Herbal Nutraceutical Formulation to Reduce Oxidative Stress, Viral and Microbial Infections, and Inflammation

Abstract

The main objective of the present invention is to develop an herbal formulation with high polyphenol concentrations. A therapeutic effect of ingredients in herbal composition with Antioxidants, Anti-inflammatories, Anti Viral & Anti Microbial properties to reduce Oxidative Stress, Inflammation, Viral and Microbial Infections. The herbal composition promotes management of diabetes, regulation of cholesterol, also immune system processes that helps to fight against viral and bacterial infections, colds, or the flu, skin ailments such as Psoriasis & eczema. Herbal Composition comprises synergistic combination of Moringa oleifera Leaf Powder, Spirulina & Aqueous extracts of Oregano Vulgare Leaf Extract, Shilajit extract, Rosemary Leaf extract, Pomegranate Fruit Preel extract, Amla fruit extract, Fenugreek Seed Extract, Curcumin Extract, Piperine extract. Therefore, the chosen herbal blend provides the body with all of the vitamins, minerals, and nutrients it needs to continuously boost health.

Claims

1. A composition for reducing oxidative stress, the composition is comprised of Moringa Oleifera leaf powder, Oregano Vulgare leaf extract, Shilajit extract, Rosemary leaf extract, Pomegranate peel extract, Spirulina, Amla fruit extract, Fenugreek seed extract, Curcuma longa extract, and Piperine.

2. The composition of claim 1, wherein said Moringa oleifera leaf powder is present in an amount between 100 mg per dose and 150 mg per dose.

3. The composition of claim 1, wherein said Oregano Vulgare leaf extract is present in an amount between 30 mg per dose and 80 mg per dose.

4. The composition of claim 1, wherein said Rosemary leaf extract is present in an amount between 30 mg per dose and 40 mg per dose.

5. The composition of claim 1, wherein said Shilajit extract is present in an amount of approximately 40 mg per dose.

6. The composition of claim 1, wherein said Pomegranate peel extract is present in an amount between 30 mg per dose and 40 mg per dose.

7. The composition of claim 1, wherein said Spirulina is present in an amount of approximately 40 mg per dose.

8. The composition of claim 1, wherein said Amla fruit extract is present in an amount between 40 mg per dose and 50 mg per dose.

9. The composition of claim 1, wherein said Fenugreek seed extract is present in an amount of approximately 60 mg per dose.

10. The composition of claim 1, wherein said Curcuma Longa extract is present in an amount of approximately 10 mg per dose.

11. The composition of claim 1, wherein said Piperine extract is present in an amount of approximately 2 mg per dose.

12. The composition of claim 1, wherein said Moringa oleifera leaf powder is present in an amount between 100 mg per dose and 150 mg per dose, wherein said Oregano Vulgare leaf extract is present in an amount between 30 mg per dose and 80 mg per dose, wherein said Rosemary leaf extract is present in an amount between 30 mg per dose and 40 mg per dose, wherein said Shilajit extract is present in an amount of approximately 40 mg per dose, wherein said Pomegranate peel extract is present in an amount between 30 mg per dose and 40 mg per dose, wherein said Spirulina is present in an amount of approximately 40 mg per dose, wherein said Amla fruit extract is present in an amount between 40 mg per dose and 50 mg per dose, wherein said Fenugreek seed extract is present in an amount of approximately 60 mg per dose, wherein said Curcuma Longa extract is present in an amount of approximately 10 mg per dose, wherein said Piperine extract is present in an amount of approximately 2 mg per dose.

13. The composition of claim 12, wherein said ingredients are contained a capsule.

14. The composition of claim 12, wherein the therapeutically effective amount of the composition is between four to six capsules.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0051] So that the manner in which the above recited features of the present invention can be understood in detail, a more particular description of the invention, briefly summarized above, may be had by reference to embodiments, some of which are illustrated in the appended drawings. It is to be noted, however, that the appended drawings illustrate only typical embodiments of this invention and are therefore not to be considered limiting of its scope, for the invention may admit to other equally effective embodiments.

[0052] FIG. 1A illustrates a graph illustrating increase in insulin secretion by the present herbal supplement composition 3, in accordance with one embodiment of the present invention;

[0053] FIG. 1B illustrates a graph illustrating increase in insulin secretion by glyburide and glucose, by present herbal composition 3 in accordance with one embodiment of the present invention;

[0054] FIG. 2A illustrates a graph illustrating increase in NBD-cholesterol uptake with U-18666A, by the present herbal supplement composition 3 in accordance with one embodiment of the present invention;

[0055] FIG. 2B illustrates a graph illustrating inhibition of NBD-cholesterol uptake with the herbal supplement composition 3, in accordance with one embodiment of the present invention;

[0056] FIG. 2C illustrates a graph illustrating effect of herbal supplement composition 3 on the viability of Caco-2 cells, in accordance with one embodiment of the present invention;

[0057] FIG. 3A illustrates a graph illustrating DPPH radical scavenging activity by the herbal supplement composition 3, in accordance with one embodiment of the present invention;

[0058] FIG. 3B illustrates a graph illustrating DPPH radical scavenging activity by the Trolox, by the herbal supplement composition 3 in accordance with one embodiment of the present invention;

[0059] FIG. 4A illustrates a graph illustrating inhibition of COX-1 activity by the herbal supplement Composition 3, in accordance with one embodiment of the present invention;

[0060] FIG. 4B illustrates a graph illustrating inhibition of COX-1 activity by Indomethacin by the herbal supplement composition 3 in accordance with one embodiment of the present invention;

[0061] FIG. 5A illustrates a graph illustrating inhibition of COX-2 activity by the by the herbal supplement composition 3, in accordance with one embodiment of the present invention;

[0062] FIG. 5B illustrates a graph illustrating inhibition of COX-2 activity by Indomethacin by the herbal supplement composition 3 in accordance with one embodiment of the present invention;

[0063] FIG. 6A illustrates a graph illustrating inhibition of LDL oxidation by the herbal supplement composition 3 in accordance with one embodiment of the present invention;

[0064] FIG. 6B illustrates a graph illustrating inhibition of LDL oxidation by Atorvastatin and ascorbic acid by the herbal supplement composition 3 in accordance with one embodiment of the present invention.

