FIELD OF INVENTION

The present invention relates to a synergistic composition comprising Kaempferol and Quercetin for prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders. The present invention also relates to a method of employing the composition for preventing and/or treating obesity, overweight, cardiovascular diseases and other metabolic disorders.

BACKGROUND OF THE INVENTION

Obesity and overweight are devastating diseases. In addition to having harmful impacts on physical health, they can also wreak havoc on the mental health of an individual. Diseases such as obesity and overweight affect self-esteem of the individual, which can ultimately affect a person's ability to have normal social interactions. Unfortunately obesity and overweight are not well understood and societal stereotypes and presumptions regarding the diseases only tend to exacerbate the psychological effects of the diseases. Because of the impact of obesity and overweight on individuals and society, a lot of effort has been expended to find ways for the treatment of the diseases, but little success has been achieved in the long-term treatment and/or prevention. Obesity and overweight can even lead to other diseases such as cardiovascular diseases and other metabolic disorders in the long run.

Obesity and overweight are generally chronic diseases that require long-term behavior modification supplemented with drug therapy. Diet-drug therapies have shown moderate benefits at best. The therapies are however, poorly tolerated by patients (Jones, S. et al. (2012) Long term weight loss withsibutramine. International Journal of Obesity and Related Metabolic Disorders, 9 (Suppl. 2), p.41.). Current weight loss therapies primarily function systemically to alter neurotransmitters for suppression of appetite. While most therapies have been moderately effective, many have systemic side effects.
Glucose and lipid metabolism play an important role in the development of obesity, overweight, cardiovascular diseases and other metabolic disorders. Excessive lipids, or dietary fat uptake has been widely accepted as one of the main causes of obesity and overweight which in turn lead to cardiovascular and other metabolic diseases. Restricted glucose and lipid uptake is considered an effective therapeutic means for treatment of obesity.

Triglycerides, or neutral lipids, are the major form of daily food fat. Triacylglyceride lipases are ubiquitous enzymes required for all aspects of fat metabolism. The lipases are found in the gastrointestinal tract and mediate the digestion of triglycerides and also their uptake by the tissues. Lipases hydrolyze the ester bonds in the triacylglycerides and release free fatty acids and mono-acylglycerols. Dietary triacylglycerides need to be cleaved into free fatty acids and monoacylglycerols for absorption by intestinal enterocytes.

Pancreatic lipase is a key enzyme involved in the digestion of dietary triglycerides. The digestion of triglycerides starts in the upper digestive tract by lipases of preduodenal origin (gastric lipase in humans) and is essentially carried out in the intestine under the action of pancreatic lipase. The latter converts the triglycerides to free fatty acids, 2-monoglycerides and more polar products of hydrolysis, which are capable of crossing the enterocyte brush border membrane, after incorporation into mixed micelles of bile salts and phospholipids. Pancreatic lipase therefore plays a role in the emergence of diseases linked to the presence of an excess amount of lipids, such as cardiovascular diseases, hyperlipemias and obesity, by allowing the assimilation of practically all the triglyercides ingested. In addition, it also promotes the intestinal absorption of cholesterol, since the solubility of cholesterol is increased in the mixed micelles, which are high in fatty acids.

In the absence of lipases, dietary triacylglycerides are not absorbed by bodily tissues and pass into the stool. Consequently, reduced absorption of digested triacylglycerides should result in weight loss. Thus, current research is increasingly directed towards products which induce inhibition of pancreatic lipases.

Many compounds known in the art are known to have pancreatic lipase inhibition activity. Orlistat is one such compound. It is a potent, specific and irreversible inhibitor of pancreatic and gastric lipases. It is also known as tetrahydrolipstatin and is a chemically synthesized derivative of lipstatin, which is naturally produced by Streptomyces toxytricini. It exerts its pharmacologic activity by forming a covalent bond with the active serine site of gastric and pancreatic lipases in the lumen of the gastrointestinal tract. This action of the compound prevents these enzymes from hydrolyzing dietary fat (in the form of triglycerides) into absorbable free fatty acids and monoglycerols. The undigested triglycerides are eliminated by the fecal route. However, lipase inhibition induced by orlistat decreases systemic absorption of dietary fat, thereby contributing to caloric deficit.

