Low-bitterness yellow rice wine and preparation method thereof

Abstract

The application provides a low-bitterness yellow rice wine and a preparation method thereof, and belongs to the technical field of yellow rice wine brewing. The preparation method of the low-bitterness yellow rice wine comprises the following steps: preparing lactic acid bacteria koji; mixing rice and water, and then adding liquefying enzyme and saccharifying enzyme in sequence to obtain saccharifying liquid; adding yellow rice wine koji, yeast activation liquid and lactic acid bacteria koji into the saccharifying liquid to perform fermentation and obtain fermentation liquid; adding enzyme preparation into the fermentation liquid to perform enzymolysis, filtering and sterilizing to obtain yellow rice wine clear liquid; adding tannin solution into the yellow rice wine clear liquid, stirring and mixing, adsorbing and filtering to obtain yellow rice wine. The prepared yellow rice wine has significantly reduced bitter substances, improved entry comfort and significantly improved sensory quality; and the content of ester substances is increased, and the yellow rice wine has mellow and rich aroma.

Description

Technical Field

[0001] This invention belongs to the field of rice wine brewing technology, specifically relating to a low-bitterness rice wine and its preparation method. Background Technology

[0002] Yellow wine, or huangjiu, is one of the oldest alcoholic beverages in the world. It is a fermented wine made primarily from rice, millet, sorghum, corn, wheat, and water, through the addition of koji (fermentation starter) and / or some enzyme preparations and yeast as saccharifying and fermenting agents. During the brewing process, microorganisms break down a large amount of protein in the grains into amino acids. The bitterness of yellow wine mainly comes from certain amino acids, tyrosol, methionine, and amines produced during fermentation. Chinese patent CN202110665333.6 discloses a method for extracting, separating, identifying, and verifying bitter peptides in yellow wine. This method establishes a method for the extraction, separation, identification, and verification of bitter peptides in yellow wine, revealing the structure and bitter characteristics of these peptides. It also discovers for the first time a bitter peptide with the amino acid sequence Leu-Pro-Thr-Leu in yellow wine. Chinese patent CN202011174297.5 discloses a method for separating and identifying bitter polypeptides in Shaoxing rice wine. This method uses a combination of methods to separate and identify the bitter polypeptides in Shaoxing rice wine, and verifies that the polypeptide contains pyroglutamic acid through enzymatic digestion with pyroglutamic acid peptidase. This provides a theoretical basis for the research of mechanized Shaoxing rice wine bitterness reduction technology. Chinese patent CN201910623896.1 discloses a method for improving the quality of mechanized Shaoxing rice wine. It uses *Aspergillus oryzae* and two strains of *Saccharomyces cerevisiae* selected from Shaoxing rice wine starter to prepare a special starter for mechanized Shaoxing rice wine, thereby improving the aroma and taste. It also uses raw wheat as raw material and *Aspergillus oryzae* SR-201, a late-spore-producing mold, to prepare a special wheat starter for mechanized Shaoxing rice wine, thereby reducing the bitterness of the mechanized Shaoxing rice wine and achieving the goal of improving its quality. The Shaoxing Jiafan wine produced by this invention meets all the national standards for Shaoxing rice wine. The content of higher alcohols and aldehydes is reduced, while the content of esters is increased, resulting in a richer aroma, significantly reduced bitterness, and significantly improved sensory quality. Chinese patent CN201910623796.9 discloses a strain of Aspergillus oryzae with high saccharification power and delayed spore production for rice wine koji making and its application. The Aspergillus oryzae SR-201 has a delayed spore production time of 1 day, with the strain starting to produce spores after 4 days and producing abundant spores after 5 days. In pure wheat koji production, compared with Su-16, the number of spores in the koji is reduced by more than 80%, and the saccharification power is increased by more than 15%, which can significantly reduce the bitterness brought to rice wine by excessive spores in cooked wheat koji.

