Application of bacillus subtilis K6 in detoxification of corn germ meal

Abstract

The invention discloses bacillus subtilis K6 which is preserved in the China Center for Type Culture Collection, and the preservation number of the bacillus subtilis K6 is CCTCC (China Center for Type Culture Collection) NO: 20231096. The invention further discloses application of the bacillus subtilis K6 in detoxification of the corn germ meal, the bacillus subtilis K6 is used for fermenting and detoxifying polluted corn germ meal, the degradation rate of zearalenone in the corn germ meal reaches 95% or above, the degradation rate of aspergillus flavus B1 in the corn germ meal reaches 90% or above, toxic substances can be effectively degraded, the detoxification activity is high, and the bacillus subtilis K6 can be applied to detoxification of the corn germ meal. And the method has good tolerance to high temperature and low pH, has stable detoxification capability, and can remove recessive zearalenone toxin contained in the cake meal at the same time. The utilization value of the corn germ meal can be improved, the nutrient components of the corn germ meal can be improved, and a good technical foundation is laid for deep processing of the corn germ meal.

Description

Technical Field

[0001] The invention relates to the technical field of microbial application, and in particular to application of Bacillus subtilis K6 in detoxification of corn germ meal. Background Art

[0002] Corn germ meal is the main byproduct obtained after corn germ oil is extracted or pressed from corn germ. Except for the relatively low fat content, other nutrients in corn germ are retained in it. The crude protein content in corn germ meal is about 17-27%, which is 2-3 times the crude protein content of corn. At the same time, the amino acid composition and composition are similar to egg protein, which is consistent with the human protein standard recommended by FAO / WHO. Corn germ meal also contains 10-20% cellulose and 35-40% hemicellulose. It is rich in polysaccharides, can promote gastrointestinal motility, prevent and treat cardiovascular diseases, etc., and can be used as a good source of dietary fiber. In recent years, researchers have gradually paid attention to the deep processing and utilization of corn germ meal. Since corn is easily contaminated by fungal toxins, corn germ cakes and meal processed from corn germ often contain fungal toxins such as zearalenone and aflatoxin B1, which limits its application. At present, the methods for removing mycotoxins from corn germ meal can be divided into physical detoxification, chemical detoxification and biological detoxification. Physical detoxification methods include activated carbon adsorption, electron irradiation, negative pressure treatment, high temperature fire, etc. Although they have certain detoxification effects, they are non-targeted methods with low specificity and can cause a certain degree of damage to the nutrients in corn germ meal; chemical detoxification methods include ozone oxidation, oxidant oxidation, alkaline or acidic sulfite treatment, etc. Although they have good detoxification effects, some chemical reagents themselves have micro-toxicity, causing secondary pollution of corn germ meal, and easily produce reagent residues, which is difficult to use on a large scale. The biological detoxification method mainly uses microbial cells to adsorb toxins and enzymes or metabolites produced in metabolism, which promotes the change of the structure of zearalenone and degrades it into less toxic or non-toxic compounds, thereby achieving the purpose of detoxification. It has the characteristics of high efficiency, greenness and safety. Compared with liquid fermentation detoxification, solid fermentation has the advantages of easy growth of microorganisms, high enzyme activity, simple equipment structure, low investment and low energy consumption. It is an efficient, low-cost process with a wider variety of available strains and a simpler process. At present, scholars have made certain progress in studying the biological detoxification technology of corn and its by-products pollution. For example, patent No. CN201810964751.3 discloses Bacillus amyloliquefaciens D-1, which can efficiently degrade zearalenone. Its storage number is CCYCCM20232468. It can degrade the initial amount of zearalenone in coix seed from 90μg / g to 2.99μg / g, and the degradation rate reaches 96.68%. In addition, the degradation mechanism of the strain and the toxicity of the degradation products are also explored. Patent No. CN202410104098.9 discloses a strain of Bacillus subtilis DYF4-3 that can degrade multiple toxins. Its storage number is CCTCCNO: M20232187. It has high bile salt tolerance and can degrade multiple mycotoxins such as zearalenone and aflatoxin. The degradation rates of vomitoxin, zearalenone and aflatoxin reached 96.63%, 90.59% and 84.57%, respectively. However, these screened detoxified strains can only achieve a high detoxification rate in an in vitro liquid culture medium environment. The detoxification rate shown in the actual detoxification of cake meal is low and there is no perfect application example. In addition, whether the hidden toxins bound to the protein in the meal are removed has not been explored. At the same time, the strain should have good high temperature resistance and adapt to the lower pH value of corn germ meal (corn germ is a by-product of corn starch wet germ extraction, and its pH value is generally 3-4). Therefore, it is urgent to invent a biological detoxification method that can be practically and specifically applied to the mycotoxin in corn germ meal, which can effectively solve the problem of zearalenone and aflatoxin B in corn germ meal. 1 Free and latent bound fungal toxins; in addition, solid-state fermentation can improve the application value of corn germ meal and lay the foundation for the deep processing of corn germ meal. Summary of the invention

