Title of Invention	A PHARAMACEUTICAL FORMULATION CONTAINING QUERCETIN FOT THE TREATMENT OF DIABETIC NEPHROPATHY
Abstract	Diabetic nephropathy is one of the complications of diabetes mellitus and is known for kidney complications. During diabetic nephropathy, the functional unit of kidney is damaged leading to increased filtration rate. Diet plays a major role in the management of diabetes and its associated complications. In this aspect, dietary antioxidants are playing an important role. Quercetin is a well-documented bioflavonoid occurring in many foods and is known to be present in higher concentrations in onions, apples, broccoli, green tea and red wine. A formulation involving quercetin was examined for its efficacy on diabetic nephropathy state. The experimental groups showed considerable improvement in ameliorating the diabetic status as assessed by urine sugar, urine volume and fasting blood sugar. The kidney damage was estimated by measuring the glomerular filtration rate in diabetic and treated rats. The quercetin fed diabetic rats showed about 50% improvement in the diabetic nephropathy state.

Title of Invention

A PHARAMACEUTICAL FORMULATION CONTAINING QUERCETIN FOT THE TREATMENT OF DIABETIC NEPHROPATHY

Abstract

Diabetic nephropathy is one of the complications of diabetes mellitus and is known for kidney complications. During diabetic nephropathy, the functional unit of kidney is damaged leading to increased filtration rate. Diet plays a major role in the management of diabetes and its associated complications. In this aspect, dietary antioxidants are playing an important role. Quercetin is a well-documented bioflavonoid occurring in many foods and is known to be present in higher concentrations in onions, apples, broccoli, green tea and red wine. A formulation involving quercetin was examined for its efficacy on diabetic nephropathy state. The experimental groups showed considerable improvement in ameliorating the diabetic status as assessed by urine sugar, urine volume and fasting blood sugar. The kidney damage was estimated by measuring the glomerular filtration rate in diabetic and treated rats. The quercetin fed diabetic rats showed about 50% improvement in the diabetic nephropathy state.