[0003] Current research on bitter substances in rice wine focuses on the material analysis level, mainly revealing the source and separation method of bitterness in rice wine, or improving the aroma and reducing bitterness by adding yeast or Aspergillus oryzae in the koji-making process. There are no other technical routes besides the application of microbial strains to specifically reduce bitter substances in rice wine. Summary of the Invention

[0004] The purpose of this invention is to overcome the shortcomings of the prior art and provide a low-bitterness rice wine and its preparation method. The rice wine prepared by this invention has significantly reduced bitterness and has a harmonious aroma and taste.

[0005] To achieve the above objectives, the technical solution adopted by the present invention is as follows: a method for preparing low-bitterness rice wine, comprising the following steps:

[0006] S1. Preparation of lactic acid bacteria koji;

[0007] S2. Mix rice and water, then add liquefying enzyme and saccharifying enzyme in sequence to obtain saccharified liquid; add rice wine starter, yeast activation liquid and lactic acid bacteria starter to the saccharified liquid and ferment to obtain fermentation liquid;

[0008] S3. Add enzyme preparation to fermentation broth for enzymatic hydrolysis, filter and sterilize to obtain clear rice wine liquid;

[0009] S4. Add tannin solution to the clear rice wine liquid, stir and mix well, adsorb and filter to obtain rice wine.

[0010] In the existing brewing stage of Shaoxing wine, a large amount of bitter substances are produced during microbial fermentation. However, there is limited research on reducing these bitter substances in Shaoxing wine. The inventors of this application, through extensive research, have reduced bitter substances in Shaoxing wine and improved its aroma and taste in three ways. First, by adding lactic acid bacteria koji, the content of lactic acid bacteria in the pre-fermentation stage of Shaoxing wine is increased, the pre-fermentation time is extended, and the content of higher alcohols in Shaoxing wine is reduced, thereby reducing bitterness and enhancing aroma. Second, enzyme preparations are selected to enzymatically hydrolyze proteins, fully degrading bitter peptides, thus reducing bitterness and improving the amino acid nitrogen content in Shaoxing wine, thereby improving its flavor. Finally, by adding tannins to complex with bitter amino acids, followed by adsorption filtration with a clarifying agent and removal through molecular weight-directed ultrafiltration, the content of bitter amino acids is reduced, thus weakening bitterness.

[0011] In a preferred embodiment of the preparation method of the low-bitter rice wine of the present invention, the lactic acid bacteria koji is prepared using Lactococcus plantarum and Lactobacillus delbrueckii subsp. lactis.

[0012] The inventors of this application have discovered that lactic acid bacteria koji prepared using two combinations of lactic acid bacteria seed liquid can rapidly enrich lactic acid bacteria; adding lactic acid bacteria koji in the early stage of rice wine fermentation can rapidly increase the lactic acid bacteria content in the pre-fermentation stage, effectively prolong the pre-fermentation time of rice wine, reduce the accumulation of higher alcohols, and thus reduce the bitterness of rice wine.

[0013] In a preferred embodiment of the preparation method of the low-bitter rice wine of the present invention, the preservation number of the *Lactococcus plantarum* is GDMCC NO.1.1033; and the preservation number of *Lactobacillus delbrueckii* subsp. *lactobacter* is GDMCC NO.1.3694.

[0014] In a preferred embodiment of the preparation method of the low-bitter rice wine of the present invention, the amount of lactic acid bacteria koji added in step S2 is 1-3% by weight of rice.

[0015] In a preferred embodiment of the method for preparing low-bitter rice wine according to the present invention, the enzyme preparation in step S3 includes aminopeptidase and flavor protease.

[0016] As a preferred embodiment of the preparation method of the low-bitter rice wine of the present invention, the enzymatic hydrolysis in step S3 specifically involves: adding aminopeptidase to the fermentation broth and hydrolyzing it at 50-60℃ for 9-15 hours, then adding flavor protease and hydrolyzing it at 50-60℃ for 9-15 hours.

[0017] The inventors of this application have discovered that a segmented enzymatic hydrolysis method is used in the post-fermentation stage of Shaoxing wine. This method involves adding aminopeptidase first, followed by enzymatic hydrolysis, and then adding flavor protease for further enzymatic hydrolysis. This fully degrades bitter peptide chains into amino acids, reducing bitterness and simultaneously increasing the amino acid nitrogen content in the Shaoxing wine, thus enhancing its umami flavor. Specifically, aminopeptidase first acts on the fermentation broth, generating many free amino acids. Simultaneously, hydrophobic amino acids move from the middle to the end of the peptide chain, reducing bitterness. Then, flavor protease further fully degrades the bitter peptides, further reducing bitterness.

