WO2010082906A1

WO2010082906A1 - Use of nerium oleander for diseases manifested with type ii diabetes, obesity, high cholesterol and triglyceride

Info

Publication number: WO2010082906A1
Application number: PCT/TR2009/000013
Authority: WO
Inventors: Ahmet Levent Bas; Sule Kaya; Nuray Yazihan; Kamil Uney
Original assignee: Ahmet Levent Bas; Sule Kaya; Nuray Yazihan; Kamil Uney
Priority date: 2009-01-15
Filing date: 2009-02-03
Publication date: 2010-07-22
Also published as: TR200900312A2

Abstract

The invention relates to a Nerium extract, the method of obtaining, and using it to be used regulation of lipid metabolism in diseases manifested with Type II diabetes, obesity, high cholesterol and triglyceride levels, and cardiovascular diseases.

Description

USE OF NERIUM OLEANDER FOR DISEASES MANIFESTED WITH TYPE Il DIABETES, OBESITY, HIGH CHOLESTEROL AND TRIGLYCERIDE

Technical Field

The invention relates to the regulation of glucose and lipid metabolisms in diseases manifested by Type Il diabetes, obesity, high cholesterol and triglyceride levels and cardiovascular diseases.

The invention relates, in particular to the use of Nerium oleander composition for the control of high blood glucose levels, the control of cholesterol and triglyceride levels and the modulation of HDL-LDL ratios seen in patients with Type Il diabetes.

State of the Art

Plant anti-diabetics and Nerium oleander in these days;

According to information obtained from Ebers papyruses the use of medicinal plants for the treatment of diabetes dates back to around 1550 B.C. (¹). In many places in the world various plants are used by traditional methods for the treatment of diabetes. Some of these traditional plant treatments used are taken into consideration in scientific circles and the World Health Organization (WHO) supports studies in this field. Even though insulin and oral anti-diabetics are used in modern medicine, in especially developing countries due to factors such as the provision, storage and application of these medicines and due to the side effects of drugs, a trend in searching for new natural or synthetic antidiabetic drugs as an alternative medicine has started (²). It is known that, applications of plants are called upon in various regions, in our country as well, for the treatment of diabetes (³); and additionally, scientific studies are being conducted on the hypoglycemic effects of medicinal plants (⁴).

Oleander (lat. Nerium oleander, NO) of the dogbane (Apocynaceae) family grows along the whole Mediterranean coast starting in southern Portugal in the west, and in Syria, and in streambeds of the Western and Southern Anatolia. Oleander is a plant which has a historical use since the antique ages up till our day. Oleander, which has a limited folkloric use in the countries of the Mediterranean basin where it grows, is basically a plant which has distinct toxic properties. The fruit of oleander, which flowers in summer and has a long flowering period, has a horse bean shape. Because it is poisonous, it is dangerous for humans and animals; it is also grown as an ornamental plant. Phase I clinical tests for NO, which was brought to the agenda approximately 15 years ago with its use in the treatment of cancer by Dr. Ziya Ozel, are currently ongoing in the USA (⁵).

Different parts of NO, also known in our country as "zakkum, poison flower, zikkim tree", are used in different sections of the Mediterranean basin among local people for edema, leprosy, eye and skin diseases. The effects of the cardiotonic content found in its leaves on antibacterial, anti-carcinogen, hemorrhage-clotting time and the central nervous system has been subject to various researches (⁶).

As the result of studies of various parts of the plant, the existence of different glycosides, triterpenes and long chained composites were identified (⁷).

In the water extract of shredded leaves of NO, there is 2.3 % polysaccharide; 67 % of the polysaccharide in the composition comprises galacturonic acid, rhamnose, arabinose, and galactose (⁸).

It was stated that the fractions obtained through methanol extraction of the fresh and dried leaves of NO have analgesic effects (⁸).

NO has a potential of positive inotropic effect on human and animal heart. This effect of oleander cardenolides is similar to the Na⁺K⁺/ATP'ase inhibition in plasmalemma, as with classical digital glycosides, in terms of its physiological effect pattern. However, oleander and digital cardenolides differ in terms of their toxicities and extra- cardiac effects (⁹).

It has been stated that, central nervous system (CNS) depression was caused on mice with 50 mg/kg dose uses of the cardenolide group substances in NO leaves, isolated subsequent to methanol extraction (⁹). Researchers have stated that 4 substances of the cardenolide group (neridiginoside, nerizoside, neritaloside and odoroside-H) in the methanol extract of fresh NO leaves cause CNS depression in mice with 25 mg/kg dose. In the roots of NO, on the other hand, there are steroidal substances such as proceragenin and neridienone (¹⁰). Another group of substances found in the leaves of NO are the triterpenes. These substances on generally by ethyl acetate extraction are of tarakasterane and ursane types. In studies relating to the identification of triterpene group of substances in the NO composition and their biological effects, the presence of in vitro anti-inflammatory and anti-carcinogen effects were evaluated. Any inhibitory activities of metal extracts of ursolic acid and oleanoic acid against the intracellular adhesion molecule (ICAM- 1 )'s induction and their effects on cell development in human cell series were established (¹¹). The similar inhibitor effects relating to ICAM-1 of the three new triterpenes isolated and their cytotoxic effects on A-549, WI-38, VA-13 and HepG2 cells were also evaluated (¹²).

Oleadrine is one of the primary substances found in NO leaves. In one study performed under in-vivo conditions (¹³), its anti-inflammatory effects and effects on tumor cell development were evaluated. The researchers have stated that the local application of 2 mg oleadrine for prophylactic purposes provided positive results against the effects of TPA (l2-O-tetradecanoylphorbol-13-acetate), which is a substance widely used on the formation of skin tumors.

In our country the form of NO obtained by keeping in alcohol for a long time is used for pains of lower extremities and in paralysis cases. Additionally, there are data that its form obtained after keeping in oil is used against rheumatoid arthritis (¹⁴). Records exist that the leaves or flowers are used in painful cases and eczema, and the sap obtained from fresh leaves are used for abscess and rheumatic cases (¹⁵). In Saudi Arabia, the fruit of NO and the sap of the plant are used as anti-rheumatic and for skin diseases (¹⁶).

