Preparation method of pit mud with specific water activity

Abstract

The invention discloses a preparation method of pit mud with specific water activity, which is characterized in that 1% of xanthan gum, 0.6% of resistant starch and 0.2% of carboxymethyl cellulose sodium are added during preparation of the pit mud, so that the prepared pit mud has obvious sticky feeling, is wet, is uniform in texture, and can be used for preparing the pit mud with the specific water activity. The water content and the wall hanging stability of the pit mud are superior to those of traditional pit mud, the specific water activity of the pit mud is 0.984, and compared with the traditional pit mud, the water retention capacity and the caproic acid production capacity of the pit mud are remarkably improved.

Description

Technical Field

[0001] The invention relates to a method for preparing cellar mud with specific water activity for producing Luzhou-flavor liquor. The cellar mud is used for liquor cellar construction and maintenance to improve the quality of liquor produced by solid-state fermentation, and belongs to the field of biotechnology. Background Art

[0002] Luzhou-flavor liquor is fermented in mud cellars. There are a large number of beneficial microorganisms involved in fermentation in the mud. These microorganisms work together with the microorganisms in the fermented grains to produce the main aroma components and flavor substances of the liquor. The quality of the mud is closely related to the output and quality of Luzhou-flavor liquor. High water activity in the mud is conducive to the growth and metabolism of microorganisms, but too high a water content in the mud will make it difficult for the mud to hang on the wall or even collapse the cellar. Therefore, the water content of the mud is an important indicator in the evaluation standard of the mud for Luzhou-flavor liquor.

[0003] Definition of water activity: the ratio of water vapor partial pressure to the saturated vapor pressure of pure water at the same temperature under equilibrium conditions. Studies have found that each microorganism has a water activity range in which it can grow, and it cannot grow normally below a certain water activity. Currently, no research on the water activity of pit mud has been reported. Summary of the invention

[0004] In view of the deficiencies in the prior art, the present invention provides a method for preparing pit mud with a specific water activity, and solves the technical problem of effectively improving the stability of the pit mud structure and the effectiveness of microbial growth metabolism, while increasing the moisture content of the pit mud without affecting its molding and wall hanging performance. After adding a water retaining agent, the flowing water can be locked to maintain moisture for a long time, and the metabolic activity of the pit mud microorganisms can be enhanced, thereby improving the quality of the Luzhou-flavor original liquor.

[0005] The first technical solution provided by the present invention is a cellar mud, wherein 1% xanthan gum, 0.6% resistant starch and 0.2% sodium hydroxymethyl cellulose are added to the raw materials of the cellar mud.

[0006] In certain embodiments, the raw materials of the cellar mud contain loess, biochar, old cellar mud, vinasse, yellow water, cellar mud mother liquor, biogas liquid, wine tail, bean cake powder, corn flour, Luzhou-flavor Daqu powder, inorganic nutrient solution and caproic acid bacteria fermentation liquid.

[0007] In certain embodiments, the raw materials of the cellar mud contain, by weight, 40-45 parts of loess, 6-10 parts of biochar, 30-35 parts of high-quality old cellar mud, 2-4 parts of distiller's grains, 2-4 parts of yellow water, 2-5 parts of cellar mud mother liquor, 1-4 parts of biogas liquid, 1-4 parts of wine tails, 0.5-2 parts of bean cake powder, 0.5-2 parts of corn flour, 0.5-3.5 parts of Luzhou-flavor Daqu powder, 1-5 parts of inorganic nutrient solution, and 1-10 parts of caproic acid bacteria fermentation liquid.

[0008] In certain embodiments, the raw materials of the cellar mud contain, by weight, 45 parts of loess, 8 parts of biochar, 35 parts of high-quality old cellar mud, 4 parts of distiller's grains, 4 parts of yellow water, 5 parts of cellar mud mother liquor, 4 parts of biogas liquid, 4 parts of wine tailings, 2 parts of bean cake powder, 2 parts of corn flour, 3.5 parts of Luzhou-flavor Daqu powder, 5 parts of inorganic nutrient solution, and 10 parts of caproic acid bacteria fermentation liquid.

[0009] In certain embodiments, the caproic acid bacteria include but are not limited to Bacillus paranthracis LLMJ-JS-1, which has been deposited in the China General Microbiological Culture Collection Center (CGMCC) on October 31, 2024, with a deposit number of CGMCC No: 32446.