Prior art literature reveals that not many studies have been conducted on natural compounds with pancreatic lipase inhibition activity. Studies in literature reveal various health benefits of flavonoids isolated from various plant sources. The potential role of flavonoids in the prevention of cancers, cardiovascular disease and treatment of inflammatory diseases has also been documented. Thousands of naturally occurring flavonoids derived from various plants have been classified according to their chemical structure. The classes are flavones, isoflavones, flavan-3-ols and anthocyanidins. Flavones are divided into four groups: (1) flavones, which includes, e.g., luteolin, apigenin and tangeritin; (2) flavonols, which includes, e.g., quercetin, kaempferol, myricetin, chrysin, rutin, rhoifolin, morin, fisetin, isorhamnetin, pachypodol and rhamnazin; (3) flavanones, which includes, e.g., galangin, hesperetin, naringenin, naringin, neohesperidin, hesperidin, narirutin, pruning, eriodictyol, homoeriodictyol; and (4) 3-hydroxyflavanones or 2,3-dihydroflavonols, which includes, e.g., dihydroquercetin and dihydrokaempferol.
Kaempferol is a natural flavonol, that has been isolated from tea, broccoli, Delphinium, Witch-hazel, grapefruit, cabbage, kale, beans, endive, leek, tomato, strawberries, grapes, brussels sprouts, apples and other plant sources. Kaempferol is a yellow crystalline solid with a melting point of 276°C -278 °C. It is slightly soluble in water but soluble in hot ethanol and diethyl ether. Numerous preclinical studies have shown wide range of pharmacological activities of that kaempferol and some glycosides of kaempferol, including antioxidant, anti-inflammatory, antimicrobial, anticancer, cardioprotective, neuroprotective,
antidiabetic, anti-osteoporotic, estrogenic/antiestrogenic, anxiolytic, analgesic and antiallergic activities. Quercetin, a flavonol, is a plant-derived flavonoid found in fruits, vegetables, leaves and grains. It also may be used as an ingredient in supplements, beverages or foods. Foods rich in quercetin include black and green tea, capers, lovage, apples, onion, especially red onion, red grapes, citrus fruit, tomato, broccoli and other leafy green vegetables, and a number of berries, including raspberry, bog whortleberry, lingonberry, cranberry, chokeberry, sweet rowan, rowanberry, sea buckthorn berry, crowberry and the fruit of the prickly pear cactus.

US2009/0325906 describes methods and compositions for modulation of hyperglycemia and symptoms of hyperglycemia. The composition comprises calcineurin inhibitor capable of inducing hyperglycemia and a blood tissue barrier (BTB) transport protein modulator, wherein the blood tissue barrier (BTB) transport protein modulator is a flavonoid or flavonoid derivative selected from the group consisting of quercetin, isoquercetin, flavon, chrysin, apigenin, rhoifolin, diosmin, galangin, fisetin, morin, rutin, kaempferol, myricetin, taxifolin, naringenin, naringin, hesperetin, hesperidin, chalcone, phloretin, phlorizdin, genistein, biochanin A, catechin, and epicatechin. However the document does not disclose the pancreatic lipase inhibition activity of the composition.

US2007/0087977 describes a pharmaceutical composition comprising an analgesic agent, a blood brain barrier (BBB) transport protein activator and a pharmaceutically acceptable excipient, wherein the analgesic agent is present in an amount sufficient to produce an analgesic effect, wherein the BBB transport protein activator is selected from the group consisting of quercetin, isoquercetin, flavon, chrysin, apigenin, rhoifolin, diosmin, galangin, fisetin, morin, rutin, kaempferol, myricetin, taxifolin, naringenin, naringin, hesperetin, hesperidin, chalcone, phloretin, phlorizdin, genistein, biochanin A, catechin, and epicatechin.

US2009/0130051 describes novel flavonoid compounds having anti-oxidant activity. The compounds and compositions have been shown to exhibit anti-oxidant properties and are particularly useful in the treatment of ischemia and reperfusion injuries. The patent document also describes a method to chemically synthesize such flavonoid compounds and to test their efficacy.

US2008/0306284 describes lipase inhibitors comprising at least one proanthocyanidin derived from teas. The invention also describes medicines, pharmaceutical compositions, foods and beverages comprising the inhibitors and their use for preventing an increase in the levels of blood triglycerides.

US2007/0254948 describes foods, beverages and pharmaceutical compositions comprising lipase inhibitors for regulating the absorption of dietary lipids and suppressing increase of triglycerides in blood. The inhibitors comprise gallotannin or ellagitannin and are isolated form teas or Tellima grandiflora.

US5 503831 describes a composition with lipase inhibiting activity useful in prevention and treatment of obesity. The composition is prepared by extracting defatted rice germ with water and can also be incorporated into food products.

US7939559 describes lipase inhibitors comprising at least one flavan-3-ol monomer derived from teas. The invention also describes pharmaceutical compositions, foods and beverages comprising the inhibitors.

US2008/0299234 describes an orally administrable medication having plant extracts and acting as a lipase inhibitor.

None of the prior art documents disclose pancreatic lipase inhibition activity of flavonoid compounds or a composition comprising flavonoids having synergistic effect for prevention and/or treatment of obesity, overweight and other metabolic disorder.

Thus, there is a need to provide a composition containing natural products, flavonoids in particular, for the treatment of obesity, overweight and other metabolic disorder, wherein the composition has pancreatic lipase inhibition activity.

SUMMARY OF THE INVENTION

An aspect of the present invention relates to a synergistic composition for management and/or treatment of obesity, overweight cardiovascular diseases and other metabolic disorders, wherein the composition comprises Kaempferol and Quercetin, analogs thereof or pharmaceutically acceptable salts thereof in a weight ratio in the range of 4:1 to 1:4.

BRIEF DESCRIPTION OF THE ACCOMPANYING DRAWINGS

The following drawings form part of the present specification and are included to further illustrate aspects of the present invention. The invention may be better understood by reference to the drawings in combination with the detailed description of the specific embodiments presented herein.

Figure 1 shows the performance of natural ingredients at 100 μ M and 500 μ M on pancreatic lipase enzyme.

