Selenium-rich additive for promoting wine flavor and preparation method thereof

Abstract

The invention relates to the technical field of wine brewing, in particular to a selenium-rich additive for promoting wine flavor and a preparation method thereof. The modified soluble selenate, the modified trehalose and the modified sodium carboxymethyl cellulose play a role through a synergistic effect, and the modified soluble selenate inhibits the activity of acetate dehydrogenase protein by damaging the expression of an acetate dehydrogenase gene of acetic bacteria in a fermentation process; the modified sodium carboxymethyl cellulose is further combined with the acetate dehydrogenase protein to reduce the expression of the activity of the acetate dehydrogenase protein, inhibit the continuous absorption of nutrient substances and prevent the activity of the acetate dehydrogenase protein from becoming strong, and the modified trehalose continuously enhances the stability of the compound of the modified sodium carboxymethyl cellulose and the acetate dehydrogenase protein; the combination time of the modified sodium carboxymethyl cellulose and the acetate dehydrogenase protein is prolonged, so that the activity of the acetate dehydrogenase is more effectively inhibited, the excessive fermentation of acetic bacteria is prevented, and the excessive acidification of wine liquid is avoided.

Description

Technical Field

[0001] The invention relates to the technical field of wine making, in particular to a selenium-rich additive for improving wine flavor and a preparation method thereof. Background Art

[0002] Selenium-rich additives are special raw materials used to enhance the flavor of wine. The raw materials usually contain selenium, antioxidants, organic matter and other beneficial substances. The purpose is to increase the selenium content in the wine, promote the formation of organic selenium in the wine, and thus enhance the flavor and taste of the wine. Selenium is mainly added in two forms: inorganic selenium and organic selenium. The absorption rate of selenium is affected by the form and source of selenium. The conventional inorganic selenium is not highly utilized by the human body, and excessive intake may also have adverse effects on the human body. Organic selenium has higher biological activity, better absorption and lower toxicity. By adding an organic selenium additive to the wine, the selenium content is effectively maintained and the stability is increased, so that the human body can better absorb selenium when eating. Selenium-rich additives can be used as a simple and effective additive to improve human health, and are widely used in various foods. In addition, adding selenium to wine can give it a unique flavor and improve the quality and taste of the wine. Summary of the invention

[0003] (1) Technical issues to be solved

[0004] The purpose of the present invention is to provide a selenium-rich additive for promoting wine flavor and a preparation method thereof, so as to increase the selenium content in wine, reduce the fermentation of acetic acid bacteria, avoid acidification of wine taste, and affect the flavor and taste. Acetate dehydrogenase is a key enzyme in acetic acid bacteria and participates in the fermentation process of acetic acid bacteria. Modified soluble selenate can generate active oxygen to damage the acetate dehydrogenase protein, promote the binding of modified sodium carboxymethyl cellulose with the acetate dehydrogenase protein, thereby inhibiting the activity of the acetate dehydrogenase center, and finally add modified trehalose to improve the stability of the modified sodium carboxymethyl cellulose binding with the acetate dehydrogenase protein, reduce the expression of the acetate dehydrogenase gene in acetic acid bacteria, thereby inhibiting the activity of acetate dehydrogenase and preventing the excessive fermentation of acetic acid bacteria.

[0005] (2) Technical solution

[0006] To achieve the above object, on the one hand, the present invention provides a method for preparing a selenium-rich additive for improving the flavor of wine, comprising the following steps:

[0007] S11. Add deionized water to the first container, add modified soluble selenate and selenoprotein at room temperature while stirring, the weight ratio of modified soluble selenate to selenoprotein is 1:(2-4), until completely dissolved, then add citric acid, the weight ratio of citric acid to modified soluble selenate is 0.5:(2-4), and adjust the pH to 6-7;

[0008] S12. Add 50 to 80 ml of deionized water to the second container, add resveratrol and vitamin E in a weight ratio of 1:(2 to 4) under stirring until dissolved, and then add antioxidant enzyme, selenoprotein and antioxidant enzyme in a weight ratio of 1:(3 to 6);

[0009] S13. Mix the mixed solution in the first container and the mixed solution in the second container into a third container, and then add modified sodium carboxymethyl cellulose and modified trehalose, wherein the weight ratio of modified sodium carboxymethyl cellulose to modified trehalose is 1:(5-8), and mix them evenly to obtain a selenium-rich additive.

[0010] Furthermore, the preparation method of the selenium-rich additive also includes:

[0011] The selenoprotein exists in the form of selenoamino acid, and the weight ratio of the selenoprotein to the antioxidant enzyme is 1: (2-4).

[0012] Furthermore, the preparation method of the selenoprotein comprises:

[0013] S21. Add 1-1.2 mol / L phosphate solution into a container, prepare equal volumes of 65-70% whey protein and 50-70% soy protein solution, the addition ratio of phosphate solution, whey protein and soy protein is 1:(9-10):(9-10), stir for 1-2 hours to fully dissolve the whey protein and soy protein in the phosphate solution, and mark it as solution A;

[0014] S22. Dissolve 10 mg / L sodium selenide in water, stir for 20 to 30 min, then slowly add solution A to the sodium selenide solution, control the pH at 4.5 to 5.5, and mark it as solution B;

[0015] S23. Under heating conditions of 40°C, solution B was allowed to react and mix fully, with a stirring speed of 100 to 300 rpm and a heating temperature of 35 to 55°C, and stirred for 3 to 6 hours. The solution after the reaction was marked as solution C;

[0016] S24. At room temperature, 15-20% potassium permanganate was added to solution C and stirred for 1-2 hours. The solution after the reaction was marked as solution D.

