阅读说明：本技术 一种葡萄糖基甜菊糖苷的制备方法 (Preparation method of glucosyl stevioside ) 是由 姜维强 李正华 李广馨 祁飞 陆晓雨 于 2020-05-25 设计创作，主要内容包括：本发明涉及一种葡萄糖基甜菊糖苷的制备方法,其特征在于：(1)甜菊提取物：环糊精按质量比1：1-2混合,配成10%-60%的混合溶液,40-80℃下,搅拌预热1-3h生成悬浊液；(2)加入环糊精糖基转移酶,控制其为环糊精质量的2-10%,40-80℃、100-300rpm/min下搅拌反应1-96h,升温至105℃以上持续30-60min；(3)反应液用脱色树脂进行脱色,脱色后溶液用阴阳离子交换树脂进行脱盐,脱盐后溶液使用大孔吸附树脂去除非二萜化合物,用水洗涤柱,用20-70％乙醇进行解吸,收集乙醇解吸液；(4)乙醇解吸液进行蒸发浓缩,控制甜菊糖质量浓度在60%以上,喷雾干燥即可。本发明优点：本发明转化率90%以上,糖基供体残留量在20%以下；反应温度较低,搅拌速率低,工艺简单能耗较低。(The invention relates to a preparation method of glucosyl stevioside, which is characterized by comprising the following steps: (1) stevia extract: the cyclodextrin is mixed according to the mass ratio of 1: 1-2, preparing a 10% -60% mixed solution, and stirring and preheating for 1-3h at 40-80 ℃ to generate a suspension; (2) adding cyclodextrin glycosyltransferase, controlling the mass of cyclodextrin to be 2-10%, stirring at 40-80 ℃ and 300rpm/min for reaction for 1-96h, and heating to 105 ℃ for 30-60 min; (3) decolorizing the reaction solution with decolorizing resin, desalting the decolorized solution with anion and cation exchange resin, removing non-diterpene compounds from the desalted solution with macroporous adsorbent resin, washing the column with water, desorbing with 20-70% ethanol, and collecting ethanol desorption solution; (4) evaporating and concentrating the ethanol desorption solution, controlling the mass concentration of the stevioside to be more than 60%, and performing spray drying. The invention has the advantages that: the conversion rate of the invention is more than 90%, and the residual quantity of glycosyl donor is below 20%; the reaction temperature is lower, the stirring speed is low, the process is simple and the energy consumption is lower.)

1. A preparation method of glucosyl stevioside is characterized by comprising the following steps:

(1) and (2) mixing the stevia extract: the cyclodextrin is mixed according to the mass ratio of 1: 1-2, adding water to prepare a mixed solution with the mass concentration of 10% -60%, then placing the mixed solution at 40-80 ℃, and preheating for 1-3 hours under the stirring condition to generate suspension;

(2) adding cyclodextrin glycosyltransferase into the suspension, controlling the mass of the cyclodextrin glycosyltransferase to be 2-10% of that of cyclodextrin, then placing the mixture at 40-80 ℃, stirring and reacting the mixture at the rotating speed of 300rpm/min of 100-80 ℃ for 1-96h, continuously heating the mixture to more than 105 ℃ for 30-60min, and inactivating and denaturing the cyclodextrin glycosyltransferase through continuous high temperature to stop the reaction;

(3) after the reaction is finished, decoloring the reaction solution by using decoloring resin, collecting the decolored solution, desalting the decolored solution by using anion and cation exchange resin, collecting the desalted solution, removing non-diterpene compounds from the desalted solution by using macroporous adsorption resin, washing a column by using distilled water or deionized water after adsorption operation, desorbing by using ethanol with the volume fraction of 20-70%, and collecting the ethanol hydrolysis imbibition containing the diterpene compounds;

(4) evaporating and concentrating the ethanol hydrolysis liquid containing diterpene compounds, controlling the mass concentration of stevioside in the concentrated solution to be more than 60%, continuously carrying out spray drying on the concentrated solution, collecting the dried product in a powder shape, and packaging to obtain the finished glucosyl glycoside product.

2. The method of claim 1, wherein the method comprises the steps of: the total glycoside content in the stevia extract obtained in the step (1) is more than or equal to 90 wt%, wherein the stevioside accounts for 40-60 wt%, and the rebaudioside A accounts for 30-50 wt%.

3. The method of claim 1, wherein the method comprises the steps of: the cyclodextrin in the step (1) is one of alpha cyclodextrin, beta cyclodextrin and gamma cyclodextrin or any mixture thereof.

4. The method of claim 1, wherein the method comprises the steps of: the reaction temperature in the step (2) is preferably 55-65 ℃, the reaction time is preferably 24-50h, and the stirring speed is preferably 180-240 rpm/min.

5. A process according to any one of claims 1 to 4, wherein the reaction mixture comprises: the macroporous adsorption resin in the step (3) is a resin product which has higher selectivity and only adsorbs diterpene compounds or a commercialized resin product which can have higher selectivity and only adsorb diterpene compounds.

Technical Field

The invention belongs to the field of biosynthesis and novel sweeteners, and particularly relates to a preparation method of glucosyl stevioside.

Background art:

recognizing that many diseases are associated with the consumption of high-sugar foods and beverages, alternatives to sucrose are receiving increasing attention. However, many artificial sweeteners, such as sodium cyclamate, cyclamate and saccharin, are banned or restricted in some countries due to safety concerns. As a result, non-caloric sweeteners of natural origin are becoming increasingly popular. Various diterpene glycosides are produced in stevia leaves, and have high-intensity sweetness and characteristics of being not easily absorbed by human bodies, so that the natural sweetener stevioside extracted from the dry leaves of stevia can be an excellent product for replacing cane sugar.