Figure 2 shows the performance of different concentrations (20 uM, 40 μ M, 60 uM, 80 μ M and 100 μ M) of Kaempferol on pancreatic lipase enzyme.

Figure 3 shows the performance of different concentrations (20 μ M, 40 μ M, 60 μ M, 80 uM and 100 μ M) of Quercetin on pancreatic lipase enzyme.

Figure 4 shows the performance of synergy of Kaempferol and Quercetin in different concentrations on pancreatic lipase enzyme.

DETAILED DESCRIPTION OF THE INVENTION

Those skilled in the art will be aware that the invention described herein is subject to variations and modifications other than those specifically described. It is to be understood that the invention described herein includes all such variations and modifications. The invention also includes all such steps, features, compositions and compounds referred to or indicated in this specification, individually or collectively, and any and all combinations of any two or more of said steps or features.

Definitions

For convenience, before further description of the present invention, certain terms employed in the specification, example and appended claims are collected here. These definitions should be read in the light of the remainder of the disclosure and understood as by a person of skill in the art. The terms used herein have the meanings recognized and known to those of skill in the art, however, for convenience and completeness, particular terms and their meanings are set forth below.

The articles "a", "an" and "the" are used to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article.

The term "plurality" means more than one.

The terms "at least two," "more than one" and "plurality" are used interchangeably.

The terms "comprise" and "comprising" are used in the inclusive, open sense, meaning that additional elements may be included. It is not intended to be construed as "consists of only".

Throughout this specification, unless the context requires otherwise the word "comprise", and variations such as "comprises" and "comprising", will be understood to imply the inclusion of a stated element or step or group of element or steps but not the exclusion of any other element or step or group of element or steps.

The term "including" is used to mean "including but not limited to". "Including" and "including but not limited to" are used interchangeably.

The phrase "pharmaceutically acceptable" refers to molecular entities and compositions that are physiologically tolerable and do not typically produce an allergic or similar untoward reaction, such as gastric upset, dizziness and the like, when administered to a human. Preferably, as used herein, the term "pharmaceutically acceptable" means approved by a regulatory agency of the Federal or a state government or listed in the U.S. Pharmacopeia or other generally recognized pharmacopeia for use in animals, and more particularly in humans.

The term "carrier" refers to a diluent, adjuvant, excipient, or vehicle with which the compound is administered. Such pharmaceutical carriers can be sterile liquids, such as water and oils, including those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil and the like. Water or aqueous solution saline solutions and aqueous dextrose and glycerol solutions are preferably employed as carriers, particularly for injectable solutions.

A "therapeutically effective amount" or "an effective amount", which are used interchangeably, is an amount sufficient to decrease or prevent the symptoms associated with the conditions disclosed herein, including diseases associated with diabetes, obesity, overweight, cardiovascular diseases and other metabolic disorders and other related conditions contemplated for therapy with the compositions of the present invention.

The term "lipase inhibitors" refers to a group of compound that inhibit the metabolic cleavage of dietary triglycerides into free fatty acids and monoglycerides. Under normal physiological conditions, lipolysis occurs via a two-step process that involves acylation of an activated serine moiety of the lipase enzyme. This leads to the production of a fatty acid-lipase hemiacetal intermediate, which is then cleaved to release a diglyceride. Following further deacylation, the lipase-fatty acid intermediate is cleaved, resulting in a free lipase, a monoglyceride and a fatty acid. The resultant free fatty acids and monoglycerides are incorporated into bile acid-phospholipid micelles, which are subsequently absorbed at the level of the brush border of the small intestine. The micelles eventually enter the peripheral circulation as chylomicrons. Accordingly, compounds, including lipase inhibitors that selectively limit or inhibit the absorption of ingested fat precursors are useful in the treatment of conditions including obesity, hyperlipidemia, hyperlipoproteinemia, Syndrome X, and the like.

Pancreatic lipase mediates the metabolic cleavage of fatty acids from triglycerides at the 1- and 3-carbon positions. The primary site of the metabolism of ingested fats by pancreatic lipase is in the duodenum and proximal jejunum. Pancreatic lipase is usually secreted in vast excess of the amounts necessary for the breakdown of fats in the upper small intestine. Since pancreatic lipase is the primary enzyme required for the absorption of dietary triglycerides, inhibitors of the enzyme have found utility in the treatment of obesity and the other related conditions.

The concept of "combination therapy" is well exploited in current medical practice. Treatment of pathology by combination of two or more agents that target the same pathogen or biochemical pathway sometimes results in greater efficacy and diminished side effects relative to the use of the therapeutically relevant dose of each agent alone. In some cases, the efficacy of the drug combination is additive (the efficacy of the combination is approximately equal to the sum of the effects of each drug alone), but in other cases the effect can be synergistic (the efficacy of the combination is greater than the sum of the effects of each drug given alone). As used herein, the term "combination therapy" means the two compounds can be delivered in a simultaneous manner, e.g. concurrently or wherein one of the compounds is administered first, followed by the second agent, e.g. sequentially. The desired result can be either a subjective relief of one or more symptoms or an objectively identifiable improvement in the recipient of the dosage

As used herein the terms "treatment of obesity" and the "management of obesity", are used interchangeably and do not necessarily mean a complete cure. It means that the symptoms or complications of the underlying disease are reduced, and/or that one or more of the underlying cellular, physiological, or biochemical causes or mechanisms causing the symptoms or complications are reduced. It is understood that "reduced", as used in this context, means relative to the untreated state of the disease, including the molecular state of the disease, not just the physiological state of the disease.
The term "treating" refers to the administration of an effective amount of a composition of the present invention to a subject, who has obesity, overweight and other metabolic disorders, or a symptom or a predisposition of such diseases, with the purpose to cure, alleviate, relieve, remedy, or ameliorate such diseases, the symptoms of them, or the predispositions towards them.