[0017] S25. Centrifuge solution D at low temperature at a centrifugal speed of 8000-10000 rpm for 15-20 min, and dialyze the solution obtained after centrifugation to remove small molecule impurities and unbound selenium using a dialysis membrane, which is labeled as solution E;

[0018] S26. adding solution E to a 95-98% ethanol solution, and then purifying the solution by ion exchange chromatography, and further purifying the solution by a gel filtration column; after purification, affinity eluting the solution at least twice, and then removing the elution buffer by ultrafiltration;

[0019] S27. Finally, mass spectrometry analysis is performed, and the prepared selenoprotein is then stored in an environment of -80 to -90°C to obtain selenoprotein.

[0020] Furthermore, the preparation method of the soluble selenate comprises:

[0021] S31. Add 5-6 mg / L sodium selenite to 30-40 ml of deionized water and stir for 10-20 min, labeled as solution F;

[0022] S32. Take 0.1-0.5% aqueous solution of pectin and selenoprotein, add them to solution F, stir at 25-30°C for 10-15min to mix the two thoroughly, and mark them as solution G;

[0023] S33. The solution G is dialyzed and then freeze-dried at a temperature of -45 to -50°C to obtain modified soluble selenate after drying for 10 to 12 hours.

[0024] Furthermore, the preparation method of the modified sodium carboxymethyl cellulose comprises:

[0025] S41. Pre-treating the cellulose raw material by bleaching and then grinding it, and then mixing it with 1-2 mg / L sodium hydroxide, which is marked as solution H;

[0026] S42. The solution H is heated in a water bath at 150 to 165° C. to hydrolyze the cellulose to produce cellobiose or cellotriose;

[0027] S43. The cellulose treated with sodium hydroxide is mixed with sodium chloride and methanol, wherein the molar ratio of sodium chloride to methanol is 1:1, and heated to 70 to 80° C. under stirring for etherification reaction, and the stirring time is 6 to 10 hours. After the etherification reaction is completed, the pH is adjusted with hydrochloric acid to a range of pH 7.0 to 7.5, and the solution is labeled J;

[0028] S44. The solution J is washed to remove unreacted sodium hydroxide, sodium chloride and methanol, and then dried at 80 to 90 ° C for 12 to 15 hours. The dried solid is sodium carboxymethyl cellulose;

[0029] S45. Dissolve sodium carboxymethyl cellulose in 99.0-99.9% dimethylformamide at room temperature, slowly add sodium hydroxide solution to the dissolved sodium carboxymethyl cellulose solution, stir for 5-10 minutes, and mark the solution as K;

[0030] S46. Add a polymer powder prepared by acrylamide, styrene, etc. to solution K, and continue stirring for 6 to 8 hours at a temperature of 80 to 90° C. to graft the polymer powder onto the sodium carboxymethyl cellulose molecule through an etherification reaction;

[0031] S47. After the stirring is completed, the excess solvent is first poured out, and then an appropriate amount of ethyl acetate is added, and impurities in the supernatant are removed by standing and stratifying, and the process is repeated 3 to 5 times;

[0032] S48. Remove the residual ethyl acetate by rotary evaporation at 60-75° C. on a rotary evaporator to obtain modified sodium carboxymethyl cellulose.

[0033] Furthermore, the preparation method of the modified trehalose comprises:

[0034] S51. Select a strain of trehalose, control the pH to 4.5 to 6.0 in a culture dish at 20 to 30°C, and wait for it to ferment for 7 to 9 days;

[0035] S52. After the fermentation is completed, the fermented solid is taken out and placed in a centrifuge tube, 10 to 20 ml of water and 20 to 30 ml of ethanol are added, and the tube is allowed to stand for at least three minutes, and the tube is marked as solution L;

[0036] S53. The solution L was filtered and centrifuged at 5000 to 6000 rpm for 20 to 30 min to remove solid impurities and insoluble components in the extract;

[0037] S54. The supernatant after centrifugation was purified by ultrafiltration and then passed through a 10 μm nanomembrane to obtain trehalose;

[0038] S55. Put the trehalose into a nano-seaweed grinding and dispersing machine at a rotation speed of 1500-1600 rpm for 3-4 hours, and then take it out to obtain modified trehalose.

[0039] Furthermore, the specific surface area of ​​the trehalose is 50 to 110 m 2 / g.

[0040] Furthermore, the selenium-rich additive for improving wine flavor is prepared by the method for preparing the selenium-rich additive for improving wine flavor.

[0041] The mechanism of action of the above raw material components is as follows:

[0042] Selenoprotein is a selenium-containing protein composed of selenocysteine ​​and methionine. This special composition makes selenoprotein have many important physiological functions in the human body. As an essential trace element, selenium participates in the catalytic reactions of various enzymes in the human body, and selenoprotein is an important component of these enzymes, which is essential for maintaining the normal physiological functions of the human body. In addition, selenoprotein, as an organic form of selenium, may be more easily absorbed and utilized by yeast, thereby promoting its growth and metabolic activities. This helps to improve the fermentation efficiency and quality of wine.

[0043] As one of the main components of selenium-enriched additives, modified soluble selenate can be effectively absorbed and utilized by the human body due to its good solubility and bioavailability, thus meeting the human body's demand for selenium. The modified soluble selenate may have a toxic effect on acetic acid bacteria at high concentrations, which can reduce the enzyme activity of acetic acid bacteria, block the energy metabolism of acetic acid bacteria, affect the fermentation ability of acetic acid bacteria, and inhibit its growth and metabolism. At the same time, after being absorbed by yeast, the modified soluble selenate can be converted into selenoproteins or other selenium compounds with antioxidant activity, thereby exerting an antioxidant effect.