[0018] In a preferred embodiment of the preparation method of the low-bitter rice wine of the present invention, the amount of aminopeptidase added is 1-3% by mass percentage of the fermentation liquid; the amount of flavor protease added is 1-3%.

[0019] In a preferred embodiment of the preparation method of the low-bitter rice wine of the present invention, the concentration of the tannin solution is 0.8-1.2%.

[0020] This invention adds tannins to the clear liquid of rice wine, which interact hydrophobically with some nonpolar hydrophobic bitter amino acids and form non-covalent interactions with some positively charged polar bitter amino acids to form complexes. These complexes are then removed by filtration through bentonite ion exchange adsorption combined with molecular weight directional ultrafiltration. This significantly reduces the content of bitter amino acids such as lysine, leucine, and isoleucine, thus weakening the bitterness of rice wine.

[0021] In a preferred embodiment of the preparation method of the low-bitter rice wine of the present invention, in step S4, bentonite and diatomaceous earth are used for adsorption, followed by filtration by pressure filtration and ultrafiltration.

[0022] A type of rice wine, prepared by the aforementioned method for preparing low-bitter rice wine.

[0023] The beneficial effects of this invention are as follows: This invention provides a low-bitterness rice wine and its preparation method. This invention involves adding lactic acid bacteria koji during the pre-fermentation stage of the rice wine, followed by segmental enzymatic hydrolysis using aminopeptidase and flavor protease after pre-fermentation, and finally adding tannins to complex bitter amino acids. The rice wine prepared by this invention has significantly reduced bitter substances, improved palatability, and significantly enhanced sensory quality; the ester content of the rice wine prepared by this invention is increased, resulting in a rich and mellow aroma; and the invention reduces bitterness by hydrolyzing bitter peptides or precipitating bitter amino acids, thus significantly improving the stability of the rice wine. Attached Figure Description

[0024] Figure 1 A flavor wheel for tasting yellow rice wine. Detailed Implementation

[0025] To more concisely and clearly demonstrate the technical solution, purpose, and advantages of the present invention, the technical solution of the present invention is described in detail below with reference to specific embodiments. Unless otherwise specified, the reagents involved in the embodiments of the present invention are all commercially available products and can be purchased through commercial channels.

[0026] The lactic acid bacteria seed solution described in the embodiments and comparative examples of this invention includes *Lactococcus plantarum* and *Lactobacillus delbrueckii* subsp. *lactobacter*; the ratio of *Lactococcus plantarum* to *Lactobacillus delbrueckii* subsp. *lactobacter* in the lactic acid bacteria seed solution is 1:1.

[0027] The *Lactococcus plantarum* specimen, with accession number GDMCC NO.1.1033, was purchased from the Guangdong Provincial Center for Microbial Culture Collection; the *Lactobacillus delbrueckii* subsp. *lactotrichum* specimen, with accession number GDMCC NO.1.3694, was also purchased from the Guangdong Provincial Center for Microbial Culture Collection.

[0028] Example 1

[0029] This embodiment provides a method for preparing low-bitterness rice wine, including the following steps:

[0030] S1. Streak the two types of purchased lactic acid bacteria onto MRS agar plates and incubate upside down at 30-37℃ for 48 hours. Pick a single colony and inoculate it into 10 mL of MRS liquid medium, then incubate statically at 37℃ for 24 hours. Take the cultured bacterial solution and transfer it to 100 mL of MRS liquid medium at a ratio of 1%, then incubate statically at 37℃ for 24 hours to obtain the lactic acid bacteria seed culture. Mix wheat bran and rice husks at a weight ratio of 10:1, add water equal to 70% of the total weight of the wheat bran and rice husks, and mix thoroughly until there is no obvious dry or clumped material. Spread the mixed material evenly in a steamer and cover it with gauze. After the water boils, place the steamer on top, and poke some holes in the surface of the material to improve its looseness. After the steam rises, cover the pot and steam for 60 minutes. After steaming, allow the material to cool naturally to 40°C, and add water until it reaches 70% of the total weight of the wheat bran and rice husks. In a clean bench, weigh out 3% of the lactic acid bacteria seed solution (1.0 × 10⁻⁶) by weight of the material. 6 Mix the CFU / g thoroughly, cover the koji material with gauze and compact it; place the inoculated koji material in a constant temperature and humidity incubator and incubate at 37℃ and 85% humidity for 7 hours. Turn off the humidifier and wait for the temperature of the koji core to drop to the temperature of the incubator before drying the koji material to obtain lactic acid bacteria koji.