[0010] Among them, the Bacillus paraanthracis LLMJ-JS-1 was screened from the bottom mud of the fermentation pit of the Luzhou-flavor liquor of Shandong Lanling Wine Co., Ltd. Bacillus paraanthracis LLMJ-JS-1 has a strong acid production ability under anaerobic conditions, and the acid production is 10.9g / L. The colony morphology of Bacillus paranthracis LLMJ-JS-1: the colonies are light yellow, with irregular edges, dry and opaque surface. The fermentation performance of Bacillus paranthracis LLMJ-JS-1: the temperature is 34-39°C, 37°C is the best; it can tolerate 1%-2.5% ethanol, and the performance is best at 2%.

[0011] The second technical solution provided by the present invention is a method for preparing pit mud, which comprises the following steps:

[0012] a. Spread 40-45 parts of loess, 6-10 parts of biochar, 30-35 parts of high-quality old cellar mud, 2-4 parts of distiller's grains, 2-4 parts of yellow water, 2-5 parts of cellar mud mother liquor, 1-4 parts of biogas liquid, 1-4 parts of wine tail, 0.5-2 parts of bean cake powder, 0.5-2 parts of corn flour, 0.5-3.5 parts of Luzhou-flavor Daqu powder, 1-5 parts of inorganic nutrient solution, 1-10 parts of caproic acid bacteria fermentation liquid, 1% xanthan gum, 0.6% resistant starch, 0.2% sodium hydroxymethyl cellulose evenly on the mud mixing field. Before using the old cellar mud, pick out the bamboo sticks and other debris in the soil, add 10-15 parts of clean water to moisten the soil for one day;

[0013] b. The next day, use a rotary tiller to mix the mud evenly and transfer it to the fermentation tank;

[0014] c. After the fermentation tank is full, cover it with plastic sheeting for fermentation.

[0015] In certain embodiments, the preparation method of the hexanoic acid bacteria fermentation broth is as follows: Bacillus paraanthrax LLMJ-JS-1 is introduced into the fermentation system at an inoculation rate of 1%-10%, and anaerobically cultured at 33-39° C. for 4-7 days.

[0016] In certain embodiments, the fermentation system contains 0.5%-1% yeast extract, 0.5%-1% anhydrous sodium acetate, 0.02%-0.05% MgSO4, 0.02%-0.05% K2HPO4, and 0.05%-0.1% (NH4)2SO4.

[0017] In certain embodiments, in step c, the fermentation temperature shall not be lower than 20° C., and the anaerobic fermentation shall be carried out for 30-40 days.

[0018] The third technical approach is a method for maintaining a specific water activity of cellar mud, wherein the method comprises adding 1% xanthan gum, 0.6% resistant starch, and 0.2% sodium hydroxymethyl cellulose to the raw materials of the cellar mud to prepare the cellar mud, wherein the water activity of the cellar mud is 0.984.

[0019] The technical effects of the present invention are as follows:

[0020] The cellar mud prepared by adding 1% xanthan gum, 0.6% resistant starch and 0.2% sodium hydroxymethyl cellulose has obvious viscosity, is moist and has uniform texture, and has better water content and wall-hanging stability than traditional cellar mud; the cellar mud has a specific water activity of 0.984, and its water retention performance and caproic acid production capacity are significantly improved compared with traditional cellar mud.

[0021] Biomaterial Deposit

[0022] A strain of Bacillus paraanthracis LLMJ-JS-1, taxonomically named Bacillus paranthracis, was deposited in the China General Microbiological Culture Collection Center (CGMCC) on October 31, 2024. The deposit address is No. 3, Yard 1, Beichen West Road, Chaoyang District, Beijing, and the deposit number is CGMCC No: 32446. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 This is the relationship between the water activity of pit mud with different activities and the amount of caproic acid produced. DETAILED DESCRIPTION

[0024] The preferred embodiments of the present invention are described below. It should be understood that the embodiments are for better explaining the present invention and are not used to limit the present invention.

[0025] Test method:

[0026] 1. Method for determining the water activity of pit mud: Use a water activity measuring instrument for determination.

[0027] 2. Hexanoic acid detection method:

[0028] (1) Sample pretreatment: Weigh 10 g of pit mud sample into a 50 ml centrifuge tube, add 10 ml of ultrapure water, mix well, add 1 ml of internal standard and 10 ml of ether, vortex for 30 s, centrifuge at 5000 r / min for 2 min, filter the supernatant through a 0.22 um microporous membrane, and take 1 ml into a gas phase injection vial for gas chromatography analysis.

