in injectable form 2 mg of cinnamic acid was dissolved in 300 μl of 100% ethanol, sonicated for 5 minutes, and then diluted to 1 ml with 5% PEG400 / 5% DMSO / 5% ethyl lactate / 1.4% propylene glycol / water to obtain a clear solution of commercially available synthetic cinnamic acid (2 mg / ml). The formulation was evaluated for stability according to ICH guidelines. The stability parameters of the injectable formulation of cinnamic acid (2 mg / ml) are shown in Tables 4c and 4d below.

[0088] Table 4c [Table 4c]

[0089] Table 4d [Table 4d]

[0090] Inference: From 4c and 4d, it is evident that the 2mg / ml injectable formulation of cinnamic acid is stable in all diluents tested for stability. The presence of 30% ethanol ensures that the cinnamic acid remains soluble in solution.

[0091] Example 4: Preparation of a pharmaceutical formulation of cinnamic acid in dry powder inhalation form: Dry mixing: 0.6 grams of micronized cinnamic acid, 4.76 grams of Respitose SV010 (first lactose), and 19.56 grams of Pharmatose 450M (inhalation grade lactose (second lactose)) were mixed using a double-lined polythene bag and sieved using 80 mesh size to obtain a drug lactose blend. Separately, 0.08 grams (gms) of magnesium stearate was mixed and added to the drug lactose blend to obtain a mixture, and the mixture was blended for 30 minutes at a speed of 15 rpm to obtain a dry powder inhalation (DPI) formulation of cinnamic acid. The thus obtained dry powder inhalation (DPI) formulation of cinnamic acid was filled into size 3 HPMC capsules using a capsule filling machine.

[0092] Example 5: In vitro aerodynamic particle size distribution of cinnamic acid Aerodynamic particle size distribution (APSD) or aerosolization performance is an essential quality attribute for the in vitro characterization of orally inhaled drug products (OINDPs).

[0093] The APSD of an aerosol determines the proportion of DPI particles that accumulate in the body, especially in the lower respiratory tract: particles in the 1-5 micron range reach the lower respiratory tract and are considered effective, particles larger than 5 microns remain in the upper respiratory tract and are more likely to impact the oropharynx and be swallowed, and particles smaller than 1 micron are removed by pulmonary clearance mechanisms.

[0094] A 25mg cinnamic acid dry powder inhalation (DPI) formulation was evaluated for aerodynamic particle size distribution (APSD). The APSD from each dose of the dry powder inhalation formulation of the present disclosure was evaluated using a next generation impactor (NGI). The cinnamic acid dry powder inhalation (DPI) formulation was placed into the inhaler for use, and a mouthpiece adapter was attached to the inhalation port. The pump was turned on at a pressure of 4 kPa pressure drop across the device. The ejection sequence was repeated four times to ensure complete ejection of the powder into the NGI port. After aerosolization, the amount of drug retained in the inhaler, the inlet port, the mouthpiece adapter, the pre-separator, and the NGI cup was extracted by washing with an appropriate amount of 90:10 methanol:water for quantitative HPLC analysis of cinnamic acid. All samples were filtered through a 0.45 μm filter and analyzed for cinnamic acid content by HPLC. Important NGI parameters such as mass mean aerodynamic diameter (MMAD), geometric standard deviation (GSD), emitted dose (ED), and fine particle fraction (FPF) were calculated using CITDAS software (COPLEY Scientific, UK). The DPI, mean FPF (≦5 μm) of cinnamic acid was roughly 35% of the nominal dose (referring to the capsule content) of cinnamic acid, while the mass mean aerodynamic diameter (MMAD) and GSD values ​​were 4.2 μm and 2.1, respectively. The total emitted amount of active ingredient was 11.081 mg. In Figure 1A and 1B, the drug distribution patterns per release of various stages and devices for the MOC of NGI are shown.

[0095] Example 6: Pulmonary accumulation of cinnamic acid in vivo The in vivo lung accumulation of the dose was evaluated by using a next generation impactor (NGI) for cinnamic acid DPI. Each capsule contained 0.6 mg of the disclosed cinnamic acid (picroside 1 dissolved without salt) and was characterized using a Pastiape apparatus with the device at a flow rate of 68 L / min (≈4 KPa).

[0096] From Figure 2, which represents the cumulative (% undersize) particle size distribution of the cinnamic acid dry powder inhalation (DPI) formulation according to the present disclosure, it is clear that the dose of cinnamic acid DPI was dispensed at different stages of the NGI (Next Generation Impactor) and the fine particle dose (respirable dose) and fine particle fraction (respirable fraction) were 0.48mg and 40.6%, respectively, while the average delivered dose was 0.497mg per actuation. These results meet the requirements of Ph. Eur. 68, which suggests that + / - 25% of the target dose be delivered from the DPI, whereas that of USP 69 suggests that + / - 15% of the target dose be delivered.

[0097] Example 7: Alkaline decomposition of pure picroside 1 180 mg of picroside 1 with a purity of 95% was mixed in 650 ml of water, followed by adding 120 ml of 0.1NaOH under stirring to obtain a mixture. The resulting mixture was kept at room temperature for 1 hour, and then neutralized with hydrochloric acid to obtain a neutralized mixture having a pH of 7. The fluid medium was removed from the neutralized mixture under vacuum at a temperature ranging from 40°C to 50°C to obtain a dry powder containing decomposed picroside 1 with salt. The dry powder was mixed with a second fluid medium to obtain a solution containing decomposed picroside 1 without salt. The solution was decanted, and the second fluid medium was subsequently evaporated from the solution at a temperature ranging from 40°C to 50°C to obtain the cinnamic acid of the present disclosure (decomposed picroside 1 without salt).

[0098] The process of preparation of cinnamic acid by alkaline decomposition of pure picroside 1 is shown in Figure 1 below. [ka] Scheme 1

[0099] Picroside 1 with salt is obtained by forced alkali hydroxide decomposition of naturally occurring Picroside 1. The salt remains intact in the decomposed form. Picroside 1 without salt is a cinnamic acid according to the present disclosure, obtained by forced alkaline hydroxide decomposition of naturally occurring picroside 1. The salt is removed in this decomposed form by using a second fluid medium (methanol).

