WO2020103798A1

WO2020103798A1 - Fermented monk fruit flavored beverage and method for preparation thereof

Info

Publication number: WO2020103798A1
Application number: PCT/CN2019/119205
Authority: WO
Inventors: 蓝福生; 李健; 钟德品
Original assignee: 桂林吉福思罗汉果有限公司
Priority date: 2018-11-19
Filing date: 2019-11-18
Publication date: 2020-05-28
Also published as: TWI718769B; CN113272414A; TW202023395A; CN113272414B

Abstract

Provided is a method for preparing a monk fruit wine, comprising the preparation and fermentation of a monk fruit glycoside-free concentrated solution, the preparation of said monk fruit glycoside-free concentrated solution comprising: (i) performing water extraction after crushing monk fruit, to prepare a monk-fruit water extract; (ii) clarifying the water extract; (iii) performing adsorption chromatography on the clarified water extract and collecting the effluent; and (iv) nanofiltering the effluent. A method for preparing monk-fruit raw vinegar comprises fermentation of the fruit wine with acetic acid bacteria. A fermented monk fruit flavored beverage contains monk fruit flavor extract, monk fruit glycoside-free fruit juice, and monk fruit raw vinegar obtained by fermentation with monk fruit glycoside-free extract, the method for preparing said monk fruit flavor extract being as follows: (a) a monk fruit extract liquid after extraction of monk fruit V glycoside; (b) using a nanofiltration membrane to filter the monk fruit extract liquid obtained in step (a) to prepare a retentate; (c) monitoring the soluble solid concentration of the retentate of step (b); (d) if the soluble solid concentration of the retentate of step (b) is 3–8 g per 100 g, then diluting the retentate of step (c) with water to prepare a diluted retentate; (e) using a nanofiltration membrane to filter the diluted retentate to prepare another retentate; (f) monitoring the soluble solid concentration of the retentate of step (e); (g) using water to dilute the retentate of step (e) to prepare another diluted retentate; (h) repeating steps (e) to (g) until the monosaccharide concentration of the retentate is less than approximately 20% on the basis of dry weight, to obtain a monk fruit flavor extract.

Description

Fermented Luo Han Guo flavored drink and preparation method thereof

Technical field

The invention relates to a fermented Luo Han Guo flavored drink, which includes Luo Han Guo flavored fruit wine and Luo Han Guo flavored fruit vinegar, and a preparation method thereof.

Background technique

Luo Han Guo is a precious medicinal and food homologous plant, which is rich in medicinal active ingredients and nutrients. Mogroside V is developed into a pure natural source of high sweetness because of its pure sweetness and 300 times sweetness. Agents, mogroside deep processing products currently on the market are also mainly based on mogrosides. However, while people are extracting and separating the mogrosides, the non-glycoside parts in the mogrosides are not effectively used. This includes rich sugar components and mogroside proteins with unique flavors or can form unique flavors, and water-soluble meals. Fibers and other components, even though some researchers have conducted research on the separation of these components, how to form these products into unique products on the application side still needs further development.

In the field of mangosteen fermented products, many researchers have tried to ferment mangosteen or mixed mangosteen with other plant parts by traditional fermentation methods to obtain more applications for mangosteen ingredients. At present, there have been more relevant research literatures or reports of patent applications, but these studies have not found and concerned that the direct fermentation of Luo Han Guo will cause the hydrolysis of glycosidic bonds and sugar chains of the glycoside components, and thus form a bitter secondary glycoside or The problem of aglycone is obviously not conducive to the retention of active ingredients and the improvement of product taste.

China Patent Application No. 201710103834.9 discloses "a method for preparing fruit vinegar from the waste column liquid in the production of mogroside". This method uses the column liquid in the production process of mogroside as the raw material for fermentation to obtain fruit vinegar. The comprehensive utilization of non-glycoside V components in Siraitia grosvenorii. However, the column flow generated in the application of mogroside V production and adsorption process still contains other glycoside components besides mogroside V. If it is directly fermented, the aforementioned glycosidic bond and sugar chain hydrolysis will still occur, resulting in the formation of secondary Glycoside or aglycone makes the product bitter, which seriously affects the taste of the product. In addition, the inventors discovered that when the de-glycosidated Luo Han Guo extract is used as a fermentation substrate, it is still rich in ingredients such as inorganic salts, which will cause a significant salty taste in the fermented fruit wine, which will be formed during the further fermentation of fruit vinegar. Unpleasant smell like soy sauce. The invention provides a new method for preparing Luo Han Guo fermented products, which partially overcomes the above problems.

Summary of the invention

The inventor discovered through research that mogrosides can be hydrolyzed into bitter components-secondary glycosides and aglycones under the action of yeast. The applicant designed that before fermentation, the mogroside was removed and then fermented, which greatly eliminated the adverse effects of bitterness on the product. In addition, the inventors further demineralized the fermentation substrate, so that the fruit wine obtained after fermentation has no salty taste, and further the fruit vinegar obtained by fermentation has no smell similar to soy sauce, and the product tastes pure.

On the one hand, this application relates to: 1. A method for preparing Luohanguo fruit wine, comprising:

a) Preparation of Luo Han Guo glycoside-free concentrate; and

b) Fermentation of Luo Han Guo glycoside-free concentrate.

In some embodiments, by adding Saccharomyces cerevisiae, wine yeast (also known as oval yeast), wine yeast (RW-type wine wine yeast, SY-type wine wine yeast, acid-reducing wine wine yeast, etc.), aromatic yeast or whisky yeast Perform fermentation.

2. The method of item 1, wherein the preparation of the Luo Han Guo glycoside-free concentrate comprises:

(i) Luo Han Guo is subjected to water extraction after crushing to prepare Luo Han Guo water extract;

(ii) Clarified water extract;

(iii) Perform adsorption chromatography on the clarified water extract and collect the effluent; and

(iv) Nanofiltration treatment of the effluent.

3. The method of

item

1 or 2, wherein the adsorption resin of the adsorption chromatography is a macroporous polymer adsorption resin.

4. The method of item 3, wherein the macroporous polymer adsorption resin is a styrene-based copolymer, a styrene-divinylbenzene copolymer or a divinylbenzene copolymer.

5. The method of any one of items 1-4, wherein the nanofiltration membrane used for the nanofiltration is 200-5,000 Daltons, 200-2,500 Daltons, 200-2,000 Daltons, 200-1,000 Daltons , 500-5,000 Daltons, 500-2,500 Daltons, 500-2,000 Daltons, or 500-1,000 Daltons.

6. The method of any one of items 1-5, wherein the content of mogroside V in the glycoside-free concentrate is less than 0.5% w / w, 0.4% w / w, 0.3% w / w, 0.2% w / w , 0.1% w / w, 0.09% w / w, 0.08% w / w, 0.07% w / w, 0.06% w / w, 0.05% w / w, 0.04% w / w, 0.03% w / w or 0.01 % W / w.

7. The method of any one of items 1-6, which includes desalting the glycoside-free concentrated liquid before fermentation. In some embodiments, the desalting treatment is performed by cation exchange chromatography.

8. The present invention relates to the Luohanguo fruit wine prepared by the method of any one of items 1-7.

9. A method for preparing raw grosvenor grosvenor vinegar, which comprises fermenting acetic acid bacteria on the grosvenor mangosteen wine prepared by the method according to any one of items 1-7. In some embodiments, the acetic acid bacteria are selected from: various strains of the genus Gluconobacter, acetic acid bacteria screened and isolated from the surface of the fruit, the surrounding soil of the fruit tree, and naturally fermented fruit vinegar (eg, mixed-pool variant acetic acid) Bacteria, Acetobacter pasteurianus) and mixed acetic acid bacteria species consisting of the above-mentioned acetic acid bacteria.

10. Mangosteen raw vinegar prepared by the method of item 9.

11. A method for preparing mangosteen fruit vinegar, which comprises preparing raw mangosteen fruit vinegar of item 10 so that the total acid concentration (% w / w) is 0.10-0.50%, 0.15 to 0.45%, 0.15 to 0.40%, 0.15 to 0.35%, 0.15 to 0.30%, 0.15 to 0.25%, 0.20 to 0.45%, 0.20 to 0.40%, 0.20 to 0.35%, 0.20 to 0.30%, 0.25 to 0.45%, 0.25 to 0.40%, 0.25 to 0.35%, 0.25 to 0.30% or 0.20% to 0.3%. In some embodiments, the raw grosvenor vinegar can be mixed with any beverage of a desired taste, for example, it can be mixed with fruit juice (eg, apple juice, orange juice, peach juice, etc.), vegetable juice, cereal extract, carbonated beverage , Water, and mixing to make the total acid concentration (% w / w) 0.10-0.50%, 0.15-0.45%, 0.15-0.40%, 0.15-0.35%, 0.15-0.30%, 0.15-0.25%, 0.20- 0.45%, 0.20 to 0.40%, 0.20 to 0.35%, 0.20 to 0.30%, 0.25 to 0.45%, 0.25 to 0.40%, 0.25 to 0.35%, 0.25 to 0.30% or 0.20% to 0.3%. In some embodiments, the Luo Han Guo raw vinegar is mixed with water.

12. A method for preparing Luo Han Guo flavored drink, comprising adding Luo Han Guo flavor extract to Luo Han Guo original vinegar of Item 10 or Luo Han Guo fruit vinegar of Item 11, wherein the method of preparing the Luo Han Guo flavor extract is as follows:

a) Mogroside extract after mogroside V extraction;

b) Nanofiltration membrane filtering step a) the obtained Luo Han Guo extract to prepare retentate;

c) monitor the soluble solid concentration of the retentate of step b);

d) When the soluble solid concentration of the retentate of step b) is 3-8g / 100g, dilute the retentate of step c) with water to prepare a diluted retentate;

e) Use nanofiltration membrane to filter the diluted retentate to prepare another retentate;

f) monitor the soluble solids concentration of the retentate of step (e);

g) Dilute the retentate of step (e) with water to prepare another diluted retentate;

h) Repeat steps (e) to (g) until the monosaccharide concentration of the retentate is less than about 20% based on dry weight to obtain Luo Han Guo flavor extract.

In some embodiments, the soluble solids concentration is 3g / 100g, 4g / 100g, 5g / 100g, 6g / 100g, 7g / 100g, or 8g / 100g.

13. The method of item 13, wherein the nanofiltration membrane has a molecular weight cut-off of 250 Daltons to 2,500 Daltons, 500-2,500 Daltons, or 500-2,000 Daltons.

14. The method of

item

12 or 13, wherein after the step h), a step of drying the retentate is further included. Preferably, the drying step is spray drying.

15. The method according to any one of items 12 to 14, further comprising the step of adding a sweet product derived from Luo Han Guo to adjust the acid-sweet ratio of the Luo Han Guo flavored drink, wherein the sweet-sour ratio is based on the weight percentage of total sugar and total acid in the drink The ratio indicates that the sour-sweet ratio is 18 to 28/1, 19 to 27/1, 20 to 26/1, 21 to 25/1, 22 to 24/1, and 23 to 24/1.