Full Text	Field of the invention: The present invention relates to a pharmaceutical composition useful for treating diabetic nephropathy. Background and prior art of the invention: Diabetic nephropathy is one of the complications of diabetes mellitus involving glomerular basement membrane thickening due to decrease in heparan sulphate and laminin and increase in type-IV collagen. Diabetic nephropathy is characterized by increased excretion of urine with micro and macromolecules (viz. proteins, glucose etc), as a result of glomerular basement membrane. During diabetic nephropathy, the functional unit of kidney, the nephrons are damaged leading to increased filtration rate. Diet plays a major role in the management of diabetes and its associated complications. In this aspect, role of many traditional practices involving medicinal plants are known to have preventive and therapeutic value in diabetes management. The bioactive principles of medicinal plants and foods of medicinal value are receiving much scientific attention in recent years. In this direction anti-oxidants are of considerable interest. Higher oxidative stress is one of the factors implicated in the development and progression of diabetic complications resulting in increased levels of free radicals or impaired anti-oxidant defense mechanisms. Epidemiological evidence has suggested that anti-oxidant rich dietary flavanoids have potential health beneficial effects. Quercetin is a well-documented bioflavonoid occurring in many foods and is known to be present in higher concentrations in onions, apples, broccoli, green tea and red wine. Hence, the polyphenolic compounds constitute an integral part of human diet though they are considered as non-nutrients. Many studies have focused towards the beneficial properties of quercetin, namely, anti-proliferative, anti-bacterial, anti-oxidant, anti-inflammatory and anti-carcinogenic. However, the anti-diabetic property of querectin is yet not reported anywhere nor studied. A formulation involving quercetin could be expected to have a significant role in combating glomerular filtration rate and progression of diabetic nephropathy Reference may be made to US patent US 6,291,533 (2001) wherein quercetin dihydrate was used as a supplement in the compositions for treating people with a specific blood type. Drawback of this composition is that it is designed for people with specific antigenic blood types and have not specified for diabetes and its associated problems. Reference may be made to US patent 20030108624 (2003) wherein Gymnema sylvestre; Fenugreek seed and Ginkgo biloba are used for prevention and treatment of chronic diseases and disorders including the complications of diabetes mellitus. Drawback of this is it fails to explain the specific role of quercetin in diabetic nephropathy. Reference may be made to US patent 20020025347 (2002) wherein aqueous extract of Salviae Miltiorrhizae Radix herb is used for the treatment of diabetic nephropathy and microalbuminuria. Drawback of this invention is the composition of the herb and antioxidant activity in treatment of diabetic nephropathy is not clear. Reference may be made to the US patent 20020192314 (2002) wherein dietary supplements of grape skin and grape seed extract can inhibit platelet aggregation or LDL cholesterol oxidation in a mammal. Drawback of this invention is that it deals with variety of polyphenolic components in prevention of oxidation but it does not specify for diabetes and its complications in particular diabetic nephropathy. Reference may be made to US patent 6572897 (2003) wherein formulation contains essential amounts of alpha lipoic acid, chromium, lutein, bioflavonoids(quercetin and rutin), mormordica charantia extract, corosolic acid, and gymnema sylvestre extract used for insulin sensitivity and blood sugar level maintenance for the prevention and treatment of diabetes. The drawback of this formulation is it does not explain the effect of this in diabetic nephropathy state and it is not a single component used for the treatment. Reference may be made to the Japanese patent 2002080362 (2002) wherein gallic ester, galloyl tannin, quercetin, catechins and epicatechins are used in regulating PPAR-(Peroxisome Proliferator Activated Receptor)-dependent gene transcription activity; an agent for obesity, diabetes, hyperglycemia, hyperlipemia and insulin resistance. Drawback of this invention is it does not explain the activity of diabetic nephropathy complications. Reference may be made to the patent WO0059522 (2000) wherein extracted products from Brickellia californica and its isolated flavonoids, including apigenin, luteolin, quercetin and dihydroxykaemferol results in lowering of blood sugar and are thereby used in the treatment of both insulin dependent and non-insulin dependent diabetes. The drawback of this is that it is an herbal extract and it does not explain the role of quercetin in diabetes or its complications. Reference may be made to the Japanese patent 3232851 (1991) wherein phenolic compounds extracted from various plants like tannic acid, pentagalloyl glucose, rutin, quercetin, epicatechin are used as therapeutic agent of various complications in diabetes mellitus such as cataract, retinopathy, kidney disease or nerve disturbance by inhibiting aldose reductase. The drawback of this invention is that it does not clearly explain the role of these compounds in glomerular filtration rate during diabetic nephropathy state. Reference may be made to US patent 2004258674 (2004) wherein nutritional supplement of quercetin to treat hypertenstion, therby preventing cardiovascular disease is disclosed. The drawback of this treatment is that it does not explain anything about diabetes and its related complications. Reference may be made to patent