[0100] Example 8: Characterization of Salt-Free Degraded Picroside 1 (Cinnamic Acid of the Disclosure) The salt-free decomposed picroside 1 (cinnamic acid of the present disclosure) was subjected to HPLC to evaluate the chromatogram, and the dried sample of the salt-free decomposed picroside 1 (cinnamic acid of the present disclosure) was also subjected to LCMS, NMR and FTIR to evaluate the structure.

[0101] HPLC conditions for salt-free decomposed picroside 1 (cinnamic acid of the present disclosure): Mobile phase: A) 0.1% orthophosphoric acid B) Acetonitrile Diluent: Methanol Sample preparation: 10 mg of sample was dissolved in 10 ml of methanol.

[0102] Chromatography conditions: [Table A]

[0103] Gradient Program: Table 5 [Table 5]

[0104] From Figures 3A and 3B, it is clear that the retention time of pure picroside 1 is 12.815, whereas the retention time of decomposed picroside 1 (cinnamic acid of the present disclosure) without salt was found to be 21.947.

[0105] LCMS (Liquid Chromatography-Mass Chromatography) Study: Mobile phase: A: 5 mM ammonium formate + 0.1% formic acid in water; B: 5 mM ammonium formate + 0.1% formic acid in methanol.

[0106] Ammonium formate was used in the mobile phase for LC-MS analysis, which formed an adduct with the salt-free degraded picroside 1 (cinnamic acid of the present disclosure) and showed a molecular weight of 210. Subtracting the mass of the peak observed at 210 m / z, the mass of the salt-free degraded picroside 1 (cinnamic acid of the present disclosure) without any adduct is 210-63=147 Daltons.

[0107] From the LCMS study and Figures 3D and 3E, it is clearly evident that the resolved picroside 1 without salt is a cinnamic acid with a molecular weight of 148 g / mol.

[0108] FTIR (Fourier Transform Infrared Spectroscopy) Study: The FTIR results of decomposed picroside 1 (cinnamic acid of the present disclosure) without salt are displayed in Table 6 below and in FIG. 3F. Table 6 [Table 6]

[0109] NMR study of decomposed picroside 1 (cinnamic acid of the present disclosure) without salt: From Figures 3G and 3H, it is clear that decomposed picroside 1 without salt showed smaller hydrogen and carbon signals due to the presence of sugars. The NMR signals of the synthesized pure cinnamic acid match the peaks observed in decomposed picroside 1 (without salt). The additional NMR peaks observed in decomposed picroside 1 (without salt) are due to the presence of sugars and do not affect the identity of decomposed picroside 1 as a cinnamic acid of the present disclosure.

[0110] Comparison of 1H MNR data [Table B]

[0111] 13 CMNR Data Comparison [Table C]

[0112] Conclusion: From the data of HPLC, LCMS, FTIR, NMR (illustrated in Figures 3A-3H), and UV spectroscopy (illustrated in Figures 3I and 3J), it is evident that the decomposed picroside 1 without salt is cinnamic acid, since it shows similar retention time and similar wavelength as cinnamic acid.

[0113] Example 9: Evaluation of the efficacy of pure picroside 1 and decomposed picroside 1 with salts in cyclophosphamide-induced neutropenia assignment: 1. Induce neutropenia in mice by administering cyclophosphamide. 2. To determine the effect of treatment with pure picroside 1 and picroside 1 dissolved with salt on total white blood cell count, neutrophil, monocyte, lymphocyte and platelet count. 3. To compare the effects of pure picroside 1 and decomposed picroside 1 with salt with standard 2 μg G-CSF.

[0114] Materials and Methods: material: Cyclophosphamide was obtained from Sigma Aldrich and G-CSF was obtained from Lupin limited, a commercial formulation, Lupifil (300). Female Swiss albino mice weighing 25-27 g were obtained from National Institute of Biological Sciences, Pune.

[0115] method: Female Swiss albino mice were randomly divided into different groups based on body weight as follows: Group 1 - Cyclophosphamide control (CP control); Group 2 - pure picroside 1 treatment, Group 3 - Decomposed picroside 1 with salt, and • Group 4-GCSF (granulocyte colony stimulating factor) treated group.

[0116] On day 0, blood was taken from all animals for basal hematological parameters. Animals in all groups were injected with cyclophosphamide at 150 mg / kg via intraperitoneal route (ip). On day 3, blood was taken from all animals for total and differential counts (neutrophils, lymphocytes, monocytes). Animals were dosed with one additional dose of cyclophosphamide at 100 mg / kg on day 4. From day 5, animals were administered their respective treatments via oral route with water as vehicle, while group 1 was administered placebo tablets for 5 consecutive days. On day 10, blood was taken from all animals for total and differential counts (neutrophils, lymphocytes, monocytes), leukocytes, platelets, and AST (aspartate aminotransferase) and ALT (alanine aminotransferase).

[0117] result: Effect of pure picroside 1 and decomposed picroside 1 with salts on neutrophil percentage in cyclophosphamide-induced neutropenia On day 0, the neutrophil percentages of all animal groups were 42-43%, and no significant differences were observed between the groups on day 0. Administration of cyclophosphamide caused a significant (p<0.001) decrease in the % of neutrophils as observed on day 3 in cyclophosphamide control, pure picroside 1, degraded picroside 1 with salt, and GCSF (granulocyte colony stimulating factor) compared to day 0. As shown in Table 7A and Figure 4A, treatment with pure picroside 1 significantly (p<0.01) reduced cyclophosphamide-induced neutropenia when compared to the placebo control group. However, treatment with degraded picroside 1 with salt did not cause a significant change in cyclophosphamide-induced neutropenia when compared to animals in the cyclophosphamide control group. The improvement in neutrophil % observed with pure picroside 1 was comparable to that of GCSF (granulocyte colony stimulating factor).

