Rebaudioside A

Rebaudioside A is a steviol glycoside, α-glucosidase inhibitor with IC50 of 35.01 μg/ml.can inhibit ATP-sensitive K+-channels.Target: α-glucosidase [1]IC 50: 35.01 ug/mLIn vitro: rebaudioside A stimulat the insulin secretion from MIN6 cells in a dose- and glucose-dependent manner. In conclusion, the insulinotropic effect of rebaudioside A is mediated via inhibition of ATP-sensitive K+-channels and requires the presence of high glucose. [2]In vivo: in vivo mouse micronucleus test at doses

After pEUGT-SUS expressionin E. coli, the cells were harvested and washed twice in potassium phosphate buffer (100 mM, pH 7.2). A lysate containing soluble proteins was obtained by sonication on an ice-water bath and subsequent clarification by centrifugation at 4 °C. The supernatant was used as the crude extract. Total protein concentrations were measured by the Bradford assay using bovine serum albumin (BSA) as a standard.23) The substratestevioside was incubated with aliquots of the crude extract at 30 °C in a reaction mixture containing varying concentrations of UDP-glucose or UDP, sucrose, and 3 mM MgCl2 in potassium phosphate buffer (100 mM, pH 7.2) for synthesis of rebaudioside A.Stevioside (200 mg) and curdlan (400 mg) was dissolved in citrate buffer (0.1 M with pH 4.5) and β-1, 3-glucanase 3.425 U/g was added. The reaction mixture was shaking in an incubator for 3 hr at 55 °C. The reaction mixture was cooled to room temperature and the enzyme was deactivated by boiling at 100 °C. Progress of the reaction was monitored by TLC using solvent system ethyl acetate/ethanol/H2O (8:2:1.2, v/v/v) (Jaitak et al., 2009a, b). The developed plate was dried and spots were visualized by spraying with 5percent sulfuric acid. Reaction mixture was passed through the Dianion HP-20, followed by washing with methanol. Methanol fraction was dried over rota vapor and recrystallized using cold ethanol to yield 150 mg of pure Rebaudioside A. Product was confirmed by, NMR, HRMS and melting point determination. Rebaudioside A: MP 245–247 °C, Rf value 0.45, ethyl acetate:ethanol:water (8:2:1.2, v/v/v), 1H NMR (400 MHz, DMSO): 5.64 (1H, d, J = 8 Hz), 5.28 (1H, d, J = 8 Hz), 5.02 (1H, t, J = 8 Hz) and 4.95 (1H, d, J = 8 Hz). 13C-NMR (100 MHz, DMSO): δc 14.89, 18.56, 19.79, 21.14, 28.04, 37.48, 38.70, 39.20, 40.03, 41.04, 41.64, 43.10, 43.20, 47.0, 53.23, 56.52, 60.56, 61.02, 61.34, 68.90, 69.48, 70.01, 70.32, 72.44, 73.59, 74.35, 75.98, 76.49, 76.88, 76.98, 77.42, 78.46, 78.79, 78.88, 79.12, 85.40, 86.27, 94.45, 96.45, 102.47, 102.98, 104.0, 152.78, 175.63. HRMS (ESI): m/z calc.for C44H70O23 [M-H]: 965.4230; found 965.5040.

1. Rebaudioside A is a glycoside from the stevia plant. Rebaudioside A is used as a sweetener with sweetness about 300 times that of sugar with negligible effect on blood glucose. As a result, Rebaudioside A is known for its application in treatment of many diseases like diabetes, high blood pressure and weight loss in some places. It is used as a food additive and in dietary supplements.
2. Natural sweetener to diabetics and others on carbohydrate-controlled diets