16. The method of item 15, wherein the sweetener derived from Luo Han Guo includes mogroside V and mogroside V-free mogroside non-glycoside juice.

17. A fermented Luo Han Guo flavored beverage, comprising Luo Han Guo flavored extract, Luo Han Guo non-glycoside juice and Luo Han Guo raw vinegar, wherein the total sugar content (% w / w) of Luo Han Guo origin in the beverage is about 3.0-9.0%, 3.5- 8.5%, 4.0 ～ 8.0%, 4.5 ～ 7.5%, 5.0 ～ 7.0% or 5.5 ～ 6.5%, the total acid content (% w / w) is 0.1 ～ 0.15 ～ 0.45%, 0.15 ～ 0.40%, 0.15 ～ 0.35%, 0.15 to 0.30%, 0.15 to 0.25%, 0.20 to 0.45%, 0.20 to 0.40%, 0.20 to 0.35%, 0.20 to 0.30%, 0.25 to 0.45%, 0.25 to 0.40%, 0.25 to 0.35%, 0.25 to 0.30%, 0.20% ~ 0.3% or 0.5% (w / w).

Item 18. The beverage of item 1, wherein the ratio (weight ratio) of the total sugar to the total acid is 18 to 28: 1, 19 to 27/1, 20 to 26/1, 21 to 25/1, 22 to 24 / 1. 23 ～ 24/1.

19. The Luo Han Guo flavored drink of

Item

1 or 2, which is Luo Han Guo flavored fruit vinegar.

20. A method for preparing a fermented Luo Han Guo flavored beverage according to any one of items 17-19, comprising preparing Luo Han Guo flavored extract, Luo Han Guo non-glycoside juice and Luo Han Guo raw vinegar, and mixing them in an appropriate ratio to make Luo Han Guo in the drink The total sugar content (% w / w) of the source is about 3.0 to 9.0%, 3.5 to 8.5%, 4.0 to 8.0%, 4.5 to 7.5%, 5.0 to 7.0% or 5.5 to 6.5%, and the total acid content (% w / w) w) 0.1 to 0.15 to 0.45%, 0.15 to 0.40%, 0.15 to 0.35%, 0.15 to 0.30%, 0.15 to 0.25%, 0.20 to 0.45%, 0.20 to 0.40%, 0.20 to 0.35%, 0.20 to 0.30%, 0.25 ～ 0.45%, 0.25 ～ 0.40%, 0.25 ～ 0.35%, 0.25 ～ 0.30%, 0.20% ～ 0.3% or 0.5% (w / w), the ratio of sweet and sour is 18 ～ 28/1, 19 ～ 27/1, 20 ～ 26/1, 21 ～ 25/1, 22 ～ 24/1 or 23 ～ 24/1.

Item 20. The method according to item 20, wherein the preparation of Luohanguo raw vinegar comprises:

a) Preparation of Luo Han Guo glycosides-free concentrate;

b) Yeast fermentation of grosvenorrhiza glycoside-free concentrated liquid, followed by acetic acid bacteria fermentation.

In some embodiments, by adding Saccharomyces cerevisiae, wine yeast (also known as oval yeast), wine yeast (RW-type wine wine yeast, SY-type wine wine yeast, acid-reducing wine wine yeast, etc.), aromatic yeast or whisky yeast Perform yeast fermentation. In some embodiments, the acetic acid bacteria fermented by the acetic acid bacteria are selected from: various strains of the genus Acetobacter spp., Acetic acid bacteria isolated from the surface of the fruit, the surrounding soil of the fruit tree, and naturally fermented fruit vinegar (eg, mixed pool Variants of acetic acid bacteria, Acetobacter pasteurianus) and mixed acetic acid bacteria species consisting of the above acetic acid bacteria.

22. The method according to item 21, wherein the preparation of the Luo Han Guo glycoside-free concentrated solution comprises:

(ii) Clarified water extract;

(iv) Nanofiltration treatment of the effluent.

23. The method of

item

21 or 22, wherein the adsorption resin of the adsorption chromatography is a macroporous polymer adsorption resin.

24. The method of item 23, wherein the macroporous polymer adsorption resin is a styrene-based copolymer, styrene-divinylbenzene copolymer or divinylbenzene copolymer.

25. The method of any one of items 21-24, wherein the nanofiltration membrane used for the nanofiltration is 200-5,000 Daltons, 200-2,500 Daltons, 200-2,000 Daltons, 200-1,000 Daltons , 500-5,000 Daltons, 500-2,500 Daltons, 500-2,000 Daltons, or 500-1,000 Daltons.

26. The method of any one of items 21-25, wherein the content of mogroside V in the glycoside-free concentrate is less than 0.5% w / w, 0.4% w / w, 0.3% w / w, 0.2% w / w , 0.1% w / w, 0.09% w / w, 0.08% w / w, 0.07% w / w, 0.06% w / w, 0.05% w / w, 0.04% w / w, 0.03% w / w or 0.01 % W / w.

27. The method of any one of items 21 to 26, which comprises desalting the glycoside-free concentrated liquid before fermentation. In some embodiments, the desalting treatment is performed by cation exchange chromatography.

28. The method of any one of items 20-27, the preparation of the Luo Han Guo flavor extract comprises:

a) Mogroside extract after mogroside V extraction;

c) monitor the soluble solid concentration of the retentate of step b);

f) monitor the soluble solids concentration of the retentate of step (e);

29. The method of item 28, wherein the nanofiltration membrane has a molecular weight cut-off of 250 Daltons to 2,500 Daltons, 500-2,500 Daltons, or 500-2,000 Daltons.

30. The method of item 28 or 29, wherein after the step h), a step of drying the retentate is further included. Preferably, the drying step is spray drying.

31. The method of any one of items 20 to 30, wherein the preparation of Luo Han Guo non-glycoside juice includes:

(ii) Clarified water extract; and

(iii) Perform adsorption chromatography on the clarified water extract and collect the effluent.

32. The method of item 31, wherein the adsorption resin of the adsorption chromatography is a macroporous polymer adsorption resin.

33. The method of item 32, wherein the macroporous polymer adsorption resin is a styrene-based copolymer, styrene-divinylbenzene copolymer or divinylbenzene copolymer.

34. The method of any one of items 31-33, wherein the preparation of the Luo Han Guo glycoside-free fruit juice further comprises the step of ion exchange chromatography. In some embodiments, the ion exchange chromatography includes cation exchange chromatography and anion exchange chromatography. 35. A mangosteen fruit wine or fruit vinegar obtained by fermenting mangosteen-extracted mogroside extract.

BRIEF DESCRIPTION

Figure 1 is an HPLC chart of 6 glycoside control substances. Among them, peak 1 is: 11-O-mogroside V; peak 2 is: mogroside V; peak 3 is: siamin I; peak 4 is: mogroside IVe; peak 5 is: mogroside III ; Peak 6 is: Mogroside IIe.

Figures 2A-2B are HPLC profiles of the glycoside-free concentrate B sample at 0 hours of fermentation time (Figure 2A) and HPLC profiles of the sample and the control at 0 hours of fermentation time (Figure 2B).

Figures 3A-3B are HPLC profiles of the glycoside-free concentrate B sample at 6 hours of fermentation time (Figure 3A) and HPLC profiles of the sample and the control at 6 hours of fermentation time (Figure 3B). Figures 4A-4B are HPLC patterns of the glycoside-free concentrate B sample at the fermentation time of 12 hours (Figure 4A) and HPLC patterns of the sample and the control at the fermentation time of 12 hours (Figure 4B). Figures 5A-5B are HPLC profiles of the sample B without glycoside concentrate (Figure 5A) when the fermentation time is 18 hours and the HPLC profiles of the sample and the control when the fermentation time is 18 hours (Figure 5B). Figures 6A-6B are HPLC profiles of the sample B without glycoside concentrate at 24 hours of fermentation time (Figure 6A) and HPLC profiles of the sample and the control at 24 hours of fermentation time (Figure 6B). Figures 7A-7B are the HPLC profiles of the glycoside-free concentrate B sample at 30 hours of fermentation time (Figure 7A) and the HPLC profiles of the sample and the control at 30 hours of fermentation time (Figure 7B). Figures 8A-8B are the HPLC profiles of the glycoside-free concentrate B sample at the fermentation time of 36 hours (Figure 8A) and the HPLC profiles of the sample and the control at the fermentation time of 36 hours (Figure 8B).

Figure 9 is a graph showing the degradation trend of six glycosides in Luo Han Guo during alcohol fermentation. With the continuation of alcohol fermentation time, the contents of 11-O-glycoside V and glycoside V continued to decrease to 0, and the content of thymidine I, glycoside IVe, glycoside III, and glycoside IIe increased first and then decreased. During the alcohol fermentation process, the above 6 glycosides will eventually be decomposed and converted into other substances as long as there is enough time, and the sugar groups on the glycosides are degraded one by one during the degradation process, forming secondary glycosides or aglycones with a bitter taste .

FIG. 10A is an HPLC chart of six glycosides after nanofiltration membrane separation treatment (grosvenorrhiza no glycoside concentrate C). As shown in the figure, after passing through the column effluent (mogroside-free glycoside concentrate B) through nanofiltration membrane separation, mogrosides will be completely removed, and the material is more pure. FIG. 10B is an HPLC chart of six glycosides after nanofiltration membrane separation treatment (grosvenorrhiza no glycoside concentrate D). As shown in the figure, after passing through the column effluent (mogroside-free glycoside concentrate B) through nanofiltration membrane separation, mogrosides will be completely removed, and the material is more pure.

FIG. 11 is a comparison chart of taste tests of products obtained by using Luo Han Guo extract A, glycoside-free concentrate B, glycoside-free concentrate C and glycoside-free concentrate D as fermentation substrates. This figure shows that the Luo Han Guo flavored fruit wine obtained by fermentation after deglycosylation treatment has greatly improved the post-bitterness, off-flavor, sourness, taste and other indicators compared to the fruit wine obtained by direct fermentation without deglycosidation.

Figures 12A-12B show the results of acidity optimization studies of Luohanguo fruit vinegar. According to the total acid concentration of 0.15%, 0.20%, 0.25%, 0.35%, and 0.45% of the total acid concentration of Luohanguo raw vinegar and water, the obtained solution was tested by the tasting team to obtain the preference and different total acid content. Under the correlation of different sweet-acid ratios.

Fig. 13 shows the results of a preferred study on the amount of Luo Han Guo flavor extract added in Luo Han Guo flavor fruit vinegar. Using Luohanguo flavored fruit vinegar with total acid content of 0.3% and sweet / acid ratio of 24/1, different levels of Luohanguo flavor extracts were added to test its taste performance. As a result, it was found that the amount of flavor extracts added was 0.2-0.6% It has a good taste.

Fig. 14 shows the comparison results of the taste test of the diluted solution of the Luohanguo flavored fruit vinegar after completion of the preparation and the unadjusted Luohanguo raw vinegar (total acid concentration 0.3%).

