Preparation method of mogroside
Through low-temperature alkaline water flash extraction, composite column chromatography and biological fermentation, the problems of many impurities and loss of nutrients during the extraction of Luohan Fruit Sweet Glycoside were solved, and high-purity Luohan Fruit Sweet Glycoside was prepared, which is suitable for food and beverage fields.
Patent Information
- Application Number
- CN202311862341.5
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2023-12-29
- Publication Date
- 2025-07-01
AI Technical Summary
During the extraction process, existing Luohanbulin V products have problems such as many impurities, loss of nutrients, impure white color and poor taste. Especially during the purification process, the nutrients of Luohanbulin itself are removed, which affects the product quality.
Fresh Luohan fruit pulp is flicked with low-temperature alkaline water, combined with composite column chromatography and biofermentation, eluting through high-active yeast fermentation and gradient solvent, and crystallization with low-temperature stirring to prepare high-purity Luohan fruit sweet glycoside.
The product of Rahan Fruit Sweet Glycoside is prepared with few impurities, high content of rhubarb V, white color and good taste, retains the nutrients of the flesh and is suitable for food and beverage fields.
Smart Images
Figure CN120230168A_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to mogroside, and particularly to a preparation method of mogroside. Background Art
[0002] Existing mogroside V products mainly include whole fresh fruits and peeled pulp. The products are mostly prepared by water or alcohol extraction, followed by treatment with macroporous resin and ion exchange resin to obtain mogroside products. Most of them are 50% light white mogroside V products. For example, a production method of mogroside V provided by CN201811250821.51 includes the following steps: 1) Pretreatment: Immediately heat the raw material of Siraitia grosvenorii by instantaneous high temperature after freezing. After freezing the raw material of Siraitia grosvenorii at -25°C to -10°C for 1 to 2 hours, immediately heat it at 105°C to 120°C for 5 to 50 seconds; 2) Extraction: Peel the pretreated Siraitia grosvenorii, crush the pulp, and perform countercurrent circulation extraction with water to obtain an extraction solution; 3) Filtration: Centrifuge the extraction solution, and filter the supernatant obtained by centrifugation through a ceramic membrane to obtain a filtrate; 4) Adsorption and desorption: After the filtrate is adsorbed by a macroporous resin column, desorb it with an ethanol solution to obtain a desorbed solution. The macroporous resin model is selected from one of T-28, HDP-100 or DA201-H; wherein the mass concentration of the ethanol solution is 55 to 70%, and the volume is 1.5 to 2.5 BV; the ethanol solution is pre-washed before desorption. First, pre-wash it with 1 to 3 BV of an alkaline solution with a mass concentration of 2 to 6‰, then pre-wash it with 1 to 3 BV of an acidic solution with a mass concentration of 2 to 6‰, and finally wash it with 2 to 4 BV of pure water until the column passing solution is neutral; 5) Decolorization: The desorbed solution passes through a cation exchange resin and an anion exchange resin in sequence, and collect the column passing solution; 6) Concentration and drying: Concentrate and dry the column passing solution to obtain mogroside V; wherein the content of mogroside V is ≥50%.
[0003] If the processed Siraitia grosvenorii products only undergo rough treatment, the taste is not very good, and there will be precipitation or color turbidity during use. If the Siraitia grosvenorii is purified, such as decolorized by ion exchange resin, some nutrients in the Siraitia grosvenorii itself are removed during the purification process, reducing its nutritional efficacy, and the color still does not reach pure white. Summary of the Invention
[0004] The present invention provides a preparation method of mogroside, which can obtain a mogroside product with less impurities and a high content of mogroside V.
[0005] The object of the present invention is achieved by the following technical solutions: A preparation method of mogroside includes the following steps:
[0006] (1) Take fresh Siraitia grosvenorii pulp and perform flash extraction with alkaline water at a temperature of 10 - 15 degrees.
[0007] (2) The pulp processed in step (1) is placed in a flash extractor and extracted twice with a solvent. The filtrates from the two extractions are combined to obtain a total filtrate.