[0031] S2. Add the materials to a pot at a rice-to-water ratio of 1:2, add 0.05% of the rice amount of liquefying enzyme, heat to 92℃, keep warm for 30 minutes, then heat to 98℃ and keep warm for 20 minutes; cool the materials to 63℃, add 0.1% of the rice amount of saccharifying enzyme, keep warm for 1 hour to prepare saccharification liquid; cool the saccharification liquid to room temperature, then add 10% of the rice amount of rice wine starter, 0.2% of the yeast activation liquid and 1% of the lactic acid bacteria starter, add water to make up to the predetermined weight (water evaporates during the replenishment process), stir evenly; ferment at 25℃ for 3 days, after being transferred to a tank, stir at least twice a day, after 3 days transfer to 15℃ for 25 days of fermentation to obtain fermentation liquid;

[0032] S3. After the pre-fermentation is completed, 1% aminopeptidase is added to the fermentation liquid in sequence and hydrolyzed at 50℃ for 15 hours. Then, 1% flavor protease is added and hydrolyzed at 50℃ for 15 hours. Perlite is used as a filter aid for filtration. The resulting sake is collected and bottled into clean glass bottles. The sake is then sterilized at 85℃ for 25 minutes to obtain clear rice wine liquid.

[0033] S4. Prepare a 1% tannin solution, then add the tannin solution to the clear rice wine liquid from step S3. Stop adding when no obvious precipitate forms, stir and mix well, and let stand for 16 hours. Adsorb the flocculated solution with bentonite and diatomaceous earth for 16 hours, stir and mix well, then filter under pressure, and then filter through a 20kDa ultrafiltration membrane to obtain rice wine.

[0034] Example 2

[0035] This embodiment provides a method for preparing low-bitterness rice wine, including the following steps:

[0036] S1. Streak the two types of purchased lactic acid bacteria onto MRS agar plates and incubate upside down at 30-37℃ for 48 hours. Pick a single colony and inoculate it into 10 mL of MRS liquid medium, then incubate statically at 37℃ for 24 hours. Take the cultured bacterial solution and transfer it to 100 mL of MRS liquid medium at a ratio of 1%, then incubate statically at 37℃ for 24 hours to obtain the lactic acid bacteria seed culture. Mix wheat bran and rice husks at a weight ratio of 10:1, add water equal to 70% of the total weight of the wheat bran and rice husks, and mix thoroughly until there is no obvious dry or clumped material. Spread the mixed material evenly in a steamer and cover it with gauze. After the water boils, place the steamer on top, and poke some holes in the surface of the material to improve its looseness. After the steam rises, cover the pot and steam for 60 minutes. After steaming, allow the material to cool naturally to 40°C, and add water until it reaches 70% of the total weight of the wheat bran and rice husks. In a clean bench, weigh out 3% of the lactic acid bacteria seed solution (1.0 × 10⁻⁶) by weight of the material. 6 Mix the CFU / g thoroughly, cover the koji material with gauze and compact it; place the inoculated koji material in a constant temperature and humidity incubator and incubate at 37℃ and 85% humidity for 7 hours. Turn off the humidifier and wait for the temperature of the koji core to drop to the temperature of the incubator before drying the koji material to obtain lactic acid bacteria koji.