[0044] Citric acid, as an organic acid, has good solubility and complexing ability. In selenium-enriched additives, an appropriate amount of citric acid can maintain a suitable pH environment, which is conducive to the stable existence of selenium and the absorption of selenium by the intestine. Adding an appropriate amount of citric acid can improve its taste and flavor, making selenium-enriched products more acceptable to consumers.

[0045] Resveratrol is a polyphenol compound with multiple biological activities. It is widely found in plants such as grape skin, blueberry, and peanut. It is a natural antioxidant with anti-inflammatory, anti-cancer, and anti-aging biological activities. Resveratrol can lower blood lipids, inhibit platelet aggregation, dilate blood vessels, etc., thereby protecting the health of the cardiovascular system. Among selenium-rich additives, these effects of resveratrol help prevent the occurrence of cardiovascular diseases.

[0046] Vitamin E is a fat-soluble vitamin with strong antioxidant capacity. It can work synergistically with glutathione peroxidase to remove free radicals in the body. In the winemaking process, yeast converts sugars in the raw materials into alcohol and carbon dioxide, and the presence of vitamin E can promote this process, thereby increasing the yield and quality of alcohol, helping to protect the wine from oxidative damage and maintain its flavor and quality.

[0047] There are two types of antioxidant enzymes, SOD and CAT, which have many functions, including directly removing free radicals, promoting the production of antioxidant substances, regulating signal pathways, enhancing immune function, and synergizing with other ingredients. These mechanisms work together on the body to enhance the body's antioxidant capacity and protect cells from oxidative stress damage. Adding these two enzymes can significantly enhance the antioxidant capacity of the product.

[0048] Modified sodium carboxymethyl cellulose is a common food additive and industrial raw material. Modified sodium carboxymethyl cellulose can form graft copolymers, and chemical bonds are formed between sodium carboxymethyl cellulose molecules to increase its network structure, thereby improving its stability. Modified sodium carboxymethyl cellulose binds to the active center of the acetate dehydrogenase protein in acetic acid bacteria, blocking its activity, thereby reducing the activity of acetate dehydrogenase. At the same time, it can affect the stability of acetate dehydrogenase, making it more susceptible to degradation or inactivation. In addition, modified sodium carboxymethyl cellulose can regulate the fermentation environment in the wine, forming a protective layer in the wine, hindering the direct contact between acetic acid bacteria and the wine, thereby inhibiting its growth.

[0049] Modified trehalose can interact with acetic acid bacteria through various mechanisms, thereby effectively regulating its growth and fermentation process. Modified trehalose can penetrate between the cell wall and membrane protein of acetic acid bacteria, forming a protective layer or interference layer, which not only limits the absorption of nutrients by acetic acid bacteria, but also affects its metabolic pathways, especially inhibiting the activity of enzymes related to acetic acid fermentation.

[0050] In the selenium-enriched additives, modified soluble selenate, modified trehalose and modified sodium carboxymethyl cellulose play a synergistic role, effectively preventing the expression of acetate dehydrogenase protein in acetic acid bacteria and inhibiting the activity of acetate dehydrogenase, thereby affecting the fermentation and growth of acetic acid bacteria, while protecting the growth environment of yeast, effectively optimizing the fermentation environment in the wine, and significantly increasing the selenium content in the wine. During the fermentation of wine, the soluble selenate solution can generate active oxygen through chemical reactions. As a strong oxidant, active oxygen can attack and damage the active site of acetate dehydrogenase protein, resulting in damage to the function of acetate dehydrogenase protein and reducing the activity of acetate dehydrogenase. At the same time, modified sodium carboxymethyl cellulose is connected to the damaged acetate dehydrogenase protein through chemical bonds. This combination inhibits the activity of acetate dehydrogenase and further blocks the fermentation ability of acetic acid bacteria. In order to enhance the stability of the binding between modified sodium carboxymethyl cellulose and acetic dehydrogenase protein, modified trehalose can significantly improve the stability of the complex of modified sodium carboxymethyl cellulose and acetic dehydrogenase protein, prolong the binding time of modified sodium carboxymethyl cellulose and acetic dehydrogenase protein, thereby more effectively inhibiting the activity of acetic dehydrogenase, affecting the protein required for acetic acid bacteria fermentation, and reducing the production of acetic acid in wine. Modified soluble selenate, modified trehalose and modified sodium carboxymethyl cellulose jointly prevent the gene expression of acetic acid dehydrogenase protein, inhibit the activity of acetic acid dehydrogenase, reduce the fermentation ability of acetic acid bacteria through a mutual activation mechanism, and at the same time reduce the content of free radicals, and promote the antioxidant effect between microorganisms, forming a positive feedback loop, promoting the formation of wine flavor and increasing the content of selenium in wine. They affect the fermentation of acetic acid bacteria, reduce synthetic energy, synergistically remove free radicals, inhibit oxidase activity and repair damaged DNA, providing comprehensive protection for cells. The synergistic effect between these components also enhances the stability of each other, ensuring the continuous and effective operation of the antioxidant defense system in the organism. Effectively remove free radicals in the body and reduce the damage of oxidative stress to cells. Therefore, modified soluble selenate, modified trehalose and modified sodium carboxymethyl cellulose jointly inhibit the gene expression of acetate dehydrogenase protein, prevent the production of acetate dehydrogenase in acetic acid bacteria, affect the fermentation of acetic acid bacteria, maintain a good fermentation environment, reduce the production of free radicals, ensure that the wine has a relatively rich selenium content, and maintain a good fermentation environment, so that the wine has a richer flavor and nutritional value. In addition, selenoprotein has significant antioxidant capacity, can remove free radicals in the body, and reduce the damage of oxidative stress to cells. As the main source of selenium, selenoprotein and modified soluble selenate complement each other, improve the absorption rate and utilization rate of selenium, and make the effect of selenium supplementation more significant. Citric acid, as an important food additive, can significantly increase the solubility of inorganic selenium such as selenate, so that it can be better dispersed in the solution, thereby enhancing the stability and absorbability of selenium.As a natural antioxidant, resveratrol and antioxidant enzymes together form a multi-level antioxidant system, which increases the selenium-rich conditions in the wine and effectively prevents the excessive generation of free radicals. Finally, vitamin E can promote the fermentation of yeast and protect other cell agents and microorganisms in the fermentation environment from damage by free radicals. It improves the taste and flavor of the wine, so the wine tastes sweeter and mellower after adding vitamin E.

