FIELD OF THE INVENTION

The present invention pertains to nutritional or pharmaceutical compositions comprising extracts or concentrates of plants and the mixtures thereof belonging to Lagerstroemia sp. with specific reference to Lagerstroemia speciosa (Banaba). The present invention further relates to extracts which are isolated from different parts of Lagerstroemia speciosa plant, the preparation of such extracts and the medicaments containing said extracts. The invention further relates to screening and characterization of extracts for their activity in maintaining body weight and lowering high levels of blood sugar to achieve the lowest possible risk. Furthermore, the invention relates to the use of the extracts as a supplement or a medicament useful for Calorie management and Diabetes.

BACKGROUND AND PRIOR ART OF THE INVENTION

Obesity is a condition of abnormal or excessive calorie (fat) accumulation in adipose tissue to the extent that health may be impaired. It is difficult to measure body fat directly but measurements of relative weight for height, such as the Body Mass Index (BMI) will indicate overweight and obesity in adults. The World Health Organization (WHO) has recognized obesity as an international health problem of staggering magnitude.

Diabetes is a metabolic disorder where in human body does not produce or properly uses insulin, a hormone that is required to convert sugar, starches, and other food into energy. Diabetes mellitus is characterized by constant high levels of blood glucose (sugar).
Human body has to maintain the blood glucose level at a very narrow range, which is done with insulin and glucagon. The function of glucagon is to release glucose from the liver to the bloodstream so that, it can be transported to body tissues and cells for the production of energy.

A. Obesity

Food is a solid or liquid material upon ingestion by higher animals provides all necessary energy and building blocks for growth and development, without harming the animals in anyway. The energy (calorie) in the food is stored in the form of carbohydrates and fats.

Digestion of the food in the gut yields calories and are used for daily functions such as breathing, digestion, and physical activities. In normal healthy people, the number of calories consumed is approximately equal to the number of calories used (activities) and any imbalance will lead to calorie storage or accumulation. Weight gain is the result of either extra calorie consumption or decreased calories used (reduced physical activity) or the both (British Nutrition Foundation 2004).

Etiology: The main factors responsible for the energy imbalance are lifestyle and environmental factors. Limiting the absorption of calories in the digestive track or limiting the number of calorie storage depots or both can address the condition of energy imbalance. Inhibiting the enzymes involved carbohydrate and fat metabolism in intestine and limiting the differentiation of pre-adipocytes will achieve this. Differentiated adipocytes acts as depots for the storage of fats.

Treatment: Two principal risk factors that lead to obesity, poor nutrition and lack of physical activity combined. Exercise increases glucose transport whereas in sedentary states there is less glucose transport. A lack of glucose transport in fat cells allows them to mature, resulting in greater body fat. Glucose transport helps to regulate intracellular glucose levels which in turn help to maintain healthy blood glucose, in part because of the greater energy provided to muscle cells which also get their energy from the transport of glucose (Health Encyclopedia - Diseases and Conditions).

B. Diabetes

Since insulin is the principal hormone that regulates uptake of glucose into most cells from the blood (primarily muscle and fat cells, but not central nervous system cells), deficiency of insulin or the insensitivity of its receptors plays a central role in all forms of diabetes mellitus. Insulin is released into the blood by beta cells (P-cells) in the pancreas in response to rising levels of blood glucose (e.g., after a meal). Insulin enables most body cells (about 2/3 is the usual estimate, including muscle cells and adipose tissue) to absorb glucose from the blood for use as fuel, for conversion to other needed molecules, or for storage. Insulin is also the principal control signal for conversion of glucose (the basic sugar used for fuel) to glycogen for internal storage in liver and muscle cells.

Reduced insulin levels result both in the reduced release of insulin from the beta cells and in the reverse conversion of glycogen to glucose when glucose levels fall, although only glucose thus recovered by the liver re-enters the bloodstream as muscle cells lack the necessary export mechanism (Encyclopedia Britannica, Inc).

Etiology : Type 1 diabetes mellitus - formerly known as insulin-dependent diabetes (IDDM), childhood diabetes, or juvenile-onset diabetes - is characterized by loss of the insulin-producing beta cells of the islets of Langerhans of the pancreas leading to a deficiency of insulin. The most common cause of beta cell loss leading to type 1 diabetes is autoimmune destruction, accompanied by antibodies directed against insulin and islet cell proteins. Type 2 diabetes mellitus - previously known as adult-onset diabetes, maturity-onset diabetes, or non-insulin dependent diabetes mellitus (NIDDM) - is due to a combination of defective insulin secretion and defective responsiveness to insulin (often termed insulin resistance or reduced insulin sensitivity), almost certainly involving the insulin receptor in cell membranes. In early stages, the predominant abnormality is reduced insulin sensitivity, characterized by elevated levels of insulin in the blood. In the early stages, hyperglycemia can be reversed by a variety of measures and medications that improve insulin sensitivity or reduce glucose production by the liver, but as the disease progresses the impairment of insulin secretion worsens, and therapeutic replacement of insulin often becomes necessary. Other factors may include aging and family history, although in the last decade it has increasingly begun to affect children and adolescents. Type 2 diabetes may go unnoticed for years in a patient before diagnosis, since the symptoms are typically milder (e.g. lack of ketoacidotic episodes) and can be sporadic. However, severe complications can result from unnoticed type 2 diabetes, including renal failure, vascular disease (including coronary artery disease), vision damage, etc.Gestational diabetes also involves a combination of inadequate insulin secretion and responsiveness, resembling type 2 diabetes in several respects. It develops during pregnancy and may improve or disappear after delivery. Even though it may be transient, gestational diabetes may damage the health of the fetus or mother, and about 20%-50% of women with gestational diabetes develop type 2 diabetes later in life.