[0037] S2. Add the materials to a pot at a rice-to-water ratio of 1:2, add 0.05% of the rice amount of liquefying enzyme, heat to 92℃, keep warm for 30 minutes, then heat to 98℃ and keep warm for 20 minutes; cool the materials to 63℃, add 0.1% of the rice amount of saccharifying enzyme, keep warm for 1 hour to prepare saccharification liquid; cool the saccharification liquid to room temperature, then add 10% of the rice amount of rice wine starter, 0.2% of the yeast activation liquid and 2% of the lactic acid bacteria starter, add water to make up to the predetermined weight (water evaporates during the replenishment process), stir evenly; ferment at 25℃ for 3 days, after being transferred to a tank, stir at least twice a day, after 3 days transfer to 15℃ for 25 days of fermentation to obtain fermentation liquid;

[0038] S3. After the pre-fermentation is completed, 2% aminopeptidase is added to the fermentation liquid in sequence and hydrolyzed at 55℃ for 12 hours. Then, 2% flavor protease is added and hydrolyzed at 55℃ for 12 hours. Perlite is used as a filter aid for filtration. The clear wine is collected and bottled into clean glass bottles. It is then sterilized at 85℃ for 25 minutes to obtain clear rice wine liquid.

[0039] S4. Prepare a 1% tannin solution, then add the tannin solution to the clear rice wine liquid from step S3. Stop adding the tannin solution when no obvious precipitate is formed, stir and mix well, and let it stand for 24 hours. Adsorb the flocculated solution with bentonite and diatomaceous earth for 24 hours, stir and mix well, filter by pressure, and then filter through a 20kDa ultrafiltration membrane to obtain rice wine.

[0040] Example 3

[0041] This embodiment provides a method for preparing low-bitterness rice wine, including the following steps:

[0042] S1. Streak the two types of purchased lactic acid bacteria onto MRS agar plates and incubate upside down at 30-37℃ for 48 hours. Pick a single colony and inoculate it into 10 mL of MRS liquid medium, then incubate statically at 37℃ for 24 hours. Take the cultured bacterial solution and transfer it to 100 mL of MRS liquid medium at a ratio of 1%, then incubate statically at 37℃ for 24 hours to obtain the lactic acid bacteria seed culture. Mix wheat bran and rice husks at a weight ratio of 10:1, add water equal to 70% of the total weight of the wheat bran and rice husks, and mix thoroughly until there is no obvious dry or clumped material. Spread the mixed material evenly in a steamer and cover it with gauze. After the water boils, place the steamer on top, and poke some holes in the surface of the material to improve its looseness. After the steam rises, cover the pot and steam for 60 minutes. After steaming, allow the material to cool naturally to 40°C, and add water until it reaches 70% of the total weight of the wheat bran and rice husks. In a clean bench, weigh out 3% of the lactic acid bacteria seed solution (1.0 × 10⁻⁶) by weight of the material. 6 Mix the CFU / g thoroughly, cover the koji material with gauze and compact it; place the inoculated koji material in a constant temperature and humidity incubator and incubate at 37℃ and 85% humidity for 7 hours. Turn off the humidifier and wait for the temperature of the koji core to drop to the temperature of the incubator before drying the koji material to obtain lactic acid bacteria koji.

[0043] S2. Add the materials to a pot at a rice-to-water ratio of 1:2, add 0.05% liquefying enzyme (based on rice weight), heat to 92℃, keep warm for 30 minutes, then heat to 98℃ and keep warm for 20 minutes; cool the materials to 63℃, add 0.1% saccharifying enzyme (based on rice weight), and keep warm for 1 hour to prepare a saccharified liquid; cool the saccharified liquid to room temperature, then add 10% rice wine starter, 0.2% yeast activation liquid, and 3% lactic acid bacteria starter, add water to make up to the predetermined weight (water evaporates during replenishment), and stir evenly; ferment at 25℃ for 3 days, stirring at least twice a day after being transferred to a tank, then transfer to 15℃ for 25 days of fermentation to obtain a fermented liquid;

[0044] S3. After the pre-fermentation is completed, add 3% aminopeptidase to the fermentation liquid in sequence and hydrolyze at 60℃ for 9 hours. Then add 3% flavor protease and hydrolyze at 60℃ for 9 hours. Use perlite as a filter aid to filter and collect the sake. Bottle the sake into clean glass bottles and sterilize at 85℃ for 25 minutes to obtain clear rice wine liquid.