The term "administration" covers oral or parenteral delivery to a subject a composition of this invention in any suitable form, e.g., food product, beverage, tablet, and capsule.

The term "parenteral" refers to subcutaneous, intracutaneous, intravenous, intramuscular, intraarticular, intraarterial, intrasynovial, intrasternal, intrathecal, intralesional, and intracranial injection, as well as various infusion techniques.

The term "oral administration" includes oral, buccal, enteral and intra-gastric administration.

The term "Flavonoids" also called bioflavonoids also collectively known as Vitamin P and citrin, are a class of plant secondary metabolites

The term "Synergistic effect/Synergize" refers to a combination of two or more treatments, which is more effective to produce advantageous results than the additive effects of these agents.

The term "synergist" refers to an agent which when present results in a greater-than-additive increase, augmentation or enhancement of the biological or functional effect of the agent. In some cases, it may be difficult to determine which compound in a mixture is of primary importance and which only secondary.

Thus, in a synergistic mixture of compounds, any of the active compounds within the mixture can be considered a synergist.

A composition comprising "synergistic activity" or a "synergistic composition" is a combination of compounds which exhibits increased biological or functional activity as a non-linear multiple of the biological or functional activity of the individual compounds. In other words, the combined biological or functional activity of two or more compounds being tested is significantly greater than the expected result based on independent effects of the compounds when tested separately. Synergism may be apparent only at some ranges or concentrations.

The term "a metabolic disorder" refers to any pathological condition resulting from an alteration in a patient's metabolism. Such disorders include those resulting from an alteration in glucose homeostasis resulting, for example, in hyperglycemia. According to this invention, an alteration in glucose levels is typically an increase in glucose levels by at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or even 100% relative to such levels in a healthy individual. Metabolic disorders include obesity and diabetes (e.g., diabetes type I, diabetes type II, MODY (Maturity Onset Diabetes of the Young) and gestational diabetes).

The term "salts" refers to inorganic and organic salts of compounds. These salts can be prepared in situ during the final isolation and purification of a compound, or by separately reacting a purified compound in its free base form with a suitable organic or inorganic acid and isolating the salt thus formed.

Representative salts include the hydrobromide, hydrochloride, sulfate, bisulfate, nitrate, acetate, oxalate, palmitiate, stearate, laurate, borate, benzoate, lactate, phosphate, tosylate, besylate, esylate, citrate, maleate, fumarate, succinate, tartrate, naphthylate, mesylate, glucoheptonate, lactobionate, and laurylsulphonate salts, and the like. These may include cations based on the alkali and alkaline earth metals, such as sodium, lithium, potassium, calcium, magnesium, and the like, as well as non-toxic ammonium, quaternary ammonium, and amine cations including, but not limited to, ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine, and the like.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the invention, the preferred methods, and materials are now described. All publications mentioned herein are incorporated herein by reference.

It is to be understood that this invention is not limited to particular methods, and experimental conditions described, as such methods and conditions may vary. It is also to be understood that the terminology used herein is for purposes of describing particular embodiments only, and is not intended to be limiting, since the scope of the present invention will be limited only in the appended claims.

It is a primary object of the present invention to provide a synergistic composition for the prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders
It is an object of the present invention to provide a synergistic composition comprising Kaempferol and Quercetin or analogs of Kaempferol and Quercetin and/or pharmaceutically acceptable salts thereof.

The present invention provides a synergistic composition of plant based ingredients which act synergistically at appropriate ratios for prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders.

The present invention provides pharmaceutical compositions useful for prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders comprising an effective amount of Kaempferol and Quercetin or analogs of Kaempferol and Quercetin and a pharmaceutically acceptable carrier.

The Kaempferol and Quercetin or analogs of Kaempferol and Quercetin described herein are present in the compositions in an amount sufficient to produce a desired effect upon obesity, overweight, cardiovascular diseases and other metabolic disorders.

The synergistic composition comprising Kaempferol and Quercetin described herein may be compounded, for example with a pharmaceutically acceptable carrier for solid compositions such as tablets, pellets or capsules, capsules containing liquids, suppositories, solutions, emulsions, suspensions or any other form suitable for use. Suitable carriers include, for example, sterile water, sterile physiological saline, gum acacia, gelatin, starch paste, talc, keratin, colloidal silica, urea and the like. In addition, auxiliary, stabilizing, thickening, lubricating and coloring agents may be used.

