Bean sour soup processing method based on soybean clear liquid fermentation

Abstract

The invention relates to the technical field of fermentation, in particular to a bean sour soup processing method based on bean clear liquid fermentation, and the processing method comprises the following steps: S1, filtering bean clear liquid, adjusting dry matter concentration, adding glucose, pasteurizing, and cooling; s2, inoculating a mixed strain of aspergillus oryzae and aspergillus sojae, and performing aerated fermentation; s3, inoculating a lactic acid bacteria mixed strain, and carrying out sealed anaerobic fermentation; s4, carrying out hot filtration, centrifugation and ultra-high temperature sterilization on the fermentation liquid, inoculating bacillus velezensis and gluconacetobacter, carrying out aeration stirring fermentation, inoculating saccharomycetes, and standing for anaerobic fermentation; and S5, carrying out solid-liquid separation on the fermentation liquor, and sterilizing to obtain the bean sour soup product. According to the method, multi-stage ordered fermentation is combined with a multi-strain synergistic effect, and parameters such as filtration, inoculation amount and temperature in each stage are optimized, so that efficient debitterizing, umami enhancement, aroma level enrichment and sour taste softening are realized, sanitation, safety and quality stability of products are guaranteed, and the taste and quality of the bean sour soup are effectively improved.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of fermentation, in particular to a processing method of bean sour soup based on bean clear liquid fermentation. BACKGROUND

[0002] As a traditional fermented food with both nutrition and flavor, the processing core of bean sour soup lies in the microbial transformation of the nutritional components in bean raw materials to form a unique sour and fragrant flavor, which has a stable consumer base in the market. In the field of bean product processing, bean clear liquid is the main by-product produced during the production of mainstream products such as tofu and soy milk, which contains a certain amount of plant protein, carbohydrates and minerals. If it is directly discharged, it not only wastes nutritional resources, but also increases environmental treatment costs. Therefore, using bean clear liquid as the fermentation raw material of bean sour soup can not only realize the high-value utilization of by-products and reduce resource loss and environmental pressure, but also provide a natural nutritional substrate for bean sour soup, which meets the current demand for green development of the food industry.

[0003] Traditional bean sour soup fermentation relies on natural inoculation fermentation or single strain fermentation mode, and the fermentation process lacks precise control. For the fermentation application of bean clear liquid raw materials, it is more inclined to simple anaerobic fermentation process - usually after basic filtration and impurity removal (or without treatment) of bean clear liquid, a single lactic acid bacteria strain is directly inoculated to complete fermentation in a sealed environment. The fermentation stages are not divided according to the nutritional characteristics of the bean clear liquid, and multiple strains are not introduced for synergistic action. The overall process is relatively extensive.

[0004] However, the traditional fermentation method has some disadvantages. Specifically, on the one hand, due to the single type of strain and the lack of stage design in the fermentation process, the types of flavor substances generated by microbial metabolism are limited, and only basic sour taste can be formed, lacking complex aroma and umami levels, resulting in monotonous taste of bean sour soup, which is difficult to meet the needs of consumers for flavor diversity. On the other hand, the protein in the bean clear liquid is not completely decomposed in the traditional fermentation, and it is easy to leave bitter peptides or free hydrophobic bitter amino acids (such as leucine and phenylalanine), which makes the product have a weak but affecting taste of bitterness, which is a key problem restricting the quality improvement and market competitiveness of bean sour soup. SUMMARY

[0005] In view of the technical defects in the background art, the present application provides a processing method of bean sour soup based on bean clear liquid fermentation, which solves the above technical problems and meets the actual needs. The specific technical scheme is as follows: A processing method of bean sour soup based on bean clear liquid fermentation, comprising the following steps: S1. Bean clear liquid pretreatment: filter the bean clear liquid to remove insoluble impurities, adjust the dry matter concentration to 5-8°Bx, add glucose as the initial carbon source, and then sterilize at 75-85°C for 15-30 min and cool to 30-35°C; S2. The first stage of enzymatic hydrolysis of fresh fermentation: inoculating Aspergillus oryzae and Aspergillus sojae mixed strains into the soybean liquid, and fermenting at 30-35℃, 0.3-0.5 m 3 / (m 3 ·h) under low-intensity ventilation conditions for 24-48h, until the content of amino acid nitrogen (AAN) in the fermentation broth reaches 0.3-0.5g / L; S3. The second stage of acid production and flavor enhancement fermentation: without sterilization, directly inoculating Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus and Paracaseicola lactis mixed strains into the first stage fermentation broth, and fermenting at 37-40℃ under sealed anaerobic conditions for 12-24h, until the pH of the fermentation broth is reduced to 4.0-4.3; S4. The third stage of debittering and flavor enhancement fermentation: centrifuging and sterilizing the second stage fermentation broth, adding 0.5%-1% of glucose, then inoculating Bacillus velezensis and Gluconacetobacter, and fermenting at 28-30℃ under ventilation and stirring conditions for 12-20h, then inoculating Kluyveromyces marxianus and Saccharomyces cerevisiae, and fermenting at 30-32℃ under static and anaerobic conditions for 20-24h; S5. After the fermentation is completed, the fermentation broth of the third stage is subjected to solid-liquid separation, heated to 85-95℃ for sterilization for 10-15min, and the soybean acid soup product is obtained.

[0006] As a further technical solution of the present application, the preparation method of the soybean liquid is: Selecting high-quality green skin soybeans with full grains and no insect damage, soaking the green skin soybeans at a water temperature of 18-22℃, the weight of the soaked green skin soybeans being about 2.0~2.2 times that before soaking, grinding the soaked green skin soybeans with water to obtain a slurry; Steam heating the obtained slurry to 40-60℃, continuing to steam heat to boiling after defoaming treatment, then slowly boiling for 3-5min, then performing extrusion separation with a filter bag for 3 times to obtain soybean milk; Adjusting the concentration of the soybean milk to 6°Bx, and cooling to 75-80℃, then mixing the cooled soybean milk with acid slurry water (prepared by sealed fermentation of soybean liquid) at a mass ratio of 3:1 until flocculent precipitates appear in the soybean milk, then heating for 1-2min, then standing for heat preservation, then rolling the coagulated soybean milk with a weight, collecting the squeezed slurry water to obtain the soybean liquid, and the total acid content of the acid slurry water is 5.0-5.5g / kg.