Gestational diabetes mellitus occurs in about 2%-5% of all pregnancies. It is temporary, and fully treatable, but, if untreated, may cause problems with the pregnancy, including macrosomia (high birth weight) of the child. Fetal/Neonatal risks associated with GDM include congenital anomalies such as cardiac, CNS, and skeletal muscle malformations. Increased fetal insulin may inhibit fetal surfactant production and cause respritory distress syndrome. Hyperbilirubinemia may result from RBC destruction. In severe cases perinatal death may occur, most commonly as a result of poor placental profusion due to vascular impairment.

Treatment: Type 1 treatment must be continued indefinitely at present. Treatment does not impair normal activities, if sufficient awareness, appropriate care, and discipline in testing and medication.Type 2 diabetes is usually first treated by changes in physical activity (usually increase), diet (generally decrease carbohydrate intake, especially glucose generating carbohydrates), and through weight loss. There are several rare causes of diabetes mellitus that do not fit into type 1, type 2, or gestational diabetes:

• Genetic defects in beta cells (autosomal or mitochondrial)

• Genetically-related insulin resistance, with or without lipodystrophy (abnormal body fat deposition)

• Diseases of the pancreas (e.g. chronic pancreatitis, cystic fibrosis)

• Hormonal defects

• Chemicals or drugs

The inclusion of botanicals in a nutritional approach presents an inexpensive means of achieving the goal of cost effective management of the said disease condition. However, the effects of the nutritional strategies recommended today are rather modest. Thus, research into novel nutritional strategies preventing Calorie management and diabetes is needed.

The management of calorie/weight and diabetes is a particularly promising area for botanicals. Most botanicals derive their effectiveness from a mixture of active molecules, acting in concert. Multiple agents attacking multiple targets simultaneously present decided advantages over conventional drugs, which are each based on one compound that produces one action. Plants hold the power to keep the increasing prevalence of diabetes and calorie management, in check, prompting the search and trial of plant extracts to develop a whole new category of natural products. Comprehensive studies on the components of the herb that are responsible for certain indications need to be undertaken to obtain effective medications from this therapy. Also, not necessarily all the ayurvedic extracts are non-toxic.

The botanical compound Banaba (Lagerstroemia speciosa L.) also has shown positive effects on the calorie management and diabetes, as its ellagitannins appear able to increase glucose uptake by adipocytes, lowering blood glucose levels. In addition, banaba extract may increase the expression of PPAR-gamma, a transcription factor that regulates the development of adipocytes.

Banaba (Lagerstroemia speciosa) been traditionally used in India and by the people of the
Philippines for maintaining healthy blood sugar. Garcia F.1941 found that although feeding extracts of the green leaves lowered high levels of blood sugar in rabbits, the fully mature or old leaves, which are red in color, held the greatest potency. Miyaji N, et al 1999 studies also support the traditional use of the banaba for maintaining healthy blood sugar. When healthy rats with normal blood sugar levels were orally administered a powder extract of the green leaves for 11 days before a single large dose of glucose, the rise in blood sugar was significantly suppressed. In another test by Suzuki Y, et al.2001, blood glucose was significantly lowered when extracts of the leaves were orally administered to normal rats after feeding them high amounts of carbohydrates (soluble starch) to raise their blood glucose levels. In another study by Chanda S, et al, 2005 normal rats of either sex were orally administered an extract of the fully mature, red leaves containing 20% tannins (Renaissance Herbs, Inc.) at the equivalent human dose of 500 mg twice a day (morning and afternoon) for 15 days. At the end of the study, the average blood sugar level of the rats was 29% lower than the control group. Administered a large, single oral dose of sucrose, the increase in blood sugar 24 hours later was significantly inhibited by 64.5%. Further evidence that banaba helps to maintain healthy blood sugar is found from significant improvements in blood sugar, both in animals (Miyaji N, et all999, Mishra Y, et al .1990 and Kakuda T, et al.1996) and people with type 2 diabetes (Ikeda Y, et al.1999 and Judy WV, et al.2003).

Adipogenesis, the differentiation and proliferation of adipocytes, is a major mechanism leading to weight gain and obesity. It is highly desirable to develop pharmaceuticals and treatments for T2D that reduce blood glucose levels without inducing adipogenesis in patients. Using glucose uptake assays and Western/Northern blot analyses as major tools and 3T3-L1 cells as a model, In 2005, Chen et al showed that the banaba extract inhibited the expression of key genes for adipogenesis and the quantitative assays were significant (P < 0.05). During 2004, Hong and Maeng studied the effects of malted barley extract and banaba extract on blood glucose levels in genetically diabetic mice. The glucose-6-phosphatase activity in kidney was significantly lower in both the MBE and banaba groups compared with the control group (PO.05), but there was no significant difference between the MBE and banaba groups. In another study Suzuki et al., in 1999 examined the anti-obesity potency of dietary banaba extract vising five-week-old female KK-Ay mice for 12 weeks. The mice fed banaba extract showed a significant decrease, to 65% of the control level in total hepatic lipid contents. This decrease was due to a reduction in the accumulation of triglyceride. These results suggest that banaba had a beneficial effect on obese female KK-Ay mice.

