Composite enzyme preparation, preparation method and application thereof in yak lung tissue tenderization

Abstract

The application belongs to the technical field of deep processing of livestock and poultry meat products, and discloses a compound enzyme preparation, a preparation method and application of the compound enzyme preparation in yak lung tissue tenderization. The raw materials of the compound enzyme preparation include: compound protease, TG enzyme microcapsules, wheat bran-soybean fermentation extract and a protective agent. The TG enzyme microcapsules are obtained by embedding TG enzyme with compound polysaccharide. The preparation method of the wheat bran-soybean fermentation extract is as follows: wheat bran and soybean are mixed, water is added for beating, Bacillus subtilis is inoculated into the slurry for fermentation, and then the filtrate is obtained by filtration, concentrated and dried. The compound enzyme preparation is used in yak lung tissue tenderization, can effectively maintain the integrity of the tissue structure while ensuring sufficient tenderization, and realizes tender and not rotten, elastic and not hard products.

Description

Technical Field

[0001] This invention relates to the field of deep processing technology for livestock and poultry meat products, specifically to a compound enzyme preparation, its preparation method, and its application in tenderizing yak lung tissue. Background Technology

[0002] Yak are a rare and unique livestock and poultry germplasm resource in my country. Their lung tissue is rich in protein, collagen, and various bioactive components, possessing high nutritional value and development potential. However, due to long-term adaptation to the low-oxygen environment of high altitudes, yak lung tissue has well-developed connective tissue, dense collagen fiber arrangement, and high elastin content, resulting in a tough texture and rough mouthfeel. This severely restricts the development and market acceptance of deep-processed yak lung products.

[0003] Currently, methods for tenderizing animal internal organs mainly include physical methods (mechanical tenderizing, ultrasonic tenderizing), chemical methods (calcium salt tenderizing, phosphate treatment), and biological enzymatic methods. Among them, enzymatic tenderizing has attracted much attention due to its mild operating conditions, high specificity, and high safety. Commonly used tenderizing enzymes include papain, bromelain, and neutral protease. These proteases can degrade proteins in muscle and connective tissues, reduce shear forces, and improve tenderness.

[0004] However, existing enzymatic tenderizing methods have obvious technical defects: First, they mostly use single proteases or simple protease combinations, resulting in a single degradation dimension and poor tenderizing effect on the dense connective tissue of yak lungs with high cross-linking and high toughness; Second, simple protease hydrolysis is prone to over-enzymatic hydrolysis, leading to loose tissue structure, softness and water leakage, which seriously affects the texture characteristics and commercial value of the product.

[0005] In view of this, this invention is hereby proposed. Summary of the Invention

[0006] The purpose of this invention is to overcome the shortcomings of the prior art and provide a compound enzyme preparation, its preparation method, and its application in the tenderization of yak lung tissue. This enzyme preparation utilizes the synergistic effect of a compound protease, TG enzyme microcapsules, and wheat bran-soybean fermentation extract to achieve efficient tenderization while repairing the loosened tissue structure caused by excessive enzymatic hydrolysis, thereby preventing the occurrence of soft, mushy, and watery tissue.

[0007] To achieve the above objectives, the first aspect of the present invention provides a compound enzyme preparation, the raw materials of which include: Complex protease, TG enzyme microcapsules, wheat bran-soybean fermentation extract, and protectant; among which, The TG enzyme microcapsules are obtained by encapsulating TG enzyme with complex polysaccharides. The preparation method of the wheat bran-soybean fermentation extract is as follows: wheat bran and soybeans are mixed with water and pulped, Bacillus subtilis is inoculated into the pulp for fermentation, and then the filtrate is concentrated and dried.

[0008] Preferably, the mass ratio of the complex protease, TG enzyme microcapsules, wheat bran-soybean fermentation extract, and protectant is 5-7:2-4:3-5:0.5-0.8.

[0009] Preferably, the complex protease is obtained by combining two or more of papain, bromelain, fig protease, flavor protease, and neutral protease.

[0010] Preferably, the complex polysaccharide comprises sodium carboxymethyl cellulose, soluble soybean polysaccharide, and sodium alginate, with a mass ratio of 4-6:3-5:1.