[0118] Effect of pure picroside 1 and picroside 1 degraded with salt on the percentage of monocytes in cyclophosphamide-induced hematological changes On day 0, all test animals had a 7-7.5% exploration percentage of monocytes, and there was no significant difference between the groups. Cyclophosphamide administration resulted in a significant (p<0.001) decrease in the % of monocytes observed on day 3 in the cyclophosphamide control group, pure picroside 1 and picroside 1 degraded with salt, and GCSF (granulocyte colony stimulating factor) groups compared to day 0. The cyclophosphamide-induced decrease in the percentage of monocytes was significantly reversed (p<0.01) by pure picroside 1 treatment when compared to the cyclophosphamide control group, as displayed in Table 7A and Figure 4A. However, picroside 1 degraded with salt treatment did not cause a significant change in the cyclophosphamide-induced decrease in the % of monocytes when compared to the animals in the cyclophosphamide control group. The reversal observed with pure picroside 1 was comparable to that of the GCSF (granulocyte colony stimulating factor)-treated group. Reversal means that the cyclophosphamide-induced decrease in monocyte percentage was significantly reversed by treatment with pure picroside 1. First, CP reduces the number of monocytes, which is then reversed by pure picroside 1.

[0119] Effect of pure picroside 1 and picroside 1 digested with salts on the percentage of lymphocytes in cyclophosphamide-induced hematological changes On day 0, the % of lymphocytes in all test animal groups was between 39-41%, and no significant differences were observed between groups on this day. All groups treated with cyclophosphamide had significantly higher (p<0.01) lymphocyte counts on day 3 than on day 0. As displayed in Table 7A and Figure 4A, a significant (p<0.01) reduction in the % of lymphocytes was observed with treatment of pure picroside 1 when compared to the cyclophosphamide control group. However, treatment with picroside 1 degraded with salt did not cause a significant change in the cyclophosphamide-induced lymphocytosis % when compared to the cyclophosphamide control group animals. GCSF (granulocyte colony-stimulating factor) significantly (p<0.001) reversed the cyclophosphamide-induced lymphocyte mobilization compared to the cyclophosphamide control group. The reversal observed with picroside 1 was comparable to that of the GCSF (granulocyte colony stimulating factor)-treated group.

[0120] Effect of pure picroside 1 and picroside 1 degraded with salt on total white blood cell count in cyclophosphamide-induced hematological changes On day 0, the total white blood cell counts of all test animal groups were between 7410 and 8085. No significant differences were observed between the groups on this day. Administration of cyclophosphamide caused a significant (p<0.001) decrease in WBC counts observed on day 3 in the cyclophosphamide control group, pure picroside 1, and picroside 1 degraded with salt, and GCSF (granulocyte colony stimulating factor) groups compared to day 0. Treatment with pure picroside 1 and GCSF (granulocyte colony stimulating factor) significantly (p<0.001) reversed the cyclophosphamide-induced decrease in WBC counts when compared to the cyclophosphamide control, as displayed in Table 7A and Figure 4A. However, picroside 1 degraded with salt significantly (p<0.001) reversed the cyclophosphamide-induced decrease in WBC counts when compared to the cyclophosphamide control. The procedure is as follows: Cyclophosphamide Compared with the control group, cyclophosphamide Dopamine-induced leukopenia did not cause any change in The reversal observed in pure picroside 1 was comparable to that of the GCSF (granulocyte colony stimulating factor)-treated group.

[0121] Effect of pure picroside 1 and picroside 1 degraded with salt on cyclophosphamide-induced hematological changes and platelet counts On day 0, the platelet counts of all test animal groups were between 732400 and 748400. No significant differences were observed between the groups on this day. Administration of cyclophosphamide caused a significant (p<0.001) decrease in platelet counts observed on day 3 in the cyclophosphamide control group, pure picroside 1, and picroside 1 degraded with salt and GCSF (granulocyte colony stimulating factor) groups compared to day 0. Treatment with picroside 1 significantly (p<0.001) improved platelet counts relative to the cyclophosphamide control group, as displayed in Table 7A and Figure 4A. However, the platelet counts of picroside 1 degraded with salt were significantly (p<0.001) improved relative to the cyclophosphamide control group. Treatment did not cause any changes in platelet counts compared to the cyclophosphamide control group. Treatment with GCSF (granulocyte colony stimulating factor) resulted in a significant increase in platelet count compared to the cyclophosphamide control group. The increase in platelet count observed in the pure picroside 1 group was comparable to that of the GCSF (granulocyte colony stimulating factor) treated group.

[0122] Table 7A: Effects of pure picroside 1 and picroside 1 degraded with salts on cyclophosphamide-induced hematological changes [Table 7A] # p<0.001 compared to day 3 * p<0.05, ** p<0.01, *** p<0.001 compared with cyclophosphamide control

[0123] Inference: From Table 7A and Figure 4A, it is clear that administration of cyclophosphamide caused a significant decrease in neutrophils%, monocytes%, total white blood cell count, and platelet count, and a significant increase in lymphocyte count. Furthermore, treatment with pure picroside 1 caused the alleviation of cyclophosphamide-induced decreases in neutrophils%, monocytes%, total white blood cell count, and the increased lymphocyte count was also alleviated. In the pure picroside 1 treatment group, there was an improvement in platelet count, which was statistically significant. Treatment with salt-dissolved picroside 1 did not significantly change neutrophils%, monocytes%, total white blood cell count, and platelet count. However, the changes observed in the pure picroside 1 treatment group were comparable to those of the GCSF (granulocyte colony-stimulating factor) group, with lower efficacy.

[0124] Effect of pure picroside 1 and degraded picroside 1 with salts on AST (aspartate aminotransferase) (IU / L) in cyclophosphamide-induced hepatotoxicity The cyclophosphamide control group showed an increase in AST (aspartate aminotransferase) levels after cyclophosphamide administration. As displayed in Table 7B and FIG. 4B, a significant (p<0.001) decrease in AST (aspartate aminotransferase) levels was observed following administration of pure picroside 1 compared to the cyclophosphamide control group. Administration of degraded picroside 1 with salt significantly (p<0.01) reduced AST (aspartate aminotransferase) compared to the cyclophosphamide control group. There was a significant (p<0.001) reduction in AST (aspartate aminotransferase) levels in the GCSF (granulocyte colony stimulating factor) treatment group when compared to the cyclophosphamide control group.

