DESC:F O R M 2
THE PATENTS ACT, 1970 (39 of 1970)
&
THE PATENTS RULES, 2003
COMPLETE SPECIFICATION
[See section 10 and rule 13]

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1. TITLE OF THE INVENTION: COMPOSITION AND METHOD
COMPRISING BEETROOT EXTRACT

2. APPLICANT (A) NAME: BIO-GEN EXTRACTS

(B) ADDRESS: BIO-GEN HOUSE, 254/1-9, 3RD
FLOOR, 11TH MAIN ROAD, 3RD PHASE, PEENYA INDUSTRIAL AREA, BANGALORE 560 058, KARNATAKA INDIA

3. NATIONALITY (C) INDIA

THE FOLLOWING SPECIFICATION PARTICULARLY DESCRIBES THE NATURE OF THIS INVENTION AND THE MANNER IN WHICH IT IS TO BE PERFORMED

PRIORITY CLAIM
[001] The instant patent application is related to and claims priority from the co-pending India provisional patent application entitled, “COMPOSITION AND METHOD COMPRISING BEETROOT EXTRACT”, Patent Application no.: 202241042216, Filed on: 23 July 2022, which is incorporated in its entirety herewith.
[002] BACKGROUND OF THE DSCLOSED EMBODIMENTS:
[003] Technical field
[004] The disclosed embodiment is in the technical field of composition and method comprising beetroot extract, more particularly, beetroot extract and anti-caking agent.
[005] Related art
[006] Nitrate (NO3) has received considerable attention in recent years and is quickly gaining traction as a health and performance enhancing nutritional supplement. Once ingested, inorganic NO3 metabolizes in vivo to bioactive nitrite (NO2) and is subsequently salvaged and circulated in human blood.
[007] Nitrate and nitrite play a key role in modulating blood pressure in both healthy and disease states. The vast arrays of surprisingly diverse mechanisms by which NO can be generated from nitrite, serve to underline its vital role in regulating vascular tone.
[008] Its physiological handling appears to be designed to deliver NO to the point of greatest need with evidence that nitrite reduction increases with increasing hypoxia and falling pH-an effect perhaps most evident in certain disease states.
[009] In addition, evidence is emerging that nitrite may have a direct vasoactive effect independent of its role as a precursor for NO. Many authors have reported that NO3- supplementation can improve finish times, elongate time to exhaustion, reduce the gross O2 cost of exercise, and increase peak power and work rate.
[010] Many of these studies have also evaluated cycling exercise performed by well-trained or elite-level athletes to improve due to intake of dietary nitrates.
[011] Beetroot is a rich source of dietary NO3- and a number of studies have investigated its potential for reducing blood pressure in humans, which appears to be more potent in men.
[012] Beetroot is as an exceptionally rich source of antioxidant compounds. The beta lain pigment in particular, has been shown through several in vitro studies to protect cellular components from oxidative injury.
[013] Based on the already available data, beetroot appears to be a powerful dietary source of health promoting agents that holds potential as therapeutic treatment for several pathological disorders.
[014] The powerful antioxidant, anti-inflammatory and vascular-protective effects offered by beetroot and its constituents have been clearly demonstrated by several in vitro and in vivo human and animal studies; hence its increasing popularity as a nutritional approach to help manage cardiovascular disease and cancer.
[015] In the human studies to date, beetroot supplementation has been reported to reduce blood pressure, attenuate inflammation, avert oxidative stress, preserve endothelial function and restore cerebrovascular hemodynamic.
[016] Furthermore, several studies have now established beetroot supplementation as an effective means of enhancing athletic performance.
[017] However, composition comprising beetroot extract with natural high content of polyphenols are challenge to obtain.
[018] Therefore, there is an urgent need to develop an antioxidant, anti-inflammatory and vascular-protective effects composition comprising beetroot extracts with a higher natural polyphenolfor improved exercise performance and cardiovascular benefits.
[019] SUMMARY OF THE DISCLOSURE:
[020] According to an aspect of the disclosed embodiment relates to a composition and method comprising beetroot extract, more particularly, beetroot extract and anti-caking agent.
[021] According to an aspect of the disclosed embodiment, a composition for increasing nitric oxide levels in a subject comprising, (i) a beet root extract in an amount between 89%-100% by weight; and (ii) an anti-caking agent in an amount between 0.1%-10% by weight, wherein the composition comprises 3% to 10% of inorganic nitrate, 1% to 5% of polyphenols, 70 to 75% of carbohydrates, 5 to 10% of dietary fiber, 8 to 12% of protein, and less than0.01% of oxalate.
According to an embodiment,wherein the anti-caking agent comprises Silicon Dioxide, Magnesium stearate, Calcium silicate, Rice hull concentrate, Starch or combinations thereof.