[0008] (3) The total filtrate is passed through a composite column chromatography and eluted with pure water. The effluent and the washing solution collected are combined and concentrated into a solution with a solids content of 7 - 10%.
[0009] (4) High - activity yeast is added to the solution obtained in step (3) and fermented at 30 - 32 °C for 4 - 6 hours. Then it is inactivated, cooled, filtered, passed through a macroporous resin chromatography, and eluted successively with pure water, acetone, and ethanol. The ethanol eluate is collected and concentrated at low temperature until the Brix of the solution is 45 - 55 BRIX to obtain a concentrated solution.
[0010] (5) Pure methanol or absolute ethanol is added to the concentrated solution obtained in step (4), sealed, and crystallized under stirring conditions for 38H - 40H. The crystals are obtained by suction filtration, washed, dissolved in pure water, concentrated to 50 - 52 BRIX, and spray - dried to obtain mogroside.
[0011] In the above - mentioned step (1), the dosage of the alkaline water is 5 - 8 times the mass of the pulp, and the mass concentration of the alkaline water is 0.5 - 1.2%. The alkaline water is a calcium hydroxide solution prepared by dissolving calcium oxide in water.
[0012] In the above - mentioned step (2), the dosage of the alkaline water is 2.5 - 4 times the mass of the pulp. The motor speed of the flash extractor is 10000 - 15000 R / min, and the extraction is carried out for 3 minutes, twice. Specifically, each extraction is for one minute, and there is a 2 - minute break between the two extractions.
[0013] In the above - mentioned step (3), the mixed resin of the composite column consists of alumina in the upper layer and cation resin in the lower layer. The mass ratio of alumina to cation resin is 1﹕2 - 3, and the total dosage of the mixed resin and the raw material weight ratio is 1﹕5 - 8. The dosage of pure water is 0.5 - 1 times the total mass of the mixed resin.
[0014] In the above - mentioned step (4), the addition amount of high - activity yeast is 0.5 - 1‰ of the raw material amount, and the solution is cooled to a temperature of 40 - 50 degrees. The mass ratio of the macroporous resin to the raw material is 1:4 - 5.
[0015] In the above - mentioned step (4), it is eluted with pure water until the effluent is clear and transparent, and then gradient fractional elution is carried out. The gradient fractional elution is as follows: first, elute with 15 - 27% acetone, and the elution volume is 1:2 - 4 of the resin dosage volume ratio; then elute with 50 - 58% ethanol, and the elution volume is 1:2 - 4 of the resin dosage volume ratio.
[0016] In the above - mentioned step (5), the addition amount of pure methanol or absolute ethanol is 3 - 4 times the volume of the concentrated solution. The pure water is 3 - 4 times the total mass of the crystals.
[0017] The raw material described in the present invention refers to Siraitia grosvenorii.
[0018] Compared with the prior art, the present invention has the following beneficial effects:
[0019] 1. For the pericarp and kernel of Siraitia grosvenorii, the proportion of total sugar and other nutrients is very small. Using the whole fruit for extraction will bring some lipid components, which will affect the subsequent purification and product color, and more importantly, will affect the taste during the use of the product. Therefore, the present invention preferably uses the pulp as the raw material.
[0020] 2. The present invention extracts the pulp of Siraitia grosvenorii with alkali at low temperature for a short time, and all the nutritional components of the pulp are retained in the obtained extract. The extract passes through composite alumina and cation exchange resin to effectively remove the bitter taste in the pulp and neutralize the pH, while also taking into account the decolorization effect.
[0021] 3. The extract of the present invention is subjected to biological fermentation, and the fermentation product is eluted by gradient with different solvents of macroporous resin. By separating the active ingredients and impurities, the purity of mogroside V in the pulp is optimized.
[0022] 4. The present invention uses low-temperature stirring crystallization to greatly increase the content of mogroside V, dissolves it in water, concentrates it, and sprays it to ensure that there is no solvent residue in the product.
