Introduction

Diabetes mellitus (DM) is a debilitating and often life threatening disease with increasing incidence throughout the world. It was postulated that DM is the most common chronic disorder affected more than 176 million people worldwide, and this global figure has been set to double by the year 2030, Tiwari \& Madhusudaan [₁]. The treatment of DM relied heavily on dietary measures, which included the use of traditional plant therapies. Several reviews on plants with known antidiabetic activity or with traditional use as antidiabetic remedies have been published, Ajganonkar [₂], Oliver-Bever [₃], Bailey \& Day [₄]. Traditional antidiabetic plants might provide a useful source for developing new oral hypoglycemic compounds as pharmaceutical entities or simple dietary adjuncts to the exiting therapies. Studying such traditional medicines might offer an alternative and natural key to unlock diabetologists’ pharmacy. On the other hand, suggested mechanisms describing therapeutic effects of several traditional medicinal plant systems are holistic, Handa et al [₅]. Most of the reported hypoglycemic plants are anecdotal, and only few have received adequate scientific evaluation. The fundamental mechanisms of these medicinal systems are still unexplainable using modern tools, Rahman \& Zaman [₆]. It is claimed that most medicinal preparations in traditional medicines contain a variety of synergistically acting phytochemicals that are thought to act on a variety of targets by various modes and mechanisms, Tiwari \& Madhusudaan [₁]. In this study, we investigated the effect of Enicostemma hyssopifolium, Maroo et al [₇], Srinivasan ^{et al [₈], vasu et al [₉], Vijayvargia [₁₀]. Eugenia jumbolana, Achrekar et al [₁₁], Grover et al [₁₂], Mallick [₁₃], Ravi et al [₁₄], Kedar et al [₁₅], Tinospora cordifolia, Gupta et al [₁₆], Stanely et al [₁₇], Wadood et al [₁₈], Raghunathan [₁₉], Gymnema sylvestre, Shanmugasundaram et al [₂₀], Baskaran et al [₂₁], Okabayashi et al [₂₂], Sughira et al [₂₃], and Trigonella foenum Khosla et al [₂₄], Ravikumar \& Anuradha [₂₅], Ribes et al [₂₆], Vats et al [₂₇], a well-documented hypoglycemic plants on the kinetic uptake of D (+)-glucose in in-vitro everted gut sac model as described by Wilson \& Wiseman [₂₈] .}

Materials and methods

Preparation of the aqueous extracts

Experimental design

Preparation of everted gut sacs

Control experiments

Data analysis

Results

Biochemical parameters of D (+)-glucose transport across rat everted small intestines in vitro are shown in graphical forms in Figures. The Km and V_max were calculated in the absence as well as in the presence of plant extracts in the mucosal solution and are shown in Table 1. Data analysis revealed that V_max for glucose uptake decreased by 40.09, 28.67, 83.67, and 34.5 µM hr^-1 in the presence of the aqueous EJ seed extract, TC stem extract, GS leaf and TF, respectively. The apparent Km remained unaltered in the all case of the studied plant extracts.

Figure 1

Table 1: Biochemical parameters obtained for the effect of plant extracts on the transport of D (+)-glucose at different concentrations (5-20 mM) across the rat everted gut sacs.

The everted gut sacs were incubated in Krebs- Henseleit buffer (pH= 7.4) at 37°C.

Figure 2

Figure 1: Effect of extract on D (+)-glucose uptake across rat everted gut sac. ns: not significant, * p < 0.05

Figure 3

Figure 2: Effect of extract on D (+)-glucose uptake across rat everted gut sac. * p < 0.05, *** p < 0.0001

Figure 4

Figure 3: Effect of extract on D (+)-glucose uptake across rat everted gut sac. ** p < 0.001.

Figure 5

Figure 4: Effect of extract on D (+)-glucose uptake across rat everted gut sac. * p < 0.05, ** p < 0.001, *** p < 0.0001.

Figure 6

Figure 5: Effect of extract on D (+)-glucose uptake across rat everted gut sac. * p < 0.05, ** p < 0.001, *** p < 0.0001.

Discussion

The most challenging goal in the management of patients with DM is to achieve blood glucose level as close to normal as possible. Unfortunately, postprandial hyperglycaemia (PPHG) or hyperinsulinaemia are independent risk factors for the development of vascular complications in DM patients Tiwari \& Madhusudaan [₁]. Mechanisms playing role in release and transport of glucose across the intestinal brush border membrane down to the blood stream have attracted much attention recently as potential targets to control PPHG. In this category, majority of recent studies reported the potential use of antidiabetic medicinal plants on inhibition of glucose transport. Drugs that reduce PPHG by suppressing the absorption of carbohydrate are effective in prevention and treatment of non-insulin dependent DM. Alpha-glucosidase inhibitors are used to establish greater glycemic control over hyperglycemia in diabetes mellitus type 2, particularly with regard to PPHG. They may be used as monotherapy (acarbose, miglitol, voglibose) in conjunction with an appropriate diabetic diet and exercise, or they may be used in conjunction with other anti-diabetic drugs. They may also be useful in patients with diabetes mellitus type 1; however, this use has not been officially approved by the Food and Drug Administration.

Our findings would tend to indicate that glucose transport was significantly decreased in the presence of the aqueous extract of EJ seeds, TC stem, GS leaf, and TF seeds which caused a decrease in the V_max. Only the aqueous extract of aerial parts of EH did not decreased the glucose absorption in vitro rather it increases the V_max. Since the aqueous extract of aerial parts of EH found to contain high amount of reducing sugars in free form it may presume that it could cause enhancement of glucose uptake in vitro. However, the Km remained unaltered in the presence as well in the absence of these extracts. This indicates that these extracts act by bringing a non-competitive type of inhibition of glucose at the level of the small intestine. Glucose transport through biological membranes requires specific transport proteins. Active transport of glucose through the apical membrane of intestinal and kidney epithelial cells depends on the presence of secondary active Na⁺/glucose symporters, SGLT-1 and SGLT-2, which concentrate glucose inside the cells, using the energy provided by co-transport of Na⁺ ions down their electrochemical gradient, Hediger \& Rhoads [₃₁]. Passive transport (facilitated diffusion) of glucose through the cellular membrane is otherwise catalyzed by glucose carriers (protein symbol GLUT), Henderson et al [₃₂]. GLUT 2, a transmembrane carrier protein of GLUT family, located in basolateral membrane of small intestine, is very efficient carrier of glucose at small intestine level. A thiazolidinedione class of drug, rosiglitazone, activated the GLUT2 mRNA level in the liver, Im et al [₃₃] and thereby reducing blood glucose level. Encouraging data are available concerning effects of flavonoids on inhibiting GLUT 2 and their hypoglycemic effect, Kwon et al [₃₄], Song et al [₃₅], Johnston et al [₃₆].

Although our findings are promising, uncertainties remain. Some investigators suggested that flavonoids decreased glucose uptake by a sodium-dependent pathway via the sodium-dependent glucose transporter 1 SGLT1. It is most probable that active phytochemicals in the seeds of EJ, stem of TC, leaf of GS and seeds of TF binds on the glucose transporters thus may lead to wash out of glucose from the body. The latter may be one of the mechanisms for the hypoglycemic phenomenon after administration of these extracts noted by various investigators in animals or subjects.

In conclusions, our study provides evidence for a biochemical mechanism which carries blood glucose lowering effect of EJ seeds, TC stem, GS leaf and TF seeds in intestine via non-competitive inhibition. However, further kinetic data on carrier-mediated transport of D (+)-glucose is needed.

Acknowledgements