Edible fungus fermented black beans based on co-fermentation of Mucor and Aspergillus and its koji making process

Abstract

The invention discloses an edible fungus fermented black beans based on co-fermentation of Mucor and Aspergillus and its koji making process, belonging to the technical field of food fermentation. The edible fungus fermented black beans are made from edible fungi or their scraps and beans as raw materials, and are obtained after fermentation of mixed fungus spore liquid containing Mucor and Aspergillus, wherein the number ratio of Mucor and Aspergillus is (1-6): (1-6). The protease activity, β-glucosidase activity, amino acid nitrogen content, soybean isoflavone content and antioxidant activity of the edible fungus fermented black beans provided by the present invention are significantly improved, and have excellent nutrient content, biological activity and health care efficacy. The koji making process of the present invention is simple to operate, low in cost, short in fermentation time, and has broad application prospects.

Description

Technical Field

[0001] The invention belongs to the technical field of food fermentation, and in particular relates to edible fungus fermented black beans based on co-fermentation of Mucor and Aspergillus and a koji making process thereof. Background Art

[0002] Douchi is one of the four traditional fermented soybean products in my country. It has many health functions such as anti-oxidation, thrombolysis, and blood pressure reduction. Generally, it can be divided into three types according to the different microorganisms involved in the fermentation: Aspergillus type, Mucor type, and bacterial type. Among them, Mucor type Douchi and Aspergillus type Douchi are unique traditional condiments in Sichuan and Chongqing and Hunan respectively. The koji making of Douchi is a key stage that determines the quality and function of Douchi. In this process, the enzyme system produced by Mucor / Aspergillus plays an important role in the accumulation of nutrients and the conversion of components. For example, protease can decompose the protein in soybeans into peptones, peptides, and free amino acids, which is more conducive to human absorption; β-glucosidase can convert glycoside-type soybean isoflavones in soybeans into aglycone-type soybean isoflavones with higher functional activity, so that Douchi has better health functions than soybeans.

[0003] The Chinese patent application with publication number CN 110169539 A discloses a method for preparing soybean and black bean mixed fermented black beans. The method uses soybean and black bean mixed in a certain proportion as raw materials, soaks and cooks, first inoculates Mucor for fermentation, then adds salt (salt content 10% to 13%) and other auxiliary materials to inoculate Aspergillus oryzae for continued fermentation, and prepares soybean and black bean mixed fermented black beans. The fermented black beans prepared by this method have better appearance and flavor than traditional soybean fermented black beans, and are more popular with people. However, on the one hand, this method has the disadvantages of complex operation and long fermentation time (up to 6 months or more); on the other hand, the addition of high salt will also inhibit the growth of Aspergillus oryzae, reduce the activity of β-glucosidase, hinder the biotransformation of soybean isoflavones, reduce the nutritional and health value of fermented black beans, and also increase the risk of cardiovascular disease, which is not conducive to human health. Therefore, a method for making koji with simpler operation and shorter fermentation time is needed to develop fermented black beans with better nutrient content, biological activity and health effects.

[0004] Edible fungi are a kind of edible large fungi with high protein, low fat, rich in polysaccharides, flavonoids, active peptides, nucleosides and other active substances. They have multiple health functions such as anti-cancer, anti-diabetes, and anti-obesity. According to statistics, in 2018, the total output of edible fungi in China reached 40 million tons, with a total output value of 274.178 billion yuan, accounting for more than 75% of the world's total output, while the output of edible fungi in Sichuan Province reached 2.2 million tons, with an output value of 12.6 billion yuan, ranking seventh in the country. At present, 80% of edible fungi in Sichuan Province are mainly used for fresh sales, and 20% are used for deep processing. In the above sales and processing processes, a large amount of edible fungi waste will be generated, including mushroom feet, mushroom slices, mushroom stems, deformed mushrooms, and fungi entity residues after functional component extraction, accounting for about 30% of the total output. Because the appearance and flavor of the waste are relatively poor and difficult to be accepted by consumers, they are often directly thrown away as animal feed or waste, which not only causes serious waste of edible fungi resources, but also brings certain environmental pollution problems.