Detailed description of the invention

The present invention relates to a method for preparing Luo Han Guo flavored beverage, including Luo Han Guo flavored wine and fruit vinegar. This method effectively avoids the traditional direct fermentation method or the prior art due to the effective separation of the mogroside components in Luo Han Guo before fermentation. The bitter secondary glycosides or aglycones formed by the fermentation process of the column fluid in the production process of middle-use glycosides are used as raw materials, resulting in a bad taste of fruit wine or fruit vinegar. In addition, the fermentation substrate is subjected to desalination treatment, such as cation exchange resin desalination treatment, so that the taste of the fruit wine obtained after fermentation of the desalted treatment is controlled, and the fruit vinegar product has no soy sauce smell and pure taste. In the product preparation, due to the addition of the appropriate proportion of Luo Han Guo glycoside concentrate and Luo Han Guo flavor extract to the fermented product, the prepared Luo Han Guo flavored beverage, such as Luo Han Guo fruit vinegar, has a unique taste and also makes non-glycoside nutrients in Luo Han Guo In order to make full use of it, it provides a new way of thinking about the value of Luohanguo resources.

The following description is intended to illustrate the methods and parameters described in the present invention by way of example, and is not intended to limit the scope of the present application.

The present application provides a method for preparing a mangosteen fermented beverage from an extract obtained from Cucurbitaceae fruits containing terpene glycosides, such as Siraitia fruits. The Siraitia fruit includes the following species of Siraitia grosvenorii: S. grosvenorii, S. siamensis, S. silomaradjae, S. sikkimensis, S. africana, S. borneensis and Taiwan Luo Han Guo (S. taiwaniana). However, most preferably, the fruit used is Siraitia grosvenorii (S.grosvenorii), formerly known as Momordica (Momordica grosvenorii)), which is a member of Cucurbitaceae. Luo Han Guo (S.grosvenorii) grows in the southeastern provinces of China, mainly in Guangxi. It has been cultivated and used as a traditional Chinese medicine for hundreds of years, used to treat cough and lung congestion, and is also used as a sweetener and flavoring agent in soups and teas.

The other fruits of Luo Han Guo and Cucurbitaceae contain terpene glycosides, which are known as mogrosides and siamin, they are present in fruits at a level of about 1%. These compounds have been described and characterized by Matsumoto et al .; Chem. Pharm. Bull., 38 (7), 2030-2032 (1990). Mogrosides according to the present invention are a general term for glycoside compounds with 1 to 6 glucose molecules connected to the triterpene main chain. The most abundant mogroside is mogroside V, which is estimated to have about 250% (by weight) sweetness of sucrose. Therefore, mogroside can be used as an excellent sugar substitute.

The “glycosides”, “glycosides” and “sweet glycosides” described in the present invention have the same meaning, and represent compounds containing glycosidic bonds, which can be used interchangeably. "Mogroside", "mogroside", and "mogroside" have the same meaning, meaning glycoside compounds derived from Mogrosides and having 1 to 6 glucose molecules connected to the triterpene main chain. These terms are used interchangeably.

In the process of extracting mogrosides, many by-products are produced, and their use is one of the subjects of research. Many researchers have tried to ferment the by-products produced in the extraction process of Mogrosides by traditional fermentation methods, but these studies have not found and paid attention to the hydrolysis of glycosidic bonds and sugar chains of the glycoside components accompanying fermentation, and thus formed a bitter taste The problem of secondary glycosides or aglycones is obviously not conducive to the retention of active ingredients and the improvement of product taste.

The invention provides a method for preparing a fermented Luo Han Guo flavored drink, which includes:

(a) Preparation of Luo Han Guo glycoside-free concentrate; and

(b) Fermentation of Luo Han Guo glycoside-free concentrate.

Preparation of glycoside-free concentrate

The Luo Han Guo glycoside-free concentrate, or simply the glycoside-free concentrate for short, is a Luo Han Guo extract concentrate that removes or substantially removes the mogroside. The glycoside-free concentrate of the present invention is extracted from cucurbitaceous fruits such as the grosvenorii, preferably S. grosvenorii, wherein the glycoside-free concentrate contains very low mogroside V and other terpene glycosides. In some embodiments, the glycoside-free concentrate contains less than 1% w / w, 0.5% w / w, 0.4% w / w, 0.3% w / w, 0.2% w / w, 0.1% w / w, 0.09 Mogroside V in% w / w, 0.08% w / w, 0.07% w / w, 0.06% w / w, 0.05% w / w, 0.04% w / w, 0.03% w / w, 0.01% w / w . In some embodiments, the glycoside-free concentrate does not contain mogroside V.

Mogroside V can be detected by HPLC method, for example, the method specified in the National Food Safety Standard "Food Additive Mogroside" (GB1886.77-2016) used in this application.

In some embodiments, the present application provides a method for preparing a glycoside-free concentrate, the method includes extracting Luo Han Jue from Luo Han Guo, contacting the extracted Luo Han Juice with an adsorption medium to reduce the content of mogroside V, and then extracting Luohan juice is in contact with the nanofiltration membrane. In such an embodiment, before the nanofiltration step, the extracted Luo Han juice is contacted with the adsorption medium. Such an adsorption medium may be an adsorption resin. In other embodiments, other methods suitable for reducing mogroside V content may be used.

In an exemplary method, Luo Han Guo is contacted with water to prepare a water extract juice. The water extract juice contains mogroside V and other terpene glycosides. The clear water extracts the juice (for example, to remove pectin and other proteins) to facilitate subsequent processing. The clarified water extraction juice is contacted with an adsorption resin, wherein at least a portion of mogroside V and other terpene glycosides in the clarified water extraction juice are adsorbed onto the resin. The effluent from the adsorption resin was mogroside juice after mogroside reduction, and it was contacted with the nanofiltration membrane. The permeate prepared from the nanofiltration step is fermented.

In some embodiments, the method includes:

The fruit of the genus Luohanguo is contacted with water to obtain water extract juice, wherein the water extract juice contains mogroside V and other terpene glycosides;

Clarified water extract juice;

Contacting the clarified water extraction juice with an adsorption medium to adsorb at least a portion of mogroside V and other terpene glycosides in the clarified water extraction juice and prepare an effluent;

Use nanofiltration membranes to filter effluent and prepare permeate; and

The permeate was fermented to prepare Luohanguo flavored wine and Luohanguo raw vinegar.

The glycoside-free concentrate obtained by the above method contains very low mogroside V and other terpene glycosides, such as less than 0.5% w / w, 0.4% w / w, 0.3% w / w, 0.2% w / w, 0.1 % W / w, 0.09% w / w, 0.08% w / w, 0.07% w / w, 0.06% w / w, 0.05% w / w, 0.04% w / w, 0.03% w / w, 0.01% w Mogroside V of / w most preferably does not contain mogroside V.

Compared with mogroside V and total terpene glycoside content naturally occurring in fruits, the effluent obtained by contact with the adsorption medium has a lower mogroside V content and a lower total terpene glycoside content. In some embodiments, the clarified juice is contacted with an adsorption medium to prepare an effluent having an amount of mogroside V two, three, or four times smaller than the amount of mogroside V naturally present in the fruit. In other embodiments, the clarified juice is contacted with an adsorption medium to prepare an effluent having an amount of mogroside V two to three times, three times less than the amount of mogroside V naturally present in the fruit To four times or twice to four times. In still other embodiments, the clarified juice is contacted with an adsorption medium to prepare an effluent having an amount of mogroside V that is at least about 95% w / w less than the amount of mogroside V naturally present in the fruit, At least about 96% w / w, at least about 97% w / w, or at least about 98% w / w. In certain embodiments, the clarified juice is contacted with an adsorption medium to prepare an effluent having an amount of mogroside V that is about 95% to about 98% w less than the amount of mogroside V naturally present in the fruit / w, about 95% to about 97% w / w, about 95% to about 96% w / w, about 96% to about 98% w / w, 96% to 97% w / w, or 97% to 98% w / w.

The above method for preparing the glycoside-free concentrated liquid of the present invention includes contacting the effluent prepared above with a nanofiltration membrane to prepare a permeate to further remove mogroside V and other terpene glycosides in the effluent.

Those skilled in the art can understand that in order to further remove mogroside V and other terpene glycosides in the effluent, those skilled in the art can use other methods besides nanofiltration, for example, adsorption chromatography, ion exchange chromatography, etc. Its purpose is to further remove mogroside V and other terpene glycosides in the effluent. These technical solutions are covered by the protection scope of the present invention.

In certain embodiments, mogrosides are contacted with water to prepare water extraction juice, and the water extraction juice is contacted with an adsorption medium to remove at least a portion of mogroside V and other terpene glycosides in the water extraction juice. The effluent coming out of contact with the adsorption medium is mogroside reduced mogroside. In some embodiments, the water extraction juice can be clarified before and / or after contact with the adsorption medium.

In other embodiments, the present application provides a method for preparing a glycoside-free concentrate, the method comprising extracting juice from Siraitia grosvenori, contacting the extracted juice with a nanofiltration membrane, and then contacting the filtered juice with an adsorption medium to reduce Mogroside V content. In such an embodiment, after the nanofiltration step, the extracted juice is contacted with the adsorption medium.

Each step of the preparation method of the glycoside-free concentrated liquid is described in more detail below.

extract

Mangosteen fruit juice can be extracted by contacting macerated fruits (such as fresh mangosteen and dried mangosteen) with normal temperature water or hot water. In some embodiments, the macerated fruit is contacted with water at a sufficient temperature for a sufficient time to obtain a water-extracted juice.

In some embodiments, the fruit is contacted with water at a temperature of at least 60 ° C, at least 70 ° C, or at least 80 ° C. In other embodiments that can be combined with the foregoing, the fruit is contacted with water for 30 minutes to 4 hours, 30 minutes to 3 hours, and 60 minutes to 2 hours.

In some embodiments, this extraction step is performed using a counter-current water extraction method.

clarify

The water extraction juice can be clarified before contact with the nanofiltration membrane. In some embodiments, the clarification step comprises ultrafiltration of water to extract juice. In certain embodiments, the clarification step includes treating the extract with pectinase under conditions that dissolve the pectin and complex sugar.

Adsorption media

The adsorption medium used in the description of the present application may be a single medium or a mixture of suitable media.

In some embodiments, the adsorption medium is an adsorption resin. In some examples, the adsorption resin is provided as a bed of adsorption resin. In embodiments where the adsorbent material is an adsorbent resin, suitable adsorbent resins may include any resin that is suitable for wettable hydrophobic matrices in contact with food, including, for example, polyvinylpolypyrrolidone (PVPP), nylon, acrylate , Styrene-divinylbenzene copolymer and divinylbenzene copolymer. In some embodiments, the adsorption resin is a macroporous polymer adsorption resin. In certain embodiments, the adsorption resin is a styrene-based copolymer resin. In other embodiments, the adsorption resin is a styrene-divinylbenzene copolymer or a divinylbenzene copolymer. Any combination of media described in this application may also be used.

The adsorption medium used in the described method may be commercially available or prepared according to any method or technique known in the art.