[0007] As a further technical solution of the present application, the filtration in step S1 uses a 100-200 mesh screen; the glucose addition amount is 1.5%-2.5% of the mass of the soybean liquid.

[0008] As a further technical scheme of the present application, the mass ratio of the Aspergillus oryzae and the Aspergillus sojae in step S2 is 8:2, the inoculation amount is 0.5‰-1.0‰ of the total mass of the bean liquid; the Aspergillus oryzae is selected from a strain with an amino peptidase activity of ≥500 U / g, and the Aspergillus sojae is selected from a strain with a protease activity of ≥300 U / g.

[0009] As a further technical scheme of the present application, in step S3, the mass ratio of the Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus and Lactobacillus paracasei is 1:1:0.2, and before use, the three strains are revived in sterile bean liquid at 37-40℃ for 1-2h, and the inoculation amount is 1.0%-2.0% of the volume of the fermentation liquid; the Lactobacillus delbrueckii subsp. bulgaricus and the Lactobacillus paracasei are selected from a strain with an acid production rate of ≥2.0 g / (L·h), and the Streptococcus thermophilus is selected from a strain with an acetaldehyde production amount of ≥5.0 mg / L.

[0010] As a further technical scheme of the present application, in step S4, when aerobic fermentation is performed, the inoculation amount of the Bacillus velezensis is 0.5%-1.0% of the volume of the fermentation liquid, the inoculation amount of the Gluconacetobacter is 0.2%-0.4% of the volume of the fermentation liquid, and the aeration and stirring conditions are specifically as follows: the aeration amount is 0.5-1.0 m 3 / (m 3 ·h), and the stirring mode is to stir at a speed of 60 rpm for 10-15 min every 2h.

[0011] As a further technical scheme of the present application, in step S4, when anaerobic fermentation is performed, the Kloeckera apiculata and the Saccharomyces cerevisiae are both activated in sterile water containing 2%-3% (mass percent) glucose at 30℃-32℃ for 2-3 hours, and the activity of the activated bacterial liquid is ≥10^8 CFU / mL, the inoculation amount of the Kloeckera apiculata is 0.5%-1.0% of the volume of the fermentation liquid, and the Saccharomyces cerevisiae is an edible strain with an amino acid assimilation rate of ≥80%, and the inoculation amount is 2.0%-3.0% of the volume of the fermentation liquid.

[0012] As a further technical scheme of the present application, in step S4, the specific operation of centrifugal separation and sterilization is as follows: the second-stage fermentation liquid is subjected to hot filtration using a sieve with a mesh size of 200-400 to collect the filtrate, the filtration temperature is 40-50℃, then the filtrate is centrifugally separated at a speed of 8000 rpm for 10 min, the supernatant is collected, the supernatant is subjected to ultra-high temperature instantaneous sterilization at 115-121℃ for 8-15s, and then rapidly cooled to 28-30℃ to obtain the initial fermentation liquid for the third-stage fermentation.

[0013] The present application has the beneficial effects of: The present application effectively solves the problem of bitterness of plant protein fermentation products through multi-stage precise fermentation, and improves the flavor quality. Through three-stage orderly fermentation, the first stage of mold releases hydrophobic bitter amino acids by the action of aminopeptidase, and the third stage of bacillus velezensis consumes these amino acids and converts them into flavor substances, fundamentally solving the problem of bitterness and enhancing umami. The first stage of aspergillus oryzae and aspergillus sojae decompose soybean supernatant proteins into free amino acids such as glutamic acid, thereby increasing the content of amino acid nitrogen. The third stage of bacillus can also produce fresh and sweet amino acids. In addition, the aroma is rich and has multiple levels. Lactic acid bacteria bring sour and incense aroma, acetic acid bacteria and yeast generate alcohol, esters and other substances to impart multiple types of aroma. And the consumption of lactic acid by Kluyveromyces marxianus makes the sour taste soft and harmonious. Finally, a soybean soup product with excellent quality is obtained. DETAILED DESCRIPTION

[0014] The embodiments of the present application will be described below in conjunction with relevant examples. The embodiments of the present application are not limited to the following examples, and the present application relates to necessary components in the related technical field, which should be regarded as known technology in the related technical field and can be known and mastered by those skilled in the art.

[0015] A soybean soup processing method based on soybean supernatant fermentation, comprising the following steps: S1. Soybean supernatant pretreatment: filter the soybean supernatant to remove insoluble impurities, adjust the dry matter concentration to 5-8°Bx, add glucose as the initial carbon source, and then pasteurize at 75-85°C for 15-30 min and cool to 30-35°C; S2. First-stage enzyme hydrolysis and umami fermentation: inoculate the soybean supernatant with a mixed strain of Aspergillus oryzae and Aspergillus sojae, and ferment at 30-35°C, 0.3-0.5 m 3 / (m 3 ·h) low-intensity ventilation conditions for 24-48 h, until the content of amino acid nitrogen (AAN) in the fermentation broth reaches 0.3-0.5 g / L; S3. Second-stage acid production and aroma enhancement fermentation: without sterilization, directly inoculate the mixed strain of Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus and Paracaseolus casei into the first-stage fermentation broth, and ferment at 37-40°C under sealed anaerobic conditions for 12-24 h, until the pH of the fermentation broth drops to 4.0-4.3; S4. Third-stage debittering and aroma enhancement fermentation: centrifuge and sterilize the second-stage fermentation broth, add 0.5%-1% glucose, then inoculate Bacillus velezensis and Gluconacetobacter, and ferment at 28-30°C under aeration and stirring conditions for 12-20 h, then inoculate Kluyveromyces marxianus and Saccharomyces cerevisiae, and ferment at 30-32°C under static and anaerobic conditions for 20-24 h; S5. After the fermentation is completed, the third-stage fermentation liquor is subjected to solid-liquid separation, heated to 85-95 DEG C and sterilized for 10-15 min to obtain a soybean sour soup product.