[0065] FIG. 07 illustrates a graph illustrating inhibition of LDL oxidation by the herbal supplement composition 2, in accordance with one embodiment of the present invention;

[0066] FIG. 08A illustrates a graph illustrating Increase in DPPH radical scavenging activity by the herbal supplement composition 2, in accordance with one embodiment of the present invention;

[0067] FIG. 08B illustrates a graph illustrating Increase in DPPH radical scavenging activity by the Trolox, in accordance with one embodiment of the by the herbal Supplement composition 2;

[0068] FIG. 9A illustrates a graph illustrating effect of Gallic acid on super oxide radicals scavenging activity by the herbal supplement composition 2 and,

[0069] FIG. 9B illustrates a graph illustrating effect of herbal Supplement composition 2 on super oxide radicals scavenging activity in accordance with one embodiment of the present invention.

[0070] While the present systems and methods have been described herein by way of example for several embodiments and illustrative drawings, those skilled in the art will recognize that the multiple embodiments disclosed hereinbelow are not limited to the embodiments or drawings described. It should be understood, that the drawings and detailed description thereto are not intended to limit embodiments to the particular form disclosed. Rather, the intention is to cover all modifications, equivalents and alternatives falling within the spirit and scope of the invention as defined by the appended claims. Any headings used herein are for organizational purposes only and are not meant to limit the scope of the description or the claims. As used herein, the word “can” and “may” is used in a permissive sense (i.e., meaning having the potential to), rather than the mandatory sense (i.e., meaning must). Similarly, the words “include”, “including”, and “includes” mean including, but not limited to.

DETAILED DESCRIPTION

Experimental Data

Experiment 1: To Evaluate ORAC (Oxygen Radical Absorbance Capacity) of Herbal Supplement Composition 1

[0071] The Objective of the experiment is to Evaluate ORAC(Oxygen Radical Absorbance Capacity) of Herbal Supplement composition. Oxygen Radical Absorbance Capacity (ORAC) tests are among the most acknowledged methods that measure antioxidant scavenging activity against oxygen radicals that are known to be involved in the pathogenesis of aging and many common diseases. ORAC 5.0 consists of five types of ORAC assays that evaluate the antioxidant capacity of a material against six primary reactive oxygen species (ROSs, commonly called “oxygen radicals”) found in humans: peroxyl radical, hydroxyl radical, superoxide anion, singlet oxygen. This is a comprehensive panel that evaluates the antioxidant capacity of a material against oxygen radicals

[0072] The ORAC tests are based on evaluating the capacity of an interested material to protect a probe (a fluorescent probe or chromagen) from its damage by ROSs. In all ORAC assays, an ROS inducer is introduced to the assay system.

[0073] The ROS inducer triggers the release of a specific ROS, which would degrade the probe and cause its emission wavelength or intensity change. When an antioxidant material presents in the environment, the antioxidant absorbs the ROS and preserves the probe from degradation. The degree of probe preservation indicates the antioxidant capacity of the material. Trolox is used as the reference standard, and the results are expressed as □mole Trolox equivalency per gram (or milliliter) of a tested material.

TABLE-US-00002 TABLE 1 ORAC Score (Oxygen Radical Absorbance Capacity) of Herbal Supplement composition 1 against free radicals Radicals Result ORAC against peroxyl radicals 5,219 μmole TE/serving size ORAC against hydroxyl 10,964 μmole TE/serving size radicals ORAC against peroxynitrite 543 μmole TE/serving size ORAC against super oxide 6,152 μmole TE/serving size anion ORAC against singlet oxygen 1,244 μmole TE/serving size

[0074] There are six predominant reactive species found in the body: peroxyl radicals, hydroxyl radicals, peroxynitrite, super oxide anion, singlet oxygen and hypochlorite. ORAC 5.0 provides a measure of the total antioxidant power of a food/nutrition product against the six predominant reactive species. The ORAC result is expressed as micromole Trolox equivalency (μmole TE) per gram. Serving size=2.17 g

Experiment 2: To Evaluate Effect of Herbal Supplement Composition 2 on 2,2-Diphenyl-1-picrylhydrazyl (DPPH) Radical Scavenging Activity

[0075] Briefly, 12.5 mg of the herbal composition 2 contents was weighed, dissolved in 312.5 μl of DMSO and the volume was made up to 10 ml with 100% methanol. Further dilutions were done in 100% methanol containing 3.125% of DMSO. The sample was found to be approximately 70% soluble upon visual inspection. For calculation purposes the sample was considered to be 100% soluble.

[0076] Gallic acid (reference inhibitor) preparation. For gallic acid, a stock of 200 μg/ml was prepared by dissolving 2 mg of gallic acid in 10 ml of methanol. Further dilutions were made as required with methanol. The sample was found to be approximately 100% soluble upon visual inspection.

[0077] DPPH assay was carried out as per the method of Vani et al [4]. In brief, the total reaction mixture contained methanol/vehicle buffer/positive control/various concentrations of test solution and DPPH to a final concentration of 0.132 mM. The reaction mixture was incubated at 25° C. for 20 minutes. Following incubation the absorbance was read at 510 nm using a micro-well plate reader (Molecular devices Versamax microplate reader). A control reaction was carried out without test samples.

[0078] The % inhibition values were calculated as follows:

[00001]$\%.Math..Math.\mathrm{inhibition}=\frac{\mathrm{Absorbance}.Math..Math.\left(\mathrm{control}\right)-\mathrm{Absorbance}.Math..Math.\left(\mathrm{test}\right)}{\mathrm{Absorbance}.Math..Math.\left(\mathrm{control}\right)}\times 100$

Where, Absorbance=Final Absorbance−Initial absorbance. IC.sub.50 was calculated using log-probit analysis.

[0079] Herbal composition 2 exhibited DPPH free radical scavenging activity in a dose dependent manner in the concentration range tested.

[0080] Herbal composition 2 exhibited an IC.sub.50 of 50.70 μg/ml in DPPH free radical scavenging assay

TABLE-US-00003 TABLE 02 IC.sub.50 data of Herbal composition 2 and reference inhibitor in DPPH radical scavenging assay IC.sub.50 (μg/ml) (95% Concentration % Inhibition Confidence Sample (μg/ml) (Mean ± SEM) Interval) Gallic Acid 0.5  8.71 ± 1.12 2.09 (Reference Inhibitor) 1.5 32.14 ± 2.42 (1.81-2.42) 2.5 60.58 ± 2.84 5 80.52 ± 0.57 Herbal composition 2 5  9.89 ± 5.79 50.70 10 13.59 ± 3.71 (42.84-59.81) 25 23.76 ± 1.56 50 41.36 ± 2.70 100 75.05 ± 3.22 250 93.13 ± 0.64 500 94.04 ± 0.78 1000 94.28 ± 0.54

Experiment 3: To Evaluate the Effect of Herbal Supplement Composition 2 on LDL Oxidation in a Cell-Free System

[0081] Briefly, 29.73 mg of the Herbal composition 2 contents was weighed, dissolved in 500 μl of DMSO and the volume was made up to 25 ml with phosphate buffered saline. The sample was found to be approximately 70% soluble upon visual inspection. The sample was filtered and the filtrate was used for the assay. For calculation purposes the sample was considered to be 100% soluble.

