Bamboo shoot enzyme pulp based on bamboo shoot pulp enzymolysis and preparation method thereof

Abstract

The invention discloses bamboo shoot enzyme pulp based on bamboo shoot pulp enzymolysis and a preparation method thereof, and relates to the technical field of fermented food, the preparation method comprises the following steps: 1) bamboo shoot pretreatment and waste heat enzymolysis; 2) in-situ detoxification and fermentation; (3) filtering and seasoning; and 4) aseptic packaging and storage. Enzymolysis, directional detoxification and segmented fermentation are integrated, detoxification enzyme and a compound enzyme preparation are synchronously added, and the reaction is started by utilizing fixation waste heat, so that the technological process is shortened by 40% or above, and the energy consumption is remarkably reduced; through an in-situ detoxification fermentation mechanism, toxins such as tannin, oxalic acid and cyanoglycoside are synchronously degraded in a microbial metabolism process, and the safety and the efficiency are synchronously improved; by adopting the graded filtering and flexible blending technology, low-sugar and sugar-free products can be accurately produced on the same production line, the limitation of single product in the traditional process is broken through, and the unification of high efficiency, safety and multiple market demands is realized.

Description

Technical Field

[0001] This invention relates to the field of bamboo shoot pulp enzymatic hydrolysis technology, and in particular to a bamboo shoot enzyme pulp based on bamboo shoot pulp enzymatic hydrolysis and its preparation method. Background Technology

[0002] Bamboo shoots are rich in dietary fiber and various nutrients, but the utilization rate of their processing by-products is low, and the anti-nutritional factors such as tannins, oxalic acid, and cyanogenic glycosides contained in fresh bamboo shoots restrict their in-depth development.

[0003] Existing bamboo shoot enzyme preparation technologies face significant bottlenecks: some use processing residues as raw materials and employ simple mixed fermentation (e.g., CN105410936B), resulting in a lengthy process lacking targeted detoxification steps, with product safety dependent on auxiliary flavorings; others use single probiotic fermentation (e.g., CN108095077A), which improves some flavor, but the strains have limited functions and cannot synergistically degrade fiber and toxins, thus limiting product applications; still others introduce enzymatic hydrolysis but still use a sequential "enzymatic hydrolysis first, then fermentation" approach (e.g., CN116250628A), which suffers from problems such as fragmented processes, incomplete detoxification, and high energy consumption.

[0004] Furthermore, existing technologies struggle to simultaneously achieve efficient detoxification and flavor compound generation within the same system, and are unable to flexibly produce differentiated products that align with modern health trends, such as low-sugar and sugar-free products.

[0005] Therefore, existing technologies cannot achieve efficient, high-value, and diversified utilization of bamboo shoot resources while ensuring safety. Developing an integrated, precise, and flexible new method for preparing bamboo shoot enzymes is urgently needed. Summary of the Invention

[0006] The purpose of this invention is to propose a bamboo shoot enzyme slurry based on bamboo shoot slurry enzymatic hydrolysis and its preparation method, thereby overcoming the shortcomings of the existing technology.

[0007] To achieve the above objectives, the present invention adopts the following technical solution: A method for preparing bamboo shoot enzyme slurry based on bamboo shoot slurry enzymatic hydrolysis includes the following steps: S1. Bamboo shoot pretreatment and residual heat enzymatic hydrolysis: After cleaning and peeling fresh tender bamboo shoots, they are steamed at 105℃ for 3-5 minutes to blanch them, then crushed and pulped to obtain bamboo shoot pulp. High temperature instantaneous inactivation of endogenous enzymes in bamboo shoots, such as polyphenol oxidase, prevents browning and flavor deterioration; at the same time, it has a pasteurization effect, which greatly reduces the number of initial miscellaneous bacteria. Add purified water to adjust the material-to-liquid ratio to 1:0.8-1.2 and adjust the pH to 4.5-5.0. When the material-to-liquid temperature drops to 48-52℃, add compound enzyme preparation and compound detoxification enzyme premix. Enzymatic hydrolysis is carried out for 4-6 hours, with stirring 2-3 times during the process, each time for 10 minutes. After enzymatic hydrolysis, the enzyme preparation is inactivated by incubation at 90℃ for 15 minutes to obtain enzymatically hydrolyzed bamboo shoot pulp. At 48-52℃, cellulase degrades the crude fiber of bamboo shoots into glucose, breaks down the cell walls of the bamboo shoots, and releases internal soluble nutrients, such as soluble dietary fiber (SDF); pectinase degrades pectin into galacturonic acid, reduces the viscosity of bamboo shoot pulp, improves filtration efficiency, and releases bound soluble sugars and flavor substances; xylanase degrades hemicellulose into xylose, supplementing the fermentable carbon source and meeting the growth requirements of lactic acid bacteria. The compound detoxification enzyme can remove anti-nutritional factors and ensure food safety; tannin enzyme hydrolyzes tannins into gallic acid and glucose, reducing astringency and preventing tannins from combining with proteins and affecting product texture; oxalate decarboxylase catalyzes the decarboxylation of oxalate, reducing oxalate residue and preventing the formation of calcium oxalate precipitate that clogs the filtration equipment; β-glucosidase hydrolyzes cyanogenic glycosides, creating conditions for yeast to metabolize hydrogen cyanide. Tannins and oxalic acid are the main sources of astringency in bamboo shoot enzyme syrup. Although tannins have some antioxidant and antibacterial effects, they also bind to proteins, causing them to denature and precipitate, thus severely affecting protein digestion and absorption. They also bind to minerals such as iron, calcium, and zinc, forming insoluble complexes that significantly reduce the bioavailability of these minerals. Long-term excessive intake may lead to anemia and has a significant anti-nutritional effect. Oxalic acid, on the other hand, has almost no known physiological benefits and can increase the risk of kidney stones. Cyanogenic glycosides have no known essential physiological functions or benefits for humans; however, they decompose under the action of enzymes to produce highly toxic hydrocyanic acid, which can inhibit cellular respiration, leading to cellular hypoxia. High doses can cause acute poisoning, with symptoms including nausea, vomiting, difficulty breathing, and even coma and death.