It has been stated that the water and ethanol extracts of NO reduce to a significant degree the abdominal contractions created in mice by p-benzoquinone (antinociceptive effect) and has a significant anti-inflammatory effect on carrageenan- induced edema on the back foot with (¹⁷). In pain trials, in applications with fresh and dried flowers of the plant the fact that gastric ulcerations were not observed in the subjects was monitored as a noteworthy finding.

Antimicrobial activities of alcohol, water and chloroform extracts of NO against 10 micro-organisms (8 different bacteria and 2 yeasts) were studied and whereas the alcohol phase products had antimicrobial activity against only Bacillus cereus and Psυdomanas aeruginosa, no antimicrobial activity was found against the studied bacteria and yeast with the products of water and chloroform phases (¹⁸).

The basic cardiac glycosides found in the composition of Anvirzel™ (sterile filtered lyophilized hot water extract of NO) are oleandrine, odorside, neritaloside and aglycone oleandrigenin. Subsequent to oral (80 μg/kg) administration of Anvirzel™ in mice, oleadrine is rapidly absorbed (C_max 20 min), the elimination phase half-life is (tv₂ β) 2.3±0.5 hours, and oral absorbrion ratio is at the level of 30 %. The liver level of oleadrine subsequent to its IV administration is approximately twice that of the heart and kidney tissues. Oleadrigenine, which is the aglicone form of oleadrine, is also found in tissues subsequent to administration at an approximately Y≥ level. 1.9% of oleadrine and 4.4% of oleadrigenine are eliminated by urine within 24 hours, their level of clearance with feces is higher (66% for both of the substances) (¹⁹).

In the Phase I studies performed regarding the use of Anvirzel™, which is obtained with the water extract of NO, basically a poisonous plant, against solid tumors it is stated that administration by IM method at a dose of 1.2 mg/m²/day is safe. It is reported that symptoms such as pain in the application area and additionally, fatigue, nausea and dyspnea were observed during the application (20).

The tendency to suppress weight gain (% ratio as compared to their weights in the beginning) after a 9 week administration (1 per week) of a non-lethal dose

(1000mg/kg bw) in alcohol of dried leaves of NO on female and male mice by SC method was determined. Multiple administrations of the determined dose have not caused significant changes in the myocardium and blood parameters (WBC, RBC,

Hb, HCT, PLT). On the other hand, the lethal dose (4000 mg/kg bw) has caused myocardial electrical activities which resulted in heart arrest (²¹). In order to determine the acute toxicity of dried NO leaves; single dosage (1 and 0.25 mg/kg bw) and a daily (0.06 g/kg bw) NO administration were studied in Nadji sheeps; and restlessness, mastication, dyspnea, gas accumulation in rumen, ataxia in movement and paresis of muscles; and death after 4-24 hours were determined (²²). Hemorrhage, cyanotic lungs, emphysema, catarrhal abomasitis and enteritis were noted by autopsy of dead animals. It was expressed that the daily administered NO caused milder symptoms and that deaths were observed in 3-14 days. In autopsy, hepatonephropathy, gelatinization in renal pelvis and mesenterium were observed. In biochemical analyses, on the other hand, it was stated that increases in AST and LDH activities, bilirubin, cholesterol and urea densities and decrease in total protein and albumin levels, anemia and leucopenia were observed.

In our country scientific studies are being conducted on hypoglycemic effects of medicinal plants. In research where the hypoglycemic effects of the extract obtained from the fruits of the Urtica pilulifera (roman nettle) plant on mice were evaluated, it was exhibited that the extract prepared had a statistically significant effect (²³). In a research, where the hypoglycemic effect of Rheum ribes (rhubarb) roots, consumed in the city of Van and its vicinity, on healthy and alloxane diabetic mice was studied, it was stated that the extract of the plant had hypoglycemic effect on mice (²⁴).

Today the interest in the use of vegetable sourced medicines and medicinal raw materials is increasing each day. Additionally, in recent years numerous biological activity tests are being performed on plants and active substances are identified and the activities are scientifically evidenced.

Publications on the anti-diabetic efficacy of oleander, which is used for different purposes in the countries where it grows and in our country, are very limited (²⁵). Researchers have evaluated oleander among anti-diabetic factors in the classification of plant substances used in Morocco among local people. There are findings regarding the use of oleander for anti-diabetic purposes in investigations performed by different researcher groups in the same region (²⁶).

Diabetes Mellitus (DM) is a widespread endocrine disorder manifested with chronic increase of blood glucose. It develops in relation with the relative or absolute deficiency of the insulin hormone. One of the most important clinical characteristics of diabetes is chronic tissue complications. In the long term, microangiopathic changes are observed in the nerves and the small veins on the eye and kidney tissues. The frequency of atherosclerosis in larger veins distinctly increases.

Diabetes is a widespread disease as a public health issue. In Europe, as well, approximately 1-2% of the population has diabetes. This ratio is 3-8% in the USA. While its worldwide prevalence is 100 million, it is estimated that this number shall double in the next 10-15 years. Diabetes is diagnosed with the presence of chronic hyperglycemia. Classical symptoms of the disease are polyuria, polydipsia, polyphagia and high venous plasma glucose along with weight loss. Diagnosis may be made in the presence of explained symptoms without requiring further examination. It is possible to have an idea of the history of blood glucose levels with HbAIc measurements.

Type Il diabetes (Non Independent Diabetes Mellitus-NIDDM) is the type which is non insulin related. Type Il diabetes generally starts in middle - or advanced ages, and is more prevalent (80 %). It is reported that the disease may be related to disorder in insulin secretion or insulin resistance in target cells.

Approximately 80 % of type Il diabetes patients are obese. In both sexes, as the weight gain progressively increases the risk of developing diabetes increases. Obesity, in the least, causes a reduction in insulin sensitivity.

Insulin resistance is frequently related to cases where biochemical and clinical characteristics known as metabolic syndrome X (insulin resistance syndrome) are found together (glucose intolerance, truncal obesity, and hypertension). In such cases low HDL, increase VLDL cholesterol and accelerated atherosclerosis are observed. While LDL and normal cholesterol are relatively normal, low HDL and high triglycerides are characteristic of type Il diabetes (dyslipidemia).