[0045] S4. Prepare a 1% tannin solution, then add the tannin solution to the clear rice wine liquid from step S3. Stop adding when no obvious precipitate forms, stir and mix well, and let stand for 32 hours. Adsorb the flocculated solution with bentonite and diatomaceous earth for 32 hours, stir and mix well, then filter under pressure, and then filter through a 20kDa ultrafiltration membrane to obtain rice wine.

[0046] Comparative Example 1

[0047] This comparative example provides a method for preparing rice wine, including the following steps:

[0048] S1. Add the materials to a pot at a rice-to-water ratio of 1:2, add 0.05% liquefying enzyme (based on rice weight), heat to 92℃, keep warm for 30 minutes, then heat to 98℃ and keep warm for 20 minutes; cool the materials to 63℃, add 0.1% saccharifying enzyme (based on rice weight), and keep warm for 1 hour to prepare a saccharified liquid; cool the saccharified liquid to room temperature, then add 10% rice wine yeast and 0.2% yeast activation liquid (based on rice weight), add water to make up to the predetermined weight (water evaporates during the replenishment process), and stir evenly; ferment at 25℃ for 3 days, stirring at least twice a day after being transferred to a tank, then transfer to 15℃ for 25 days of fermentation to obtain a fermented liquid;

[0049] S2. After the pre-fermentation is completed, 1% aminopeptidase is added to the fermentation broth in sequence and hydrolyzed at 50℃ for 15 hours. Then, 1% flavor protease is added and hydrolyzed at 50℃ for 15 hours. Perlite is used as a filter aid for filtration. The resulting sake is collected and bottled into clean glass bottles. The sake is then sterilized at 85℃ for 25 minutes to obtain clear rice wine liquid.

[0050] S3. Prepare a 1% tannin solution, then add the tannin solution to the clear rice wine liquid from step S3. Stop adding when no obvious precipitate forms, stir and mix well, and let stand for 16 hours. Adsorb the flocculated solution with bentonite and diatomaceous earth for 16 hours, stir and mix well, and then filter under pressure to obtain clear rice wine. Filter the obtained clear rice wine through a 20kDa ultrafiltration membrane to obtain rice wine.

[0051] Comparative Example 2

[0052] S1. Streak the two types of purchased lactic acid bacteria onto MRS agar plates and incubate upside down at 30-37℃ for 48 hours. Pick a single colony and inoculate it into 10 mL of MRS liquid medium, then incubate statically at 37℃ for 24 hours. Take the cultured bacterial solution and transfer it to 100 mL of MRS liquid medium at a ratio of 1%, then incubate statically at 37℃ for 24 hours to obtain the lactic acid bacteria seed culture.

[0053] S2. Add the materials to a pot at a rice-to-water ratio of 1:2, add 0.05% liquefying enzyme (based on rice weight), heat to 92℃, keep warm for 30 minutes, then heat to 98℃ and keep warm for 20 minutes; cool the materials to 63℃, add 0.1% saccharifying enzyme (based on rice weight), and keep warm for 1 hour to prepare a saccharified liquid; cool the saccharified liquid to room temperature, then add 10% rice wine yeast, 0.2% yeast activation liquid, and 2% lactic acid bacteria seed liquid (based on rice weight), add water to make up to the predetermined weight (water evaporates during replenishment), and stir evenly; ferment at 25℃ for 3 days, stirring at least twice a day after being transferred to a tank, then transfer to 15℃ for 25 days of fermentation to obtain a fermented liquid;

[0054] S3. After the pre-fermentation is completed, 1% aminopeptidase is added to the fermentation liquid in sequence and hydrolyzed at 50℃ for 15 hours. Then, 1% flavor protease is added and hydrolyzed at 50℃ for 15 hours. Perlite is used as a filter aid for filtration. The resulting sake is collected and bottled into clean glass bottles. The sake is then sterilized at 85℃ for 25 minutes to obtain clear rice wine liquid.