[0082] Trolox (reference inhibitor) preparation, For trolox, a stock of 8 mM was prepared by dissolving 2 mg of trolox in 2 ml of phosphate buffered saline. The sample was found to be approximately 100% soluble upon visual inspection. Further dilutions were made as required with phosphate buffered saline.

[0083] The assay was carried out as per the method of Hodgson et al [3] with modifications. In brief, the reaction mixture contained phosphate buffered saline/vehicle buffer/positive control/test sample of various concentrations and serum from fasting human. Copper sulphate solution was added to the reaction mixture to start the reaction. The reaction mix was mixed and read immediately at 234 nm using a UV-VIS spectrophotometer, for the initial reading, and incubated. After the completion of incubation period, the final absorbance was measured at 234 nm. A control reaction was carried out without test samples.

[0084] The % inhibition values were calculated as follows:

[00002]$\%.Math..Math.\mathrm{inhibition}=\frac{\mathrm{Absorbance}.Math..Math.\left(\mathrm{control}\right)-\mathrm{Absorbance}.Math..Math.\left(\mathrm{test}\right)}{\mathrm{Absorbance}.Math..Math.\left(\mathrm{control}\right)}\times 100$

Where, Absorbance=Final Absorbance−Initial absorbance. IC.sub.50 was calculated using log-probit analysis.

[0085] Herbal composition 2 was tested in LDL oxidation inhibition assay at concentrations ranging from 37.03 μg/ml to 1000 μg/ml. The sample exhibited inhibition of LDL oxidation in a dose dependent manner in the concentration range tested. The IC.sub.50 of Herbal composition in LDL oxidation inhibition assay was found to be 77.75 μg/ml.

TABLE-US-00004 TABLE 03 IC.sub.50 data of positive control and test herbal composition 2 in LDL oxidation inhibition assay IC.sub.50 % Inhibition (95% Confidence Sample Concentration (Mean ± SEM) Interval) Trolox 0.25 μM  48.80 ± 6.21  0.18 μM (Reference Inhibitor) 0.5 μM  73.07 ± 3.39  (0.06-0.29) 1.0 μM  76.33 ± 0.07  2.0 μM  79.04 ± 0.64  Herbal Composition 2 37.03 μg/ml  43.11 ± 12.62 77.75 μg/ml 111.11 μg/ml  45.93 ± 0.09  (57.09 ± 99.95) 333.33 μg/ml  75.23 ± 0.59  1000 μg/ml 100.00 ± 0.00 

Experiment 4: To Evaluate the Effect of Herbal Supplement Composition 2 on Super Oxide Radical Scavenging Activity

[0086] Briefly, 8 mg of the Herbal composition 2 contents was weighed, dissolved in 200 μl of DMSO and the volume was made up to 2 ml with 0.1M phosphate buffer pH 7.4. Further dilutions were done in 0.1M phosphate buffer pH 7.4 containing 10% of DMSO. The sample was found to be approximately 70% soluble upon visual inspection. For calculation purposes the sample was considered to be 100% soluble.

[0087] Gallic acid (reference inhibitor) preparation, For gallic acid, a stock of 500 μg/ml was prepared by dissolving 5 mg of gallic acid in 10 ml of 0.1M phosphate buffer pH 7.4. The sample was found to be approximately 100% soluble upon visual inspection. The assay was carried out as per the method of Yen et al. [5] with modifications. In brief, the reaction mixture contained 0.1M phosphate buffer pH 7.4/vehicle buffer/positive control/test sample of various concentrations, 292.5 μM NADH, 37.5 μM NBT and 15 μM PMS. The plate was mixed and incubated at 25° C. for 5 minutes. The absorbance was measured at 560 nm in a micro-plate reader (Versamax, Molecular devices). A control reaction was carried out without test samples.

[0088] The % inhibition values were calculated as follows:

[00003]$\%.Math..Math.\mathrm{inhibition}=\frac{\mathrm{Absorbance}.Math..Math.\left(\mathrm{control}\right)-\mathrm{Absorbance}.Math..Math.\left(\mathrm{test}\right)}{\mathrm{Absorbance}.Math..Math.\left(\mathrm{control}\right)}\times 100$

[0089] IC.sub.50 was calculated using log-probit analysis.

[0090] Herbal composition 2 was tested in super oxide radical scavenging assay at concentrations ranging from 5 μg/ml to 1000 μg/ml. The sample exhibited scavenging of super oxide radicals activity in a dose dependent manner in the concentration range tested. The IC.sub.50 of Herbal composition 2 in super oxide radical scavenging assay was found to be 68.37 μg/ml.

TABLE-US-00005 TABLE 04 IC.sub.50 data of positive control and test herbal composition 2 in super oxide radical scavenging assay IC.sub.50 (μg/ml) Concentration % Inhibition (95% Confidence Sample (μg/ml) (Mean ± SEM) Interval) Gallic acid 5 27.00 ± 3.37 17.86 (Reference Inhibitor) 10 32.89 ± 3.26 (14.53-21.76) 20 47.80 ± 4.26 40 69.50 ± 1.00 80 83.67 ± 0.00 Herbal Composition 2 5  4.09 ± 1.72 68.37 10  7.33 ± 2.63 (57.45-81.33) 25 28.33 ± 1.71 50 52.50 ± 1.11 100 62.37 ± 0.63 250 75.19 ± 0.14 500 87.24 ± 0.37 1000 93.66 ± 2.26

Experiment 4: To Evaluate ORAC(Oxygen Radical Absorbance Capacity) of Herbal Supplement Composition 1