[0029] (2) Chromatographic conditions: Agilent 7820A gas chromatograph (Agilent Technologies, Inc.) was used, equipped with an Agilent DB-WAX UI chromatographic column (30m*0.25mm*0.5μm; Agilent Technologies, Inc.); the initial column temperature of the chromatographic analysis was set at 40°C, then increased to 80°C at a rate of 3°C / min, then increased to 200°C at a rate of 5°C / min, and maintained at this temperature for 30 minutes; the initial column pressure was set to 8psi, maintained for 5 minutes, then increased to 17psi at a rate of 3psi / min, and maintained at this pressure for 57 minutes. The detector used was a flame ionization detector (FID), and its temperature was maintained at 250°C; the injection port temperature was set to 250°C and the pressure was maintained at 8psi.

[0030] The raw materials used in the embodiment:

[0031] Liquid culture medium: 1% yeast extract, 0.5% anhydrous sodium acetate, 0.02% MgSO4, 0.04% K2HPO4, 0.05% (NH4)2SO4, sterilized at 121°C for 20 min. Add 2% ethanol and 1% dry heat sterilized CaCO3 before use.

[0032] Solid culture medium: Add 2% agar powder to liquid culture medium before sterilization.

[0033] The raw materials used in the culture medium were purchased from Sinopharm Chemical Reagent Co., Ltd.

[0034] Example 1 Preparation of caproic acid fermentation broth

[0035] 1. Screening and identification of caproic acid bacteria

[0036] 1.1 Strain screening

[0037] (1) Enrichment culture-preliminary screening: Take 10g of pit mud sample, place it in a sterile bag containing 90mL of sterile water, and shake it at 180r / min for 30min to make the pit mud sample evenly distributed in the sterile water. Place the shaken pit mud mixture in a constant temperature water bath at 85℃ for 10min to kill the bacteria. After the water bath, inoculate it into the liquid culture medium at a 10% inoculation rate, seal it with a sealing film, place it in an anaerobic culture bag containing an anaerobic gas production bag, and place it in a 37℃ incubator for enrichment culture for 4-7d, and enrich it three times in a row. After the enrichment culture is completed, perform a copper sulfate color development test on the enrichment solution, and select the culture solution with a darker blue-green color for the next step of screening and separation.

[0038] (2) Isolation and culture of microorganisms: The enriched solution with obvious color development is diluted in a gradient manner and then spread on a solid culture medium. The culture is then anaerobically cultured at 37°C for 4-7 days. A single colony is picked and purified by streaking more than 3 times.

[0039] (3) Rescreening of caproic acid-producing bacteria: The purified strains were inoculated into liquid culture medium and cultured anaerobically at 37°C for 4-7 days. The culture medium was extracted and quantitatively analyzed by gas chromatography.

[0040] 1.2 Strain identification

[0041] The strain with the highest hexanoic acid production was selected and inoculated into a pasteurized synthetic solid medium and cultured until colonies grew out. The morphological characteristics of the colonies were observed. The colonies of the strain were light yellow, with irregular edges, dry and opaque surfaces. The whole genome was sequenced, and after gene sequence alignment and combined with the construction of a phylogenetic tree, the strain was identified as Bacillus paranthracis and named LLMJ-JS-1.

[0042] 1.3 Strain preservation

[0043] This strain was deposited in the China General Microbiological Culture Collection Center on October 31, 2024, with the deposit number: CGMCC No: 32446.

[0044] 2. Preparation of caproic acid fermentation broth

[0045] Fermentation was performed using Bacillus paraanthrax LLMJ-JS-1, the fermentation medium formula was: 1% sodium acetate, 0.1% yeast extract, 0.05% urea, 0.04% dipotassium hydrogen phosphate, 0.02% magnesium sulfate, sterilized at 121°C for 30 minutes, and 2% ethanol was added after cooling. A 50L fermentation tank was used, the inoculation amount was 10%, and anaerobic culture was carried out at 35°C for 7 days to obtain caproic acid fermentation liquid.

[0046] Example 2

[0047] The preparation method of this embodiment comprises the following steps:

[0048] a. 45 parts of loess, 8 parts of biochar, 35 parts of high-quality old cellar mud, 4 parts of lees, 4 parts of yellow water, 5 parts of cellar mud mother liquor, 4 parts of biogas liquid, 4 parts of wine tail, 2 parts of bean cake powder, 2 parts of corn flour, 3.5 parts of Luzhou-flavor Daqu powder, 5 parts of inorganic nutrient solution, 10 parts of caproic acid bacteria fermentation liquid. On this basis, xanthan gum (0.6%-1%), resistant starch (0.4%-0.8%) and sodium hydroxymethyl cellulose (0.2%-0.6%) are added according to different formula proportions, as shown in Table 1. After mixing evenly, spread it on the mud mixing field. Before using the old cellar mud, the bamboo sticks and other debris in the soil need to be removed, and 13 parts of clean water should be added to moisten the soil for one day.