WO2004037015 (2004) wherein a method of treating Crohn's disease, arthritis, leukemia, lowering cholesterol levels or blood pressure using antioxidant compositions of various plant products soy isoflavones, taurine, sugar beet pectin fiber, or a ginko biloba extract are described. The drawback is that it includes variety of antioxidants and does not treat diabetes and its associated complications. Reference may be made to US patent 2004014686 (2004) wherein a combination of catechin and quercetin possessing a potential antioxidant activity is used as pharmaceutical and dietary formulation. The drawback of this is that a combination of antioxidants is used and individual effects on the disease condition particularly in diabetic nephropathy are not explained. Reference may be made to Korean patent 2003015815 (2003) wherein quercetin is used as an inhibitor of gene expression of vascular endothelial growth factor (vegf) and erythropoietin (epo). The drawback of this invention is that it fails to explain the role of quercetin in diabetic nephropathy. Novelty: Novelty of this invention is that quercetin (0.1%) helps to minimize the complications of kidney damage that take place during diabetic nephropathy state. Objects of the invention: The main objective of the present invention is to provide a process for the preparation of pharmaceutical formulation containing quercetin for treating diabetic nephropathy. Another objective of the invention is to provide a pharmaceutical formulation containing quercetin for treating diabetic nephropathy. Summary of the invention: This invention relates to a pharmaceutical formulation, useful for treating diabetic nephropathy, comprising of mixing a number of different ingredients at a temperature of 24-27°C for a period of 3 to 45 minutes, to obtain the desired formulation. In the final formulation, the carbohydrate content ranges from 65 to 75 percent, proteins range from 15 to 20 percent and fat content is in the range of 5 to 15 percent. The said formulation is useful for controlling diabetic nephropathy and is administered by oral feeding to mammals. Accordingly, the present invention provides a pharmaceutical composition useful for treating diabetic nephropathy comprising of the following ingredients: Ingredients % by weight in (g) [a] Corn starch 65-70 [b] Casein 15-20 [c] Vegetable Fat 05-15 [d] Mineral mix 02-05 [e] Vitamin Mix 01-05 [f] Quercetin 0.005 - 0.02 Composition of component [d] mineral (g/kg mixture: mixture) Calcium phosphate (dibasic) 500 Sodium chloride 74 Potassium citrate monohydrate 220 Potassium sulphate 52 Magnesium oxide 24 Manganous carbonate 3.5: Ferric citrate 6 Zinc carbonate 1.6 Cupric carbonate 0.3 Potassium iodate 0.01 Sodium selenite 0.01 Chromium potassium sulphate 0.55 Corn starch was added to make the total amount 1000g Composition of component [e] vitamin (g/kg mix) mixture Thiamin hydrochloride 0.6 Riboflavin 0.6 Pyridoxine hydrochloride 0.7 Nicotinic acid 3 D-calcium pantothenate 1.6 Folic acid 0.2 D-biotin 0.02 Cyanocobalamine (vit B-12) 0.001 Retinyl acetate (400000 I. U) 0.4 DL- DD tocopherols acetate (5000 I.U) 7.7 Menadione 0.005 Corn starch was added to make the total amount 1000g Detailed description of the invention: The present invention relates to a design of a pharmaceutical formulation useful for diabetic nephropathy comprising of antioxidant - quercetin. The said pharmaceutical formulation is useful for diabetic nephropathy and is comprising of the following ingredients in the given range of percent, by weight in gram: Ingredients % by weight in (g) Corn starch 65-70 Casein 15-20 Vegetable Fat 05-15 Mineral mix 02-05 Vitamin Mix 01 -05 Quercetin 0.01 Composition of mineral mixture: (g/kg mixture) Calcium phosphate (dibasic) 500 Sodium chloride 74 Potassium citrate monohydrate 220 Potassium sulphate 52 Magnesium oxide 24 Manganous carbonate 3.5: Ferric citrates Zinc carbonate 1.6 Cupric carbonate 0.3 Potassium iodate 0.01 Sodium selenite 0.01 Chromium potassium sulphate 0.55 Corn starch was added to make 1000g Composition of vitamin mixture (g/kg mix) Thiamin hydrochloride 0.6 Riboflavin 0.6 Pyridoxine hydrochloride 0.7 Nicotinic acid 3 D-calcium pantothenate 1.6 Folic acid 0.2 D-biotin 0.02 Cyanocobalamine (vit B-12) 0.001 Retinyl acetate (400000 I. U) 0.4 DL- 00 tocopherols acetate (5000 I.U) 7.7 Menadione 0.005 The corn starch is added to make the entire thing 1000g. The above said ingredients are mixed manually in the above said order at a temperature of 24-27°C for a period of 3- 45 minutes to obtain the desired formulation. The composition involving quercetin was examined for its efficacy on diabetic nephropathy state. The experimental groups showed considerable improvement in ameliorating the diabetic status as assessed by urine sugar, urine volume and fasting blood sugar. The kidney damage was estimated by measuring the glomerular filtration rate in diabetic and treated rats. The quercetin fed diabetic rats showed about 50% improvement in the diabetic nephropathy state. In an embodiment of the present invention, the final content of nutrients in the pharmaceutical formulation is in the range of, carbohydrates ranging from 65 to 75 percent, proteins ranging from 15 to 20 percent and fat content in the range of 5 to 15 percent. In another embodiment of the present invention, a method for controlling diabetic nephropathy progression is presented, wherein the said method comprises of administering a composition comprising quercetin to the mammals. The following examples are given by way of illustration of the present invention and therefore should not be construed to limit the scope of the present invention. Example 1 Diabetes was induced in male Wistar rats weighing around 120 g using streptozotocin at 55 mg / kg body weight and the rats were