[0003] Technical problem to be solved: In view of the above technical problem, the purpose of the present invention is to disclose a Bacillus subtilis K6, which is preserved by China Center for Type Culture Collection with a preservation number of CCTCC NO: 20231096. The present invention also discloses an application of Bacillus subtilis K6 in corn germ meal detoxification. Bacillus subtilis K6 ferments and detoxifies contaminated corn germ meal, and the degradation rate of zearalenone in corn germ meal is more than 95%, and the degradation rate of Aspergillus flavus B is 1.5%, 2.5%, 3.5%, 4.5%, 5.5%, 6.5%, 7.5%, 8.5%, 9.5%, 10.5%, 11.5%, 12.5%, 13.5%, 14.5%, 15.5%, 16.5%, 17.5%, 18.5%, 19.5%, 20.5%, 21.5%, 22.5%, 23.5%, 24.5%, 25.5%, 26.5%, 27.5%, 28.5%, 29.5%, 30.5%, 31.5%, 32.5%, 33.5%, 34.5%, 35.5%, 36.5%, 37.5%, 38.5%, 39.5%, 40.5%, 41.5%, 42.5%, 43.5%, 44.5%, 45.5%, 46.5%, 47.5%, 48.5%, 49.5%, 50.5%, 51.5%, 52.5%, 53.5%, 54.5%, 55.5%, 56.5%, 57.5%, 58.5%, 59.5%, 5 1 The degradation rate is over 90%, and it can effectively degrade toxic substances, has high detoxification activity, and has good tolerance to higher temperatures and lower pH values. It has stable detoxification ability and can remove latent zearalenone toxins contained in the meal. It can not only improve the utilization value of corn germ meal, but also improve the nutritional components of corn germ meal, laying a good technical foundation for the deep processing of corn germ meal.