[0091] The Objective of the experiment is to Evaluate ORAC(Oxygen Radical Absorbance Capacity) of Herbal Supplement composition. Oxygen Radical Absorbance Capacity (ORAC) tests are among the most acknowledged methods that measure antioxidant scavenging activity against oxygen radicals that are known to be involved in the pathogenesis of aging and many common diseases. ORAC 5.0 consists of five types of ORAC assays that evaluate the antioxidant capacity of a material against six primary reactive oxygen species (ROSs, commonly called “oxygen radicals”) found in humans: peroxyl radical, hydroxyl radical, superoxide anion, singlet oxygen, peroxynitrite and hypochlorite. This is a comprehensive panel that evaluates the antioxidant capacity of a material against oxygen radicals

[0092] The ORAC tests are based on evaluating the capacity of an interested material to protect a probe (a fluorescent probe or chromagen) from its damage by ROSs. In all ORAC assays, an ROS inducer is introduced to the assay system. The ROS inducer triggers the release of a specific ROS, which would degrade the probe and cause its emission wavelength or intensity change. When an antioxidant material presents in the environment, the antioxidant absorbs the ROS and preserves the probe from degradation. The degree of probe preservation indicates the antioxidant capacity of the material. Trolox is used as the reference standard, and the results are expressed as □mole Trolox equivalency per gram (or milliliter) of a tested material.

TABLE-US-00006 TABLE 5 ORAC Score (Oxygen Radical Absorbance Capacity) of Herbal Supplement composition 1 against free radicals Radicals Result ORAC against peroxyl radicals 3,857 μmole TE/serving size ORAC against hydroxyl 10,860 μmole TE/serving size radicals ORAC against peroxynitrite 480 μmole TE/serving size ORAC against super oxide 3,924 μmole TE/serving size anion ORAC against singlet oxygen 392 μmole TE/serving size

Experiment 5: Effect of Herbal Supplement Composition 3 on Insulin Release in Rat Islets

[0093] The objective of the experiment is to evaluate the effect of herbal supplement test composition on insulin secretion in rat pancreatic islets. Insulin, a peptide hormone, is secreted by pancreatic β-cells and is a key regulator in glucose homeostasis. Insulin deficiency leads to insulin-dependent (type 1) diabetes, whereas resistance to insulin action is common in non-insulin-dependent diabetes (type 2), obesity, and endocrine dysfunctions. The pancreatic β-cells are known to be sensitive to the glucose concentration in the blood. When the blood glucose levels are high they secrete insulin into the blood; when the levels are low they cease their secretion of this hormone into the general circulation. Insulin released by the β-cells is the main regulator of glucose levels. Stimulation of insulin release from β-cells is one of the major mechanisms by which natural products exhibit an anti-diabetic effect.

[0094] Firstly, the herbal supplement composition is dissolved in DMSO with sonication for 15 min and centrifuged at 10,000 rpm for 10 minutes at room temperature to remove insoluble debris. Glyburide (CAS: 10238-21-8) and Glucose are used as reference compounds in the present experiment. Glyburide (CAS: 10238-21-8) is dissolved in DMSO and used at 3 μM. Glucose is dissolved in water and used at 2.5 mM and 11.5 mM as a control. The rat pancreatic islets are isolated using standard in-house procedures. After culturing overnight, islets are starved in starvation media (KRBH buffer containing 0.01% BSA, no glucose) for about 1 hr. After starvation, islets are selected and added to 20 μl of KRBH buffer containing 0.01% BSA to each well of a 96 well plate (4 islets/well). The supplement composition is then added to each well at the indicated concentrations. Glucose is also added to control wells at a concentration of 2.5 mM for no insulin secretion and 11.5 mM for maximum glucose-stimulated insulin secretion, and the plate is incubated for 1 hr at 37° C. at 5% CO.sub.2.

[0095] At the end of the 1 hr incubation period, 20 μl of supernatant is gently transferred to another plate. Insulin secretion is measured using the Mercodia Rat Insulin ELISA kit according to the manufacturer's protocol. The DMSO concentration in the assay is maintained at less than equal to 1.0%. The data is fit to a non-linear regression using Graph pad Prism (version 5.0). The half maximal effective concentration (EC.sub.50) is obtained from the sigmoidal dose response and variable slope curve is generated. Statistical analyses are carried out using a one-way ANOVA followed by a Dunnett's multiple comparison test. The data represents five replicates for each condition.

[0096] FIG. 1A illustrates a graph 100 illustrating increase in insulin secretion by the present herbal Supplement composition, in accordance with one embodiment of the present invention. FIG. 1B illustrates a graph 150 illustrating increase in insulin secretion by glyburide and glucose, in accordance with one embodiment of the present invention. The supplement composition significantly increases insulin secretion in rat pancreatic islets in a dose-dependent manner with an EC.sub.50 of 47 μg/mL as illustrated in FIG. 1A of the present invention. However, a significant increase in insulin secretion is also observed with 3 μM Glyburide and 11.5 mM glucose. The error bars in FIG. 1A and FIG. 1B represent Mean±SEM (Standard Error of the Mean) where, *** P<0.001, ** P<0.01, * P<0.05, and one-way ANOVA is followed by Dunnett's test. Table 1 below provides data related to Insulin secretion (ng/ml) as a function of supplement composition concentration.

TABLE-US-00007 TABLE 6 Insulin secretion (ng/ml) as a function of supplement composition 3 concentration Supplement Composition, Insulin Secretion, μg/ml ng/ml (Mean ± SEM) 0  67.41 ± 36.77 12.5  89.43 ± 6.97  25 109.57 ± 22.67 50 152.31 ± 26.4  100 190.67 ± 30.03 200 211.89 ± 51.15

Experiment 6: To Evaluate Effect of Herbal Supplement Composition 3 on Cholesterol Uptake in Caco-2 Cells (Human Epithelial Colorectal Adenocarcinoma Cells)