fed with formulation comprising quercetin. After three days, diabetic status was assessed by measuring fasting blood glucose level in plasma and urine sugar levels. Fasting blood glucose level was measured in the plasma of the rats by glucose oxidase method. The reaction mixture containing plasma and reagent was incubated for 15 minutes and absorbance was measured at 505nm. Urine was collected under the layer of toluene for 24 hrs and urine sugar (reducing) levels were measured by Dinitrosalicylic acid method. The reaction mixture was kept in boiling water bath for 10 minutes and colour developed was measured at 540 nm. Formulation devoid of quercetin was not effective in controlling the increased levels of fasting blood glucose, urine volume (Tablel), urine sugar (Tablel) during diabetes. Formulation involving antioxidant - quercetin was effective in controlling these changes as evident in Table 1. These results clearly indicate that during diabetes, progression of diabetic nephropathy was considerably controlled with the formulation comprising quercetin (Tablel). Table 1. Effect of quercetin on urine sugar, urine volume and fasting blood sugar in control and diabetic rats (Table Removed) Values are mean + SEM of 6 rats in control groups and 8 rats in diabetic groups, a. Statistically significant when compared to SFC at p b. Statistically significant when compared to SFD at p SFC-Starch fed control, SFD-Starch fed diabetic, QFC- Quercetin fed control and QFD-Quercetin fed diabetic. Example 2 Diabetes was induced in male Wistar rats weighing around 110-120 g using streptozotocin at 55 mg/kg body weight and the rats were fed with formulation comprising of quercetin. After three days diabetic status was assessed by measuring fasting blood glucose level in plasma and urine sugar levels. Fasting blood glucose level was measured in the plasma of the rats by glucose oxidase method. The reaction mixture containing plasma and reagent was incubated for 15 minutes and absorbance was measured at 505nm. Urine was collected under the layer of toluene and urine sugar (reducing) levels, was measured by dinitrosalicylic acid. The reaction mixture was kept in boiling water bath for 10 minutes and colour developed was measured at 540 nm. Jrinary creatinine was estimated by alkaline picrate reagent methi mixture was incubated at room temperature for 15 minutes and a measured at 520 nm. The plasma was deproteinised using tungstic creatinine estimation. The glomerular filtration rate (GFR) was determined at the end of the experiment by measuring urinary and plasma creatinine. The formula used was- Urinary creatinine (mg/dL) X urine volume (ml) GFR = —ml/min Plasma creatinine (mg/dL) X 1440 (min) As evident from the results formulation devoid of quercetin deteriorated kidney filtration during diabetes. Formulation involving antioxidant - quercetin was effective in controlling the kidney filtration during diabetes. Results clearly indicate that during diabetes, progression of diabetic nephropathy was considerably controlled with the formulation comprising quercetin (Table2). (Table Removed) Effect of quercetin on glomerular filtration rate in control and diabetic rats i ne main advantages of the present invention are: 1. A pharmaceutical composition involving quercetin is provided for treating diabetic nephropathy. 2. Quercetin (0.1%) helps to minimize the complications of kidney damage that take place during diabetic nephropathy state. 3. The disclosed pharmaceutical composition helps in reducing urine volume and urine sugar in patients suffering from diabetic nephropathy. 4. The composition of the present invention helps in reducing glomerular filtration rate in patients suffering from diabetic nephropathy. We claim: 1. A pharmaceutical composition useful for treating diabetic nephropathy comprising of the following ingredients: Ingredients % by weight in (g) [a] Corn starch 65-70 [b] Casein 15-20 [c] Vegetable Fat 05-15 [d] Mineral mix 02-05 [e] Vitamin Mix 01-05 [f] Quercetin 0.005 - 0.02 Composition of component [d] mineral (g/kg mixture: mixture) Calcium phosphate (dibasic) 500 Sodium chloride 74 Potassium citrate monohydrate 220 Potassium sulphate 52 Magnesium oxide 24 Manganous carbonate 3.5: Ferric citrate 6 Zinc carbonate 1.6 Cupric carbonate 0.3 Potassium iodate 0.01 Sodium selenite 0.01 Chromium potassium sulphate 0.55 Corn starch was added to make the total amount 1000g Composition of component [e] vitamin (g/kg mix) mixture Thiamin hydrochloride 0.6 Riboflavin 0.6 Pyridoxine hydrochloride 0.7 Nicotinic acid 3 D-calcium pantothenate 1.6 Folic acid 0.2 D-biotin 0.02 Cyanocobalamine (vit B-12) 0.001 Retinyl acetate (400000 I. U) 0.4 DL- DD tocopherols acetate (5000 I.U) 7.7 Menadione 0.005 Corn starch was added to make the total amount 1000g 2. A composition as claimed in claim 1, wherein the content of carbohydrates is in the range of 65 to 70 percent. 3. A composition as claimed in claim 1, wherein the content of proteins is in the range of 15 to 20 percent. 4. A composition as claimed in claim 1, wherein the content of fats is in the range of 5 to 15 percent. 5. A composition as claimed in claim 1, wherein the composition is used in controlling diabetic nephropathy in mammals. 6. A composition as claimed in claiml, wherein the said formulation is administered by oral feeding. 7. A process for the preparation of pharmaceutical composition as claimed in claim 1, wherein the process comprises of mixing the said ingredients manually in the above said order at a temperature of 20 - 35°C for a period of 3- 45 minutes to obtain the desired formulation. 8. A pharmaceutical composition useful for treating diabetic nephropathy and a process for the preparation thereof substantially as herein described with references to the foregoing examples.