[0016] The present application realizes systematic optimization of product taste, flavor and quality in the processing of soybean sour soup by constructing a multi-stage ordered fermentation process system, and the processing method can realize efficient debittering and taste improvement. The bitterness problem of plant protein fermentation is solved by stage-by-stage synergistic effect, the first stage of inoculation of mold can produce specific enzyme system, which releases the hydrophobic amino acid prone to produce bitterness in the protein decomposition process from the end of the peptide chain, laying a foundation for subsequent debittering; the third stage introduces functional strains, which can efficiently consume these free bitter amino acids and convert them into flavor substances, fundamentally eliminating the source of bitterness, avoiding the problems of residual bitterness or incomplete debittering in traditional processing, and significantly improving the palatability of the product.

[0017] In terms of umami enhancement, the process design of the present application shows clear progression and synergy. The mold strain selected in the first stage has high enzyme production capacity, which can fully decompose the protein in the soybean juice into free amino acids, among which the accumulation of umami amino acids provides the core source of product umami; and the strains inoculated in the third stage can further generate fresh sweet amino acids, which complement the umami accumulated in the previous stage, making the product umami more rich and natural, and the umami substances formed during the fermentation process are more easily perceived by the human body than exogenous addition, and have stronger flavor coordination.

[0018] In addition, the process effectively enriches the product aroma levels through multi-strain functional synergy, the acid aroma and milk aroma produced by the metabolism of lactic acid bacteria in the second stage lay the foundation for the product flavor; the third stage of acetic acid bacteria and yeast respectively contribute to characteristic flavor substances and alcohol, ester, aldehyde and other aromatic compounds, and various aroma components gradually accumulate and integrate in the process, forming a complex aroma system with acid aroma, milk aroma, fruit aroma and wine aroma, breaking through the limitation of single aroma of traditional soybean sour soup. At the same time, the regulation of sour taste quality also shows the advantage of the process. The introduction of yeast in the third stage can specifically consume part of the lactic acid, effectively reducing the sharp sour taste commonly found in traditional fermented products, making the sour taste more tender and mellow; realizing the synergistic improvement of soybean sour soup taste and flavor.

[0019] Further, the preparation method of the soybean juice of the present application is: Select high-quality green skin beans with full particles and no insect damage, soak the green skin beans at a water temperature of 18-22 DEG C, the weight of the soaked green skin beans is about 2.0-2.2 times that of the unsoaked green skin beans, grind the soaked green skin beans with water to obtain a slurry; The ground slurry is heated to 40-60℃ by steam, defoamed, and then heated to boiling by steam, and then boiled for 3-5min, and then separated by squeezing for 3 times by filter bag to obtain soybean milk; The concentration of the soybean milk is adjusted to 6°Bx, and then cooled to 75-80℃, and then mixed with acid slurry water (prepared by sealed fermentation of soybean dreg) at a mass ratio of 3:1 until flocculent precipitate appears in the soybean milk, and then heated for 1-2min, and then kept still to obtain coagulated soybean milk, and then the coagulated soybean milk is rolled by weight, and then the squeezed slurry water is collected to obtain soybean dreg, and the total acid content of the acid slurry water is 5.0-5.5g / kg.

[0020] Specifically, in the present application, the mixing temperature of the cooled soybean milk and the acid slurry water is 76℃, and the total acid content of the acid slurry water is 5.3g / kg.

[0021] As one of the preferred embodiments of the present application, the filter in step S1 adopts a 100-200 mesh screen; and the added amount of glucose is 1.5%-2.5% of the mass of the soybean dreg.

[0022] Specifically, the filter adopts a 150 mesh screen, and the added amount of glucose is 2% of the mass of the soybean dreg. The selection of the 150 mesh screen can efficiently trap insoluble impurities (such as small soybean dregs and raw material debris) in the soybean dreg, avoid the residues affecting the uniformity of subsequent inoculation of strains and metabolic efficiency, and reduce the loss of water-soluble nutrients caused by too fine screen, so as to ensure the purity and nutritional integrity of the fermentation substrate. The glucose added amount of 2% can serve as an initial carbon source to meet the carbon source demand of the initial growth and reproduction of the subsequent inoculated strains (Aspergillus oryzae and Aspergillus sojae), promote the strains to quickly adapt to the environment and start the enzyme production process, and avoid the fermentation delay caused by insufficient carbon source.

[0023] As one of the preferred embodiments of the present application, the mass ratio of the Aspergillus oryzae and the Aspergillus sojae in step S2 is 8:2, and the inoculation amount is 0.5‰-1.0‰ of the total mass of the soybean dreg; the Aspergillus oryzae selected is a strain with an ammonia peptidase activity of ≥500U / g, and the Aspergillus sojae selected is a strain with a protease activity of ≥300U / g.

[0024] The mass ratio of the Aspergillus oryzae and the Aspergillus sojae is 8:2, the Aspergillus oryzae has high ammonia peptidase activity as a core advantage, and the Aspergillus sojae focuses on efficient production of protease. This ratio can balance the output of the two key enzyme systems. The ammonia peptidase dominated by the Aspergillus oryzae can effectively release the hydrophobic bitter amino acids at the end of the peptide chain, laying a foundation for subsequent debittering; the protease of the Aspergillus sojae assists in the decomposition of proteins in the soybean dreg, and the two can synergistically accelerate the protein degradation process to create conditions for umami substance accumulation.

