A method for preparing health-promoting red yeast rice wine

Abstract

This invention relates to a method for preparing health-promoting red yeast rice wine, comprising: (1) preparing a liquid fermentation broth of *Pseudomonas ulmoides*; (2) preparing a starter culture, wherein the starter culture is obtained by inoculating *Rhizopus* into a starter culture made from glutinous rice, soybeans, banana peel, vitamin B2, vitamin C, and sterile water; (3) steaming and cooling glutinous rice and adding *Pseudomonas ulmoides* fermentation broth, *Monascus purpureus* expanded culture broth, decanoic acid, and *Lactobacillus plantarum* expanded culture broth, and saccharifying and fermenting to obtain a first fermented product; (4) adding starter culture to the first fermented product, stirring evenly, and continuing saccharification to obtain a second fermented product; (5) pumping the second fermented product into a fermentation tank for alcoholic fermentation; and (6) after the fermentation in step (4) is completed, clarifying, filtering, sterilizing, and bottling the finished product. By brewing rice wine using this method, vitamins and various amino acids from *Pseudomonas ulmoides* can be extracted into the rice wine, and various specific amino acids can reduce the yield of citrinin in red yeast rice under certain fermentation conditions, resulting in high-quality red yeast rice wine.

Description

Technical Field

[0001] This invention relates to the field of rice wine brewing technology, and more specifically, to a method for preparing a health-promoting red yeast rice wine. Background Technology

[0002] Rice wine is a low-alcohol fermented beverage. Modern nutrition science believes that the nutritional value of rice wine lies in the sugars, organic acids, proteins, amino acids, vitamins, and minerals contained in the liquid, excluding water and ethanol. Sweet rice wine is a traditional fermented food in my country. In addition to alcohol and esters, sweet rice wine also contains elements such as calcium, phosphorus, and iron, as well as various nutrients such as sugars, dextrin, and glycerol. Some sweet rice wines also contain trace elements such as zinc, copper, manganese, and selenium.

[0003] Monascus purpureus is a genus of small, filamentous saprophytic fungi belonging to the Ascomycota, Class Undariaceae, Order Monasceales, Family Monasceae. It is found in trees, soil, and sediments. It grows well on malt extract agar, with colonies initially white, turning pale pink, purple, or grayish-black as they mature, often resulting in a red color. Red yeast rice can be used as a coloring and flavoring agent in brewing, vinegar making, and fermented bean curd, and it is also used in traditional Chinese medicine. In recent years, it has been discovered that the active substances produced by red yeast rice have cholesterol-lowering, blood pressure-lowering, and disease-preventing and therapeutic functions. During its growth, Monascus purpureus produces highly active saccharifying enzymes and proteases. Due to the medicinal and edible properties of red yeast rice, its use in brewing fermentation allows various active substances from red yeast rice metabolism to dissolve into the wine, enhancing its nutritional and health benefits.

[0004] However, the discovery of the fungal toxin citrinin has challenged the safety of functional red yeast rice. Citrinin is a secondary metabolite produced during the growth of Monascus purpureus, exhibiting nephrotoxicity, teratogenicity, carcinogenicity, and gene mutation induction. The kidney is the target organ for citrinin; ingestion causes nephrotoxicity in various experimental animals, characterized by enlarged kidneys and ultimately kidney failure. Studies have also found that citrinin has teratogenic effects, impairs liver metabolism, has carcinogenic and mutagenic effects, and may have inhibitory effects on the central nervous system. The presence of citrinin has become a bottleneck for the application and export of functional red yeast rice in my country. Therefore, finding fermentation processes that reduce the citrinin content in red yeast rice wine products is key to solving these problems. Summary of the Invention

[0005] To address the problems existing in the prior art, this invention proposes a method for preparing health-promoting red yeast rice wine. By brewing rice wine using this method, not only can vitamins and various amino acids from *Pseudomonas aeruginosa* be extracted into the rice wine, but more importantly, various specific amino acids from *Pseudomonas aeruginosa* can effectively control the metabolism of *Monascus purpureus* under certain fermentation conditions, reducing the yield of citrinin in red yeast rice and obtaining high-quality red yeast rice wine.

[0006] To achieve the above objectives, the present invention is implemented through the following technical solution:

[0007] The purpose of this invention is to provide a method for preparing health-promoting red yeast rice wine, comprising the following steps:

[0008] (1) Preparation of liquid fermentation broth of Eclipta prostrata;

[0009] (2) Prepare the yeast starter, which is prepared by weight of 20-30 parts glutinous rice, 5-10 parts soybean, 3-7 parts banana peel, 1-5 parts vitamin B2, 0.1-1.0 parts vitamin C and 8-14 parts sterile water into a yeast starter group and then inoculated with Rhizopus mold for culture.

[0010] (3) After steaming the glutinous rice, rinse it with water to disperse the rice grains. When the temperature is cooled to 32-35℃, add 10-16% of the liquid fermentation liquid of Eriocaulon elm, 0.8%-1.5% of the expanded culture liquid of Monascus purpureus, 0.1%-0.5% of the decanoic acid and 0.2%-0.5% of the expanded culture liquid of Lactobacillus plantarum. Saccharify at 33-35℃ for 40-48 hours to obtain the first fermentation product.