[022] According to another embodiment, wherein the composition comprises about 0.24 g of fat, about 1.0 mg of Cholesterol, about 2104.0 mg of sodium, about 73.3 g of carbohydrate, about 10.92 g of protein, about 2.0 g of vitamin D, about 269.0 mg of calcium, about 14.80 mg of iron, and about 2771.0 mg of Potassium per 100 grams of the composition.
[023] According to further embodiment, wherein the subject is administered with a dosage of 400mg to 600mg of the composition once a day for 6 days for increasing the plasma nitrate [NO3-] of the subject by about 33% to 187% after performing a physical exercise for about two hours.
[024] According to an aspect of the disclosed embodiment, a method for preparing a composition for increasing nitric oxide levels in a subject, a) heating shredded beetroot with water at a temperature between 60 degree Celsius to 80 degree Celsius for about 6-7 days, (ii) reducing the temperature to about 40 degree Celsius and filtering by Gravity filtration method to obtain a filtrate, (iii) distilling the filtrate at a temperature between 70 degree Celsius to 80 degree Celsius to obtain a concentrate, (iv) spraying drying the concentrate; and (v) adding an anti-caking agent to the spray dried concentrate and blending to obtain the composition for increasing nitric oxide levels in the subject, wherein the composition comprises 3% to 10% of inorganic nitrate, 1% to 5% of polyphenols, 70 to 75% of carbohydrates, 5 to 10% of dietary fiber, 8 to 12% of protein, less than 0.01% of oxalate.
[025] According to another embodiment,wherein the anti-caking agent comprises Silicon Dioxide, Magnesium stearate, Calcium silicate, Rice hull concentrate, Starch or combinations thereof.
[026] According to a further embodiment, wherein the composition comprises about 0.24 g of fat, about 1.0 mg of Cholesterol, about 2104.0 mg of sodium, about 73.3 g of carbohydrate, about 10.92 g of protein, about 2.0 g of vitamin D, about 269.0 mg of calcium, about 14.80 mg of iron, and about 2771.0 mg of Potassium per 100 gram of the composition.
[027] According to yet another embodiment, wherein the subject is administered with a dosage of 400mg to 600mg of the composition once a day for 6 days for increasing the plasma nitrate [NO3-] of the subject by about 33% to 187%after performing a physical exercise for about two hours.
[028] According to the embodiment, wherein mean reduction of Peripheral Blood Pressure of the subject comprises from baseline of 1.7% to 2.5%, and mean reduction of Central Blood Pressure of the subject comprises from baseline of 2% to 3.9%, wherein Flow Mediated Dilation (FMD) of the subject comprises an improvement from baseline of 25% to 49%.
[029] According to the embodiment, wherein muscle deoxyhaemoglobin concentration [HHb] during the 60 s end-sprint following 2 hours moderate intensity exercises of the subject is reduced by about 10.95%
[030] According to the embodiment, wherein mean power output during the 60 s end-sprint following 2 hours moderate intensity exercises of the subject is increased by about 6.4%.
[031] Several aspects of the disclosed embodiments are described below with reference to examples for illustration. However, one skilled in the relevant art will recognize that the invention can be practiced without one or more of the specific details or with other methods, components, materials and so forth. In other instances, well-known structures, materials, or operations are not shown in detail to avoid obscuring the features of the invention. Furthermore, the features/aspects described can be practiced in various combinations, though only some of the combinations are described herein for conciseness.
[032] BRIEF DESCRIPTION OF THE DRAWINGS:
[033] Example embodiments of the disclosed embodiments will be described with reference to the accompanying drawings briefly described below.
[034] FIG.1 illustrates thesteps in manufacturing of Beetroot extract which uses water extraction method to arrive at product A, according to the aspects of the disclosed embodiment.
[035] FIG.2A illustrates the flowchart of further manufacturing steps involving addition of Silicon dioxide and processing of product A, to obtain final product, according to the aspects of the disclosed embodiment.
[036] FIG.2B illustrates the flowchart for the composition of the beetroot extract composition, to obtain final product, according to the aspects of the disclosed embodiment
[037] FIG.3 illustrates test products used in the experiment for exercise economy & performance, according to the aspects of the disclosed embodiment.
[038] FIG.4 illustrates plasma [NO3-] and [NO2-] concentration pre- and post-exercise in the active arm and placebo, according to the aspects of the disclosed embodiment.
[039] FIG.5 illustrates mean power during the 60 s end-sprint following 2 hrs, according to the aspects of the disclosed embodiment.
[040] FIG.6 illustrates mean power between 30-60 s of end-sprint following 2 hrs moderate intensity exercises. Dashed lines represent individual responses, according to the aspects of the disclosed embodiment.
[041] FIG. 7 illustrates muscle deoxyhaemoglobin concentration [HHb] during the 60 s end-sprint following 2 h moderate intensity exercises. Data are expressed as the change (?) from baseline, according to the aspects of the disclosed embodiment.