[0049] Table 1 Specific formula of adding water retaining agent to active pit mud

[0050]

[0051]

[0052] b. The next day, use a rotary tiller to mix the mud evenly and transfer it to the fermentation tank.

[0053] c. After the fermentation tank is full, cover it with plastic sheeting for fermentation. Use after 30 days.

[0054] After fermentation, the water activity of the pit mud with different recipes was measured using a water activity meter, and the caproic acid production of the pit mud with different recipes was measured using a gas chromatograph (Table 2).

[0055] Table 2 Water activity values ​​of pit mud with different formulas

[0056]

[0057] When xanthan gum, resistant starch and sodium hydroxymethyl cellulose were added to the pit mud separately, the effects on the water activity and caproic acid production in the pit mud were also different. Among them, xanthan gum was most effective in maintaining water activity and maintaining the pit mud hanging on the wall, while resistant starch was more helpful in increasing caproic acid production. Furthermore, by accurately mixing the three additives, the performance of the pit mud was optimized, thereby improving the overall quality of the pit mud and the efficiency of brewing.

[0058] Example 2

[0059] The invention discloses artificial pit mud for making Luzhou-flavor liquor with a specific water activity. The artificial pit mud is prepared by the following raw material proportions in parts by weight: 45 parts of loess, 8 parts of biochar, 35 parts of high-quality old pit mud, 4 parts of lees, 4 parts of yellow water, 5 parts of pit mud mother liquor, 4 parts of biogas liquid, 4 parts of liquor tail, 2 parts of bean cake powder, 2 parts of corn flour, 3.5 parts of Luzhou-flavor Daqu powder, 5 parts of inorganic nutrient solution, and 10 parts of caproic acid bacteria fermentation liquid.

[0060] The preparation method of this embodiment comprises the following steps:

[0061] a. Add 45 parts of loess, 8 parts of biochar, 35 parts of high-quality old cellar mud, 4 parts of distiller's grains, 4 parts of yellow water, 5 parts of cellar mud mother liquor, 4 parts of biogas liquid, 4 parts of wine tail, 2 parts of bean cake powder, 2 parts of corn flour, 3.5 parts of Luzhou-flavor Daqu powder, 5 parts of inorganic nutrient solution, and 10 parts of caproic acid bacteria fermentation liquid. On this basis, add xanthan gum (0.6%-1%), resistant starch (0.4%-0.8%) and sodium hydroxymethyl cellulose (0.2%-0.6%) according to different formula proportions, as shown in Table 3. After mixing evenly, spread it on the mud mixing field. Before using the old cellar mud, the bamboo sticks and other debris in the soil need to be removed, and 13 parts of clean water should be added to moisten the soil for one day;

[0062] Table 3 Specific formula of adding water retaining agent to active pit mud

[0063]

[0064] b. The next day, use a rotary tiller to mix the mud evenly and transfer it to the fermentation tank.

[0065] c. After the fermentation tank is full, cover it with plastic sheeting for fermentation. Use after 30 days.

[0066] After fermentation, the water activity of the pit mud with different recipes was measured using a water activity meter, and the caproic acid production of the pit mud with different recipes was measured using a gas chromatograph (Table 4).

[0067] Table 4 Water activity values ​​of pit mud with different formulas

[0068]

[0069]

[0070] From the water activity values ​​of pit mud with different formulas in Table 4, it can be seen that there is a certain correlation between water activity and caproic acid production. The caproic acid production of pit mud under different water activities is further analyzed ( Figure 1 ), and found that when the water activity of the active pit mud was 0.984, that is, under the formula of 1% xanthan gum, 0.6% resistant starch, and 0.2% sodium hydroxymethyl cellulose, the pit mud produced the highest amount of caproic acid, reaching 839 mg / kg. This result shows that although xanthan gum, resistant starch, and sodium hydroxymethyl cellulose have certain water-retaining components, inappropriate addition will cause the water activity to be too high or too low, and affect the synthesis of caproic acid. In addition, this study also found that there is no obvious linear relationship between the addition amount of xanthan gum, resistant starch, and sodium hydroxymethyl cellulose and the water activity of the pit mud and the amount of caproic acid produced by the pit mud.