[0011] (4) Add 0.8%-1.5% of the weight of glutinous rice to the first fermentation product, stir evenly, and continue to saccharify at a constant temperature of 33-35℃ for 10-15 hours until the rice grains float to obtain the second fermentation product.

[0012] (5) The second fermentation material is put into the fermentation tank and 1.5-3.0 times the weight of glutinous rice is added with sterile water to continue fermentation to obtain the original wine. The fermentation temperature is 28-32℃ and the fermentation time is 60-72 hours.

[0013] (6) After the fermentation in step (5) is completed, the raw wine is clarified, filtered, sterilized and then bottled as finished product.

[0014] Furthermore, the method for preparing the liquid fermentation broth of *Elaeagnus angustifolia* includes:

[0015] (1) Prepare a culture medium comprising the following components in parts by weight: 0.1-1.0 parts selenium-enriched corn flour, 0.2-2.0 parts glucose, 0.2-0.5 parts peptone, 0.05-0.1 parts potassium dihydrogen phosphate, 0.01-0.03 parts magnesium sulfate, 0.02-0.05 parts L-arginine, 0.03-0.1 parts xylooligosaccharides, and 1000 ml water;

[0016] (2) Sterilize the culture medium obtained in step (1), and then inoculate the elm spp. onto the sterile culture medium;

[0017] (3) After the inoculated culture medium is left to stand for 24 hours, it is cultured in a shaker at 30℃ and 200r / min for 100-120 hours until the mycelial balls are filled with the fermentation liquid. Then the fermentation is terminated to obtain the liquid fermentation liquid of Elm Yellow Fungus.

[0018] Furthermore, the mass of the *Euphorbia ulmoides* inoculated in step (2) is 0.2%-0.6% of the weight of the sterile culture medium.

[0019] Further, the preparation method of the yeast is as follows: (1) Soak glutinous rice and soybeans in water for 24 hours, drain, crush, pass through a 100-mesh sieve, sterilize at 100-110℃ to obtain a mixed powder of glutinous rice and soybeans; (2) Dissolve vitamin B2 and vitamin C completely in sterile water to obtain a vitamin solution; (3) Crush washed and sterilized banana peel; (4) Mix the mixed powder, vitamin solution and banana peel evenly, and knead into yeast dough; (5) Inoculate Rhizopus on the cooled yeast dough and ferment the yeast dough at 25-35℃ for 2-3 days. When the mycelium covers the surface of the yeast dough, ventilate and air dry until the temperature drops to 20-25℃. When the moisture content is no more than 10%, it is ready.

[0020] Furthermore, in step (5), the amount of Rhizopus inoculated is 0.2%-1.0% of the weight of the yeast starter.

[0021] Furthermore, the preparation method of the Lactobacillus plantarum expanded culture medium is as follows: Lactobacillus plantarum is inoculated into Lactobacillus plantarum seed culture medium, and the inoculated culture medium is allowed to stand for 24 hours. Then, it is cultured in a shaker at 30°C and 200 r / min for 100-120 hours until the mycelial balls are filled with the fermentation broth. The fermentation is then terminated to obtain the Lactobacillus plantarum expanded culture medium. The mass of the Lactobacillus inoculation is 0.2% of the mass of the Lactobacillus plantarum seed culture medium.

[0022] Furthermore, the Lactobacillus plantarum seed culture medium consists of: 10.0g casein peptone, 10.0g beef extract, 5.0g yeast powder, 5.0g glucose, 5.0g sodium acetate, 2.0g diammonium citrate, 1.0g Tween 80, 2.0g K2HPO4, 0.2g MgSO4·7H2O, 0.05g MnSO4·H2O, 20.0g Na2CO3, 15.0g agar, 1.0L distilled water, and pH 6.5.

[0023] Furthermore, in step (5), HSZQB5A-1 clarifying agent is used, and the amount added is 0.02% to 0.1% of the original wine weight.

[0024] Compared with the prior art, the present invention has the following beneficial technical effects:

[0025] (1) The red yeast rice wine prepared by the present invention combines the fermentation liquid of Eriobotrya elm containing vitamins and various amino acids with specially made yeast, and co-fermented with glutinous rice in combination with relevant fermentation process. This not only extracts vitamins and various amino acids from Eriobotrya elm into the rice wine, but more importantly, various specific amino acids in Eriobotrya elm can effectively control the metabolism of Monascus purpureus in a certain fermentation environment, thereby reducing the yield of citrinin in red yeast rice. Therefore, the red yeast rice wine prepared by the present invention is nutritious, safe, and has a bright and glossy color.

[0026] (2) In the traditional fermentation process of rice wine, Rhizopus is used as the saccharifying bacteria and yeast is used as the starter culture. However, this invention uses Monascus purpureus as the saccharifying bacteria and specially made Rhizopus as the starter culture. During the saccharification process, the various amino acids and decanoic acid in the fermentation liquid give Monascus purpureus a strong fermentation power and esterification power, which reduces the yield of citric acid without affecting the saccharification efficiency. Experiments show that adding the starter culture to the fermentation liquid 10-15 hours before the fermentation can significantly increase the alcohol content without extending the fermentation time or increasing the amount of starter culture.