[0097] The objective of the experiment is to evaluate the effect of the supplement composition on cholesterol uptake in Caco-2 (human epithelial colorectal adenocarcinoma) cells. Cholesterol is both an important structural component of cell membranes and an early intermediate in hormone and bile acid biosynthesis. The supplement composition is dissolved in Dimethyl Sulphoxide (DMSO) with sonication for 15 min and centrifuged at 10,000 rpm for 10 min at room temperature to remove insoluble debris. The reference compound used in the present experiment is U-18666A (CAS: 3039-71-2), which increases cholesterol uptake by inhibiting trafficking of synthesized cholesterol. The U-18666A is dissolved in DMSO and used at a concentration of about 1.25, 2.5, 5 and 10 μM. Moreover, U-18666A is used as a positive control. The reagent used is 3-hexanoyl-NBD Cholesterol. Caco-2 cells are maintained at 60-70% confluence in complete growth medium EMEM containing 20% FBS and required antibiotic. For the assay, cells are harvested using 0.25% trypsin EDTA. The trypsinized cells are centrifuged at 1200 rpm for 5 minutes and re-suspended in growth medium. Cells are seeded at a density of 10,000 cells per well in complete growth medium (EMEM+20% FBS+antibiotic) in poly D-lysine coated black plates, and incubated overnight at 37° C. in 5% CO.sub.2. After incubation, the plates are washed with Phosphate Buffered Saline (PBS) and incubated with 20 μg/ml NBD cholesterol in the presence of the test supplement composition and positive control U-18666A in PBS at pH 7.2. The cells are incubated for 48 hrs at 37° C. in 5% CO.sub.2. At the end of the incubation period, the plates are washed twice with PBS and cholesterol uptake is calculated by measuring NBD cholesterol fluorescence (Ex 485 nm, Em 535 nm). Cell viability studies are carried out using an XTT kit with varying concentration of supplement composition using the manufacturer's recommended protocol.

[0098] The DMSO concentration in the assay is maintained at less than equal to 1.0%. The data is fit to a non-linear regression using Graph pad Prism (version 5.0). The half maximal effective concentration (EC.sub.50) is obtained from the sigmoidal dose response and variable slope curve is generated. Statistical analyses are carried out using a one-way ANOVA followed by a Dunnett's multiple comparison test. The data represent five replicates for each condition.

[0099] FIG. 2A illustrates a graph 200 illustrating increase in NBD-cholesterol uptake with U-18666A, in accordance with one embodiment of the present invention. The results illustrate positive control standard U-18666A increased NBD-cholesterol uptake by up to 25% in Caco-2 cells. FIG. 2B illustrates a graph 250 illustrating inhibition of NBD-cholesterol uptake with the supplement composition, in accordance with one embodiment of the present invention. Particularly, the supplement composition significantly inhibited NBD-cholesterol uptake into Caco-2 cells in a dose-dependent manner.

[0100] FIG. 2C illustrates a graph 270 illustrating effect of supplement composition on the viability of Caco-2 cells, in accordance with one embodiment of the present invention. The decrease in NBD-cholesterol uptake is nota function of the effect of the test compound supplement composition on cell viability, as no effects are observed in the XTT assay up to 200 μg/mL. The Error bars in FIG. 2A, FIG. 2B, & FIG. 2C represent Mean±SEM where, *** P<0.001, ** P<0.01, * P<0.05, and one-way ANOVA followed by Dunnett's test. Table 2 below illustrates data related to cholesterol uptake (% activity) as a function of supplement concentration.

TABLE-US-00008 TABLE 7 Cholesterol uptake (% activity) as a function of Herbal supplement composition 3. Supplement Composition, % Activity μg/mL (Mean ± SEM) 0   100 ± 2.35 0.78 77.67 ± 9.94 1.56 86.86 ± 8.65 3.13 69.84 ± 3.44 6.25 68.65 ± 5.75 12.5 57.82 ± 3.46 25 46.02 ± 2.55 50 42.51 ± 4.22 100 42.45 ± 3.67 200 41.36 ± 2.13

Experiment 7: To Evaluate Effect of Herbal Supplement Composition 3 on 2,2-Diphenyl-1-Picrylhydrazyl (DPPH) Radical Scavenging Activity

[0101] The objective of this experiment is to evaluate the effect of the supplement concentration on DPPH radical scavenging activity in a cell-free assay. The supplement concentration is dissolved in DMSO with sonication for 15 min and centrifuged at 10,000 rpm for 10 min at room temperature to remove insoluble debris. 2,2-Diphenyl-1-picrylhydrazyl (DPPH) is used as a reagent for this experiment. DPPH is an antioxidant reagent that is used as a free radical trap or scavenger. The color of DPPH changes from red to yellow as its scavenging activity increases. This is measured by absorbance at 517 nm. The reference compound used is Trolox (CAS: 53188-07-1). The Trolox is dissolved in DMSO and used at different concentrations. Trolox is used as positive control.

[0102] A reaction mix is made in 200 μL methanol containing 200 μM of DPPH and various concentrations of the supplement. The mixture is mixed gently with a pipette and left to stand for 30 min in the dark, after which absorbance readings are taken at 517 nm. The change in DPPH absorbance is calculated relative to the absorbance of DPPH alone i.e. without the supplement. The DMSO concentration in the assay is maintained at less than equal to 1.0%. The data is fit to a non-linear regression using Graph pad Prism (version 5.0). The half maximal effective concentration (EC.sub.50) is obtained from the sigmoidal dose response and a vertical slope curve is generated. The data represent three replicates for each condition.

[0103] FIG. 3A illustrates a graph illustrating DPPH radical scavenging activity by the supplement composition, and FIG. 3B illustrates a graph illustrating DPPH radical scavenging activity by the Trolox, in accordance with one embodiment of the present invention. Particularly, the supplement composition illustrates dose-dependent DPPH scavenging activity, with an EC.sub.50 of 48 μg/mL. An increase in DPPH scavenging activity is also observed with Trolox, with an EC.sub.50 of 28 μg/mL. Particularly, the error bars in FIG. 3A and FIG. 3B represent Mean±SEM. Table 3 below illustrates DPPH scavenging (% activity) as a function of Herbal supplement concentration.

TABLE-US-00009 TABLE 8 DPPH scavenging (% activity) as a function of Herbal Supplement Composition 3. Supplement, μg/mL % Activity (Mean ± SEM) 0     0 ± 0.5    0.78  9.86 ± 0.13   1.56 11.88 ± 0.12   3.13 10.53 ± 30.075 6.3  13.8 ± 0.2    12.5 15.27 ± 1.12   25 32.03 ± 0.28   50 45.77 ± 3.33   100 78.57 ± 0.18   200  94.8 ± 1.85   400 83.92 ± 1.72  

Experiment 8: To Evaluate Effect of Herbal Supplement Composition 3 on COX-1 Enzyme Activity

[0104] The objective of this experiment is to evaluate the effect of the supplement composition on COX-1 activity in a biochemical assay. Cyclooxygenase (COX), also known as prostaglandin-endoperoxide synthase (PTGS, EC 1.14.99.1), is a bi functional enzyme that shows both cyclooxygenase and peroxidase activity. Two isoforms of this enzyme are COX-1 and COX-2. Particularly, COX-1 is constitutively expressed in many tissues. COX-2 is not expressed under normal conditions in most cells, but elevated during disease conditions. Cyclooxygenase converts arachidonic acid to hydroperoxy endoperoxide PGG2, and COX's peroxidase activity further converts PGG2 to the corresponding alcohol, PGH2 with a loss of oxygen radical. The released oxygen can be quantitatively measured using the Ampiflu (ADHP) substrate and in turn the activity of the enzyme is calculated from fluorescence measurements.