[0025] The inoculation amount is preferably 0.75‰ of the total mass of the bean juice, which can ensure that the strains quickly form a dominant flora in the fermentation system, inhibit the growth of miscellaneous bacteria, and avoid fermentation abnormalities, and can also avoid excessive initial nutrient competition and unbalanced accumulation of metabolic products, thereby ensuring a stable and controllable fermentation process.

[0026] Meanwhile, the enzyme activity indicators (Aspergillus oryzae aminopeptidase activity ≥ 500 U / g and soy sauce koji protease activity ≥ 300 U / g) of the strains can ensure the fermentation effect, the aminopeptidase activity can ensure the release efficiency of bitter amino acids, and the protease activity can ensure that the proteins are fully degraded into small molecular peptides and free amino acids, thereby ultimately promoting the content of amino acid nitrogen (AAN) in the first-stage fermentation liquor to stably reach the target range, thereby providing a fermentation basis for the subsequent acid production and flavor enhancement and debittering and flavor enhancement stages.

[0027] As one of the preferred embodiments of the present application, in step S3, the mass ratio of Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus and Lactobacillus paracasei is 1:1:0.2, and the strains are revived in sterile bean juice at 37-40°C for 1-2h before use, and the inoculation amount is 1.0%-2.0% of the volume of the fermentation liquor; the Lactobacillus delbrueckii subsp. bulgaricus and Lactobacillus paracasei are selected to be strains with an acid production rate ≥2.0 g / (L·h), and the Streptococcus thermophilus is selected to be a strain with an acetaldehyde production amount ≥5.0 mg / L.

[0028] The mass ratio of Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus and Lactobacillus paracasei 1:1:0.2 is based on the complementary selection of their metabolic characteristics: the first two strains in a 1:1 ratio can synergistically produce acid and flavor, Lactobacillus delbrueckii subsp. bulgaricus is the main acid-producing strain, and Streptococcus thermophilus assists in acid production and contributes to characteristic flavors; Lactobacillus paracasei is added at a ratio of 0.2, which can further enhance the stability of acid production, and form a metabolic synergy with the first two strains to avoid monotonous flavors caused by single-strain fermentation.

[0029] Before use, the mixed strains are revived in sterile bean juice at 38.5°C for 2h, and the revival temperature is within the subsequent fermentation temperature range, which can reduce the adaptation time of the strains after inoculation; the nutrients in the sterile bean juice can provide the energy and substances required for the revival of the strains, and compared with sterile water revival, the strains are more likely to recover high activity, which can ensure that the metabolism is quickly started after inoculation and avoid fermentation delay.

[0030] The inoculation amount is preferably 1.5% of the fermentation liquid volume, which can ensure that the bacteria quickly form a dominant flora in the system, inhibit the potential breeding of residual miscellaneous bacteria in the first stage fermentation, avoid excessive inoculation amount leading to intense nutrient competition and unbalanced accumulation of metabolic products, maintain the stability of the fermentation process, and ensure that the Lactobacillus delbrueckii subsp. bulgaricus and Lactobacillus paracasei have an acid production rate of ≥2.0 g / (L·h) and the Streptococcus thermophilus has an acetaldehyde production amount of ≥5.0 mg / L, which can effectively ensure the fermentation effect of the second fermentation stage, and the acid production rate of the Lactobacillus delbrueckii subsp. bulgaricus and the Lactobacillus paracasei can ensure that the pH of the fermentation liquid is reduced to 4.0-4.3 on time, and the acetaldehyde production amount can impart a fresh milk fragrance to the product.

[0031] As one of the preferred embodiments of the present application, in step S4, when aerobic fermentation is performed, the inoculation amount of the Bacillus velezensis is 0.5%-1.0% of the fermentation liquid volume, the inoculation amount of the Gluconacetobacter is 0.2%-0.4% of the fermentation liquid volume, and the aeration and stirring conditions are as follows: the aeration amount is 0.5-1.0 m 3 / (m 3 ·h), and the stirring mode is to stir at a speed of 60 rpm for 10-15 min every 2 h.

[0032] Specifically, the inoculation amount of the Bacillus velezensis is 0.75% of the fermentation liquid volume, the inoculation amount of the Gluconacetobacter is 0.3% of the fermentation liquid volume, and the aeration and stirring conditions are as follows: the aeration amount is 0.75 m 3 / (m 3 ·h), and the stirring mode is to stir at a speed of 60 rpm for 12 min every 2 h.

[0033] The inoculation amount of the Bacillus velezensis is 0.75%, and the inoculation amount of the Gluconacetobacter is 0.3%. The above inoculation amount is combined with the metabolic characteristics of the two strains, i.e., the Bacillus velezensis consumes free bitter amino acids and generates fresh and sweet amino acids, and the Gluconacetobacter needs to be inoculated moderately to convert organic acids into characteristic flavors, which can avoid excessive inoculation amount leading to intense nutrient competition or insufficient metabolic efficiency caused by low inoculation amount, and maximize the synergistic effect of functional strains.

[0034] The aeration amount of 0.75 m 3 / (m 3 ·h) can stably provide an aerobic environment to meet the aerobic metabolism needs of the two strains, and avoid excessive aeration causing water loss and temperature fluctuation of the fermentation liquid; regular stirring for 12 min every 2 h can realize uniform contact of the bacteria and the fermentation substrate through a moderate stirring speed of 60 rpm, prevent local nutrient deficiency or accumulation of metabolic products, reduce the influence of continuous stirring on the activity of the strains, ensure the stability of the metabolic process, and accumulate high-quality flavor substrates for the subsequent anaerobic fermentation stage.