[0027] (3) Experiments show that the Lactobacillus plantarum added during the saccharification process will affect the flavor of the finished rice wine. The present invention uses modified Rhizopus as the starter culture, which can increase the acid production to a certain extent and enhance the flavor of the rice wine. Detailed Implementation

[0028] The examples provided illustrate certain embodiments of the invention and should not be construed as limiting the scope of the invention. Improvements can be made to the disclosed content of this invention in terms of materials, methods, and reaction conditions, and all such improvements should fall within the spirit and scope of the invention. Unless otherwise specified, the techniques used in the examples are conventional methods well known to those skilled in the art. Unless otherwise specified, the reagents and biological materials mentioned are commercially available.

[0029] In this application, *Lactobacillus plantarum* and *Elaeagnus pungens* (i.e., *Elaeagnus pungens*) were purchased from the China Industrial Microbial Culture Collection Center.

[0030] Example 1:

[0031] 1. Isolation and purification of Monascus purpureus and preparation of Monascus purpureus culture medium (for specific methods, refer to Wu Genfu's "Experimental Guide to Fermentation Engineering", Higher Education Press).

[0032] Weigh 5g of red yeast rice, add 45ml of sterile water, shake well, and incubate in a 60℃ water bath for 30 minutes to kill heat-sensitive bacteria and yeast. Separate the supernatant spore suspension using the dilution plating method, and incubate at 30℃ for 5 days. Pick single colonies of the mold that produce red pigment and examine them under a microscope to confirm their purity. After confirmation, take pure red yeast rice culture and aseptically transfer it to an agar slant culture medium. Incubate at 30℃ for 4-5 days. Once the slant cells turn purplish-red, store in a refrigerator at 4℃.

[0033] Red Monascus purpureus culture medium expansion: Place 100ml of bean sprout juice culture medium and 5 glass beads in a 500ml Erlenmeyer flask, seal with 8 layers of gauze, wrap with kraft paper, and sterilize at 0.08 MPa for 30 minutes. After cooling, inoculate each flask with 1 / 2 branch of red Monascus purpureus slant culture medium and incubate at 180 rpm in a 30℃ constant temperature shaker for 3-5 days, until the culture medium turns deep red.

[0034] 2. Preparation of Lactobacillus plantarum culture medium

[0035] Lactobacillus plantarum seed culture medium: 10.0g casein peptone, 10.0g beef extract, 5.0g yeast powder, 5.0g glucose, 5.0g sodium acetate, 2.0g diammonium citrate, 1.0g Tween 80, 2.0g K2HPO4, 0.2g MgSO4·7H2O, 0.05g MnSO4·H2O, 20.0g NaCO3, 15.0g agar, 1.0L distilled water, pH 6.5, sterilized.

[0036] Take 2.15g of Lactobacillus plantarum and inoculate it into Lactobacillus plantarum seed culture medium. After the inoculated culture medium is left to stand for 24 hours, it is cultured in a shaker at 30℃ and 200r / min for 100-120h until the mycelial balls are filled with the fermentation broth. Then stop the fermentation to obtain the Lactobacillus plantarum expanded culture medium.

[0037] 3. Preparation of liquid fermentation broth for *Syngonium elm*

[0038] (1) Prepare the culture medium, which includes the following components by weight: 1.0g selenium-enriched corn flour, 2.0g glucose, 2.0g peptone, 0.5g potassium dihydrogen phosphate, 0.1g magnesium sulfate, 0.5g L-arginine, 0.7g xylooligosaccharide and 1000ml water, pH 5.3, sterilize at 110℃ for 20min.

[0039] (2) Take 2.14g of Elm Yellow Fungus and inoculate it onto the culture medium without bacteria in step (1).

[0040] (3) After the inoculated culture medium is left to stand for 24 hours, it is cultured in a shaker at 30℃ and 200r / min for 100-120 hours until the mycelial balls are filled with the fermentation liquid. Then the fermentation is terminated to obtain the liquid of *Eriocaulon elm*.

[0041] 4. Preparation of yeast starter

[0042] The yeast starter ingredients include 20g glutinous rice, 8g soybeans, 3g banana peel, 3.5g vitamin B2, 0.1g vitamin C, and 8g sterile water.

[0043] The preparation method is as follows: (1) Soak glutinous rice and soybeans in water for 24 hours, drain, crush, pass through a 100-mesh sieve, and sterilize at 100-110℃ to obtain a mixed powder of glutinous rice and soybeans; (2) Dissolve vitamin B2 and vitamin C completely in sterile water to obtain a vitamin solution; (3) Crush washed and sterilized banana peels; (4) Mix the mixed powder, vitamin solution and banana peel evenly, and knead into a yeast dough; (5) Inoculate the cooled yeast dough with Rhizopus mold and ferment the yeast dough at 25-35℃ for 2-3 days. When the mycelium covers the surface of the yeast dough, ventilate and air dry until the temperature drops to 20-25℃. When the moisture content is no more than 10%, it is ready. The inoculation amount of Rhizopus mold is 85.2 mg (0.2% of the weight of the yeast dough).