Nanofiltration

In some embodiments, the nanofiltration membrane selectively removes mogroside V and other terpene glycosides in the water extraction juice or adsorption chromatography effluent. In some embodiments, the nanofiltration membranes used in the methods described herein have a molecular weight cut-off of 200-5,000 Daltons, 200-2,500 Daltons, 200-2,000 Daltons, 200-1,000 Daltons, 500- 5,000 Daltons, 500-2,500 Daltons, 500-2,000 Daltons, 500-1,000 Daltons.

Nanofiltration membranes can be obtained from commercially available sources.

When the water extraction juice or adsorption chromatography effluent comes into contact with the nanofiltration membrane, retentate and permeate are generated. The retentate contains components that cannot pass through the nanofiltration membrane. The permeate contains components that can pass through the membrane based on the molecular weight cut-off of the membrane, such as monosaccharides.

In some embodiments, the soluble solids content of the retentate is monitored. Water can be added to the retentate, the diluted retentate is in contact with the nanofiltration membrane, and the process is repeated. For example, the process can be repeated until the retentate reaches a low level of monosaccharides and / or the retentate has no sourness.

In some embodiments, the soluble solids in the retentate may include monosaccharides, disaccharides, polysaccharides, inorganic salts, minerals or proteins, or any combination thereof. The soluble solid content can be measured using any suitable method and technique known in the art. For example, the soluble solids content can be measured using a brix refractometer. The soluble solids content can be expressed in any suitable unit. For example, the soluble solids content can be expressed as a weight ratio (w / w, grams of soluble solids / grams of sample), a percentage based on dry weight (% w / w), or Brix. The percentage of soluble substances of the present invention is a percentage based on dry weight (% w / w).

Preparation of Luohanguo flavor wine and Luohanguo raw vinegar

The glycoside-free concentrated liquid is fermented to prepare Luohanguo flavored fruit wine. "Fruit wine" is the use of fruit or fruit ingredients as raw materials, through natural fermentation or artificial addition of yeast to break down the sugar in the fruit to brew a low-alcohol beverage wine with an alcohol flavor. The glucose and fructose in the Luo Han Guo glycoside-free concentrated solution of the present invention can be directly utilized by alcohol fermentation under the action of yeast, and sucrose and maltose generate glucose and fructose in the fermentation process to participate in alcohol fermentation. Yeast is the main microorganism for fruit wine fermentation. Any yeast known in the art capable of fermenting fruits may be used, such as Saccharomyces cerevisiae, wine yeast (also known as oval yeast), wine wine yeast (RW type wine wine yeast, SY type wine wine yeast, acid-reducing wine wine yeast) Etc.), raw fragrance yeast, whisky yeast, etc.

"Original vinegar" is an acidic beverage or condiment made from fruit or fruit wine as raw materials and fermented by acetic acid bacteria. Acetic acid bacteria can be any acetic acid bacteria commonly used in the art to prepare fruit vinegar, such as various strains in the genus Gluconobacter, such as Ga. Europaeus, or can be selected from the surface of the fruit, the surrounding soil of the fruit tree and the naturally fermented fruit vinegar Isolated acetic acid bacteria. For example, mixed pool variants of acetic acid bacteria, acetic acid bacteria of Pasteurella or P. pastoris and mixed acetic acid bacteria composed of the above-mentioned acetic acid bacteria.

In some embodiments, although the above-mentioned concentrated solution of Luo Han Guo glycosides after separation treatment by adsorption resin or nanofiltration membrane reduces or avoids the hydrolysis of glycoside components during fermentation to form bitter components, but the fermentation substrate-Luo Han Guo glycoside concentrated solution It still contains a large amount of inorganic salts and other ingredients, which will produce a significant salty taste in the fermented mangosteen-flavored fruit wine and form a soy sauce-like odor in the resulting mangosteen-flavored fruit vinegar product. Although the soy sauce odor and salty taste are beneficial for fruit vinegar products in seasoned vinegar, as beverage wine and vinegar, these two flavors obviously reduce the competitiveness of the product. For this reason, in some embodiments, a desalination step is added, for example, a cation exchange resin is used to desalt the fermentation substrate, the salty taste of the fruit wine obtained after fermentation of the processed product is controlled, and the fruit vinegar product has no soy sauce-like smell and taste Pure.

Because the pH value of the material treated by the cation exchange resin is low, it is not conducive to the fermentation of fruit wine. If alkali metal alkalis (such as NaOH, KOH, etc.) are used to adjust the pH, metal ions will be re-introduced into salt to produce salty taste, so ammonia is used. As a pH adjusting agent is a feasible method. Another method for adjusting the pH value is to connect an anion exchange resin in series when the cation exchange resin is desalted to control the appropriate ratio of positive and negative resins, so that the effluent can obtain a pH suitable for fruit wine fermentation while desalting. The pH suitable for fruit wine fermentation can be routinely determined by those skilled in the art.

Preparation of Luo Han Guo Flavor Extract

Luo Han Guo flavor extract is a Luo Han Guo extract that retains the flavor of Luo Han Guo and is partially or substantially eliminated by Mogroside V. The invention relates to a method for preparing Luo Han Guo flavor extract. The method comprises:

a) Extract (for example, water extract) Luo Han juice, and remove mogroside V in Luo Han juice;

b) Filtration of the Luohan juice obtained in step a) using a nanofiltration membrane to prepare a retentate, wherein the molecular weight cut-off of the nanofiltration membrane is 250 Daltons to 2,500 Daltons;

c) Monitor the soluble solids concentration of the retentate of step (b);

d) When the soluble solid concentration of the retentate of step (c) is about 3-8g / 100g (eg 3g / 100g, 4g / 100g, 5g / 100g, 6g / 100g, 7g / 100g, 8g / 100g), Dilute the retentate of step (c) with water to prepare diluted retentate;

f) monitor the soluble solids concentration of the retentate of step (e);

The applicant found that when the monosaccharide concentration of the above retentate is less than about 20% (greater than about 13%) based on dry weight, the retentate can maintain both a good Luo Han Guo flavor and a good sugar-acid ratio and mouthfeel. .

In step a) of the above method for preparing Luo Han Guo flavor extract, any method of removing Luo Han Guo glycoside V in the art can be used to remove Luo Han Guo glycoside V in Luo Han Guo juice. For example, the aqueous extract of Luo Han Guo can be contacted with an adsorption resin to make At least a portion of mogroside V in the extracted juice is combined with an adsorption resin to remove mogroside V.

After the step h), the method for preparing the Luo Han Guo flavor extract may further include step i): drying (for example, spray drying) the concentrated retentate to prepare a dry extract. When the monosaccharide concentration of the retentate prepared in step h) above is less than about 20% (more than about 13%) based on dry weight, the retentate is easier to dry. The dried Luo Han Guo flavor extract is easier to store and use. In another embodiment, the present application provides a method of Luo Han Guo flavor extract as follows, the method includes:

a) Filter the water extract juice obtained from fruits using a nanofiltration membrane to prepare a retentate, wherein the molecular weight cut-off of the nanofiltration membrane is 250 Daltons to 2,500 Daltons;

b) Monitor the soluble solids concentration of the retentate of step (a);

c) When the soluble solid concentration of the retentate of step (b) is about 3-8g / 100g (eg 3g / 100g, 4g / 100g, 5g / 100g, 6g / 100g, 7g / 100g, 8g / 100g) Dilute the retentate of step (b) with water to prepare a diluted retentate;

d) Use nanofiltration membrane to filter the diluted retentate to prepare another retentate;

e) Monitor the soluble solids concentration of the retentate of step (d);

f) Dilute the retentate of step (d) with water to prepare another diluted retentate;

g) Repeat steps (d) to (f) until the monosaccharide concentration of the retentate is less than about 20% based on dry weight;

h) After step (g), the retentate is contacted with the adsorption resin, wherein the resin is combined with terpene glycoside and mogroside V to remove mogroside V.

In the above solution, the adsorption resin includes any resin suitable for wettable hydrophobic substrates in contact with food, and the resin includes, for example, polyvinylpolypyrrolidone (PVPP), nylon, acrylate, styrene-divinylbenzene copolymer And divinylbenzene copolymer. In some embodiments, the adsorption resin is a macroporous polymer adsorption resin. In certain embodiments, the adsorption resin is a styrene-based copolymer resin. In other embodiments, the adsorption resin is a styrene-divinylbenzene copolymer or a divinylbenzene copolymer. Any combination of media described in this application may also be used.

The retentate may contain fruit acids naturally present in the fruit. Therefore, the total acid content and pH of the retentate can be determined. In certain embodiments, the concentrated retentate has a total acid content of less than about 5% w / w, less than about 3% w / w, or less than about 2.5% w / w. In other embodiments, the pH of the concentrated retentate is a pH greater than about 4, for example, a pH of about 4-5.5, a pH of 4-5. In certain embodiments, the pH of the concentrated retentate is a pH greater than or equal to 4 to 4.5.

The retentate may contain monosaccharides, disaccharides and polysaccharides. In some embodiments, the total sugar content of the retentate is less than the total sugar content in Luo Han Guo water extract juice.

Sweets from Luo Han Guo

The sweetness derived from Luohanguo can be added to the raw vinegar of Luohanguo prepared in this application to adjust the ratio of sweet and sour to improve the taste. The sweet product may be any sweet-tasting substance derived from Luo Han Guo, such as Luo Han Guo V and Luo Han Guo extract from which Luo Han Guo V is removed (also referred to as Luo Han Guo glycoside-free fruit juice).

Preparation of Luo Han Guo Non-Glycoside Juice

Mogroside juice is partly or basically cleared mogroside V. The partial or basic removal of mogroside V means that the content of mogroside V in the juice is less than 0.5% w / w, 0.4% w / w, 0.3% w / w, 0.2% w / w, 0.1% w / w , 0.09% w / w, 0.08% w / w, 0.07% w / w, 0.06% w / w, 0.05% w / w, 0.04% w / w, 0.03% w / w or 0.01% w / w. In some embodiments, the present application provides a method for preparing Luo Han Guo glycoside-free fruit juice, the method comprising extracting Luo Han Ju fruit juice from Luo Han Guo, and contacting the extracted Luo Han fruit juice with an adsorption medium to reduce the Mogroside V content. In the above embodiment, optionally, first extract the Luo Han Juice from Luo Han Guo for ion exchange chromatography, and then contact the Luo Han Juice after ion exchange chromatography with the adsorption medium to reduce the content of mogroside V; preferably, in After the above adsorption chromatography step, a mogroside glycoside-free concentrated liquid is obtained, and ion-exchange chromatography is performed on the mogroside glycoside-free concentrated liquid to remove proteins, amino acids and the like. In the above method, the adsorption medium may be an adsorption resin, including any wettable hydrophobic matrix resin suitable for contact with food, the resin including, for example, polyvinyl polypyrrolidone (PVPP), nylon, acrylate, styrene- Divinylbenzene copolymer and divinylbenzene copolymer. In some embodiments, the adsorption resin is a macroporous polymer adsorption resin. In certain embodiments, the adsorption resin is a styrene-based copolymer resin. In other embodiments, the adsorption resin is a styrene-divinylbenzene copolymer or a divinylbenzene copolymer. Any combination of media described in this application may also be used. In other embodiments, other methods suitable for reducing mogroside V content may be used to prepare mogroside non-glycoside juice.