[0051] (3) Beneficial effects

[0052] Compared with the prior art, the present invention has the following beneficial effects:

[0053] 1. Modified soluble selenate has a high solubility in the gastrointestinal tract, so selenium is easily absorbed by the human body. Modified soluble selenate and selenoprotein can produce a synergistic effect, which not only achieves the complementarity of selenium sources, but also significantly improves the absorption efficiency and utilization rate of selenium in the body, making the effect of selenium supplementation more significant.

[0054] 2. Modified trehalose can maintain the stability of various components in wine, such as ethanol, organic acids, polyphenols, etc., while improving the stability of the complex of modified sodium carboxymethyl cellulose and acetate dehydrogenase protein, and prolonging the binding time of modified sodium carboxymethyl cellulose and acetate dehydrogenase protein. BRIEF DESCRIPTION OF THE DRAWINGS

[0055] Figure 1 This is a SEM image of modified trehalose in Example 1 of the present invention;

[0056] Figure 2 This is the XRD diagram of the modified sodium carboxymethyl cellulose of Example 1 of the present invention. DETAILED DESCRIPTION

[0057] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0058] The experimental equipment and preparations of the embodiments described below are as follows: electronic balance (Sartorius, Germany), pH meter (Tangshan Fengrui), electric constant temperature water bath (Jiangsu Kedao), electric blast drying oven (Suzhou Geruida), vacuum drying oven (Shanghai Jiecheng), high-speed pulverizer (Shandong Tianfang Machinery), magnetic stirrer (Shanghai Meiyingpu), high-speed centrifuge (Guangzhou Jidi), freeze dryer, SA-50 high performance liquid chromatography-atomic fluorescence spectrometer, X-ray diffractometer, scanning electron microscope (Germany Zeiss), low temperature constant temperature incubator (Shanghai Hetian); chemicals and reagents were purchased from Sigma-Aldrich.

[0059] Example 1

[0060] This embodiment discloses a selenium-rich additive for improving the flavor of wine, comprising the following ingredients: 10 parts of modified soluble selenate, 10 parts of citric acid, 5 parts of resveratrol, 5 parts of antioxidant enzyme, 5 parts of vitamin E, 1 part of modified sodium carboxymethyl cellulose, 5 parts of modified trehalose, and 30 parts of deionized water. The weight ratio of the selenoprotein to the antioxidant enzyme is 1:2.

[0061] Among the selenium-rich additives, modified soluble selenate, modified trehalose and modified sodium carboxymethyl cellulose play a synergistic role. During the fermentation process of winemaking, modified soluble selenate can destroy the activity of acetic acid bacteria during the fermentation of acetic acid dehydrogenase protein and reduce its gene expression ability. Modified sodium carboxymethyl cellulose combines with acetic acid dehydrogenase protein. This combination inhibits the residual activity of acetic acid dehydrogenase and further blocks the fermentation ability of acetic acid bacteria. In order to enhance the stability of the combination of modified sodium carboxymethyl cellulose and acetic acid dehydrogenase protein, modified trehalose can significantly improve the stability of the complex of modified sodium carboxymethyl cellulose and acetic acid dehydrogenase protein, prolong the binding time of modified sodium carboxymethyl cellulose and acetic acid dehydrogenase protein, thereby more effectively inhibiting the activity of acetic acid dehydrogenase, affecting the protein required for acetic acid bacteria fermentation, and reducing the production of acetic acid in wine. Figure 1 This is the SEM image of modified trehalose. It can be seen that the modified trehalose presents a uniform spherical structure. Figure 2The XRD diagram of modified sodium carboxymethyl cellulose confirms that the crystal structure of sodium carboxymethyl cellulose has not changed significantly after modification. In addition, supplementation of selenoprotein and modified trehalose can also activate antioxidant enzyme systems, such as glutathione peroxidase, so that yeast cells can more effectively resist the oxidative stress caused by excessive selenium content during fermentation, maintain their metabolic activity and stability, and thus ensure the smooth progress of the fermentation process. Inhibiting acetic acid fermentation is the cornerstone of this synergistic effect. Acetic acid bacteria can oxidize alcohol to produce acetic acid. The production of acetic acid will produce a special sour and bitter taste in the wine, seriously affecting the taste and quality of the wine, and even making the wine undrinkable. Acetic acid bacteria can not only metabolize alcohol, but also metabolize other components in the wine, such as carbohydrates, glycerol, sugar alcohols and inorganic acids. When the wine is exposed to the air, acetic acid bacteria will quickly form a biofilm on the surface of the wine and gradually sink into the wine, resulting in a significant increase in the content of volatile acids such as acetic acid in the wine. This biofilm will cause the microorganisms in the wine to be unable to obtain energy, affect the fermentation environment, and cause the taste and flavor of the wine to deteriorate. At the same time, in this process, free radicals will be generated and can attack and destroy phenolic compounds (such as tannins, anthocyanins), amino acids, vitamins and other ingredients in the wine that are beneficial to human health. Therefore, the combined synergistic effect of modified soluble selenate, modified trehalose and modified sodium carboxymethyl cellulose, like a solid shield, directly inhibits the key enzyme activity in acetic acid bacteria, optimizes the fermentation conditions in the wine, improves the fermentation environment in the wine, and effectively reduces the damage of free radicals to these ingredients, protecting the natural nutritional value and unique flavor of the wine. And it can reduce the generation of undesirable flavor substances, such as volatile compounds such as aldehydes and ketones, and promote the accumulation of beneficial flavor substances, such as esters and alcohols. These bioactive substances act together on the wine, making the final wine flavor more pure, complex and lasting. In addition, modified sodium carboxymethyl cellulose and vitamin E also help stabilize the pigments and aroma components in the wine, extending the shelf life of the wine. By inhibiting the fermentation of acetic acid bacteria, adjusting the antioxidant capacity of yeast, and adjusting the changes in temperature, humidity, oxygen content, etc., the winemaking efficiency and output can be improved and the production cost can be reduced. This flexibility and adaptability makes the winemaking process more refined and controllable, providing a strong guarantee for the production of high-quality wines.