[0105] The reagents used for this experiment are Cyclooxygenase 1 from sheep (Cat #C0733), Tris-HCl, EDTA, Hematin (Cat #51280), Arachidonic acid (Cat #A3611) and ADHP (10-Acetyl-3, 7-dihydroxyphenoxazine; Cat #90101) procured from Sigma-Aldrich. The supplement composition is dissolved in DMSO with sonication for 15 min and centrifuged at 10,000 rpm for 10 min at room temperature to remove insoluble debris. The reference compound used is Indomethacin (CAS: 53-86-1) and is dissolved in DMSO and used at 0.5, 5 and 50 μM concentrations. The assay is run in a 384 well format in a final volume of 20 μL. The enzyme is taken in Tris-HCl buffer (pH 8.0) containing EDTA (3 μM) and Hematin (15 μM). The enzyme is treated with or without different concentrations of the supplement or the standard Indomethacin and incubated for 10 min. At the end of 10 minutes, Arachidonic acid (100 μM) and ADHP (30 μM) are mixed and added to the reaction mixture.

[0106] After 5 minutes, the enzyme activity is calculated from the fluorescence measurements (Ex 535 nm, Ex 587 nm). The DMSO concentration in the assay is maintained at less than equal to 1.0%. The data is fit to a non-linear regression using Graph pad Prism (version 5.0). The half maximal inhibitory concentration (IC.sub.50) is obtained from the sigmoidal dose response and variable slope curve is generated. Statistical analyses are carried out using a one-way ANOVA followed by a Dunnett's multiple comparison test.

[0107] FIG. 4A illustrates a graph illustrating inhibition of COX-1 activity by the supplement composition, in accordance with one embodiment of the present invention. Particularly, the test supplement showed significant inhibition of COX-1 activity with an IC.sub.50 of 17 μg/mL. FIG. 4B illustrates a graph illustrating inhibition of COX-1 activity by Indomethacin, in accordance with one embodiment of the present invention. The standard inhibitor, Indomethacin, showed significant inhibition of COX-1 activity at 0.5, 5 and 50 μM. The error bars in FIG. 4A and FIG. 4B represent Mean±SEM, where *** P<0.001, ** P<0.01, * P<0.05, and one-way ANOVA followed by Dunnett's test. Table 4 below illustrates COX-1 activity (% activity) as a function of test supplement concentration.

TABLE-US-00010 TABLE 9 COX-1 activity (% activity) as a function of Herbal Supplement Composition 3 Supplement, μg/mL % Activity (Mean ± SEM) 0   100 ± 6.2  0.41 107.4 ± 4.97 1.23   100 ± 6.2  3.7 85.48 ± 12.1 11.1 75.16 ± 7.07 33.3 19.84 ± 3.39 100  5.81 ± 4.84

Experiment 9: To Evaluate Effect of Herbal Supplement Composition 3 on COX-2 Enzyme Activity

[0108] The objective of the experiment is to evaluate the effect of the Herbal supplement composition on COX-2 activity in a biochemical assay. The reagents used for the evaluation are recombinant Cyclooxygenase 2 (Cat #C0858), Tris-HCl, EDTA, Hematin (Cat #51280), Arachidonic acid (Cat #A3611) and ADHP (10-Acetyl-3, 7-dihydroxyphenoxazine; Cat #90101), which are procured from Sigma-Aldrich.

[0109] Firstly, the supplement is dissolved in DMSO with sonication for 15 min and centrifuged at 10,000 rpm for 10 min at room temperature to remove insoluble debris. The reference compound used is Indomethacin (CAS: 53-86-1) and the same is dissolved in DMSO and used at 0.5, 5 and 50 μM concentrations. The assay is run in a 384 well format in a final volume of 20 μL. The enzyme is taken in Tris-HCl buffer (pH 8.0) containing EDTA (3 μM) and Hematin (15 μM). The enzyme is treated with or without different concentrations of the test supplement or the standard Indomethacin and incubated for 10 min. At the end of 10 minutes, Arachidonic acid (100 μM) and ADHP (30 μM) are mixed and added to the reaction mixture. After 5 minutes, enzyme activity is calculated from the fluorescence measurements (Ex 535 nm, Em 587 nm). The DMSO concentration in the assay is maintained at less than equal to 1.0%. The data is fit to a non-linear regression using Graph pad Prism (version 5.0). The half maximal inhibitory concentration (IC.sub.50) is obtained from the sigmoidal dose response and a variable slope curve is generated. Statistical analyses are carried out using a one-way ANOVA followed by a Dunnett's multiple comparison test.

[0110] FIG. 5A illustrates a graph illustrating inhibition of COX-2 activity by supplement composition, in accordance with one embodiment of the present invention. The supplement composition illustrates significant inhibition of COX-2 activity with an IC.sub.50 of 14 μg/mL. FIG. 5B illustrates a graph illustrating inhibition of COX-2 activity by Indomethacin, in accordance with one embodiment of the present invention. The standard inhibitor Indomethacin illustrates significant inhibition of COX-2 activity at 0.5, 5 and 50 μM. The error bars represent Mean±SEM where, *** P<0.001, ** P<0.01, * P<0.05, and one-way ANOVA is followed by Dunnett's test. Table 5 below illustrates COX-2 activity (% activity) as a function of Herbal supplement concentration.

TABLE-US-00011 TABLE 10 COX-2 activity (% activity) as a function of Herbal Supplement Composition 3 Supplement, μg/mL % Activity (Mean ± SEM) 0   100 ± 0.88 2.1 97.67 ± 1.9  6.2 79.48 ± 5.18 18.5 40.96 ± 0.52 55.6 18.16 ± 1.3  166.7   5.1 ± 0.15 500  2.25 ± 1.1 

Experiment 10: To Evaluate the Effect of Herbal Supplement Composition 3 LDL Oxidation in a Cell-Free System

[0111] The objective of the experiment is to evaluate the effect of the test supplement on LDL oxidation in a cell-free system. Cardiovascular disease (CVD) is the leading cause of death in developed countries. Atherosclerosis is a response of blood vessels to injury at the beginning of the formation of an atherosclerotic plaque. Low-density lipoprotein (LDL) is a major cholesterol carrier in the bloodstream, and the concentration of LDL cholesterol is directly correlated with the incidence of CVD. Increased oxidized LDL (OxLDL) acts as an atherogenic factor by triggering an inflammatory process. OxLDL can induce cell activation, secretion of inflammatory mediators, and expression of adhesion molecules. The reagents used are Lipoproteins, Low Density, Human Plasma and Atorvastatin.