[0011] Preferably, the mass ratio of wheat bran to soybeans is 2-3:1.

[0012] Preferably, the inoculum amount of Bacillus subtilis is 2-5%.

[0013] Preferably, the fermentation temperature is 30-37 ℃ and the time is 48-60 h.

[0014] A second aspect of this invention provides a method for preparing the above-mentioned complex enzyme preparation, comprising the following steps: The complex protease was added to water, mixed evenly, and then a protectant was added. After homogenization, the mixture was dried to obtain the pretreated protease. A compound enzyme preparation was obtained by combining pretreated protease with TG enzyme microcapsules and wheat bran-soybean fermentation extract.

[0015] The third aspect of this invention provides the application of the above-mentioned compound enzyme preparation in the tenderization of yak lung tissue.

[0016] Preferably, the amount of the compound enzyme preparation is 0.1-0.3% of the mass of yak lung tissue.

[0017] The beneficial effects of this invention are as follows: The compound enzyme preparation provided by this invention, with the synergistic effect of the compound protease and the protease in the wheat bran-soybean fermentation extract, can effectively achieve high-efficiency tenderization of yak lung tissue. At the same time, the cellulase, hemicellulase and other enzyme systems in the fermentation extract act simultaneously on the composite polysaccharide wall material of the TG enzyme microcapsules, allowing the embedded TG enzyme to be slowly released, catalyzing the cross-linking of small molecule proteins and peptides produced by enzymatic hydrolysis, repairing the loose tissue structure, and thus achieving a product that is tender but not mushy, and elastic but not hard.

[0018] This invention integrates a multi-enzyme synergistic system into a compound formulation, which requires only a single addition and does not require step-by-step control. It is simple to operate, has mild conditions, and is easy to achieve continuous industrial production. Detailed Implementation

[0019] To make the objectives, technical solutions, and advantages of this invention clearer, the following detailed description of the invention is provided in conjunction with embodiments. Those skilled in the art will understand that the following embodiments are merely exemplary and not intended to limit the scope of the invention. Furthermore, in the following description, unless specific conditions are specified in the embodiments, conventional conditions or conditions recommended by the manufacturer are followed. Reagents or instruments whose manufacturers are not specified are all commercially available conventional products.

[0020] The first embodiment of the present invention provides a compound enzyme preparation, the raw materials of which include: a compound protease, TG enzyme microcapsules, wheat bran-soybean fermentation extract, and a protective agent; wherein, The TG enzyme microcapsules are obtained by encapsulating TG enzyme with complex polysaccharides. The preparation method of the wheat bran-soybean fermentation extract is as follows: wheat bran and soybeans are mixed with water and pulped, Bacillus subtilis is inoculated into the pulp for fermentation, and then the filtrate is concentrated and dried.

[0021] In this embodiment, the complex protease is used to hydrolyze myofibril proteins and connective tissues (collagen, elastin, etc.) in yak lung tissue, causing the tough muscle fiber structure to break down, thereby reducing shear force and toughness, and achieving tenderization. To ensure the sufficiency and balance of the tenderizing effect, the complex protease is obtained by blending two or more of papain, bromelain, fig protease, flavor protease, and neutral protease in an appropriate ratio.

[0022] The TG enzyme is encapsulated with a complex polysaccharide. This serves two purposes: firstly, to prevent premature inactivation or premature cross-linking of the substrate when coexisting with the complex protease, which would negatively impact the tenderizing effect; and secondly, to achieve controlled, sustained release of the TG enzyme. The microcapsule wall material gradually absorbs water and swells in the aqueous system, decreasing its density, allowing the encapsulated TG enzyme to slowly diffuse into the system through the pores. Simultaneously, the polysaccharide-degrading enzyme in the wheat bran-soybean fermentation extract further promotes the loosening and disintegration of the wall material structure, accelerating the release of the TG enzyme. Under the combined effect of these mechanisms, the release of TG enzyme is relatively low in the early stages of tenderization (when the tenderizing effect is primarily proteolytic), but the release rate gradually increases as the tenderizing process progresses (as the cross-linking repair effect gradually strengthens), thus achieving a temporal synergy between tenderization and cross-linking repair.