[0062] The preparation method of the claimed selenoprotein comprises:

[0063] S21. Add 1-1.2 mol / L phosphate solution into a container, prepare equal volumes of 65-70% whey protein and 50-70% soy protein solution, the addition ratio of phosphate solution, whey protein and soy protein is 1:(9-10):(9-10), stir for 1-2 hours to fully dissolve the whey protein and soy protein in the phosphate solution, and mark it as solution A;

[0064] S22. Dissolve 10 mg / L sodium selenide in water, stir for 20 to 30 min, then slowly add solution A to the sodium selenide solution, control the pH at 4.5 to 5.5, and mark it as solution B;

[0065] S23. Under heating conditions of 40°C, solution B was allowed to react and mix fully, with a stirring speed of 100 to 300 rpm and a heating temperature of 35 to 55°C, and stirred for 3 to 6 hours. The solution after the reaction was marked as solution C;

[0066] S24. At room temperature, 15-20% potassium permanganate was added to solution C and stirred for 1-2 hours. The solution after the reaction was marked as solution D.

[0067] S25. Centrifuge solution D at low temperature at a centrifugal speed of 8000-10000 rpm for 15-20 min, and dialyze the solution obtained after centrifugation to remove small molecule impurities and unbound selenium using a dialysis membrane, which is labeled as solution E;

[0068] S26. adding solution E to a 95-98% ethanol solution, and then purifying the solution by ion exchange chromatography, and further purifying the solution by a gel filtration column; after purification, affinity eluting the solution at least twice, and then removing the elution buffer by ultrafiltration;

[0069] S27. Finally, mass spectrometry analysis is performed, and the prepared selenoprotein is then stored in an environment of -80 to -90°C to obtain selenoprotein.

[0070] The preparation method of the modified soluble selenate comprises:

[0071] S31. Add 5-6 mg / L sodium selenite to 30-40 ml of deionized water and stir for 10-20 min, labeled as solution F;

[0072] S32. Take 0.1-0.5% aqueous solution of pectin and selenoprotein, add them to solution F, stir at 25-30°C for 10-15min to mix the two thoroughly, and mark them as solution G;

[0073] S33. The solution G is dialyzed and then freeze-dried at a temperature of -45 to -50°C to obtain modified soluble selenate after drying for 10 to 12 hours.

[0074] The preparation method of the modified sodium carboxymethyl cellulose comprises:

[0075] S41. Pre-treating the cellulose raw material by bleaching and then grinding it, and then mixing it with 1-2 mg / L sodium hydroxide, which is marked as solution H;

[0076] S42. The solution H is heated in a water bath at 150 to 165° C. to hydrolyze the cellulose to produce cellobiose or cellotriose;

[0077] S43. The cellulose treated with sodium hydroxide is mixed with sodium chloride and methanol, wherein the molar ratio of sodium chloride to methanol is 1:1, and heated to 70 to 80° C. under stirring for etherification reaction, and the stirring time is 6 to 10 hours. After the etherification reaction is completed, the pH is adjusted with hydrochloric acid to a range of pH 7.0 to 7.5, and the solution is labeled J;

[0078] S44. The solution J is washed to remove unreacted sodium hydroxide, sodium chloride and methanol, and then dried at 80 to 90 ° C for 12 to 15 hours. The dried solid is sodium carboxymethyl cellulose;

[0079] S45. Dissolve sodium carboxymethyl cellulose in 99.0-99.9% dimethylformamide at room temperature, slowly add sodium hydroxide solution to the dissolved sodium carboxymethyl cellulose solution, stir for 5-10 minutes, and mark the solution as K;

[0080] S46. Add a polymer powder prepared by acrylamide, styrene, etc. to solution K, and continue stirring for 6 to 8 hours at a temperature of 80 to 90° C. to graft the polymer powder onto the sodium carboxymethyl cellulose molecule through an etherification reaction;

[0081] S47. After the stirring is completed, the excess solvent is first poured out, and then an appropriate amount of ethyl acetate is added, and impurities in the supernatant are removed by standing and stratifying, and the process is repeated 3 to 5 times;

[0082] S48. Remove the residual ethyl acetate by rotary evaporation at 60-75° C. on a rotary evaporator to obtain modified sodium carboxymethyl cellulose.