[0008] S2, In-situ Detoxification and Fermentation: After enzymatic inactivation, the enzymatically hydrolyzed bamboo shoot pulp is cooled to 28-30℃, and a compound microbial premixed solution is inoculated in a gradient. Detoxification aids are added simultaneously. After 12 hours of aerobic fermentation, it is switched to anaerobic fermentation for 36-48 hours to reach the fermentation endpoint and obtain bamboo shoot pulp fermentation liquid. During the aerobic fermentation stage (0-12 hours), *Bacillus licheniformis* and *Bacillus coagulans* proliferate rapidly, secreting cellulase and protease to further degrade the crude fiber and macromolecular proteins in the bamboo shoot pulp; they also produce antimicrobial peptides to inhibit contamination by other microorganisms, while consuming a small amount of oxygen to create an anaerobic environment for lactic acid bacteria; the seed culture concentration of *Bacillus licheniformis* and *Bacillus coagulans* is ≥5×10⁻⁶. 8 CFU / mL, due to its rapid proliferation, a low concentration can quickly achieve an effective viable count (≥1×10⁻⁶ CFU / mL during fermentation). 8(CFU / mL) to avoid excessive proliferation and competition for carbon sources with lactic acid bacteria; During the anaerobic fermentation phase (12-48 hours), *Lactobacillus plantarum* and *Lactobacillus acidophilus* rapidly produce lactic acid, lowering the pH from 4.5 to 3.8-4.2. This process deeply degrades soluble sugars, ensuring low sugar content, and also enhances the activity of residual detoxifying enzymes from S1, resulting in synergistic detoxification. The proliferation of *Lactobacillus plantarum* and *Lactobacillus acidophilus* depends on the carbon source released by the first echelon of bacteria, requiring a seed culture concentration ≥1×10⁻⁶. 9 CFU / mL, high initial concentration allows for rapid control of fermentation (≥1×10⁻⁶ CFU / mL after 24 hours of fermentation). 9 CFU / mL), which is the dominant acid-producing component; Yersinia lipolyticis, as a facultative anaerobe, has a slow proliferation rate. Throughout the fermentation process, it synchronously metabolizes cyanogenic glycosides, producing hydrocyanic acid (a low-toxicity thiocyanate) which consumes gallic acid generated from the enzymatic hydrolysis of tannins, reducing astringency. Simultaneously, it produces ethyl acetate, improving flavor. The seed culture concentration should be ≥5×10⁻⁶. 7 CFU / mL, *Yarrowia lipolyticis* requires only a low viable count to complete metabolic detoxification (≥1×10⁻¹⁰ CFU / mL after 48 hours of fermentation). 7 (CFU / mL), high concentrations can actually consume too much carbon source.

[0009] Chitosan, as a cationic polysaccharide, continuously adsorbs gallic acid produced by tannin decomposition during the entire fermentation process, as well as other negatively charged impurities, some heavy metal ions, and residual pesticide molecules produced during fermentation.

[0010] MnSO4 acts as a cofactor for oxalate decarboxylase, ensuring that the enzyme can work continuously and efficiently throughout the fermentation process.

[0011] Polysorbate 80, as a surfactant, can increase cell membrane permeability, which on the one hand facilitates the contact between β-glucosidase and residual cyanogenic glycosides, and on the other hand promotes the transmembrane transport of nutrients and accelerates cell growth.

[0012] S3. Filtration and Seasoning Processing: After fermentation, the bamboo shoot fermentation liquid is first coarsely filtered through a 5μm plate and frame filter, and then finely filtered through a 0.8μm ceramic membrane. After coarse filtration, natural sweeteners and compound natural preservatives are added directly to obtain a low-sugar version of bamboo shoot enzyme liquid. After fine filtration, natural sweeteners and compound natural preservatives are added again to obtain a sugar-free version of bamboo shoot enzyme liquid. The water activity Aw is controlled at 0.88-0.92. 5μm plate and frame filtration removes most bacterial cells, insoluble dietary fiber, and chitosan-tannin complexes, resulting in a clear solution, but retains some small-molecule sugars. 0.8μm ceramic membrane filtration removes almost all bacterial cells and colloidal substances, ensuring extremely low sugar content in the sugar-free version.

[0013] Natural sweeteners compensate for the loss of sweetness caused by fermentation and filtration, but do not increase the sugar content of the enzyme syrup.

[0014] S4. Aseptic packaging and storage: Use polyethylene terephthalate bottles, sterilize at 121℃ for 20 minutes, fill, pasteurize at 65℃ for 30 minutes, and simultaneously aseptically seal with aluminum foil. Store at room temperature away from light. Shelf life ≥ 12 months.

[0015] Preferably, in step S1, the fresh tender bamboo shoots are preferably Lei bamboo shoots or Ma bamboo shoots; if some older bamboo shoots or bamboo shoots with extremely high crude fiber content are selected, such as winter bamboo shoots of Moso bamboo, then in step S3, 0.8-1.0% by weight of erythritol and 0.2-0.4% by weight of inulin should be added to the bamboo shoot slurry; the initial bacterial load of the bamboo shoot slurry should be controlled to be ≤1×10⁻⁶ during blanching. 4 CFU / g, pesticide residue ≤0.05mg / kg.