[0044] 5. Red yeast rice wine brewing

[0045] (1) After steaming 1000g of glutinous rice, rinse it with water to disperse the rice grains. When the temperature is cooled to 32-35℃, add 100g of elm yellow fungus liquid fermentation liquid, 8g of red mold expansion culture liquid, 5g of decanoic acid and 2g of plant lactobacillus expansion culture liquid. Saccharify at 33-35℃ for 40-48 hours to obtain the first fermentation product.

[0046] In this embodiment, after removing impurities, the glutinous rice is rinsed and soaked in cold water for 10-12 hours, and then steamed in a rice steamer. The steam pressure of the rice steamer is controlled at 0.8-1.0 MPa, the steam temperature at 110-120°C, and the steaming time at 15-30 minutes. After the glutinous rice is steamed at high temperature, the rice should be free of white core, loose and not mushy, translucent but not mushy, and uniform, which lays the foundation for the subsequent saccharification and fermentation effect.

[0047] (2) Add 8g of yeast to the first fermentation product, stir well, and continue to saccharify at a constant temperature of 33-35℃ for 12 hours until the rice grains float to obtain the second fermentation product.

[0048] (3) Put the second fermented material into the fermentation tank and add 2.2 times the amount of sterile water of glutinous rice to continue fermentation. The fermentation temperature is 28-32℃ and the fermentation time is 65 hours.

[0049] (4) After the fermentation in step (3) is completed, the raw wine is clarified, filtered, sterilized and then bottled as finished product.

[0050] After fermentation, add 0.02% of HSZQB5A-1 clarifying agent by weight of the original wine, stir at 80 rpm for 5 minutes at room temperature, let stand at room temperature for 24 hours, and centrifuge at 6000 rpm for 25 minutes to obtain the clarified liquid.

[0051] The clarified liquid was filtered through a 300-mesh filter cloth, and the filtrate was ultrafiltered to obtain the wine. The wine was then sterilized with 5 Gy of 60Co-γ rays for 8 hours to obtain the high-alcohol rice wine product. The ultrafiltration conditions were: working pressure 0.25 MPa, working temperature 15℃, and filter membrane pore size 0.2 μm.

[0052] Example 2:

[0053] 1. The isolation, purification, and preparation of Monascus purpureus culture medium were the same as in Example 1.

[0054] 2. The preparation of Lactobacillus plantarum culture medium is the same as in Example 1.

[0055] 3. Preparation of liquid fermentation broth for *Syngonium elm*

[0056] (1) Prepare the culture medium, which includes the following components by weight: 5.0g selenium-enriched corn flour, 20g glucose, 5.0g peptone, 0.75g potassium dihydrogen phosphate, 0.2g magnesium sulfate, 0.35g L-arginine, 0.3g xylooligosaccharide and 1000ml water, pH 5.3, sterilize at 110℃ for 20min.

[0057] (2) Take 4.13g of Elm Yellow Fungus and inoculate it onto the culture medium without bacteria in step (1).

[0058] (3) After the inoculated culture medium is left to stand for 24 hours, it is cultured in a shaker at 30℃ and 200r / min for 100-120 hours until the mycelial balls are filled with the fermentation liquid. Then the fermentation is terminated to obtain the liquid of *Eriocaulon elm*.

[0059] 4. Preparation of yeast starter

[0060] The yeast starter ingredients include 25g glutinous rice, 10g soybeans, 5g banana peel, 5g vitamin B2, 0.5g vitamin C, and 11g sterile water.

[0061] The preparation method is as follows: (1) Soak glutinous rice and soybeans in water for 24 hours, drain, crush, pass through a 100-mesh sieve, and sterilize at 100-110℃ to obtain a mixed powder of glutinous rice and soybeans; (2) Dissolve vitamin B2 and vitamin C completely in sterile water to obtain a vitamin aqueous solution; (3) Crush washed and sterilized banana peels; (4) Mix the mixed powder, vitamin aqueous solution and banana peel evenly, and knead into a yeast dough; (5) Inoculate the cooled yeast dough with Rhizopus mold and ferment the yeast dough at 25-35℃ for 2-3 days. When the mycelium covers the surface of the yeast dough, ventilate and air dry until the temperature drops to 20-25℃. When the moisture content is no more than 10%, it is ready. The inoculation amount of Rhizopus mold is 339mg (0.6% of the weight of the yeast dough).

[0062] 5. Red yeast rice wine brewing

[0063] (1) After steaming 1000g of glutinous rice, rinse it with water to disperse the rice grains. When the temperature is cooled to 32-35℃, add 140g of elm yellow fungus liquid fermentation liquid, 15g of red mold expansion culture liquid, 1g of decanoic acid and 3.5g of plant lactobacillus expansion culture liquid. Saccharify at 33-35℃ for 45 hours to obtain the first fermentation product.