[0004] Technical solution: Application of Bacillus subtilis K6 in detoxification of corn germ meal. Furthermore, the deposit number of the Bacillus subtilis K6 is CCTCC NO: 20231096, and the deposit date is June 25, 2023. The strain has been deposited in the China Center for Type Culture Collection, and the deposit address is Wuhan University, Wuhan City, Hubei Province. Furthermore, the method for removing corn germ meal toxins from Bacillus subtilis K6 comprises the following steps: S1. Take corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. The crushed corn germ meal was placed in a container and sterilized at 121°C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, inoculated with Bacillus subtilis K6 bacterial solution, sterile water was added to the corn germ meal until the moisture content was 80-120%, to obtain a solid medium of corn germ meal; S4. Fermenting the corn germ meal solid medium at 32-47°C to obtain fermented corn germ meal; S5. The fermented corn germ meal is dried at 60° C. for 48 hours to obtain detoxified corn germ cake. Furthermore, the inoculation amount of the Bacillus subtilis K6 bacterial solution in step S3 is 4-12%. Furthermore, the preparation process of the Bacillus subtilis K6 bacterial solution in step S3 is as follows: S1. Slowly thaw the frozen Bacillus subtilis K6 strain on ice, then take a small amount of bacterial solution into LB solid medium, spread it and seal it, then invert it in a 37℃ incubator until colonies grow; S2. The colony was inoculated into LB liquid medium, and the seed solution of Bacillus subtilis K6 was obtained when the absorbance at 600 nm was 0.6-0.8; S3. The Bacillus subtilis K6 seed liquid is transferred to the fermentation medium for the second time at a 5% inoculation rate to obtain the Bacillus subtilis K6 bacterial liquid. Furthermore, the fermentation time in step S4 is 48-84 hours. Beneficial effects: 1. The Bacillus subtilis K6 of the present invention produces a variety of active substances during the fermentation process, which can accurately degrade zearalenone and aflatoxin B in corn germ meal 1 , has good targeting and specificity, and can destroy zearalenone and aflatoxin B 1 The molecular structure of the toxic lactone ring hydrolyzes the lactone bond in the ring structure to make zearalenone and aflatoxin B 1The toxicity is lost and converted into non-toxic or less toxic substances, which finally disappear through bacterial metabolism. At the same time, the active substances secreted by Bacillus subtilis K6 can also release latent zearalenone bound to the protein, thereby achieving the purpose of improving the detoxification effect. The total degradation rates can reach more than 95% and more than 90% respectively, with high detoxification activity and excellent detoxification effect. 2. The Bacillus subtilis K6 in the present invention has good tolerance to pH and temperature, can still grow and maintain activity under the conditions of pH 4-9 and temperature 25-60°C, and can adapt well to the environment and substrate characteristics. 3. After the invention adopts Bacillus subtilis K6 fermentation and detoxification, the nutritional components of corn germ meal are significantly improved, the crude protein content increases by 4.78%, and the total phenol content increases by 45.24%, which greatly improves the nutritional composition and functional characteristics of corn germ meal. 4. The present invention adopts Bacillus subtilis K6 solid-state fermentation detoxification, which belongs to the category of biological detoxification, has the characteristics of safety, convenience, green environmental protection, strong specificity, high detoxification efficiency, and has wide practicality; the corn germ meal detoxified by solid-state fermentation has comprehensive nutrition, improves the biological functional characteristics of the cake, reduces anti-nutritional components, and greatly improves bioavailability. The present invention provides new technical support for the deep processing and utilization of corn germ meal, and has broad market application prospects. DETAILED DESCRIPTION The present invention will be further described below in conjunction with embodiments, which are explanations of the present invention and are not limited to the following embodiments: Information on the strain used in the present invention: Bacillus subtilis K6 (Bacillus subtilis K6), the preservation number is CCTCC NO: 20231096, the preservation date is June 25, 2023, the preservation unit is China Center for Type Culture Collection, and the preservation address is Wuhan University, Wuhan City, Hubei Province. Example 1 Preparation technology of Bacillus subtilis K6 bacterial liquid S1. Slowly thaw the frozen Bacillus subtilis K6 strain on ice, then take a small amount of bacterial solution into LB solid medium, spread it and seal it, then invert it in a 37℃ incubator until colonies grow; S2. The colony was inoculated into 40 ml LB liquid medium, and the Bacillus subtilis K6 seed solution was obtained when the absorbance at 600 nm was 0.6-0.8; S3. The Bacillus subtilis K6 seed liquid was transferred to the fermentation medium for the second time at a 5% inoculation rate to obtain the Bacillus subtilis K6 bacterial liquid. Example 2 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 30g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 4% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 80% to obtain a solid fermentation medium for corn germ meal; S4. Fermenting the corn germ meal solid fermentation medium at 37° C. for 60 h to obtain fermented corn germ meal; S5. Dry the fermented corn germ meal at 60° C. for 48 hours to obtain detoxified corn germ cake. Example 3 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 6% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 80% to obtain a solid fermentation medium for corn germ meal; S4. The corn germ meal solid fermentation medium was fermented at 37°C for 60h to obtain; S5. Dry the fermented corn germ meal at 60° C. for 48 hours to obtain detoxified corn germ cake. Example 4 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 30g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 8% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 80% to obtain a corn germ meal solid fermentation medium; S4. fermenting the corn germ meal mixed solution at 37° C. for 60 h to obtain fermented corn germ meal; S5. The fermented corn germ meal is dried at 60° C. for 48 hours to obtain detoxified corn germ cake. Example 5 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 35g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 10% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 80% to obtain a solid fermentation medium for corn germ meal; S4. Fermenting the corn germ meal solid fermentation medium at 37° C. for 60 h to obtain fermented corn germ meal; S5. The fermented corn germ meal is dried at 60° C. for 48 hours to obtain detoxified corn germ cake. Example 6 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 35g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 12% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 80% to obtain a corn germ meal solid fermentation medium; S4. Fermenting the corn germ meal solid fermentation medium at 37° C. for 60 h to obtain fermented corn germ meal; S5. Dry the fermented corn germ meal at 60° C. for 48 hours to obtain detoxified corn germ cake. Example 7 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 45g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 8% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 100% to obtain a solid fermentation medium of corn germ meal; S4. Fermenting the corn germ meal solid fermentation medium at 37° C. for 60 h to obtain fermented corn germ meal; S5. Dry the fermented corn germ meal at 60° C. for 48 hours to obtain detoxified corn germ cake. Example 8 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 30g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 8% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 120% to obtain a solid fermentation medium for corn germ meal; S4. Fermenting the corn germ meal solid fermentation medium at 37° C. for 60 h to obtain fermented corn germ meal; S5. Dry the fermented corn germ meal at 60° C. for 48 hours to obtain detoxified