[0023] In a specific implementation, to ensure the encapsulation effect of TG enzyme and its slow release in the later stage, the complex polysaccharide includes sodium carboxymethyl cellulose, soluble soybean polysaccharide and sodium alginate, with a mass ratio of 4-6:3-5:1. More specifically, the complex polysaccharides are added to water and stirred at 45-55 ℃ until the three polysaccharides are fully swollen and dissolved. After standing for 30-45 min to remove bubbles, a wall material solution with a total polysaccharide mass concentration of 6-10% is obtained. TG enzyme powder is added to water to prepare a TG enzyme solution with a mass concentration of 15-20%, which is then added dropwise to the above wall material solution. The dry basis mass ratio of TG enzyme to complex polysaccharide wall material is controlled at 1:4-6. After stirring at 40-45 ℃ for 1.5-2 h, spray drying and encapsulation are performed (inlet air temperature 160-180 ℃, outlet air temperature 75-85 ℃, feed rate 8-12 mL / min). The dried microcapsule powder is passed through an 80-120 mesh sieve to remove large particles and adhering substances, thus obtaining TG enzyme microcapsules with the desired uniform particle size.

[0024] Bacillus subtilis is a bacterial strain capable of fermenting and producing enzyme systems such as cellulase, hemicellulase, and protease. Fermenting wheat bran and soybean with Bacillus subtilis yields a wheat bran-soybean fermentation extract containing a significant amount of these enzymes. These enzymes hydrolyze the TG enzyme microcapsule wall material, particularly sodium carboxymethyl cellulose and soluble soybean polysaccharides, during the tenderizing process, thus slowly releasing the TG enzyme. Furthermore, the wheat bran-soybean fermentation extract also contains a small amount of protease, which can synergistically enhance the tenderizing effect; it also contains fermentation-produced small molecule peptides and free amino acids, serving as flavor precursors and improving the overall flavor of the product.

[0025] In a specific implementation, wheat bran and soybeans are mixed at a mass ratio of 2-3:1, and 4-6 times their total mass of water is added to make a pulp. Then, Bacillus subtilis (with a viable count ≥10) is inoculated at a rate of 2-5%. 8 The seed liquid (CFU / g) is fermented at 30-37 ℃ for 48-60 h. The filtrate is filtered and concentrated to 1 / 4-1 / 3 of its original volume, then spray-dried. The inlet air temperature is controlled at 160-180 ℃, the outlet air temperature at 75-85 ℃, and the feed rate at 8-12 mL / min. The dried powder is passed through an 80-120 mesh sieve to obtain the desired wheat bran-soybean fermentation extract. It should be noted that the wheat bran used in this embodiment is edible wheat bran conforming to GB / T 42225-2022 "Wheat Bran" standard, and the Bacillus subtilis used is Bacillus subtilis DE111, which is suitable for food use.

[0026] Protectants are used to maintain the activity stability of complex proteases during storage and use, and to prevent the enzyme protein from denaturing and becoming inactive. Their composition and ratio can be adjusted according to actual production and application needs. For example, two or more of trehalose, sodium chloride, and maltodextrin can be selected and compounded in an appropriate ratio.

[0027] The mass ratio of the above-mentioned compound protease, TG enzyme microcapsules, wheat bran-soybean fermentation extract, and protectant is 5-7:2-4:3-5:0.5-0.8. This ratio can achieve full degradation of the coarse and hard muscle fibers and connective tissue of yak lung tissue, ensuring the tenderizing effect, while completing appropriate protein cross-linking repair, resulting in yak lung tissue that is tender but not rotten and elastic but not hard.

[0028] The second embodiment of the present invention provides a method for preparing the above-mentioned complex enzyme preparation, comprising the following steps: Add the complex protease to 2.5-3.5 times its weight of water, mix well, then add a protectant, homogenize 1-2 times under 25-30 MPa pressure, and then spray dry under the following conditions: inlet air temperature 150-170℃, outlet air temperature 70-80℃, and feed rate 10-15 mL / min to obtain the pretreated protease. A compound enzyme preparation was prepared by combining pretreated protease with TG enzyme microcapsules and wheat bran-soybean fermentation extract. The preparation was sealed in aluminum foil bags and stored in a cool, dry place.