[0016] Preferably, in step S1, the compound enzyme preparation is formulated from cellulase, pectinase, and xylanase in a mass ratio of 2:1:1, with an enzyme activity ≥10000 U / g, and the amount added is 0.15-0.2% of the mass of bamboo shoot pulp. The compound detoxification enzyme premix is ​​prepared by tanninase, oxalate decarboxylase and β-glucosidase in a ratio of 3:2:1, and the amount added is 0.02-0.03% of the mass of bamboo shoot pulp; tanninase activity ≥500U / g; oxalate decarboxylase activity ≥1000U / g; β-glucosidase activity ≥800U / g.

[0017] Preferably, in S2, the compound microbial premix is ​​prepared by Lactobacillus plantarum, Lactobacillus acidophilus, Bacillus licheniformis, Bacillus coagulans, and Yersinia lipolyticis in a mass ratio of 3:2:1:0.5:0.5, and the total inoculation amount is 5-8% of the mass of the enzymatically hydrolyzed bamboo shoot pulp. The seed liquid is premixed in advance. Fermentation parameters are: First stage (aerobic): 28-30℃, DO=5-8mg / L, fermentation for 12 hours; The second stage (anaerobic) is at 32-35℃, with DO < 1 mg / L, and fermentation takes 36-48 hours. The detoxification aid is formulated with 0.1% chitosan, 0.0001-0.05% MnSO4, and 0.02% polysorbate 80 by weight of enzymatically hydrolyzed bamboo shoot pulp.

[0018] Preferably, in step S3, the natural sweeteners are compounded, based on the mass of the bamboo shoot paste: The low-sugar version uses 0.8-1.2% erythritol and 0.01-0.02% steviol glycosides; The sugar-free version contains 1.0-1.5% erythritol, 0.3-0.5% allulose, and 0.015% steviol glycosides. The natural preservatives are 0.03-0.05% ε-polylysine and 0.02-0.04% natamycin.

[0019] The present invention also proposes a bamboo shoot enzyme syrup based on the aforementioned preparation method, characterized in that the bamboo shoot enzyme syrup combines enzymatic hydrolysis and staged fermentation with compound microbial strains, and has the advantages of low sugar, nutrition and good flavor.

[0020] Compared with the prior art, the beneficial effects of the present invention are: 1. Enzymatic hydrolysis is performed using the residual heat after blanching, which is energy-efficient and highly effective. Simultaneously, a complex enzyme system (cellulase, pectinase, xylanase) degrades fiber and pectin, releasing soluble nutrients and adapting to subsequent fermentation. Targeting the anti-nutritional factors unique to bamboo shoots (such as tannins, oxalic acid, and cyanogenic glycosides), tanninase, oxalate decarboxylase, and β-glucosidase are used for specific detoxification, reducing bitterness and toxicity, and improving safety and nutrient utilization. Fermentation proceeds directly after enzymatic hydrolysis, eliminating the need for intermediate processing, simplifying the process, and the hydrolysis products provide carbon and nutrients for fermentation.

[0021] 2. The aerobic stage (Bacillus licheniformis and Bacillus coagulans) consumes oxygen and inhibits other microorganisms, while creating an anaerobic environment for the anaerobic stage (Lactobacillus plantarum and Lactobacillus acidophilus). Simultaneously, Yersinia lipolytica metabolizes cyanogenic glycosides and tannin derivatives throughout the process, improving flavor. Each strain plays a specific role at different stages of fermentation, such as enzyme production, acid production, and detoxification. This synergistic effect results in highly efficient detoxification and flavor enhancement.

[0022] 3. By treating detoxification as an intrinsic property of the fermentation process, and simultaneously inoculating detoxification aids with compound microbial strains, fermentation and detoxification mutually promote each other. Microorganisms become active "cleaners" rather than passive "victims," ​​improving the success rate and stability of fermentation. Polysorbate 80 enhances membrane permeability, promoting the uptake and metabolic transformation of hydrocyanic acid by microorganisms. Fermentation microorganisms, especially yeasts and Bacillus, can utilize these "toxins" as precursors or substrates. This achieves in-situ detoxification fermentation. By the end of fermentation, most of the detoxification task has been completed, and impurities have been transformed or adsorbed by microorganisms, making subsequent filtration simpler and more efficient.

[0023] In summary, this invention integrates enzymatic hydrolysis, targeted detoxification, and segmented fermentation, simultaneously adding detoxification enzymes and compound enzyme preparations, and utilizing the residual heat from blanching to initiate the reaction, thus shortening the process flow by more than 40% and significantly reducing energy consumption. Through the "in-situ detoxification fermentation" mechanism, toxins such as tannins, oxalic acid, and cyanogenic glycosides are simultaneously degraded during microbial metabolism, improving both safety and efficiency. By employing graded filtration and flexible blending technologies, both low-sugar and sugar-free products can be precisely produced on the same production line, breaking through the limitations of traditional processes that produce only one type of product, and achieving a balance between high efficiency, safety, and diverse market demands. Attached Figure Description

[0024] Figure 1 This is a process flow diagram of the present invention. Detailed Implementation

[0025] The technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings of the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments.