[042] In the drawings, like reference numbers generally indicate identical, functionally similar, and/or structurally similar elements. The drawing in which an element first appears is indicated by the leftmost digit(s) in the corresponding reference number.
[043] DETAILED DESCRIPTION OF THE DISCLOSED EMBODIMENTS
[044] It is to be understood that the present disclosure is not limited in its application to the details of construction and the arrangement of components set forth in the following description or illustrated in the drawings. The present disclosure is capable of other embodiments and of being practiced or of being carried out in various ways. Also, it is to be understood that the phraseology and terminology used herein is for the purpose of description and should not be regarded as limiting.
[045] The use of “including”, “comprising” or “having” and variations thereof herein is meant to encompass the items listed thereafter and equivalents thereof as well as additional items. The terms “a” and “an” herein do not denote a limitation of quantity, but rather denote the presence of at least one of the referenced items. Further, the use of terms “first”, “second”, and “third”, and the like, herein do not denote any order, quantity, or importance, but rather are used to distinguish one element from another.
[046] As used herein, the singular forms “a”, “an”, and “the” include both singular and plural referents unless the context clearly dictates otherwise. By way of example, “a dosage” refers to one or more than one dosage.
[047] The terms “comprising”, “comprises” and “comprised of” as used herein are synonymous with “including”, “includes” or “containing”, “contains”, and are inclusive or open-ended and do not exclude additional, non-recited members, elements or method steps.
[048] All documents cited in the present specification are hereby incorporated by reference in their totality. In particular, the teachings of all documents herein specifically referred to are incorporated by reference.
[049] Example embodiments of the disclosed embodiment are described with reference to the accompanying figures.
[050] In the drawings, like reference numbers generally indicate identical, functionally similar, and/or structurally similar elements. The drawing in which an element first appears is indicated by the leftmost digit(s) in the corresponding reference number.
[051] The spray drying process involves the atomization of a solution, slurry, or emulsion containing one or more components of the desired product into droplets by spraying followed by the rapid evaporation of the sprayed droplets into solid powder by hot air at a certain temperature and pressure.
[052] DEFINITIONS
[053] The term ‘Silicon dioxide’ refers to a natural chemical mix of silicon and oxygen that has uses in many food products as an anticaking agent.
[054] The term ‘Magnesium stearate’ refers to the magnesium salt of the fatty acid, stearic acid. It has been widely used for many decades in the food industry as an emulsifier, binder and thickener, as well as an anticaking, lubricant, release, and antifoaming agent.
[055] The term ‘Calcium silicate (Ca2SiO4)’, also known as calcium silicon oxide, is a product derived from limestone and diatomaceous earth, and is used in the pharmaceutical industry as an anticaking agent.
[056] The term ‘Rice hull’ and rice straw are also used in various industrial applications, such as building materials, particle boards, insulation boards, human food and animal feed, cosmetics, medicines, biopolymers, and fine chemicals.
[057] The term ‘Nitrates’ (NO3-) are one of nitrogen forms. Useful for plant growth, they are of natural or non-natural origin (mineral nitrogen fertilisers). Nitrate is a compound combining the nitrate ion (NO3-) with one or more cations.
[058] The term ‘Plasma nitrate’ refers to the frequently considered a measure of the individual overall exposure to nitrate derived from various individual (patho-) physiological pathways and exogenous (ie, dietary) sources, as well as the individual capacity to eliminate nitrate.
[059] The term ‘Gravity filtration’ refers to a method of filtering impurities from solutions by using gravity to pull liquid through a filter. The two main kinds of filtration used in laboratories are gravity and vacuum/suction.
[060] The term ‘Deoxyhemoglobin’ refers to the form of hemoglobin without oxygen, the predominant protein in red blood cells. Hemoglobin forms an unstable, reversible bond with oxygen. In its oxygen-loaded form it is oxyhemoglobin and is bright red.
[061] The term ‘vastus lateralis’ refers to a muscle located on the lateral, or outside, part of your thigh. The muscle is one of the four quadriceps muscles and is the largest muscle of that group. The function of the vastus lateralis is to work with the other quad muscles to help extend your knee joint.
[062] The term ‘placebo’ refers to any treatment that has no active properties, such as a sugar pill. There are many clinical trials where a person who has taken the placebo instead of the active treatment has reported an improvement in symptoms. Belief in a treatment may be enough to change the course of a person's physical illness.
[063] The term ‘sphygmomanometer’ refers to an instrument used to measure blood pressure which is also known as a blood pressure meter or blood pressure gauge or blood pressure monitor.
[064] 1. EMBODIMENTS OF THE DISCLOSURE
[065] The disclosed embodiment relates to a composition and method comprising beetroot extract, more particularly, beetroot extract and anti-caking agent.
[066] Athletes and active adults are leaning more towards nutrient-dense foods to improve athletic performance. Diets rich in vegetables like beetroots are shown to have a significant impact on body functions during exercise.