Blood glucose displays fluctuations in type I diabetics. Therefore, a single glucose measurement provides very little information. In type II, on the other hand, blood glucose levels are more stable. It is possible to have an acceptable idea about the fasting glucose.

The starting point of treatment is the arrangements in diet and lifestyle. Diet is important in reducing body weight, lowering blood glucose concentration, increasing insulin sensitivity and reducing cardiovascular risk factors such as abnormal lipid profile and hypertension.

Same diets are recommended for both type I and type Il diabetics. Instead of saturated fatty acids, unsaturated fatty acids should be increased and carbohydrates with complex fibers should be consumed. Diet and exercise have been found to be effective in achieving glycemic control in 20 % of type Il diabetes cases. After this step, oral anti-diabetic agents should be recommended. Sulphonylurea group substances which are among the first invented medicines stimulate insulin secretion. Biguanide compositions (metformin etc.) increase insulin secretion and inhibit hepatic gluconeogenesis. While metformin is used with obese diabetics with marked insulin resistance, sulphonylureas are generally used with obese patients with whom the major defect is believed to be disrupted insulin secretion. Metformin may also reduce appetite and increase weight loss.

It may cause nausea and diarrhea in 25% of the cases. However, these can be reduced with decreased dose and food ingest. Lactic acidosis is rare and NO is contraindicated in those with renal, hepatic and cardiac diseases.

These agents used may, after some time, become insufficient due to worsening of beta cell functions and/or insulin resistance. In such cases metformin and sulphonylurea combination or alpha-glucosidase inhibitors are tried. Many patients may require insulin in the end.

Among alpha-glucosidase inhibitors, acarbose reduces the absorption of carbohydrates by inhibiting disaccharidase enzyme in the intestinal tract. Particularly postprandial blood glucose is lowered. Its side effects are gas, abdominal tension and diarrhea.

Glimepiride, among new oral agents, is similar to sulphonylurea derivative drugs. It causes sparing use of insulin by extra-pancreatic effect. It has a risk of low hypoglycemia. The important of insulin resistance in the pathophysiology of type Il diabetes was caused the development of other new drugs. The first among Thiazolidines that entered the practice was triglitazone in 1997. Even though it was removed from use in the UK due to hepatic damage, it is still in use in the USA. During its use liver functional tests should be checked monthly. The most effective among these groups of drugs is Roziglitazone. Hepatotoxicity was not reported after clinical trials. Roziglitazone improves glucose control, corrects hyperinsulinemia by overcoming insulin resistance. These agents also reduce triglycerides and free fatty acids. Diet, anti-diabetic agent, 2- , 3- combinations are used in the treatment of type Il diabetes. If glycemic control cannot be achieved with these treatment protocols; insulin injections daily are started usually.

During the use of oral anti-diabetic agents, certain important side effects may occur. While Metformin is used with obese diabetics with marked insulin resistance, sulphonylureas are generally used with non-obese patients considered to have disrupted insulin secretion. Metformin may also reduce appetite and increase weight loss. Metformin may cause nausea and diarrhea in 25% of the cases. It is contraindicated for those with renal, hepatic and cardiac diseases. These agents used may, after some time, become insufficient due to worsening of beta cell functions and/or insulin resistance. In such cases metformin and sulphonylurea combination or alpha-glucosidase inhibitors are tried. Many patients may require insulin in the end. The primary side effects of alpha-glucosidase inhibitors are gas, abdominal tension and diarrhea. During the use of oleander extract, obese/non- obese limitation is not in question, gastrointestinal problems have not been observed is animals used in trial. During application weight loss has been prevented in the animals. The fact that weight loss was prevented while blood glucose level was decreasing indicates to "insulin like effect" with the use of oleander extract, rather than a worsening of beta cell functions and/or insulin resistance.

During the use of oleander extract as well as not causing sudden fluctuations in glucose, a controlled reduction was obtained, and marked liver and kidney damage was not formed. In addition to glycemic control, the control of triglycerides, cholesterol and HDL/LDL were achieved in Type Il diabetes induced rats fed a high fat diet where 58 % of metabolic energy was provided with animal fat content. During this time the necessity of administration of combined drug and insulin with the use of more than one anti-diabetic drugs was not in question.

Consequently, the presence of a need for the use of Nerium oleander composition for controlling high blood glucose levels observed is patients with type Il diabetes, for the control of cholesterol and triglyceride levels and the modulation of HDL-LDL ratios, and the inadequacy of existing solutions has made it necessary to bring an improvement in the related technical filed. Object of the Invention

The object of the invention is; to eliminate the disadvantages with the state of the art, one object of the invention is to; regulate the glucose and lipid metabolisms in diseases manifested with Type Il diabetes, obesity, high cholesterol and triglyceride levels, and the glucose and lipid metabolisms in cardiovascular diseases, using Nerium oleander composition.

Another object of the invention; is to enable the control of high blood glucose levels, which are observed in patients with type Il diabetes, the control of cholesterol and triglyceride levels and the modulation of HDL-LDL ratios.

Another object of the invention is to; enable its use, as supportive treatment in patients who have type Il diabetes, for the purpose of keeping lipid profile and blood glucose under control.

Another object of the invention is; to provide for the control of high HbAIc levels, which are observed in patients with type Il diabetes, using Nerium oleander composition.

Another object of the invention is to; enable its use, as supportive treatment in patients who have type Il diabetes, for the purpose of keeping high HbAIc levels under control.

Another object of the invention is to; reduce the cardiotoxicity risk which may develop in relation with these substances, since less glycosides and flavonoid, but more polysaccharides shall be present with the implementation of hot water extraction during obtaining the oleander extract.

Another object of the invention is to; enable its use, in the control and modulation of leptin levels in Type Il diabetes and obesity for the treatment and prophylactic purposes.

Another object of the invention is to; enable its use, in the supportive treatment for the purpose of keeping high lipid profiles under control in patients with cardiovascular diseases, with some diseases related with dyslipidemia and in obesity. The invention may be used in diabetes and especially in etiopathogenesis as an immunomodulator, by affecting the regulation of release of proinflammatory cytokines (IL-1 , TNF-alpha etc.) in atherosclerotic cases, other cardiovascular pathologies and inflammatory processes where acute or chronic inflammation is present.

Another object of the invention is to; enable low side effects.

Another object of the invention is to; offer a low cost solution in the treatment of the disease.