corn germ cake. Example 9 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 30g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 8% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 80% to obtain a corn germ meal solid fermentation medium; S4. Fermenting the corn germ meal solid fermentation medium at 37° C. for 48 h to obtain fermented corn germ meal; S5. Dry the fermented corn germ meal at 60° C. for 48 hours to obtain detoxified corn germ cake. Example 10 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 35g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 8% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 80% to obtain a corn germ meal solid fermentation medium; S4. Fermenting the corn germ meal solid fermentation medium at 37° C. for 72 h to obtain fermented corn germ meal; S5. The fermented corn germ meal is dried at 60° C. for 48 hours to obtain detoxified corn germ cake. Embodiment 11 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 35g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 8% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 80% to obtain a corn germ meal solid fermentation medium; S4. Fermenting the corn germ meal solid fermentation medium at 37° C. for 84 h to obtain fermented corn germ meal; S5. Dry the fermented corn germ meal at 60° C. for 48 hours to obtain detoxified corn germ cake. Example 12 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 35g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 8% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 80% to obtain a corn germ meal solid fermentation medium; S4. Fermenting the corn germ meal solid fermentation medium at 32° C. for 60 h to obtain fermented corn germ meal; S5. Dry the fermented corn germ meal at 60° C. for 48 hours to obtain detoxified corn germ cake. Embodiment 13 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 40g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 8% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 80% to obtain a corn germ meal solid fermentation medium; S4. Fermenting the corn germ meal solid fermentation medium at 42° C. for 60 h to obtain fermented corn germ meal; S5. Dry the fermented corn germ meal at 60° C. for 48 hours to obtain detoxified corn germ cake. Embodiment 14 A method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take 35g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, 8% Bacillus subtilis K6 culture was inoculated and sterile water was added to a moisture content of 80% to obtain a corn germ meal solid fermentation medium; S4. Fermenting the corn germ meal solid fermentation medium at 47° C. for 60 h to obtain fermented corn germ meal; S5. Dry the fermented corn germ meal at 60° C. for 48 hours to obtain detoxified corn germ cake. Comparative Example 1 The difference between this comparative example and Example 11 is that 10% hydrogen peroxide is used for detoxification, as follows: S1. Take 35g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15g of crushed cake and treat it with 10% hydrogen peroxide at 80°C for 16 hours. The zearalenone content in the zearalenone solution decreased by 83.90%, and the residual amount of hidden zearalenone in the zearalenone solution was 386.88ppb. Comparative Example 2 The difference between this comparative example and Example 11 is that microwave heating is used for detoxification, as follows: S1. Take 35g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15g of the crushed cake and heat it in a microwave at 160℃ for 8min. Aflatoxin B 1 The content decreased by 54%. Performance Testing (1) Detoxification efficiency The toxins were extracted from the detoxified cake obtained in Example 2-14, blown dry with concentrated nitrogen, reconstituted with 1 ml of mobile phase, filtered with a 0.45 μL organic filter membrane, and detected by HPLC. The HPLC detection conditions were as follows: mobile phase (acetonitrile: water: methanol = 46:46:8), chromatographic column was 150 mm × 4.6 mm C18 column, flow rate was 1 mL / min, column temperature was 30°C, injection volume was 20 μL, excitation wavelength was 274 nm, and emission wavelength was 440 nm. Table 1 Detoxification efficiency of Examples 2-14 and Comparative Example 1 and content of hidden zearalenone after detoxification The total zearalenone content in corn germ meal is 2200ppb-2500ppb, of which the hidden zearalenone content is 570-630ppb. The hidden zearalenone in Examples 2-14 is completed in the detoxification step. As shown in Table 1, the detoxification efficiency of Example 11 is the highest, which is higher than the detoxification efficiency of Comparative Example 1, and the hidden zearalenone after detoxification is significantly lower than that of Comparative Example 1, indicating that Bacillus subtilis K6 can effectively degrade the zearalenone in corn germ meal, and can also remove the hidden zearalenone tightly bound to macromolecular substances such as protein and starch in corn germ meal, convert it into non-toxic or less toxic substances, and then remove it through metabolism, thereby achieving the purpose of efficient detoxification. (2) Aflatoxin B 1 content Aflatoxin B of Example 11 and Comparative Example 2 was detected according to GB / T 36858-2018 1 content. Aflatoxin B of Example 11 1 The removal rate of aflatoxin B in comparative example 2 was 91.47%. 1 The removal rate of Example 11 was 54.00%, and the removal rate of Example 11 was significantly higher than that of Comparative Example 2, indicating that Bacillus subtilis K6 can significantly remove aflatoxin B from corn germ meal. 1 . Comparative Example 3 The difference between this comparative example and Example 4 is that Bacillus subtilis K6 was not used for fermentation, and the details are as follows: S1. Take 30g corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. Weigh 15 g of crushed corn germ meal and place it in a 100 mL volumetric flask, sterilize it at 120 ° C for 20 min to obtain sterilized corn germ meal; S5. The sterilized corn germ meal is dried at 60° C. for 48 hours to obtain corn germ cake. Performance Testing (1) Determination of crude protein content The crude protein content of corn germ meal before and after fermentation was determined according to GB5009.5-2016 method. (2) Determination of total phenol content The Folin-Ciocalteu reagent colorimetric method was used. Take 0.2g corn germ meal, soak it twice with 10mL 70% methanol, take 0.2mL of the extract and add 1mL Folin phenol solution, then add 0.8mL 7.5% sodium carbonate solution, mix well and react at 25℃ in the dark for 1h, measure the absorbance at 765nm. The total phenol content was calculated using the gallic acid standard curve. Table 2 Crude protein content and total phenol content of Example 4 and Comparative Example 3 Crude protein content (%) Total phenol content (mg / g) Example 4 16.80 6.50 Comparative Example 2 21.67 11.87 As can be seen from Table 2, Comparative Example 3 did not use Bacillus subtilis K6 for fermentation, and its crude protein content and total phenol content were lower than those of Example 4. The crude protein content and total phenol content of Example 4 increased by 4.78% and 45.24%, respectively, indicating that Bacillus subtilis K6 can utilize the material components in corn germ meal to metabolize and produce protein through fermentation, and dissociate and release phenolic substances, thereby increasing the crude protein content and total phenol content in the fermented corn germ meal. The above is only a preferred embodiment of the present invention and does not limit the present invention in any form. Any technician familiar with the art can make many possible changes and modifications to the technical solution of the present invention by using the above disclosed methods and technical contents without departing from the spirit and technical solution of the present invention, or modify it into an equivalent embodiment of equivalent changes. Therefore, any simple modification, equivalent replacement, equivalent change and modification made to the above embodiments according to the technical essence of the present invention without departing from the content of the technical solution of the present invention still falls within the scope of protection of the technical solution of the present invention.