[0125] Effect of pure picroside 1 and degraded picroside 1 with salts on ALT (alanine aminotransferase) (IU / L) in cyclophosphamide-induced hepatotoxicity Administration of cyclophosphamide in the cyclophosphamide control group reduced ALT (Alanine Aminotransferase) Administration of pure picroside 1 to animals caused a significant increase in ALT levels when compared to the cyclophosphamide control group, as shown in Table 7B and FIG. 4B. (Alanine aminotransferaseAdministration of degraded picroside 1 with salt caused a significant reversal (p<0.001) in ALT levels when compared with the cyclophosphamide control group. (Alanine Aminotransferase) No significant changes in ALT levels were observed in pure picroside 1. (Alanine Aminotransferase) The reduction in ALT was achieved upon treatment with GCSF (granulocyte colony stimulating factor). (Alanine Aminotransferase) levels were comparable (p<0.001).

[0126] Table 7B: Effect of pure picroside 1 and degraded picroside 1 with salts on cyclophosphamide-induced hepatotoxicity [Table 7B] *** p<0.001 compared to CP controls

[0127] Inference: From Table 7B and Figure 4B, it is clear that administration of cyclophosphamide caused a significant increase in AST (aspartate aminotransferase) and ALT (alanine aminotransferase). Furthermore, in the treatment with pure picroside 1, there was a significant decrease in the levels of AST (aspartate aminotransferase) and ALT (alanine aminotransferase). Therefore, from Tables 7A, 7B and Figures 4A and 4B, it is evident that degraded picroside 1 with its salt was unable to correct the neutropenic condition of the animals.

[0128] Example 10: Evaluation of the efficacy of commercially available synthetic cinnamic acids in cyclophosphamide-induced neutropenia assignment: 1. Induce neutropenia by administering cyclophosphamide. 2. To determine the effect of low dose treatment with commercially available synthetic cinnamic acid (0.6 mg / tablet) and high dose treatment with commercially available synthetic cinnamic acid (6 mg / tablet) on total white blood cell count, neutrophils, monocytes, lymphocytes, platelet count, liver parameters SGOT (serum glutamic oxaloacetic transaminase), SGPT (serum glutamic pyruvic transaminase), renal parameters (serum creatinine, blood urea nitrogen, uric acid).

[0129] Materials and Methods: Materials: Female Swiss albino mice weighing between 25-27 g were obtained from Global Bioresearch Solutions Pvt Ltd., Pune.

[0130] Methods: Female Swiss albino mice were randomly divided into seven groups of six mice each based on body weight as follows: Group 1 - Placebo group (plain water / observational control), ● Group 2 - Cyclophosphamide control (CP control), Group 3 - GCSF (granulocyte colony stimulating factor) 2μg via subcutaneous route (scroute) Group 4: Treatment with low doses of commercially available synthetic cinnamic acid (0.6 mg / tablet); Group 5: High-dose treatment with commercially available synthetic cinnamic acid (6 mg / tablet); Group 6 Pure picroside 1, and • Group 7. Salt-free degraded picroside 1 (cinnamic acid of the present disclosure) treatment group.

[0131] On day 0, blood was taken from all animals for baseline hematological parameters. All animal groups except the placebo group were administered cyclophosphamide 150 mg / kg via intraperitoneal route (ip), and group 1 was injected ip with water for injection. On day 3, blood was taken from all animals for total and differential counts (neutrophils, lymphocytes, and monocytes). Animals from groups 2 to 7 were boosted once with cyclophosphamide 100 mg / kg on day 4. From day 5, animals from groups 4 and 5 were administered with commercial synthetic cinnamic acid (0.6 mg / tablet) low dose treatment and commercial synthetic cinnamic acid (6 mg / tablet) high dose treatment, while groups 6 and 7 were administered pure picroside 1 and salt-free decomposed picroside 1 (cinnamic acid of the present disclosure) for 5 consecutive days. The cyclophosphamide control group was injected with water for injection from day 5 to day 9. On day 10, blood was collected from all animals to measure total and differential counts (neutrophils, lymphocytes, monocytes), platelets, liver parameters AST (aspartate aminotransferase), ALT (alanine aminotransferase), and kidney parameters (serum creatinine, blood urea nitrogen, and uric acid).

[0132] Dosage preparation and administration: ● Pure picroside 1-1 tablet was dissolved in 0.1 ml of sterile water for injection and 0.1 ml was injected as a subcutaneous injection. One tablet of salt-free dissolved picroside 1 (cinnamic acid of the present disclosure) was dissolved in 0.1 ml of sterile water for injection, and 0.1 ml was injected as a subcutaneous injection. Cinnamic acid low dose (0.6 mg / tablet) treatment tablets were dissolved in 0.1 ml of sterile water for injection and 0.1 ml was injected as a subcutaneous injection. Cinnamic acid high dose treatment (6 mg / tablet) tablets were dissolved in 0.1 ml of sterile water for injection and 0.1 ml was injected as a subcutaneous injection.

[0133] result: Effect of low-dose (0.6mg / tablet) and high-dose (6mg / tablet) synthetic cinnamic acid on neutrophil percentage in cyclophosphamide-induced neutropenia On day 0, the neutrophil percentages of all test groups of animals were between 38-41%, and no significant differences were observed between the groups on day 0. Administration of cyclophosphamide caused a significant (p<0.001) decrease in neutrophil% observed on day 3 by all groups receiving cyclophosphamide compared to the placebo group. Commercially available synthetic cinnamic acid (0.6 mg / tablet) low-dose treatment and commercially available synthetic cinnamic acid (6 mg / tablet) high-dose treatment significantly (p<0.001) attenuated cyclophosphamide-induced neutropenia when compared to the CP control group, as displayed in Table 8A and Figure 5A. This observed improvement in neutrophil% was superior to that observed with commercially available synthetic cinnamic acid (0.6 mg / tablet) low-dose treatment, and comparable to that of GCSF with commercially available synthetic cinnamic acid (6 mg / tablet) high-dose treatment.