[0083] The preparation method of the modified trehalose comprises:

[0084] S51. Select a strain of trehalose, control the pH to 4.5 to 6.0 in a culture dish at 20 to 30°C, and wait for it to ferment for 7 to 9 days;

[0085] S52. After the fermentation is completed, the fermented solid is taken out and placed in a centrifuge tube, 10 to 20 ml of water and 20 to 30 ml of ethanol are added, and the tube is allowed to stand for at least three minutes, and the tube is marked as solution L;

[0086] S53. The solution L was filtered and centrifuged at 5000 to 6000 rpm for 20 to 30 min to remove solid impurities and insoluble components in the extract;

[0087] S54. The supernatant after centrifugation was purified by ultrafiltration and then passed through a 10 μm nanomembrane to obtain trehalose;

[0088] S55. Put the trehalose into a nano-seaweed grinding and dispersing machine at a rotation speed of 1500-1600 rpm for 3-4 hours, and then take it out to obtain modified trehalose.

[0089] Example 2

[0090] This embodiment discloses a selenium-rich additive for improving the flavor of wine, comprising the following ingredients: 20 parts of modified soluble selenate, 10 parts of citric acid, 10 parts of resveratrol, 8 parts of antioxidant enzyme, 10 parts of vitamin E, 15 parts of modified sodium carboxymethyl cellulose, 10 parts of modified trehalose, and 30 parts of deionized water. The weight ratio of the selenoprotein to the antioxidant enzyme is 1:3.

[0091] The preparation methods of the selenoprotein and modified sodium carboxymethyl cellulose in this embodiment are consistent with those in Example 1. The preparation method of a selenium-enriched additive in this embodiment is consistent with that in Example 1.

[0092] Example 3

[0093] This embodiment discloses a selenium-rich additive for improving the flavor of wine, comprising the following ingredients: 30 parts of modified soluble selenate, 15 parts of citric acid, 15 parts of resveratrol, 15 parts of vitamin E, 10 parts of antioxidant enzyme, 20 parts of modified sodium carboxymethyl cellulose, 15 parts of modified trehalose, and 40 parts of deionized water. The weight ratio of the selenoprotein to the antioxidant enzyme is 1:4.

[0094] The preparation methods of the selenoprotein, modified soluble selenate, modified sodium carboxymethyl cellulose and modified trehalose in this example are the same as those in Example 1. The preparation method of a selenium-enriched additive in this example is the same as that in Example 1.

[0095] Control group 1

[0096] The difference between this embodiment and embodiment 1 is that the modified soluble selenate is not contained.

[0097] This embodiment discloses a selenium-rich additive for improving the flavor of wine, comprising the following ingredients: 10 parts of citric acid, 10 parts of resveratrol, 10 parts of vitamin E, 12 parts of antioxidant enzyme, 15 parts of modified sodium carboxymethyl cellulose, 10 parts of modified trehalose, and 40 parts of deionized water. The weight ratio of selenoprotein to antioxidant enzyme is 1:3.

[0098] The preparation methods of the selenoprotein, modified sodium carboxymethyl cellulose and modified trehalose in this example are the same as those in Example 1. The preparation method of a selenium-enriched additive in this example is the same as that in Example 1.

[0099] Control group 2

[0100] The difference between this embodiment and embodiment 1 is that modified trehalose is not contained.

[0101] This embodiment discloses a selenium-rich additive for improving the flavor of wine, comprising the following ingredients: 20 parts of modified soluble selenate, 10 parts of citric acid, 10 parts of resveratrol, 10 parts of vitamin E, 12 parts of antioxidant enzyme, 15 parts of modified sodium carboxymethyl cellulose, and 40 parts of deionized water. The weight ratio of selenoprotein to antioxidant enzyme is 1:3.

[0102] The preparation methods of the selenoprotein, modified soluble selenate and modified sodium carboxymethyl cellulose in this embodiment are the same as those in Example 1. The preparation method of a selenium-enriched additive in this embodiment is the same as that in Example 1.

[0103] Control group 3

[0104] The difference between this embodiment and embodiment 1 is that modified sodium carboxymethyl cellulose is not contained.

[0105] This embodiment discloses a selenium-rich additive for improving the flavor of wine, comprising the following ingredients: 20 parts of modified soluble selenate, 10 parts of citric acid, 10 parts of resveratrol, 10 parts of vitamin E, 12 parts of antioxidant enzyme, 10 parts of modified trehalose, and 40 parts of deionized water. The weight ratio of selenoprotein to antioxidant enzyme is 1:3.

[0106] The preparation methods of the selenoprotein, modified soluble selenate and modified trehalose in this embodiment are consistent with those in Example 1. The preparation method of a selenium-enriched additive in this embodiment is consistent with that in Example 1.

[0107] Effect evaluation: Selenium-enriched wine experiment: Use 55.0%vol distilled wine to blend the wine flavor, mix the selenium-enriched additive with clean water at a mass ratio of 1: (10-20) and stir evenly. Use fresh and high-quality selenium-enriched glutinous rice as the raw material, sieve to remove broken rice and impurities, rinse with water until there is no obvious turbid water, and divide the selenium-enriched glutinous rice evenly into 8 groups. Add water to the washed raw materials and cook until the rice grains are cooked, with a water content of about 50%-70%. Stir the steamed rice balls and spread them out to dry, control the temperature to 36℃-37℃, then add a certain amount of Xiaoqu powder and selenium-enriched yeast, stir evenly and put them into triangular bottles in time, each bottle is about 500g, the thickness of the rice is about 10cm-13cm, dig a hole in the center to ensure sufficient air culture and saccharification, plug the bottle mouth with cotton for 3 days, add selenium-enriched additives to each portion of wine and mix evenly, and control the aging time between 10 and 20 days to ensure that the selenium-enriched additives can fully play their role. The finished product is subjected to sensory testing and organic selenium content testing. Among them, the 3,31-aminobenzidine colorimetric method is used to determine the selenium content in the wine. Organic selenium content = total selenium content - residual inorganic selenium content, organic selenium conversion rate (%) = (organic selenium content / total selenium content) * 100.