[067] Beetroots are rich in inorganic nitrates and when consumed, convert into nitric oxide (NO). Research shows that NO can increase blood flow and improve lung function, thereby enhancing endurance and performance.
[068] 2. METHOD OF THE DISCLOSED EMBODIMENT
[069] The steps in manufacturing of Beetroot extract which uses water extraction method to arrive at product “A” is illustrated in FIG.1.
[070] The beetroot extract was produced through a water extraction process, (102) beginning with thorough cleaning (104), peeling, and shredding (106) of the beetroot.
[071] Subsequently, water was added (108), and the temperature was raised and maintained between 60°C to 80°C (110)for 72hours (112).
[072] The material's temperature was then lowered to 40°C (114), and gravity filtration (116) was employed to remove any residue (118).
[073] After filtration, the filtrate underwent distillation/concentration at temperatures between 70°C to 80°C, (120) resulting in the creation of product A (122).
[074] The further manufacturing steps involves addition of Silicon dioxide and processing of product A, to obtain final productis illustrated in FIG.2A.
[075] Additional manufacturing steps included introducing Silicon dioxide (204) and utilizing spray drying (206) on product “A”(202).
[076] The blend (208) was then packaged (210) to yield the final product, Beetroot Extract composition (212).
[077] This process yielded a neutral-tasting and water-soluble powder.
[078] The Beetroot Extract was standardized to contain no less than 6% inorganic nitrates.
[079] 3. COMPOSITION OF THE DISCLOSED EMBODIMENT
[080] The flowchart for the composition of the beetroot extract composition is illustrated in FIG. 2B.
[081] The beetroot extract composition comprises of:
[082] Comprising 89%-100% beetroot extract and 0.1%-10% anti-caking agent by weight(214).
[083] The beetroot extract composition comprising 3%-10% inorganic nitrate, 1%-5% polyphenols, 70%-75% carbohydrates, 5%-10% dietary fiber, 8%-12% protein, and approximately 0.01% oxalate(216).
[084] Silicon Dioxide - 0.1% to 10%, preferably 2%.
[085] In further embodiment, the composition of beetroot extract is explained.
[086] Inorganic nitrate, commonly found in foods like beetroot juice, has gained attention in sports for its potential to enhance exercise efficiency and endurance by improving blood flow and oxygen delivery to muscles. It can reduce the oxygen cost of exercise, potentially allowing athletes to sustain higher-intensity efforts for longer periods.
[087] Polyphenols, acts as antioxidants to reduce exercise-induced oxidative stress and inflammation, potentially aiding in recovery and reducing muscle soreness.
[088] Carbohydrates are vital for as they serve as the primary source of energy, ensuring endurance and performance during exercise. Adequate carbohydrate intake replenishes glycogen stores in muscles and the liver, delaying fatigue, and supporting quick recovery after workouts. Additionally, carbohydrates play a key role in maintaining stable blood glucose levels, enhancing mental focus, and preserving lean muscle mass, all of which contribute to optimal athletic performance and overall well-being.
[089] Dietary fiber is crucial as it supports digestive health, helps regulate blood sugar levels, and provides sustained energy. It contributes to feelings of fullness, aiding in weight management and promoting cardiovascular health. Additionally, fiber can reduce inflammation and support a healthy gut microbiome, which can enhance recovery and overall well-being for active individuals.
[090] Protein is essential as it supports muscle repair, growth, and exercise performance. It provides the building blocks necessary for muscle recovery and helps athletes maintain and build lean muscle mass. Additionally, protein aids in weight management, supports immune function, and is crucial for injury recovery.
[091] A very low Oxalate content is primarily important forthe composition.
[092] Silicon dioxide is primarily used as an anti-caking agent and may be used to improve the flow of powders. Its function as an additive to maintain product quality and consistency.
[093] POLYPHENOL CONTENTOF COMPOSITION
[094] Table 1 describes polyphenol content of the composition in different batches.
[095] Table. 1: Polyphenol content of the composition
[096]
Batch No TEST PARAMETERS RESULTS

Batch 1 Total Polyphenol content 2.35%
Batch2 Total Polyphenol content 2.42%
Batch 3 Total Polyphenol content 2.47%
Batch 4 Total Polyphenol content 2.39%
Batch 5 Total Polyphenol content 3.26%
Batch 6 Total Polyphenol content 2.70%
Batch 7 Total Polyphenol content 2.83%
Batch 8 Total Polyphenol content 3.15%
[097] NUTRITIONAL DETAILS OF COMPOSITION
[098] Table 2 describes nutritional information of the composition.