Solid dosage forms for oral administration include capsules, tablets, powders, and granules. In such solid dosage forms, the active compound is admixed with at least one inert customary excipient (or carrier) such as sodium citrate or dicalcium phosphate or (a) fillers or extenders, as for example, starches, lactose, sucrose, mannitol, and silicic acid; (b) binders, as for example, carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, and acacia; (c) humectants, as for example, glycerol; (d) disintegrating agents, as for example, agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain complex silicates, and sodium carbonate; (e) solution retarders, as for example, paraffin; (f) absorption accelerators, as for example, quaternary ammonium compounds; (g) wetting agents, as for example, cetyl alcohol and glycerol monostearate; (h) adsorbents, as for example, kaolin and bentonite; and/or (i) lubricants, as for example, talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, or mixtures thereof. In the case of capsules and tablets, the dosage forms may also comprise buffering agents.

Solid dosage forms such as tablets, dragee, capsules, and granules can be prepared with coatings and shells, such as enteric coatings and others well known in the art. They may also contain pacifying agents, and can also be of such composition that they release the active compound or compounds in a delayed manner. Examples of embedding compositions that can be used are polymeric substances and waxes. The active compounds can also be in micro-encapsulated form, if appropriate, with one or more of the above-mentioned excipients.

Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs. In addition to the active compounds, the liquid dosage form may contain inert diluents commonly used in the art, such as water or other solvents, solubilizing agents and emulsifiers, as for example, ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3-butyIene glycol, dimethylformamide, oils, in particular, cottonseed oil, groundnut oil, corn germ oil, olive oil, castor oil, and sesame seed oil, glycerol, tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, or mixtures of these substances, and the like. Besides such inert diluents, the composition can also include adjuvants, such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.

Compositions for oral administration may be in the form of tablets, troches, lozenges, aqueous or oily suspensions, granules or powders, emulsions, capsules, syrups or elixirs. Orally administered compositions may contain one or more agents, such as sweetening agents such as fructose, aspartame or saccharin; flavoring agents such as peppermint, oil of wintergreen, or cherry, coloring agents and preserving agents to provide a pharmaceutically palatable preparation.

Further, compositions in tablet form may be coated to delay disintegration and absorption in the gastrointestinal tract thereby providing a sustained action over an extended period of time. Selectively permeable membranes surrounding an osmotically active driving compound are also suitable orally administered compositions. In these later platforms, fluid from the environment surrounding the capsule is imbibed by the driving compound, which swells to displace the agent or agent composition through an aperture.

These compositions may also contain adjuvants such as preserving, wetting, emulsifying, and dispersing agents. Prevention of microorganism contamination of the compositions can be ensured by various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, and the like. It may also be desirable to include isotonic agents, for example, sugars, sodium chloride, and the like. Prolonged absorption of injectable pharmaceutical compositions can be brought about by the use of agents capable of delaying absorption, for example, aluminum monostearate and gelatin.

The additional anti-obesity agents are preferably selected from but not limited to the group consisting of a p3-adrenergic receptor agonist, a cholecystokinin-A agonist, a monoamine reuptake inhibitor, a sympathomimetic agent, a serotoninergic agent, a dopamine agonist, a melanocyte-stimulating hormone receptor agonist or mimetic, a melanocyte-stimulating hormone receptor analog, a cannabinoid receptor antagonist, a melanin concentrating hormone antagonist, leptin, a leptin analog, a leptin receptor agonist, a galanin antagonist, a lipase inhibitor, a bombesin agonist, a neuropeptide-Y antagonist such as NPY-1 or NPY-5, a thyromimetic agent, dehydroepiandrosterone or an analog thereof, a glucocorticoid receptor agonist or antagonist, an orexin receptor antagonist, a urocortin binding protein antagonist, a glucagon-like peptide-1 receptor agonist, and a ciliary neurotrophic factor.

The synergistic composition of the present invention described herein can be used in the form of a food additive, food supplement, dietary supplement for example, in solid, semisolid or liquid form, which contains comprising Kaempferol and Quercetin or analogs of Kaempferol and Quercetin.

The synergistic composition of the present invention may be incorporated into food product alone or in combination with another anti-obesity, anti-hyperglycemic (blood glucose lowering), or anti-lipidemic
compound, in admixture with a carrier or an excipient suitable for oral administration.

Compositions for oral administration may be in the form of food product comprising the compositions of this invention.

Any conventional food processing technique may be used to achieve a product comprising the effective amount of the composition described herein. There is much information on the art and technology of the various conventional food processing techniques and their practices in both the pet food and food industries, and it is accordingly assumed that the general principals of these techniques are understood by the person skilled in the art.

The compounds identified by the present invention and other pharmaceutically active compounds, if desired, can be administered to a patient either orally, rectally, parenterally, (for example, intravenously, intramuscularly, or subcutaneously) intracisternally, intravaginally, intraperitoneally, intravesically, locally (for example, powders, ointments or drops), or as a buccal or nasal spray. They are advantageously effective when administered orally.

Compositions suitable for parenteral injection may comprise physiologically acceptable sterile aqueous or nonaqueous solutions, dispersions, suspensions, or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Examples of suitable aqueous and nonaqueous carriers, diluents, solvents, or vehicles include water, ethanol, polyols (propylene glycol, polyethylene glycol, glycerol, and the like), suitable mixtures thereof, triglycerides, including vegetable oils such as olive oil, and injectable organic esters such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants.