In an exemplary method, Luo Han Guo is contacted with water to prepare a water extract juice. The water extract juice contains mogroside V and other terpene glycosides. The clear water extracts the juice (for example, to remove pectin and other proteins) to facilitate subsequent processing. The clarified water extraction juice is contacted with an adsorption resin, wherein at least a portion of mogroside V and other terpene glycosides in the clarified water extraction juice are adsorbed onto the resin. The effluent from the adsorption resin is mogroside juice after mogroside reduction, and it is contacted with ion exchange resin, preferably cation exchange chromatography first, followed by anion exchange chromatography.

In some embodiments, the method includes:

Clarified water extract juice;

The effluent is subjected to ion exchange chromatography (preferably cation exchange chromatography followed by anion exchange chromatography) and the effluent is collected.

In certain embodiments, mogrosides are contacted with water to prepare water extraction juice, and the water extraction juice is contacted with an adsorption medium to remove at least a portion of mogroside V and other terpene glycosides in the water extraction juice. The effluent coming out of contact with the adsorption medium is mogroside juice after mogroside reduction. In some embodiments, the water extraction juice can be clarified before and / or after contact with the adsorption medium.

In some embodiments, Mangosteen fruit juice can be extracted by contacting the macerated fruit with hot water or water at an elevated temperature. In some embodiments, the macerated fruit is contacted with water at a sufficient temperature for a sufficient time to obtain a water-extracted juice.

In some embodiments, the fruit is contacted with water at a temperature of at least 60 ° C, at least 70 ° C, or at least 80 ° C. In other embodiments that can be combined with the foregoing, the fruit is contacted with water for 30 to 60 minutes or 90 to 120 minutes.

This application also provides extracts prepared according to any of the methods described in this application.

Preparation of Luohanguo flavored fruit vinegar

The invention relates to Luo Han Guo flavored fruit vinegar, which includes Luo Han Guo flavored extract, Luo Han Guo non-glycoside juice and Luo Han Guo raw vinegar. The total sugar content (% w / w) of Luo Han Guo in the drink is about 3.0-9.0%, 3.5-8.5%, 4.0-8.0%, 4.5-7.5%, 5.0-7.0%, 5.5-6.5%.

In some embodiments, the total acid content (% w / w) of the Luo Han Guo flavored fruit vinegar is 0.1 to 0.5%, 0.15 to 0.45%, 0.15 to 0.40%, 0.15 to 0.35%, 0.15 to 0.30%, 0.15 to 0.25 %, 0.20 to 0.45%, 0.20 to 0.40%, 0.20 to 0.35%, 0.20 to 0.30%, 0.25 to 0.45%, 0.25 to 0.40%, 0.25 to 0.35%, 0.25 to 0.30%, 0.20% to 0.3%.

In some embodiments, the total sugar and total acid ratio (sweet acid ratio) of the Luo Han Guo flavored fruit vinegar has a higher relative preference at 18 to 28/1, for example, 19 to 27/1, 20 to 26 / 1. Sweet and sour ratio of 21-25 / 1, 22-24 / 1, 23-24 / 1. Among them, the sweet-acid ratio is the ratio of the total sugar content (weight percent) of Luohanguo flavored fruit vinegar to the total acid content (weight percent) of Luohanguo flavored fruit vinegar, for example, when the total sugar content is 3%, the total acid content When it is 0.1%, the sweet-acid ratio is 3% / 0.1% = 30/1.

In some embodiments, the addition amount of Luo Han Guo flavor extract (expressed as a percentage based on dry weight (% w / w)) in the above Luo Han Guo flavor fruit vinegar can make the product have a good Luo Han Guo flavor mouthfeel at 0.2-0.6%, The addition amount is preferably 0.2 to 0.6%, 0.3 to 0.5%, and 0.3 to 0.4%. The introduction of Luo Han Guo flavor extract has a significant effect on the taste preference of Luo Han Guo flavor vinegar.

Unless otherwise stated, the percentages of the present invention are percentages by weight.

Sample taste test method of the invention

Sample taste test method

Sensory test requirements:

Sensory laboratory: requires good ventilation, lighting suitable for sensory evaluation, indoor color and temperature, and sufficient space.

Sensory evaluator: Can objectively evaluate the product's characteristic smell, taste, etc., has good communication skills and language expression ability.

Evaluation requirements: Do not eat, drink or smoke at least half an hour before the evaluation, and do not use perfume on your body. Focus on the spirit and take the evaluation seriously. After evaluating a sample, rinse your mouth with pure water and clean your mouth and rest thoroughly before you evaluate the next sample.

Evaluation steps:

Sample tasting temperature

The sample tasting temperature is about 50 ° C, and the heat preservation is maintained.

Evaluation and data processing

Prepare samples according to the above requirements, please sensory appraisers to taste and score the indicators in Table 1 below according to the standard of Table 2, statistic the score data of each appraiser, effective evaluation data is not less than 10 groups, draw sensory analysis radar based on the data Figure.

For the sensory test indexes shown in Table 1, the sensory test results are defined on a 0-9 scale.

Table 1 Luo Han Guo product sensory test index

Table 2 Scoring criteria for sensory tests

Method for measuring product parameters of the present invention

It should be understood that the amount of mogroside V in the product of the present invention is expressed as a weight percentage based on the dry weight of the product of the present invention. The amount of mogroside V in the product of the present invention can be determined by high performance liquid chromatography (HPLC). It should also generally be understood that the content of mogroside V in the products of the present invention can be determined by any suitable method or technique known in the art.

It should also be understood that the sugar content of the product of the present invention can be determined by any suitable method or technique known in the art. The monosaccharides in the product of the present invention are naturally present in the Luo Han Guo, from which the product of the present invention is obtained. Such monosaccharides can include, for example, glucose and fructose. In some embodiments, the product of the present invention further comprises a disaccharide, which is naturally present in the Luo Han Guo from which the product of the present invention is obtained. Such disaccharides can include, for example, sucrose. It should also be understood that the disaccharide content of the product of the present invention can be determined by any suitable method or technique known in the art. In some embodiments, the products of the present invention further comprise proteins or fragments or derivatives thereof, which are naturally present in the Luo Han Guo from which the products of the present invention are obtained. In some embodiments, the product of the present invention has the following weight percent content of protein or fragment or derivative thereof: at least about 10%, at least about 11%, at least about 12%, at least about 13%, at least about 14%, at least about 15%, at least about 16%, at least about 17%, or at least about 18%. In certain embodiments, the amount of protein or fragments or derivatives thereof in the product of the invention does not exceed about 18%, about 17%, about 16%, or about 15% based on dry weight. In other embodiments, the products of the present invention have the following levels of protein or fragments or derivatives thereof: about 10% to about 18%, about 12% to about 18%, about 14% to about 18%, about 10% to about 16%, about 12% to about 16%, about 14% to about 16%, about 10% to about 14%, or about 12% to about 14%.

It should be understood that the amount of protein or fragments or derivatives thereof in the product of the present invention is expressed as a weight percentage based on the dry weight of the product of the present invention. In some of the foregoing embodiments, the amount of protein or fragments or derivatives thereof in the product of the present invention is determined by the Kjeldahl method. It should also generally be understood that the content of protein or fragments or derivatives thereof in the products of the present invention can be determined by any suitable method or technique known in the art.

In some embodiments, the product of the present invention further comprises fruit acid, which is naturally present in the Luo Han Guo from which the product of the present invention is obtained. Such naturally occurring fruit acids may include, for example, ascorbic acid, citric acid, and lactic acid.

In some embodiments, the total acid content of the product of the present invention is expressed as a weight percentage, based on the dry weight of the product of the present invention. In one embodiment, the total acid content is determined by the acid-base titration method. According to the acid-base neutralization principle, the acid in the test solution is titrated with an alkali drop, and the titration end point is determined using phenolphthalein as an indicator. Calculate the total acid content in the product according to the consumption of lye, that is, titrate the acidity, which is calculated based on the measurement of the conversion coefficient of citric acid.

In some embodiments, the product of the present invention further comprises other terpene glycosides (ie, terpene glycosides other than mogroside V), which are naturally present in the Luo Han Guo from which the product of the present invention is obtained. Such terpene glycosides include mogrosides other than mogroside V (e.g. mogrosides I, II, III, IV, and VI) and siamin, and their chemical structures are known in the art.

The physical and chemical indexes of the samples in the examples of the present invention are measured by the following methods.

1. Mogroside V content detection

Using the standardized content of mogroside V as the standard product, the mogroside was detected by the high-performance liquid chromatography (HPLC) peak area external standard method in the national food safety standard "food additive mogroside" (GB1886.77-2016) V content.

2. Detection of total acid

Refer to the standard GB / T 12456-2008 "Determination of Total Acids in Food" for determination.

3. Detection of total protein content of plants

According to the first method of GB 5009.5-2010 "Determination of Protein in Food", Kjeldahl method is used for determination.

4. Determination of sucrose, glucose, fructose

Refer to the standard GB / T 22221-2008 "Determination of Fructose, Glucose, Sucrose, Maltose, Lactose in Foods by High Performance Liquid Chromatography" for the determination.

5. Determination of total sugar and monosaccharide

The total sugar content defined herein is the sum of the three sugar contents of sucrose, glucose and fructose.

The detection method of each sugar is tested according to the fourth detection method.

The percentages of the above product parameters of the present invention are based on the weight percentage of dry weight.

It should be understood that the above product parameters of the present invention include the numerical values at both ends of the above parameter range and any positive integers and decimals between the two ends. The above product parameters of the present invention and their variation ranges can be combined with each other. These combinations are included in the scope of the present disclosure. Although they are not listed one by one, they all belong to the content disclosed in this application, just like each combination is individually List the same.

It should be understood that "about" values or parameters cited in this application include (and describe) embodiments directed to the values or parameters themselves. For example, a description related to "about x" includes a description about "x" itself. In other cases, the term "about" is used in conjunction with other measurements or to modify a numerical value, unit, constant, or series of values. In some embodiments, about refers to a +/- 20% change. In other embodiments, about refers to a +/- 15% change. In one embodiment, about refers to a +/- 10% change. In one variation, about refers to a +/- 5% change. In another variation, about refers to a +/- 1% change.

Examples

Example 1 Preparation of Mogrosides Concentrated Solution

Luo Han Guo Extract A:

Take 60kg of fresh Luo Han Guo, crush it into small particles by a crusher, and then place it in the countercurrent extraction equipment, add 200L of hot water with a temperature of 90 ° C for extraction, remove the pomace, and after centrifugation, clarify it by ultrafiltration with a ceramic membrane of 200nm pore size. Filtrate.