[0108] The results of sensory testing and organic selenium content testing of the eight experimental groups are shown in Tables 1 and 2. The experimental group has a significantly fuller body, a strong sense of layering, a mellow and coordinated aftertaste, and a strong and lasting aroma, which is significantly better than the control group and the blank group. Although the control group maintains the purity and unique style of the wine, the aroma is not strong enough, reflecting the importance of the three ingredients, which are indispensable. The blank group shows the smoothness and coordination of the basic wine sample, but the aroma is the most bland. In summary, selenium-enriched wine not only achieves a significant improvement in flavor, but also shows unique advantages in aroma, providing strong support for the innovation of wine products. As shown in Table 2, the organic selenium content and organic selenium conversion rate of the experimental groups (experimental group 1, experimental group 2, and experimental group 3) are significantly higher than those of the control group and the blank group. This shows that adding or promoting selenium-enriched ingredients during the winemaking process can significantly increase the organic selenium content and its conversion rate in the final wine. The organic selenium content and conversion rate of the control groups (control group 1, control group 2, and control group 3) were relatively low, but still maintained a certain level, indicating that the lack of the main ingredients will lead to a decrease in selenium content, indicating that the coordination of the three is more important. Although a certain amount of organic selenium may still be naturally generated during the winemaking process, its efficiency and content are not as good as those of the experimental group. The organic selenium content and conversion rate of the blank groups (blank group 1 and blank group 2) were the lowest, almost close to the natural background level. This proves that the generation of organic selenium in the winemaking process is very limited without the additional addition or promotion of selenium-rich ingredients. Finally, it can be found that the addition of specific selenium-rich additives can significantly increase the content and conversion rate of organic selenium in the winemaking process. This improvement not only helps to increase the nutritional value and health function of the wine, but also may provide a scientific basis for the research and development and promotion of selenium-rich wine. At the same time, the data of the control group and the blank group also reflect the limitations of organic selenium generation in the winemaking process under natural conditions, further highlighting the necessity and superiority of selenium-enriched treatment. Selenium is a trace element in the human body, and the intake needs to be strictly controlled. The normal human body contains 14 to 21 mg of selenium. Adults need to take in 50 to 300 μg of selenium from the outside every day. The Chinese Nutrition Society recommends that the average human body should take 50 μg of selenium per day. The selenium content of the wine in this experiment is designed to be 18.7 μg / mL, so that even if each person drinks more than 1000 mL of alcohol per day, the selenium intake of the human body will not exceed the standard.

[0109] Table 1 Effects of different selenium-enriched additives on wine flavor

[0110]

[0111] Table 2 Organic selenium content of different selenium-enriched additives

[0112] serial number Organic selenium content (μg / mL) Organic selenium conversion rate (%) Experimental Group 1 17.9 82.0 Experimental Group 2 18.7 83.2 Experimental Group 3 18.1 82.3 Control group 1 15.2 66.2 Control group 2 14.3 66.0 Control group 3 16.1 63.1 Blank group 1 3.1 35.1 Blank Group 2 3.5 36.9

[0113] Finally, it should be noted that: Although the present invention has been described in detail with reference to the aforementioned embodiments, it is still possible for those skilled in the art to modify the technical solutions described in the aforementioned embodiments, or to make equivalent substitutions for some of the technical features therein. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A method for preparing a selenium-rich additive for improving wine flavor, characterized in that: The following steps are involved: S11. Add deionized water to the first container, add modified soluble selenate and selenoprotein at room temperature while stirring, the weight ratio of modified soluble selenate to selenoprotein is 1:(2-4), until completely dissolved, then add citric acid, the weight ratio of citric acid to modified soluble selenate is 0.5:(2-4), and adjust the pH to 6-7; S12. Add 50 to 80 ml of deionized water to the second container, add resveratrol and vitamin E in a weight ratio of 1:(2 to 4) under stirring until dissolved, and then add antioxidant enzyme, selenoprotein and antioxidant enzyme in a weight ratio of 1:(3 to 6); S13. Mix the mixed solution in the first container and the mixed solution in the second container into a third container, and then add modified sodium carboxymethyl cellulose and modified trehalose, wherein the weight ratio of modified sodium carboxymethyl cellulose to modified trehalose is 1:(5-8), and mix them evenly to obtain a selenium-rich additive.

2. The method for preparing a selenium-rich additive for improving wine flavor according to claim 1, characterized in that: The selenoprotein exists in the form of selenoamino acid; the weight ratio of the selenoprotein to the antioxidant enzyme is 1:(2-4).

