A novel botanical formula prevents diabetes by improving insulin resistance

Abstract

Background: Type 2 diabetes mellitus (T2DM) is a major risk factor for cardiovascular disease, and the prevalence has increased significantly in recent decades to epidemic proportions in China. Individually, fenugreek (Trigonella foenum graecum) seed, mulberry (Morus alba L.) leaf and American ginseng (Panax quinquefolius) root can improve glycemia in various animal models and humans with impaired glucose metabolism and T2DM. The aim of this study was to design an optimized botanical formula containing these herbal extracts as a nutritional strategy for the prevention of insulin resistance and T2DM.

Methods: Cell-free α-amylase and α-glucosidase enzyme assays were used to determine inhibitory potential of extracts. Glucose uptake was examined in differentiated human adipocytes using radiolabeled 2-deoxyglucose. Male Sprague Dawley rats were divided and glycemia balanced into 5 groups: two controls (naïve and model) and three doses of the botanical test formula containing standardized fenugreek seed, mulberry leaf and American ginseng extracts (42.33, 84.66 and 169.33 mg/kg BW). Insulin resistance and T2DM was induced by feeding animals a high fat diet and with an alloxan injection. Glucose tolerance was examined by measuring serum glucose levels following an oral glucose load.

Results: Fenugreek seed and mulberry leaf dose dependently inhibited α-amylase (IC50 = 73.2 μg/mL) and α-glucosidase (IC50 = 111.8 ng/mL), respectively. All three botanical extracts improved insulin sensitivity and glucose uptake in human adipocytes, which lead to the design of an optimized botanical test formula. In a rat model of insulin resistance and T2DM, the optimized botanical test formula improved fasting serum glucose levels, fasting insulin resistance and the development of impaired glucose tolerance. The reduction in epididymal adipose tissue GLUT4 and PDK1 expression induced by high fat diet and alloxan was blunted by the botanical test formula.

Conclusions: A novel botanical formula containing standardized extracts of mulberry leaf, fenugreek seed and American ginseng at a ratio of 1:1.3:3.4 prevented the development of insulin resistance, impaired glucose tolerance and T2DM. Given the rising need for effective non-drug targeting of insulin resistance and progression to T2DM, complementary and alternative nutritional strategies without intolerable side effects could have meaningful impact on metabolic health and diabetes risks.

Keywords: Alloxan; American ginseng; Botanical test formula; Fenugreek seed; GLUT4; High fat diet; Insulin resistance; Mulberry leaf; Type 2 diabetes mellitus.

Fig. 1

The effect of fenugreek seed (FS) and mulberry leaf (ML) on carbohydrate digestive enzymes. The effect of fenugreek seed on α-amylase (a) and mulberry leaf on α-glucosidase (b) activity were determined in a cell-free, enzyme-substrate assay model. Data were fitted with log (inhibitor) vs. response, variable slope (four parameters) equation in Prism Graph Pad 6.00. Data are presented as mean ± SD of triplicates

Fig. 2

The individual (a) and synergetic (b) effect of fenugreek seed (FS), mulberry leaf (ML) and American ginseng (AG) extracts on insulin sensitivity in differentiated primary human subcutaneous adipocytes. After 20 h of pre-treatment with extracts or botanical test formula, glucose uptake was initiated with the addition of a cocktail containing 2-DOG and ³H-2-DOG with 1 nM insulin and incubate for 2 h. Change in insulin sensitivity was determined by setting 1 nM insulin as 0% and 100 nM as 100%, and calculating the percent change in the delta between 1 nM and 100 nM insulin. Data are presented as mean ± SD of triplicates. ^* P < 0.05 vs. 1:1:1 ratio group

Fig. 3

The effect of the botanical test formula on fasting glycemia status. Fasting (3 h) serum glucose (a), insulin (b) and HOMA-IR (c) in a chronic HFD fed and an acute alloxan induced model of insulin resistance and T2DM treated with the botanical test formula comprised of fenugreek seed, mulberry leaf and American ginseng for 33 d. Naïve control (n = 10); model control (n = 10); 1× (n = 10); 2× (n = 9); 4× (n = 7). Data are presented as mean ± SD. ^### P < 0.001, ^## P < 0.01 and ^# P < 0.05 vs. naïve control group, ^*** P < 0.001 and ^** P < 0.01 vs. model control group

Fig. 4

The effect of the botanical test formula on oral glucose tolerance. Serum glucose levels (a) and AUC (b) in a chronic HFD fed and an acute alloxan induced model of insulin resistance and T2DM treated with the botanical test formula comprised of fenugreek seed, mulberry leaf and American ginseng for 33 d. Animals were fasted 3 h followed by an administration of an oral glucose load (2.5 g/kg) and blood was collected at 0, and 0.5 and 2 h post challenge. Naïve control (n = 10); model control (n = 10); 1× (n = 10); 2× (n = 9); 4× (n = 7). Data are presented as mean ± SD. ^### P < 0.001 vs. naïve control group, ^*** P < 0.001, ^** P < 0.01 and ^* P < 0.05 vs. model control group

Fig. 5

The effect of the botanical test formula on fasting lipids. Fasting (3 h) triglycerides (a) and total cholesterol (b) levels in a chronic HFD fed and an acute alloxan induced model of insulin resistance and T2DM treated with a test formula comprised of fenugreek seed, mulberry leaf and American ginseng for 33 d. Naïve control (n = 10); model control (n = 10); 1× (n = 10); 2× (n = 9); 4× (n = 7). Data are presented as mean ± SD. ^### P = 0.0015 vs. naïve control group, ^# P < 0.05 vs. naïve control group

Fig. 6

The effect of the test formula (1X dose) on adipose tissue expression of PDK1 and GlUT4. Western blot (a) and densitometry analysis of PDK1 (b) and GlUT4 (c) in epididymal adipose tissue in a chronic HFD fed and an acute alloxan induced model of insulin resistance and T2DM treated with the botanical test formula comprised of fenugreek seed, mulberry leaf and American ginseng for 33 d. GAPDH was used as the loading control. Quantitated data are presented as mean ± SD of pool group samples run in triplicate. Model control (M); Naïve control (N); Test Formula (1×) (F). ^### P < 0.001 and ^# P < 0.05 vs. naïve control group, ^** P < 0.01 vs. model control group