[0064] In this embodiment, after removing impurities, the glutinous rice is rinsed and soaked in cold water for 10-12 hours, and then steamed in a rice steamer. The steam pressure of the rice steamer is controlled at 0.8-1.0 MPa, the steam temperature at 110-120°C, and the steaming time at 15-30 minutes. After the glutinous rice is steamed at high temperature, the rice should be free of white core, loose and not mushy, translucent but not mushy, and uniform, which lays the foundation for the subsequent saccharification and fermentation effect.

[0065] (2) Add 12g of yeast to the first fermentation product, stir well, and continue to saccharify at a constant temperature of 33-35℃ for 15 hours until the rice grains float to obtain the second fermentation product.

[0066] (3) Put the second fermented material into the fermentation tank and add 3.0 times the weight of the glutinous rice with sterile water to continue fermentation. The fermentation temperature is 28-32℃ and the fermentation time is 60 hours.

[0067] (4) After the fermentation in step (3) is completed, the raw wine is clarified, filtered, sterilized and then bottled as finished product.

[0068] After fermentation, add 0.06% of HSZQB5A-1 clarifying agent by weight of the original wine, stir at 80 rpm for 5 minutes at room temperature, let stand at room temperature for 24 hours, and centrifuge at 6000 rpm for 25 minutes to obtain the clarified liquid.

[0069] The clarified liquid was filtered through a 300-mesh filter cloth, and the filtrate was ultrafiltered to obtain the wine. The wine was then sterilized with 5 Gy of 60Co-γ rays for 8 hours to obtain the high-alcohol rice wine product. The ultrafiltration conditions were: working pressure 0.25 MPa, working temperature 15℃, and filter membrane pore size 0.2 μm.

[0070] Example 3:

[0071] 1. The isolation, purification, and preparation of Monascus purpureus culture medium were the same as in Example 1.

[0072] 2. The preparation of Lactobacillus plantarum culture medium is the same as in Example 1.

[0073] 3. Preparation of liquid fermentation broth for *Syngonium elm*

[0074] (1) Prepare the culture medium, which includes the following components by weight: 10.0g selenium-enriched corn flour, 12g glucose, 3.5g peptone, 1.0g potassium dihydrogen phosphate, 0.3g magnesium sulfate, 0.2g L-arginine, 1.0g xylooligosaccharide and 1000ml water, pH 5.3, sterilize at 110℃ for 20min.

[0075] (2) Take 6.17g of Elm Yellow Fungus and inoculate it onto the culture medium without bacteria in step (1).

[0076] (3) After the inoculated culture medium is left to stand for 24 hours, it is cultured in a shaker at 30℃ and 200r / min for 100-120 hours until the mycelial balls are filled with the fermentation liquid. Then the fermentation is terminated to obtain the liquid of *Eriocaulon elm*.

[0077] 4. Preparation of yeast starter

[0078] The yeast starter ingredients include 30g glutinous rice, 5g soybeans, 7g banana peel, 1g vitamin B2, 1.0g vitamin C, and 14g sterile water.

[0079] The preparation method is as follows: (1) Soak glutinous rice and soybeans in water for 24 hours, drain, crush, pass through a 100-mesh sieve, and sterilize at 100-110℃ to obtain a mixed powder of glutinous rice and soybeans; (2) Dissolve vitamin B2 and vitamin C completely in sterile water to obtain a vitamin solution; (3) Crush washed and sterilized banana peels; (4) Mix the mixed powder, vitamin solution and banana peel evenly, and knead into a yeast dough; (5) Inoculate Rhizopus on the cooled yeast dough and ferment the yeast dough at 25-35℃ for 2-3 days. When the mycelium covers the surface of the yeast dough, ventilate and air dry until the temperature drops to 20-25℃. When the moisture content is no more than 10%, it is ready. The inoculation amount of Rhizopus is 580mg (1.0% of the weight of the yeast dough).

[0080] 5. Red yeast rice wine brewing

[0081] (1) After steaming 1000g of glutinous rice, rinse it with water to disperse the rice grains. When the temperature is cooled to 32-35℃, add 160g of elm yellow fungus liquid fermentation liquid, 11g of red mold expansion culture liquid, 2.5g of decanoic acid and 5.0g of plant lactobacillus expansion culture liquid. Saccharify at 33-35℃ for 40-48 hours to obtain the first fermentation product.

[0082] In this embodiment, after removing impurities, the glutinous rice is rinsed and soaked in cold water for 10-12 hours, and then steamed in a rice steamer. The steam pressure of the rice steamer is controlled at 0.8-1.0 MPa, the steam temperature at 110-120°C, and the steaming time at 15-30 minutes. After the glutinous rice is steamed at high temperature, the rice should be free of white core, loose and not mushy, translucent but not mushy, and uniform, which lays the foundation for the subsequent saccharification and fermentation effect.

[0083] (2) Add 15g of yeast to the first fermentation product, stir well, and continue to saccharify at a constant temperature of 33-35℃ for 10 hours until the rice grains float to obtain the second fermentation product.

[0084] (3) Put the second fermented material into the fermentation tank and add 1.5 times the amount of sterile water of glutinous rice to continue fermentation. The fermentation temperature is 28-32℃ and the fermentation time is 72 hours.

[0085] (4) After the fermentation in step (3) is completed, the raw wine is clarified, filtered, sterilized and then bottled as finished product.