[0029] The third embodiment of the present invention provides the application of the above-mentioned compound enzyme preparation in the tenderization of yak lung tissue.

[0030] The compound enzyme preparation of this invention can be used in the deep processing of yak lung tissue to achieve uniform tenderization of yak lung tissue, while repairing some of the loosened tissue structure caused by enzymatic hydrolysis, thus preventing the occurrence of soft, mushy, and watery tissue. The specific application method is as follows: Fresh yak lung tissue is cleaned and then cut into blocks or strips with a thickness of 1-3 cm. Weigh 0.1-0.3% of the compound enzyme preparation by weight of yak lung tissue and 3-5 times the amount of water. Mix the two evenly and add them to the cut yak lung tissue. Let it stand at 25-30 ℃ for 50-65 min for enzymatic hydrolysis. After enzymatic hydrolysis, the yak lung tissue is retrieved and subjected to enzyme inactivation treatment to obtain high-quality tenderized yak lung tissue. The enzyme inactivation treatment conditions can be routinely adjusted according to the actual enzyme system composition and enzyme dosage used, with the aim of completely inactivating all enzyme components in the system. As an example, the enzyme inactivation treatment temperature is 95-100 ℃, and the time is 5-10 min.

[0031] It should be noted that the compound enzyme preparation of this embodiment is not only suitable for tenderizing yak lung tissue, but can also be applied to other animal offal rich in dense connective tissue, such as beef tripe, pig lungs, and sheep tripe. It can effectively prevent softness and water loss while ensuring tenderness, and improve the texture and water retention of the product.

[0032] The following will disclose specific embodiments for implementing the present invention, as well as corresponding comparative examples to demonstrate the relevant technical effects of the compound enzyme preparation of the present invention.

[0033] All raw materials used in the following examples were purchased commercially; the wheat bran used was edible wheat bran, conforming to GB / T 42225-2022 "Wheat Bran" standard; the proteases and TG enzymes used were food-grade enzyme preparations, wherein the enzyme activities of flavor protease and neutral protease were both 5×10⁻⁶. 4 U / g, the enzyme activities of papain, bromelain, and fig protease were all 10. 5 The enzyme activity of TG enzyme is 5 × 10 U / g. 4 U / g; Bacillus subtilis is essentially a seed culture of Bacillus subtilis DE111, with a viable count of 10. 8 CFU / g.

[0034] Example 1 Experimental group: Preparation of compound enzyme preparations (1) Preparation of complex protease A complex protease was obtained by mixing papain and neutral protease at a mass ratio of 2:1.

[0035] (2) Preparation of TG enzyme microcapsules Sodium carboxymethyl cellulose, soluble soybean polysaccharide, and sodium alginate were mixed in a mass ratio of 4:3:1, added to water, and stirred at 45 °C until the three polysaccharides were fully swollen and dissolved. After standing for 30 min to remove bubbles, a wall material solution with a total polysaccharide mass concentration of 6% was obtained. Add TG enzyme powder to water to prepare a TG enzyme solution with a mass concentration of 15%. The above-mentioned TG enzyme solution was added dropwise to the wall material solution, and the dry weight ratio of TG enzyme to composite polysaccharide wall material was controlled at 1:4. After stirring at 40 ℃ for 1.5 h, spray drying and encapsulation were carried out. The spray drying conditions were: inlet air temperature 160 ℃, outlet air temperature 75 ℃, and feed rate 12 mL / min. The dried microcapsule powder was passed through an 80-mesh sieve to remove large particles and adhering substances, and TG enzyme microcapsules were obtained.