[0134] Effects of low dose treatment with commercially available synthetic cinnamic acid (0.6mg / tablet) and high dose treatment with commercially available synthetic cinnamic acid (6mg / tablet) on monocyte percentage in cyclophosphamide-induced hematological changes On day 0, all test animals had a monocyte percentage between 6 and 6.82, and there was no significant difference between groups. Cyclophosphamide administration resulted in a significant (p<0.001) decrease in monocyte% on day 3 by all cyclophosphamide-treated groups compared to the placebo group. The cyclophosphamide-induced decrease in monocyte percentage was significantly (p<0.001) reversed by commercial synthetic cinnamic acid (0.6 mg / tablet) low dose treatment and commercial synthetic cinnamic acid (6 mg / tablet) high dose treatment and GCSF (granulocyte colony stimulating factor) treatment when compared to the cyclophosphamide control group, as displayed in Table 8A and Figure 5A. The reversal observed in the commercial synthetic cinnamic acid (0.6 mg / tablet) low dose treatment and the commercial synthetic cinnamic acid (6 mg / tablet) low dose treatment was better than pure picroside 1, salt-free degraded picroside 1 (cinnamic acid of the present disclosure), and comparable to that of the GCSF (granulocyte colony stimulating factor) treatment group.

[0135] Effects of low-dose commercial synthetic cinnamic acid (0.6mg / tablet) and high-dose commercial synthetic cinnamic acid (6mg / tablet) on lymphocyte percentage in hematological changes induced by cyclophosphamide On day 0, the lymphocyte % of all test animal groups was between 43-46%, and no significant differences were observed between the groups on this day. All groups treated with cyclophosphamide had significantly (p<0.001) higher lymphocyte counts when compared to the placebo group. As displayed in Table 8A and Figure 5A, a significant (p<0.001) decrease in lymphocyte % was observed with commercial synthetic cinnamic acid (0.6 mg / tablet) low dose treatment and commercial synthetic cinnamic acid (6 mg / tablet) low dose treatment and GCSF (granulocyte colony stimulating factor) treatment when compared to the CP control (cyclophosphamide) group. GCSF (granulocyte colony stimulating factor) significantly (p<0.001) reversed cyclophosphamide-induced lymphocyte mobilization compared to the CP control group. The reversal observed in low dose treatment of commercial synthetic cinnamic acid (0.6 mg / tablet) and high dose treatment of commercial synthetic cinnamic acid (6 mg / tablet) was better than pure picroside 1, salt-free degraded picroside 1 (cinnamic acid of the present disclosure), and comparable to that of the GCSF (granulocyte colony stimulating factor) treatment group.

[0136] Effects of low-dose commercial synthetic cinnamic acid (0.6mg / tablet) and high-dose commercial synthetic cinnamic acid (6mg / tablet) on total leukocyte counts in cyclophosphamide-induced hematological changes On day 0, the total white blood cell counts of all test groups of animals ranged between 7139 and 7644. No significant differences were observed between the groups on this day. Administration of cyclophosphamide caused a significant (p<0.001) decrease in white blood cell counts on day 3 by all groups receiving cyclophosphamide when compared to the placebo group. Treatment with low dose of commercially available synthetic cinnamic acid (0.6 mg / tablet) and high dose of commercially available synthetic cinnamic acid (6 mg / tablet) and treatment with GCSF (granulocyte colony stimulating factor) significantly (p<0.001) reversed the cyclophosphamide-induced decrease in white blood cell counts when compared to the cyclophosphamide control, as displayed in Table 8A and Figure 5A. The reversal observed in the low dose commercial synthetic cinnamic acid (0.6 mg / tablet) treatment and the high dose commercial synthetic cinnamic acid (6 mg / tablet) treatment was comparable to that of the GCSF (granulocyte colony stimulating factor) treatment group, and better than pure picroside 1, salt-free degraded picroside 1 (cinnamic acid of the present disclosure).

[0137] Effects of low-dose commercial synthetic cinnamic acid (0.6mg / tablet) and high-dose commercial synthetic cinnamic acid (6mg / tablet) on platelet counts in cyclophosphamide-induced hematological changes On day 0, the platelet counts of all test groups of animals were between 735,700 and 772,500. No significant differences were observed between the groups on this day. Administration of cyclophosphamide caused a significant (p<0.05) decrease in platelet counts in all of the cyclophosphamide-treated groups compared to the placebo group. The low-dose commercial synthetic cinnamic acid (0.6 mg / tablet) treatment and the high-dose commercial synthetic cinnamic acid (6 mg / tablet) treatment did not cause a significant change in platelet counts on day 10 when compared to the cyclophosphamide control group, as displayed in Table 8A and Figure 5A; however, there was also no significant change in the low-dose commercial synthetic cinnamic acid (0.6 mg / tablet) treatment and the high-dose commercial synthetic cinnamic acid (6 mg / tablet) treatment when compared to the placebo group. Treatment with GCSF (granulocyte colony-stimulating factor) caused a significant (p<0.05) increase in platelet counts compared to the cyclophosphamide control group.

[0138] Table 8A: Effects of low-dose commercial synthetic cinnamic acid (0.6 mg / tablet) treatment and high-dose commercial synthetic cinnamic acid (6 mg / tablet) treatment on hematological parameters in cyclophosphamide-induced neutropenia. [Table 8A-1] [Table 8A-2] When compared with the placebo group, $ p<0.05, $$ p<0.01, # p<0.001 * p<0.01, *** p<0.001 compared with CP controls

[0139] Inference: From Table 8A, it is clear that the test compounds (pure picroside 1, decomposed picroside without salt (cinnamic acid of the present disclosure)), commercial cinnamic acid (0.6 mg / tablet) low dose treatment, and commercial cinnamic acid (6 mg / tablet) high dose treatment were administered as one tablet per animal per day for 5 days starting from day 5; CP control was administered with a placebo tablet. Furthermore, from Table 8A, it is clear that pure picroside 1 and decomposed picroside 1 without salt (cinnamic acid of the present disclosure) show better antineutropenic activity. Commercial synthetic cinnamic acid (0.6 mg / tablet) low dose treatment and commercial synthetic cinnamic acid (6 mg / tablet) high dose treatment are also active in combating neutropenia.