The natural sweeteners steviol glycosides are a class of extracts with a common aglycone (steviol) and differ by the number and type of carbohydrate residues at the C13 and C19 positions, the major glycosides found in stevia leaves being rebaudioside a (2-10%), stevioside (2-10%) and rebaudioside C (1-2%). Other glycosides, such as rebaudioside B, D, E and F, steviolbioside and rubusoside, are found at much lower levels (approximately 0-1%).

Rebaudioside a in the natural sweetener steviol glycoside principal has the least astringency, least bitterness and the least lingering aftertaste and therefore has the most favorable sensory attributes, however, even in a highly purified state, rebaudioside a still has undesirable taste attributes such as bitterness, sweet aftertaste, licorice flavor, and the like. These undesirable taste attributes in stevia sweeteners have been one of the major obstacles in commercialization, affecting the taste quality of stevia, and limiting its broader use.

The glucosyl stevioside is prepared by using the natural stevioside as a raw material and carrying out polymerization reaction on components such as stevioside, rebaudioside A and the like in the stevioside and glucosyl by methods such as enzyme catalysis and the like, the sweetness multiple of the stevioside is higher than that of sucrose by 100-250 times, bad mouthfeel such as bitter taste, sweet aftertaste, liquorice taste and the like of the natural stevioside is greatly improved, and the stevioside has excellent taste quality and is widely applied.

At present, a biological enzyme method is used for producing glucosyl stevioside, but because the stevioside raw material extracted from plants has low purity, and the influence of the variety, structure and the like of glycosyl donors causes the enzyme modified stevioside to generally have the defects of low conversion rate, more glycosyl donor residues, poor taste and color, complex operation steps, resource waste and the like.

Disclosure of Invention

The invention aims to overcome the defects of low conversion rate, more glycosyl donor residues, poor taste and color, complicated operation steps, resource waste and the like in the process of producing glucosyl stevioside by using the conventional biological enzyme method.

In order to achieve the purpose, the technical scheme adopted by the invention is as follows:

a preparation method of glucosyl stevioside is characterized by comprising the following steps:

(1) and (2) mixing the stevia extract: the cyclodextrin is mixed according to the mass ratio of 1: 1-2, adding water to prepare a mixed solution with the mass concentration of 10% -60%, then placing the mixed solution at 40-80 ℃, and preheating for 1-3 hours under the stirring condition to generate suspension;

(2) adding cyclodextrin glycosyltransferase (CGTase) into the suspension, controlling the mass of the cyclodextrin glycosyltransferase to be 2-10% of that of cyclodextrin, then placing the mixture at 40-80 ℃ and stirring the mixture at the rotating speed of 100-300rpm/min for reaction for 1-96h, continuously heating the mixture to more than 105 ℃ for 30-60min, and inactivating the cyclodextrin glycosyltransferase (CGTase) at a continuous high temperature to stop the reaction;

(3) after the reaction is finished, decoloring the reaction liquid by using decoloring resin (Weihua SHD-806), collecting the decolored solution, desalting the decolored solution by using anion-cation exchange resin (001 × 8 macroporous strong acid styrene cation exchange resin; 201 × 7 macroporous strong base styrene anion exchange resin) continuously, collecting the desalted solution, removing non-diterpene compounds (such as a plurality of sequentially connected columns filled with macroporous adsorption resin) by using macroporous adsorption resin, washing the columns by using distilled water or deionized water after the adsorption operation, desorbing by using 20-70% volume fraction ethanol, and collecting the ethanol hydrolysis imbibition containing diterpene compounds (stevioside);

(4) evaporating and concentrating the ethanol hydrolysis liquid containing diterpene compounds (stevioside), controlling the mass concentration of the stevioside in the concentrated solution to be more than 60%, continuously carrying out spray drying on the concentrated solution, controlling the inlet temperature (110-.

Further, the total glycoside content in the stevia extract in the step (1) is more than or equal to 90 wt%, wherein the stevioside accounts for 40-60 wt%, and the rebaudioside A accounts for 30-50 wt%.

Further, the cyclodextrin in the step (1) is one of alpha cyclodextrin, beta cyclodextrin and gamma cyclodextrin or any mixture thereof.

Further, the reaction temperature in the step (2) is preferably 55-65 ℃, the reaction time is preferably 24-50h, and the stirring speed is preferably 180-240 rpm/min.

Further, the macroporous adsorption resin in the step (3) is a resin product with higher selectivity for adsorbing diterpene compounds (stevioside) only or a resin product which is commercialized and can adsorb diterpene compounds (stevioside) only with higher selectivity.

The invention has the beneficial effects that:

1. the invention has higher conversion rate to the stevioside raw material, and the conversion rate of components such as stevioside, rebaudioside A and the like in the stevioside raw material can reach more than 90 percent through one-time reaction;

2. the residue of glycosyl donor in the system after the reaction is less and is below 20 percent of mass fraction;

3. the invention has the advantages of lower reaction temperature, low stirring speed, simple process and lower energy consumption, can realize concentration, recovery and cyclic utilization of the used solvent (ethanol and the like), realizes lower environmental pollution and higher economic benefit, and provides a reliable method for producing high-quality glucosyl stevioside.

Detailed Description

A preparation method of glucosyl stevioside comprises the following specific implementation steps: