CLIAMS:I/We claim:
1.A composition for management of Type 2 Diabetes Mellitus, said composition comprises:
a. a xanthonoid; and
b. cholecalciferol
wherein, said composition shows a synergistic increases in glucose uptake in skeletal muscle cells.
2.The composition as claimed in claim 1, wherein the xanthanoid is mangiferin.
3.The composition as claimed in claim 2, wherein the mangiferin is extracted from the bark of Mangifera indica.
4.The composition as claimed in claim 3, wherein the concentration of mangiferin in the Mangifera indica bark extract is 80%.
5.The composition as claimed in claim 1, wherein the w/w ratio of Mangifera indica extract to cholecalciferol is in the range of 70:1 to 350:1
6.The composition as claimed in claim 1, wherein the w/w ratio of Mangifera indica extract to cholecalciferol is 325:1.
7.A food supplement comprising the composition as claimed in claim 1.
8.A method for management of Type 2 Diabetes Mellitus, wherein the method comprises oral administration to a subject in need thereof an effective amount of said food supplement as claimed in claim 8.
9.The method as claimed in claim 8, wherein oral administration can be in the form of a solid or a liquid.
10.The method as claimed in claim 8, wherein the subject is a mammal.
,TagSPECI:FIELD OF INVENTION
[0001] The present disclosure relates to a composition comprising of Mangifera indica extract and cholecalciferol and methods for management of Type II Diabetes Mellitus (T2DM).
BACKGROUND OF THE INVENTION
[0002] The incidence of diabetes has steadily increased worldwide in the last few years affecting all ages across the population. Diabetes Mellitus type II is a chronic disorder primarily characterized by defects in glucose metabolism. The elevated level of glocuse in the blood, known as “hyperglycemia”, is hallmark of diabetes. Hyperglycemia can be due to insulin resistance or insulin deficiency.
[0003] Environmental factors such as imbalanced diet, lack of adequate physical activity, and imbalanced micronutrient intake can all contribute to manifestation of diabetic condition (Murea et al., Rev Diabet Stud., 2012, 9(1), 6–22).
[0004] Uncontrolled chronic hyperglycemia produces systemic effects including severe complications such as retinopathy, nephropathy, neuropathy and cardiovascular complications (Aronson, Adv Cardiol., 2008, 45, 1-16).
[0005] Skeletal muscles are one of the major insulin targets for blood glucose homeostasis. Glucose uptake defects in skeletal muscle cells are regularly seen in T2DM (Nedachi et al., Am J Physiol Endocrinol Metab., 2006, 291, E817–E828).
[0006] It has been reported that there is a direct association between cholecalciferol/vitamn D and T2DM. Vitamin D levels are reduced in the patient population diagnosed with T2DM. Supplementation of vitamin D improves the glucose uptake by skeletal muscles even under insulin resistance (Zhou et al., Diabetes Metab Res Rev, 2008, Sep;24(6), 459-64). Vitamin D supplementation has also been shown to help maintain blood glucose homeostasis (Alvarez et al., International Journal of Endocrinology, 2009, 1-18).
[0007] Mangiferia indica, commonly known as mango is found extensively in many tropical and sub-tropical regions worldwide. Mangifera indica is rich in phytochemicals including phenolic acids, phenolic esters, flavan-3-ols and mangiferin. Mangiferin is the predominant component of Mangifera indica extract. Extracts from the mango plant have diverse biological indications such as analgesic, anti-viral, anti-fungal, anti-bacterial, anti-parasitic, anti-allergic, anti-tumor, anti-inflammatory, anti-diabetic, radioprotective, immunomodulatory, lipolytic, antibone resorption, monoamine oxidase inhibiting and antioxidant properties (Shah et al., Pharmacogn Rev., 2010, 4(7), 42–48).
[0008] US 2006/0159784 A1 discloses pharmaceutical or veterinary or nutritional compositions of polyherbal extracts containing norathyriol and cholecalciferol.
[0009] Given the rising incidence of metabolic disorders such as diabetes among the population worldwide and increased awareness of natural products that combat such metabolic disorders, there is a need to develop high efficacy compositions that provide enhanced long-term management.
[0010] Natural products with known beneficial effects apart from its intended function have fewer side effects in contrast to synthetic formulations and enjoy wider acceptance among the healthy and target population.
SUMMARY OF INVENTION
[0011] This summary is provided to introduce concepts related to a composition for management of T2DM comprising of Mangifera indica extract, and cholecalciferol.