[0035] As one of the preferred embodiments of the present application, in step S4, when anaerobic fermentation is carried out, the K. marxianus and the S. cerevisiae are activated by using sterile water containing 2%-3% (mass percentage) of glucose, and the activation is carried out at 30-32℃ for 2-3 hours. After the activation, the activity of the bacterial liquid is greater than or equal to 10^8 CFU / mL. The inoculation amount of the K. marxianus is 0.5%-1.0% of the volume of the fermentation liquid, and the S. cerevisiae is an edible strain with an amino acid assimilation rate greater than or equal to 80%, and the inoculation amount is 2.0%-3.0% of the volume of the fermentation liquid.

[0036] Specifically, the K. marxianus and the S. cerevisiae are activated by using sterile water containing 2.5% (mass percentage) of glucose, and the activation is carried out at 31℃ for 2.5 hours. The inoculation amount of the K. marxianus is 0.75% of the volume of the fermentation liquid, and the inoculation amount of the S. cerevisiae is 2.5% of the volume of the fermentation liquid.

[0037] In terms of the inoculation amount, the K. marxianus is 0.75%, and the S. cerevisiae is 2.5%. This setting fully adapts to the functional positioning of the two strains. The K. marxianus consumes the accumulated lactic acid in the early fermentation, weakens the sharp sour taste, and gives softness to the sour taste. The S. cerevisiae can fully assimilate free amino acids (including bitter amino acids), which not only helps to remove bitterness, but also generates alcohol and ester aromatic substances. This inoculation amount can not only ensure that the two strains quickly form a dominant flora and avoid the growth of miscellaneous bacteria, but also avoid excessive inoculation that causes metabolic imbalance due to nutrient competition, and ultimately ensure the stable realization of flavor improvement and taste optimization in the anaerobic fermentation stage.

[0038] As one of the preferred embodiments of the present application, in step S4, the specific operation of centrifugal separation and sterilization is as follows: the second-stage fermentation liquid is subjected to hot filtration using a 200-400 mesh screen and the filtrate is collected, the filtration temperature is 40-50℃, then the filtrate is centrifugally separated at a speed of 8000 rpm for 10 min, the supernatant is collected, the supernatant is subjected to ultra-high temperature instantaneous sterilization at 115-121℃ for 8-15 s, then it is rapidly cooled to 28-30℃, and the initial fermentation liquid for the third-stage fermentation is obtained.

[0039] Specifically, the second-stage fermentation liquid is subjected to hot filtration using a 300 mesh screen and the filtrate is collected, the filtration temperature is 45℃, then the filtrate is centrifugally separated at a speed of 8000 rpm for 10 min, the supernatant is collected, the supernatant is subjected to ultra-high temperature instantaneous sterilization at 118℃ for 11 s, then it is rapidly cooled to 28℃, and the initial fermentation liquid for the third-stage fermentation is obtained.

[0040] It should be noted that after the lactic acid is consumed through the above fermentation, the ph of the third-stage fermentation liquid is increased to 4.7-4.9.

[0041] The 300-mesh screen is selected to intercept lactic acid bacteria cell fragments, small undigested substrates and other impurities, so as to avoid the influence of the residues on subsequent strain metabolism. The filtration temperature of 45°C can maintain the flowability of the fermentation broth to improve the filtration efficiency, while avoiding the destruction of the organic acids and small-molecule flavor precursors accumulated in the early stage by high temperature, and protecting the substrate nutrients and flavor basis.

[0042] Centrifugation at 8000 rpm for 10 min can further separate the small particles and colloidal substances that are not removed by hot filtration, so that the supernatant is more clear, the interference of impurities on the growth of the third stage strain (such as Bacillus velezensis and yeast) is eliminated, and the uniform contact between the strain and nutrients is ensured, laying a foundation for stable metabolism.

[0043] The ultra-high temperature instantaneous sterilization at 118°C for 11 s can quickly kill the residual lactic acid bacteria and potential bacteria in the supernatant, prevent them from competing with the third stage inoculated strain for nutrients or interfering with metabolism, and at the same time, the "instant high temperature" characteristic can minimize the thermal decomposition of nutrients such as amino acids and small-molecule peptides, retaining the nutritional value of the substrate; after sterilization, the substrate is quickly cooled to 28°C, which can avoid the growth of intermediate temperature zone bacteria, and directly adapt the substrate temperature to the optimal growth conditions of the initial strain in the third stage, ensuring the rapid and stable start of subsequent fermentation.

[0044] The application will be further described below through examples and comparative examples.

[0045] Example 1: A soy acid soup processing based on soy broth fermentation S1. Soy broth pretreatment: filter the soy broth using a 150-mesh screen to remove insoluble impurities, adjust the dry matter concentration of the soy broth to 6.5°Bx, and add 2% glucose based on the mass of the soy broth as the initial carbon source; the above system is subjected to 80°C pasteurization for 20 min, and then cooled to 32°C to obtain the pretreated soy broth.

[0046] S2. First-stage enzyme hydrolysis and flavor enhancement fermentation: inoculate the pretreated soy broth with 0.75‰ of mixed strains (mass ratio of Aspergillus oryzae to Aspergillus sojae 8:2, wherein the amylolytic enzyme activity of Aspergillus oryzae is 550 U / g, and the protease activity of Aspergillus sojae is 350 U / g) based on the total mass of the soy broth; ferment at 32°C, 0.4 m 3 / (m 3 h) low-intensity aeration conditions for 36 h, until the amino acid nitrogen (AAN) content in the fermentation broth reaches 0.4 g / L, to obtain the first-stage fermentation broth.

[0047] S3. Second-stage acid-producing and flavor-enhancing fermentation: without sterilization, 1.5% of mixed lactic acid bacteria (Lactobacillus delbrueckii subsp. bulgaricus: Streptococcus thermophilus: Lactobacillus paracasei in a mass ratio of 1:1:0.2) were inoculated into the first-stage fermentation liquor; the mixture was incubated at 38°C under sealed anaerobic conditions for 18 h until the pH of the fermentation liquor dropped to 4.0-4.3, thereby obtaining a second-stage fermentation liquor.

