CN115536715A - Method for extracting stevioside RD - Google Patents

Abstract

A method for extracting rebaudioside RD comprises the following steps: providing a solution comprising steviol glycoside RD, the glycoside solution having a pH of 5 to 6; contacting the macroporous neutral adsorption resin with a glucoside solution derived from stevia at the temperature of 20-25 ℃, keeping the temperature for 5-15min, then slowly heating to the temperature of 50-60 ℃, keeping the temperature for 15-20min, and then slowly cooling to the temperature of 30-40 ℃; contacting the macroporous neutral adsorption resin with adsorbed glycoside from step S3 with the elution solvent of ethanol and water to obtain an enriched solution enriched in steviol glycoside RD; separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD; crystallizing the crystallized wet solid in the S6 by taking a methanol solution with the methanol volume fraction of 40-70% as a crystallization solvent, and filtering and collecting crystals; and drying the crystal at low temperature under reduced pressure, and then crushing and filtering to obtain the high-content stevioside RD powder. Through the scheme, the stevioside RD is produced by enrichment, so that the operation steps in the RD production process are greatly reduced, and the production efficiency is improved.

Description

A method for extracting steviol glycoside RD

technical field

The invention belongs to the technical field of sweetener purification, and in particular relates to a method for extracting steviol glycoside RD.

Background technique

Stevia (scientific name: Stevia rebaudiana (Bertoni) Hemsl.) is a perennial herb of the Compositae family and the genus Stevia, native to the alpine grasslands at the border of Paraguay and Brazil in South America. Stevia likes to grow in a warm and humid environment and is sensitive to light. Its leaves contain 6-12% of steviol glycosides (by dry weight). High-purity steviol glycosides are white or light yellow powder, which is a low-calorie It is a natural sweetener with high sweetness and is one of the most potential natural sweeteners in the food industry in the future. Extracted from the leaves and stems of the Compositae plant stevia, it is generally a mixture of several steviol glycosides, mainly composed of Stevioside (St) stevioside and Rebaudioside A (RA) rebaudioside A; stevioside Compared with sucrose, its sweetness is 200~350 times that of sucrose, its calories are 1/300 of that of sucrose, and its sweetness is relatively pure. It is an ideal substitute for sucrose. Steviol glycoside RA, which is currently used in the largest amount on the market, has a refreshing and pure sweetness, and has a very wide range of applications in the food industry. However, its further market expansion is limited due to its slightly bitter aftertaste and metallic taste.

As the third sugar source, steviol glycoside has the irreplaceable advantages of cane sugar and beet sugar: pure natural safety, heat stability, zero calories, no effect on blood sugar, non-fermentable, acid-base stability, and it is more stable than ordinary sucrose 150~300 times sweeter, no browning reaction, no fat and carbohydrates, anti-caries. It also has special medicinal and health benefits, so it is favored by people at home and abroad, especially those with obesity, diabetes, and metabolic syndrome. Due to the increase in consumer demand and applications, it has achieved considerable development.

Rebaudioside D (referred to as RD) is one of dozens of steviol glycosides naturally occurring in stevia rebaudiana, and its content accounts for 0.3%~0.8% of steviol glycosides. , also has the advantage of pure taste, and is the component of steviol glycosides that tastes closest to sucrose. The extract obtained from fresh leaves contains a large amount of impurities. In the prior art, the extract can be purified by resin adsorption, recrystallization and other methods. The method of adsorption and analysis can enrich but the purity is not high. The purity of the crystal is high, but the operation is complex and consumes a lot of solvents. It is used in industrial production, which greatly reduces the economic benefits of the product.

Contents of the invention

In order to solve the above technical problems, the present invention provides a method for extracting steviol glycoside RD. In the present invention, the temperature rise and temperature drop operations are performed during the adsorption process, so that the solute is adsorbed by the resin to shift the adsorption-analysis equilibrium, reducing the adsorption of some impurities, adjusting the structure of molecular adsorption, and increasing the concentration of steviol glycosides in the enrichment solution. content, it provides better raw materials for crystallization, and the purity of primary crystallization can reach 96.7%.

Concrete technical scheme of the present invention is as follows:

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5 to 6;

S2: Provide macroporous neutral adsorption resin;

S3: Contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 20~25°C, heat it for 5~15 minutes, then slowly raise the temperature to 50~60°C, keep it for 15~20 minutes, and then slowly cool down to 25~ 40°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 20-35% w/w ethanol and 65-80% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 with methanol alcohol solution with a volume fraction of 40-70% of methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In the above-mentioned technical scheme of the present invention, first at a lower temperature, the macroporous neutral resin is contacted with the solution containing steviol glycoside RD to promote the occurrence of adsorption, and then in the heating and cooling process, part of the heating process The adsorbed solute will be decomposed. At this pH 5~6, the binding ability of steviol glycoside RD to the resin is stronger, so the impurities with low binding strength and part of steviol glycoside RD will be desorbed; then the temperature is lowered so that the solute is adsorbed by the resin Afterwards, the adsorption-analysis balance is shifted, so that the resin continues to adsorb part of the steviol glycoside RD; through this method, the adsorption of some impurities is reduced, the structure of molecular adsorption is adjusted, and the content of the steviol glycoside in the enriched solution is increased, providing a good source for crystallization. For better raw materials, the purity of primary crystallization can reach up to 96.7%.