Luo Han Guo Glycoside Concentrate B:

Apply Luohanguo extract A to a chromatography column (120 * 1200mm) equipped with 10L adsorption resin (SEPABEADS SP700) and perform adsorption chromatography to adsorb mogroside. Collect the column effluent 120L that is not adsorbed by the resin column and vacuum. Concentrate to obtain 6.5L of Mogrosides Concentrate B with solid content Brix = 40%. Physical and chemical testing showed that the pH was 4.31, the total sugar content was 22.94% (wt.%, The same below), and the total acid was 5.24% (calculated as citric acid, wt.%, The same below).

Luo Han Guo Glycoside Concentrate C:

Take Luo Han Guo glycosides-concentrated solution B20L, conduct membrane separation and concentration through a 500 Dalton nanofiltration membrane (NFW1812), and collect the membrane permeate. When Brix = 18% of the side material to be retained, the separation is stopped and the membrane permeate is removed. Carry out vacuum concentration to obtain 1.1L solid content of Brix = 30% Mogrosides Concentrate C. The physical and chemical indicators detected its pH4.58, total sugar content 23.23%, total acid 2.69%.

Luo Han Guo Glycoside Concentrate D:

Take Luo Han Guo glycosides-concentrated solution B20L, dilute it with purified water to a solid content of Brix = 8%, use a 1000 Dalton nanofiltration membrane (GE1812C34D) for membrane separation and concentration, collect membrane permeate, and wait for the side material to be retained. = 15%, add 5L of water to wash the concentrated liquid, and continue to separate. When the concentration of the material on the intercepting side reaches Brix = 15% again, the separation is stopped, and the membrane permeate is concentrated in vacuo to obtain 2.4L solid content Brix = 16.5% Luo Han Guo Concentrate D without glycosides. The physicochemical index detected the pH of 4.6, the total sugar content was 13.6%, and the total acid was 1.3%.

Example 2: Preparation of Luo Han Guo flavor extract (E)

1) Take 25L of the above-mentioned column effluent, which is not adsorbed by the resin column and obtained by the preparation method of Luo Han Guo glycoside-free concentrated solution B, and clarify it by ultrafiltration of a 100k Dalton ceramic membrane, and then use a nanofiltration membrane with a molecular weight cut-off of 500 ( NFW1812), carry out membrane separation and concentration, control the inlet pressure ≤1MPa, after 3.5h, the solid content of the concentrated solution on the interception side Brix = 13.0%, volume 3.20L, add 1.6L of water to the interception side concentrated solution to dilute, continue separation and concentration, wait until The solid content of the concentrated solution on the retention side is Brix = 11%, and then diluted with the same amount of water, and washed and concentrated twice. The third concentration is until the solid content is ≥ 10%. The concentration is stopped, and the concentrated solution on the retention side is released. The volume is 1.65L. The content is Brix = 11.8%. The released concentrated liquid on the intercepting side was concentrated under reduced pressure to obtain a Luo Han Guo flavor extract with a solid content of 33.0% and a volume of 580 mL. The concentrated solution was spray-dried to obtain 180 g (E1) of Luo Han Guo flavor extract powder.

2) Take 30L of the adsorbed column effluent obtained according to the preparation method of Luo Han Guo Glycoside Concentrate B above, clarify by ultrafiltration of 100k Dalton ceramic membrane, and then use a molecular membrane with molecular weight cutoff of 1000 (membrane specification is 1812 type) , Perform membrane separation and concentration, control the inlet pressure ≤1MPa, after 3.3h, the solid content of the concentrated solution on the retention side is Brix = 10.7%, the volume is 3.95L, add 2.0L of water to the concentrated solution to dilute, continue to separate and concentrate, and concentrate the solution on the retention side Solid content Brix% = 10, then dilute with the same amount of water, wash and concentrate twice, concentrate for the third time to solid content ≥10%, stop concentration, release concentrated liquid, volume 1.73L, solid content Brix% = 10.7 . The concentrated solution was then concentrated under reduced pressure to obtain a solid substance content of 35.2%, a volume of 525ml of Luo Han Guo flavor extract, the concentrate was spray dried to obtain 215g (E2) of Luo Han Guo flavor extract powder.

The Luo Han Guo flavor extract obtained by the above method can be used for flavoring of Luo Han Guo flavor fermented drinks directly or after drying to form a solid powder.

Example 3: Preparation of Luo Han Guo non-glycoside juice (F)

Take 10L Mogroside Concentrate B, dilute it to Brix = 5%, filter and clarify with a ceramic membrane with a pore size of 200nm, and the clear liquid enters into a resin column group composed of 1L cation exchange resin and 1L anion exchange resin respectively, and the flow rate through the column is controlled At 1 L / h, the column effluent was collected and concentrated in vacuo to obtain 423 g of Luo Han Guo non-glycoside juice with a solid content of Brix = 66%. The physical and chemical indicators were tested at pH 5.2 and the total sugar content was 56.25%.

Example 4: Preparation of Luo Han Guo flavored wine

Preparation of Luohanguo flavored fruit wine (G)

Luo Han Guo flavor wine G1:

Take Luo Han Guo extract A and concentrate it under reduced pressure until the solid content of the concentrated solution is 20%. The concentration of mogroside V in the concentrated solution is 0.59%, which is equivalent to the total sugar concentration of about 11.76%.

Weigh 1.0g of wine yeast (Shandong Shengqi Biological Co., Ltd.), dissolve it in 10mL of Brix = 5% Luo Han Guo glycosides-free juice (F), stir well, and place Luo Han Guo glycosides-free juice (F) in 35-37 Incubate for 30 min at ℃, then inoculate the activated yeast solution into the dilution of 1000 mL of Luo Han Guo extract A to be fermented, stir evenly, control the fermentation temperature of 26-30 ℃, and the total sugar in the fermentation broth after 24 hours of fermentation When the content drops below 1%, the fermentation is stopped. The fermentation broth was clarified by filtration, and the filtrate was placed in a sterilization tank and heated to 80-85 ° C for 30 minutes, then cooled to room temperature, and sampled for testing. The physical and chemical test results of Luo Han Guo flavored wine: alcoholic degree 5.3%, total sugar content 0.86%, pH 4.4.

Luohanguo fruit wine G2:

Take Luo Han Guo glycoside concentrate B as the fermentation substrate and dilute with water to obtain a solid content of 18%, which is equivalent to a total sugar concentration of about 10.32%. The solution was measured at pH 4.3.

Weigh 1.0g of wine yeast (Shandong Shengqi Biological Co., Ltd.), dissolve it in 10mL of Brix = 5% Luo Han Guo non-glycoside juice solution (F), stir well, and incubate the solution at 35-37 ℃ for 30min, Then inoculate the activated yeast solution into the dilution of 1000 mL of Luo Han Guo Glycoside Concentrate B to be fermented, stir evenly, control the fermentation temperature of 26-30 ℃, after fermentation continues for 24h, sampling and monitoring every 2h, when the fermentation broth When the total sugar content drops below 1%, the fermentation is stopped. The fermentation broth was clarified by filtration, and the filtrate was placed in a sterilization tank and heated to 80-85 ° C for 30 minutes, then cooled to room temperature, and sampled for testing. Fruit wine physical and chemical test results: alcohol 5.3%, total sugar content 0.86%, pH4.4.

Luohanguo fruit wine G3:

Take Luo Han Guo Glycoside Concentrate B as the fermentation substrate, dilute with water to make the solid content about 16%, equivalent to the total sugar concentration of about 9.18%, the solution was measured pH5.1.

Measure 1000mL of fermentation substrate, weigh 1.4g of Saccharomyces cerevisiae (RW type, Angel Yeast Co., Ltd.), dissolve it in 10mL of Brix = 5% Luo Han Guo non-glycoside juice solution (F), stir evenly, and The solution was incubated at 35 ° C for 30 min, and then the activated cider yeast solution was inoculated into the 1000 mL of Luo Han Guo glycoside-free concentrate B to be fermented, stirred evenly, and the fermentation temperature was controlled at 26-30 ° C. After the fermentation continued for 20 h, the fermentation The total sugar content in the liquid is 0.04g / mL, and the fermentation is stopped. The fermentation broth was clarified by filtration, and the filtrate was placed in a sterilization tank and heated to 80-85 ° C for 30 minutes, then cooled to room temperature, and sampled for testing. Fruit wine physicochemical test results: alcohol content 4.3%, total sugar content 0.04%, pH 5.47.

Luohanguo fruit wine G4:

Take Luo Han Guo Glycoside Concentrate B as the fermentation substrate and dilute with water so that the solid content is about 18%, which is equivalent to the total sugar concentration of about 10.23%, and the solution has a measured pH of 4.4.

Measure 1000mL of fermentation substrate, weigh 1.5g of yeast dedicated to fruit wine (Yantai Disba Yeast Co., Ltd.), dissolve it in 10mL of Brix = 5% Luo Han Guo non-glycoside juice solution (F), stir well, and place the solution Incubate at 35 ° C for 30 minutes, then inoculate the activated fruit wine yeast solution into the 1000 mL of diluent of Mogrosides Concentrate B to be fermented, stir evenly, control the fermentation temperature at 28 ° C, and the fermentation process lasts for 20 hours. The sugar content was 0.017g / mL and the fermentation was stopped. The fermentation broth was clarified by filtration, and the filtrate was placed in a sterilization tank and heated to 80-85 ° C for 30 minutes, then cooled to room temperature, and sampled for testing. Fruit wine physicochemical test results: alcohol degree 5.8%, total sugar content 0.02%, pH 5.72.

Luohanguo fruit wine G5:

Take Luo Han Guo Glycoside Concentrate C as the fermentation substrate, dilute with water to make the solid content 16.5%, the pH of the solution is 6.15, and take 1000 mL to be fermented.

Weigh 1.5g of wine yeast (Yantai Dishibo Yeast Co., Ltd.), dissolve it in 10mL of Brix = 5% Luohanguo non-glycoside juice solution (F), stir evenly, and incubate the solution at 35 ° C for 30min, then Inoculate the activated fruit wine yeast solution into the dilution of 1000 mL of Luo Han Guo Glycoside Concentrate C to be fermented, stir evenly, control the fermentation temperature at 29 ° C, and the fermentation continues for 54 h, the total sugar content in the fermentation broth is 0.001 g / mL, Stop fermentation. The fermentation broth was clarified by filtration, and the filtrate was placed in a sterilization tank and heated to 80-85 ° C for 30 minutes, then cooled to room temperature, and sampled for testing. Fruit wine physical and chemical test results: alcohol 8.0%, total sugar content 0.001%, pH5.30.

Luohanguo fruit wine G6:

Take Luo Han Guo Glycoside Concentrate C as the fermentation substrate and dilute with water to obtain a solid content of 15%, which is equivalent to a total sugar concentration of 11.65%, and the solution was measured at pH 4.3.