The inhibition of weight gain by banaba may be partly attributed to its ability to inhibit the in vitro activity of enzymes (Alpha-glucosidase and Alpha-amylase) involved in the digestion of starches and sugars (carbohydrates) and that allow increased levels of sugar in the blood following the digestion of carbohydrates (Suzuki Y, et al 2001 and Hosoyama H, et al 2003). Banaba also inhibits the maturation of fat cells, (Liu F, et al. 2001) an effect produced by tannins in the leaves. 15 A particular group of tannins, known as ellagitannins, also enhance glucose transport in fat cells (Hayashi T, et al.2002 and Hattori K, et al.2003) Among the ellagitannins in banaba, lagerstroemin ('lager-streemin') dose dependently activated glucose transport, (Hayashi T, et al.2002) and is found in higher amounts in the red, fully mature leaves than the green leaves.

This background information is provided for the purpose of making known information believed by the applicant to be of possible relevance to the present invention. No admission is necessarily intended, nor should be construed, that any of the preceding information constitutes prior art against the present invention.

OBJECTIVES OF THE PRESENT INVENTION

The main objective of the present invention is to obtain an extract of Lagerstroemia species optionally along with pharmaceutically acceptable additive.

Another main objective of the present invention is to develop a process for obtaining extract from Lagerstroemia species.

Still another main objective of the present invention is to develop a method for management of calorie/weight and diabetes in a subject in need thereof, said method comprising step of administering pharmaceutically acceptable amount of extract of Lagerstroemia species optionally along with pharmaceutically acceptable additives to the subject.

The principle object of the present invention is to provide an active standardized extract of banaba obtained from the leaves of Lagerstroemia speciosa plant.

Another object of the invention is to provide the reproducibility of plant material extractions from batch to batch variation of bioactive, which is used to ensure the quality of bioactive. The preparation of such extracts, evaluating bioenhancing/bioavailability of Lagerstroemia speciosa extract or bioactive fraction in combination with nutraceuticals or herbal drugs/products to evaluate the Lagerstroemia plant extracts, capable in Calorie/Weight management and Diabetes in more than one mode of action.

Yet another object of the invention is Cell-based assay to monitor the extract for their insulin mimicking activity and also synergistic activity at four different concentrations. The toxicity of the extract was also tested at all the concentrations.

Yet another object of the invention is to provide composition comprising active principles of Lagerstroemia speciosa, and the use of these extracts and constituents for the preparation of nutritional and nutraceutical application.

Still another object of the present invention is to provide Lagerstroemia speciosa plant extract, which is easily and safely administrable to children and adults.

SUMMARY OF THE INVENTION

The present invention relates to an extract of Lagerstroemia species optionally along with pharmaceutically acceptable additive; a process for obtaining extract from Lagerstroemia species, said process comprising steps of: (a) powdering plant material of Lagerstroemia species; (b) extracting and refluxing the powder with solvents to obtain extract; and (c) concentrating the extract followed by drying to obtain the extract of Lagerstroemia species and a method for management of calorie/weight and diabetes in a subject in need thereof, said method comprising step of administering pharmaceutically acceptable amount of extract of Lagerstroemia species optionally along with pharmaceutically acceptable additives to the subject.

BRIEF DESCRIPTION OF ACCOMPANYING DRAWINGS

Figure 1: HPLC chromatogram of Banopta 70% extract at 254nm

Figure 2: HPLC chromatogram of Banopta 20% extract at 254nm

Figure 3: Comparative overlay of HPLC Run of Banopta 70% extract from three
replicate run at 254nm

Figure 5:Comparative overlay of HPLC Run of Banopta 20% extract from three
replicate run at 254nm

Figure 6: Cytotoxic effect of Banopta 70% on tritiated glucose uptake in C2C12 cell
lines after 5 days differentiation.

Figure 7: Cytotoxic effect of Banopta 20% on tritiated glucose uptake in C2C12 cell lines
after 5 days differentiation.

Figure 8: Insulin mimicking effect of Banopta 70% on tritiated glucose uptake in C2C12
cell lines after 5 days differentiation.

Figure 9: Insulin mimicking effect of Banopta 20% on tritiated glucose uptake in C2C12
cell lines after 5 days differentiation.

Figure 10:Insulin synergistic effect of Banopta 70% on tritiated glucose uptake in
C2C12 cell lines after 5 days differentiation.

Figure 11: Insulin synergistic effect of Banopta 20% on tritiated glucose uptake in
C2C12 cell lines after 5 days differentiation.

Figure 12: Cytotoxic effect of Banopta 70% on tritiated glucose uptake in HepG2 cell
lines after 4 hrs of serum starvation.

Figurel3: Cytotoxic effect of Banopta 20% on tritiated glucose uptake in HepG2 cell
lines after 4 hrs of serum starvation.

Figure 14: Insulin mimicking effect of Banopta 70% on tritiated glucose uptake in
HepG2 cell lines after 4 hrs of serum starvation.

Figure 15: Insulin mimicking effect of Banopta 20% on tritiated glucose uptake in
HepG2 cell lines after 4 hrs of serum starvation.

Figure 16: Insulin synergistic activity of Banopta 70% on tritiated glucose uptake in
HepG2 cell lines after 4 hrs of serum starvation.

Figure 17:Insulin synergistic activity of Banopta 20% on tritiated glucose uptake in
HepG2 cell lines after 4 hrs of serum starvation.

DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to an extract of Lagerstroemia species optionally along with pharmaceutically acceptable additive.

In another embodiment of the present invention, said Lagerstroemia species is Lagerstroemia speciosa.