[099] Table. 2: The nutritional information of the tested composition
Nutritional information for 100 g (or 100ml)
Energy (kCal) 339.20 kcal/ 100g
Total Fat
Saturated fat
Trans fat
0.24 g/ 100g
0.23 g/ 100g
<0.10 g/ 100g
Cholesterol <1.0 mg/ 100g
Sodium 2104.0 mg/ 100g
Total carbohydrates
Dietary fibers
Total sugars 73.3 g/ 100g
7.10 g/ 100g
20.96 g/ 100g
Protein 10.92 g/ 100g
Vitamin D <2.0 µg/100g
Calcium 269.0 mg/ 100g
Iron 14.80 mg/ 100g
Potassium 2771.0 mg/ 100g
[0100] The composition comprises about 0.24 g of fat, less than 1.0 mg of Cholesterol, about 2104.0 mg of sodium, about 73.3 g of carbohydrate, about 10.92 g of protein, about 2.0 g of vitamin D, about 269.0 mg of calcium, about 14.80 mg of iron, and about 2771.0 mg of Potassium per 100 grams of the composition.
[0101] SYNERGISTIC EFFECT
[0102] Anti-caking:Anti-caking agent helps in free-flowing properties of the beetroot extract composition. Without anti-caking agentthe composition will cake or turn into a hard mass.
[0103] Polyphenols and nitrate
[0104] Dietary polyphenols play a crucial role in the reduction of nitrite to nitric oxide (NO) within the human stomach, providing significance to an anion traditionally considered an end product of NO metabolism. Phenolics have the potential to mediate NO-dependent biological processes through the nitrate-nitrite-NO pathway. This pathway involves intricate chemical interactions occurring under acidic gastric conditions (pO2, pH).
[0105] In the stomach, nitrite undergoes protonation, forming nitrous acid (HNO2). HNO2 can follow various chemical pathways:
[0106] The major pathway involves reduction to NO, especially in the presence of reductants like polyphenols.
[0107] It may yield NO2, serving as a nitrating agent.
[0108] Alternatively, it can combine with NO to produce N2O3, functioning as a nitrosating agent.
[0109] Therefore, presence of polyphenols in food particularly helps in plasma [NO3-] concentration, muscle oxygenation that in turn affect endurance, and exercise performance.
[0110] 4. EXPERIMENTAL EMBODIMENTS
[0111] The disclosed embodiment will be further described in the following examples. It should be understood that these examples are for illustrative purposes only and are not to be construed as limiting this disclosed embodiment in any manner.
[0112] EXAMPLE 1: EFFECTS OF BEETROOT COMPOSITION ON EXERCISE PERFORMANCE
[0113] STUDY OBJECTIVE
[0114] A12-subject,2-arm,randomized,double-blind,cross-over, placebo-controlled study was performed to assess the impact of supplementation of beetroot extracton plasma nitrate [NO3-],plasma nitrite [NO2-], muscle oxygenation andenduranceexerciseperformanceover7daysinhealthyadultsubjects(Vo2 peak49.0±5.1 ml/kg/min).
[0115] The test products used in the experiment for exercise economy & performance is illustrated in FIG. 3.
[0116] EXPERIMENTAL DESIGN
[0117] Thesubjectswereassignedtoreceivebeetroot extractandplacebo in a randomized, counter-balanced order.Between days 1-6 of the supplementation period,subjectsconsumedthetestproducts(400mg to 600mg)mixedwith= 250mlofwater.Onday7,thesubjectsconsumedthetestproducts2hourspriortotheexperimentaltrial.During the experimental trial, the subjects completed2-hour moderate-intensity cycling exercise whichimmediately transitioned into a 60 second end-sprintperformancetest.
[0118] TEST PRODUCTS
[0119] The test products (FIG.3) used were Product “A” (Beetroot extract 8mmol nitrate) and Product “B” (<1 mmol nitrate).
[0120] EXAMPLE. 2: RESULTS OBTAINED
[0121] PLASMA [NO3- ] AND [NO2- ]
[0122] The experiment showed that plasma [NO3-] was higher pre- and post-exercise in the active arm compared to placebo (P<0.001). There was no difference between plasma [NO3-] pre-to-post exercise in the placebo arm (P>0.05). However, post- exercise plasma [NO3-] was higher than pre-exercise plasma [NO3-] in the active arm (P<0.001) [Supplement x time interaction effect].
[0123] Plasma [NO2-] was higher pre- and post- exercise in the active arm compared to placebo (P<0.001). However, there was no supplement x time interaction effect for plasma [NO2-].
[0124] The plasma [NO3-] and [NO2-] concentration pre- and post-exercise in the active arm and placebo is illustrated in FIG. 4.
[0125] ENDURANCE EXERCISE PERFORMANCE
[0126] The experiment showed that time to peak power and peak power were not different between placebo and active arm (P=0.493 & P=0.606). Mean power between 0-60 s was not signi?cantly different (P=0.058).
[0127] The mean power during the 60 s end-sprint following 2 hrs is illustrated in FIG. 5.
[0128] It explains that mean power was signi?cantly greater in the active arm between 30-60 s (P<0.001) and 50-60 s (P<0.05).