The present invention provides methods and compositions comprising Kaempferol and Quercetin, analogs of Kaempferol and Quercetin or a pharmaceutically acceptable salt thereof for modification and regulation of lipid metabolism to reduce obesity, overweight, cardiovascular disease and/or for the improvement of metabolism disorders, especially those associated with obesity.

The subject method of the present invention includes the administration, to an animal, of a synergistic composition comprising two pancreatic lipase inhibitors in a specific ratio, which are able to inhibit the activity of pancreatic lipase thereby reducing breakdown of fats.

The present invention relates to two specific natural ingredients namely Kaempferol and Quercetin. Kaempferol and Quercetin belong to class of flavonoids. The present invention provides a novel synergistic combination of two natural products of plant origin, such as Kaempferol and Quercetin or their analogs in a ratio in the range of 1:99 to 99:1 preferably, 1:4 for inhibition of pancreatic lipase enzyme.
The synergistic composition of the present invention comprises a unique combination of natural ingredients of plant origin which facilitate enhanced inhibition of pancreatic lipase. The present invention also provides an optimized synergistic combination of these natural ingredients at a desired ratio and molar concentration to inhibit pancreatic lipase enzyme for prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders.

The synergistic composition of the present invention having Kaempferol and Quercetin in the weight ratio of 1:4 shows enhanced inhibition activity of pancreatic lipase.

The synergistic composition of the present invention may comprise additional anti-obesity agents preferably selected from but not limited to the group consisting of a p> adrenergic receptor agonist, a cholecystokinin-A agonist, a monoamine reuptake inhibitor, a sympathomimetic agent, a serotoninergic agent, a dopamine agonist, a melanocyte-stimulating hormone receptor agonist or mimetic, a melanocyte-stimulating hormone receptor analog, a cannabinoid receptor antagonist, a melanin concentrating hormone antagonist, leptin, a leptin analog, a leptin receptor agonist, a galanin antagonist, a lipase inhibitor, a bombesin agonist, a neuropeptide-Y antagonist such as NPY-1 or NPY-5, a thyromimetic agent, dehydroepiandrosterone or an analog thereof, a glucocorticoid receptor agonist or antagonist, an orexin receptor antagonist, a urocortin binding protein antagonist, a glucagon-like peptide-1 receptor agonist, and a ciliary neurotrophic factor.

The present invention demonstrates an enhanced pancreatic lipase inhibition using a synergistic composition comprising Kaempferol and Quercetin in the weight ratio of 1:4. It is a surprise finding that at a given ratio, the effect is synergistic for its inhibition.

This work will have significance, especially to develop wellness and healthy indulgence food products for the management of obesity, overweight, cardiovascular diseases and other metabolic disorders.
The present invention provides a synergistic composition for the prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders comprising Kaempferol and Quercetin, analogs thereof and/or pharmaceutically acceptable salts thereof, in a weight ratio of Kaempferol and Quercetin in the range of 4:1 and 1:4.

In another embodiment of the present invention there is provided a synergistic composition for the prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders comprising Kaempferol and Quercetin, analogs thereof and/or pharmaceutically acceptable salts thereof, in a weight ratio of Kaempferol and Quercetin 1:4.

In yet another embodiment of the present invention, there is provided a synergistic composition for the prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders comprising Kaempferol and Quercetin, analogs thereof and/or pharmaceutically acceptable salts thereof, in a weight ratio of Kaempferol and Quercetin between 4:1 and 1:4, wherein the analogs are natural analogs or synthetic analogs.

In an embodiment of the present invention, there is provided a synergistic composition for the prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders comprising Kaempferol and Quercetin, analogs thereof and/or pharmaceutically acceptable salts thereof, in a weight ratio of Kaempferol and Quercetin between 4:1 and 1:4, wherein the composition comprises a pharmaceutically acceptable additive or a carrier.

In still another embodiment of the present invention, there is provided a synergistic composition for the prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders comprising Kaempferol and Quercetin, analogs thereof and/or pharmaceutically acceptable salts thereof, in a weight ratio of Kaempferol and Quercetin between 4:1 and 1:4, wherein the composition optionally comprises one or more other therapeutic agents selected from a group consisting of pancreatic lipase inhibitor, calcineurin inhibitor, blood tissue barrier (BTB) transport protein modulator, a-glucosidase inhibitor, insulin, Ml receptor antagonists, prolactin inhibitors, agents acting on an ATP-dependent channel of P-cells, metformin, an intestinal glucose transporter inhibitor, a glycation inhibitor, a nitric oxide production inhibitor, an aldose reductase inhibitor, a PPAR agonist, an adipocytokine activator, a glucose uptake enhancer, a dipeptidyl peptidase-IV inhibitor, and a thermogenesis enhancer.