Full Text

Field of the invention:
The present invention relates to a pharmaceutical composition useful for treating diabetic nephropathy.
Background and prior art of the invention:
Diabetic nephropathy is one of the complications of diabetes mellitus involving glomerular basement membrane thickening due to decrease in heparan sulphate and laminin and increase in type-IV collagen. Diabetic nephropathy is characterized by increased excretion of urine with micro and macromolecules (viz. proteins, glucose etc), as a result of glomerular basement membrane. During diabetic nephropathy, the functional unit of kidney, the nephrons are damaged leading to increased filtration rate.
Diet plays a major role in the management of diabetes and its associated complications. In this aspect, role of many traditional practices involving medicinal plants are known to have preventive and therapeutic value in diabetes management. The bioactive principles of medicinal plants and foods of medicinal value are receiving much scientific attention in recent years. In this direction anti-oxidants are of considerable interest. Higher oxidative stress is one of the factors implicated in the development and progression of diabetic complications resulting in increased levels of free radicals or impaired anti-oxidant defense mechanisms. Epidemiological evidence has suggested that anti-oxidant rich dietary flavanoids have potential health beneficial effects.
Quercetin is a well-documented bioflavonoid occurring in many foods and is known to be present in higher concentrations in onions, apples, broccoli, green tea and red wine. Hence, the polyphenolic compounds constitute an integral part of human diet though they are considered as non-nutrients. Many studies have focused towards the beneficial properties of quercetin, namely, anti-proliferative, anti-bacterial, anti-oxidant, anti-inflammatory and anti-carcinogenic. However, the anti-diabetic property of querectin is yet not reported anywhere nor studied. A formulation involving quercetin could be expected to have a significant role in combating glomerular filtration rate and progression of diabetic nephropathy

Reference may be made to US patent US 6,291,533 (2001) wherein quercetin dihydrate was used as a supplement in the compositions for treating people with a specific blood type. Drawback of this composition is that it is designed for people with specific antigenic blood types and have not specified for diabetes and its associated problems.
Reference may be made to US patent 20030108624 (2003) wherein Gymnema sylvestre; Fenugreek seed and Ginkgo biloba are used for prevention and treatment of chronic diseases and disorders including the complications of diabetes mellitus. Drawback of this is it fails to explain the specific role of quercetin in diabetic nephropathy.
Reference may be made to US patent 20020025347 (2002) wherein aqueous extract of Salviae Miltiorrhizae Radix herb is used for the treatment of diabetic nephropathy and microalbuminuria. Drawback of this invention is the composition of the herb and antioxidant activity in treatment of diabetic nephropathy is not clear.
Reference may be made to the US patent 20020192314 (2002) wherein dietary supplements of grape skin and grape seed extract can inhibit platelet aggregation or LDL cholesterol oxidation in a mammal. Drawback of this invention is that it deals with variety of polyphenolic components in prevention of oxidation but it does not specify for diabetes and its complications in particular diabetic nephropathy.
Reference may be made to US patent 6572897 (2003) wherein formulation contains essential amounts of alpha lipoic acid, chromium, lutein, bioflavonoids(quercetin and rutin), mormordica charantia extract, corosolic acid, and gymnema sylvestre extract used for insulin sensitivity and blood sugar level maintenance for the prevention and treatment of diabetes. The drawback of this formulation is it does not explain the effect of this in diabetic nephropathy state and it is not a single component used for the treatment.