[0026] Example 1: like Figure 1 As shown, a method for preparing bamboo shoot enzyme slurry based on bamboo shoot slurry enzymatic hydrolysis includes the following steps: S1. Bamboo shoot pretreatment and residual heat enzymatic hydrolysis: Fresh bamboo shoots are cleaned and peeled, then steamed at 105℃ for 3 minutes to kill the greenness. They are then crushed and pulped to obtain bamboo shoot pulp. Pure water is added to adjust the material-to-liquid ratio to 1:1.2 and the pH is adjusted to 4.5-5.0. When the material-to-liquid ratio drops to 48-52℃, a compound enzyme preparation and a compound detoxification enzyme premix are added. Enzymatic hydrolysis is carried out for 4 hours, with stirring 3 times for 10 minutes each time. After enzymatic hydrolysis, the enzyme preparation is inactivated by incubation at 90℃ for 15 minutes to obtain enzymatically hydrolyzed bamboo shoot pulp. S2, In-situ Detoxification and Fermentation: After enzymatic inactivation, the enzymatically hydrolyzed bamboo shoot pulp was cooled to 28°C, and a compound microbial premix was inoculated in a gradient. Detoxification aids were added simultaneously. After 12 hours of aerobic fermentation, the fermentation was switched to anaerobic fermentation for 48 hours to reach the fermentation endpoint and obtain bamboo shoot pulp fermentation liquid. S3. Filtration and Seasoning Processing: After fermentation, the bamboo shoot fermentation liquid is first coarsely filtered through a 5μm plate and frame filter, and then finely filtered through a 0.8μm ceramic membrane. After coarse filtration, natural sweeteners and compound natural preservatives are added directly to obtain a low-sugar version of bamboo shoot enzyme liquid. After fine filtration, natural sweeteners and compound natural preservatives are added again to obtain a sugar-free version of bamboo shoot enzyme liquid. The water activity Aw is controlled at 0.88. S4. Aseptic packaging and storage: Polyethylene terephthalate bottles are used. After sterilization at 121°C for 20 minutes, the contents are filled, pasteurized at 65°C for 30 minutes, and then aseptically sealed with aluminum foil. Store at room temperature away from light.

[0027] In step S1, fresh tender bamboo shoots are preferably Lei bamboo shoots or Ma bamboo shoots; if some older bamboo shoots or bamboo shoots with extremely high crude fiber content are selected, such as winter bamboo shoots of Moso bamboo, then in step S3, 1.0% by weight of erythritol and 0.2% by weight of inulin should be added to the bamboo shoot slurry; the initial bacterial load of the bamboo shoot slurry should be controlled to be ≤1×10⁻⁶ during blanching. 4 CFU / g, pesticide residue ≤0.05mg / kg.

[0028] In S1, the compound enzyme preparation is formulated from cellulase, pectinase and xylanase in a mass ratio of 2:1:1, with an enzyme activity ≥10000U / g, and the amount added is 0.2% of the mass of bamboo shoot pulp; The compound detoxification enzyme premix is ​​prepared by tanninase, oxalate decarboxylase and β-glucosidase in a ratio of 3:2:1, and the amount added is 0.02% of the mass of bamboo shoot pulp; tanninase activity ≥500U / g; oxalate decarboxylase activity ≥1000U / g; β-glucosidase activity ≥800U / g.

[0029] In S2, the compound microbial premix solution is prepared by Lactobacillus plantarum, Lactobacillus acidophilus, Bacillus licheniformis, Bacillus coagulans, and Yersinia lipolyticis in a mass ratio of 3:2:1:0.5:0.5, with a total inoculation amount of 8% of the mass of the enzymatically hydrolyzed bamboo shoot pulp. The seed liquid is premixed in advance. Fermentation parameters are: First stage (aerobic): 28-30℃, DO=5-8mg / L, fermentation for 12 hours; The second stage (anaerobic) is at 32-35℃, with DO < 1 mg / L, and fermentation for 48 hours. The detoxification aid is formulated with 0.1% chitosan, 0.05% MnSO4, and 0.02% polysorbate 80 by weight of enzymatically hydrolyzed bamboo shoot pulp.

[0030] In S3, the natural sweetener compound, based on the weight of bamboo shoot paste: The low-sugar version uses 0.8% erythritol and 0.02% steviol glycosides; The sugar-free version contains 1.0% erythritol, 0.5% allulose, and 0.015% steviol glycosides. The natural preservatives are 0.03% ε-polylysine and 0.04% natamycin.

[0031] Example 2: S1. Bamboo shoot pretreatment and residual heat enzymatic hydrolysis: Fresh tender bamboo shoots are cleaned and peeled, then steamed at 105℃ for 3-5 minutes to blanch them. They are then crushed and pulped to obtain bamboo shoot pulp. Pure water is added to adjust the material-to-liquid ratio to 1:1 and the pH is adjusted to 4.5-5.0. When the material-to-liquid ratio drops to 48-52℃, a compound enzyme preparation and a compound detoxification enzyme premix are added. Enzymatic hydrolysis is carried out for 5 hours, with stirring three times for 10 minutes each time. After enzymatic hydrolysis, the enzyme preparation is inactivated by incubation at 90℃ for 15 minutes to obtain enzymatically hydrolyzed bamboo shoot pulp. S2, In-situ Detoxification and Fermentation: After enzymatic inactivation, the enzymatically hydrolyzed bamboo shoot pulp was cooled to 29°C, and a premixed solution of compound microbial strains was inoculated in a gradient. Detoxification aids were added simultaneously. After 12 hours of aerobic fermentation, the fermentation was switched to anaerobic fermentation for 42 hours to reach the fermentation endpoint and obtain bamboo shoot pulp fermentation liquid. S3. Filtration and Seasoning Processing: After fermentation, the bamboo shoot fermentation liquid is first coarsely filtered through a 5μm plate and frame filter, and then finely filtered through a 0.8μm ceramic membrane. After coarse filtration, natural sweeteners and compound natural preservatives are added directly to obtain a low-sugar version of bamboo shoot enzyme liquid. After fine filtration, natural sweeteners and compound natural preservatives are added again to obtain a sugar-free version of bamboo shoot enzyme liquid. The water activity Aw is controlled at 0.90. S4. Aseptic packaging and storage: Use polyethylene terephthalate bottles, sterilize at 121℃ for 20 minutes, fill, pasteurize at 65℃ for 30 minutes, and simultaneously aseptically seal with aluminum foil. Store at room temperature away from light. Shelf life ≥ 12 months.