In still another embodiment of the present invention, there is provided a synergistic composition for the prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders comprising Kaempferol and Quercetin, analogs thereof and/or pharmaceutically acceptable salts thereof, in a weight ratio of Kaempferol and Quercetin between 4:1 and 1:4, wherein the composition is in a form selected from the group consisting of an immediate-release composition, a controlled-release composition, sustained-release orally-administrable compositions, topically-administrable compositions, liquid solutions, liquid sprays, lozenges, throat sprays, ointments, solutions, foams, cough drops, dissolvable strips, a jelly, a mouthwash; a gargle, a lollipop, a gum, aqueous or oily suspensions, dispersible powders or granules, a syrup, an elixir, emulsions, a cream, a paste, a gel, a lotion, impregnated dressings, occularly-administrable compositions, inhalable particles, inhalable solutions, droplets, and aerosols.

The present invention further provides a process for the preparation of the synergistic composition, wherein the process comprising mixing Kaempferol and Quercetin or analogs thereof in DMSO in a weight ratio of Kaempferol and Quercetin in the range between 4:1 and 1:4; and optionally adding a pharmaceutically acceptable carrier to obtain the composition.

An embodiment of the present invention provides a food, beverage or feed supplement comprising the synergistic composition of the present invention, wherein the synergistic composition comprises Kaempferol and Quercetin, analogs thereof and/or pharmaceutically acceptable salts thereof, in a weight ratio of Kaempferol and Quercetin between 4:1 and 1:4.

Another embodiment of the present invention provides a food, beverage or feed product comprising the synergistic composition of the present invention, wherein the synergistic composition comprises Kaempferol and Quercetin, analogs thereof and/or pharmaceutically acceptable salts thereof, in a weight ratio of Kaempferol and Quercetin between 4:1 and 1:4.

In yet another embodiment of the present invention, there is provided a method of management and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders, wherein the method comprises administering to a subject in need thereof a therapeutically effective amount of a synergistic composition comprising Kaempferol and Quercetin, analogs thereof or pharmaceutically acceptable salts thereof, in a weight ratio in the range of 4:1 to 1:4; wherein the effective amount is sufficient to treat obesity and overweight but not sufficient to suppress the immune system of the subject.

In yet another embodiment of the present invention, there is provided a method of management and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders, wherein the method comprises administering to a subject in need thereof a therapeutically effective amount of a synergistic composition comprising Kaempferol and Quercetin, analogs thereof or pharmaceutically acceptable salts thereof, in a weight ratio in the range of 4:1 to 1:4; wherein the synergistic composition is administered orally.

The composition of the present invention may be compounded with foods such as, but not limited to dairy products, grains, breads, meats, fruits, vegetables, rice and the like. The composition of the present invention can be compounded with additional ingredients. For example, the composition can be mixed with carbohydrates lipids, polypeptides, fatty acids, phytochemicals, and combinations thereof.

Industrial Applicability
The present invention provides a useful synergistic composition for prevention and/or treatment and management of patients suffering from obesity, overweight, cardiovascular diseases and other metabolic disorders. The synergistic composition of the present invention for prevention and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders comprising Kaempferol and Quercetin or their analogs as the main components is highly safe, useful and can be administered easily and over a long term.

Further, the composition of the present invention can be used for development of anti-obesity food, feed, and nutraceutical composition and for development of satiety enhancement foods.

EXAMPLES
It should be understood that the following examples described herein are for illustrative purposes only and that various modifications or changes in light of the specification will be suggestive to person skilled in the art and are to be included within the spirit and purview of this application and the scope of the appended claims.

Example 1
Screening of Natural ingredients as pancreatic lipase Inhibitors

The natural ingredients belonging to the group: Kaempferol, Aloin, Mangiferin, Capsaicin, Quercetin and Caffeine were selected for the pancreatic lipase inhibition assay. The assay is based on the action of pancreatic lipase enzyme which catalyses hydrolytic cleavage of triglycerides into free fatty acids and monoglyceride. The substrate used in the assay is para-nitrophenol tagged laurate molecule. The enzyme acts on the substrate and p-nitrophenol is released, the absorbance of which is measured colorimetrically at 410 nm. The pancreatic lipase inhibition assay was carried out in a total reaction volume of 200 μ L in 96-well microwell plate comprising assay buffer, emulsifier, substrate, inhibitor, and enzyme. 0.1M Tris-HCl buffer at pH 7.5 was used as the assay buffer as well as the solvent for enzyme and emulsifier solutions. The substrate was prepared in isopropanol. The assay components were added in their respective volumes and the reaction was started by adding the enzyme. The reaction was carried out at 37°C for one hour with the absorbance readings being measured every one minute. Each assay was performed (in duplicates) against its ingredient blank comprising assay buffer, emulsifier, substrate and inhibitor or ingredient (at lOOuM and 500|iM concentration).

Enzyme preparation was carried out using human pancreatic lipase procured from Sigma. The enzyme was diluted to 200uL with assay buffer to obtain a final concentration of 50U/200uT. The enzyme prepared was stored at 4°C till use.

Orlistat was used at 5 uM as a positive control in the pancreatic liapse inhibition assay. Percentage inhibition was calculated as, % Inhibition = (Absorbance control -Absorbance Test)/ Absorbance control x 100. Activity of natural ingredient of plant origin was assayed (Figure 1-4).