Reference may be made to the Japanese patent 2002080362 (2002) wherein gallic ester, galloyl tannin, quercetin, catechins and epicatechins are used in regulating PPAR-(Peroxisome Proliferator Activated Receptor)-dependent gene transcription activity; an agent for obesity, diabetes, hyperglycemia, hyperlipemia and insulin resistance. Drawback of this invention is it does not explain the activity of diabetic nephropathy complications.
Reference may be made to the patent WO0059522 (2000) wherein extracted products from Brickellia californica and its isolated flavonoids, including apigenin, luteolin, quercetin and dihydroxykaemferol results in lowering of blood sugar and are thereby used in the treatment of both insulin dependent and non-insulin dependent diabetes. The drawback of this is that it is an herbal extract and it does not explain the role of quercetin in diabetes or its complications.
Reference may be made to the Japanese patent 3232851 (1991) wherein phenolic compounds extracted from various plants like tannic acid, pentagalloyl glucose, rutin, quercetin, epicatechin are used as therapeutic agent of various complications in diabetes mellitus such as cataract, retinopathy, kidney disease or nerve disturbance by inhibiting aldose reductase. The drawback of this invention is that it does not clearly explain the role of these compounds in glomerular filtration rate during diabetic nephropathy state.
Reference may be made to US patent 2004258674 (2004) wherein nutritional supplement of quercetin to treat hypertenstion, therby preventing cardiovascular disease is disclosed. The drawback of this treatment is that it does not explain anything about diabetes and its related complications.
Reference may be made to patent WO2004037015 (2004) wherein a method of treating Crohn's disease, arthritis, leukemia, lowering cholesterol levels or blood pressure using antioxidant compositions of various plant products soy isoflavones, taurine, sugar beet pectin fiber, or a ginko biloba extract are described. The drawback is that it includes variety of antioxidants and does not treat diabetes and its associated complications.

Reference may be made to US patent 2004014686 (2004) wherein a combination of catechin and quercetin possessing a potential antioxidant activity is used as pharmaceutical and dietary formulation. The drawback of this is that a combination of antioxidants is used and individual effects on the disease condition particularly in diabetic nephropathy are not explained.
Reference may be made to Korean patent 2003015815 (2003) wherein quercetin is used as an inhibitor of gene expression of vascular endothelial growth factor (vegf) and erythropoietin (epo). The drawback of this invention is that it fails to explain the role of quercetin in diabetic nephropathy.
Novelty:
Novelty of this invention is that quercetin (0.1%) helps to minimize the complications of kidney damage that take place during diabetic nephropathy state.
Objects of the invention:
The main objective of the present invention is to provide a process for the preparation of pharmaceutical formulation containing quercetin for treating diabetic nephropathy.
Another objective of the invention is to provide a pharmaceutical formulation containing quercetin for treating diabetic nephropathy.
Summary of the invention:
This invention relates to a pharmaceutical formulation, useful for treating diabetic nephropathy, comprising of mixing a number of different ingredients at a temperature of 24-27°C for a period of 3 to 45 minutes, to obtain the desired formulation. In the final formulation, the carbohydrate content ranges from 65 to 75 percent, proteins range from 15 to 20 percent and fat content is in the range of 5 to 15 percent. The said formulation is useful for controlling diabetic nephropathy and is administered by oral feeding to mammals. Accordingly, the present invention provides a pharmaceutical composition useful for
treating diabetic nephropathy comprising of the following ingredients:

Ingredients % by weight in (g)
[a] Corn starch 65-70
[b] Casein 15-20
[c] Vegetable Fat 05-15
[d] Mineral mix 02-05
[e] Vitamin Mix 01-05
[f] Quercetin 0.005 - 0.02
Composition of component [d] mineral (g/kg
mixture: mixture)
Calcium phosphate (dibasic) 500
Sodium chloride 74
Potassium citrate monohydrate 220
Potassium sulphate 52
Magnesium oxide 24
Manganous carbonate 3.5:
Ferric citrate 6
Zinc carbonate 1.6
Cupric carbonate 0.3
Potassium iodate 0.01
Sodium selenite 0.01
Chromium potassium sulphate 0.55
Corn starch was added to make the total amount 1000g
Composition of component [e] vitamin (g/kg mix)
mixture
Thiamin hydrochloride 0.6
Riboflavin 0.6
Pyridoxine hydrochloride 0.7
Nicotinic acid 3
D-calcium pantothenate 1.6

Folic acid 0.2
D-biotin 0.02
Cyanocobalamine (vit B-12) 0.001
Retinyl acetate (400000 I. U) 0.4
DL- DD tocopherols acetate (5000 I.U) 7.7
Menadione 0.005
Corn starch was added to make the total amount 1000g
Detailed description of the invention:
The present invention relates to a design of a pharmaceutical formulation useful for diabetic nephropathy comprising of antioxidant - quercetin. The said pharmaceutical formulation is useful for diabetic nephropathy and is comprising of the following ingredients in the given range of percent, by weight in gram:
Ingredients % by weight in (g)
Corn starch 65-70
Casein 15-20
Vegetable Fat 05-15
Mineral mix 02-05
Vitamin Mix 01 -05
Quercetin 0.01
Composition of mineral mixture: (g/kg mixture)
Calcium phosphate (dibasic) 500
Sodium chloride 74
Potassium citrate monohydrate 220
Potassium sulphate 52
Magnesium oxide 24 Manganous carbonate 3.5: Ferric citrates