[0112] The Herbal supplement composition is dissolved in DMSO with sonication for about 15 min and centrifuged at 10,000 rpm for 10 min at room temperature to remove insoluble debris. The reference compound for the present experiment is Atorvastatin (CAS: 134523-03-8) and is dissolved in DMSO. Atorvastatin is used at different concentrations and used as positive control. LDL samples of about 50 μg protein/mL are pre-incubated at 37° C. in a medium containing 10 mM phosphate buffer having pH 7.4 with different supplement concentrations.

[0113] After 5 minutes oxidation is initiated by the addition of CUSO.sub.4 at 25 μM. The results are monitored by measuring the increase in absorbance at 234 nm due to conjugated diene (CD) formation for 5 hours in kinetic mode. The DMSO concentration in the assay is maintained at less than equal to 1.0%. The data is fit to a non-linear regression using Graph pad Prism (version 5.0). The half maximal inhibitory concentration (IC.sub.50) is obtained from the sigmoidal dose response and variable slope curve is generated. Statistical analyses are carried out using a one-way ANOVA followed by a Dunnett's multiple comparison test. Moreover, the data represent five replicates for each condition.

[0114] FIG. 6A illustrates a graph illustrating inhibition of LDL oxidation by the Herbal supplement composition, in accordance with one embodiment of the present invention. As illustrated in the graph, the supplement composition significantly inhibited LDL oxidation caused by Cu.sup.2+ ions in a dose-dependent manner with an EC.sub.50 of 15 μg/mL.

[0115] FIG. 6B illustrates a graph illustrating inhibition of LDL oxidation by Atorvastatin and ascorbic acid, in accordance with one embodiment of the present invention. A significant inhibition of LDL oxidation is also observed with Atorvastatin and ascorbic acid. The error bars in FIG. 6A & FIG. 6B represent Mean±SEM where, *** P<0.001, ** P<0.01, * P<0.05, and one-way ANOVA is followed by Dunnett's test. Table 11 below illustrates data related to LDL oxidation (% activity) as a function of supplement concentration.

TABLE-US-00012 TABLE 11 LDL oxidation (% activity) as a function of Herbal Supplement Composition 3 Supplement, μg/mL % Activity (Mean ± SEM) 0  103 ± 4   6.25 84.5 ± 1.5 12.5   59 ± 3   25 25.5 ± 3.5 50   10 ± 6   100   −1 ± 2  

Experiment 11: To Evaluate ORAC(Oxygen Radical Absorbance Capacity) of Herbal Supplement Composition 3

[0116] The Objective of the experiment is to Evaluate ORAC(Oxygen Radical Absorbance Capacity) of Herbal Supplement composition. Oxygen Radical Absorbance Capacity (ORAC) tests are among the most acknowledged methods that measure antioxidant scavenging activity against oxygen radicals that are known to be involved in the pathogenesis of aging and many common diseases. ORAC 6.0 consists of six types of ORAC assays that evaluate the antioxidant capacity of a material against six primary reactive oxygen species (ROSs, commonly called “oxygen radicals”) found in humans: peroxyl radical, hydroxyl radical, superoxide anion, singlet oxygen, peroxynitrite and hypochlorite. This is a comprehensive panel that evaluates the antioxidant capacity of a material against oxygen radicals

[0117] The ORAC tests are based on evaluating the capacity of an interested material to protect a probe (a fluorescent probe or chromagen) from its damage by ROSs. In all ORAC assays, an ROS inducer is introduced to the assay system. The ROS inducer triggers the release of a specific ROS, which would degrade the probe and cause its emission wavelength or intensity change. When an antioxidant material presents in the environment, the antioxidant absorbs the ROS and preserves the probe from degradation. The degree of probe preservation indicates the antioxidant capacity of the material. Trolox is used as the reference standard, and the results are expressed as □mole Trolox equivalency per gram (or milliliter) of a tested material.

TABLE-US-00013 TABLE 12 ORAC Score (Oxygen Radical Absorbance Capacity) of Herbal Supplement composition 3 against free radicals Radicals Result ORAC against peroxyl radicals 3,930 μmole TE/serving size ORAC against hydroxyl 14,246 μmole TE/serving size radicals ORAC against peroxynitrite 597 μmole TE/serving size ORAC against super oxide 6,243 μmole TE/serving size anion ORAC against singlet oxygen 20,691 μmole TE/serving size ORAC against hypochlorite 4,449 μmole TE/serving size

[0118] There are six predominant reactive species found in the body: peroxyl radicals, hydroxyl radicals, peroxynitrite, super oxide anion, singlet oxygen and hypochlorite. ORAC 6.0 provides a measure of the total antioxidant power of a food/nutrition product against the six predominant reactive species. The ORAC result is expressed as micromole Trolox equivalency (μmole TE) per gram. Serving size=2.17 g

[0119] Therefore, as may be seen, embodiments of the present invention a herbal composition provide a health supplement for total body health needs. The present supplement can be consumed by anyone who has attained the age of 17 years. In-vivo and in-vitro studies conducted on the herbal composition 3 shows efficacy. The present herbal composition supplement has high antioxidant activity and insulin resistance activity through ORAC study and DPPH radical scavenging activity. Therefore, the present composition has been found to be useful towards improving health and enhancing the ability to overcome various disease conditions naturally. Moreover, the present supplement bypasses the need of intake of different dietary supplements to reap the same type of health benefits. The present invention is a broad spectrum antioxidant, antiviral, and antibacterial herbal composition supplement.

[0120] Accordingly, while there has been shown and described the preferred embodiment of the invention is to be appreciated that the invention may be embodied otherwise than is herein specifically shown and described and, within said embodiment, certain changes may be made in the form and arrangement of the parts without departing from the underlying ideas or principles of this invention within the scope of the claims appended herewith.

[0121] In one embodiment, the herbal composition 1 is provided to reduce oxidative stress, comprising therapeutically effective amounts of Moringa oleifera Leaf Powder, Oregano Vulgare Leaf Extract, Shilajit extract, Spirulina, Amla fruit extract, and Piperine extract.