[0055] S4. Prepare a 1% tannin solution, then add the tannin solution to the clear rice wine liquid from step S3. Stop adding when there is no obvious precipitation, stir and mix well, and let stand for 16 hours. Adsorb the flocculated solution with bentonite and diatomaceous earth for 16 hours, stir and mix well, then filter under pressure, and then filter through a 20kDa ultrafiltration membrane to obtain rice wine.

[0056] Comparative Example 3

[0057] The difference between this comparative example and Example 1 lies only in step S2. Step S2 of this comparative example is as follows:

[0058] S2. Add the materials to a pot at a rice-to-water ratio of 1:2, add 0.05% of the rice amount of liquefying enzyme, heat to 92℃, keep warm for 30 minutes, then heat to 98℃ and keep warm for 20 minutes; cool the materials to 63℃, add 0.1% of the rice amount of saccharifying enzyme, keep warm for 1 hour to prepare saccharification liquid; cool the saccharification liquid to room temperature, then add 10% of the rice amount of rice wine starter and 0.2% of the yeast activation liquid, add water to make up to the predetermined weight (water evaporates during the replenishment process), stir evenly; ferment at 25℃ for 3 days, after being placed in a tank, stir at least twice a day, after 3 days transfer to 15℃ and add 2% lactic acid bacteria starter, ferment for 25 days to obtain fermentation liquid.

[0059] Comparative Example 4

[0060] The only difference between this comparative example and Example 1 is step S3, which is as follows:

[0061] After the initial fermentation is completed, 1% aminopeptidase and 1% flavor protease are added simultaneously for enzymatic hydrolysis at 50℃ for 15 hours. Perlite is used as a filter aid for filtration, and the resulting sake is collected. The sake is then bottled into clean glass bottles and sterilized at 85℃ for 25 minutes to obtain clear rice wine liquid.

[0062] Comparative Example 5

[0063] The only difference between this comparative example and Example 1 is step S4. Step S4 of this comparative example is as follows: S4. Bentonite and diatomaceous earth are used to adsorb the clear liquid of rice wine for 16 hours, stir and mix well, filter, and then filter through a 20kDa ultrafiltration membrane to obtain rice wine.

[0064] Example of effect

[0065] 1. The physicochemical properties of the rice wines prepared in Examples 1-3 and Comparative Examples 1-5 were tested and sensory evaluations were performed. The results are shown in Table 1.

[0066] Turbidity

[0067] Destructive testing method: The sample is frozen at -18℃ for one day, thawed at 50℃, and then frozen at -18℃ for one day. This freeze-thaw cycle is repeated 7 times, and the turbidity of the sample is then measured.

[0068] The sensory rating method is as follows:

[0069] like Figure 1 As shown, a flavor wheel for huangjiu (yellow wine) evaluation is constructed based on smell and taste. The specific method is as follows: Huangjiu samples are placed in dry, odorless stemmed glasses according to random numbers. Evaluators conduct sensory evaluations by smelling and tasting in a well-lit environment free of external noise and odors. Evaluators should be in good physical condition; those with colds, nasal congestion, or other conditions that may affect the evaluation results should not participate. Maintaining good personal hygiene and avoiding perfumes or strongly scented cosmetics can also negatively impact the sensory evaluation. When pouring samples, the volume of each sample should be consistent (approximately 30 mL). If there are significant differences in sample color, the evaluation glass should be wrapped with aluminum foil to prevent color from affecting the evaluation results. Scoring should be completed within three minutes of pouring the samples to prevent aroma loss due to prolonged exposure. The number of samples in the same evaluation group should be limited to five. If the number of samples is too large, multiple rounds of evaluation can be conducted in groups. A single assessment should ideally consist of no more than two rounds, with a 10-minute break in between. This process should be repeated three times to prevent sensory fatigue from causing inaccurate assessment results.