[0012] Yet another aspect of the present disclosure relates to a method for management of T2DM comprising administering an effective amount of the composition as described in the present disclosure.
[0013] Another aspect of the present disclosure relates to a food supplement comprising Mangifera indica extract, and cholecalciferol.
[0014] Yet another aspect of the present disclosure relates to a method for management of T2DM, said method comprising oral administration to a subject in need thereof an effective amount of a food supplement comprising Mangifera indica extract, and cholecalciferol.
BRIEF DESCRIPTION OF THE ACCOMPANYING DRAWINGS
[0015] The following drawings form part of the present specification and are included to further illustrate aspects of the present disclosure. The disclosure may be better understood by reference to the drawings in combination with the detailed description of the specific embodiments presented herein.
[0016] Figure 1 shows a graphical representation of the effect of various natural extracts on glucose uptake by murine C2C12 skeletal muscle cells, in accordance with an embodiment of the present disclosure.
[0017] Figure 2 shows a graphical representation of the effect of various micronutrients, specifically vitamins on glucose uptake by murine C2C12 skeletal muscle cells, in accordance with an embodiment of the present disclosure.
[0018] Figure 3 shows a graphical representation of the effect of various concentrations of cholecalciferol on glucose uptake by murine C2C12 skeletal muscle cells, in accordance with an embodiment of the present disclosure.
[0019] Figure 4 shows a graphical representation of the effect of various concentrations of Mangifera indica extract on glucose uptake by murine C2C12 skeletal muscle cells, in accordance with an embodiment of the present disclosure.
[0020] Figure 5 shows a graphical representation of the effect of various combinations of cholecalciferol and norathyriol on glucose uptake by murine C2C12 skeletal muscle cells, in accordance with an embodiment of the present disclosure.
[0021] Figure 6 shows a graphical representation of the synergistic effect of a composition comprising cholecalciferol and Mangifera indica extract on glucose uptake by murine C2C12 skeletal muscle cells, in accordance with an embodiment of the present disclosure.
[0022] Figure 7 shows a graphical representation of the synergistic effect of a composition comprising of cholecalciferol and Mangifera indica extract on total number of murine C2C12 skeletal muscle cells taking up 6-NBDG, in accordance with an embodiment of the present disclosure.
DETAILED DESCRPTION OF THE INVENTION
[0023] Those skilled in the art will be aware that the present disclosure is subject to variations and modifications other than those specifically described. It is to be understood that the present disclosure includes all such variations and modifications. The disclosure also includes all such steps, features, compositions and compounds referred to or indicated in this specification, individually or collectively, and any and all combinations of any or more of such steps or features.
Definitions
[0024] For convenience, before further description of the present disclosure, certain terms employed in the specification, and examples are collected here. These definitions should be read in the light of the remainder of the disclosure and understood as by a person of skill in the art. The terms used herein have the meanings recognized and known to those of skill in the art, however, for convenience and completeness, particular terms and their meanings are set forth below.
[0025] The articles “a”, “an” and “the” are used to refer to one or to more than one (i.e., to at least one) of the grammatical object of the article.
[0026] The terms “comprise” and “comprising” are used in the inclusive, open sense, meaning that additional elements may be included. It is not intended to be construed as “consists of only”.
[0027] Throughout this specification, unless the context requires otherwise the word “comprise”, and variations such as “comprises” and “comprising”, will be understood to imply the inclusion of a stated element or step or group of element or steps but not the exclusion of any other element or step or group of element or steps.
[0028] The term “including” is used to mean “including but not limited to”. “Including” and “including but not limited to” are used interchangeably.
[0029] The term “cholecalciferol’ and ‘vitamin D’ are used interchangeably.
[0030] As used herein, the term “vitamin D” refers specifically to vitamin D3.
[0031] As used herein, the term “management” does not necessarily mean a complete cure. It means that the symptoms or complications of the underlying disease are reduced, and/or that one or more of the underlying cellular, physiological, or biochemical causes or mechanisms causing the symptoms or complications are reduced. It is understood that “reduced”, as used in this context, means relative to the untreated state of the disease, including the molecular state of the disease, not just the physiological state of the disease.
[0032] Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the disclosure, the preferred methods, and materials are now described. All publications mentioned herein are incorporated herein by reference.