[0122] In one embodiment, the herbal composition 1 includes 40% to 42% by weight of Moringa oleifera Leaf Powder, 21% to 23% by weight of Oregano Vulgare Leaf Extract, 10% to 12% by weight of Shilajit Extract, 10% to 12% by weight of Spiriluna, 12% to 14% by weight of Amla Extract, and 0.5% to 1% by weight of Piperine Extract.

[0123] In one embodiment, the herbal composition 1 further includes about 41.43% by weight of Moringa oleifera Leaf powder.

[0124] In one embodiment, the herbal composition 1 further includes about 22% by weight of Oregano Vulgare Extract powder.

[0125] In one embodiment, the herbal composition 1 further includes about 11% by weight of Shilajit Extract powder.

[0126] In one embodiment, the herbal composition 1 further includes about 11% by weight of Spirulina powder.

[0127] In one embodiment, the herbal composition 1 further includes about 13.81% by weight of Amla fruit Extract powder.

[0128] In one embodiment, the herbal composition 1 further includes about 0.55% by weight of Piperine Extract powder.

[0129] In one embodiment, the herbal composition as discussed herein is administered orally.

[0130] In accordance with an embodiment of the present invention, the herbal composition as discussed herein in the form of a capsule.

[0131] In one embodiment, the herbal composition 1 further includes a high Oxygen Radical Absorbance Capacity (ORAC) value, wherein the ORAC value of a serving of the composition against peroxyl radicals 5,219 μmole TE/2 g.

[0132] In one embodiment, the herbal composition 1 further includes a high Oxygen Radical Absorbance Capacity (ORAC) value, wherein the ORAC value of a serving of the composition against hydroxyl radicals 10,964 μmole TE/2 g.

[0133] In one embodiment, the herbal composition 1 further includes a high Oxygen Radical Absorbance Capacity (ORAC) value, wherein the ORAC value of a serving of the composition against peroxynitrite radicals 543 μmole TE/2 g.

[0134] In one embodiment, the herbal composition 1 further includes a high Oxygen Radical Absorbance Capacity (ORAC) value, wherein the ORAC value of a serving of the composition against super oxide anion radicals 3,924 μmole TE/2 g.

[0135] In one embodiment, the herbal composition 1 further includes a high Oxygen Radical Absorbance Capacity (ORAC) value, wherein the ORAC value of a serving of the composition against singlet oxygen radicals 392 μmole TE/2 g.

[0136] In one embodiment, the herbal composition 1 further includes therapeutically effective amounts of Moringa oleifera Leaf Powder, Oregano Vulgare Leaf Extract, Shilajit extract, Rosemary Leaf Extract, Pomegranate Peel Extract Spirulina, Amla fruit extract, Piperine extract.

[0137] In one embodiment, the herbal composition 1 further includes 27% to 28% by weight of Moringa oleifera Leaf Powder, 13% to 15% by weight of Oregano Vulgare Leaf Extract, 6% to 8% by weight of Rosemary Leaf Extract, 10% to 12% by weight of Pomegranate Peel Extract, 10% to 12% by weight of Shilajit Extract, 10% to 12% by weight of Spiriluna, 13% to 15% by weight of Amla Extract, 0.5% to 1% by weight of Piperine Extract.

[0138] In one embodiment, the herbal composition 1 further includes about 27.6% by weight of Moringa oleifera Leaf powder.

[0139] In one embodiment, the herbal composition 1 further includes about 13.81% by weight of Oregano Vulgare Extract powder.

[0140] In one embodiment, the herbal composition 1 further includes about 11% by weight of Shilajit Extract powder.

[0141] In one embodiment, the herbal composition 1 further includes about 11% by weight of Spirulina powder.

[0142] In one embodiment, the herbal composition 1 further includes about 13.81% by weight of Amla fruit Extract powder.

[0143] In one embodiment, the herbal composition 1 further includes about 11% by weight of Rosemary Leaf Extract powder.

[0144] In one embodiment, the herbal composition 1 further includes about 11% by weight of Pomegranate Peel Extract powder.

[0145] In one embodiment, the herbal composition 1 further includes about 0.52% by weight of Piperine Extract powder.

[0146] In accordance with an embodiment of the present invention, a herbal composition 3 to reduce oxidative stress and Inflammation, includes therapeutically effective amounts of Moringa oleifera Leaf Powder, Oregano Vulgare Leaf Extract, Shilajit extract, Rosemary Leaf Extract, Pomegranate Peel Extract Spirulina, Amla fruit extract, Fenugreek seed extract, Curcuma longa extract, Piperine extract.

[0147] In one embodiment, the herbal composition 3 further includes 26% to 28% by weight of Moringa oleifera Leaf Powder, 6% to 8% by weight of Oregano Vulgare Leaf Extract, 6% to 8% by weight of Rosemary Leaf Extract, 6% to 8% by weight of Pomegranate Peel Extract, 10% to 12% by weight of Shilajit Extract, 10% to 12% by weight of Spiriluna, 10% to 12% by weight of Amla Extract, 15% to 16% by weight of Fenugreek seed Extract, 2% to 3% by weight of Curcuma longa Extract, 0.5% to 1% by weight of Piperine Extract.

[0148] In one embodiment, the herbal composition 3 further includes about 26.17% by weight of Moringa oleifera Leaf powder.

[0149] In one embodiment, the herbal composition 3 further includes about 7.8% by weight of Oregano Vulgare Extract powder.

[0150] In one embodiment, the herbal composition 3 further includes about 10.4% by weight of Shilajit Extract powder.

[0151] In one embodiment, the herbal composition 3 further includes about 10.4% by weight of Spirulina powder.

[0152] In one embodiment, the herbal composition 3 further includes about 10.4% by weight of Amla fruit Extract powder.

[0153] In one embodiment, the herbal composition 3 further includes about 7.8% by weight of Rosemary Leaf Extract powder.

[0154] In one embodiment, the herbal composition 3 further includes about 7.8% by weight of Pomegranate Peel Extract powder.

[0155] In one embodiment, the herbal composition 3 further includes about 15.7% by weight of Fenugreek seed Extract powder.

[0156] In one embodiment, the herbal composition 3 further includes about 2.6% by weight of Curcuma Longa Extract powder.

[0157] In one embodiment, the herbal composition 3 further includes about 0.52% by weight of Piperine Extract powder.