[0048] S4. Third-stage bitterness-removing and flavor-enhancing fermentation: Centrifugal separation and sterilization: the second-stage fermentation liquor was hot-filtered at 45°C using a 300-mesh screen and the filtrate was collected; the filtrate was centrifugally separated at a speed of 8000 rpm for 10 min and the supernatant was collected; the supernatant was subjected to ultra-high temperature instantaneous sterilization at 118°C for 11 s, and then rapidly cooled to 28°C, thereby obtaining a third-stage initial fermentation liquor; Aerobic fermentation: 0.75% of glucose was added to the third-stage initial fermentation liquor, followed by inoculation of 0.75% of Bacillus velezensis and 0.3% of Gluconacetobacter sp.; the mixture was incubated at 29°C under aeration and agitation conditions (aeration amount: 0.75 m 3 / (m 3 ·h, agitation speed: 60 rpm, agitation time: 12 min, agitation interval: 2 h) for 16 h; Anaerobic fermentation: mixed yeasts (0.75% of Kluyveromyces marxianus and 2.5% of Saccharomyces cerevisiae in terms of the volume of the fermentation liquor; both of the yeasts were activated at 31°C for 2.5 h using sterile water containing 2.5% of glucose, and the activity of the activated bacteria was ≥10^8 CFU / mL, and the amino acid assimilation rate of Saccharomyces cerevisiae was 85%) were inoculated into the system after aerobic fermentation; the mixture was incubated at 31°C under static anaerobic conditions for 22 h, thereby obtaining a third-stage fermentation liquor (pH increased to 4.6-4.8).

[0049] S5. Post-treatment: the third-stage fermentation liquor was subjected to solid-liquid separation, the liquid phase was collected and sterilized at 90°C for 12 min, and then filled after sterilization, thereby obtaining a soybean sour soup product.

[0050] Example 2 Example 2 differs from Example 1 in that: in S2, the strain of Aspergillus oryzae has an aminopeptidase activity of 600 U / g, and the strain of Aspergillus sojae has a protease activity of 400 U / g; in S3, the acid-producing rate of Lactobacillus delbrueckii subsp. bulgaricus and Lactobacillus paracasei is both ≥2.5 g / (L·h), and the acetaldehyde-producing amount of Streptococcus thermophilus is ≥6.0 mg / L; in S4, the strain of Saccharomyces cerevisiae has an amino acid assimilation rate of ≥90%; and the parameters of the remaining steps are the same as those in Example 1.

[0051] Example 3 The difference between Example 3 and Example 1 is that the S2 first-stage enzymatic freshening fermentation time is shortened to 28 h; the S3 second-stage acid production and flavor enhancement fermentation temperature is 37℃, and the fermentation time is shortened to 14 h; in S4, the aerobic fermentation time is shortened to 12 h, and the anaerobic fermentation time is shortened to 20 h; the remaining step parameters are the same as those in Example 1.

[0052] Comparative Example 1 The difference between Comparative Example 1 and Example 1 is that the S2 first-stage enzymatic freshening fermentation step is completely omitted, and the S1 pretreated bean juice is directly subjected to the S3 second-stage acid production and flavor enhancement fermentation (the strain and fermentation parameters of S3 are the same as those in Example 1), and the remaining step (S1, S3-S5) parameters are the same as those in Example 1.

[0053] Comparative Example 2 The difference between Comparative Example 2 and Example 1 is that the S4 third-stage debittering and flavor enhancement fermentation step is completely omitted, and after the S3 second-stage fermentation is completed, the second-stage fermentation liquid is directly subjected to the S5 solid-liquid separation and sterilization (the S5 parameters are the same as those in Example 1), and the remaining step (S1-S3) parameters are the same as those in Example 1.

[0054] Comparative Example 3 The difference between Comparative Example 3 and Example 1 is that the S4 third-stage debittering and flavor enhancement fermentation only has an aerobic fermentation step (i.e., the Bacillus velezensis and Gluconacetobacter bacteria are inoculated and fermented for 16 h, and the parameters are the same as those in Example 1), and after the aerobic fermentation is completed, the subsequent anaerobic fermentation step is omitted, and the remaining step (S1-S3, S5) parameters are the same as those in Example 1.

[0055] Comparative Example 4 The difference between Comparative Example 4 and Example 1 is that the S4 third-stage debittering and flavor enhancement fermentation only has an anaerobic fermentation step (i.e., the initial fermentation liquid after centrifugal sterilization is directly inoculated with mixed yeast and fermented for 22 h, and the parameters are the same as those in Example 1), and the previous aerobic fermentation step is omitted, and the remaining step (S1-S3, S5) parameters are the same as those in Example 1.

[0056] Comparative Example 5 The difference between Comparative Example 5 and Example 1 is that only the S1 bean juice pretreatment and S3 lactic acid fermentation steps are retained, and the S3 fermentation time is extended to 36 h; the S1 pretreatment parameters are the same as those in Example 1, the strain ratio, revival conditions, and inoculation amount of the mixed lactic acid bacteria in S3 are the same as those in Example 1; after the S3 fermentation is completed, the S5 solid-liquid separation and sterilization are directly performed (the parameters are the same as those in Example 1), and the S2 and S4 steps are omitted.

[0057] The above examples and comparative examples are tested for relevant items, and the test results are shown in the following table:

[0058] From the comparison of examples and comparative examples, it can be seen that the complete multi-stage orderly fermentation system is the key to guarantee the comprehensive quality of the bean acid soup. Examples 1, 2 and 3 all contain the first-stage enzyme resolution freshening fermentation, the second-stage acid production and flavor enhancement fermentation and the third-stage debittering and flavor enhancement fermentation, and all show obvious advantages in various core indicators: the pH value is stabilized at 4.7-4.8, the total acid content is controlled in the moderate range of 8.2%-8.5%, the acid taste is soft and not sharp, avoiding the sharp acid feeling commonly seen in traditional fermented products; the amino acid nitrogen content reaches 0.10-0.15 g / L, the umami substances are sufficient and natural, compared with the exogenous added umami ingredients, they are more easily perceived by the human body and have stronger flavor coordination; in the volatile flavor substances, the ester content accounts for as high as 52.1%-56.2%, and the alcohol content also maintains at 28.9%-30.5%, forming a complex aroma system with acid aroma, milk aroma, fruit aroma and wine aroma, greatly enriching the product aroma levels and breaking through the limitation of single aroma of traditional bean acid soup.