Weigh 1.0g of wine yeast (Shandong Shengqi Biological Co., Ltd.), dissolve it in 10mL of Brix = 5% Luo Han Guo non-glycoside juice solution (F), stir well, and incubate the solution at 35-37 ℃ for 30min, After that, the activated yeast solution was inoculated into the diluted solution of 1000 mL of Luo Han Guo glycosides concentrate C to be fermented, stirred evenly, and the fermentation temperature was controlled at 26-30 ° C. After the fermentation continued for 27 hours, the fermentation was stopped. The fermentation broth was clarified by filtration, and the filtrate was placed in a sterilization tank and heated to 80-85 ° C for 30 minutes, then cooled to room temperature, and sampled for testing. Fruit wine physical and chemical test results: alcohol 5.9%, total sugar content 0.046%, pH5.5.

Luohanguo fruit wine G7:

Take 1000mL of Luo Han Guo glycosides-free concentrated liquid D as a fermentation substrate, weigh 1.0g of wine yeast (Shandong Shengqi Biological Co., Ltd.), dissolve it with Brix = 5% Luo Han Guo glycosides-free juice solution (F) 10mL, stir well, Incubate the solution at 35-37 ° C for 30min, then inoculate the activated yeast solution into the fermentation substrate to be fermented, stir it evenly, control the fermentation temperature at 26-30 ° C, and the fermentation stops after 27h of fermentation. The fermentation broth was clarified by filtration, and the filtrate was placed in a sterilization tank and heated to 80-85 ° C for 30 minutes, then cooled to room temperature, and sampled for testing. Fruit wine physical and chemical test results: alcohol 6.9%, total sugar content 0.022%, pH5.5.

Luohanguo fruit wine G8:

Take Luo Han Guo Glycoside Concentrate B as the fermentation substrate, dilute with water to 25% solid content, and perform desalination treatment according to the cation exchange resin with net weight of solid 1: 1 (m / V). The obtained desalting liquid has a solid content of about 18 %, Equivalent to a total sugar concentration of about 10.03%, the solution was measured at pH 4.1.

Measure 1000mL of fermentation substrate, weigh 1.5g of yeast dedicated to fruit wine (Yantai Disba Yeast Co., Ltd.), dissolve it in 10mL of Brix = 5% Luo Han Guo non-glycoside juice solution (F), stir well, and place the solution Incubate at 35 ° C for 30 min, then inoculate the activated fruit wine yeast solution into the 1000 mL of diluent of Luo Han Guo without glycoside concentrate A to be fermented, stir evenly, and control the fermentation temperature at 28 ° C. After the fermentation lasts for 20 h, the total amount in the fermentation broth The sugar content was 0.017g / mL and the fermentation was stopped. The fermentation broth was clarified by filtration, and the filtrate was placed in a sterilization tank and heated to 80-85 ° C for 30 minutes, then cooled to room temperature, and sampled for testing. Fruit wine physicochemical test results: alcohol content 5.2%, total sugar content 0.02%, pH 5.12.

Luohanguo fruit wine G9:

Take Luo Han Guo Glycoside Concentrate C as fermentation substrate, dilute with water, dilute with water to 25% solid content, perform desalination treatment according to cation exchange resin with net weight of solid 1: 2 (m / V), adjust solid content to 16.5%, Adjust the solution to pH 4.2 with food-grade ammonia, and measure 1000mL, to be inoculated and fermented.

Weigh 1.5g of wine yeast (Yantai Dishibo Yeast Co., Ltd.), dissolve it in 10mL of Brix = 5% Luohanguo non-glycoside juice solution (F), stir evenly, and incubate the solution at 35 ° C for 30min, then Inoculate the activated fruit wine yeast solution into the dilution of 1000 mL of Luo Han Guo Glycoside Concentrate A to be fermented, stir evenly, control the fermentation temperature at 31 ° C, and the fermentation continues for 48h, the total sugar content in the fermentation broth is 0.001g / mL, Stop fermentation. The fermentation broth was clarified by filtration, and the filtrate was placed in a sterilization tank and heated to 80-85 ° C for 30 minutes, then cooled to room temperature, and sampled for testing. Fruit wine physical and chemical test results: alcohol 8.0%, total sugar content 0.001%, pH5.30.

Example 5: Comparative study of fermentation products

In this example, the contents and changes of six glycosides at different time points in the fermentation process of the above-mentioned Luo Han Guo glycoside-free concentrates A and B were studied. Figure 1 shows the HPLC profiles of 6 glycosides: 11-O-mogroside V; mogroside V; saligenin I; mogroside IVe; mogroside III and mogroside IIe. Among them, peak 1 is: 11-O-mogroside V; peak 2 is: mogroside V; peak 3 is: siamin I; peak 4 is: mogroside IVe; peak 5 is: mogroside III ; Peak 6 is: Mogroside IIe. The following figures are the same. Figure 2 to Figure 8 show the HPLC profile of the sample B without glycoside concentrate and the comparison of the HPLC profile of the sample and the control when the fermentation time is 0, 6, 12, 18, 24, 30, and 36 hours, respectively.

As a result of the study, a small amount of mogrosides remained in the above-mentioned glycoside-free concentrated solution B (see FIGS. 2A-2B). With the continuation of alcohol fermentation time, the contents of 11-O-glycoside V and glycoside V continued to decrease to 0, and the content of thymidine I, glycoside IVe, glycoside III, and glycoside IIe increased first and then decreased. During the alcohol fermentation process, the above 6 glycosides will eventually be decomposed and converted into other substances as long as there is enough time, and the sugar groups on the glycosides are degraded one by one during the degradation process, forming secondary glycosides or aglycones with a bitter taste (See Figure 9).

Mogroside Glycoside Concentrate B is a by-product in the current mogroside V production process. Fermenting it directly will form a bitter taste secondary glycoside or aglycone.

The taste test of products obtained by using Luo Han Guo extract A, glycoside-free concentrates B, C and D as fermentation substrates, respectively, found that the fermented wine after deglycosidation treatment was directly fermented without deglycosidation. Compared with fruit wine, after bitterness, off-flavor, sourness, taste and other indicators have been greatly improved (see Figure 11). Compared with the direct fermentation of Luo Han Guo extract, the remaining glycosides in the effluent from the column are greatly reduced, the bitter taste components formed by fermentation are reduced, and the resulting bitter taste of the wine is reduced, but it is still significant. The sweetness in the fermentation substrate after membrane treatment The glycosides are completely separated, and the bitter taste of the fruit wine obtained by fermentation is significantly reduced. It is an effective way to greatly improve the product quality by processing Luo Han Guo extract or column effluent and then fermenting to prepare fruit wine.

Example 6: Fermentation preparation of Luo Han Guo raw vinegar (H)

Mangosteen Raw Vinegar H1

Place 500mL Luohanguo fruit wine G2 in a fermenter, heat 80 ℃, keep it warm for 30min, cool to 33 ℃, weigh 2.5g of active acetic acid bacteria (AS.1.41) into the fermentor, airtight, let in sterile air, and the ventilation rate is 130mL / h, continuous stirring fermentation, controlling fermentation temperature 33-35 ℃, monitoring the total acid change in the fermentation broth, continuous fermentation, the total acid in the fermentation broth was 4.41% and 4.25% on day 6 and 6.5, respectively, but the total acid in the fermentation broth was not Increase again, stop the fermentation, directly heat the fermentation broth to 85 ℃, keep it warm for 30 minutes, cool to room temperature, filter and clarify, the obtained filtered clear liquid is Luo Han Guo raw vinegar H1. The total acid content of raw vinegar is 4.20%, and the conversion rate of alkyd is 67.9%.

Mangosteen Raw Vinegar H2

Place 500mL Luohanguo fruit wine G4 in the fermentor, dilute it with 80mL of purified water to reduce the alcohol content of the solution to 5.0%, heat at 80 ℃, keep for 30min, cool to 33 ℃, weigh 2.5g of active acetic acid bacteria (AS.1.41) into the fermentation In the tank, airtight, with sterile air, ventilation rate of 130mL / h, continuous stirring fermentation, control fermentation temperature 33-35 ℃, monitoring the total acid change of fermentation broth, continuous fermentation, total fermentation broth on days 6.5 and 7 The acid was 3.91% and 3.89% respectively. The total acid in the fermentation broth was no longer elevated. The fermentation was stopped. The fermentation broth was directly heated to 85 ° C, kept for 30 minutes, cooled to room temperature, and clarified by filtration. The resulting filtered clear liquid was Luo Han Guo raw vinegar H2. The total acid content of raw vinegar is 3.89%, and the conversion rate of alkyd is 71.9%.

Mangosteen Raw Vinegar H3

Place 500mL Luohanguo fruit wine G5 in a fermentation tank, add 300mL of purified water to dilute the alcohol content of the solution to 5.0%, heat at 80 ℃, keep for 30min, cool to 33 ℃, weigh 2.5g of active acetic acid bacteria (AS.1.41) into fermentation In the tank, airtight, with sterile air, ventilation rate of 130mL / h, continuous stirring fermentation, control fermentation temperature 33-35 ℃, monitoring the total acid change of fermentation broth, continuous fermentation, total fermentation broth on days 6.5 and 7 The acid was 4.01% and 3.98% respectively. The total acid in the fermentation broth was no longer elevated. The fermentation was stopped. The fermentation broth was directly heated to 85 ° C, held for 30 minutes, cooled to room temperature, and clarified by filtration. The resulting filtered clear liquid was Luo Han Guo raw vinegar H3. The total acid content of raw vinegar is 3.98%, and the conversion rate of alkyd is 72.2%.

Mangosteen Raw Vinegar H4

Place 500mL Luohanguo fruit wine G7 in the fermentor, add 190mL of purified water to dilute the alcohol content of the solution to 5.0%, heat at 80 ℃, hold for 30min, cool to 33 ℃, weigh 2.5g of active acetic acid bacteria (AS.1.41) into fermentation In the tank, airtight, with sterile air, ventilation rate of 130mL / h, continuous stirring fermentation, control fermentation temperature 33-35 ℃, monitoring the total acid change of fermentation broth, continuous fermentation, total fermentation broth on days 6.5 and 7 The acid was 3.71% and 3.70% respectively. The total acid in the fermentation broth was no longer elevated. The fermentation was stopped. The fermentation broth was directly heated to 85 ° C, kept at room temperature for 30 minutes, cooled to room temperature, and clarified by filtration. The obtained filtered clear liquid was Luo Han Guo raw vinegar H4. The total acid content of raw vinegar is 3.70%, and the conversion rate of alkyd is 68.2%.

Mangosteen Raw Vinegar H5

Place 500mL Luohanguo fruit wine G8 in a fermenter, heat at 80 ℃, keep it warm for 30min, cool to 30 ℃, weigh 2.5g of active acetic acid bacteria (AS.1.41) into the fermentor, seal it, and let in sterile air, the ventilation rate is 170mL / h, continuous stirring fermentation, controlling fermentation temperature 30-35 ℃, monitoring the total acid change in the fermentation broth, continuous fermentation, the total acid in the fermentation broth at 22h and 26h is 4.33% and 4.01%, respectively, the total acid in the fermentation broth no longer rises High, stop the fermentation, directly heat the fermentation broth to 85 ℃, keep it warm for 30 minutes, cool to room temperature, filter and clarify, the obtained filtered clear liquid is Luo Han Guo raw vinegar H5. The total acid content of raw vinegar is 4.00%, and the conversion rate of alkyd is 71.54%.