[0140] Effects of low-dose commercial synthetic cinnamic acid (0.6mg / tablet) treatment and high-dose commercial synthetic cinnamic acid (6mg / tablet) treatment on SGOT (serum glutamic oxaloacetic transaminase) (IU / L) in cyclophosphamide-induced neutropenia The cyclophosphamide control group showed significantly (p<0.001) elevated SGOT (serum glutamic oxaloacetic transaminase) levels after cyclophosphamide administration when compared to the placebo group. As shown in Table 8B and FIG. 5B, a significant (p<0.001) decrease in SGOT (serum glutamic oxaloacetic transaminase) levels was observed following administration of commercial synthetic cinnamic acid (0.6 mg / tablet) low dose treatment and commercial synthetic cinnamic acid (6 mg / tablet) high dose treatment when compared to the cyclophosphamide control group. There was a significant (p<0.001) decrease in SGOT (serum glutamic oxaloacetic transaminase) in the GCSF (granulocyte colony stimulating factor) treatment group when compared to the cyclophosphamide control group. However, this decrease was not superior to the decrease observed in treatment with pure picroside 1 and salt-free decomposed picroside 1 (cinnamic acid of the present disclosure).

[0141] Effects of low-dose commercial synthetic cinnamic acid (0.6mg / tablet) treatment and high-dose commercial synthetic cinnamic acid (6mg / tablet) treatment on SGPT (serum glutamic pyruvic transaminase) (IU / L) in cyclophosphamide-induced neutropenia Administration of cyclophosphamide significantly reduced the incidence of SGPT in the cyclophosphamide control group compared with the placebo group. (Serum glutamic pyruvic transaminase) Administration of low dose commercial synthetic cinnamic acid (0.6 mg / tablet) and high dose commercial synthetic cinnamic acid (6 mg / tablet) to animals significantly increased SGPT levels compared to the cyclophosphamide control group, as shown in Table 8B and Figure 5B. (Serum glutamic pyruvic transaminase) The low dose of commercially available synthetic cinnamic acid (0.6 mg / tablet) and the high dose of commercially available synthetic cinnamic acid (6 mg / tablet) caused a significant reversal in the levels of SGPT (p<0.001). (serum glutamic pyruvic transaminase) The reduction was comparable to that of SGPT (serum glutamic pyruvic transaminase) levels achieved in treatment with GCSF (granulocyte colony stimulating factor) (p<0.001). However, this reduction was not superior to that observed in treatment with pure picroside 1 and salt-free decomposed picroside 1 (cinnamic acid of the present disclosure).

[0142] Effects of low-dose commercial synthetic cinnamic acid (0.6 mg / tablet) and high-dose commercial synthetic cinnamic acid (6 mg / tablet) on serum creatinine levels (mmol / L) in cyclophosphamide-induced neutropenia Administration of cyclophosphamide caused significantly (p<0.001) elevated levels of creatinine when compared to the placebo group. As shown in Table 8B and FIG. 5B, the commercially available synthetic cinnamic acid (0.6 mg / tablet) increased creatinine levels compared to the cyclophosphamide control group. Low-dose treatment and commercially available synthetic cinnamic acid (6 mg / tablet). High-dose treatment A significant (p<0.001) reduction in creatinine values ​​was observed following administration of cyclophosphamide. There was a significant (p<0.001) reduction in creatinine levels in the GCSF-treated group when compared to the cyclophosphamide control group. Commercially available synthetic cinnamic acid (0.6 mg / tablet) Low-dose treatment and commercially available synthetic cinnamic acid (6 mg / tablet). High-dose treatment The reduction in creatinine levels by was comparable to and better than GCSF, pure picroside 1, and salt-free degraded picroside 1 (cinnamic acid of the present disclosure) treatment groups.

[0143] Effect of low dose treatment with commercially available synthetic cinnamic acid (0.6 mg / tablet) and high dose treatment with commercially available synthetic cinnamic acid (6 mg / tablet) on blood urea nitrogen (mmol / L) in cyclophosphamide-induced nephrotoxicity. Administration of cyclophosphamide caused a significant (p<0.001) increase in blood urea nitrogen (BUN) levels when compared to the placebo group. As shown in Table 8B and Figure 5B, the commercially available synthetic cinnamic acid (0.6 mg / tablet) increased blood urea nitrogen (BUN) levels significantly compared to the cyclophosphamide control group. Low-dose treatment and commercially available synthetic cinnamic acid (6 mg / tablet). High-dose treatment There was a significant (p<0.001) decrease in BUN levels following administration of. There was a significant (p<0.001) decrease in BUN levels in the GCSF (granulocyte colony stimulating factor) treated group when compared to the cyclophosphamide control group. Commercially available synthetic cinnamic acid (0.6 mg / tablet) Low-dose treatment and commercially available synthetic cinnamic acid (6 mg / tablet). High-dose treatment The reduction in BUN levels in GCSF (granulocyte colony stimulating factor), pure picroside 1, and salt-free degraded picroside 1 (cinnamic acid of the present disclosure) was similar in each group.

[0144] Effect of low dose treatment with commercially available synthetic cinnamic acid (0.6 mg / tablet) and high dose treatment with commercially available synthetic cinnamic acid (6 mg / tablet) on uric acid (mg / L) in cyclophosphamide-induced nephrotoxicity. Administration of cyclophosphamide caused a significant increase (p<0.001) in serum uric acid levels in the cyclophosphamide control group (p<0.001). Low-dose treatment and commercially available synthetic cinnamic acid (6 mg / tablet). High-dose treatment was administered to the animals, which resulted in a significant improvement (p<0.001) in uric acid levels compared to the cyclophosphamide control group, as shown in Table 8B and Figure 5B. Treatment with GCSF (granulocyte colony stimulating factor) also caused a significant decrease (p<0.001) in uric acid levels when compared to the cyclophosphamide control group. Commercially available synthetic cinnamic acid (0.6 mg / tablet) Low-dose treatment and commercially available synthetic cinnamic acid (6 mg / tablet). High-dose treatment The reduction in serum uric acid levels by was comparable to and better than GCSF (granulocyte colony stimulating factor), pure picroside 1, and salt-free degraded picroside 1 (cinnamic acid of the present disclosure) treatment groups.