As an optimization of the above technical solution of the present invention, in step S3, the heating process lasts for 10-15 minutes, and the cooling process lasts for 10-30 minutes.

As a preference of the above-mentioned technical solution of the present invention, the eluting solvent in step S4 contains 25-35% w/w ethanol.

As a preference of the above-mentioned technical solution of the present invention, the crystallization solvent in step S7 includes 55-65% methanol.

As a preference of the above-mentioned technical solution of the present invention, the method further includes intentionally adjusting the pH of the glycoside solution to 5-6.

As a preference of the above-mentioned technical solution of the present invention, in step S4, the pH of the eluent in the above-mentioned medium is 3-4.

As a preference of the above-mentioned technical solution of the present invention, in step S1, the glycoside solution is selected from stevia extract after extracting stevioside RA and ST.

As a preference of the above-mentioned technical solution of the present invention, in step S1, it includes adding an appropriate amount of salt to the glycoside solution.

In the above technical solution of the present invention, the inventor found that some steviol glycoside RD remained in the purified solution, which would cause the product yield to not increase. The inventor did a lot of experiments, first by reducing the final temperature of the cooling process temperature, the structure shows that although the method improves the yield, it reduces the purity of the product to a certain extent, which is not ideal; finally the inventors found that by adding a certain amount of salt to the glycoside solution, it can be very good. The yield of purifying glycoside RD in the glycoside solution can obtain better purity.

As a preference of the above-mentioned technical solution of the present invention, the salt is an inorganic salt selected from one of sodium chloride, sodium sulfate, and ammonium sulfate.

As a preference of the above-mentioned technical solution of the present invention, the addition amount of the inorganic salt is 1% to 5% of the mass of the detection solution.

In summary, the present invention has the following effects:

1. The present invention makes the adsorption-analysis equilibrium shift after the solute is adsorbed by the resin by performing temperature-raising and cooling-down operations during the adsorption process, which reduces the adsorption of some impurities, adjusts the structure of molecular adsorption, and improves the concentration of the enriched solution. The content of steviol glycosides provides better raw materials for crystallization, and the purity of one crystallization can reach 96.7%;

2. The process uses water, ethanol, etc. as solvents, without the use of any volatile organic solvents, and all DES and water can be recovered and recycled, so it fully meets the requirements of green environmental protection and clean technology.

3. After the RD content of the product during the extraction and separation process is increased, better raw materials are provided for the further production of commercial high-purity stevioside, which can significantly increase the yield and reduce costs, and the economic effect is remarkable;

4. The present invention increases the yield of glycoside RD without reducing the purity of RD by adding inorganic salts to the glycoside solution.

detailed description

In order to make the purpose, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Those who do not indicate the specific conditions in the examples are carried out according to the conventional conditions or the conditions suggested by the manufacturer. The reagents or instruments used were not indicated by the manufacturer, and they were all conventional products that could be purchased from the market. The method for extracting stevioside provided by the embodiments of the present invention will be described in detail below.

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a glycoside solution containing steviol glycoside RD, the glycoside solution is obtained by dissolving the dry mother liquor powder with ethanol aqueous solution, wherein the dry mother liquor powder is obtained by drying the stevia extract after extracting most of the steviol glycosides RA and ST; The pH of the glycoside solution is between 5 and 6. If it is found that the pH range is not within this range after dissolving, it can be adjusted by adding a small amount of acid, such as hydrochloric acid; in the following examples, the method for obtaining the glycoside solution can be Directly weigh 10g of mother liquor dry powder, dissolve it in 100ml ethanol aqueous solution, then add inorganic salt to the solution and adjust the pH of the solution with hydrochloric acid;

S2: Provide macroporous neutral adsorption resin, you can choose divinylbenzene macroporous resin, such as SP70, SP710, etc., the following are SP70 resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 35°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent is 4, which reduces the adsorption strength of RD by the resin, improves the efficiency of analysis, and improves the adsorption strength of other components at the same time, making it difficult to contain 35% w/ w ethanol and 65% w/w water were eluted.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution, and by adding inorganic salts to the glycoside solution, the adsorption capacity of the resin can be improved without reducing the impurities of the purified product To ensure the yield of the product.