[0005] So far, there are few research reports on the preparation method of edible fungus fermented black beans. Among them, the Chinese patent application with publication number CN103053952A discloses a process for producing fermented black beans by edible fungus fermentation method, in which edible fungus is used as the only strain to ferment soybeans to make fermented black beans, and the process flow is: soybean washing, soaking → bottling → sterilization → cooling → inoculation → spawning culture → making finished products. Because edible fungi are rich in amino acids, the fermented products taste delicious. However, on the one hand, in this process, soybeans are mainly used as the fermentation nutrient matrix of edible fungi to provide nutrition for edible fungi mycelium, while ignoring the important nutritional active ingredient of soybeans itself-the biotransformation of soybean isoflavones, so that the obtained fermented black beans have low amino acid nitrogen content, insufficient fermented black bean flavor, and low antioxidant activity, which is very different from traditional fermented black beans; on the other hand, this process can only use fresh and complete edible fungi as strains, and cannot effectively use edible fungi waste such as mushroom feet, mushroom slices, mushroom stems, deformed mushrooms and fungi entity residues after functional component extraction, resulting in resource waste.

[0006] In summary, in order to overcome the shortcomings of the existing technology, it is urgent to develop a koji making method with simpler operation, shorter fermentation time and more resource-saving to prepare fermented black beans with better nutrient content, biological activity and health benefits. Summary of the invention

[0007] The purpose of the present invention is to provide fermented black beans with excellent nutrient content, biological activity and health-care effects, as well as a koji-making process with simple operation, low cost and short fermentation time.

[0008] The invention provides an edible fungus fermented black beans, which is prepared by fermenting edible fungi or their waste materials and beans with a mixed fungus spore liquid containing Mucor and Aspergillus, wherein the quantity ratio of Mucor to Aspergillus in the mixed fungus spore liquid is (1-6):(1-6).

[0009] Furthermore, the volume mass ratio of the mixed bacterial spore liquid to the raw material is 2%-12% mL / g, preferably 6% mL / g;

[0010] And / or, the concentration of the mixed bacterial spore solution is 1×10 6 -1×10 8 CFU / mL, preferably 1×10 7 CFU / mL.

[0011] Further, the Mucor is Mucor racemosus, and the Aspergillus is Aspergillus oryzae;

[0012] And / or, the ratio of the number of Mucor and Aspergillus is 1:1.

[0013] Furthermore, in the raw materials, the mass ratio of edible fungi or their residues to beans is (5-35):100, preferably (20-21):100.

[0014] Further, the fermentation time is 24-96 hours, preferably 63 hours;

[0015] The fermentation temperature is 20-30°C, preferably 28°C;

[0016] The relative humidity of the fermentation is 70%-90%, preferably 80%.

[0017] Furthermore, the beans are one or a mixture of two or more of soybeans, mung beans, and peas, preferably soybeans.

[0018] Furthermore, the edible fungi waste materials include one or a mixture of two or more of the following: mushroom feet, mushroom slices, mushroom stems, deformed mushrooms, and fungi residues after functional components are extracted from edible fungi;

[0019] And / or, the edible fungi are one or both of Pleurotus eryngii and Pleurotus ostreatus.

[0020] The invention also provides a koji making process for the edible fungus fermented black beans, which comprises the following steps: soaking beans in water, draining, uniformly mixing with edible fungi or their wastes, high pressure steaming, cooling, inoculating mixed fungus spore liquid, and fermenting to obtain the edible fungus fermented black beans.

[0021] Furthermore, the mass volume ratio of the beans to water is 1:(2-5) g / mL, preferably 1:3 g / mL, the soaking temperature is 24-45° C., preferably 35° C., and the soaking time is 2-5 h, preferably 4 h.

[0022] Furthermore, the high-pressure cooking temperature is 110-130° C., preferably 121° C., the pressure is 0.05-0.2 MPa, preferably 0.1 MPa, and the time is 15-35 min, preferably 25 min.

[0023] Compared with the prior art, the present invention has achieved the following beneficial effects:

[0024] (1) The present invention is the first to use edible fungus waste and soybeans as a common substrate for fermentation, effectively solving the problems of waste of edible fungus waste resources and low added value;

[0025] (2) The present invention adopts mixed fungi of Mucor racemosus and Aspergillus oryzae to make koji and ferment, which shortens the koji making time (≤3 days), improves the fermentation efficiency, and solves the problem that the traditional process has a long natural fermentation time and is easily contaminated by miscellaneous bacteria;

[0026] (3) The protease activity, β-glucosidase activity, amino acid nitrogen content, soybean isoflavone content and antioxidant activity of the mixed fermented edible fungus Douchiqu of the present invention are significantly improved, and it has excellent nutrient content, biological activity and health care effect.