[0086] After fermentation, add 0.1% of HSZQB5A-1 clarifying agent by weight of the original wine, stir at 80 rpm for 5 minutes at room temperature, let stand at room temperature for 24 hours, and centrifuge at 6000 rpm for 25 minutes to obtain the clarified liquid.

[0087] The clarified liquid was filtered through a 300-mesh filter cloth, and the filtrate was ultrafiltered to obtain the wine. The wine was then sterilized with 5 Gy of 60Co-γ rays for 8 hours to obtain the high-alcohol rice wine product. The ultrafiltration conditions were: working pressure 0.25 MPa, working temperature 15℃, and filter membrane pore size 0.2 μm.

[0088] Comparative example:

[0089] 1. Comparative Examples 1-7 investigated the effects of adding *Eriocaulon elm* liquid fermentation broth, *Monascus purpureus* expanded culture broth, decanoic acid, and *Lactobacillus plantarum* expanded culture broth on the saccharification effect and citrus erythromycin content of rice wine. The formulations are shown in Table 1, and other steps are the same as in Example 2.

[0090] Table 1

[0091]

[0092] 2. Comparative Examples 8-12 investigated the effect of the improved yeast starter on the fermentation effect of rice wine. The experimental methods are shown in Table 2, and the other steps are the same as in Example 2.

[0093] Table 2

[0094]

[0095]

[0096] 3. Comparative Example 13 investigated the effect of the timing of adding yeast on the fermentation effect. The brewing method of red yeast rice wine in Comparative Example 13 is as follows, with other steps the same as in Example 2.

[0097] (1) After steaming 1000g of glutinous rice, rinse it with water to disperse the rice grains. When the temperature is cooled to 32-35℃, add 140g of elm yellow fungus liquid fermentation liquid, 15g of red mold expansion culture liquid, 1g of decanoic acid and 3.5g of plant lactobacillus expansion culture liquid. Saccharify at 33-35℃ for 60 hours to obtain the first fermentation product.

[0098] In this embodiment, after removing impurities, the glutinous rice is rinsed and soaked in cold water for 10-12 hours, and then steamed in a rice steamer. The steam pressure of the rice steamer is controlled at 0.8-1.0 MPa, the steam temperature at 110-120°C, and the steaming time at 15-30 minutes. After the glutinous rice is steamed at high temperature, the rice should be free of white core, loose and not mushy, translucent but not mushy, and uniform, which lays the foundation for the subsequent saccharification and fermentation effect.

[0099] (2) Put the first fermentation material into the fermentation tank and add 12g of yeast and 3.0 times the weight of glutinous rice of sterile water. Stir well and continue fermentation. The fermentation temperature is 28-32℃ and the fermentation time is 60 hours.

[0100] (3) After the fermentation in step (2) is completed, the raw wine is clarified, filtered, sterilized and then bottled as finished product.

[0101] After fermentation, add 0.06% of HSZQB5A-1 clarifying agent by weight of the original wine, stir at 80 rpm for 5 minutes at room temperature, let stand at room temperature for 24 hours, and centrifuge at 6000 rpm for 25 minutes to obtain the clarified liquid.

[0102] The clarified liquid was filtered through a 300-mesh filter cloth, and the filtrate was ultrafiltered to obtain the wine. The wine was then sterilized with 5 Gy of 60Co-γ rays for 8 hours to obtain the high-alcohol rice wine product. The ultrafiltration conditions were: working pressure 0.25 MPa, working temperature 15℃, and filter membrane pore size 0.2 μm.

[0103] Test Example 1:

[0104] 1. The color value of pigments and citric acid in the second fermentation products and raw wines of Examples 1-3 and Comparative Examples 1-7 were determined.

[0105] Method for determining the color value of alcohol-soluble pigments: The Monascus purpureus fermentation broth was centrifuged at 5000 r / min for 10 min, the supernatant was collected, diluted with 70% ethanol by a certain factor, and then incubated in a water bath at 60℃ for 1 h, and filtered; the filtrate was then diluted with 70% ethanol by a certain factor, and the OD505, OD465, and OD410 were measured respectively. The color values ​​of alcohol-soluble red, orange, and yellow pigments were calculated according to the formula: color value = OD × dilution factor / volume of fermentation broth obtained.

[0106] Method for determining the color value of water-soluble pigments: Centrifuge the Monascus fermentation broth at 5000 r / min for 10 min, take the supernatant, dilute it with distilled water by a certain factor, incubate it in a water bath at 60℃ for 1 h, and filter it; dilute the filtrate with distilled water by a certain factor again, and measure OD505, OD465, and OD410 respectively. Calculate the color value of water-soluble red, orange, and yellow pigments according to the formula: color value = OD × dilution factor / volume of fermentation broth obtained.

[0107] The total color value is the sum of the color values ​​of alcohol-soluble pigments and water-soluble pigments.

[0108] Citrulline determination: Take 10 ml of the sample to be tested and incubate at 5000 r / min for 10 min. Add 20 ml of 70% ethanol solution to the supernatant and mix well. Filter through a 0.2 μm microporous membrane and perform high performance liquid chromatography (HPLC) detection according to national standard GB / T 5009, 222-2008. The detection results are shown in Table 3.