Another object of the invention is to; provide a combined effect (combined effect on glucose, cholesterol, triglyceride and HDL/LDL ).

Another object of the invention is to; be suitable for oral use.

Another object of the invention is to; enable the use of a single dose per day to be sufficient.

Another object of the invention is to; not to have a requirement to diet as with the use of atorvastatin.

Another object of the invention is to; prevent the loss of body weight in patients with type Il diabetes.

Another object of the invention is to; enable the loss of body weight in healthy individuals.

In order to achieve the preceding advantages, the invention;

Relates to; a Nerium extract to be used for the regulation of lipid metabolism in diseases manifested with type Il diabetes, obesity, high cholesterol and triglyceride levels, and cardiovascular diseases.

Nerium relates to; Nerium oleander.

Relates to a method of obtaining Nerium oleander extract characterized in that it comprises the process steps of;

- Cleaning by washing the Nerium oleanders collected - Subsequent to the washing process cutting the Nerium oleanders into small pieces,

- Obtaining a mixture by adding chopped Nerium oleanders in distilled water,

Putting the mixture in a container and applying heat to said mixture,

- Subsequent to the heat application on the mixture, without covering the lid of the container until the liquid starts to evaporate, and after the evaporation starts, by closing the system and collecting the vapor formed in a separate container by causing it to come in contact with a surface cooled with cold water,

collecting the Nerium oleanders from among new shoots during the March- September period,

subsequent to the washing process cutting the Nerium oleanders into small pieces of approximately 1-3 cm length,

chopping up of approximately 400-700 gr (preferably 500 gr),

obtaining a mixture by adding chopped Nerium oleanders in 5-10 It (preferably 8 It) distilled water,

placing the mixture in a heat resistant container and applying heat to this mixture for approximately 30-50 min (preferably 40 min),

performing the process of collecting the vapor formed subsequent to the application of heat to the mixture, in a separate container by causing it to come in contact with a surface cooled with cold water for a period of 3-6 hours (preferably 5 hours) on taking care not to burn the mixture,

keeping the obtained Nerium oleander extract in a clean glass container at room temperature until use.

Relates to; the oral use of the magistral and specialty form of Nerium oleander extract. Relates to; the rectal use of the magistral and specialty form of Nerium oleander extract.

Relates to; the intravenous use of the magistral and specialty form of Nerium oleander extract.

Relates to; the intramuscular use of the magistral and specialty form of Nerium oleander extract.

Relates to; the subcutaneous use of the magistral and specialty form of Nerium oleander extract.

Relates to; the oral spray use of the magistral and specialty form of Nerium oleander extract.

Relates to; the nasal spray use of the magistral and specialty form of Nerium oleander extract.

Relates to; the use of Nerium oleander extract in the treatment of Type Il diabetes and obesity.

Relates to; the use of Nerium oleander extract in the prevention of loss of body weight in patients with Type Il diabetes.

Relates to; the use of Nerium oleander extract in enabling loss of body weight in healthy individuals.

Relates to; the use of Nerium oleander extract in the control of high blood glucose levels seen in patients with Type Il diabetes.

Relates to; the use of Nerium oleander extract in the control of cholesterol and triglyceride levels.

Relates to; the use of Nerium oleander extract for the modulation of HDL-LDL ratios.

Relates to; the use of Nerium oleander extract in patients with Type Il diabetes as supportive treatment for keeping the lipids profile and blood glucose under control. Relates to; the use of Nerium oleander extract in the control of high HbAIc levels seen in patients with Type Il diabetes.

Relates to; the use of Nerium oleander extract in patients with Type Il diabetes as supportive treatment for the purpose of keeping high HbAIc levels under control.

Relates to; the conversion of Nerium oleander extract to "specialty" form.

Relates to; the use of Nerium oleander extract as a food additive.

Relates to; the use of Nerium oleander extract for autoimmune diseases and in pathologies manifested with proinflammatory cytokine activation in oral, IV, IM, intraarticular form or as food additive.

Relates to; the local use of Nerium oleander extract for ulcerations, inflammations where inflammation is dominant and/or is the inducer.

Relates to; the particularly local use of Nerium oleander extract for skin diseases such as dermatitis, eczema, psoriasis advancing with dysregulation in cell proliferation and increased inflammation.

Relates to; the use of Nerium oleander extract in the control and modulation of leptin levels in Type Il diabetes and obesity for the treatment and prophylactic purposes.

Relates to; the use of Nerium oleander exctract in the supportive treatment for the purpose of keeping high lipid profiles under control in patients with cardiovascular diseases, with some diseases related with dyslipidemia and in obesity.

The composition and characteristic properties of the invention and all its advantages shall be more clearly comprehended with the figures given below and the detailed description written by reference to those figures and therefore, the evaluation needs to be made taking into consideration those figures and the detailed description. Figures;

Figure 1. Fasting blood glucose levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1: Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 2. Fasting blood HbAIc levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml

NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat,

YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 3. Fasting blood Triglyceride levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD)₁ Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 4. Fasting blood cholesterol levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD). Figure 5. Fasting blood Cholesterol, HDL and LDL levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01: Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 6. Body Weight levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 7. Fasting blood HDL levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 8. Fasting blood LDL levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1: Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD). Figure 9. Fasting blood HDL/LDL ratios obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 10. Fasting blood atherogenic index levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 11. Fasting blood AST levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 12. Fasting blood ALT levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD). Figure 13. Fasting blood ALP levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 14. Creatinine levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 15. Fasting blood Total Protein levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet. 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01 : Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD).

Figure 16. Fasting blood Albumin levels obtained in the groups at the end of the experimental period (SK: Healthy rat, Normal diet; KZ10: Healthy rat, Normal diet, 10 ml NO extract; YD: Healthy rat; High fat diet (YYD), Z01: Diabetic rat, YYD, 0.1 ml NO extract; Z1 : Diabetic rat, YYD, 1 ml NO extract; Z10: Diabetic rat, YYD, 10 ml NO extract; DK: Diabetic rat, YYD; GK: Glibenclamide (0.6 mg/kg, bw), Diabetic rat, YYD; AT: Atorvastatin (25 mg/kg, bw), Diabetic rat, YYD; FF: Fenofibrate (30 mg/kg, bw), Diabetic rat, YYD). Tables;

Table. 1 High fat feed composition

Detailed Description of the Invention

The invention relates to; the use and obtaining of Nerium oleander composition for the control of high blood glucose levels manifested with Type Il diabetes, obesity and cardiovascular diseases, the control of cholesterol and triglyceride levels, and the modulation of HDL-LDL ratios.