[0027] (4) The protease activity, β-glucosidase activity and amino acid nitrogen content of the mixed fermented edible fungi fermented black bean koji of the present invention are greater than the sum of the fermented black bean koji of Mucor racemosus edible fungi of comparative example 1 and the fermented black bean koji of Aspergillus oryzae edible fungi of comparative example 2, indicating that the preparation process of the present invention has played a synergistic effect.

[0028] In summary, the present invention uses edible fungus scraps and soybeans as main raw materials, inoculates racemosus and Aspergillus oryzae for mixed fermentation to prepare an edible fungus fermented bean curd, and the fermentation process of the present invention has the advantages of simple operation, low cost, and short fermentation time. The fermented bean curd prepared by the present invention has a strong fermented bean curd aroma, outstanding edible fungus flavor, high nutritional value (amino acid nitrogen, soybean isoflavones), strong antioxidant activity, and high industrial application value. The present invention solves the problem of waste of edible fungus scraps resources, broadens the types of edible fungus fermented bean curd products, and meets the increasing demand of residents for health functional foods.

[0029] Obviously, according to the above contents of the present invention, in accordance with common technical knowledge and customary means in the art, without departing from the above basic technical ideas of the present invention, other various forms of modification, replacement or change may be made.

[0030] The above contents of the present invention are further described in detail below through specific implementation methods in the form of embodiments. However, this should not be understood as the scope of the above subject matter of the present invention being limited to the following examples. All technologies realized based on the above contents of the present invention belong to the scope of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 .Effect of inoculation amount on neutral protease activity and β-glucosidase activity of edible fungus Douchi.

[0032] Figure 2 .Effect of the ratio of Aspergillus oryzae to Mucor racemosus on the neutral protease activity and β-glucosidase activity of the edible fungus Douchi.

[0033] Figure 3 .Effect of the amount of dry powder of edible fungi waste added on the neutral protease activity and β-glucosidase activity of edible fungi fermented black beans.

[0034] Figure 4 .Effect of fermentation time on neutral protease activity and β-glucosidase activity of edible fungus Douchi. DETAILED DESCRIPTION

[0035] Unless otherwise specified, the methods used in the present invention are conventional methods in the art. Unless otherwise specified, the reagents and equipment used in the present invention can be obtained by purchasing commercial products.

[0036] Source of raw materials: Soybeans were harvested in 2021 and purchased from Heilongjiang Changhe Agricultural Products Processing Co., Ltd. Edible fungus waste powder (60 mesh) was from Chengdu Huiguyuan Biotechnology Co., Ltd., which was composed of deformed mushrooms of Pleurotus eryngii and Pleurotus ostreatus. Mucor racemosus, Actinomucor elegans, Mucorwutungkiao, and Aspergillus oryzae 41736 were purchased from the China Industrial Microbiological Collection Center (CICC).

[0037] Embodiment 1: Koji making process of mixed bacteria fermented edible fungus fermented black beans of the present invention

[0038] Weigh 100g of soybeans without mildew and with full grains, wash the soybeans with clean water, add 300mL of distilled water at a material-liquid ratio of 1:3 (m:V), and soak in a 35℃ constant temperature water bath for 4h (more than 90% of the beans are wrinkle-free). Drain the soaked soybeans and mix them with 21% (21g) of edible fungus waste dry powder, and steam them in a 121℃ 0.1MPa high pressure sterilizer for 25min. When cooled to room temperature, transfer the mixture to a sterile culture dish in an ultra-clean workbench in 20g portions, and quickly inoculate 1.2mL of mixed bacterial spore solution (concentration: 1×10 7 CFU / mL, wherein the ratio of Aspergillus oryzae 41736 to Mucor racemosus is 1:1), and finally fermented at a constant temperature (28°C) for 63 hours in an incubator with a relative humidity of 80% to obtain mixed fermented edible fungus Douchi Koji.