[0023] 5. For the product prepared by the present invention, mogroside V ≥ 85%, off-white, retaining the nutritional components of the pulp, also taking into account the pure taste and good color, and can be widely applied in various aspects, such as food, beverages and other products.
[0024] 6. The process provided by the present invention is safe, environmentally friendly throughout the whole process. The pollution is less. The process is simple, has good adaptability, and is suitable for large-scale industrial production. Description of the Drawings
[0025] Figure 1 It is the HPLC chromatogram of Example 1.
[0026] Figure 2 It is the HPLC chromatogram of Example 2.
[0027] Figure 3 It is the HPLC chromatogram of Example 3. Detailed Embodiments
[0028] The specific embodiments of the present invention will be described in more detail below. These embodiments are provided to enable a more thorough understanding of the present invention and to fully convey the scope of the present invention to those skilled in the art.
[0029] Example 1
[0030] (1) Take 5000 g of fresh Siraitia grosvenorii (0.4% mogroside V), peel and remove the seeds.
[0031] (2) Only take the pulp and perform flash extraction with 6 times the amount of 0.5% alkaline water at a temperature of 10 degrees. The alkaline water is made by dissolving calcium oxide in water.
[0032] (3) Put the pulp processed in step (2) into a flash extractor, add 3 times the amount of solvent of alkaline water each time, the motor speed is 10000 R / min, extract at room temperature for 3 minutes, and extract 2 times. Each time is 1 minute. Take a 2-minute break in the middle after each extraction. Separate the filtrate, and combine the filtrates extracted 2 times to obtain the total filtrate.
[0033] (4) Pass the total filtrate through a 1000 g composite column chromatography and collect the effluent. Among them, the composite column consists of an upper layer and a lower layer. The upper layer is alumina, and the lower layer is cation resin. The mass ratio of alumina to cation resin is 1:3. After the total filtrate is fed in, wash it with 500 ML of pure water, and collect the washing solution. Combine the collected effluent and the washing solution, and concentrate it into a solution with a solid content of 7% using a vacuum concentration device.
[0034] (5) The solid content of the concentrated solution is 7%. Add 2.5 g of high-activity yeast and ferment at 30 °C for 4 hours. After fermentation is completed, heat the material to 98 °C and keep it for 5 minutes to inactivate.
[0035] (6) Then cool the material to a temperature of 40 degrees, pass it through a 200-mesh filter cloth, and then pass it through a 1250 g macroporous resin column (LK2MG of Amyco Biopharmaceutical Co., Ltd.) for chromatography. After the material enters the column, wash it with pure water until the effluent is clear and transparent, and then perform gradient fractional elution. Specifically, for gradient fractional elution, first elute with 15% acetone for 2500 ml, and then elute with 50% ethanol for 2500 ml, and collect the ethanol eluate.
[0036] (7) Using a low-temperature concentrator, under the condition of a temperature < 60 degrees, concentrate the ethanol eluate until there is no alcohol, and concentrate it to 45 BRIX. Then add 3 volumes of anhydrous ethanol, seal it, keep stirring at 4 degrees for 38 H to crystallize, filter the crystals, wash the crystals with a small amount of pure water, dissolve the crystals with 4 times the amount of pure water, concentrate to 50 BRX, remove the solvent residue, and spray dry. Obtain 12.5 g of sample, and the mogroside V is detected by HPLC to be 85.57%, see Figure 1 .
[0037] Example 2
[0038] (1) Take 5000 g of fresh Siraitia grosvenorii (0.4% mogroside V), peel and remove the seeds.
[0039] (2) Only take the pulp and perform flash extraction with 5 times the amount of alkaline water at a temperature of 15°C and a concentration of 1.2%. The alkaline water is made by dissolving calcium oxide in water.