In yet another embodiment of the present invention, said additive is selected from a group comprising granulating agents, binding agents, lubricating agents, disintegrating agents, sweetening agents, coloring agents, flavoring agents, coating agents, plasticizers, preservatives, suspending agents, emulsifying agents and spheronization agents or a combination thereof.

In still another embodiment of the present invention, said extract is formulated as cosmetics and dosage forms selected from a group comprising tablet, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsion hard or soft gel capsules, syrups, elixirs.

The present invention also relates to a process for obtaining extract from Lagerstroemia species, said process comprising steps of:

(a) powdering plant material of Lagerstroemia species;

(b) extracting and refluxing the powder with solvents to obtain extract; and

(c) concentrating the extract followed by drying to obtain the extract of
Lagerstroemia species.

In another embodiment of the present invention, said Lagerstroemia species is Lagerstroemia speciosa.

In yet another embodiment of the present invention, said plant material is selected from a group comprising leaves, seeds, roots, stems, flowers, fruits, or various combinations thereof, preferably leaves.

In still another embodiment of the present invention, said extract is optionally centrifuged followed by lyophilization to obtain the extract of Lagerstroemia species.

In still another embodiment of the present invention, said solvents are selected from a group comprising water, alcohol and combinations thereof.

The present invention also relates to a method for management of calorie/weight and diabetes in a subject in need thereof, said method comprising step of administering pharmaceutically acceptable amount of extract of Lagerstroemia species optionally along with pharmaceutically acceptable additives to the subject.

In still another embodiment of the present invention, said subject is animal including human being.

Accordingly, the present invention provides Lagerstroemia speciosa plant extract for Calorie/ Weight management and Diabetes and/ or related disorders in a subject in need thereof, said method comprising step of administering pharmaceutically acceptable amount of standardized Lagerstroemia speciosa plant composite extract, optionally along with pharmaceutically acceptable additives, to the subject; and a process for enhancing calorie consumption and diabetes management of & Lagerstroemia speciosa plant extract, said process comprising steps of (a) size-reducing plant parts to obtain powder; (b) extracting the bioactives with a solvent and/or combination of solvents by heating at temperature ranging from 210 to 1050 C to obtain a mixture; (c) clarifying the mixture to arrive at clear liquid; (d) concentrating the clear liquid to achieve a concentrated extract; (e) solubilizing the concentrated extract in a solvent and re-concentrating it to obtain further concentrated extract, followed by drying the treated extract to obtain the plant bioactives; (f) the reproducibility of plant material extractions from batch to batch were tested using in-house metabolite fingerprinting QA/QC method based on HPLC equipped with PDA, which takes care the batch-to-batch variation of bioactives in more precise way and is used to ensure the quality of bioactives; (g) Cell Based Assays: Banopta 70% and 20% was tested for its efficacy in glucose uptake in two different cell lines; (C2C12) and (HepG2). The extract was monitored for their insulin mimicking activity and also synergistic activity at four different concentrations. The toxicity of the extract was also tested at all the concentrations. The invention further provides for uses of the extract to manufacture a medicament for multiple therapeutic uses, as well as other healthful benefits.

The present invention is in relation to efficiency of the bioactive component of the plant extract for therapeutic use, wherein said extract from Lagerstroemia speciosa optionally along with healthful or for nutritional and nutraceutical applications.

In one aspect of the invention, there is a provided a prophylactic method for preventing the occurrence of a disease state in a mammal which comprises administering to the said mammal an effective non-toxic amount of an extract from Lagerstroemia speciosa as defined herein in the preparation of a comestible (foodstuff) for prophylaxis on weight management and diabetes. Preferably the mammal is human and the said extract comprises a single extract from a plant part of Lagerstroemia speciosa or a combination of extracts there from as detailed herein. Thus the present invention further relates to extracts, which may be isolated from leaves of the Lagerstroemia speciosa plant, the preparation of such extracts, medicaments comprising such extracts, and the use of these extracts and constituents for the preparation of a medicament.

Some of the embodiments of the present invention will include pharmacognostic evaluation of the botanicals as per the conventional pharmacopoeial standards for ascertaining the quality, purity and efficacy of the herbs. These tests will comprise determination of physico-chemical standards like total ash, water and alcohol soluble extractives, foreign organic matter, moisture content and screening of the plant material for total microbial count (Total Bacterial Count and Total Fungal Count) in order to meet the most stringent quality regulations. Chromatographic finger printing of the herbs for their general profile or marker compounds using Thin Layer Chromatography will also form part of this study.

In another aspect of the present invention, extracts are isolated from seeds of the Lagerstroemia speciosa, using conventional inorganic and organic solvent extraction and supercritical fluid extraction technology. Generally, extracts of the invention capable of functioning in a prophylactic or therapeutic manner as outlined herein can be extracted from any Lagerstroemia speciosa plant, depending on the end purpose that is required of the extract.

In some of the embodiments of the present invention there is provided a process for preparing extracts of the invention from plant parts of Lagerstroemia speciosa that comprises:

a. Obtaining plant material from one or more parts of the plants of claim 1.

b. Obtaining an extract from the plant material by contacting the plant
material with an aqueous, an ethanolic or an organic solvent, or a
combination thereof, optionally for a defined period of time thereby
providing one or more plant extracts.

c. Removing the plant material from the supernatant obtained in step b.

d. Optionally, lyophilizing said supernatant.

e. Analyzing the plant extracts for efficacy and presence of inhibitory
activity for Weight management and Diabetes.

f. Selecting plant extracts having one or both of these activities.

The choice of selected plant material may be of any type but is preferably the leaves of the Lagerstroemia speciosa plant.