[0039] The neutral protease activity of the obtained fermented black bean curd sample was determined by referring to the SB / T 23527-2009 method and was 792.03±9.76U / g. The β-glucosidase activity was determined by the p-nitrophenol colorimetric method and was 1162.00±35.95U / g. The amino acid nitrogen content was determined by formaldehyde titration and was 0.80±0.01%. The aglycone-type soy isoflavone content was determined by HPLC and was 512.32±6.45mg / kg. The antioxidant activity was evaluated by DPPH scavenging ability assay and the DPPH value was 472.72±2.83μmolTrolox / g (as shown in Table 1).

[0040] The following is the koji making process of the control fermented black beans.

[0041] Comparative Example 1: Koji making process of edible fungus fermented black beans with Mucor racemosus

[0042] Weigh 100g of soybeans without mildew and with full grains, wash the soybeans with clean water, add 300mL of distilled water at a material-liquid ratio of 1:3 (m:V), and soak in a 35℃ constant temperature water bath for 4h (more than 90% of the beans are wrinkle-free). Drain the soaked soybeans and mix them with 21% (21g) of edible fungus waste powder, and steam them in a 121℃ 0.1MPa autoclave for 25min. When cooled to room temperature, transfer the mixture to a sterile culture dish in a clean bench in portions of 20g each, and quickly inoculate 1.2mL of Mucor racemosus spore solution (concentration: 1×10 7 CFU / mL), and finally fermented at a constant temperature (28°C) for 63 h in an incubator with a relative humidity of 80% to obtain the edible fungus Mucor racemosus in fermented black beans (ZS).

[0043] According to the testing method of Example 1, the protease activity of the edible fungus Douchiqu of Mucor racemosus was measured to be 106.77±4.74U / g, the β-glucosidase activity was 781.50±20.92U / g, the amino acid nitrogen content was 0.16±0.03%, the aglycone-type soy isoflavone content was 326.38±2.95mg / kg, and the DPPH scavenging ability was 161.06±2.83μmol Trolox / g, which were reduced by 86.52%, 32.75%, 80%, 36.30% and 65.93% respectively compared with the edible fungus Douchiqu of Example 1 (as shown in Table 1).

[0044] Comparative Example 2: Koji making process of Aspergillus oryzae edible fungus fermented black beans

[0045] Weigh 100g of soybeans without mildew and with full grains, wash the soybeans with clean water, add 300mL of distilled water at a solid-liquid ratio of 1:3 (m:V), and soak in a 35℃ constant temperature water bath for 4h (more than 90% of the beans are wrinkle-free). Drain the soaked soybeans and mix them with 21% (21g) of edible fungus waste powder, and steam them in a 121℃ 0.1MPa autoclave for 25min. When cooled to room temperature, transfer the mixture to a sterile culture dish in a clean bench in 20g portions, and quickly inoculate 1.2mL of Aspergillus oryzae 41736 spore solution (concentration: 1×10 7 CFU / mL), and finally fermented at a constant temperature (28°C) for 63h in an incubator with a relative humidity of 80% to obtain the Aspergillus oryzae edible fungus fermented with black beans.

[0046] According to the test method of Example 1, the protease activity of the Aspergillus oryzae edible fungus Douchiqu was measured to be 564.29±12.23U / g, the β-glucosidase activity was 266.81±8.65U / g, the amino acid nitrogen content was 0.53±0.04%, the aglycone-type soybean isoflavone content was 313.57±1.47mg / kg, and the DPPH scavenging ability was 376.06±2.08μmol Trolox / g, which were respectively reduced by 28.75%, 77.04%, 33.75%, 38.79% and 20.45% (as shown in Table 1) compared with the edible fungus Douchiqu of Example 1.

[0047] Comparative Example 3: Koji making process of fermented black beans without adding edible fungi

[0048] Weigh 100g of soybeans without mildew and with full grains, wash the soybeans with clean water, add 300mL of distilled water at a solid-liquid ratio of 1:3 (m:V), and soak in a 35℃ constant temperature water bath for 4h (more than 90% of the beans are wrinkle-free). Drain the soaked soybeans and steam them directly in a 0.1MPa autoclave at 121℃ for 25min without adding edible fungus powder. When cooled to room temperature, transfer 20g of soybeans to sterile culture dishes in a clean bench, and quickly inoculate 1.2mL of mixed bacterial spore solution (concentration: 1×10 7 CFU / mL, wherein the ratio of Aspergillus oryzae 41736 to Mucor racemosus was 1:1), and finally the mixture was fermented at a constant temperature (28°C) for 63 h in an incubator with a relative humidity of 80% to obtain edible fungus-free fermented black bean curd (ZM).