Regarding the research conducted; with the use of NO (Nerium oleander) extract in the trial on Type Il diabetic rats, fed with a high fat diet where 58 % of the metabolic energy was provided by animal fat content, significant positive developments were obtained in the blood glucose, triglyceride, cholesterol, HbAIc and HDL/LDL levels, which are primary clinical and laboratory findings. Loss of body weight was prevented in the groups on which the NO extract was administered. A more interesting finding is the occurrence of body weight loss in the KZ10 group, which was the group in which the highest dose of NO extract was used on healthy animals, and in this sense the observation that elements relating to the achievability of an important advantage in obesity control was made. When the difficulties of administration of the classical treatment, its side effect incidence and cost factors are taken into consideration, it can be contrived that, even with the present form of the NO extract, a possible alternative treatment protocol can be propounded.

The stage of in vitro evaluation of the biological material obtained from the experimental part of the research has been initiated and currently necessary practices are in progress.

The findings obtained in the process of the research until now show that the product can be used in its present form, for the purpose of keeping the lipids profile and blood glucose under control within the scope supportive treatment, in patients with Type Il diabetes. Particularly the positive developments obtained on the lipids profile display the impression that the product may also be helpful in obesity cases. The findings obtained show that the product, as well as being used in its present form as a "magistral drug" in the areas briefly described above, may also be converted to the "specialty" form by pursuing the analysis of active substance.

In the research, the rats (Sprague-Dawley, 3-4 months, male) obtained from the Experimental Research Center of the Akdeniz University Faculty of Medicine were used. After necessary healthy controls of the animals were completed, the method proposed by Srinivasan et. al. (²⁷) was carried out, in order to form an experimental type Il diabetes model. In this method, after the animals were fed for two weeks with a high fat diet, in which 58 % of the metabolic energy is provided from animal fat content, and the composition of which is stated below, by administering a low dose Streptozotocin (STZ, 35 mg/kg, IP), those animals with a postprandial blood glucose level of 300 mg/dl> were accepted to be Type Il diabetic.

Table. 1 High fat feed composition

NUTRITIONAL

FEED TO BE GIVEN % Ratio SUBSTANCE RATION

Vegetable oil 3.00 KM,% 94.4

Wheat flour, bonkalite - HP,% 22.3

Animal fat 37.00 ME,kcal/kg 5387

Corn, yellow 30.50 Ca, % 1.16

Casein, dried 20.00 P,% 0.62

Soya pulp, 48% 4.50 Na,% 0.21

Dicalcium phosphate 1.70 Met+Sis,% 0.94

Dl-methionine 0.20 Lysine, % 1.81

Lime stone 1.60 HS,% 0.89

Salt 0.50 HY,% 40.34

Vitamin-mineral 1.00 Linoleic a.% 3.40 A. Control Groups:

1. Healthy Control-Normal Diet (SK): Was formed with healthy rats with suitable lipid profiles and were not subjected to any process. They were fed with normal diet throughout the duration of the experiment.

2. Healthy Control-High Fat Diet (YD): Was formed with healthy rats with suitable lipid profiles and were not subjected to any process. They were fed with the high fat diet throughout the duration of the experiment.

3. Healthy Control-Nerium oleander (KZ10): Was formed with healthy rats with suitable lipid profiles and were not subjected to any process. The maximum amount of active substance, which is administered in the test group, was administered on the animals throughout the duration of the experiment. They were fed with normal diet throughout the duration of the experiment.

4. Negative Control (DK): Rats with the experimental Type Il diabetes model formed were used. The animals were not subjected to any process. They were fed with the high fat diet throughout the duration of the experiment.

5. Positive Control (GK): Rats with the experimental Type Il diabetes model was formed were used. Glibenclamide (0.6mg/kg bw) were administered once a day by gavage to the animals. They were fed with the high fat diet throughout the duration of the experiment.

B. Test Groups: Appropriate dose levels were obtained by condensing the oleander extract 10 times through lyophilization. Test groups were administered lyophilized material corresponding to 0.1 , 1 and 10 ml of extract. They were fed with the high fat diet throughout the duration of the experiment.

1. Nerium oleander 0.1ml (Z-0.1): Rats with the experimental Type Il diabetes model formed were used. The animals were administered, once a day by gavage, the lyophilized material corresponding to 0.1 ml of the original oleander extract by diluting with 0.5 ml of physiological saline.

2. Nerium oleander 1ml (Z-1): Rats with the experimental Type Il diabetes model formed were used. The animals were administered, once a day by gavage, the lyophilized material corresponding to 1 ml of the original oleander extract by diluting with 0.5 ml of physiological saline.

3. Nerium oleander 10 ml (Z-10): Rats with the experimental Type Il diabetes model formed were used. The animals were administered, once a day by gavage, the lyophilized material corresponding to 10 ml of the original oleander extract by diluting with 0.5 ml of physiological saline.

In view of enabling a healthier comparison of the results obtained, two additional test groups were formed by administering atorvastatin (AT, 25 mg/kg) and fenofibrate (FF, 30 mg/kg) to those rats in which the experimental type Il diabetes model was formed and which were subjected to the diet described above throughout the experiment. In the AT and FF groups rats in which the experimental type Il diabetes model was formed were used. They were fed with the high fat diet throughout the duration of the experiment. Stated active substances were administered once a day by gavage.

Blood and urine samples were taken from the animals in the test groups at 15 day intervals. Fasting Blood Glucose, Total cholesterol, Triglyceride, HDL, LDL, Creatinine, Urea, Total Protein, Albumin, HbAIc, Amylase, Lipase, CK-MB, ALT, AST, ALP, Na, Ca, and Phosphate levels were measured in blood samples. In the urine, on the other hand, Creatinine, Na, K, Cl, Ca, GGT, Amylase, Total Protein, and Glucose levels were determined.