[0129] The mean power between 30-60 s of end-sprint following 2 hrs moderate intensity exercises is illustrated in FIG. 6.
[0130] EXAMPLE 3:MUSCLE OXYGENATION
[0131] The experiment explained the muscle oxygenation status. Near-infrared spectroscopy (NIRS) was used to continuously assess changes in the oxygenation status of the M.vastuslateralis of the right leg during exercise. During the end-sprint the muscle deoxyhaemoglobin [HHb] was lower in the active arm compared to the placebo (6.5 ± 1.4 s vs. 7.3 ± 1.4 s; p<0.001). This demonstrates better oxygen extraction to the muscles.
[0132] The muscle deoxyhaemoglobin concentration [HHb] during the 60 s end-sprint following 2 hrs moderate intensity exercises is illustrated in FIG. 7.
[0133] CONCLUSION:
[0134] Vital signs and laboratory data showed that both test products were safe and did not interfere with the normal functioning of the subjects. The clinical study demonstrated signi?cantly higher Plasma [NO3-] and [NO2-] values when compared to the placebo.
[0135] While the peak power & time to peak power were not signi?cantly different between the two arms, the mean peak power during the end sprint between 30-60s and 50-60s was signi?cantly higher in the active arm when compared to the placebo.
[0136] This shows better endurance exercise performance, and is representative of real-world endurance-based events. This may be attributed towards better muscle oxygenation or oxygen extraction in the active arm when compared to the placebo as demonstrated in the study.
[0137] EXAMPLE 4:EFFCTES OF BEETROOT EXTRACT ON NITRIC OXIDE BIOMARKERS AND CARDIOVASCULAR HEALTH
[0138] Cardiovascular disease (CVD) is the leading cause of mortality globally and places a significant burden on healthcare services.
[0139] In an effort to attenuate CVD morbidity cost-effectively, many governments actively promote the consumption of a diet rich in fruit and vegetables based on evidence that adhering to such diets can mitigate the risks associated with CVD morbidity.
[0140] However, emerging evidence indicates that the cardioprotective effect afforded by vegetable consumption outweighs that of fruit consumption and that leafy green vegetables rich in inorganic nitrate (NO3-) might be particularly effective at improving the health of the cardiovascular system.
[0141] The cardiovascular benefits associated with NO3-rich vegetable consumption has been ascribed to the stepwise reduction of NO3- to nitrite (NO2-) and then nitric oxide (NO).
[0142] The gaseous molecule, nitric oxide (NO), regulates an array of physiological processes, but is perhaps best known for its vasodilatory and cardio protective properties.
[0143] It has been demonstrated that NO can be generated through the O2-independent reduction of nitrite (NO2-) to complement O2-dependent NO generation through the NO synthases.
[0144] The circulating plasma [NO2-] can be increased through dietary supplementation with inorganic nitrate (NO3-) and is associated with a reduction in blood pressure and arterial stiffness and improved endothelial function, important predictors of future adverse cardiovascular events.
[0145] In addition, NO3- supplementation can improve vascular function in healthy older adults and some clinical populations including patients with peripheral artery disease and heart failure.
[0146] Increasing dietary NO3- intake, therefore, appears to confer cardioprotective effects and might hold promise as a nutritional intervention to lower the societal and economic burden of cardiovascular diseases.
[0147] There is evidence to suggest that the antihypertensive effect of dietary NO3- supplementation is greater in individuals with a higher baseline blood pressure.
[0148] Therefore, the purpose of the proposed project is to conduct a series of investigations to assess the effect of dietary NO3- supplementation, administered as beetroot extract which contains >6% inorganic nitrate, on cardiovascular function and health in individuals with elevated cardiometabolic disease risk factors, and in healthy individuals exposed to dietary and environmental stimuli that strain the cardiovascular system.
[0149] Only the study on healthy individuals is completed. The study on individuals with cardiometabolic risk is ongoing. Since there was a positive impact on healthy individuals, it is hypothesized that it would be more effective on individuals with cardiometabolic risk.
[0150] Summary of the study on healthy individuals:
[0151] Number of subjects – 18
[0152] Study – Randomized, double-blind, placebo-controlled, cross-over study supplementing placebo, 400mg and 800mg nitrate.
[0153] Endpoints:
[0154] Peripheral Blood Pressure – Mean reduction from baseline of 1.7% to 2.5%
[0155] Central Blood Pressure – Mean reduction from baseline of 2% to 3.9%
[0156] Flow Mediated Dilation (FMD) – Improvement from baseline of 25% to 49%
[0157] Study shows that beetroot extract composition improves cardiovascular function in healthy individuals, and will attenuate the vascular strain in healthy individuals exposed to dietary and environmental stimuli that strain the cardiovascular system.