[0040] (3) Put the pulp processed in step (2) into a flash extractor. Add 4 times the amount of solvent of alkaline water each time, with the motor speed at 15000R / min, extract at room temperature for 3 minutes, and extract 2 times, 1 minute each time. Take a 2-minute break in between each extraction. Separate the filtrate and combine the filtrates from the 2 extractions to obtain the total filtrate.
[0041] (4) Pass the total filtrate through a 1000g composite column chromatography and collect the effluent. Among them, the composite column consists of an upper layer and a lower layer. The upper layer is alumina and the lower layer is cation resin. The mass ratio of alumina to cation resin is 1:2.5. After the total filtrate has been fed in completely, elute with 500ML of pure water and collect the washing solution. Combine the collected effluent and the washing solution and concentrate them into a solution with a solid content of 7% using a vacuum concentration device.
[0042] (5) The solid content of the concentrated solution is 7%. Add 2.5g of highly active yeast and ferment at 32°C for 4 hours. After fermentation is completed, heat the material to 98°C and keep it for 5 minutes to inactivate.
[0043] (6) Then cool the material to a temperature of 50°C, pass it through a 200-mesh filter cloth, and then pass it through a 1250g macroporous resin column (LK2MG of Emmagen Biopharmaceuticals Co., Ltd.) for chromatography. After the material has been fed into the column completely, elute with pure water until the effluent is clear and transparent, and then perform gradient fractional elution. Specifically, for gradient fractional elution, first elute with 15% acetone for 5000ml, and then elute with 50% ethanol for 5000ml, and collect the ethanol eluate.
[0044] (7) Using a low-temperature concentrator, under the condition of a temperature <60°C, concentrate the ethanol eluate until there is no alcohol and concentrate it to 55BRIX. Then add 4 volumes of absolute ethanol, seal it, keep stirring at 4°C for 38H to crystallize, filter the crystals by suction, wash the crystals with a small amount of pure water, dissolve the crystals in 4 times the amount of pure water, concentrate to 50BRX, remove the solvent residue, and perform spray drying. Obtain 12.7g of sample. After HPLC detection, mogroside V is 86.71%, see Figure 2 .
[0045] Example 3
[0046] (1) Take 5000g of fresh Siraitia grosvenorii (0.4% mogroside V), remove the skin and seeds.
[0047] (2) Only take the pulp and perform flash extraction with 8 times the amount of alkaline water at a temperature of 12°C and a concentration of 0.8%. The alkaline water is made by dissolving calcium oxide in water.
[0048] (3) Put the pulp processed in step (2) into a flash extractor. Add 2.5 times the amount of solvent of alkaline water each time. The motor speed is 12,500 R / min. Extract at room temperature for 3 minutes, and extract 2 times, 1 minute each time. Take a 2-minute break in the middle after each extraction. Separate the filtrate, and combine the filtrates extracted 2 times to obtain the total filtrate.
[0049] (4) Pass the total filtrate through a 1000 g composite column chromatography and collect the effluent. Among them, the composite column consists of an upper layer and a lower layer. The upper layer is alumina, and the lower layer is cation resin. The mass ratio of alumina to cation resin is 1:3. After the total filtrate is fed in, wash it with 500 ML of pure water, and collect the washing solution. Combine the collected effluent and the washing solution, and concentrate it into a solution with a solid content of 7% using a vacuum concentration device.
[0050] (5) The solid content of the concentrated solution is 7%. Add 5 g of highly active yeast and ferment at 30 °C for 6 hours. After fermentation is completed, heat the material to 98 °C and keep it for 5 minutes to inactivate.
[0051] (6) Then cool the material to 45 °C, filter it through a 200-mesh filter cloth, and then pass it through a 1250 g macroporous resin column (LK2MG of Amerigen Biopharmaceuticals Co., Ltd.) for chromatography. After the material is fed into the column, wash it with pure water until the effluent is clear and transparent, and then perform gradient fractional elution. Specifically, for gradient fractional elution, first elute with 27% acetone for 5000 ml, and then elute with 58% ethanol for 5000 ml, and collect the ethanol eluate.