The subjects in all test groups were euthanized under general anesthesia subsequent to the final sampling time after the experiment with a duration of three months. Liver, muscle, fat tissue, pancreas and blood samples were taken and kept under required conditions. Studies relating to insulin, leptin and IL-1 in the blood samples taken, and relating to caspase-3, tissue cytokine level and tissue mRNA expressions in the tissue samples taken are still ongoing.

The first stage of the diabetic predisposition subsequent to the administration of high fat diet is the increase in lipid stores related to consumption of foods with a high fat ratio, and the impairment of metabolic balance in subsequent stages. The increased lipid stores cause defense mechanisms to be activated in the organism and, lead to insulin resistance in time. During this stage which is defined as pre-diabetes, hyperinsulinemia is formed in order to withdraw the hyperglycemia formed within normal limits. Consequently, these developments result in the development of type Il diabetes which is a disease characterized by hypoinsulinemia and/or insulin resistance due to the damage or functional deficiency on pancreas cells. The metabolic response of insulin deficiency and/or receptor resistance, on the other hand, is obtaining energy from stored fats due to inadequate glucose intake by the cells and/or failures in the storage of energy sources in related cells as triglycerides, raised lipid levels in blood (dyslipidemia) hyperglycemia with accompanying atherosclerosis and cardiovascular diseases, seen in relation with the producing of LDL molecules in smaller molecules, and additionally, secondary complications such as nephropathy and neuropathy.

In the study it was observed that; at the end of trial, the fasting blood glucose levels of rats which were fed with high fat diet during the 3 months test period following the formation of experimental type Il diabetes, and which were administered 3 different doses of oleander (NO-Nerium oleander) extract (Z-01 , Z- 1 and Z-10) by daily gavage, were significantly lower than the fasting blood glucose levels of rats , with type Il diabetes formed, and which were fed under the same conditions, in the diabetes control (DK) group which were not subjected to any process, and than those rats in the (GK, AT and FF) groups in which anti-diabetic, anti-lipidemic, anti- cholesterolemic other substances were administered (Figure 1 ). The most important result in terms of the fasting below glucose is that, the level in the groups which were administered oleander, exhibited a level which was close to the data from the healthy rats fed with normal diet, and that this condition was confirmed with the HbAIc levels obtained from the above mentioned groups (Figure 2).

The fact that the fasting blood sugar level, in the group with healthy rats which were administered the highest dose of NO extract (KZ10), was reduced quantitatively as compared to the level of healthy rats (SK), was noted as another important finding which reflected the effect of the NO extract in reducing the level of blood glucose. The effect on healthy rats (YD), of administering a fatty diet, in forming a predisposition for type Il diabetes was also confirmed by the elevated blood glucose data observed in this group (Figure 1 ) While in the diabetes control (DK) group a noteworthy numerical increase in triglyceride levels was observed, no significant differences among groups developed. While thriglyceride values obtained from especially the atorvastatin (AT) and glibenclamide (GK) administered groups were close to DK, the most positive contribution was observed with the administration of fenofibrate (FF). The trigilyceride levels were clearly reduced in NO groups gradually and dose dependent, but the reduction in the level was not sitatisticaly significant.

The positive effect of NO extract on the triglyceride level was clearly observed in the KZ-10 group, by the lower levels of triglyceride as compared to SK (Figure 3).

Important changes in cholesterol levels were observed in the experimental groups which were administered NO. Particularly in DK, the cholesterol which increased due to the effect of diabetes, could not be prevented by administering AT and FF. The most noteworthy effect among the drugs used as a positive control was obtained with GK. While the cholesterol levels formed in the groups administered NO extract displayed a significant similarity with SK, they were found to be significantly lower than DK; additionally, a cleared positive effect was determined in NO groups as compared to all three drugs (AT, FF, GK). As with the parameters stated above, the fact that the cholesterol level obtained in group KZ-10 was numerically lower than SK was noted as a striking finding (Figure 4,5).

In addition to all these positive developments, the increase in body weight, in the groups administered NO extract (Z-01 , Z-1 and Z-10) as accompanied to the high fat diet in the period following the formation of experimental diabetes, is another important matter which needs emphasis (Figure 6). The contribution of a high fat diet in weight increase is a known condition (YD); the obesity which developed in relation with this practice is among the first concerns in diabetes etiology. The fact that, while body weight was significantly reduced in DK, in groups which were administered NO extract an increase in body weight was recorded may give rise to the view that, together with the positive contribution formed on the lipid profile, NO may be effective in the storage of blood lipids in fat cells subsequent to an insulin-like effect profile and afterwards it may prevent cardiovascular complications which may develop in relation with dyslipidemia. When the HDL and LDL cholesterol levels are examined, it was seen that HDL increased subsequent to the administration of NO extract, as compared to SK, and that LDL followed a development close to SK. Negative developments on HDL and LDL values observed with diabetes markedly decreased with the administration of NO extract. The increase in the HDL value in DK is caused by the existing high total cholesterol level. It was seen that, atorvastatin with anti-cholesterolemic effect, and fenofibrate with anti-lipidemic effect, were not as effective in reducing the amount of LDL in diabetics who have a high fat diet, as the 1 ml and 10 ml (Z- 1 and Z-10) doses of NO. The positive effect of the NO administration in increasing HDL level and decreasing LDL level, which are very important in terms of reducing cardiovascular diseases, was also confirmed with. the evaluation of HDL/LDL ratios in the groups (Figure 7, 8, 9).

In individuals with type Il diabetes, metabolic syndrome, and the combined dyslipidemia, cardiovascular risk is increased by a clustering of risk factors such as abdominal obesity, impaired fasting glucose, increased blood pressure, low HDL- cholesterol, increased triglycerides, and an increase in small, dense LDL particles. Although insulin resistance is crucial to the pathogenesis of the disease, the associated atherogenic lipoprotein phenotype considerably enhances the risk. Hence there is an ongoing intense search for a medication capable of modifying the atherogenic lipid profile as well as lowering glucose (²⁸). Insulin resistance is often associated with increased triglyceride and decreased HDL-cholesterol concentrations and increased small LDL particles. The Atherogenic Index has recently been proposed as a marker of plasma atherogenicity because it is increased in people at higher risk for coronary heart disease and is inversely correlated with LDL particle size (²⁹). For this purpose, atherogenic index were calculated using the appropriate formula (Atherogenic index (Al): ([Total Cholesterol] - [HDL-Cholesterol])/[HDL- Cholesterol] ) as others explained (³⁰). NO extract was found to be an effective against atherogenic process in Z01 and Z1 groups but especially Z-10 concentration of NO extract showed statistically significant decreased atherogenic index when compared to DK, GK, AT and FF groups (Figure 10).