[0036] (3) Preparation of wheat bran-soybean fermentation extract Wheat bran and soybeans were mixed at a mass ratio of 2:1, and water was added in four times their total mass to form a pulp. Then, Bacillus subtilis was inoculated at a 2% inoculation rate, and fermented at 30 °C for 48 h. The filtrate was filtered and concentrated to 1 / 4 of its original volume, and then spray-dried. The inlet air temperature was controlled at 160 °C, the outlet air temperature at 75 °C, and the feed rate at 12 mL / min. The dried powder was passed through an 80-mesh sieve to obtain the wheat bran-soybean fermented extract.

[0037] (4) Preparation of protective agent Trehalose and sodium chloride are mixed in a mass ratio of 2:1 to obtain a protective agent.

[0038] (5) Preparation of compound enzyme preparations Take the above-mentioned compound protease, TG enzyme microcapsules, wheat bran-soybean fermentation extract and protectant in a mass ratio of 5:2:3:0.5; The complex protease was added to 2.5 times its weight of water, mixed evenly, and then a protectant was added. The mixture was homogenized once under a pressure of 25 MPa, and then spray-dried under the following conditions: inlet air temperature 150 ℃, outlet air temperature 70 ℃, and feed rate 10 mL / min to obtain the pretreated protease. A compound enzyme preparation was prepared by combining pretreated protease with TG enzyme microcapsules and wheat bran-soybean fermentation extract. The preparation was then sealed in aluminum foil bags and stored in a cool, dry place.

[0039] Based on the above experimental groups, the following control group was set up: Control group 1: No TG enzyme microcapsules or wheat bran-soybean fermentation extract were added; the pretreated protease was the compound enzyme preparation.

[0040] Control group 2: No TG enzyme microcapsules added.

[0041] Control group 3: No wheat bran-soybean fermented extract added.

[0042] Control group 4: TG enzyme powder was added directly without using TG enzyme microcapsules, and the amount added was consistent with the TG enzyme dry matter content in the TG enzyme microcapsules of the experimental group.

[0043] Control group 5: No sodium alginate was added during the preparation of TG enzyme microcapsules.

[0044] The compound enzyme preparations from the above experimental and control groups were used to treat yak lung tissue. The specific treatment method was as follows: Fresh yak lung tissue is cleaned and cut into strips with a thickness of 1 cm; Weigh out 0.1% of the compound enzyme preparation by weight of yak lung tissue and 3 times the amount of water. Mix the two evenly and add them to the cut yak lung tissue. Let it stand at 25 ℃ for 50 min for enzymatic hydrolysis. After enzymatic hydrolysis, the yak lung tissue is removed and treated with enzyme inactivation at 95 ℃ for 5 min to obtain tenderized yak lung tissue.

[0045] The following indicators were measured on the lung tissue of tenderized yak treated with various groups of compound enzyme preparations: (1) Sensory evaluation: A 9-point scoring system was used, with the sensory evaluation team scoring the tenderness, elasticity, juiciness, and flavor of the samples. The sensory evaluation team consisted of 10 trained food professionals (aged 20-45, half male and half female). The tenderized yak lung tissue samples from each treatment group were steamed until the core temperature reached 75°C, then cut into uniform small pieces, randomly numbered, and presented to the evaluation team members. Evaluators were required to rinse their mouths with purified water before evaluation to eliminate residual taste, and there was a 30-second interval between each two samples. Fresh yak lung tissue without tenderization treatment was used as a blank control. The evaluation criteria are shown in Table 1, and the results are shown in Table 2.

[0046] Table 1 Sensory Evaluation Criteria for Yak Lung Tissue .

[0047] Table 2 Sensory evaluation results of tenderized yak lung tissue .

[0048] (2) Water holding capacity determination: The centrifugation method was used for determination. A sample of tenderized yak lung tissue (mass denoted as M1) was weighed and placed in a 50 mL centrifuge tube, and centrifuged at 3000 r / min for 15 min at 4℃. After centrifugation, the sample was removed, and the surface free water was gently absorbed with filter paper before weighing (denoted as M2). The water holding capacity was then calculated according to the formula: Water holding capacity (%) = M2 / M1 × 100%. Fresh yak lung tissue without tenderization treatment was used as a blank control. The results are shown in Table 3.