Example 1

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 35°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Example 2

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 30°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Example 3

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 6;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 30°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Example 4

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 30°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 with methanol alcohol solution with a volume fraction of 40% as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

Example 5

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 35°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Example 6

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 32.5°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 with methanol alcohol solution with a volume fraction of 40% as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Example 7

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 27.5°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 with methanol alcohol solution with a volume fraction of 40% as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Example 8

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 25°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Example 9

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 50°C, keep it for 15 minutes, and then slowly cool down to 30°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Example 10

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from Stevia rebaudiana at 25°C, heat it for 15 minutes, then slowly raise the temperature to 60°C, keep it for 15 minutes, and then slowly cool down to 30°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Example 11

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 30°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium chloride accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Example 12

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 25°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding ammonium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Example 13

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 30°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 1% of the mass of the glycoside solution to the glycoside solution.

Example 14

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 30°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 2% of the mass of the glycoside solution to the glycoside solution.

Example 15

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 30°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 3% of the mass of the glycoside solution to the glycoside solution.

Example 16

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 30°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 5% of the mass of the glycoside solution to the glycoside solution.

Comparative example 1

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: At 25°C, the macroporous neutral adsorption resin is contacted with a glycoside solution derived from stevia for adsorption;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Comparative example 2

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 60°C, keep it for 15 minutes, and then slowly cool down to 22.5°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 4% of the mass of the glycoside solution to the glycoside solution.

Comparative example 3

A method for extracting steviol glycoside RD, comprising the following steps:

S1: Provide a solution containing steviol glycoside RD, the pH of the glycoside solution is 5.5;

S2: Provide macroporous neutral adsorption resin;

S3: contact the macroporous neutral adsorption resin with the glycoside solution derived from stevia at 25°C, heat it for 15 minutes, then slowly raise the temperature to 55°C, keep it for 15 minutes, and then slowly cool down to 30°C;

S4: providing at least one elution solvent comprising a mixture of ethanol and water, the elution solvent comprising 35% w/w ethanol and 65% w/w water;

S5: contacting the macroporous neutral adsorption resin with adsorbed glycosides from step S3 with the elution solvent of ethanol and water to obtain an enrichment solution enriched in steviol glycoside RD;

S6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

S7: Crystallize the crystalline wet solid in S6 by using methanol alcohol solution with a volume fraction of 40% methanol as the crystallization solvent, and collect the crystals by filtration; the crystals are dried under reduced pressure at low temperature, crushed and filtered to obtain high-content steviol glycoside RD powder.

In step S3, the heating process lasts for 15 minutes, and the cooling process lasts for 10 minutes.

In step S4, the pH of the eluent in the said medium is 4.

In step S1, it also includes adding sodium sulfate accounting for 6% of the mass of the glycoside solution to the glycoside solution.

For the above-obtained steviol glycoside RD, the inventor carried out a purity test and a yield. The specific operation process is as follows:

1. Purity test:

Chromatographic conditions: C18 column reverse phase chromatography column (250mm×4.6mm), particle size 5μm; column temperature: room temperature 40°C; UV detector: 210nm detection wavelength; mobile phase: acetonitrile: sodium phosphate buffer = 32:68; Flow rate: 1.0mL/min; injection volume: 2μL~10μL.

Preparation of mixed standard solution: weigh appropriate amount of stevioside, rebaudioside A, rebaudioside B, rebaudioside C, rebaudioside C, rebaudioside D, rebaudioside F, The standard substances of dulcoside A, rubusoside, and steviolbioside were placed in the same volumetric flask, dissolved completely with acetonitrile aqueous solution and then constant volume to obtain a mixed standard solution. The mixed standard solution was used to determine the relative retention times of the 9 glycosides.

Preparation of standard solution: Weigh 0.05g of stevioside standard and rebaudioside A standard respectively, accurate to 0.001g, place them in 50mL volumetric flasks, dissolve with acetonitrile aqueous solution and dilute to the mark to obtain stevioside standard solution and rebaudioside A standard solution.

Preparation of sample solution: Weigh 0.1g sample (the mother liquor test is the dried product of the mother liquor), accurate to 0.001g, put it in a 50mL volumetric flask, dissolve it with acetonitrile aqueous solution and dilute to the mark to obtain a sample solution .

Determination: Under the reference chromatographic conditions, carry out chromatographic analysis on the mixed standard solution, standard solution and sample solution respectively. Compare the chromatogram of the sample solution with the chromatogram of the mixed standard solution to determine the peaks corresponding to each component in the chromatogram of the sample solution. Record stevioside, rebaudioside A, rebaudioside B, rebaudioside C, rebaudioside D, rebaudioside F, dulcoside A, rubusoside, stevioside in the sample solution chromatogram The peak area of glycoside and the peak area of stevioside and rebaudioside D in the standard solution chromatogram.