[0033] A composition comprising “synergistic activity” or a “synergistic composition” is a combination of compounds which exhibits increased biological or functional activity as a non-linear multiple of the biological or functional activity of the individual compounds. In other words, the combined biological or functional activity of two or more compounds being tested is significantly greater than the expected result based on independent effects of the compounds when tested separately. Synergy may be apparent only at some ranges or concentrations.
[0034] The present disclosure is not to be limited in scope by the specific embodiments described herein, which are intended for the purposes of exemplification only. Functionally-equivalent products, compositions, and methods are clearly within the scope of the disclosure, as described herein.
[0035] In an embodiment of the present disclosure, there is provided a composition for management of Type 2 Diabetes Mellitus, said composition comprising a xanthonoid and cholecalciferol.
[0036] In an embodiment of the present disclosure, the xanthonoid is mangiferin.
[0037] In an embodiment of the present disclosure, mangiferin is in the form of an extract from the bark of Mangifera indica.
[0038] In an embodiment of the present disclosure, the concentration of mangiferin in the extract from the bark of Mangifera indica is 80%.
[0039] In an embodiment of the present disclosure, the Mangifera indica extract enhances the uptake of glucose by murine C2C12 skeletal muscle cells.
[0040] In an embodiment of the present disclosure, cholecalciferol enhances the uptake of glucose by murine C2C12 skeletal muscle cells.
[0041] In an embodiment of the present disclosure, there is provided a composition comprising Mangifera indica extract, and cholecalciferol, said composition synergistically enhances the uptake of glucose by murine C2C12 skeletal muscle cells.
[0042] In an embodiment of the present disclosure, there is provided a composition comprising Mangifera indica extract, and cholecalciferol, wherein the w/w ratio of Mangifera indica extract to cholecalciferol is in the range of 70:1 to 350:1.
[0043] In a preferred embodiment of the present disclosure, there is provided a composition comprising Mangifera indica extract, and cholecalciferol, wherein the w/w ratio of Mangifera indica extract to cholecalciferol is 325:1.
[0044] In an embodiment of the present disclosure, there is provided a composition comprising Mangifera indica extract, and cholecalciferol, wherein the concentration of Mangifera indica extract in said composition is in the range of 25µg/mL to 60µg/mL.
[0045] In a preferred embodiment of the present disclosure, there is provided a composition comprising Mangifera indica extract, and cholecalciferol, wherein the concentration of mangiferin in said composition is 60µg/mL.
[0046] In an embodiment of the present disclosure, there is provided a composition comprising Mangifera indica extract, and cholecalciferol, wherein the concentration of cholecalciferol in said composition is in the range of 0.19258µg/mL to 0.308µg/mL.
[0047] In a preferred embodiment of the present disclosure, there is provided a composition comprising Mangifera indica extract, and cholecalciferol, wherein the concentration of cholecalciferol is 0.1925µg/mL.
[0048] In a preferred embodiment of the present disclosure, there is provided a composition comprising Mangifera indica extract, and cholecalciferol, wherein the concentration of Mangifera indica extract and cholecalciferol is 60µg/mL and 0.1925µg/mL respectively.
[0049] In an embodiment of the present disclosure, there is provided a food supplement comprising a composition consisting of Mangifera indica extract and cholecalciferol for management of T2DM.
[0050] In an embodiment of the present disclosure, there is provided a food supplement comprising a composition consisting of Mangifera indica extract and cholecalciferol for management of T2DM, wherein the w/w ratio of Mangifera indica extract to cholecalciferol is 325:1.
[0051] In an embodiment of the present disclosure, there is provided a food supplement comprising a composition consisting of Mangifera indica extract and cholecalciferol for management of T2DM, wherein the concentration of Mangifera indica extract and cholecalciferol is 60µg/mL and 0.1925µg/mL respectively.
[0052] The composition of the present disclosure may be compounded with foods such as, but not limited to dairy products, grains, breads, meats, fruits, vegetables, rice and the like. The composition according to the present disclosure can be compounded with additional ingredients. For example, the composition can be mixed with carbohydrates lipids, polypeptides, fatty acids, phytochemicals, and combinations thereof.
[0053] Any conventional food processing technique may be used to achieve a product comprising the effective amount of the composition described herein. There is much information on the art and technology of the various conventional food processing techniques and their practices in both the pet food and food industries, and it is accordingly assumed that the general principals of these techniques are understood by the person skilled in the art.
[0054] In an embodiment of the present disclosure, the food supplement is in a solid form.
[0055] In an embodiment of the present disclosure, the food supplement is in a liquid form.
[0056] In an embodiment of the present disclosure, the food supplement is in a semi-solid form.
[0057] In an embodiment of the present disclosure, there is provided a method of management of T2DM, wherein said method comprises administering to a subject in need thereof a therapeutically effective amount of a composition comprising Mangifera indica extract, and cholecalciferol, wherein the effective amount is insufficient to suppress natural physiology at local or at a systemic level.
[0058] In another embodiment of the present disclosure, the subject in need thereof a therapeutically effective amount of the composition comprising Mangifera indica extract, and cholecalciferol is a mammal.
[0059] The present disclosure provides a useful composition for prevention and/or management of T2DM. The composition of the present disclosure comprises Mangifera indica extract, and cholecalciferol, wherein the composition is highly safe for consumption, useful and can be administered easily over a long term.
[0060] In an embodiment of the present disclosure, there is provided a composition comprising Mangifera indica extract, and cholecalciferol, said composition enhances the transcript levels of GLUT4 mRNA in target cells.
[0061] Although the subject matter has been described in considerable detail with reference to certain preferred embodiments thereof, other embodiments are possible.
EXAMPLES
[0062] The disclosure will now be illustrated with working examples, which is intended to illustrate the working of disclosure and not intended to take restrictively to imply any limitations on the scope of the present disclosure. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art to which this disclosure belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice of the disclosed methods and compositions, the exemplary methods, devices and materials are described herein. It is to be understood that this disclosure is not limited to particular methods, and experimental conditions described, as such methods and conditions may vary.
Example 1
Preparation of solutions of test compounds used for screening for evaluation of glucose uptake
[0063] Preparation of Mangifera indica extract: Mangifera indica extract was sourced from Natural remedies. A 134 mg/mL stock solution of Mangifera indica extract was prepared by dissolving the extract in a solvent, preferably dimethyl sulfoxide (DMSO). A 30 -250 µg/mL of Mangifera indica extract working solution was prepared by dissolving stock solution in Dulbecco’s modified Eagle’s medium (DMEM)(Gibco, NY, USA).
[0064] Preparation of vitamin D: vitamin D was sourced from Sigma (catalog number C9756). A 0.39 mg/ml stock solution of vitamin D was prepared by dissolving the powder in a solvent, preferably Ethanol. A 0.385 µg/mL - 0.0385 ng/mL working solution was prepared from the stock solution by dissolving in DMEM (Gibco, NY, USA).
[0065] Preparation of Diadzein: Diadzein was sourced from Sigma (D7802). A 7.0 mg/ml stock solution was prepared Diadzein powder in solvent, preferably DMSO. A 30 µg/mL working solution of Diadzein was prepared by dissolving stock solution in DMEM (Gibco, NY, USA).
[0066] Preparation of Echinacea extract: Echinacea extract was sourced from Indena S.A.S. (catalog number 9045000). A 126.0 mg/ml stock solution was prepared by dissolving the extract powder in a solvent, preferably H20: DMSO (2:3). A 400 µg/mL working solution of Echinacea extract was prepared by dissolving stock solution in DMEM (Gibco, NY, USA).
[0067] Preparation of Oleuropein: Oleuropein was sourced from Sami Labs (C110827). A 51.0 mg/ml stock solution was prepared by Oleuropein powder in a solvent, preferably Ethanol. A 100 µg/mL working solution of Oleuropein was prepared by dissolving stock solution in DMEM (Gibco, NY, USA).
[0068] Preparation of Morus alba extract: Morus alba extract was sourced from Natural remedies. A 128 mg/ml stock solution was prepared by dissolving Morus alba extract powder in a solvent, preferably H2O:DMSO (5:1). A 460µg/mL working solution of Morus alba extract was prepared by dissolving stock solution in DMEM (Gibco, NY, USA).
[0069] Preparation of Catharanthus roseus extract: Catharanthus roseus extract was sourced from ITC Limited, Bangalore, India. A 124mg/mL stock solution was prepared by dissolving Catharanthus roseus extract powder in a solvent, preferably DMSO. A 75.2µg/mL working solution of Catharanthus roseus extract was prepared by dissolving stock solution in DMEM (Gibco, NY, USA).
[0070] Preparation of Pterostilbene: Pterostilbene was sourced from Sigma (P1499). A 55.5mg/mL stock solution was prepared by dissolving Pterostilbene powder in a solvent, preferably Ethanol. A 33.3µg/mL working solution of Pterostilbene was prepared by dissolving stock solution in DMEM (Gibco, NY, USA).
Example 2
Cell culture and culture media
[0071] The cell culture media for C2C12 cells was prepared by adding 445mL of DMEM(Gibco, N.Y., USA) with 50mL Fetal Bovine Serum (FBS)(Invitrogen, NZ) and 5mL 100U/mL penicillin / 0.1mg/mL streptomycin (Sigma, USA). The solution was filtered through a 0.2 micron filter unit.
[0072] C2C12 murine skeletal muscle precursor cells (myoblasts) were obtained from American Type Culture Collection (Manassas, VA, USA) and maintained in proliferation media, consisting of DMEM supplemented with 10% FBS. Myoblasts were induced to differentiate into skeletal muscle myotubes by culturing for six-days in differentiation media, consisting of DMEM supplemented with 2% horse serum (HS) (Invitrogen, NZ). Differentiation media was replenished every 48 hours.
Example 3
Glucose uptake assay by fluorimetry
[0073] The C2C12 cells were allowed to get 70% confluent before differentiation was initiated. The C2C12 cells were treated with DMEM supplemented with 2 % (v/v) HS. The cells were subsequently re-fed every 48 hours with 2% HS supplemented DMEM.
[0074] At the 6th day of differentiation, the culture media were removed and replaced with serum-free DMEM containing no glucose and incubated at 37 °C with 5% CO2 for one-hour. The C2C12 cells were subsequently re-fed for one-hour with serum free no glucose DMEM with or without treatments. Subsequently, serum free media with or without treatment were removed and cells were incubated at 37 °C with 5% CO2 for 30 minutes with 250µM of fluorescent analogue of glucose, 6-(N-(7-Nitrobenz-2-oxa-1,3-diazol-4-yl)amino)-2-deoxyglucose (6-NBDG) in serum free no glucose DMEM. The 6-NBDG uptake reaction was stopped by removing the incubation medium and washing the cells twice with pre-chilled phosphate buffered saline (PBS pH 7.4). Cells in each well were subsequently re-suspended in pre-chilled fresh Opti-MEM (Invitrogen, NZ) and maintained at 4°C for fluorimetry analysis performed within 30 minutes. The experiments were conducted in 96-well plates. All the treatment groups were done in triplicates.
Example 4
Glucose uptake assay by flow cytometry
[0075] The C2C12 cells were allowed to get 70% confluent and differentiation was initiated. The C2C12 cells were treated with DMEM supplemented with 2 % (v/v) HS. The cells were subsequently re-fed every 48 hours with DMEM supplemented with 2% HS.
[0076] At the 6th days of differentiation, the culture media were removed and replaced with serum free DMEM containing no glucose and incubated at 37 °C with 5% CO2 for one-hour. The C2C12 cells were subsequently re-fed for one-hour with serum free no glucose DMEM with or without treatments. Then serum free media with and without treatment was removed and cells were incubated at 37 °C with 5% CO2 for 30 minutes with 250µM of fluorescent analogue of 6-NBDG in serum free no glucose DMEM. The 6-NBDG uptake reaction was stopped by removing the incubation medium and the cells were washed twice with pre-cold phosphate buffered saline (PBS). Cells in each well were subsequently re-suspended in pre-cold fresh optimum medium and maintained at 4°C for flow cytometry analysis performed within 30 minutes. For each measurement, data from 5000 single cell events was collected using a FACS calibur (Becton Dickinson Immunocytometry Systems, SanJose, CA) flow cytometer. The experiments were conducted in 96-well plates. All the treatment groups were done in triplicates.
Example 5
Effect of natural extracts on glucose uptake by murine C2C12 skeletal muscle cells
[0077] Figure 1 shows the effect of various natural extracts on glucose uptake by murine C2C12 skeletal muscle cells. A 250µg/mL Mangifera indica extract with 80% mangiferin concentration shows maximal efficacy in positively modulating glucose uptake by C2C12 cells. This dose of Mangifera indica extract was found to be safe based on ¬in-vitro cyto-toxicity studies.
[0078] Stained control: Relative fluorescence intensity of 6-NBDG uptake in C2C12 cells without any treatment.