[0071] Although the present invention has been disclosed as above in the form of a preferred embodiment, it is not intended to limit the present invention. Anyone familiar with this technology can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the scope of protection of the present invention should be based on the definition of the claims.

Claims

1. A pit mud, characterized in that: 1% xanthan gum, 0.6% resistant starch and 0.2% sodium hydroxymethylcellulose are added to the raw materials of pit mud.

2. The pit mud according to claim 1, characterized in that: The raw materials of the cellar mud include loess, biochar, old cellar mud, vinasse, yellow water, cellar mud mother liquor, biogas liquid, wine tail, bean cake powder, corn flour, Luzhou-flavor Daqu powder, inorganic nutrient solution and caproic acid bacteria fermentation liquid.

3. The pit mud according to claim 1 or 2, characterized in that: By weight, the raw materials of the cellar mud contain 40-45 parts of loess, 6-10 parts of biochar, 30-35 parts of high-quality old cellar mud, 2-4 parts of wine lees, 2-4 parts of yellow water, 2-5 parts of cellar mud mother liquor, 1-4 parts of biogas liquid, 1-4 parts of wine tail, 0.5-2 parts of bean cake powder, 0.5-2 parts of corn flour, 0.5-3.5 parts of Luzhou-flavor Daqu powder, 1-5 parts of inorganic nutrient solution, and 1-10 parts of caproic acid bacteria fermentation liquid.

4. The pit mud according to claim 2, characterized in that: In terms of weight, the raw materials of the cellar mud contain 45 parts of loess, 8 parts of biochar, 35 parts of high-quality old cellar mud, 4 parts of distiller's grains, 4 parts of yellow water, 5 parts of cellar mud mother liquor, 4 parts of biogas liquid, 4 parts of wine tail, 2 parts of bean cake powder, 2 parts of corn flour, 3.5 parts of Luzhou-flavor Daqu powder, 5 parts of inorganic nutrient solution, and 10 parts of caproic acid bacteria fermentation liquid.

5. The pit mud according to any one of claims 2 to 4, characterized in that: The caproic acid bacteria include but are not limited to Bacillus paranthracis LLMJ-JS-1. The Bacillus paranthracis LLMJ-JS-1 has been deposited in the China General Microbiological Culture Collection Center (CGMCC) on October 31, 2024, with a deposit number of CGMCC No: 32446.

6. A method for preparing pit mud, characterized in that: The method comprises the following steps: a. Spread 40-45 parts of loess, 6-10 parts of biochar, 30-35 parts of high-quality old cellar mud, 2-4 parts of distiller's grains, 2-4 parts of yellow water, 2-5 parts of cellar mud mother liquor, 1-4 parts of biogas liquid, 1-4 parts of wine tail, 0.5-2 parts of bean cake powder, 0.5-2 parts of corn flour, 0.5-3.5 parts of Luzhou-flavor Daqu powder, 1-5 parts of inorganic nutrient solution, 1-10 parts of caproic acid bacteria fermentation liquid, 1% xanthan gum, 0.6% resistant starch, 0.2% sodium hydroxymethyl cellulose evenly on the mud mixing field. Before using the old cellar mud, pick out the bamboo sticks and other debris in the soil, add 10-15 parts of clean water to moisten the soil for one day; b. The next day, use a rotary tiller to mix the mud evenly and transfer it to the fermentation tank; c. After the fermentation tank is full, cover it with plastic sheeting for fermentation.

7. The preparation method according to claim 6, characterized in that: The preparation method of the caproic acid bacteria fermentation liquid is as follows: Bacillus paraanthrax LLMJ-JS-1 is introduced into the fermentation system at an inoculation amount of 1%-10%, and anaerobically cultured at 33-39° C. for 4-7 days.

8. The preparation method according to claim 7, characterized in that: The fermentation system contains 0.5%-1% yeast extract, 0.5%-1% anhydrous sodium acetate, 0.02%-0.05% MgSO4, 0.02%-0.05% K2HPO4, and 0.05%-0.1% (NH4)2SO4.

9. The preparation method according to claim 6, characterized in that: In step c, the fermentation temperature shall not be lower than 20°C, and the anaerobic fermentation shall be carried out for 30-40 days.

10. A method for maintaining a specific water activity of pit mud, characterized in that: The method comprises the following steps: adding 1% xanthan gum, 0.6% resistant starch and 0.2% sodium hydroxymethyl cellulose to the raw material of cellar mud to prepare the cellar mud, wherein the water activity of the cellar mud is 0.984.

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