It was found that out of all the natural ingredients used for the assay, Kaempferol and Quercetin were able to inhibit above 50% pancreatic lipase enzyme at a concentration of 100 uM with respect to Orlistat (a known inhibitor of pancreatic lipase enzyme) which was taken as positive control. Figure 1 illustrates the performance of natural ingredients at 100 μ M and 500 μ M on pancreatic lipase enzyme.

Based on the results obtained, pancreatic lipase inhibition assay was performed using Kaempferol and Quercetin at a concentration in the range from 20 uM to 100 uM (Figure 2-3). Figure 2 illustrates the performance of Kaempferol at different concentrations (20 uM, 40 uM, 60 |j.M, 80 uM and 100 uM) on pancreatic lipase. Figure 3 illustrates the performance of different concentrations of Quercetin (20 uM, 40 \iM, 60 uM, 80 uM and 100 uM) on pancreatic lipase.

It was inferred from Figure 2 that maximum % inhibition of pancreatic lipase enzyme was obtained with 100 μ M concentration of Kaempferol with respect to Orlistat which was taken as positive control. The % inhibition at 100 μ M was calculated to be 52%.

It was inferred from Figure 3 that maximum % inhibition of pancreatic lipase enzyme was obtained with 100 μ M concentration of Quercetin with respect to Orlistat which was taken as positive control. The % inhibition at 100 μ M was calculated to be 63%.

Example 2
Synergy of Kaempferol and Quercetin for enhanced pancreatic lipase inhibition

Based on the results obtained, Kaempferol and Quercetin were selected for synergy studies and pancreatic lipase inhibition assay was performed. The amount of Kaempferol and Quercetin were taken as provided in Table 1.

Table 1: Amount of Kaempferol and Quercetin taken for pancreatic lipase inhibition assay

The above test ingredient ratios were incorporated in the assay well and pancreatic lipase assay was performed as described earlier. Figure 4 illustrates the performance of synergy of Kaempferol and Quercetin at different concentrations on pancreatic lipase. It was inferred from Figure 4 that maximum % inhibition of pancreatic lipase with respect to Orlistat was obtained in composition comprising 20 μ M Kaempferol and 80 μ M Quercetin (weight ratio of 20:80). The % inhibition was calculated to be 82%.
Thus, it was demonstrated that Kaempferol and Quercetin in a particular combination in the weight ratio of 20:80 (1:4) ratio shows enhanced inhibitory activity of pancreatic lipase as compared to Kaempferol and Quercetin alone. These results suggest that the synergistic effects of Kaempferol and Quercetin in inhibiting pancreatic lipase activity can be used for development of pharmaceutical compositions for the treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders, for development of food and feed products for patients suffering from such disease and for development of satiety enhancement foods and nutraceutical composition.

I/We Claim:

1. A synergistic composition for management and/or treatment of obesity, overweight cardiovascular diseases and other metabolic disorders, wherein the composition comprises Kaempferol and Quercetin, analogs thereof or pharmaceutically acceptable salts thereof, in a weight ratio in the range of 4:1 to 1:4.

2. The synergistic composition as claimed in claim 1, wherein weight ratio of Kaempferol to Quercetin is 1:4.

3. The synergistic composition as claimed in claim 1, wherein the analogs are natural analogs or synthetic analogs.

4. The synergistic composition as claimed in claim 1, wherein the composition optionally comprises pharmaceutically acceptable additives or a carrier.

5. The synergistic composition as claimed in claim 1, wherein the composition optionally comprises one or more therapeutic agents selected from the group consisting of calcineurin inhibitor, blood tissue barrier (BTB) transport protein modulator, a-glucosidase inhibitor, insulin, Ml receptor antagonists, prolactin inhibitors, agents acting on an ATP-dependent channel of p-cells, metformin, an intestinal glucose transporter inhibitor, a glycation inhibitor, a nitric oxide production inhibitor, an aldose reductase inhibitor, a PPAR agonist, an adipocytokine activator, a glucose uptake enhancer, a dipeptidyl peptidase-IV inhibitor, and a thermogenesis enhancer.

6. The synergistic composition as claimed in claim 1, wherein the composition is in a form selected from the group consisting of an immediate-release composition, a controlled-release composition, sustained-release orally-administrable compositions, topically-administrable compositions, liquid solutions, liquid sprays, lozenges, throat sprays, ointments, solutions, foams, cough drops, dissolvable strips, a jelly, a mouthwash, a gargle, a lollipop, a gum, aqueous or oily suspensions, dispersible powders or granules, a syrup, an elixir, emulsions, a cream, a paste, a gel, a lotion, impregnated dressings, occularly-administrable compositions, inhalable particles, inhalable solutions, droplets, and aerosols.

7. A food, beverage or feed supplement comprising the composition as claimed in claim 1.

8. A food, beverage or feed product comprising the composition as claimed in claim 1.

9. A method of management and/or treatment of obesity, overweight, cardiovascular diseases and other metabolic disorders, wherein the method comprises administering to a subject in need thereof a therapeutically effective amount of the synergistic composition as claimed in claim 1, wherein the effective amount is sufficient to treat obesity and overweight but not sufficient to suppress the immune system of the subject.

10. The method as claimed in claim 9, wherein the synergistic composition is administered orally.