[0079] AICAR positive control: Relative fluorescence intensity of 6-NBDG uptake in C2C12 cells treated with 120nM of Insulin
Example 6
Effect of micronutrients on glucose uptake by murine C2C12 skeletal muscle cells
[0080] Figure 2 shows the effect of various micronutrients on glucose uptake by murine C2C12 skeletal muscle cells. Vitamin D3/cholecalciferol shows a significant positive effect on uptake of glucose by C2C12 cells. Cholecalciferol was chosen for further studies as cholecalciferol may be less toxic than vitamin D2 and a lower dose of cholecalciferol is more effective at inducing glucose uptake in C2C12 cells than vitamin D2 as inferred from Figure 2. Additionally, cholecalciferol is highly stable and is a more potent form of vitamin D.
Example 7
Dose response of cholecalciferol on glucose uptake by murine C2C12 skeletal muscle cells
[0081] Figure 3 shows the effect of various concentrations of cholecalciferol on glucose uptake by murine C2C12 skeletal muscle cells. It can be inferred that 0.385µg/mL dose of cholecalciferol is able to significantly enhance glucose uptake by C2C12 cells. A higher concentration was chosen for combination studies over ng/mL concentrations as ng/mL concentrations are deemed too low in target population with vitamin D deficiency.
Example 8
Dose response of Mangifera indica extract on glucose uptake by murine C2C12 skeletal muscle cells
[0082] Figure 4 shows the effect of various concentrations of a Mangifera indica extract on glucose uptake by murine C2C12 skeletal muscle cells. While 250µg/mL concentration of extract shows maximal effect on glucose uptake by C2C12 cells, lower concentration were chosen for further studies to uncover potential synergistic response when used in a combination with cholecalciferol.
Example 9
Effect of a combination of norathyriol and cholecalciferol on glucose uptake by murine C2C12 skeletal muscle cells
[0083] Figure 5 shows the effect of various concentrations of a composition comprising cholecalciferol and a mangiferin metabolite, norathyriol on glucose uptake by murine C2C12 skeletal muscle cells. It is inferred from Figure 5 that no synergistic effect of the composition can be seen on glucose uptake compared to individual components of the composition alone. The ratio of cholecalciferol to norathyriol is given in Table 1. It can be inferred from Figure 5 that norathyriol, an active metabolite of mangiferin, does not offer any enhanced synergistic effect on glucose uptake by C2C12 cells.
Table 1
Cholecalciferol Norathyriol Ratio
0.385µg/mL 0 µg/mL 1 : 0
0.308µg/mL 10 µg/mL 1 : 33
0.231µg/mL 20 µg/mL 1 : 87
0.1925µg/mL 25 µg/mL 1 : 130
0.154µg/mL 30 µg/mL 1 : 195
0.077µg/mL 40 µg/mL 1 : 520
0 µg/mL 50 µg/mL 0 : 1

Example 10
Effect of a combination of Mangifera indica extract and cholecalciferol on glucose uptake by murine C2C12 skeletal muscle cells
[0084] Figure 6 shows the effect of various concentrations of a composition comprising cholecalciferol and a Mangifera indica extract containing 80% mangiferin on glucose uptake by murine C2C12 skeletal muscle cells. Table 2 lists the ratio of mangiferin to cholecalciferol in the composition used. It can be inferred from Figure 6 that a combination of Mangifera indica extract and cholecalciferol in a w/w ratio of 325:1 provides a synergistic effect on glucose uptake relative to the effect of the Mangifera indica extract or cholecalciferol alone. MIE stands for Mangifera indica extract.
Table 2
Mangifera indica extract Cholecalciferol Ratio
125 µg/mL 0 µg/mL 1 : 0
100 µg/mL 0.077µg/mL 1299 : 1
75 µg/mL 0.154µg/mL 487 : 1
62.5 µg/mL 0.1925µg/mL 325 : 1
50 µg/mL 0.231µg/mL 217 : 1
25µg/mL 0.308µg/mL 81 : 1
0 µg/mL 0.385µg/mL 0 : 1

[0085] Figure 7 shows the relative fluorescence intensity of 6-NBDG uptake in murine C2C12 skeletal muscle cells treated with different ratios of Mangifera indica extract and cholecalciferol (Table 2). It can be inferred from Figure 7 that a maximal synergistic effect of the composition can be seen at a specific ratio of 325:1 (Mangifera indica extract: cholecalciferol), where the concentration of Mangifera indica extract is 62.5µg/mL and concentration of cholecalciferol is 0.1925µg/mL.