[0052] (7) Using a low-temperature concentrator, under the condition of a temperature < 60 °C, concentrate the ethanol eluate until it is alcohol-free and concentrate it to 55 BRIX. Then add 3 volumes of anhydrous ethanol, seal it, keep stirring at 4 °C for 40H to crystallize, filter the crystals, wash the crystals with a small amount of pure water, dissolve the crystals in 4 times the amount of pure water, concentrate to 50 BRX, remove the solvent residue, and spray dry. Obtain 12.9 g of sample. The mogroside V is detected by HPLC to be 85.02%, see Figure 3 .
Claims
1. A method for preparing mogroside, characterized in that, It includes the following steps: (1) Take fresh Momordica grosvenori pulp and perform flash extraction with alkaline water at a temperature of 10 - 15°C; (2) Place the pulp processed in step (1) into a flash extractor, extract twice in a solvent, and combine the filtrates from the two extractions to obtain the total filtrate; (3) Pass the obtained total filtrate through a composite column chromatography, elute with pure water, combine the collected effluent and washing solution, and concentrate to a solution with a solids content of 7 - 10%; (4) Add high - activity yeast to the solution obtained in step (3) at 30 - 32°C, ferment for 4 - 6 hours, then inactivate, cool, filter, pass through a macroporous resin chromatography, elute successively with pure water, acetone, and ethanol, collect the ethanol eluate, and concentrate it at low temperature to a Brix of 45 - 55 BRIX to obtain the concentrated solution; (5) Add pure methanol or absolute ethanol to the concentrated solution obtained in step (4), seal it, crystallize for 38H - 40H under stirring conditions, filter by suction to obtain crystals, wash them, dissolve them in pure water, concentrate to 50 - 52 BRIX, and perform spray drying to obtain mogrosides.
2. The preparation method of mogroside according to claim 1, characterized in that, In step (1), the dosage of the alkaline water is 5 - 8 times the mass of the pulp, and the mass concentration of the alkaline water is 0.5 - 1.2%.
3. The preparation method of mogroside according to claim 1, characterized in that, The alkaline water is a calcium hydroxide solution prepared by dissolving calcium oxide in water.
4. The preparation method of mogroside according to claim 1, characterized in that, In step (2), the dosage of the alkaline water is 2.5 - 4 times the mass of the pulp, the motor speed of the flash extractor is 10000 - 15000 R / min, extract for 3 minutes, and extract twice.
5. The preparation method of mogroside according to claim 4, characterized in that Extract for one minute each time, and take a 2 - minute break in the middle after each extraction.
6. The preparation method of mogroside according to claim 1, characterized in that, In step (3), the mixed resin of the composite column consists of alumina in the upper layer and cation resin in the lower layer; the mass ratio of alumina to cation resin is 1﹕2 - 3, the total dosage of the mixed resin and the raw material weight ratio is 1﹕5 - 8; the dosage of pure water is 0.5 - 1 times the total mass of the mixed resin.
7. The preparation method of mogroside according to claim 1, characterized in that, In step (4), the addition amount of high - activity yeast is 0.5 - 1‰ of the raw material amount, and the solution is cooled to a temperature of 40 - 50°C. The mass ratio of the macroporous resin to the raw material is 1:4 - 5.
8. The preparation method of mogroside according to claim 1, characterized in that, In step (4), elute with pure water until the effluent is clear and transparent, and then perform gradient fractional elution.
9. The preparation method of mogroside according to claim 8, characterized in that, The gradient fractional elution is as follows: first elute with 15 - 27% acetone, and the elution volume is 1:2 - 4 of the resin volume, then elute with 50 - 58% ethanol, and the elution volume is 1:2 - 4 of the resin volume.
10. The preparation method of mogroside according to claim 1, characterized in that, In step (5), the addition amount of pure methanol or absolute ethanol is 3 - 4 times the volume of the concentrated solution; the pure water is 3 - 4 times the total mass of the crystals.