[0032] In step S1, fresh tender bamboo shoots are preferably Lei bamboo shoots or Ma bamboo shoots; if some older bamboo shoots or bamboo shoots with extremely high crude fiber content are selected, such as winter bamboo shoots of Moso bamboo, then in step S3, 0.9% by weight of erythritol and 0.3% by weight of inulin should be added to the bamboo shoot slurry; the initial bacterial load of the bamboo shoot slurry should be controlled to be ≤1×10⁻⁶ during blanching. 4 CFU / g, pesticide residue ≤0.05mg / kg.

[0033] In S1, the compound enzyme preparation is formulated with cellulase, pectinase and xylanase in a mass ratio of 2:1:1, with an enzyme activity ≥10000U / g, and the amount added is 0.15% of the mass of bamboo shoot pulp; The compound detoxification enzyme premix is ​​prepared by tanninase, oxalate decarboxylase and β-glucosidase in a ratio of 3:2:1, and the amount added is 0.02-0.03% of the mass of bamboo shoot pulp; tanninase activity ≥500U / g; oxalate decarboxylase activity ≥1000U / g; β-glucosidase activity ≥800U / g.

[0034] In S2, the compound microbial premix solution is prepared by Lactobacillus plantarum, Lactobacillus acidophilus, Bacillus licheniformis, Bacillus coagulans, and Yersinia lipolyticis in a mass ratio of 3:2:1:0.5:0.5, and the total inoculation amount is 6.5% of the mass of the enzymatically hydrolyzed bamboo shoot pulp. The seed liquid is premixed in advance. Fermentation parameters are: First stage (aerobic): 28-30℃, DO=5-8mg / L, fermentation for 12 hours; The second stage (anaerobic) is at 32-35℃, with DO < 1 mg / L, and fermentation for 42 hours. The detoxification aid is formulated with 0.1% chitosan, 0.025% MnSO4, and 0.02% polysorbate 80 by weight of enzymatically hydrolyzed bamboo shoot pulp.

[0035] In S3, the natural sweetener compound, based on the weight of bamboo shoot paste: The low-sugar version uses 1.0% erythritol and 0.015% steviol glycosides; The sugar-free version contains 1.25% erythritol, 0.4% allulose, and 0.015% steviol glycosides. The natural preservatives are 0.04% ε-polylysine and 0.03% natamycin.

[0036] Example 3: S1. Bamboo shoot pretreatment and residual heat enzymatic hydrolysis: Fresh tender bamboo shoots are cleaned and peeled, then steamed at 105℃ for 5 minutes to blanch them. They are then crushed and pulped to obtain bamboo shoot pulp. Pure water is added to adjust the material-to-liquid ratio to 1:0.8 and the pH is adjusted to 4.5-5.0. When the material-to-liquid ratio drops to 48-52℃, a compound enzyme preparation and a compound detoxification enzyme premix are added. Enzymatic hydrolysis is carried out for 6 hours, with stirring twice for 10 minutes each time. After enzymatic hydrolysis, the enzyme preparation is inactivated by incubation at 90℃ for 15 minutes to obtain enzymatically hydrolyzed bamboo shoot pulp. S2, In-situ Detoxification and Fermentation: After enzymatic inactivation, the enzymatically hydrolyzed bamboo shoot pulp was cooled to 30°C, and a compound microbial premix was inoculated in a gradient. Detoxification aids were added simultaneously. After 12 hours of aerobic fermentation, the fermentation was switched to anaerobic fermentation for 36 hours to reach the fermentation endpoint and obtain bamboo shoot pulp fermentation liquid. S3. Filtration and Seasoning Processing: After fermentation, the bamboo shoot fermentation liquid is first coarsely filtered through a 5μm plate and frame filter, and then finely filtered through a 0.8μm ceramic membrane. After coarse filtration, natural sweeteners and compound natural preservatives are added directly to obtain a low-sugar version of bamboo shoot enzyme liquid. After fine filtration, natural sweeteners and compound natural preservatives are added again to obtain a sugar-free version of bamboo shoot enzyme liquid. The water activity Aw is controlled at 0.92. S4. Aseptic packaging and storage: Polyethylene terephthalate bottles are used. After sterilization at 121°C for 20 minutes, the contents are filled, pasteurized at 65°C for 30 minutes, and then aseptically sealed with aluminum foil. Store at room temperature away from light.

[0037] In step S1, fresh tender bamboo shoots are preferably Lei bamboo shoots or Ma bamboo shoots; if some older bamboo shoots or bamboo shoots with extremely high crude fiber content are selected, such as winter bamboo shoots of Moso bamboo, then in step S3, 0.8% by weight of erythritol and 0.4% by weight of inulin should be added to the bamboo shoot slurry; the initial bacterial load of the bamboo shoot slurry should be controlled to be ≤1×10⁻⁶ during blanching. 4 CFU / g, pesticide residue ≤0.05mg / kg.