[0158] EXAMPLE 5: EFFECT OF ACUTE AND SHORT-TERM SUPPLEMENTATION WITH NITRATE-RICH BEETROOT EXTRACT ON VASCULAR FUNCTION IN INDIVIDUALS WITH ELEVATED CARDIOMETABOLIC DISEASE RISK FACTORS
[0159] There is evidence to suggest that some of the vascular benefits attained following dietary NO3- supplementation are greater in individuals with elevated cardiometabolic risk factors, for those who have not yet started taking medicines to manage their condition.
[0160] Therefore, this study will evaluate the potential for a single serving of beetroot extract and short-term supplementation with beetroot extract to improve vascular function and health in individuals with poor cardiometabolic health. This question is important to address given the increasing prevalence and burden of poor cardiometabolic health on healthcare services.
[0161] Therefore, if a cost-effective and natural dietary intervention can lower cardiovascular risk factors in this population, this will be of potential benefit to population health and the strain on healthcare services.
[0162] 5. EXPERIMENTAL DESIGN
[0163] A study to evaluate the effect of acute and short-term supplementation with nitrate-rich beetroot extract on vascular function in individuals with elevated cardiometabolic disease risk factors has been designed where 15 will be participants recruited with elevated cardiometabolic risk factors (overweight, elevated resting blood pressure and elevated fasting blood glucose concentration) to participate in this study.
[0164] Participants will have to report to the laboratory on 5 occasions to complete the study. The initial visit will serve as a preliminary session to confirm the participants meet the entry criteria of having heightened cardiometabolic risk factors.
[0165] Participants will who meet the entry criteria will be asked to return to the laboratory on 4 further occasions to complete the study.
[0166] The study will employ a randomized, double-blind, placebo-controlled, crossover experimental design. On visit 2, participants will consume beetroot extract providing a NO3- dose of 8 mmol or beetroot extract depleted of NO3-.
[0167] Participants then continue to supplement with the beetroot extract composition provided on visit 2 for 5 additional days before returning to the laboratory for visit 3 on the following day to repeat the session completed in visit 2. Visits 2 and 3 will then be repeated on visits 4 and 5 with the opposite beetroot extract supplement.
[0168] MEASURES
[0169] The following measures will be complete prior 2hrs and 3 hrs following the ingestion of the beetroot extract supplement to coincide with peak plasma [NO3-] after NO3- ingestion:
[0170] A venous blood sample will be obtained for nitric oxide and oxidative stress biomarkers.
[0171] Peripheral blood pressure will be assessed using an automated sphygmomanometer.
[0172] Central blood pressure and arterial stiffness markers will be assessed using applanation tonometry.
[0173] Flow mediated dilation will be used to assess endothelial function using flow mediated dilation.
[0174] Heart rate variability will be assessed to evaluate cardiac autonomic function.
[0175] 6. ADVANTAGES OF THE DISCLOSURE EMBODIMENT
[0176] The technology and product developed herein is novel in terms of its application and usage.
[0177] Beetroots are rich in inorganic nitrates and when consumed, convert into nitric oxide (NO). Research shows that NO can increase blood flow and improve lung function, thereby enhancing endurance and performance. Beetroot extract in this contains >6% inorganic nitrates.
[0178] Vital signs and laboratory data showed that test products were safe and did not interfere with the normal functioning of the subjects.
[0179] It is manufactured using a water extraction process, resulting in a neutral-tasting and water-soluble powder, suitable for a variety of end-product applications.
[0180] 7. USES, APPLICATIONS AND BENEFITS OF THE INVENTION
[0181] It is hypothesized that this beetroot extract supplementation will improve cardiovascular function in individuals with elevated cardiometabolic disease risk factors, and will attenuate the vascular strain in healthy individuals exposed to dietary and environmental stimuli that strain the cardiovascular system.
[0182] The beetroot extract will provide improved exercise performance and cardiovascular benefits.
[0183] 8. BEST MODE TO PRACTICE
[0184] Best mode to practicethe disclosure is through commercialization of this invention through the distributors, e-commerce sites, and marketing to reach private gyms/exercise centers, end users and hospitals.
[0185] According to aspects of the disclosed embodiment, the extract may be administered 2 hours prior to performing exercises thereby enhancing endurance and performance.
[0186] It is extremely versatile and easy to use in applications such as stick packs, energy bars, powdered drink mixes & beverages.
[0187] Merely for illustration, only representative number/type of graph, chart, block, and sub-block diagrams were shown. Many environments often contain many more block and sub-block diagrams or systems and sub-systems, both in number and type, depending on the purpose for which the environment is designed.
[0188] While specific embodiments of the invention have been shown and described in detail to illustrate the inventive principles, it will be understood that the invention may be embodied otherwise without departing from such principles.
[0189] Reference throughout this specification to “one embodiment”, “an embodiment”, or similar language means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one embodiment of the disclosed embodiment. Thus, appearances of the phrases “in one embodiment”, “in an embodiment” and similar language throughout this specification may, but do not necessarily, all refer to the same embodiment.