[0049] (3) Texture determination: including hardness, elasticity, cohesiveness and chewiness, were determined using a texture analyzer. The determination conditions were: probe P / 50, pre-test speed 2.0 mm / s, test speed 1.0 mm / s, post-test speed 1.0 mm / s, compression ratio 50%, trigger force 5.0 g, interval between two compressions 5.0 s, and data acquisition rate 200 pps. Each sample was measured in parallel at least 6 times, and the average value was taken. Fresh yak lung tissue without tenderization treatment was used as a blank control. The results are shown in Table 3.

[0050] (4) Shear force determination: The shear force was determined using a texture analyzer. The determination conditions were as follows: probe model HDP / BSW (V-shaped shear blade), pre-test speed 2.0 mm / s, test speed 1.0 mm / s, post-test speed 10.0 mm / s, trigger force 5.0 g, and shearing distance 30 mm. The tenderized yak lung tissue was placed on the texture analyzer test platform, and the blade was used to cut perpendicular to the muscle fiber direction. At least 6 parallel samples were measured for each group of samples, and the average value was taken. Fresh yak lung tissue without tenderization treatment was used as a blank control. The results are shown in Table 3. The smaller the shear force value, the better the tenderness of the sample.

[0051] Table 3. Results of water holding capacity, texture and shear force determination of lung tissue from tenderized yak. .

[0052] According to the results in Tables 2 and 3, the experimental group was significantly better than all control groups in terms of elasticity (sensory score 8.2, texture 0.89), juiciness (8.4), water retention (86.7%), and cohesion (0.75), which characterize tissue integrity and water retention capacity. Although control groups 1 and 2 were slightly better than the experimental group in tenderness score and shear force, their elasticity, water retention, and cohesion were significantly lower than those of the experimental group. This indicates that simply pursuing extreme tenderness will lead to excessively loose tissue structure and severe juice loss, failing to achieve the ideal texture quality. Control group 3 lacked the wall-degrading enzymes provided by wheat bran-soybean fermentation extract, making it difficult for the TG enzyme microcapsules to disintegrate and hindering the cross-linking repair function, resulting in unsatisfactory water retention and tenderness. Control group 4 used direct addition of free TG enzyme, which acted synchronously with protease in the early stages of enzymatic hydrolysis. The dense network formed by cross-linking severely hindered the penetration of protease into the tissue, resulting in a product that was hard on the outside and tough on the inside, with uneven tenderization. Control group 5 lacked sodium alginate in its microcapsule wall material, leading to decreased film-forming properties and structural stability. The TG enzyme was released too quickly and unevenly, resulting in a product with significantly inferior overall quality compared to the experimental groups. In contrast, this invention effectively maintained the integrity of the tissue structure while ensuring sufficient tenderization, achieving a good balance between tenderness and texture. The sample was tender but not mushy, and elastic but not hard, with the best overall quality.

[0053] Example 2 (1) Preparation of complex protease A complex protease was obtained by mixing papain, bromelain, and flavor protease in a mass ratio of 1:1:1.

[0054] (2) Preparation of TG enzyme microcapsules Sodium carboxymethyl cellulose, soluble soybean polysaccharide, and sodium alginate were mixed in a mass ratio of 5:5:1, added to water, and stirred at 50 °C until the three polysaccharides were fully swollen and dissolved. After standing for 40 min to remove bubbles, a wall material solution with a total polysaccharide mass concentration of 8% was obtained. Add TG enzyme powder to water to prepare a TG enzyme solution with a mass concentration of 18%. The above-mentioned TG enzyme solution was added dropwise to the wall material solution, and the dry basis mass ratio of TG enzyme to composite polysaccharide wall material was controlled at 1:5. After stirring at 42 ℃ for 1.8 h, spray drying and encapsulation were carried out. The spray drying conditions were: inlet air temperature 170 ℃, outlet air temperature 85 ℃, and feed rate 10 mL / min. The dried microcapsule powder was passed through a 100-mesh sieve to remove large particles and adhering substances, and TG enzyme microcapsules were obtained.