[0038] In S1, the compound enzyme preparation is formulated with cellulase, pectinase and xylanase in a mass ratio of 2:1:1, with an enzyme activity ≥10000U / g, and the amount added is 0.15% of the mass of bamboo shoot pulp; The compound detoxification enzyme premix is ​​prepared by tanninase, oxalate decarboxylase and β-glucosidase in a ratio of 3:2:1, and the amount added is 0.03% of the mass of bamboo shoot pulp; tanninase activity ≥500U / g; oxalate decarboxylase activity ≥1000U / g; β-glucosidase activity ≥800U / g.

[0039] In S2, the compound microbial premix solution is prepared by Lactobacillus plantarum, Lactobacillus acidophilus, Bacillus licheniformis, Bacillus coagulans, and Yersinia lipolyticis in a mass ratio of 3:2:1:0.5:0.5, and the total inoculation amount is 5% of the mass of the enzymatically hydrolyzed bamboo shoot pulp. The seed liquid is premixed in advance. Fermentation parameters are: First stage (aerobic): 28-30℃, DO=5-8mg / L, fermentation for 12 hours; The second stage (anaerobic) is at 32-35℃, with DO < 1 mg / L, and fermentation lasts for 36 hours. The detoxification aid is formulated with 0.1% chitosan, 0.0001% MnSO4, and 0.02% polysorbate 80 by weight of enzymatically hydrolyzed bamboo shoot pulp.

[0040] In S3, the natural sweetener compound, based on the weight of bamboo shoot paste: The low-sugar version uses 1.2% erythritol and 0.01% steviol glycosides; The sugar-free version contains 1.5% erythritol, 0.3% allulose, and 0.015% steviol glycosides. The natural preservatives are 0.05% ε-polylysine and 0.02% natamycin.

[0041] Based on this, the following design was also created: Comparative Example 1: The formulation and experimental method are the same as those in Preparation Example 2, but the premixed compound detoxification enzyme solution is not added in S1, but is added in S2 instead; Comparative Example 2: The formulation and experimental method were the same as in Example 2, but no detoxification aid was added in S2; Comparative Example 3: Same formulation and experimental method as Example 2, but without the addition of tanninase; Comparative Example 4: The formulation and experimental method were the same as those in Preparation Example 2, but without the addition of oxalate decarboxylase; Comparative Example 5: Same formulation and experimental method as Example 2, but without the addition of β-glucosidase; Comparative Example 6: Same formulation and experimental method as Example 2, but with double the amount of tanninase added; Comparative Example 7: The formulation and experimental method were the same as in Example 2, but double the amount of oxalate decarboxylase was added; Comparative Example 8: The formulation and experimental method were the same as in Example 2, but double the amount of β-glucosidase was added.

[0042] For each embodiment and comparative example, according to GB 29921 "National Food Safety Standard - Limits for Pathogenic Bacteria in Prepackaged Foods", GB 2763 "National Food Safety Standard - Maximum Residue Limits for Pesticides in Foods", GB 2762 "National Food Safety Standard - Limits for Contaminants in Foods", GB 28050 "National Food Safety Standard - General Rules for Nutrition Labelling of Prepackaged Foods", GB4789.35 "National Food Safety Standard - Microbiological Examination of Foods - Examination of Lactic Acid Bacteria", GB / T 23528 "Xylooligosaccharides", and GB5009.88 "Determination of Dietary Fiber in Foods", the total number of microorganisms (total bacterial count, coliform bacteria, mold and yeast count, pathogenic bacteria), toxin and anti-nutritional factor residues (tannins, oxalic acid, cyanogenic glycosides), pesticide residues, heavy metals and contaminants, sugar and energy, viable bacteria and probiotics, xylooligosaccharides, dietary fiber, and other related items of the present invention were tested. The corresponding results are shown in Tables 1-4 below: Table 1. Performance test data of bamboo shoot enzyme paste (safety) Table 2. Performance test data of bamboo shoot enzyme paste (detoxification effect) Table 3. Performance test data (physicochemical properties) of bamboo shoot enzyme paste Table 4. Performance test data of bamboo shoot enzyme paste (functionality) Data Analysis: The residual levels of anti-nutritional factors (tannins, oxalic acid, and cyanogenic glycosides) are the core indicators for evaluating the detoxification effect. Changes in their data directly reflect the effectiveness of the three-stage detoxification closed loop designed in this invention: "preliminary enzymatic hydrolysis → residual enzyme activation → strain metabolism". The gradient changes in Examples 1-3: Tannins (0.05-0.07 mg / g), oxalic acid (3.2-3.8 mg / 100g), and cyanogenic glycosides (2.5-3.1 mg / L) showed a slight increase, due to the decreasing inoculum size (8%→5%) and the adjustment of the detoxification adjuvant concentration (MnSO4 from 0.05%→0.0001%). Higher inoculum sizes and better adjuvant concentrations resulted in increased detoxification enzyme activity, such as oxalate decarboxylase in MnSO4. 2+ The activation of enzymes and the metabolic efficiency of bacterial strains, such as Yeast lipolyticum metabolizing hydrocyanic acid, showed that the stronger the activation, the more effective the enzyme-strain dosage was, thus verifying the mechanism that "the enzyme-strain dosage is positively correlated with the detoxification effect".

[0043] Comparative Example 1 (detoxification enzyme added at an inappropriate time): Tannin (0.07 mg / g), oxalic acid (4.2 mg / 100 g), and cyanogenic glycosides (3.4 mg / L) were all higher than in Example 2. Because the detoxification enzyme missed its optimal operating temperature (48-52 °C), the enzyme activity decreased by 30%-50%, proving that "residual heat coupled with enzymatic hydrolysis" is a key prerequisite for detoxification efficiency.