[0190] It should be understood that the figures and/or screen shots illustrated in the attachments highlighting the functionality and advantages of the disclosed embodiment are presented for example purposes only. The disclosed embodiment is sufficiently flexible and configurable, such that it may be utilized in ways other than that shown in the accompanying figures.
[0191] It should be understood that the examples and embodiments described herein are for illustrative purposes only and that various modifications or changes in light thereof will be suggested to persons skilled in the art and are to be included within the spirit and purview of this application and scope of the appended claims. All publications, patents, and patent applications cited herein are hereby incorporated by reference in their entirety for all purposes.

[000] REFERENCE
1) Majeed, Muhammed & Majeed, Shaheen & Lad, Prachi & Vuppala, Kiran. (2016). Standardized Beetroot Extract in Cardiovascular and Exercise Performance. International Journal of Innovative Research in Medical Science. 01. 376-383.

,CLAIMS:I/WE CLAIM:

1. A composition for increasing nitric oxide levels in a subject comprising, (i) a beet root extract in an amount between 89%-100% by weight; and (ii) an anti-caking agent in an amount between 0.1%-10% by weight, wherein the composition comprises 3% to 10% of inorganic nitrate, 1% to 5% of polyphenols, 70% to 75% of carbohydrates, 5% to 10% of dietary fiber, 8% to 12% of protein, and less than0.01% of oxalate.
2. The composition as claimed in claim 1, wherein the anti-caking agent comprises Silicon Dioxide, Magnesium stearate, Calcium silicate, Rice hull concentrate, Starch or combinations thereof.
3. The composition as claimed in claim 1, wherein the composition comprises about 0.24 g of fat, less than1.0 mg of Cholesterol, about 2104.0 mg of sodium, about 73.3 g of carbohydrate, about 10.92 g of protein, about 2.0 g of vitamin D, about 269.0 mg of calcium, about 14.80 mg of iron, and about 2771.0 mg of Potassium per 100 grams of the composition.
4. The composition as claimed in claim 1, wherein the subject is administered with a dosage of 400mg to 600mg of the composition once a day for 6 days for increasing the plasma nitrate [NO3-] of the subject by about 33% to 187% after performing a physical exercise for about two hours.
5. A method for preparing a composition for increasing nitric oxide levels in a subject, a) heating shredded beetroot with water at a temperature between 60 degree Celsius to 80 degree Celsius for about 6-7 days, (ii) reducing the temperature to about 40 degree Celsius and filtering by Gravity filtration method to obtain a filtrate, (iii) distilling the filtrate at a temperature between 70 degree Celsius to 80 degree Celsius to obtain a concentrate, (iv) spraying drying the concentrate; and (v) adding an anti-caking agent to the spray dried concentrate and blending to obtain the composition for increasing nitric oxide levels in the subject, wherein the composition comprises 3% to 10% of inorganic nitrate, 1% to 5% of polyphenols, 70 to 75% of carbohydrates, 5 to 10% of dietary fiber, 8 to 12% of protein, and about 0.01% of oxalate.
6. The method as claimed in claim 5, wherein the anti-caking agent comprises Silicon Dioxide, Magnesium stearate, Calcium silicate, Rice hull concentrate, Starch or combinations thereof.
7. The composition as claimed in claim 5, wherein the composition comprises about 0.24 g of fat, about 1.0 mg of Cholesterol, about 2104.0 mg of sodium, about 73.3 g of carbohydrate, about 10.92 g of protein, about 2.0 g of vitamin D, about 269.0 mg of calcium, about 14.80 mg of iron, and about 2771.0 mg of Potassium per 100 grams of the composition.
8. The method as claimed in claim 5, wherein the subject is administered with a dosage of 400mg to 600mg of the composition once a day for 6 days for increasing the plasma nitrate [NO3-] of the subject by about 33% to 187%after performing a physical exercise for about two hours.
9. The method as claimed in claim 8, wherein mean reduction of Peripheral Blood Pressure of the subject comprises from baseline of 1.7% to 2.5%, and mean reduction of Central Blood Pressure of the subject comprises from baseline of 2% to 3.9%, wherein Flow Mediated Dilation (FMD) of the subject comprises an improvement from baseline of 25% to 49%.
10.The method as claimed in claim 8, wherein muscle deoxyhaemoglobin concentration [HHb] during the 60 s end-sprint following 2 hours moderate intensity exercises of the subject is reduced by about 10.95%.
11. The method as claimed in claim 8, wherein mean power output during the 60 s end-sprint following 2 hours moderate intensity exercises of the subject is increased by about 6.4%.

Dated this 23rd day of September, 2023
Signature.......................................................
(LIPIKA SAHOO)
Registration Number: IN/PA-2467
Agent for Applicant
This document is signed with the digital signature of Patent Agent for the Applicant
LIPIKA SAHOO (IN/PA-2467)