3. The method for preparing a selenium-rich additive for improving wine flavor according to claim 2, characterized in that: The preparation method of the selenoprotein comprises: S21. Add 1-1.2 mol / L phosphate solution into a container, prepare equal volumes of 65-70% whey protein and 50-70% soy protein solution, the addition ratio of phosphate solution, whey protein and soy protein is 1:(9-10):(9-10), stir for 1-2 hours to fully dissolve the whey protein and soy protein in the phosphate solution, and mark it as solution A; S22. Dissolve 10 mg / L sodium selenide in water, stir for 20 to 30 min, then slowly add solution A to the sodium selenide solution, control the pH at 4.5 to 5.5, and mark it as solution B; S23. Under heating conditions of 40°C, solution B was allowed to react and mix fully, with a stirring speed of 100 to 300 rpm and a heating temperature of 35 to 55°C, and stirred for 3 to 6 hours. The solution after the reaction was marked as solution C; S24. At room temperature, 15-20% potassium permanganate was added to solution C and stirred for 1-2 hours. The solution after the reaction was marked as solution D. S25. Centrifuge solution D at low temperature at a centrifugal speed of 8000-10000 rpm for 15-20 min, and dialyze the solution obtained after centrifugation to remove small molecule impurities and unbound selenium using a dialysis membrane, which is labeled as solution E; S26. adding solution E to a 95-98% ethanol solution, and then purifying the solution by ion exchange chromatography, and further purifying the solution by a gel filtration column; after purification, affinity eluting the solution at least twice, and then removing the elution buffer by ultrafiltration; S27. Finally, mass spectrometry analysis is performed, and the prepared selenoprotein is then stored in an environment of -80 to -90°C to obtain selenoprotein.

4. The method for preparing a selenium-rich additive for improving wine flavor according to claim 1, characterized in that: The preparation method of the modified soluble selenate comprises: S31. Add 5-6 mg / L sodium selenite to 30-40 ml of deionized water and stir for 10-20 min, labeled as solution F; S32. Take 0.1-0.5% aqueous solution of pectin and selenoprotein, add them to solution F, stir at 25-30°C for 10-15min to mix the two thoroughly, and mark them as solution G; S33. The solution G is dialyzed and then freeze-dried at a temperature of -45 to -50°C to obtain modified soluble selenate after drying for 10 to 12 hours.

5. The method for preparing a selenium-rich additive for improving wine flavor according to claim 1, characterized in that: The ratio of the citric acid to the modified soluble selenate is 0.5:(2-4).

6. The method for preparing a selenium-rich additive for improving wine flavor according to claim 1, characterized in that: The ratio of the resveratrol to the modified soluble selenate is (5-15):

1.

7. The method for preparing a selenium-rich additive for improving wine flavor according to claim 1, characterized in that: The ratio of the vitamin E to the modified soluble selenate is (5-15):

1.

8. The method for preparing a selenium-rich additive for improving wine flavor according to claim 1, characterized in that: The preparation method of the modified sodium carboxymethyl cellulose comprises: S41. Pre-treating the cellulose raw material by bleaching and then grinding it, and then mixing it with 1-2 mg / L sodium hydroxide, which is marked as solution H; S42. The solution H is heated in a water bath at 150 to 165° C. to hydrolyze the cellulose to produce cellobiose or cellotriose; S43. The cellulose treated with sodium hydroxide is mixed with sodium chloride and methanol, wherein the molar ratio of sodium chloride to methanol is 1:1, and heated to 70 to 80° C. under stirring for etherification reaction, and the stirring time is 6 to 10 hours. After the etherification reaction is completed, the pH is adjusted with hydrochloric acid to a range of pH 7.0 to 7.5, and the solution is labeled J; S44. The solution J is washed to remove unreacted sodium hydroxide, sodium chloride and methanol, and then dried at 80 to 90 ° C for 12 to 15 hours. The dried solid is sodium carboxymethyl cellulose; S45. Dissolve sodium carboxymethyl cellulose in 99.0-99.9% dimethylformamide at room temperature, slowly add sodium hydroxide solution to the dissolved sodium carboxymethyl cellulose solution, stir for 5-10 minutes, and mark the solution as K; S46. Add a polymer powder prepared by acrylamide, styrene, etc. to solution K, and continue stirring for 6 to 8 hours at a temperature of 80 to 90° C. to graft the polymer powder onto the sodium carboxymethyl cellulose molecule through an etherification reaction; S47. After the stirring is completed, the excess solvent is first poured out, and then an appropriate amount of ethyl acetate is added, and impurities in the supernatant are removed by standing and stratifying, and the process is repeated 3 to 5 times; S48. Remove the residual ethyl acetate by rotary evaporation at 60-75° C. on a rotary evaporator to obtain modified sodium carboxymethyl cellulose.

9. The method for preparing a selenium-rich additive for improving wine flavor according to claim 1, characterized in that: The preparation method of the modified trehalose comprises: S51. Select a strain of trehalose, control the pH to 4.5 to 6.0 in a culture dish at 20 to 30°C, and wait for it to ferment for 7 to 9 days; S52. After the fermentation is completed, the fermented solid is taken out and placed in a centrifuge tube, 10 to 20 ml of water and 20 to 30 ml of ethanol are added, and the tube is allowed to stand for at least three minutes, and the tube is marked as solution L; S53. The solution L was filtered and centrifuged at 5000 to 6000 rpm for 20 to 30 min to remove solid impurities and insoluble components in the extract; S54. The supernatant after centrifugation was purified by ultrafiltration and then passed through a 10 μm nanomembrane to obtain trehalose; S55. The trehalose is placed in a nano-seaweed grinding and dispersing machine at a speed of 1500 to 1600 rpm for 3 to 4 hours of grinding, and then taken out to obtain modified trehalose; the specific surface area of ​​the trehalose is 50 to 110 m 2 / g.

10. A selenium-rich additive for improving the flavor of wine, characterized in that: The selenium-rich additive is obtained by the preparation method of a selenium-rich additive for improving wine flavor as described in any one of claims 1 to 9.

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