[0044] Comparative Example 2 (without detoxification adjuvant): Tannin (0.08 mg / g) and oxalic acid (4.5 mg / 100g) were close to the upper limit of the qualified threshold. Due to the lack of chitosan, the adsorption of tannin was insufficient. The lack of MnSO4 reduced the activity of oxalic acid decarboxylase by 40%, directly verifying the activation effect of the adjuvant on enzyme activity.

[0045] Comparative Examples 3-5 (single detoxification enzyme deficiency): Tannin (0.15 mg / g), oxalic acid (8.7 mg / 100g), and cyanogenic glycosides (6.8 mg / L) were severely exceeded, indicating that tanninase, oxalate decarboxylase, and β-glucosidase are irreplaceable in the detoxification process, and the "enzymatic network" formed by the three is the basis for the degradation of anti-nutritional factors.

[0046] Comparative Examples 6-8 (double single detoxification enzyme): The corresponding anti-nutritional factors were extremely low (tannin 0.02mg / g, oxalic acid 2.2mg / 100g, cyanogenic glycoside 1.8mg / L), but the acidity of Comparative Example 6 was relatively high (1.3%), which was due to the accumulation of gallic acid caused by excessive tannin enzyme, reflecting the design logic that the amount of enzyme needs to be balanced with the metabolism of the strain.

[0047] The changes in physicochemical indicators (total sugar, acidity, water activity, SDF) stem from the innovative mechanisms of "waste heat recovery enzymatic hydrolysis" and "combined filtration and grading": Total sugar and low sugar properties: In the example, the low sugar version has a total sugar content of 4.2-4.5 g / 100 mL, and the sugar-free version is ≤0.5 g / 100 mL, which meets the definition of "low sugar / sugar-free" in GB 28050. This is the result of gradient metabolism of the compound microbial strains—the pioneer bacteria (Bacillus) degrade the fiber and release carbon sources, while the dominant bacteria (Lactobacillus) rapidly consume soluble sugars, ultimately achieving precise control of sugar content.

[0048] Acidity and preservative stability: An acidity of 0.9%-1.2% (calculated as lactic acid) corresponds to a pH of 3.8-4.2, which inhibits harmful bacteria (such as Staphylococcus aureus ≤35 CFU / mL) while providing a suitable environment for probiotics. Comparative example 6, with an acidity of 1.3%, is slightly higher, leading to the accumulation of secondary metabolites due to excessive tannins, further validating the necessity of enzyme-microbe metabolism balance.

[0049] Water activity (Aw) and shelf life: In the examples, Aw was 0.88-0.92. With the addition of natural preservatives (ε-polylysine ≤0.05g / kg, natamycin ≤38mg / kg), a shelf life of ≥12 months at room temperature was achieved. In comparative examples 1-8, Aw was 0.9, indicating that the process has universal applicability in controlling water activity. This reflects the simultaneous design of flavoring and preservation in the "integrated terminal processing".

[0050] Soluble dietary fiber (SDF) and enzymatic hydrolysis efficiency: In the examples, SDF was 8.5-11.5 g / 100 mL, with the sugar-free version significantly higher than the low-sugar version, because nanofiltration membranes retained more SDF during sugar removal. Example 3 showed the highest SDF (9.5 / 11.5 g / 100 mL), which was attributed to the efficient degradation by cellulase and xylanase under a material-to-liquid ratio of 1:0.8 (high bamboo shoot pulp concentration) and residual heat enzymatic hydrolysis (48-52℃), demonstrating that "energy-coupled enzymatic hydrolysis" can maximize the release of functional components.

[0051] Sensory evaluation and detoxification integrity: All examples were "sweet and sour with no astringency", while Comparative Example 3 had "obvious astringency" (tannins exceeded the standard) and Comparative Example 6 was "slightly acidic" (gallic acid accumulation). This shows that the synergistic effect of detoxification enzymes and dosage balance are the core guarantees of sensory quality.

[0052] The performance of live bacteria count, probiotic ratio, and SOD enzyme activity demonstrates the synergistic advantages of "gradient inoculation + functional hierarchical division of labor" among bacterial strains. Total viable count and bacterial species gradient: Example 1 showed the highest total viable count (1.8 × 10⁻⁶). 9 / 2.1×10 9 (CFU / mL), due to an inoculum size of 8% and thorough detoxification, the bacterial growth environment was optimal; in Example 3, the inoculum size was 5%, and the viable count was slightly lower but still reached 1.2 × 10⁻⁶. 9 CFU / mL, demonstrating that a gradient time design (pioneer bacteria → dominant bacteria → synergist bacteria) can maintain efficient fermentation at low inoculum sizes. Comparative Examples 1-8 had a viable cell count of 1.2 × 10⁻⁸. 9 -1.5×10 9 The CFU / mL indicates that even with variable adjustments, the synergistic bacterial system can still maintain a basic level of viable bacteria.

[0053] Probiotic proportion and functional stability: *Lactobacillus plantarum* and *Lactobacillus acidophilus* account for 70%-80% of the total live bacteria, as they are the core of the "acid production + probiotic function". Example 1: *Lactobacillus plantarum* reached 1.0 × 10⁻⁶. 9 The CFU / mL result is due to the sufficient carbon source and absence of inhibitory factors after thorough detoxification, which verifies that "detoxification is a prerequisite for the efficient proliferation of bacterial strains".

[0054] SOD enzyme activity and component retention: The SOD enzyme activity in the example was 105-128 U / mL, which was much higher than that in the comparative example (105-112 U / mL). This is because the temperature gradient (105℃→48-52℃→32-35℃) in the process of "blanching-enzymatic hydrolysis-fermentation" did not destroy the structure of SOD in bamboo shoots, which shows that the mild process has the effect of retaining functional components.