The solvent extraction process may be selected from direct types such as extraction from plant parts in reflux extractor apparatus or in flasks at room temperature or at higher temperature with polar and/or non-polar solvent(s). Typically, the extraction process is as outlined herein. In another embodiment of the invention, the compositions for preventing, treating, or managing Calorie and Diabetes, comprises of direct composite extract of plant species with alcohol, water and hydroalcohol solvent and successive extract with solvents from non-polar to polar range. The compositions/medicaments may contain a pharmaceutically acceptable carrier, excipient, or diluent.

It will be apparent to the skilled addressee that the selection of solvent, or mixtures of solvents for each step in the isolation of extracts of the invention showing activity can be guided by results of bioassay analysis of separate fractions.

Some of the embodiments of the invention will describe the HPLC profiles and Mass spectrums of direct and successive solvent extracts of 70% and 20% Lagerstroemia speciosa plant parts thereby giving each extract an identity of itself.

The plants selected for the isolation of therapeutically relevant extracts/molecules to be used in the calorie management and diabetes, will be subjected to both targeted and non-targeted screening procedures. The ongoing-targeted screening procedures, which feature a comprehensive metabolite profiling of multitudes of phytoextracts, are envisaged in the study to facilitate the creation of a metabolite grid.

The successive extraction from plant extract will be carried out using soxhlet extractor. The solvents used, will be based on their sequential polarity starting from non-polar to polar, wherein, various classes of metabolites will be extracted viz. petroleum ether (phytosterols, fixed oils and fats), benzene (fixed oils and fats), chloroform (alkaloids), acetone (phytosterols, phenolics and tannins) ethanol (alkaloids, carbohydrates, glycosides, phytosterols, saponins, phenolics, tannins, proteins and amino acids) and water (alkaloids, carbohydrates, glycosides, saponins, phenolics, tannins, proteins, amino acids, gums and mucilage) at 65°C. These fractions will be lyophilized and stored in amber colored bottles at 4°C.

Phytochemical investigations will be also carried out on these extracts using various tests like Mayer's and Dagendorf s tests for alkaloids; Molisch, Fehling and Benedict tests for carbohydrates; Lieberman Buchard's test for phytosterols and triterpenes; spot test for fixed oils and fats; Ferric chloride and Lead acetate test for phenolic compounds and tannins; Ninhydrin and Biuret tests for protein and aminoacids; alcoholic precipitation followed by Molisch test for gum and mucilages.

The extracted fractions will be subjected to HPLC using \i bondapak C i8 column (Waters Alliance 2695 Separation Module) to separate the constituent metabolites. The fractions are eluted using a combination (80:20, 60:40, 50:50, 40:60, 20:80) of methanol: water / acetonitrile: water. The gradient run will also be carried out wherever required. 5-l0ul of sample is injected with flow rate of 1 ml/min and HPLC run will be performed for 30 minutes. The detection is done on photodiode array and the analysis of the results will be done with the help of Millennium™ software.

The invention further describes the biotherapeutic potential of various extracts of Lagerstroemia speciosa as described above, by studying their performance in cell based assay models.

In some of the embodiments of the present invention, mammalian cell based efficacy tests are conducted by growing Human hepatoblastoma cell line (Hep G2) and in striated muscle (C2C12), in a flask with Eagle's Minimum Essential Medium (EMEM) containing 10% Fetal Bovine Serum (FBS), 1% glutamine-penicillin-streptomycin and 1% fungizone in a humidified incubator at 37°C in an atmosphere of 5% C02 and 95%

air. It is further subcultured when cell become 80% confluent they are subjected to treatment with bioactive under investigation. The incubation is followed by estimating levels of efficacy in glucose uptake between the bioactive treated and untreated sets.

In another aspect of the invention there is provided a method for treating a disease in a mammal, which comprises administering to the said mammal an effective non-toxic amount of at least an extract from Lagerstroemia speciosa as defined herein. Preferably the mammal is a human being. The skilled addressee will appreciate that "treating a disease" in a mammal means treating, that is to say, alleviating symptoms of the disease and may also mean managing a disease in the sense of preventing such a disease state either advancing i.e. getting worse or becoming more invasive, or slowing down the rate of advance of a disease.

The compositions/medicaments may contain a pharmaceutically acceptable carrier, excipient, or diluent. The compositions can be included as unit dosage suitable for parenteral, oral, or intravenous administration to a human. Alternatively, the compositions are dietary supplements, food compositions or beverage compositions suitable for human or animal consumption.

The invention is further elaborated with the help of following examples; however, these examples should not be construed to limit the scope of the instant invention.

Example 1: Extraction of Lagerstroemia speciosa:

Extraction of Lagerstroemia speciosa plant parts was carried out with alcohol, water and hydroalcohol solvent in reflux extractor apparatus or at room temperature under agitation followed by lyophilization under vacuum.

The detailed process is given below:

Reflux Extraction

Powdered Lagerstroemia speciosa plant material was weighed into the round bottom
flask. Various concentrations of alcohol, water and hydro-alcohol was added in to the round-bottomed flask and placed on the mantle along with few (3-4) ceramic chips. The reflux condenser was then placed on the flask. Cold water was allowed to circulate continuously in the condenser from the tap. The mantle was switched on and the temperature was set to the boiling point of the solvent. The vapors of the solvent from the flask passed through the inlet of the extractor and condenses. The condensed (distilled) solvent thus extracting the compounds from it. This process is continuous as long as there is stable heat and water circulation. The extraction was continued for 2 hours at room temperature. After 2 hours the mantle was switched off and the water flow was stopped.