Mangosteen Raw Vinegar H6

Put 500mL Luohanguo fruit wine G9 in the fermentor, heat 80 ℃, keep it warm for 30min, cool to 30 ℃, weigh 2.5g active acetic acid bacteria (AS.1.41) into the fermentor, close it, pass the sterile air, the ventilation rate is 150mL / h, continuous stirring fermentation, controlling fermentation temperature 30-35 ℃, monitoring the total acid change of fermentation broth, continuous fermentation, total acid in fermentation broth at 4.77% and 4.70% at 40h and 44h, total acid in fermentation broth no longer rises High, stop the fermentation, directly heat the fermentation broth to 85 ℃, keep it warm for 30 minutes, cool to room temperature, filter and clarify, the obtained filtered clear liquid is Luo Han Guo raw vinegar H6. The total acid content of raw vinegar is 4.70%, and the conversion rate of alkyd is 66.11%.

Example 7: Effect of desalting treatment on products

Although the deglycosylated Luo Han Guo extract after separation by adsorption resin or membrane reduces or avoids the hydrolysis of glycosides during fermentation to form bitter components, the fermentation substrate--de-glycosylated Luo Han Guo extract still contains a lot of inorganic Salt and other ingredients, these ingredients will produce a significant salty taste in the fermented fruit wine, and will form a soy sauce-like smell in the resulting fruit vinegar product. Although soy sauce odor and salty taste are beneficial for vinegar products in seasoned vinegar, as beverage wine and vinegar, these two flavors obviously reduce the competitiveness of the product. To this end, the inventors used a cation exchange resin to treat the fermentation substrate, the taste of the fruit wine obtained after fermentation of the processed product was controlled, and the fruit vinegar product had no odor of soy sauce and pure taste.

The column effluent is used as a fermentation substrate. When the ash content is close to 2%, the fruit wine produced by fermentation begins to have a salty taste. With the increase in the amount of resin treatment, the ash content in the product gradually increases, and the salty taste is significant. increase. The corresponding soy sauce flavor of fruit vinegar is similar to that of fruit wine's salty taste.

Because the pH value of the material treated by the cation exchange resin is low, it is not suitable for the fermentation of fruit wine. If alkali metal alkalis (such as NaOH, KOH, etc.) are used to adjust the pH, metal ions will be re-introduced into salt to produce salty taste, so ammonia is used It is a feasible method to use as a PH regulator. Another way to adjust the pH value is to connect anion exchange resin in series with the cation exchange resin during desalination, control the appropriate ratio of positive and negative resins and the amount of feed, so that the effluent can obtain a suitable pH for fruit wine fermentation while desalting. The pH value can be controlled without adding any regulator.

Example 8 Preparation of Luohanguo flavored fruit vinegar

1. The optimal study of the acidity of Luohanguo fruit vinegar

The ratio of the design of Luo Han Guo raw vinegar H3 and water was investigated according to the total acid concentration of 0.05%, 0.10%, 0.15%, 0.20%, 0.25%, 0.35%, 0.45%, 0.5%, 0.55%. Then at each total acid concentration, different sweet and acid ratios (ie, total sugar content to total acid content ratio) were adjusted with Luo Han Guo non-glycoside juice, and the obtained solution was tested by the tasting team. , Softness, refreshing degree, comprehensive preference, etc. The samples are evaluated according to 0-9 grades, the correlation between the obtained preference and different sweet-acid ratios under different total acid content, to obtain the best total acid content and sweetness of the drink Acid ratio.

It was found that when the total acid level was 0.05%, the maximum preference of the solution under different sweet-acid ratios was only 5.5 points. The main taste defect was the weak representation of vinegar beverages. When the total acid level reaches 0.55%, because the total acid level is higher, even if the sweet acid ratio is increased, it still has a high irritancy, and the softness of the product decreases significantly.

As shown in FIG. 12A, the total acid of Luohanguo fruit vinegar beverage has a higher relative preference (more than 6 points) when it is in the range of 0.10-0.50%, preferably the total acid content is 0.20% to 0.3%. The corresponding sweet acid is shown in the figure. 12B.

Based on the determination of the total acid of the fruit vinegar beverage at 0.15 to 0.45%, the parameters of the total sugar and total acid content ratio of Luohanguo fruit vinegar and Luohanguo non-glycoside juice were investigated, and the obtained solution was tasted by the team Tests were conducted and it was found that the sweet-acid ratio of Luohanguo flavored fruit vinegar has a higher relative preference at 18-28 / 1, preferably a sweet-acid ratio of 20-26 / 1.

When the total acidity is 0.3%, and the sweetness-acid ratio is 24/1, the highest preference is obtained in the mouth of the taste tester, and the product's fruit vinegar flavor characteristics and sweet-sweet coordination are optimized.

2. Study on the optimization of the amount of Luo Han Guo flavor extract in Luo Han Guo flavor fruit vinegar

The fruit vinegar beverage with a total acid content of 0.3% and a sweet-acid ratio of 24/1 was added with different levels of Luo Han Guo flavor extracts to test its taste performance. As a result, it was found that the amount of flavor extracts added as dry products was in quality At a ratio of 0.2 to 0.6%, it has a good mouthfeel, and preferably an added amount of 0.3 to 0.4% (see FIG. 13). The introduction of Luo Han Guo flavor extract has significantly improved the taste preference of Luo Han Guo vinegar beverage, mainly reflected in the product aroma, flavor coordination and palatability of vinegar beverage has been significantly improved, when the added amount reaches 0.4g / 100mL, The taste of fruit vinegar drinks is the best. Afterwards, with the gradual increase of the amount of addition, the taste of the tasters decreased significantly, mainly due to the increase in flavor substances, the aroma of Luo Han Guo charcoal in the substances is too rich, covering the unique aroma of vinegar drinks , And the astringency and softness decreased.

Take Luo Han Guo raw vinegar H4 prepared in Example 6: 100mL, Luo Han Guo non-glycoside juice F in Example 3: 157.9mL, Luo Han Guo flavor extract E1 in Example 2: 4.9g, dilute with purified water to 1233.3mL, stir well , Filled after filtering and sterilizing to get Luo Han Guo flavored fruit vinegar drink.

Fig. 14 shows the comparison of the taste test results of the fruit vinegar drink after completion of the preparation and the unadjusted original vinegar dilution H4 (total acid concentration 0.3%).

As shown in the figure, at the same total acid concentration (0.3g / 100mL), the blended fruit vinegar is compared with the original vinegar dilution H4 in terms of appearance, flavor recognition, acidity, sweetness, mouthfeel and preference. On the former, the former is much higher than the latter. It means that the fruit vinegar beverage obtained by directly diluting the original vinegar cannot achieve good sensory quality, and needs to be formulated before it can become a vinegar beverage product that people are willing to accept.

Claims

A method for preparing mangosteen fruit wine includes:

a) Preparation of Luo Han Guo glycoside-free concentrate; and

b) Fermentation of Luo Han Guo glycoside-free concentrate.
The method of claim 1, wherein the preparation of the Luo Han Guo glycoside-free concentrate comprises:

(i) Luo Han Guo is subjected to water extraction after crushing to prepare Luo Han Guo water extract;

(ii) Clarified water extract;

(iii) Perform adsorption chromatography on the clarified water extract and collect the effluent; and

(iv) Nanofiltration treatment of the effluent.
The method of claim 1 or 2, wherein the adsorption resin of the adsorption chromatography is a macroporous polymer adsorption resin.
The method of claim 3, wherein the macroporous polymer adsorption resin is a styrene-based copolymer, a styrene-divinylbenzene copolymer or a divinylbenzene copolymer.
The method according to any one of claims 1 to 4, wherein the nanofiltration membrane used for the nanofiltration is 500-

5000 Daltons.
The method of any one of claims 1 to 5, wherein the content of mogroside V in the glycoside-free concentrate is less than 0.5% (w / w).
The method according to any one of claims 1 to 6, which comprises desalting the glycoside-free concentrated liquid before fermentation.
Luohanguo fruit wine prepared by the method of any one of claims 1-7.
A method for preparing raw grosvenor grosvenor vinegar, which comprises fermenting grosvenori fruit wine prepared by the method according to any one of claims 1-7 with acetic acid bacteria.
Luohanguo raw vinegar prepared by the method of claim 9.
A method for preparing mangosteen fruit vinegar comprises blending mangosteen raw vinegar of claim 10 so that the total acid concentration is 0.10-0.50% (w / w), preferably 0.20% -0.3% (w / w).
A method for preparing mangosteen flavored drink, comprising adding mangosteen flavor extract to mangosteen raw vinegar of claim 10 or mangosteen fruit vinegar of claim 11, wherein the preparation method of mangosteen flavor extract is as follows:

a) Mogroside extract after mogroside V extraction;

b) Nanofiltration membrane filtering step a) the obtained Luo Han Guo extract to prepare retentate;

c) monitor the soluble solid concentration of the retentate of step b);

d) When the soluble solid concentration of the retentate of step b) is 3-8g / 100g, dilute the retentate of step c) with water to prepare a diluted retentate;

e) Use nanofiltration membrane to filter the diluted retentate to prepare another retentate;

f) monitor the soluble solids concentration of the retentate of step (e);

g) Dilute the retentate of step (e) with water to prepare another diluted retentate;

h) Repeat steps (e) to (g) until the monosaccharide concentration of the retentate is less than about 20% based on dry weight to obtain Luo Han Guo flavor extract.
The method of claim 13, wherein the nanofiltration membrane has a molecular weight cut-off of 250 Daltons to 2,500 Daltons.
The method of claim 12 or 13, wherein after the step h), a step of drying the retentate is further included.
The method according to any one of claims 12 to 14, further comprising the step of adding a sweetener derived from Luo Han Guo to adjust the acid-sweet ratio of Luo Han Guo flavored beverage, wherein the ratio of sweet and sour is the weight percentage of total sugar and total acid in the beverage It shows that the sweet-sweet ratio is 18 ～ 28/1.
The method of claim 15, wherein the sweetener derived from Luo Han Guo comprises mogroside V and mogroside V-free mogroside non-glycoside juice.
A fermented Luo Han Guo flavored drink, comprising Luo Han Guo flavored extract, Luo Han Guo non-glycoside juice and Luo Han Guo raw vinegar, wherein the total sugar content of Luo Han Guo origin in the drink is about 3.0-9.0% (w / w), and the total acid content is 0.1 ~ 0.5% (w / w).
The beverage of claim 1, wherein the ratio of the total sugar to the total acid is 18-28: 1 (weight ratio).
The Luo Han Guo flavored drink according to claim 1 or 2, which is a Luo Han Guo flavored fruit vinegar.
A kind of mangosteen fruit wine or fruit vinegar is obtained by fermenting mangosteen extract from which mogroside is removed.