[0145] Table 8B: Effects of low dose commercial synthetic cinnamic acid (0.6 mg / tablet) treatment and high dose commercial synthetic cinnamic acid (6 mg / tablet) treatment on hepatic and renal parameters in cyclophosphamide-induced neutropenia. [Table 8B] $ p<0.05, $$ p<0.01, # p<0.001 compared with placebo group * p<0.01, *** p<0.001 compared with CP controls

[0146] Inference: From Table 8B, it is clear that the test compounds (pure picroside 1, decomposed picroside without salt (cinnamic acid of the present disclosure)), commercial cinnamic acid (0.6 mg / tablet) low dose, and commercial cinnamic acid (6 mg / tablet) high dose were administered one tablet per animal per day for 5 days starting from day 5, and CP control (cyclophosphamide) was administered together with placebo tablets. Furthermore, from Table 8B, it is clear that pure picroside 1 and decomposed picroside 1 without salt (cinnamic acid of the present disclosure) show improved liver and kidney parameters in neutropenic animal models compared to the known antineutropenic protein recombinant drug GCSF (granulocyte colony stimulating factor).

[0147] Experiment 11: Comparison of the lethal dose of the disclosed cinnamic acid (decomposed picroside 1 without salt) with the amount required for efficacy in alleviating neutropenia The lethal dose (LD50) of commercially available synthetic cinnamic acid was 5000 mg / kg, but the effective dose of cinnamic acid was found to be 24 mg / kg. The animal efficacy doses of cinnamic acid prepared according to the present disclosure were 0.6 mg / mouse, or 0.6 mg / 25 g mouse, or 6 mg / 250 g mouse, or 24 mg / kg mouse.

[0148] The human dose of cinnamic acid prepared according to the present disclosure is the AED (animal equivalent dose) divided by 12.3 = 24 / 12.3 = 1.95 mg / kg HED (human equivalent dose). For an individual weighing 65 kg = 60 x 1.95 mg = 117 mg / dose

[0149] The regimen was administered for 5 days. Hence, the total HED (human equivalent dose) for patients undergoing chemotherapy would be 1117mg x 5 = 585mg / dose taken by cancer patients with lower stage cancer.

[0150] The lethal dose (LD50) of commercially available synthetic cinnamic acid was 5000mg / kg in rats, while the effective dose for antineutropenic activity of the disclosed cinnamic acid (degraded picroside 1 without salt) was found to be 24mg / kg. Thus, the effective animal dose of 24mg / kg, when administered to the human equivalent dose of 2mg / kg, is clearly non-toxic when administered orally to neutropenic patients.

[0151] The effective animal dose of cinnamic acid prepared according to the present disclosure was 6 mg / 25 g mouse or 60 mg / 250 g mouse, or 240 mg cinnamic acid / kg. The human dosage of cinnamic acid prepared according to the present disclosure is the AED (animal equivalent dose) divided by 12.3, 240 / 12.3=19.51 mg / kg HED (human equivalent dose). For an individual weighing 65 kg = 60 x 19.51 mg = 1170 mg / dose

[0152] The dosage was administered for 5 days. Hence, the total HED (human equivalent dose) for chemotherapy patients is 1170mg x 5 = 5850mg / dose, taken by cancer patients with more advanced stage cancer.

[0153] Technical advancement and economic significance The above disclosure provides several technical advantages, including but not limited to the realization of pharmaceutical formulations and the process of preparation of cinnamic acid: Effectively improves neutrophil counts as well as other blood cell counts without serious adverse events; • It may be used in cases of cyclophosphamide-induced neutropenia; ● Improved patient compliance; and - may be suitably administered via oral, injection and inhalation routes; has.

[0154] Throughout this specification the word "comprises" or variations such as "comprises" or "comprises" will be understood to imply the inclusion of stated elements, integers or steps, or groups of elements, integers or steps, but not the exclusion of other elements, integers or steps, or groups of elements, integers or steps.

[0155] The use of the phrase "at least" or "at least one" suggests the use of one or more elements, ingredients, or amounts to achieve one or more desired objectives or results in the disclosed embodiments.

[0156] Any discussion of documents, acts, materials, devices, articles, or the like which has been included in this specification is for the purpose of providing a context for the disclosure only, and should not be construed as an admission that any or all of those matters form part of the prior art base or existed as common general knowledge in the field relevant to the disclosure prior to the priority date of this application.

[0157] Numerical values ​​mentioned for various physical parameters, dimensions or quantities are approximate only, and values ​​higher / lower than the numerical values ​​assigned to the parameters, dimensions or quantities are assumed to be within the scope of the disclosure, unless specifically stated to the contrary in the specification.

[0158] While considerable emphasis has been placed herein on the components and component parts of the preferred embodiments, it will be understood that many embodiments may be made and many changes may be made in the preferred embodiments without departing from the principles of the disclosure. These and other variations in the preferred embodiments as well as other embodiments of the present disclosure will be apparent to those skilled in the art from the disclosure herein, and it should be undoubtedly understood that the foregoing description is to be construed as merely illustrative of the present disclosure and not as limiting.

Claims

1. below: a) cinnamic acid as a pharma- ceutical active agent in an amount ranging from 0.1% to 10% by weight relative to the total weight of the formulation; and b) A pharmaceutical formulation comprising at least one excipient in an amount ranging from 90% to 99.9% by weight relative to the total weight of the formulation.

2. The pharmaceutical formulation according to claim 1 , wherein the excipient is at least one selected from the group consisting of a solvent, a colorant, a lubricant, a diluent and a disintegrant.

3. 3. The pharmaceutical formulation of claim 2, wherein the solvent is at least one selected from water, ethanol, and isopropyl alcohol; the colorant is selected from iron oxide yellow NF; dye brilliant blue supra; FD&C Green No. 3; and sunset yellow; the lubricant is at least one selected from magnesium stearate, talc, silica, and stearic acid; the disintegrant is at least one selected from carboxymethylcellulose and hydroxypropylmethylcellulose; and the diluent is selected from polyethylene glycol, dimethylsulfoxide, ethyl lactate, and a combination mixture of D-mannitol-xylitol-microcrystalline cellulose-crospovidone-dicalcium phosphate anhydrous.

4. 10. The pharmaceutical formulation of claim 1, wherein the formulation is in a form selected from an oral formulation, an injectable formulation, a dry powder inhalation formulation, a metered dose inhalation formulation, an ointment, a gel, a patch, an ophthalmic formulation, and a spray.

5. 5. The pharmaceutical formulation of claim 4, wherein the oral formulation comprises cinnamic acid in an amount ranging from 0.1% to 5% by weight, based on the total weight of the formulation, and at least one excipient in an amount ranging from 95% to 99.9% by weight, based on the total weight of the formulation, the excipient being at least one selected from the group consisting of solvents, colorants, lubricants, diluents and disintegrants.