After cooling the extract was collected separately and centrifuged.

The extract was concentrated by fitting the flask containing the extract with the empty soxhlet extractor body that in turn was fitted tightly with the water-cooled condenser. Continuous water flow was maintained and the flask was heated till the solvent from the flask was distilled and collected in the extractor body up to a level (One inch below the inlet). The temperature was reduced to avoid charring as the volume of the solvent reduced in the flask. The distilled solvent collected in the extractor was transferred to the solvent bottles and label appropriately. The process was continued till only very little solvent was left in the flask and no charring had occurred. Further concentration was done in the rotovapour apparatus to remove the solvent completely. The extract in the flask were swirled and were dried under vacuum. Storage and labeling of the extract was done to obtain the Extract ID.

Room Temperature Extraction

Powdered Lagerstroemia speciosa plant material was weighed into conical flask. Various concentrations of alcohol, water and hydro-alcohol was added in to the conical flask and placed under agitation at room temperature for 2 hrs. After 2 hrs, centrifuge at 4500 rpm for 15 minutes at 4°C. After centrifugation supernatant is taken and concentration is done in rotor evaporator. Further concentration is done in lyophilizer. The % yield of the extract obtained is calculated.

Calculations:

Calculate the percentage yield of the extract with respect to the initial weight of the plant material taken before extraction.

% Yield = wt. of lyophilized extract (after drying) * 100 Wt. of dry Plant material (initial)
The extract was concentrated by fitting the flask containing the extract with the empty soxhlet extractor body that in turn was fitted tightly with the water-cooled condenser. Continuous water flow was maintained and the flask was heated till the solvent from the flask was distilled and collected in the extractor body up to a level (One inch below the inlet). The temperature was reduced to avoid charring as the volume of the solvent reduced in the flask. The distilled solvent collected in the extractor was transferred to the solvent bottles and label appropriately. The process was continued till only very little solvent was left in the flask and no charring had occurred. Further concentration was done in the rotovapour apparatus to remove the solvent completely. The extract in the flask were swirled and were dried under vacuum. Storage and labeling of the extract was done as 70% and 20% Banopta extract.

Example 2: HPLC analysis:

HPLC analysis of Banopta 70% was done using Shimadzu LC using the parameters given below: Instrument initiation

Instrument was switched on and connected to the data station through communication bus module. Each line was purged with the respective solvents for five minutes. Further the column was washed with 100% methanol for 30 minutes and then equilibrated with the initial run conditions of the corresponding LC time program.

Instrument parameters Data acquisition

LC stops time: 50 mins

Acquisition time (PDA) Sampling: 3.125 Hz Start time: Omins End time: 40mins Time constant 0.64sec

Column used: Xterra® RP 18.3 5um 4.6x150mm.
Mobile phase

Mobile phase B: 0.1% TFA acidified acetonitrile
Mobile phase D: HPLC grade water

Auto sampler (SIL 20A)

Sample rack: 1.5ml standsrd

Rinsing volume: 200ul

Needle stroke: 52mm

Control vial needle stroke: 52mm

Rinsing speed: 35ul/sec

Sampling speed: 15ul/sec

Purge time: 25 min

Rinse mode: Before and after aspiration

Rinse dip time: l0Sec Auto purge

Purge order

Mobile phase A: 5 mins Mobile phase B: 5 mins Mobile phase C: 5 mins Mobile phase D: 5 mins Auto sampler: 5 mins Total pump A flow: 1ml/min

Samples analyzed

Banopta 70%

Run conditions

No of runs performed for each sample: 5

Injection volume: 5ul

LC time program used: Banopta 70% MD

Sample preparation

20mg of the lyophilized extract was weighed out into sterile eppendorf and 2ml of 50% methanol is added to it and sonicated for 15 minutes to ensure that the extract dissolves completely. The extract was centrifuged at 12,000rpm for 20 minutes at room temperature and the supernatant was filtered through 0.2u filters and collected into sterile recovery vials before analysis was performed.

The 70% and 20% extract of Banopta separately were injected at a constant volume of 5ul and its metabolite profiling monitored at 254nm. (Fig 1.1, Table. 1, Fig. 1.2, Table. 2) A comparative profiling of 70% and 20% extract of Banopta was performed between the three replicates runs/extract for the reproducibility of the extract profiles (Figl.3,1.4)

Example 2: Cell Based Assays

Insulin- stimulated glucose uptake in striated muscle (C2C12) and hepatocytes (Hep-G2) is critical for reducing post prondial blood glucose concentration and the dysregulation of this process is one hallmark of Type -II Diabetes mellitus [Non Insulin dependent]. Deoxyglucose is recognised by the Glucose transporter GLUT1 and is phosphorylated by hexokinase, since deoxyglucose labelled with Tritium (2-Deoxyglucose containing 2-deoxy 3H glucose) lacks a hydroxyl group, which prevents its further metabolism and facilitates intracellular accumulation of deoxyglucose. Banopta 70% was tested for its efficacy in glucose uptake in two different cell lines. Insulin- stimulated glucose uptake in striated muscle (C2C12) and hepatocytes (HepG2) is critical for reducing post prondial blood glucose concentration and the dysregulation of this process is one hallmark of Type -II Diabetes mellirus [Non Insulin dependent]. Deoxyglucose is recognised by the glucose transporter GLUT1 and is phosphorylated by hexokinase, since deoxyglucose labelled with Tritium (2-Deoxyglucose containing 2-deoxy 3H glucose) lacks a hydroxyl group, which prevents its further metabolism and facilitates intracellular accumulation of deoxyglucose. The extract was monitored for their insulin mimicking activity and also synergistic activity at four different concentrations. The toxicity of the extract was also tested at all the concentrations.