[0049] According to the testing method of Example 1, the protease activity of the edible fungus-free fermented black bean curd was measured to be 587.03±17.31U / g, the β-glucosidase activity was 524.78±25.02U / g, the amino acid nitrogen content was 0.25±0.02%, the aglycone-type soy isoflavone content was 339.84±3.94mg / kg, and the DPPH scavenging ability was 418.28±2.36μmol Trolox / g, which were respectively reduced by 25.88%, 54.84%, 68.75%, 33.67% and 11.52% compared with the edible fungus fermented black bean curd of Example 1 (as shown in Table 1).

[0050] Comparative Example 4: Production process of unfermented edible fungus soybeans

[0051] Weigh 100g of soybeans without mildew and with full grains, wash the soybeans with clean water, add 300mL of distilled water at a material-liquid ratio of 1:3 (m:V), and soak in a 35℃ constant temperature water bath for 4h (more than 90% of the beans are wrinkle-free). Drain the soaked soybeans and mix them with 21% (21g) of edible fungus waste dry powder, steam them in a 121℃ 0.1MPa high pressure sterilizer for 25min to obtain unfermented edible fungus soybeans (CN).

[0052] According to the test method of Example 1, it was measured that the unfermented edible fungus soybean had no protease activity and β-glucosidase activity, the amino acid nitrogen content was 0.04±0.02, the aglycone type soybean isoflavone content was 37.88±0.08 mg / kg, of which daidzein was 3.86±0.17 mg / kg, and the DPPH scavenging ability was 113.83±4.76 μmol Trolox / g, which were respectively reduced by 95%, 92.61% and 75.92% compared with the edible fungus fermented black bean koji in Example 1 (as shown in Table 1).

[0053] Table 1. Comparison of enzyme activity, active ingredients and antioxidant activity of the products obtained in the examples and comparative examples

[0054]

[0055]

[0056] As can be seen from Table 1, compared with Comparative Examples 1-4, the protease activity, β-glucosidase activity, amino acid nitrogen content, soy isoflavone content and antioxidant activity of the edible fungus Douchiqu prepared by co-fermentation of Aspergillus oryzae and Mucor racemosus prepared in Example 1 of the present invention are significantly improved. Specifically, compared with the fermented bean curd with Mucor racemosus edible fungus of comparative example 1, the fermented bean curd with Aspergillus oryzae edible fungus of comparative example 2, and the fermented bean curd without adding edible fungi of comparative example 3, the protease activity of the mixed bacteria fermented edible fungus fermented bean curd in Example 1 of the present invention was respectively increased by 641.81%, 213.30% and 34.92%, the β-glucosidase activity was respectively increased by 48.69%, 335.52% and 121.43%, the amino acid nitrogen content was respectively increased by 400.00%, 50.94% and 220.00%, the soy isoflavone content was respectively increased by 56.97%, 63.38% and 50.75%, and the DPPH scavenging ability was respectively increased by 193.51%, 25.70% and 13.02%.

[0057] In addition, it can be seen from Table 1 that the protease activity, β-glucosidase activity and amino acid nitrogen content of the mixed fermented edible fungi fermented black bean koji in Example 1 of the present invention are greater than the sum of the edible fungi fermented black bean koji of Mucor racemosus 1 and the edible fungi fermented black bean koji of Aspergillus oryzae 2 of Comparative Example 2, indicating that the process based on the co-fermentation of Aspergillus oryzae and Mucor racemosus in Example 1 of the present invention has played a synergistic role.

[0058] The above experiments show that the protease activity, β-glucosidase activity, amino acid nitrogen content, soybean isoflavone content and antioxidant activity of the mixed fermented edible fungus Douchiqu of the present invention are significantly improved, and it has excellent nutrient content, biological activity and health care effect.

[0059] The beneficial effects of the present invention are described below through test examples.