6. 6. The pharmaceutical formulation of claim 5, wherein the solvent is at least one selected from water, ethanol, and isopropyl alcohol; the colorant is selected from iron oxide yellow NF; dye brilliant blue supra; FD&C Green No. 3; and sunset yellow; the lubricant is at least one selected from magnesium stearate, talc, silica, and stearic acid; the disintegrant is at least one selected from carboxymethylcellulose and hydroxypropylmethylcellulose; and the diluent is selected from polyethylene glycol, dimethylsulfoxide, ethyl lactate, and a combination mixture of D-mannitol-xylitol-microcrystalline cellulose-crospovidone-dicalcium phosphate anhydrous.

7. 5. The pharmaceutical formulation of claim 4, wherein the injectable formulation comprises cinnamic acid in an amount ranging from 1% to 10% by weight, based on the total weight of the formulation, and at least one excipient selected from the group consisting of polyethylene glycol, water, ethanol, propylene glycol, ethyl lactate and dimethyl sulfoxide, in an amount ranging from 90% to 99% by weight, based on the total weight of the formulation.

8. The dry powder inhalation formulation comprises: a. micronized cinnamic acid having a particle size ranging from 0.1 μm to 10 μm, in an amount ranging from 1% to 5% by weight relative to the total weight of the formulation; b. a first lactose having a particle size ranging from 20 μm to 300 μm, in an amount ranging from 10% to 30% by weight relative to the total weight of the formulation; c. a second lactose having a particle size ranging from 0.1 μm to 10 μm, in an amount ranging from 75% to 85% by weight relative to the total weight of the formulation; and d. excipients, in an amount ranging from 0.1% to 1% by weight relative to the total weight of the formulation; Including, The pharmaceutical formulation according to claim 4, wherein the mass ratio of the first lactose to the second lactose is in the range of 1:2 to 1:

5.

9. 9. The pharmaceutical formulation of claim 8, wherein the excipient is magnesium stearate.

10. 9. The pharmaceutical formulation of claim 8, wherein the mass median aerodynamic diameter (MMAD) of the dry powder inhalation formulation is in the range of 2 μm to 5 μm.

11. 9. The pharmaceutical formulation of claim 8, wherein the average particle size (d10) of the first lactose is in the range of 35 μm to 65 μm; the average particle size (d50) of the first lactose is in the range of 95 μm to 125 μm; and the average particle size (d90) of the first lactose is in the range of 160 μm to 190 μm.

12. 9. The pharmaceutical formulation of claim 8, wherein the average particle size (d10) of the second lactose is in the range of 0.01 μm to 1 μm; the average particle size (d50) of the second lactose is in the range of 1 μm to 5 μm; and the average particle size (d90) of the second lactose is in the range of 1 μm to 10 μm.

13. 2. The pharmaceutical formulation of claim 1, wherein the cinnamic acid is selected from trans-cinnamic acid and cis-cinnamic acid.

14. 2. The pharmaceutical formulation of claim 1, wherein the cinnamic acid is administered at a dose ranging from 1.5 mg / kg body weight to 30 mg / kg body weight.

15. 2. The pharmaceutical formulation of claim 1, wherein the formulation has antineutropenic activity, antituberculous activity, antimalarial activity, antiviral activity against COVID-19, antibacterial activity and cardiovascular activity.

16. 10. The pharmaceutical formulation of claim 1 for use in a method of treating neutropenia, tuberculosis, malaria, viral infections, bacterial infections and cardiovascular disease in a mammal comprising administering to the mammal a therapeutically effective amount of cinnamic acid in an amount ranging from 1.5 mg / kg to 30 mg / kg of body weight.

17. 17. The pharmaceutical formulation of claim 16, wherein the mammal is a human.

18. 17. The pharmaceutical formulation of claim 16, wherein the therapeutically effective amount of cinnamic acid is in the range of 15 mg / kg body weight to 25 mg / kg body weight.

19. 10. The pharmaceutical formulation of claim 1 for use in the treatment of neutropenia, tuberculosis, malaria, viral infections, bacterial infections and cardiovascular diseases.

20. A process for the preparation of cinnamic acid comprising the following steps: a) mixing picroside 1 in a first fluid medium, followed by adding at least one alkali hydroxide under stirring to obtain a mixture; b) maintaining the mixture at a temperature ranging from 25° C. to 35° C. for a predetermined period of time, and subsequently neutralizing the mixture with hydrochloric acid to obtain a neutralized mixture; c) removing the fluid medium from the neutralized mixture under vacuum at a temperature ranging from 40° C. to 50° C. to obtain a dry powder containing decomposed picroside 1 along with salts; d) mixing a second fluid medium with the dry powder to obtain a solution containing salt-free decomposed picroside 1; and e) decanting the solution followed by evaporation of the second fluid medium from the solution at a temperature ranging from 40°C to 50°C to obtain cinnamic acid (decomposed picroside 1 without salt); The process comprising:

21. 21. The process of claim 20, wherein the first fluid medium is water and the second fluid medium is methanol, ethanol, propanol and butanol.

22. 21. The process of claim 20, wherein the alkali hydroxides are sodium hydroxide, potassium hydroxide and calcium hydroxide.

23. 21. The process of claim 20, wherein the ratio of picroside 1 to alkali hydroxide is 0.01:

50.

24. 21. The process of claim 20, wherein the predetermined period of time is in the range of 30 minutes to 2 hours.

25. 21. Cinnamic acid prepared as described in claim 20 for use in the treatment of neutropenia, tuberculosis, malaria, viral infections, bacterial infections and cardiovascular diseases, having a therapeutically effective amount ranging from 1.5 mg / kg body weight to 30 mg / kg body weight.

26. 21. A cinnamic acid prepared as claimed in claim 20 for use in the treatment of neutropenia, tuberculosis, malaria, viral infections, bacterial infections and cardiovascular diseases, wherein said cinnamic acid is present in an amount ranging from 0.1% to 10% by weight, together with at least one excipient in an amount ranging from 90% to 99.9% by weight, relative to the total weight of the formulation.

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