Claims

1. An application of Bacillus subtilis K6 in detoxification of corn germ meal.

2. The use of Bacillus subtilis K6 in corn germ meal detoxification according to claim 1, characterized in that: The preservation number of the Bacillus subtilis K6 is CCTCC NO: 20231096, and the preservation date is June 25, 2023. The strain has been deposited in the China Center for Type Culture Collection, and the preservation address is Wuhan University, Wuhan City, Hubei Province.

3. The use of Bacillus subtilis K6 in corn germ meal detoxification according to claim 1, characterized in that: The method for removing corn germ meal toxins by using Bacillus subtilis K6 comprises the following steps: S1. Take corn germ meal into a grinder, crush it through a 40-mesh sieve to obtain crushed corn germ meal; S2. The crushed corn germ meal was placed in a container and sterilized at 121°C for 20 min to obtain sterilized corn germ meal; S3. Under sterile conditions, inoculated with Bacillus subtilis K6 bacterial solution, sterile water was added to the corn germ meal until the moisture content was 80-120%, to obtain a solid medium of corn germ meal; S4. Fermenting the corn germ meal solid medium at 32-47°C to obtain fermented corn germ meal; S5. The fermented corn germ meal is dried at 60° C. for 48 hours to obtain detoxified corn germ cake.

4. The method for removing corn germ meal toxin by Bacillus subtilis K6 according to claim 3, characterized in that: In step S3, the inoculation amount of the Bacillus subtilis K6 bacterial solution is 4-12%.

5. The use of Bacillus subtilis K6 in corn germ meal detoxification according to claim 3, characterized in that: The preparation process of Bacillus subtilis K6 bacterial solution in step S3 is as follows: S1. Slowly thaw the frozen Bacillus subtilis K6 strain on ice, then take a small amount of bacterial solution into LB solid medium, spread it and seal it, then invert it in a 37℃ incubator until colonies grow; S2. The colony was inoculated into LB liquid medium, and the seed solution of Bacillus subtilis K6 was obtained when the absorbance at 600 nm was 0.6-0.8; S3. The Bacillus subtilis K6 seed liquid was transferred to the fermentation medium for the second time at a 5% inoculation rate to obtain the Bacillus subtilis K6 bacterial liquid.

6. The use of Bacillus subtilis K6 in corn germ meal detoxification according to claim 3, characterized in that: The fermentation time in step S4 is 48-84 hours.

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