Zinc carbonate 1.6
Cupric carbonate 0.3
Potassium iodate 0.01
Sodium selenite 0.01
Chromium potassium sulphate 0.55
Corn starch was added to make 1000g
Composition of vitamin mixture (g/kg mix)
Thiamin hydrochloride 0.6
Riboflavin 0.6
Pyridoxine hydrochloride 0.7
Nicotinic acid 3
D-calcium pantothenate 1.6
Folic acid 0.2
D-biotin 0.02
Cyanocobalamine (vit B-12) 0.001
Retinyl acetate (400000 I. U) 0.4
DL- 00 tocopherols acetate (5000 I.U) 7.7
Menadione 0.005
The corn starch is added to make the entire thing 1000g. The above said ingredients are mixed manually in the above said order at a temperature of 24-27°C for a period of 3- 45 minutes to obtain the desired formulation.
The composition involving quercetin was examined for its efficacy on diabetic nephropathy state. The experimental groups showed considerable improvement in ameliorating the diabetic status as assessed by urine sugar, urine volume and fasting blood sugar. The kidney damage was estimated by measuring the glomerular filtration rate in diabetic and treated rats. The quercetin fed diabetic rats showed about 50% improvement in the diabetic nephropathy state.

In an embodiment of the present invention, the final content of nutrients in the pharmaceutical formulation is in the range of, carbohydrates ranging from 65 to 75 percent, proteins ranging from 15 to 20 percent and fat content in the range of 5 to 15 percent.
In another embodiment of the present invention, a method for controlling diabetic nephropathy progression is presented, wherein the said method comprises of administering a composition comprising quercetin to the mammals.
The following examples are given by way of illustration of the present invention and therefore should not be construed to limit the scope of the present invention.
Example 1
Diabetes was induced in male Wistar rats weighing around 120 g using streptozotocin at 55 mg / kg body weight and the rats were fed with formulation comprising quercetin. After three days, diabetic status was assessed by measuring fasting blood glucose level in plasma and urine sugar levels. Fasting blood glucose level was measured in the plasma of the rats by glucose oxidase method. The reaction mixture containing plasma and reagent was incubated for 15 minutes and absorbance was measured at 505nm. Urine was collected under the layer of toluene for 24 hrs and urine sugar (reducing) levels were measured by Dinitrosalicylic acid method. The reaction mixture was kept in boiling water bath for 10 minutes and colour developed was measured at 540 nm.
Formulation devoid of quercetin was not effective in controlling the increased levels of fasting blood glucose, urine volume (Tablel), urine sugar (Tablel) during diabetes. Formulation involving antioxidant - quercetin was effective in controlling these changes as evident in Table 1. These results clearly indicate that during diabetes, progression of diabetic nephropathy was considerably controlled with the formulation comprising quercetin (Tablel).

Table 1. Effect of quercetin on urine sugar, urine volume and fasting blood sugar in control and diabetic rats

(Table Removed) Values are mean + SEM of 6 rats in control groups and 8 rats in diabetic groups,
a. Statistically significant when compared to SFC at p b. Statistically significant when compared to SFD at p SFC-Starch fed control, SFD-Starch fed diabetic, QFC- Quercetin fed control and QFD-Quercetin fed diabetic.
Example 2
Diabetes was induced in male Wistar rats weighing around 110-120 g using streptozotocin at 55 mg/kg body weight and the rats were fed with formulation comprising of quercetin. After three days diabetic status was assessed by measuring fasting blood glucose level in plasma and urine sugar levels. Fasting blood glucose level was measured in the plasma of the rats by glucose oxidase method. The reaction mixture containing plasma and reagent was incubated for 15 minutes and absorbance was measured at 505nm. Urine was collected under the layer of toluene and urine sugar (reducing) levels, was measured by dinitrosalicylic acid. The reaction mixture was kept in boiling water bath for 10 minutes and colour developed was measured at 540 nm.
Jrinary creatinine was estimated by alkaline picrate reagent methi
mixture was incubated at room temperature for 15 minutes and a
measured at 520 nm. The plasma was deproteinised using tungstic
creatinine estimation.
The glomerular filtration rate (GFR) was determined at the end of the experiment by
measuring urinary and plasma creatinine. The formula used was-
Urinary creatinine (mg/dL) X urine volume (ml)