[0059] Further analysis of the comparative examples missing different fermentation stages showed that each fermentation stage has irreplaceable functions, and the absence of any stage will cause the product quality to be significantly unbalanced. Comparative Example 1 omitted the first-stage enzymatic fresh-taste-enhancing fermentation and directly subjected the pretreated bean liquid to the second-stage acid-producing and flavor-enhancing fermentation. Due to the lack of specific enzymes produced by A. oryzae and A. sojae for protein decomposition, the protein could not be fully degraded into free amino acids, which not only resulted in an amino acid nitrogen content of only 0.05 g / L, far lower than that of the example group, and a severe lack of umami, but also caused the ester content in the volatile flavor substances to decrease to 30.5%, and the aldehyde content to increase to 12.5%, fully demonstrating that the first-stage fermentation is the key to releasing umami amino acids and laying the foundation for subsequent flavor formation. Comparative Example 2 omitted the third-stage debittering and flavor-enhancing fermentation and directly subjected the fermented liquid to post-treatment after the second-stage fermentation. Due to the lack of the synergistic effect of B. licheniformis, G. oxydans and yeast, lactic acid was not effectively consumed, resulting in a pH of 4.1 and a total acid content of 9.2%, a sharp and unbearable sour taste, and a significant bitter taste due to the inability to consume and convert bitter amino acids. The acid content in the volatile flavor substances increased to 52.48%, and the ester content decreased to only 15.2%, resulting in a single aroma, highlighting the core value of the third-stage fermentation in debittering, softening the sour taste and enriching the aroma. Comparative Example 3 only retained the third-stage aerobic fermentation and omitted the anaerobic fermentation. Although debittering was achieved to some extent by B. licheniformis and G. oxydans, the consumption of lactic acid by yeast and the generation of alcohol and ester aromatic compounds were lacking, resulting in a total acid content of 8.9%, a still irritating sour taste, an ester content of only 48.3%, and a lack of thickness and wine and fruit aromas, indicating that the third-stage anaerobic fermentation is crucial for sour taste softening and ester aroma generation. Comparative Example 4 only retained the third-stage anaerobic fermentation and omitted the aerobic fermentation. Due to the lack of debittering by B. licheniformis, free bitter amino acids could not be efficiently consumed, resulting in poor debittering, and G. oxydans could not participate in the metabolism to generate characteristic flavor substances, resulting in a single product flavor, an amino acid nitrogen content of only 0.09 g / L, and a further confirmation of the necessity of the third-stage aerobic fermentation for debittering and flavor foundation.

[0060] Compared with the traditional single lactic acid fermentation process, the multi-stage fermentation system of the present application has more obvious advantages. Comparative Example 5 only retains the soybean supernatant pretreatment and lactic acid fermentation steps, even if the fermentation time is extended to 36 h, the product pH value is still as low as 4.0, the total acid content is as high as 9.5%, and the sharpness of the sour taste is far higher than that of other groups; the amino acid nitrogen content is only 0.03 g / L, and the umami is almost missing; among the volatile flavor substances, the proportion of acids is as high as 61.6%, and the proportion of esters is only 8.5%, the aroma is single and the sour and astringent feeling is prominent, and the comprehensive quality is the worst, which fully confirms the great advantages of the multi-stage fermentation system in improving the taste, strengthening the umami and enriching the aroma compared with the traditional single fermentation process.

[0061] In addition, the strain activity and fermentation time also have certain influence on the product quality, but this influence needs to be based on the complete process system. In Example 2, higher activity strains are selected, such as the increase of the aspergillus oryzae aminopeptidase activity to 600 U / g, the aspergillus sojae protease activity to 400 U / g, the lactic acid bacteria acid production rate to ≥2.5 g / (L·h), and the saccharomyces cerevisiae amino acid assimilation rate to ≥90%, so that the amino acid nitrogen content reaches 0.15 g / L, and the ester proportion rises to 56.2%, which is slightly better than Example 1, indicating that high-activity strains can further improve the umami and flavor of the product on the basis of the optimal process parameters. In Example 3, the fermentation time of each stage is appropriately shortened, the first-stage fermentation time is shortened from 36 h to 28 h, the second-stage fermentation time is shortened from 18 h to 14 h, and the aerobic fermentation and anaerobic fermentation times are also shortened to 12 h and 20 h respectively, although the indicators are slightly lower than those of Example 1, the amino acid nitrogen content is still 0.10 g / L, the ester proportion is 52.1%, and the total acid content is 8.2%, which are all much better than all the comparative examples, indicating that under the complete process framework, appropriately shortening the fermentation time can provide the possibility for optimizing the production efficiency on the premise of ensuring the product quality, and will not cause a substantial decline in quality.

[0062] The above only describes the preferred embodiments of the present application, and it should be noted that for ordinary skilled persons in the art, without departing from the principles of the present application, a number of improvements and refinements can be made, and these improvements and refinements should also be considered as the protection scope of the present application.