[0055] (3) Preparation of wheat bran-soybean fermentation extract Wheat bran and soybeans were mixed at a mass ratio of 2.5:1, and water was added in 5 times their total mass to make a pulp. Then, Bacillus subtilis was inoculated at a 3% inoculation rate, and fermented at 33 ℃ for 60 h. The filtrate was filtered and concentrated to 1 / 4 of its original volume, and then spray-dried. The inlet air temperature was controlled at 170 ℃, the outlet air temperature at 85 ℃, and the feed rate at 10 mL / min. The dried powder was passed through a 100-mesh sieve to obtain the wheat bran-soybean fermented extract.

[0056] (4) Preparation of protective agent Trehalose and maltodextrin were mixed at a mass ratio of 2:1 to obtain a protective agent.

[0057] (5) Preparation of compound enzyme preparations Take the above-mentioned compound protease, TG enzyme microcapsules, wheat bran-soybean fermentation extract and protectant in a mass ratio of 7:3:4:0.7; The complex protease was added to three times its weight of water, mixed evenly, and then a protectant was added. The mixture was homogenized twice under a pressure of 28 MPa, and then spray-dried under the following conditions: inlet air temperature 160 ℃, outlet air temperature 75 ℃, and feed rate 15 mL / min to obtain the pretreated protease. A compound enzyme preparation was prepared by combining pretreated protease with TG enzyme microcapsules and wheat bran-soybean fermentation extract. The preparation was then sealed in aluminum foil bags and stored in a cool, dry place.

[0058] The obtained compound enzyme preparation was used to treat yak lung tissue. The specific treatment method was as follows: Fresh yak lung tissue is cleaned and cut into strips 2 cm thick; Weigh out 0.2% of the compound enzyme preparation by weight of yak lung tissue and 4 times the amount of water, mix them evenly, add them to the cut yak lung tissue, and let it stand at 28 ℃ for 65 min for enzymatic hydrolysis. After enzymatic hydrolysis, the yak lung tissue is removed and treated with enzyme inactivation at 98 ℃ for 10 min to obtain tenderized yak lung tissue.

[0059] Sensory evaluation, water holding capacity, texture and shear force were performed on the tenderized yak lung tissue obtained above. The tenderness, elasticity, juiciness and flavor scores were 8.7, 8.3, 8.5 and 8.4, respectively. The water holding capacity was 87.3%, the hardness was 16.18 N, the elasticity was 0.9, the cohesion was 0.76 and the shear force was 17.74 N.

[0060] Example 3 (1) Preparation of complex protease A complex protease was obtained by mixing papain, fig protease, bromelain, and neutral protease in a mass ratio of 1:1:1:1.

[0061] (2) Preparation of TG enzyme microcapsules Sodium carboxymethyl cellulose, soluble soybean polysaccharide, and sodium alginate were mixed in a mass ratio of 6:4:1, added to water, and stirred at 55 °C until the three polysaccharides were fully swollen and dissolved. After standing for 45 min to remove bubbles, a wall material solution with a total polysaccharide mass concentration of 10% was obtained. Add TG enzyme powder to water to prepare a TG enzyme solution with a mass concentration of 20%. The above-mentioned TG enzyme solution was added dropwise to the wall material solution, and the dry basis mass ratio of TG enzyme to composite polysaccharide wall material was controlled at 1:6. After stirring at 45 ℃ for 2 h, spray drying and encapsulation were carried out. The spray drying conditions were: inlet air temperature 180 ℃, outlet air temperature 80 ℃, and feed rate 8 mL / min. The dried microcapsule powder was passed through a 120-mesh sieve to remove large particles and adhering substances, and TG enzyme microcapsules were obtained.

[0062] (3) Preparation of wheat bran-soybean fermentation extract Wheat bran and soybeans were mixed at a mass ratio of 3:1, and water was added in six times their total mass to form a pulp. Then, Bacillus subtilis was inoculated at a 5% inoculum and fermented at 37 ℃ for 54 h. The filtrate was filtered and concentrated to 1 / 3 of its original volume before being spray-dried. The inlet air temperature was controlled at 180 ℃, the outlet air temperature at 80 ℃, and the feed rate at 8 mL / min. The dried powder was passed through a 120-mesh sieve to obtain the wheat bran-soybean fermented extract.