[0109] Table 3

[0110]

[0111]

[0112] Table 3 shows that the fermentation process of Monascus purpureus produces a variety of natural pigments, which gives the rice wine produced in this application a bright and vivid color. This experiment also further verifies that the rice wine is stable when stored at 4-28℃ and does not change color within 6 months.

[0113] As can be seen from Comparative Example 1, when Monascus purpureus fermentation broth is used alone as koji, although a large amount of pigment is produced, Monascus purpureus metabolism produces a large amount of the toxic substance citrinin. Moreover, when Monascus purpureus fermentation broth and Lactobacillus plantarum expanded culture broth are used as koji (Comparative Example 4), not only is the problem of high yield of the toxic substance citrinin not significantly improved, but the pigment yield is also reduced. This shows that adding Lactobacillus plantarum expanded culture broth alone cannot control the metabolism of Monascus purpureus to reduce the yield of citrinin.

[0114] Comparative Examples 2-3 and 5 show that when the liquid fermentation broth of *Eriocaulon buergerianum* and decanoic acid, either separately or together, are mixed with the fermentation broth of *Monascus purpureus* to ferment glutinous rice, the pigment yield does not change significantly compared to the comparative examples. The yield of the toxic substance citrinin is reduced, but still significantly higher than that in Examples 1-3. Adding components of the *Lactobacillus plantarum* culture medium to the comparative examples 2-3 did not significantly affect the pigment or citrinin yields.

[0115] As shown in Table 3, the fermentation broth of *Eriocaulon elm* is rich in various amino acids, and decanoic acid can control the metabolism of *Monascus purpureus*, thereby reducing the yield of citrinin to a certain extent. The expanded culture broth of *Lactobacillus plantarum* promotes this control. In the saccharification process of this invention, the fermentation broth of *Eriocaulon elm*, the expanded culture broth of *Monascus purpureus*, decanoic acid, and the expanded culture broth of *Lactobacillus plantarum* exhibit a synergistic effect among the raw materials used to prepare the starter culture. The interaction of these multiple raw materials enables the starter culture to have better saccharification ability, better pigment production ability, and the ability to reduce citrinin yield in the fermentation environment of this application.

[0116] Test Example 2:

[0117] The yield, reducing sugar, alcohol content, and total acid content of the raw wines from Examples 1-3 and Comparative Examples 8-12 were tested (the method for detecting reducing sugar content was based on GB5009.7, the method for detecting alcohol content was based on GB5009.225, and the method for detecting total acid content was based on GB / T 12456). The results are shown in Table 4.

[0118] Table 4. Yield, reducing sugar, alcohol content, and total acid content of the raw spirit.

[0119]

[0120] As shown in Table 4, after improving the yeast formula, the yield, alcohol content, and total acid content of the wine were significantly increased compared with pure Rhizopus (Comparative Example 8). Compared with pure activated brewing yeast (Comparative Example 9), the yield and alcohol content were comparable, but the total acid content was higher. Rhizopus has a certain acid-producing ability. On the one hand, the presence of a certain amount of acid in the raw wine is beneficial for preserving the flavor of rice wine. On the other hand, maintaining a certain acidity in the mash is beneficial for inhibiting the growth of miscellaneous bacteria. This is consistent with the fact that in Examples 1-3 of this application, when the total acid was 0.79-0.84 mg / L, the yield and alcohol content were higher than those of the comparative example.

[0121] This application utilizes Comparative Example 8 to extend the fermentation time under existing conditions. Specifically, the second fermentation product was added to a fermentation tank along with 3.0 times the weight of the glutinous rice in sterile water for continued fermentation to obtain the original wine. The fermentation temperature was 28–32°C, and the fermentation times were 70, 80, 90, 95, 100, 105, 110, 115, and 120 hours, respectively. Experiments showed that with extended fermentation time, when using pure Rhizopus mold as the starter culture, both the yield and alcohol content increased, reaching a level comparable to Example 2 after 117–118 hours of fermentation.

[0122] Therefore, it can be seen that the improved yeast formula of this invention helps to enhance the growth and metabolism of Rhizopus, accelerate the fermentation rate, increase sugar consumption, increase alcohol accumulation, and correspondingly improve the alcohol content and yield of rice wine.

[0123] Test Example 3:

[0124] The yield, reducing sugar, alcohol content, and total acid content of the raw wines from Example 2 and Comparative Example 13 were tested (the method for detecting reducing sugar content was based on GB5009.7, the method for detecting alcohol content was based on GB 5009.225, and the method for detecting total acid content was based on GB / T 12456). The results are shown in Table 5.

[0125] Table 5

[0126] Group Alcohol yield reducing sugars Alcohol content (%) Total acid (mg / L) Example 2 75％ 14.9 9.6 0.79 Comparative Example 13 55％ 22.6 5.3 0.46

[0127] Experiments show that adding yeast to the fermentation liquid 10-15 hours before the start of fermentation can significantly increase the alcohol content without extending the fermentation time or increasing the amount of yeast used.