[0070] Table 1

[0071]

[0072] Table 1 shows that the rice wine prepared in Examples 1-3 exhibits reduced higher alcohol content and increased ester content, reduced bitter amino acid content, and improved aroma and palatability. After destructive testing, the turbidity was ≤1.0 NTU, indicating high clarity. Comparing Example 1 with Comparative Example 1, the rice wine prepared in Comparative Example 1 has a higher higher alcohol content and a lower volatile ester content, indicating that adding lactic acid bacteria koji during rice wine preparation can reduce the higher alcohol content and enhance the ester aroma. Comparing Example 1 with Comparative Example 2, directly adding lactic acid bacteria seed liquid for fermentation has a poor effect on reducing the higher alcohol content in rice wine. Comparing Example 1 with Comparative Example 3, the timing of lactic acid bacteria koji addition during rice wine preparation significantly affects the reduction of higher alcohol content and the increase of esters. Lactic acid bacteria koji must be added only in the early stages of rice wine fermentation to rapidly increase the lactic acid bacteria content in the pre-fermentation stage, effectively prolonging the pre-fermentation time and reducing the accumulation of higher alcohols. As shown in Example 1 and Comparative Example 4, a segmented enzymatic hydrolysis method was adopted in the post-fermentation stage of rice wine. First, aminopeptidase was added to increase the content of free hydrophobic amino acids. Simultaneously, the hydrophobic amino acids moved from the middle to the end of the peptide chain, thus reducing bitterness. Then, flavor protease was added to further enzymatically hydrolyze bitter peptides, thereby fully degrading them into amino acids and further reducing bitterness. This also helped increase the amino acid nitrogen content in the rice wine, improving its flavor. Compared to adding flavor protease and aminopeptidase simultaneously, adding them sequentially was more effective, presumably because simultaneous addition might lead to incomplete hydrolysis and low protein utilization. Comparing Example 1 and Comparative Example 5, the rice wine prepared in Comparative Example 5 had a higher content of bitter amino acids. This indicates that adding tannins to the rice wine liquid complexes with bitter amino acids, reducing the content of bitter amino acids such as lysine, leucine, and isoleucine, thus weakening bitterness.

[0073] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and are not intended to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can be made to the technical solutions of the present invention without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A method for preparing a low-bitterness rice wine, characterized in that, Includes the following steps: S1. Preparation of lactic acid bacteria koji; S2. Mix rice and water, then add liquefying enzyme and saccharifying enzyme in sequence to obtain saccharified liquid; add rice wine starter, yeast activation liquid and lactic acid bacteria starter to the saccharified liquid and ferment to obtain fermentation liquid; S3. Add enzyme preparation to fermentation broth for enzymatic hydrolysis, filter and sterilize to obtain clear rice wine liquid; S4. Add tannin solution to the clear rice wine liquid, stir and mix well, adsorb and filter to obtain rice wine; The enzymatic hydrolysis in step S3 specifically involves adding aminopeptidase to the fermentation broth and hydrolyzing it at 50-60℃ for 9-15 hours, followed by adding flavor protease and hydrolyzing it at 50-60℃ for 9-15 hours.

2. The method for preparing low-bitterness rice wine according to claim 1, characterized in that, The lactic acid bacteria starter was prepared using Lactococcus plantarum and Lactobacillus delbrueckii subsp. lactis.

3. The method for preparing low-bitterness rice wine according to claim 2, characterized in that, The preservation number of *Lactococcus plantarum* is GDMCC NO.1.1033; the preservation number of *Lactobacillus delbrueckii* subsp. *lactobacter* is GDMCC NO.1.3694.

4. The method for preparing low-bitterness rice wine according to claim 1, characterized in that, The amount of lactic acid bacteria starter added in step S2 is 1-3% based on the mass percentage of rice.

5. The method for preparing low-bitterness rice wine according to claim 1, characterized in that, The amount of aminopeptidase added is 1-3% by mass percentage of the fermentation broth, and the amount of flavor protease added is 1-3%.

6. The method for preparing low-bitterness rice wine according to claim 1, characterized in that, The concentration of the tannin solution is 0.8-1.2%.

7. The method for preparing low-bitterness rice wine according to claim 1, characterized in that, In step S4, bentonite and diatomaceous earth are used for adsorption, followed by filtration by pressure filtration and ultrafiltration.

8. A type of rice wine, characterized in that, It is prepared by the method for preparing low-bitter rice wine according to any one of claims 1 to 7.

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