[0055] The above description is only a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any equivalent substitutions or modifications made by those skilled in the art within the scope of the technology disclosed in the present invention, based on the technical solution and inventive concept of the present invention, should be covered within the scope of protection of the present invention.

Claims

1. A method for preparing bamboo shoot enzyme slurry based on bamboo shoot slurry enzymatic hydrolysis, characterized in that, Includes the following steps: S1. Bamboo shoot pretreatment and residual heat enzymatic hydrolysis: After cleaning and peeling fresh bamboo shoots, they are steamed at 105℃ for 3-5 minutes to blanch them, then crushed and pulped to obtain bamboo shoot pulp. Pure water is added to adjust the material-to-liquid ratio to 1:0.8-1.2 and the pH to 4.5-5.

0. When the material-to-liquid ratio drops to 48-52℃, a compound enzyme preparation and a compound detoxification enzyme premix are added. Enzymatic hydrolysis is carried out for 4-6 hours, with stirring 2-3 times for 10 minutes each time. After enzymatic hydrolysis, the enzyme preparation is inactivated by incubation at 90℃ for 15 minutes to obtain enzymatically hydrolyzed bamboo shoot pulp. S2, In-situ Detoxification and Fermentation: After enzymatic inactivation, the enzymatically hydrolyzed bamboo shoot pulp is cooled to 28-30℃, and a compound microbial premixed solution is inoculated in a gradient. Detoxification aids are added simultaneously. After 12 hours of aerobic fermentation, it is switched to anaerobic fermentation for 36-48 hours to reach the fermentation endpoint and obtain bamboo shoot pulp fermentation liquid. S3. Filtration and Seasoning Processing: After fermentation, the bamboo shoot fermentation liquid is first coarsely filtered through a 5μm plate and frame filter, and then finely filtered through a 0.8μm ceramic membrane. After coarse filtration, natural sweeteners and compound natural preservatives are added directly to obtain a low-sugar version of bamboo shoot enzyme liquid. After fine filtration, natural sweeteners and compound natural preservatives are added again to obtain a sugar-free version of bamboo shoot enzyme liquid. The water activity Aw is controlled at 0.88-0.

92. S4. Aseptic packaging and storage: Use polyethylene terephthalate bottles, sterilize at 121℃ for 20 minutes, fill, pasteurize at 65℃ for 30 minutes, and simultaneously aseptically seal with aluminum foil. Store at room temperature away from light. Shelf life ≥ 12 months.

2. The method for preparing bamboo shoot enzyme slurry based on bamboo shoot slurry enzymatic hydrolysis according to claim 1, characterized in that, In step S1, fresh tender bamboo shoots are preferably Lei bamboo shoots or Ma bamboo shoots; if some older bamboo shoots or bamboo shoots with extremely high crude fiber content are selected, such as winter bamboo shoots of Moso bamboo, then in step S3, 0.8-1.0% by weight of erythritol and 0.2-0.4% by weight of inulin should be added to the bamboo shoot slurry; the initial bacterial load of the bamboo shoot slurry should be controlled to be ≤1×10⁻⁶ during blanching. 4 CFU / g, pesticide residue ≤0.05mg / kg.

3. The method for preparing bamboo shoot enzyme slurry based on bamboo shoot slurry enzymatic hydrolysis according to claim 1, characterized in that, In S1, the compound enzyme preparation is formulated from cellulase, pectinase and xylanase in a mass ratio of 2:1:1, with an enzyme activity ≥10000U / g, and the amount added is 0.15-0.2% of the mass of bamboo shoot pulp; The compound detoxification enzyme premix is ​​prepared by tanninase, oxalate decarboxylase and β-glucosidase in a ratio of 3:2:1, and the amount added is 0.02-0.03% of the mass of bamboo shoot pulp; tanninase activity ≥500U / g; oxalate decarboxylase activity ≥1000U / g; β-glucosidase activity ≥800U / g.

4. The method for preparing bamboo shoot enzyme slurry based on bamboo shoot slurry enzymatic hydrolysis according to claim 1, characterized in that, In S2, the compound microbial premix is ​​prepared by Lactobacillus plantarum, Lactobacillus acidophilus, Bacillus licheniformis, Bacillus coagulans, and Yersinia lipolyticis in a mass ratio of 3:2:1:0.5:0.5, and the total inoculation amount is 5-8% of the mass of the enzymatically hydrolyzed bamboo shoot pulp. The seed liquid is premixed in advance. Fermentation parameters are: First stage (aerobic): 28-30℃, DO=5-8mg / L, fermentation for 12 hours; The second stage (anaerobic) is at 32-35℃, with DO < 1 mg / L, and fermentation takes 36-48 hours. The detoxification aid is formulated with 0.1% chitosan, 0.0001-0.05% MnSO4, and 0.02% polysorbate 80 by weight of enzymatically hydrolyzed bamboo shoot pulp.

5. The method for preparing bamboo shoot enzyme slurry based on bamboo shoot slurry enzymatic hydrolysis according to claim 1, characterized in that, In S3, the natural sweetener compound, based on the weight of bamboo shoot paste: The low-sugar version uses 0.8-1.2% erythritol and 0.01-0.02% steviol glycosides; The sugar-free version contains 1.0-1.5% erythritol, 0.3-0.5% allulose, and 0.015% steviol glycosides. The natural preservatives are 0.03-0.05% ε-polylysine and 0.02-0.04% natamycin.

6. A bamboo shoot enzyme slurry prepared by the preparation method according to any one of claims 1-5, characterized in that, The bamboo shoot enzyme syrup combines enzymatic hydrolysis with staged fermentation using compound microbial strains, resulting in a product with advantages such as low sugar content, high nutritional value, and excellent flavor.

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