When liver enzymes were examined no noteworthy change among groups was observed other than the lower AST in the Z-10 group as compared to DK. ALT and ALP, on the other hand, were found to be high in the diabetic group as compared to the values of healthy rats. However, while in NO extract administered groups, no significant increases in said enzyme levels as compared to healthy rats (SK) were observed, subsequent to fenofibrate administration, marked increases in ALT, and in particular, in ALP values were recorded. These changes are important in exhibiting that, NO extract does not have a negative effect in terms of liver enzymes (Figure 11 , 12, 13)

While elevated creatinine level, which defines muscle wasting and/or kidney filtration failure, was found to be the highest with DK, the levels of other diabetic groups were found to be close to healthy control data. Additionally, the creatinine value which was nearest the value of rats fed with normal diet was obtained in the Z-10 group (Figure 14).

Because increased total protein and albumin levels, which could be an indication of dehydration, developed similar to healthy rats in groups administered with NO extract, it was seen that the negative effect for diabetes could be reduced in these groups with the effect of oleander (Figure 15, 16).

Because in alternative extraction application performed by using oleander, cold water and alcohol are used, substances that dissolve in water and alcohol and heat sensitive glycosides (oleandrine, oleandigenine etc.) are including in the composition of the extract. Cold extract comprises more glycosides and flavonoides and less polysaccharides as compared to hot extract. Naturally for the hot extraction process, the opposite of this expression is also applicable. Because less glycosides and flavonoides, however, more polysaccharides shall be present during the obtaining of the oleander extract which is the subject matter of this patent application, the risk of cardiotoxicity which may develop in relation with these substances is lesser. Obtaining the Oleander Extract:

The following process steps are used in obtaining the oleander (Nerium oleander) extract:

- Collection of the oleander plant in March-September period from among new shoots,

- Cleaning by washing the plant collected in an adequate amount,

- Cutting the fresh shoots collected into small pieces of approximately 1-3 cm length subsequent to the washing process,

- Obtaining a mixture by adding approximately 400-700 gr (preferably 500 gr) of chopped oleanders in 5-10 It (preferably 8 It) distilled water,

- Placing the mixture in a heat resistant container and applying heat to this mixture for approximately 30-50 min (preferably 40 min),

- Subsequent to the heat application on the mixture, without covering the lid of the container until the liquid starts to evaporate, and after the evaporation starts, by closing the system and collecting the vapor formed in a separate container by causing it to come in contact with a surface cooled with cold water, the process is continued for a period of 3-6 hours (preferably 5 hours). Care is taken that the mixture in the container is not burned.

- Keeping the obtained Nerium oleander extract in a clean glass container at room temperature until use.

The Nerium oleander extract obtained is used in the following areas:

— the use for autoimmune diseases and in pathologies manifested with proinflammatory cytokine activation in oral, IV, IM, intra-articular form or as food additive,

— the local use for ulcerations, inflammations where inflammation is dominant and/or is the inducer,

— particularly the local use for skin diseases such as dermatitis, eczema, psoriasis advancing with dysregulation in cell proliferation and increased inflammation,

— the oral use of the magistral and specialty forms of Nerium oleander extract,

— the oral spray use of the magistral and specialty forms,

— the nasal spray use of the magistral and specialty forms,

— the rectal use of the magistral and specialty forms,

— the intravenous use of the magistral and specialty forms,

— the intramuscular use of the magistral and specialty forms,

— the subcutaneous use of the magistral and specialty forms,

— the use in treatment of type Il diabetes and obesity,

— the use in the control of high blood glucose levels seen in patients with type Il diabetes,

— the use in the control of cholesterol and triglyceride levels,

— the use in the modulation of HDL-LDL ratios,

— the use in the in patients with type Il diabetes as supportive treatment for keeping the lipids profile and blood glucose under control, — the use in the control of high HbAIc levels seen in patients with type Il diabetes,

— the use in patients with Type Il diabetes as supportive treatment for the purpose of keeping high HbAIc levels under control

— the use in the prevention of loss of body weight in patients with Type Il diabetes,

— the use in enabling of loss of body weight in healthy individuals,

— its conversion into "specialty" form,

— the use as a food additive.

— the use of Nerium oleander extract in the control and modulation of leptin levels in Type Il diabetes and obesity for the treatment and prophylactic purposes.

— the use of Nerium oleander exctract in the supportive treatment for the purpose of keeping high lipid profiles under control in patients with cardiovascular diseases, with some diseases related with dyslipidemia and in obesity.

The scope of protection of this application is stated in the claims section and cannot be limited to those described above, given absolutely for the sake of example. It is evident that a person specialized in the technology can produce the innovation introduced with the invention by altering the parts in form and/or apply this embodiment to other fields with same purposes in the related technology. Therefore it is also evident that such embodiments shall be bereft of the innovation criterion. References:

1. Pushparaj et. al., 2000,

2. Maries and Farnsworth, 1995,

3. Bozan et. al., 1997; Erol et. al., 1997; Ozbek, 2002, 4. Akev et. al., 1991, Kavalah et. al., 1998, Ozbek et. al., 2002,

5. Smith et. al., 2001, Erdemoglu et. al.2003,

6. Manjunath, 1966, Nadkarni, 1976, Nasir and AIi, 1982, Begum et. al.1999, Zia et. al.1995,

7. Siddiqui et. al.1995, Begum et. al.1999; Zia et. al.1995, 8. Mulleret. al., 1991,

9. Siddiqui et. al..1997,

10.Huqet. al., 1999,

H.Fu et. al.2005,

12.Zhaoet. al.2006, 13.Afaqet. al.,2004,

14.Yesilada, 2002,

15. Erdemoglu et. al.2003,

16.Adamet. al.2001,

17. Erdemoglu et. al.2003, 18.Ceteet. al., 2005,

19.Ni et. al.,2002,

20. Mekhail et. al., 2006,

21.Haebaet. al.,2002,

22.Adaet. al.2001, 23.Kavalahet. al..1998,

24. Ozbek et. al..2002,

25.Bellakhdaret. al..1991

26. Jouad et. al..2001, Bnouham et. al.2002, Eddouks et. al..2002, Tahraoui et. al..2006 27.Srinivasan et. al.2005.

28.Dobiasova2004.