[0060] Experimental Example 1: Optimization of fermentation process of edible fungi and fermented black beans and koji and analysis of antioxidant activity

[0061] 1. Single factor experiment on preparation technology of fermented black bean curd

[0062] Select soybeans that are free of mold and have full grains, and wash them with clean water. Then add distilled water at a ratio of 1:3 (m:V) between soybeans and water, and soak at a constant temperature of 35°C for 4 hours (more than 90% of the beans are wrinkle-free). Drain the soaked soybeans and mix them with a certain amount of edible fungus waste powder, and steam them in a 0.1MPa high-pressure sterilizer at 121°C for 30 minutes. When cooled to room temperature, transfer the mixture to a sterile culture dish in an ultra-clean workbench in 20g portions, and quickly inoculate a certain amount of mixed spore liquid of Aspergillus oryzae 41736 and Mucor racemosus (concentration: 1×10 7CFU / mL), and finally fermented at a constant temperature (28°C) in an incubator with a relative humidity of 80% for a certain period of time to obtain fermented black bean curd.

[0063] According to the above preparation method, the effects of the following four factors on the neutral protease activity and β-glucosidase activity of Douchiqu were studied respectively. The preparation method of Douchiqu is as follows, and each treatment is repeated 3 times:

[0064] (1) The fermented bean curd making process of Example 1 was followed, except that the inoculation amount of the mixed spore solution was replaced with 2%, 4%, 6%, 8%, 10% or 12% (mL / g).

[0065] (2) Referring to the fermented black bean koji making process of Example 1, the only difference is that the quantitative ratio of Aspergillus oryzae to Mucor racemosus in the mixed spore liquid is modified to 6:1, 4:1, 2:1, 1:1, 1:2, 1:4 or 1:6 (v / v).

[0066] (3) Referring to the fermented black bean koji making process of Example 1, the only difference is that the amount of edible fungus waste dry powder added is modified to 0, 5%, 10%, 15%, 20%, 25%, 30% or 35% (g / g) of the mass of soybeans;

[0067] (4) Referring to the koji making process of fermented black beans in Example 1, the only difference is that the fermentation time is modified to 24h, 36h, 48h, 54h, 68h, 72h or 96h.

[0068] The neutral protease activity of the obtained fermented black bean curd samples was determined according to the SB / T 23527-2009 method, the β-glucosidase activity was determined by the p-nitrophenol colorimetric method, the amino acid nitrogen content was determined by the formaldehyde titration method, the aglycone-type soy isoflavone content was determined by HPLC, and the antioxidant activity was determined by the FRAP method. The TEAC value was expressed as μmol Trolox / g DW, and the FRAP value was expressed as μmol Fe2+ / DW.

[0069] The results are as follows Figure 1-Figure 4 As shown. It can be seen that at an inoculum concentration of 6% ( Figure 1 As shown), the ratio of Aspergillus oryzae to Mucor racemosus is 1:1 (as shown Figure 2 As shown), the amount of edible fungus dry powder added is 20% (as shown Figure 3 As shown), the fermentation time is 60h (as shown Figure 4 As shown in the figure, the neutral protease activity and β-glucosidase activity of edible fungus Douchiqu were the highest overall.

[0070] 2. Response surface optimization experiment

[0071] According to the above single factor test results, the ratio of Aspergillus oryzae to Mucor racemosus was kept at 1:1, and the three factors of inoculation amount (X1), edible fungus powder addition amount (X2) and fermentation time (X3) that had a significant impact on the neutral protease activity and β-glucosidase activity of edible fungi Douchiqu were selected as independent variables. The neutral protease activity and β-glucosidase activity of edible fungi Douchiqu were used as evaluation indicators (Y). The central composite design method was used to design a three-factor three-level analysis test. The experimental design factors and levels are shown in Table 2, and the experimental design scheme and results are shown in Table 3.

[0072] Table 2. Factor levels and codes of Box-Benhnken design experiment for pre-fermentation conditions of edible fungi and fermented black beans

[0073]

[0074] Table 3. Response surface design and results of edible fungus fermentation conditions

[0075]

[0076]

[0077] The software Design-Expert 7.0 was used to perform quadratic polynomial regression fitting on the experimental data in Table 3, and a quadratic multivariate regression equation was established:

[0078] Y1=791.65-7.47A+12.28B+26.43C-1.79AB-6.39AC-9.90BC-39.37A 2 -12.62B 2 -78.73C 2 ;

[0079] Y2=1160.73+7.63A+6.59B+121.51C+28.05AB-0.8662AC-16.31BC-56.46A 2 -82.57B 2 -192.71C 2 .