Claims

1. A method for processing soy-sour soup based on fermentation of soybean whey, characterized in that, Includes the following steps: S1. Pretreatment of soybean liquid: Filter soybean liquid to remove insoluble impurities, adjust the dry matter concentration to 5-8°Bx, add glucose as the initial carbon source, pasteurize at 75-85℃ for 15-30 min and then cool to 30-35℃. S2. First-stage enzymatic hydrolysis and flavor-enhancing fermentation: Inoculate the soybean liquid with a mixed strain of Aspergillus oryzae and Aspergillus soysarum, and ferment at 30-35℃ for 0.3-0.5 m. 3 / (m 3 • h) Ferment for 24-48 hours under low-intensity aeration conditions until the amino acid nitrogen (AAN) content in the fermentation broth reaches 0.3-0.5 g / L; S3. Second stage of acid-producing and aroma-enhancing fermentation: Without sterilization, a mixed strain of Lactobacillus delbrueckii subsp. bulgaricus, Streptococcus thermophilus, and Lactobacillus paracasei is directly inoculated into the first stage fermentation broth. Fermentation is carried out at 37-40℃ under sealed anaerobic conditions for 12-24 hours until the pH of the fermentation broth drops to 4.0-4.

3. S4. Third stage debittering and aroma-enhancing fermentation: Centrifuge and sterilize the second stage fermentation liquid, add 0.5%-1% glucose, then inoculate with Bacillus berberis and Bacillus succinate, ferment at 28-30℃ with aeration and stirring for 12-20 hours, then inoculate with Kluyveromyces martensii and Saccharomyces cerevisiae, and ferment at 30-32℃ under static anaerobic conditions for 20-24 hours; S5. After fermentation, the fermentation liquid of the third stage is separated into solid and liquid components, heated to 85-95℃ for 10-15 minutes for sterilization, and the fermented liquid is obtained as soy soup product.

2. The method for processing soy milk soup based on fermentation of soybean whey as described in claim 1, characterized in that: The preparation method of the soybean whey is as follows: Select plump, pest-free, high-quality green beans and soak them in water at a temperature of 18-22℃. The weight of the soaked green beans should be about 2.0 to 2.2 times that before soaking. Grind the soaked green beans with water to obtain a paste. The slurry obtained from grinding is steam-heated to 40-60℃, defoamed, and then steam-heated to boiling. After boiling, it is simmered for 3-5 minutes, and then squeezed and separated three times with a filter bag to obtain soy milk. Adjust the concentration of the soy milk to 6°Bx and cool it to 75-80℃. Mix the cooled soy milk with the fermented soy milk at a mass ratio of 3:1 until flocculent precipitate appears in the soy milk. Then heat for 1-2 minutes, keep warm and let stand to obtain coagulated soy milk. Press the coagulated soy milk with a heavy object and collect the squeezed-out liquid to obtain soy milk liquid. The fermented soy milk liquid is made by sealed fermentation of the soy milk liquid. The total acid content of the fermented soy milk liquid is 5.0-5.5g / kg.

3. The method for processing sour soybean soup based on fermentation of soybean whey as described in claim 1, characterized in that: The filtration in step S1 uses a 100-200 mesh sieve; the amount of glucose added is 1.5%-2.5% of the mass of the soybean extract.

4. The method for processing sour soybean soup based on fermentation of soybean whey as described in claim 1, characterized in that: In step S2, the mass ratio of Aspergillus oryzae to Aspergillus saurum is 8:2, and the inoculation amount is 0.5‰-1.0‰ of the total mass of soybean extract. The Aspergillus oryzae strain is selected with aminopeptidase activity ≥500U / g, and the Aspergillus saurum strain is selected with protease activity ≥300U / g.

5. The method for processing soy milk soup based on fermentation of soybean whey as described in claim 1, characterized in that: In step S3, the mass ratio of *Lactobacillus delbrueckii* subsp. bulgaricus, *Streptococcus thermophilus*, and *Lactobacillus paracasei* is 1:1:0.

2. Before use, the bacteria are reactivated in sterile soybean whey at 37-40℃ for 1-2 hours, and the inoculation amount is 1.0%-2.0% of the fermentation liquid volume. The *Lactobacillus delbrueckii* subsp. bulgaricus and *Lactobacillus paracasei* strains are selected with an acid production rate ≥2.0 g / (L·h), and the *Streptococcus thermophilus* strain is selected with an acetaldehyde production rate ≥5.0 mg / L.

6. The method for processing sour soybean soup based on fermentation of soybean whey as described in claim 1, characterized in that: In step S4, during aerobic fermentation, the inoculum size of *Bacillus belye* is 0.5%-1.0% of the fermentation broth volume, and the inoculum size of *Gluconobacterium tumefaciens* is 0.2%-0.4% of the fermentation broth volume. The specific aeration and stirring conditions are: an aeration rate of 0.5-1.0 m³ / s. 3 / (m 3 •h), the stirring method is to stir at 60rpm for 10-15min every 2h.

7. The method for processing sour soybean soup based on fermentation of soybean whey as described in claim 1, characterized in that: In step S4, during anaerobic fermentation, both the *Kluyveromyces martensii* and *Saccharomyces cerevisiae* are activated using sterile water containing 2%-3% (by mass) glucose at 30℃-32℃ for 2-3 hours. After activation, the bacterial activity of the culture is ≥10^8 CFU / mL. The inoculum size of the *Kluyveromyces martensii* is 0.5%-1.0% of the fermentation liquid volume, and the *Saccharomyces cerevisiae* is an edible strain with an amino acid assimilation rate ≥80%, with an inoculum size of 2.0%-3.0% of the fermentation liquid volume.

8. The method for processing sour soybean soup based on fermentation of soybean whey as described in claim 1, characterized in that: In step S4, the specific operations for centrifugation and sterilization are as follows: the second-stage fermentation broth is hot-filtered using a 200-400 mesh sieve and the filtrate is collected. The filtration temperature is 40-50℃. Then, the filtrate is centrifuged at 8000 rpm for 10 minutes, and the supernatant is collected. The supernatant is then subjected to ultra-high temperature instantaneous sterilization at 115-121℃ for 8-15 seconds, and then rapidly cooled to 28-30℃ to obtain the initial fermentation broth for the third-stage fermentation.

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