Studies On Methods Of Determination Of Stevia Glycosides And Their Application

Stevia glycosides, a kind of mixture which contains eight diterpene glycosides present in stevia rebaudiana Bertoni. As a new natural sweetener, it is low caloric, high sweet and notoxicty, so it is a very perfect substitute for sucrose. Reliable separation and determination of the stevia glycoside is crucial, because the taste of each Stevia glycoside is different. This paper studied the method to detect and separate the stevia glycosides, a new capillary zone electrophoresis (CZE) separation method was developed for determination and under the capillary zone electrophoresis (CZE) method developed, the concentration of stevia elution by nanofiltration and the foculation of stevia leaves were studied. The main contents of the sduty were given as follows:1. A capillary zone electrophoresis (CZE) separation was developed for simultaneous determination of rebaudioside A and stevioside which are the main component in Stevia glycoside. The effects of separation voltage, injection time, the kind of running buffer, concentration and pH of the buffer and modifiers were investigated. Single-factor tests were carried out to screen the critical influencing factors for the separation and the optimal conditons were obtained by a central composite design. The best separation of rebaudioside A and stevioside could be obtained using 50 mM borate buffer (pH=10.10), containing 23% acetonitrile and 7% methanol as modifiers.2. Under the optimum conditions, RA and St could be separated within 15 min. The calibration curves had good linearity in the range of 0.05~ 10. 0 mg/mL for RA and 0.06~10.0 mg/mL for St, with the detection limits of 0.01 mg/mL for RA and 0.02mg/mL for St. The method was successfully used in the analysis of food samples. The results were given as follows: 2.8% RA and 10.2% St in the stevia leaves, the average recovery(n=3) is 95.2% for RA and 101.7% for Sto The Tianwang stevia sample contains 1.31 mg/g RA and 3.57 mg/g St, the average recovery(n=3) is 98.2% for RA and 96.3% for St. The Pu'er stevia sample contains 1.53 mg/g St, the average recovery(n=3) is 97.5% for St.3. The concentration of stevia elution by nano filtration were studied. The nanofiltration was used firstly to ameliorate technics of stevia glucoside. MWCO 200 nanofiltration membrane was used to concentrate Stevia glycosides, the permeation can be recycled to elute stevia glycosides in the adsorption column, and the inspissation go to the next technics. The average retention coefficient of stevia glycosides is 99.8% and of methanol is 1.6%, with the pressures vary from 0.2Mpa to 0.7Mpa.The stevia eluent containing l%(w%) stevia was concentrated to 2.1% at 0.7 MPa with an average permeation flux of 19.5 L/m2 ? h. The recovery of methanol is 45%.4. The capillary zone electrophoresis (CZE) method was used to detect samples in the process of flocculation. Ferric chloride was use as flocculant, The best floccultate craft is : 70°C, 40 min in heated water, pH=5.0, the dosage of Ferric chloride is 5.2 mg/mL.