Cytotoxicity on C2C12:

BanOpta 70% was tested at doses of 0.1, 1,10,100 and 500 ug/ml. It did not show any significant cytotoxicity at all the above studied doses. (Fig2:l)

BanOpta 20% was tested at doses of 0.1, 1,10,100 and 500 ug/ml. It did not show any significant cytotoxicity at all the above studied doses.(Fig2:2)

Mimicking effect on C2C12:

BanOpta 70% was tested for insulin mimicking activity in C2C12 cells at doses of 0.1-500 ug/ml. It exhibited a dose dependent increase in the glucose uptake level at the higher doses of 0.1-10 ug/ml showing maximum of 4 fold increase at 10 ug/ml when compared to insulin showing 11 fold increase at 250nm concentration. However it had no effect at higher doses of 100-500 ug/ml. (Fig3:l)

BanOpta 20% was tested for insulin mimicking activity in C2C12 cells at doses of 0.1-500 ug/ml. It exhibited a dose dependent increase in the glucose uptake level at the higher doses of 0.1-10 =g/ml showing maximum of 10 fold increase at 10 |ag/ml when compared to insulin showing 21 fold increase at 250um concentration. However it had no effect at higher doses of 100-500 ug/ml. (Fig3:2)

Synergistic effect on C2C12:
BanOpta 70% was tested for insulin synergistic activity in C2C12 cells at doses of 0.1-500 ug/ml. It exhibited a statistically significant increase in the glucose uptake level at the lowest dose of 0.1 ug/ml showing maximum of 5-fold increase.(Fig4:l)

BanOpta 20% was tested for insulin synergistic activity in C2C12 cells at doses of 0.1-500 ug/ml. It exhibited a statistically significant increase in the glucose uptake level at the lowest dose of 0.1 ug/ml showing maximum of 3-fold increase.(Fig4:2)
Cytotoxicity on HepG2:

BanOpta 70% was tested at doses of 0.1, 1,10,100 and 500 ug/ml. It did not show any significant cytotoxicity at all the above studied doses.(Fig5:l)

BanOpta 20% was tested at doses of 0.1, 1,10,100 and 500 ug/ml. It did not show any significant cytotoxicity at all the above studied doses.(Fig5:2)

Mimicking effect on HepG2:

BanOpta 70% was tested for insulin mimicking activity in HepG2 cells at doses of 0.1-500 ug/ml. It exhibited a statistically significant dose dependent increase in the glucose uptake level at the doses of 0.1-10 ug/ml showing maximum of 11 fold increase at 1 ug/ml when compared to insulin showing 9 fold increase at 250uM concentration. (Fig6:l)

BanOpta 20% was tested for insulin mimicking activity in HepG2 cells at doses of 0.1-500 ug/ml. It exhibited a statistically significant dose dependent increase in the glucose uptake level at the doses of 0.1-1 (xg/ml showing maximum of 12 fold increase at 1 ug/ml when compared to insulin showing 6 fold increase at 250uM concentration. (Fig6:2)

Synergistic effect on HepG2:

BanOpta 70% was tested for insulin synergistic activity in HepG2 cells at doses of 0.1-
500 ug/ml. It exhibited a statistically significant increase in the glucose uptake level at the lower doses of 0.1-1 ug/ml showing maximum of 3 fold increase at 0.1 ug/ml when compared to insulin showing 2 fold increase at 50 uM concentration. (Fig7:l)

BanOpta 20% was tested for insulin synergistic activity in HepG2 cells at doses of 0.1-500 ug/ml. It exhibited a statistically significant increase in the glucose uptake level at the lower doses of 0.1-1 ug/ml showing maximum of 1.3 fold increase at 0.1 ug/ml when compared to insulin showing 2 fold increase at 50 uM concentration. (Fig7:2)

Modes of administration:

For administration to a mammal, the therapeutic composition can be formulated as a pharmaceutical or naturopathic formulation such as phytoceuticals or nutraceuticals, for oral, topical, rectal or parenteral administration or for administration by inhalation or spray. The phytoceutical or naturopathic formulation may comprise the one or more plant extracts in dosage unit formulations containing the conventional non-toxic physiologically acceptable carriers, adjuvants and vehicles. The term parenteral as used herein includes subcutaneous injections, intravenous, intramuscular, intrathecal, intrasternal injections or infusion techniques.

The pharmaceutical or naturopathic formulations may be in a form suitable for oral use, for example, as tablets, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsion hard or soft capsules, or syrups or elixirs. The therapeutic compositions of the invention may be formulated as phytoceuticals, or nutraceuticals.

Phytoceuticals may optionally comprise other plant-derived components and can therefore be delivered by such non-limiting vehicles as teas, tonics, juices or syrups. Nutraceuticals contemplated by the present invention may provide nutritional and/or supplemental benefits and therefore be delivered, for example as foods, dietary supplements, extracts, beverages or the like. Phytoceutical and nutraceuticals can be administered in accordance with conventional treatment programs and/or may be a part of the dietary or supplemental program.

Formulations intended for oral use may be prepared according to methods known in art for the manufacture of pharmaceutical compositions and may contain one or more agents selected from the group of sweetening agents, flavoring agents, coloring agents and preserving agents in order to provide palatable preparations.

