Enzymatic Transglycosylation Of Stevioside And Application Of Steviol Glycosides

Stevioside(St)is a natural,high-intensity and non-nutritive sweetener,but the bitter aftertaste limits its application,enzymatic transglycosylation could improve the edulcorant quality.In this paper,transglycosylation of St was carried out with cyclodextrin glucosyltransferase Toruzyme 3.0L,the effects of glycosyl donor,organic solvent and microwave irradiation on the glycosylation reaction were investigated.At the same time,combination of steviol glycosides and soybean protein(SPI)was carried out and the interface and emulsifying properties of the complex systems were investigated to study the application of steviol glycosides.The specific research contents and conclusions are as follows:1.The substrate specificity was investigated by measuring St conversion and product yiled.It showed that waxy corn starch,corn starch,soluble starch,β-cyclodextrin(CD)and α-CD could be used for transglycosylation with a successive increase of St conversion and a successive decrease of mono-grafted product yiled.By studying the effect of starch branching degree on the transglycosylation and the affinity of Toruzyme 3.0L on the three kinds of starches,it showed that with the decrease of starch branching degree,St conversion increased,the yield of low-grafted product decreased,and the yield of high-grafted product rised.2.The effect of organic solvent on the transglycosylation was tested,it was found that the organic solvent could change the secondary structure of Toruzyme 3.0L,which had some influences on transglycosylation of St.When ethanol,tert-butanol and ethylacetate were added,St conversion decreased,the mono-grafted product of better taste increased,bi-grafted and high-grafted products were reduced.When petroleum ether was added,St conversion increased,however the mono-grafted product decreased and high-grafted products were improved.3.By analyzing the glycosylation of St with α/β-CD and starch,the pathways of glycosylation under conventional heating and microwave heating were investigated.Under conventional heating,when CD was used as glycosyl donor,CD was first coupled to form a reducing sugar,and then directly reacted with St;with starch as glycosyl donor,it showed that reducing sugar produced from both CD coupling and starch hydrolysis could react with St.Microwave irradiation did not dramatically affect the transglycosylation pathway of starch with St,but it remarkably affected the transglycosylation pathway of α/β-CD onto St.Unlike the case under conventional heating,the transformation between α-CD and β-CD was dramatically declined by microwave.In addition,it was the ring-opened intermediate cyclodextrin instead of the(intermediate of)reducing sugar performed as the main glycosyl donor.4.Different heating methods had effects on the reaction rate and product composition.The reaction under microwave irradiation was accelerated up to 9.3 times compared to that under the conventional heating.The product under microwave irradiation was also mainly composed of mono-and di-glucosylated stevioside,which had better edulcorant quality.Microwave irradiation did not enhance the formation of high grafted steviosides unless a very high enzyme loading(1000 U/g stevioside)applied,which only enhanced the formation of tri-glucosylated steviosides.With α/β-CD as glycosyl donor,highly grafted products diminished under microwave irradiation.5.With natural and enzymatically modified St as model compound,the interface properties of steviol glycosides and SPI mixtures were determined,and the effect of mixtures on the stability of emulsion was investigated thorugh zeta potential and average particle size.The mixtures could effectively reduce the oil-water interfacial tension and air-water surface tension,and enhance the foaming performance and foam stability.Compared to a single SPI or steviol glycosides,the mixtures could significantly increase the thermal stability of the emulsion,reduce the particle size,and enhance the p H stability and electrolyte stability to a certain extent.Substantial changes did not occur during the 120 days at room temperature,the stable emulsion system is characterized by ease of preparation and high storage stability.