[0128] The above description is only a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any equivalent substitutions or modifications made by those skilled in the art within the scope of the technology disclosed in the present invention, based on the technical solution and inventive concept of the present invention, should be covered within the scope of protection of the present invention.

Claims

1. A method for preparing a health-promoting red yeast rice wine, characterized in that, Includes the following steps: (1) Preparation of liquid fermentation broth of Elm Yellow Fungus; (2) Preparation of yeast, wherein the yeast is prepared by weight of 20-30 parts glutinous rice, 5-10 parts soybean, 3-7 parts banana peel, 1-5 parts vitamin B2, 0.1-1.0 parts vitamin C and 8-14 parts sterile water into yeast clumps and then inoculated with Rhizopus mold for culture; (3) After steaming the glutinous rice, rinse it with water to disperse the rice grains. When it is cooled to 32-35℃, add 10-16% of the liquid fermentation liquid of Eriocaulon elm, 0.8%-1.5% of the expanded culture liquid of Monascus purpureus, 0.1%-0.5% of the decanoic acid and 0.2%-0.5% of the expanded culture liquid of Lactobacillus plantarum. Saccharify at 33-35℃ for 40-48 hours to obtain the first fermentation product. (4) Add 0.8%-1.5% of the glutinous rice mass of yeast to the first fermentation product, stir evenly, and continue to saccharify at a constant temperature of 33-35℃ for 10-15 hours until the rice grains float to obtain the second fermentation product; (5) The second fermentation material is put into the fermentation tank and 1.5-3.0 times the weight of glutinous rice is added with sterile water to continue fermentation to obtain the original wine. The fermentation temperature is 28-32℃ and the fermentation time is 60-72 hours. (6) After the fermentation of the raw wine in step (5) is completed, it is clarified, filtered and sterilized and then bottled as finished product; The preparation method of the liquid fermentation broth of *Elaeagnus angustifolia* includes: (1) Prepare a culture medium comprising the following components by weight: 0.1-1.0 parts selenium-enriched corn flour, 0.2-2.0 parts glucose, 0.2-0.5 parts peptone, 0.05-0.1 parts potassium dihydrogen phosphate, 0.01-0.03 parts magnesium sulfate, 0.02-0.05 parts L-arginine, 0.03-0.1 parts xylooligosaccharide, and 1000 ml water; (2) The culture medium obtained in step (1) is sterilized, and then *Euphorbia ulmoides* is inoculated onto the sterile culture medium; (3) After the inoculated culture medium is left to stand for 24 hours, it is cultured in a shaker at 30℃ and 200r / min for 100-120h until the mycelial balls are filled with the fermentation broth. Then the fermentation is terminated to obtain the liquid fermentation broth of Elm Yellow Fungus. The mass of *Euphorbia ulmoides* inoculated in step (2) is 0.2%-0.6% of the weight of the sterile culture medium.

2. The method for preparing a health-promoting red yeast rice wine according to claim 1, characterized in that, The preparation method of the yeast is as follows: (1) Soak glutinous rice and soybeans in water for 24 hours, drain, crush, pass through a 100-mesh sieve, sterilize at 100-110℃ to obtain a mixed powder of glutinous rice and soybeans; (2) Dissolve vitamin B2 and vitamin C completely in sterile water to obtain a vitamin solution; (3) Crush the washed and sterilized banana peel; (4) Mix the mixed powder, vitamin solution and banana peel evenly, and knead into a yeast dough; (5) Inoculate the cooled yeast dough with Rhizopus and ferment the yeast dough at 25-35℃ for 2-3 days. When the mycelium covers the surface of the yeast dough, ventilate and air dry until the temperature drops to 20-25℃. When the moisture content is no more than 10%, it is ready.

3. The method for preparing a health-promoting red yeast rice wine according to claim 1, characterized in that, In step (5), the amount of Rhizopus inoculated is 0.2%-1.0% of the weight of the yeast starter.

4. The method for preparing a health-promoting red yeast rice wine according to claim 1, characterized in that, The preparation method of the Lactobacillus plantarum expanded culture medium is as follows: Lactobacillus plantarum is inoculated into Lactobacillus plantarum seed culture medium, and the inoculated culture medium is allowed to stand for 24 hours. Then, it is cultured in a shaker at 30°C and 200 r / min for 100-120 hours until the mycelial balls are filled with the fermentation broth. The fermentation is then terminated to obtain the Lactobacillus plantarum expanded culture medium. The mass of Lactobacillus plantarum inoculated is 0.2% of the mass of Lactobacillus plantarum seed culture medium.

5. The method for preparing a health-promoting red yeast rice wine according to claim 4, characterized in that, The Lactobacillus plantarum seed culture medium consists of: 10.0 g casein peptone, 10.0 g beef extract, 5.0 g yeast extract, 5.0 g glucose, 5.0 g sodium acetate, 2.0 g diammonium citrate, 1.0 g Tween 80, 2.0 g K₂HPO₄, 0.2 g MgSO₄·7H₂O, 0.05 g MnSO₄·H₂O, 20.0 g Na₂CO₃, 15.0 g agar, 1.0 L distilled water, and pH 6.5.

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