2. Yield detection

Weigh the quality of steviol glycoside RD powder and then calculate its yield by the following formula:

Yield = RD powder mass × RD powder content / (mother liquid dry powder mass × dry powder RD content) × 100%.

The test results are as follows:

The detection data of embodiment 1,2,3,4 and comparative example 1 are as follows:

It is not difficult to see from the above table that the enrichment effect of Examples 1-4 on the target product RD is better than that of the comparative example, and the purity after crystallization is also higher; it can also be seen that adding inorganic salts can indeed reduce the purity of the product without affecting the purity of the product. Improve the yield of RD, so purifying stevioside RD through this technical scheme can indeed reduce the steps of operation and improve the efficiency of production.

The detection data of embodiment 2,5,6,7,8 and comparative example 2 are as follows:

It is not difficult to find out from the above table that along with the reduction of the final temperature in the cooling process in Examples 2, 5, 6, 7, 8 and Comparative Example 2, the yield of product RD will rise to a certain extent, but the product The purity of RD powder will drop a lot.

The detection data of embodiment 2,9,10 are as follows:

It is not difficult to see from the above table that when the maximum temperature in the heating process changes from 50 to 60°C, it has little effect on the product yield and purity, and the maximum temperature of yield and purity is the highest at 55°C.

The detection data of embodiment 2,11,12 are as follows:

As can be seen from the above table, among sodium chloride, ammonium sulfate and sodium sulfate, the purity and yield of the product obtained by using sodium sulfate are the highest.

The detection data of embodiment 2,13,14,15,16 and comparative example 3 are as follows:

The above description is only a preferred embodiment of the present invention, and does not limit the present invention in any form. Although the present invention has been disclosed as above with preferred embodiments, it is not intended to limit the present invention. Anyone familiar with this field Those skilled in the art, without departing from the scope of the technical solution of the present invention, can use the technical content disclosed above to make some changes or modify them into equivalent embodiments with equivalent changes, but if they do not depart from the technical solution of the present invention, according to the technical content of the present invention Technical Essence Any simple modifications, equivalent changes and modifications made to the above embodiments still fall within the scope of the technical solution of the present invention.

Claims (10)

1. A method for extracting rebaudioside RD comprises the following steps:

s1: providing a glycoside solution comprising steviol glycoside RD, the glycoside solution having a pH of from 5 to 6;

s2: providing macroporous neutral adsorption resin;

s3: contacting macroporous neutral adsorption resin with a glucoside solution derived from stevia at the temperature of between 20 and 25 ℃ for adsorption, keeping the temperature for 5 to 15min, then slowly heating to between 50 and 60 ℃, keeping the temperature for 15 to 20min, and then slowly cooling to between 25 and 40 ℃;

s4: providing at least one elution solvent comprising a mixture of ethanol and water;

s5: contacting the macroporous neutral adsorption resin with adsorbed glycoside from step S3 with the elution solvent of ethanol and water to obtain an enriched solution enriched in steviol glycoside RD;

s6: separating the enriched solution in S5 to obtain a crystalline wet solid containing steviol glycoside RD;

s7: crystallizing the crystallized wet solid in the S6 by taking a methanol solution with the volume fraction of 40-70% as a crystallization solvent, and filtering and collecting crystals; and drying the crystals at low temperature under reduced pressure, and crushing and filtering to obtain the high-content stevioside RD powder.

2. The method for extracting the stevioside RD according to the claim 1, wherein in the step S3, the temperature rising process lasts for 10 to 15min, and the temperature lowering process lasts for 10 to 30min.

3. A method of extracting steviol glycoside RD according to claim 1, characterized in that the elution solvent in step S4 contains 25 to 35% w/w ethanol.

4. A method of extracting stevioside RD as claimed in claim 1, characterized in that the crystallization solvent in step S7 contains 55 to 65% methanol.

5. A method of extracting steviol glycoside RD according to claim 1, characterized in that it further comprises deliberately adjusting the pH of the glycoside solution to 5 to 6.

6. The method for extracting rebaudioside RD according to claim 1, wherein in step S4, the pH of the eluent is 3 to 4.

7. A method of extracting stevioside RD as claimed in claim 1, wherein in step S1, the glucoside solution is selected from stevia extract after extraction of the over-sweet inulin RA, ST.

8. A method of extracting steviol glycoside RD according to claim 1, comprising in step S1 adding an appropriate amount of salt to the glycoside solution.

9. A method of extracting steviol glycoside RD according to claim 8, wherein the salt is an inorganic salt selected from one of sodium chloride, sodium sulfate, ammonium sulfate.

10. A method of extracting steviol glycoside RD according to claim 9, wherein the amount of inorganic salt added is 1% to 5% by mass of the bouquet solution.