[0080] The response surface model analysis predicted that the optimal conditions for mixed edible fungus fermentation of fermented black beans were as follows: inoculation amount 5.908%, edible fungus powder addition amount 20.65%, koji making time 62.753 h, predicted protease activity value 794.997 U / g, and predicted β-glucosidase activity value 1176.816 U / g.

[0081] 3. Verification of mixed fermentation process of edible fungi, fermented black beans and koji

[0082] According to the above process investigation results, combined with actual production, the optimal fermentation process was determined to be a mixed spore solution with a ratio of 1:1 between Aspergillus oryzae and Mucor racemosus, an inoculation amount of 6%, an edible fungus powder addition amount of 21%, and a fermentation time of 63h. According to the above process, 6 batches of edible fungus Douchi Koji were prepared for verification experiments, and the measured values ​​of protease activity and β-glucosidase activity were 792.03±9.76U / g and 1162.00±35.95U / g, which were close to the predicted values ​​of 794.997U / g and 1176.816U / g.

[0083] The above experimental results show that in the koji making process of the mixed fermented edible fungus fermented black beans of the present invention, when a mixed fungus spore liquid with a ratio of Aspergillus oryzae to Mucor racemosus of 1:1 is used, the inoculation amount is 6%, the edible fungus powder addition amount is 21%, and the fermentation time is 63h, the protease activity and β-glucosidase activity of the obtained edible fungus fermented black beans koji are significantly improved.

[0084] In summary, the present invention provides an edible fungus fermented black beans based on co-fermentation of Mucor and Aspergillus and a koji making process thereof. The protease activity, β-glucosidase activity, amino acid nitrogen content, soybean isoflavone content and antioxidant activity of the edible fungus fermented black beans provided by the present invention are significantly improved, and the edible fungus fermented black beans provided by the present invention have excellent nutrient content, biological activity and health care effects. The koji making process of the present invention is simple to operate, low in cost, short in fermentation time, and has broad application prospects.

Claims

1. A koji making process for edible fungus fermented black beans, characterized in that: The process comprises the following steps: soaking beans in water, draining, mixing evenly with edible fungus waste, steaming under high pressure, cooling, inoculating mixed fungus spore liquid containing Mucor and Aspergillus, and fermenting to obtain edible fungus fermented black beans; using edible fungus waste and beans as raw materials, the volume mass ratio of the mixed fungus spore liquid to the raw materials is 6% mL / g, and the concentration of the mixed fungus spore liquid is 1×10 6 -1×10 8 CFU / mL; in the mixed spore liquid, the Mucor is Mucor racemosus, the Aspergillus is Aspergillus oryzae, and the quantity ratio of the Mucor to the Aspergillus is 1:1; in the raw materials, the mass ratio of edible fungi scraps and beans is (20-21):100; the fermentation time is 63 hours; the fermentation temperature is 20-30°C; the relative humidity of the fermentation is 70%-90%; the beans are soybeans, and the edible fungi scraps include one or a mixture of two or more of the mushroom feet, mushroom slices, mushroom stems, deformed mushrooms, and bacterial residues after functional component extraction of edible fungi; the edible fungi are one or two of Pleurotus eryngii and Oyster mushroom; the mass volume ratio of beans to water is 1: (2-5) g / mL, the soaking temperature is 24-45°C, and the soaking time is 2-5h.

2. The koji making process according to claim 1, characterized in that: The concentration of the mixed spore solution is 1×10 7 CFU / mL.

3. The koji making process according to claim 1, characterized in that: The fermentation temperature was 28° C., and the fermentation relative humidity was 80%.

4. The koji making process according to claim 1, characterized in that: The mass volume ratio of the beans to water is 1:3 g / mL, the soaking temperature is 35° C., and the soaking time is 4 hours.

5. The koji making process according to claim 1, characterized in that: The high pressure cooking is performed at a temperature of 110-130° C., a pressure of 0.05-0.2 MPa, and a time of 15-35 min.

6. The koji making process according to claim 5, characterized in that: The high pressure cooking was carried out at a temperature of 121° C., a pressure of 0.1 MPa and a time of 25 min.

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