[0063] (4) Preparation of protective agent Trehalose, sodium chloride, and maltodextrin were mixed in a mass ratio of 1:1:1 to obtain a protective agent.

[0064] (5) Preparation of compound enzyme preparations Take the above-mentioned compound protease, TG enzyme microcapsules, wheat bran-soybean fermentation extract and protectant in a mass ratio of 6:4:5:0.8; The complex protease was added to 3.5 times its weight of water, mixed evenly, and then a protectant was added. The mixture was homogenized once under a pressure of 30 MPa, and then spray-dried under the following conditions: inlet air temperature 170 ℃, outlet air temperature 80 ℃, and feed rate 12 mL / min to obtain the pretreated protease. A compound enzyme preparation was prepared by combining pretreated protease with TG enzyme microcapsules and wheat bran-soybean fermentation extract. The preparation was then sealed in aluminum foil bags and stored in a cool, dry place.

[0065] The obtained compound enzyme preparation was used to treat yak lung tissue. The specific treatment method was as follows: Fresh yak lung tissue is cleaned and cut into strips with a thickness of 3 cm. Weigh out 0.3% of the compound enzyme preparation of yak lung tissue and 5 times the amount of water, mix them evenly and add them to the cut yak lung tissue, and let it stand at 30 ℃ for 55 min for enzymatic hydrolysis. After enzymatic hydrolysis, the yak lung tissue is removed and treated with enzyme inactivation at 100 ℃ for 8 min to obtain tenderized yak lung tissue.

[0066] Sensory evaluation, water holding capacity, texture and shear force were performed on the tenderized yak lung tissue obtained above. The tenderness, elasticity, juiciness and flavor scores were 8.6, 8.4, 8.6 and 8.5, respectively. The water holding capacity was 88.5%, the hardness was 15.37 N, the elasticity was 0.91, the cohesion was 0.77 and the shear force was 17.14 N.

[0067] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features therein. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A compound enzyme preparation, characterized in that, Raw materials include: Complex protease, TG enzyme microcapsules, wheat bran-soybean fermentation extract, and protectant; among which, The TG enzyme microcapsules are obtained by encapsulating TG enzyme with complex polysaccharides. The preparation method of the wheat bran-soybean fermentation extract is as follows: wheat bran and soybeans are mixed with water and pulped, Bacillus subtilis is inoculated into the pulp for fermentation, and then the filtrate is concentrated and dried.

2. The compound enzyme preparation as described in claim 1, characterized in that, The mass ratio of the compound protease, TG enzyme microcapsules, wheat bran-soybean fermentation extract, and protectant is 5-7:2-4:3-5:0.5-0.

8.

3. The compound enzyme preparation as described in claim 1, characterized in that, The complex protease is obtained by combining two or more of papain, bromelain, fig protease, flavor protease, and neutral protease.

4. The compound enzyme preparation according to claim 1, characterized in that, The complex polysaccharide comprises sodium carboxymethyl cellulose, soluble soybean polysaccharide, and sodium alginate, with a mass ratio of 4-6:3-5:

1.

5. The compound enzyme preparation as described in claim 1, characterized in that, The mass ratio of wheat bran to soybeans is 2-3:

1.

6. The compound enzyme preparation according to claim 1, characterized in that, The inoculation amount of Bacillus subtilis is 2-5%.

7. The compound enzyme preparation according to claim 1, characterized in that, The fermentation temperature is 30-37℃ and the time is 48-60 h.

8. The method for preparing the complex enzyme preparation according to any one of claims 1-7, characterized in that, Includes the following steps: The complex protease was added to water, mixed evenly, and then a protectant was added. After homogenization, the mixture was dried to obtain the pretreated protease. A compound enzyme preparation was obtained by combining pretreated protease with TG enzyme microcapsules and wheat bran-soybean fermentation extract.

9. The application of the compound enzyme preparation as described in any one of claims 1-7 in the tenderization of yak lung tissue.

10. The application as described in claim 9, characterized in that, The dosage of the compound enzyme preparation is 0.1-0.3% of the weight of yak lung tissue.