GFR =

—ml/min

Plasma creatinine (mg/dL) X 1440 (min)
As evident from the results formulation devoid of quercetin deteriorated kidney filtration during diabetes. Formulation involving antioxidant - quercetin was effective in controlling the kidney filtration during diabetes. Results clearly indicate that during diabetes, progression of diabetic nephropathy was considerably controlled with the formulation comprising quercetin (Table2).
(Table Removed) Effect of quercetin on glomerular filtration rate in control and diabetic rats

i ne main advantages of the present invention are:
1. A pharmaceutical composition involving quercetin is provided for treating diabetic
nephropathy.
2. Quercetin (0.1%) helps to minimize the complications of kidney damage that take
place during diabetic nephropathy state.
3. The disclosed pharmaceutical composition helps in reducing urine volume and
urine sugar in patients suffering from diabetic nephropathy.
4. The composition of the present invention helps in reducing glomerular filtration
rate in patients suffering from diabetic nephropathy.

We claim:
1. A pharmaceutical composition useful for treating diabetic nephropathy comprising of
the following ingredients:
Ingredients % by weight in (g)
[a] Corn starch 65-70
[b] Casein 15-20
[c] Vegetable Fat 05-15
[d] Mineral mix 02-05
[e] Vitamin Mix 01-05
[f] Quercetin 0.005 - 0.02
Composition of component [d] mineral (g/kg
mixture: mixture)
Calcium phosphate (dibasic) 500
Sodium chloride 74
Potassium citrate monohydrate 220
Potassium sulphate 52
Magnesium oxide 24
Manganous carbonate 3.5:
Ferric citrate 6
Zinc carbonate 1.6
Cupric carbonate 0.3
Potassium iodate 0.01
Sodium selenite 0.01
Chromium potassium sulphate 0.55
Corn starch was added to make the total amount 1000g
Composition of component [e] vitamin (g/kg mix)
mixture

Thiamin hydrochloride 0.6
Riboflavin 0.6
Pyridoxine hydrochloride 0.7
Nicotinic acid 3
D-calcium pantothenate 1.6
Folic acid 0.2
D-biotin 0.02
Cyanocobalamine (vit B-12) 0.001
Retinyl acetate (400000 I. U) 0.4
DL- DD tocopherols acetate (5000 I.U) 7.7
Menadione 0.005
Corn starch was added to make the total amount 1000g
2. A composition as claimed in claim 1, wherein the content of carbohydrates is in
the range of 65 to 70 percent.
3. A composition as claimed in claim 1, wherein the content of proteins is in the
range of 15 to 20 percent.
4. A composition as claimed in claim 1, wherein the content of fats is in the range of
5 to 15 percent.
5. A composition as claimed in claim 1, wherein the composition is used in
controlling diabetic nephropathy in mammals.
6. A composition as claimed in claiml, wherein the said formulation is administered
by oral feeding.
7. A process for the preparation of pharmaceutical composition as claimed in claim
1, wherein the process comprises of mixing the said ingredients manually in the
above said order at a temperature of 20 - 35°C for a period of 3- 45 minutes to
obtain the desired formulation.
8. A pharmaceutical composition useful for treating diabetic nephropathy and a
process for the preparation thereof substantially as herein described with
references to the foregoing examples.

Documents:

http://ipindiaonline.gov.in/patentsearch/GrantedSearch/viewdoc.aspx?id=53nl6bQTxLzY/ulBeqay5g==&loc=+mN2fYxnTC4l0fUd8W4CAA==

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Patent Number

279549

Indian Patent Application Number

313/DEL/2006

PG Journal Number

04/2017

Publication Date

27-Jan-2017

Grant Date

25-Jan-2017

Date of Filing

03-Feb-2006

Name of Patentee

COUNCIL OF SCIENTIFIC & INDISTRIAL RESEARCH

Applicant Address

ANUSANDHAN BHAWAN, RAFI MARG, NEW DELHI-110001, INDIA

Inventors:

#	Inventor's Name	Inventor's Address
1	PARAMAHANSA V. SALIMATH	DEP CFTRI Mysore - 570020.
2	KARI SAMBAIAH	DEP. CFTRI Mysore - 570020.

PCT International Classification Number

A61K 31/07

PCT International Application Number

N/A

PCT International Filing date

PCT Conventions:

#	PCT Application Number	Date of Convention	Priority Country
1			NA