29.Tanetal 2004.

30. Coetzee et al 2007, Jeon et al 2005, Yamajaki and Murata 1994.

Claims

1. The invention is a Nerium extract, wherein, it is to be used regulation of lipid metabolism in diseases manifested with type Il diabetes, obesity, high cholesterol and triglyceride levels, and cardiovascular diseases.

2. A composition according to Claim 1 , wherein, said Nerium is Nerium oleander.

3. The invention is a method of obtaining Nerium extract according to Claim 1 , wherein, the Nerium to be extracted with purpose in Claim 1 is extracted by cold and/or hot extraction.

4. A method according to Claim 3, wherein, said nerium is Nerium oleander.

5. A method of obtaining Nerium extract according to Claims 3 and 4, wherein, it comprises the process steps of;

- cleaning by washing the Nerium oleanders collected,

- subsequent to the washing process cutting the Nerium oleanders into small pieces,

- obtaining a mixture by adding chopped Nerium oleanders in distilled water,

- placing the mixture in a container and applying heat to said mixture,

- Subsequent to the heat application on the mixture, without covering the lid of the container until the liquid starts to evaporate, and after the evaporation starts, by closing the system and collecting the vapor formed in a separate container by causing it to come in contact with a surface cooled with cold water.

6. A method according to any one of the preceding Claims, wherein, by using hot extraction, the extract obtained includes less glycosides and flavonoides, however, more polysaccharides as compared to that obtained by cold extraction,

7. A method according to any one of the preceding Claims, wherein, due to the fact that the extract obtained includes less glycosides and flavonoides, however, more polysaccharides it does not pose risk of cardiotoxicity.

8. A method according to any one of the preceding Claims, wherein, the Nerium oleanders collected, are collected from among new shoots during the March- September period.

9. A method according to any one of the preceding Claims, wherein, subsequent to the washing process the Nerium oleanders are cut into small pieces of approximately 1-3 cm length.

10. A method according to any one of the preceding Claims, wherein, the Nerium oleanders are cut up in an amount of approximately 400-700 gr (preferably 500 gr).

11. A method according to any one of the preceding Claims, wherein, a mixture is obtained by adding the chopped Nerium oleanders in 5-10 It (preferably 8 It) distilled water.

12. A method according to any one of the preceding Claims, wherein, the mixture is placed in a heat resistant container and heat is applied to this mixture for approximately 30-50 min (preferably 40 min).

13. A method according to any one of the preceding Claims, wherein, the process of collecting the vapor formed subsequent to the application of heat to the mixture, in a separate container by causing it to come in contact with a surface cooled with cold water is performed for a period of 3-6 hours (preferably 5 hours), taking care not to burn the mixture.

14. A method according to any one of the preceding Claims, wherein, the Nerium oleander extract obtained is kept in a clean glass container at room temperature until use.

15. The invention relates to the use of nerium extract according to Claim 1 , which it is obtained by a method according to Claims 3 to 14.

16. A use according to Claim 15, wherein, said nerium is Nerium oleander.

17. A use according to any one of the preceding Claims, wherein, the magistral and specialty form of said Nerium oleander extract is used orally.

18. A use according to any one of the preceding Claims, wherein, the magistral and specialty form of said nerium oleander extract is used rectally.

19. A use according to any one of the preceding Claims, wherein, the magistral and specialty form of said nerium oleander extract is used intravenously.

20. A use according to any one of the preceding Claims, wherein, the magistral and specialty form of said nerium oleander extract is used intramuscularly.

21. A use according to any one of the preceding Claims, wherein, the magistral and specialty form of said nerium oleander extract is used subcutaneously.

22.A use according to any one of the preceding Claims, wherein, the magistral and specialty form of said nerium oleander extract is used nasal spray.

23.A use according to any one of the preceding Claims, wherein, the magistral and specialty form of said nerium oleander extract is used oral spray.

24.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used in the treatment of Type Il diabetes and obesity.

25.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used in the control of high blood glucose levels found in patients with Type Il diabetes.

26.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used in the control of cholesterol and triglyceride levels.

27.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used the modulation of HDL-LDL ratios.

28.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used in patients with Type Il diabetes as supportive treatment for keeping the lipids profile and blood glucose under control.

29.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract can be converted to "specialty" form.

30. A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used as a food additive.

31. A use according to any one of the preceding Claims, wherein, said Nerium oleander extract can be used in diabetes and especially in ethiopathogenesis as an immunomodulator, by affecting the regulation of release of proinflammatory cytokines (IL-I , TNF-alpha etc.) in atherosclerotic cases, other cardiovascular pathologies and inflammatory processes where inflammation is present.

32.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used for autoimmune diseases and in pathologies manifested with proinflammatory cytokine activation in oral, IV, IM, intra-articular form or as food additive.

33.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used locally for ulcerations, inflammations where inflammation is dominant and/or is the inducer.

34.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used in particular locally for skin diseases such as dermatitis, eczema, psoriasis advancing with dysregulation in cell proliferation and increased inflammation.

35.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used in the control of high HbAIc levels seen in Type Il diabetes patients.

36.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used in patients with Type Il diabetes as supportive treatment for the purpose of keeping high HbAIc levels under control.

37.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used in the prevention of loss of body weight in patients with Type Il diabetes.

38.A use according to any one of the preceding Claims, wherein, said Nerium oleander extract is used in providing loss of body weight in healthy individuals.

39. A use according to any one of the preceding Claims, wherein, the use of Nerium oleander extract in the control and modulation of leptin levels in Type Il diabetes and obesity for the treatment and prophylactic purposes.

40. A use according to any one of the preceding Claims, wherein, the use of Nerium oleander exctract in the supportive treatment for the purpose of keeping high lipid profiles under control in patients with cardiovascular diseases, with some diseases related with dyslipidemia and in obesity.