Tablets contain the active ingredient in admixture with suitable non-toxic physiologically acceptable excipients including, for example, inert diluents, such as calcium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, such as corn starch, or alginic acid, binding agents, such as starch, gelatine or acacia, and lubricating agents, such as magnesium stearate, stearic acid or talc. The tablets can be uncoated, or they may be coated by known techniques in order to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period.

Various additives or carriers can be incorporated into the orally delivered pharmaceutical naturopathic formulations or the invention. Optional additives of the present composition include, without limitation, phospholipids, such as phosphatidyl glycerol, phosphotidyl inositol, phosphotidyl serine, phosphotidyl choline, phosphotidyl ethanolamine as well as phosphatidic acids, ceramide, cerebrosides, sphingomyelins and cardiolipins.

Pharmaceutical or naturopathic formulations for oral use may also be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatine capsules wherein the active ingredient is mixed with water or an oil based medium such as peanut oil, liquid paraffin or olive oil.

A syrup may be made by adding the active extract to a concentrated, aqueous solution of a sugar, for example sucrose, to which may also be added any necessary ingredients. Such accessory ingredient (s) may include flavorings, an agent to retard crystallisation of the sugar or an agent to increase the solubility of any other ingredients, such as polyhydric alcohol for example glycerol or sorbitol.

Oily suspensions may be formulated by suspending the plant extract(s) in a vegetable oil, for example, arachis oil, olive oil, sesame oil or coconut oil, or in a mineral oil such as liquid paraffin. The oily suspensions may contain a thickening agent, for example beeswax, hard paraffin or acetyl alcohol. Sweetening agents and/or flavoring agents may be added to provide palatable oral preparations. These formulations can be preserved by the addition of an anti-oxidant such as ascorbic acid. Dispersible powders and granules suitable for preparation suitable for an aqueous suspension by the addition of water provide the active ingredient in admixture with dispersing or wetting agent, suspending agent and one or more preservatives. Suitable dispersing or wetting agents and suspending agents,sweetening, flavoring and coloring agents may also be present.

In a further aspect of the invention there is provided a comestible, that is to say, a foodtuff comprising at least an extract of the invention, typically in dried form, such as in a lyophilised form. The skilled addressee will appreciate that such cosmetibles may contain more than one extract of the invention and may be used. Such foodstuffs may be used in a prophylactic manner and may contain further extracts having a similar function to the first added extract or further added extracts may be added that have a different prophylactic function. Thus a foodstuff could either comprise extracts that provide for a comestible having a single functional aspect, or a comestible may have a multi-functional prophylactic effect against two or more disease types. It is thought that a multi-functional role could be assigned to pharmaceutical formulations comprising two or more extracts possessing dissimilar therapeutic or prophylactic properties desgined either for prophylaxis or for the treatment of more than one disease(s) in a mammal, particularly in a human.

The type of foodstuff or comestible to which at least an extract of the invention may be added includes any processed food such as confectionaries, baked products including breads such as loafs, and flat breads such as pitta bread, naan bread and the like, cakes, snack foods such as muesli bars, compressed dried fruit bars, biscuits, dairy products such as yoghurts, milk and milk-based products such as custards, cream, cheese, butter and creme fraiche, simulated dairy food product such as Elmlea products, fruits and vegetable juices, water, aerated drinks, such as carbonated soft drinks and non-aerated drinks such as squashes, soya milk, rice milk and coconut milk and the like, pastas, noodles, vegetables, seed and nut oils, fruited oils such as sunflower oil, rapeseed oil, olive oil, walnut, hazelnut, and sesame seed oil and the like, and frozen confectionaries such as ice cream, iced yoghurts and the like.

We Claim:

1. An extract of Lagerstroemia species optionally along with pharmaceutically acceptable additive.

2. The extract as claimed in claim 1, wherein said Lagerstroemia species is Lagerstroemia speciosa.

3. The extract as claimed in claim 1, wherein said additive is selected from a group comprising granulating agents, binding agents, lubricating agents, disintegrating agents, sweetening agents, coloring agents, flavoring agents, coating agents, plasticizers, preservatives, suspending agents, emulsifying agents and spheronization agents or a combination thereof.

4. The extract as claimed in claim 1, wherein said extract is formulated as cosmetics and dosage forms selected from a group comprising tablet, troches, lozenges, aqueous or oily suspensions, dispersible powders or granules, emulsion hard or soft gel capsules, syrups, elixirs.

5. A process for obtaining extract from Lagerstroemia species, said process comprising steps of:

(a) powdering plant material of Lagerstroemia species;

(b) extracting and refluxing the powder with solvents to obtain extract; and

(c) concentrating the extract followed by drying to obtain the extract of Lagerstroemia species.

6. The process as claimed in claim 5, wherein said Lagerstroemia species is Lagerstroemia speciosa.

7. The process as claimed in claim 5, wherein said plant material is selected from a group comprising leaves, seeds, roots, stems, flowers, fruits, or various combinations thereof, preferably leaves.

8. The process as claimed in claim 5, wherein said extract is optionally centrifuged
followed by lyophilization to obtain the extract of Lagerstroemia species.

9. The process as claimed in claim 1, wherein said solvents are selected from a
group comprising water, alcohol and combinations thereof.

10. A method for management of calorie/weight and diabetes in a subject in need thereof, said method comprising step of administering pharmaceutically acceptable amount of extract of Lagerstroemia species optionally along with pharmaceutically acceptable additives to the subject.

11. The method as claimed in claim 1, wherein the subject is animal including human
being.

12. The extract of Lagerstroemia species, the process and the method as substantially
described herein with reference to accompanying examples and drawings.