A method for degrading kitchen waste protein by mechanical chemical enzyme coupling

Abstract

The present invention discloses a method for degrading kitchen waste protein by mechanochemical enzyme coupling, comprising the steps of: mixing a kitchen waste stock solution with a solid-phase reagent and placing the mixture in a polytetrafluoroethylene ball mill; adding stainless steel beads; setting the ball mill speed to 100-500 rpm; and performing a milling reaction for 6-12 cycles. After the ball milling reaction, the residue is filtered and removed to obtain a kitchen waste protein degradation solution. The present invention has the advantages of a short process route, low reagent usage, simple operation, low cost, low pollution, high protein degradation rate, high amino acid yield, and good degradation efficiency. Furthermore, the method can be used as a fertilizer to better promote plant growth, making it a technology with promising application prospects.

Description

Technical Field

[0001] The present invention relates to the technical field of protein degradation, and in particular to a method for degrading kitchen waste protein by mechanical chemical enzyme coupling. Background Art

[0002] Food waste protein, a byproduct of the food and catering industries, holds enormous economic and environmental potential, but its degradation and high-value utilization face numerous challenges. Compared to the established applications of hemicellulose and cellulose, the utilization rate of food waste protein is low, primarily due to its complex molecular structure and difficulty in degradation. The stability of peptide bonds and tertiary structures, as well as certain natural antimicrobial activities, complicate the degradation process, necessitating specialized enzymatic or chemical methods. Currently, protein degradation technologies are evolving, including enzymatic and chemical degradation. Enzymatic degradation is favored for its environmental friendliness and high efficiency. Using specific proteases, such as trypsin and papain, it effectively degrades proteins into amino acids and small peptides. Chemical degradation, using metal-based catalysts, such as those derived from metal-organic frameworks (MOFs), enables peptide bond cleavage under mild conditions, yielding high-value amino acids and peptides. However, cost control, product selectivity, and scalable production remain bottlenecks in the degradation and resource utilization of food waste protein. Future research will focus on developing more efficient and economical degradation technologies, combining the strengths of enzyme engineering, chemical catalysis, and biotechnology to achieve high-value utilization of food waste protein, promoting both its sustainability and economic value. Through these efforts, food waste protein will be transformed from a waste product into a valuable resource, bringing new development opportunities to the food, pharmaceutical, and chemical industries.

[0003] Research has revealed that despite the diverse variety of proteins found in food waste, their complex structures are all rich in peptide bonds. These peptide bonds form the protein backbone and are crucial for chemical degradation. Therefore, employing appropriate methods to achieve complete hydrolysis and even targeted cleavage of peptide bonds is crucial for the efficient conversion of food waste into amino acids. Numerous biological and chemical methods have been employed for protein degradation. For example, Khan et al. used papain as a catalyst to hydrolyze proteins to oxidize side chains and cleave peptide bonds. They found that under suitable pH and temperature conditions, protein side chains undergo oxidative cleavage under the action of the enzyme, leading to direct degradation into amino acids and small peptides. Li et al. used hydrogen peroxide as an oxidant to chemically oxidize protein side chains and cleave their structure. Experimental results showed that protein side chains undergo oxidative cleavage under the action of the oxidant, degrading them into a series of oxygenated derivatives, such as amino acid oxidation products and peptide cleavage products. Zhang et al. used copper ions as a catalyst to achieve side chain oxidation and peptide bond cleavage through air oxidation. Research has found that protein side chains undergo oxidative cleavage under metal catalysis, directly degrading into amino acids and their derivatives, as well as the oxidative cleavage products of some peptides. Wang et al. employed photocatalytic technology, using titanium dioxide as a photocatalyst, to achieve protein side chain oxidation and peptide bond cleavage under ultraviolet light irradiation. The experimental results showed that protein side chains underwent oxidative cleavage under photocatalysis, degrading into amino acids, peptides, and a series of oxidation products. Liu et al. used specific microbial strains to achieve protein side chain oxidation and peptide bond cleavage through biodegradation. The study found that protein side chains underwent oxidative cleavage under microbial action, degrading into amino acids, small peptides, and some microbial metabolites, demonstrating the potential of biodegradation in protein degradation.

[0004] Through comparison and analysis of the above studies, it can be found that both chemical and biological degradation methods face certain problems. First of all, chemical degradation is efficient and fast, but it usually requires a large amount of inorganic reagents and produces a large amount of saline waste liquid. For agricultural reuse, inorganic salts have a great impact on crop growth, which makes the product difficult to use. Compared with chemical explanations, enzymatic degradation and bacterial degradation of protein have less pollution, but the price of efficient enzyme catalysts is too high, and the use of bacterial degradation is difficult to avoid the consumption of amino acids by bacteria, and the biodegradation reaction time is very long, which seriously affects production. In short, the core of the current degradation and reuse of kitchen waste protein is to introduce an efficient, convenient and low-cost reaction intensification technology that combines the advantages of biodegradation and chemical degradation technologies.

[0005] In recent years, mechanochemistry, a reaction optimization technology, has gained industrial favor due to its unique reaction mechanism, higher yields and selectivity, and the convenience of solvent-free or minimally solvent-free reactions. Many metal-catalyzed reactions that previously required prolonged, high temperatures and large amounts of high-boiling-point organic solvents can now be efficiently performed at room temperature, achieving even higher yields and selectivities. It can even utilize raw materials that are unavailable in solution reactions. In particular, for chemical reactions between poorly soluble macromolecules, mechanochemistry can directly promote complexation, neutralization, degradation, and grafting reactions in the solid phase, regardless of solvent selection. Currently, it has found extensive application in the processing of polymers such as cellulose, pectin, lignin, cyclodextrins, and cucurbiturils. However, the application of mechanochemistry in protein degradation remains largely unexplored. Therefore, leveraging the mechanism of mechanochemistry to promote solid-phase reactions between macromolecules, enhance the protein degradation process, reduce wastewater pressure, and ensure sufficient protein degradation, could further enable the efficient degradation and resource utilization of food waste proteins. Summary of the Invention

[0006] In response to the above problems, the purpose of the present invention is to overcome the problems existing in existing protein degradation processes, such as large amounts of chemical reagents, high reaction temperatures, long reaction times, and complicated processes, and to provide a simple, efficient, and environmentally friendly method for degrading kitchen waste proteins by mechanochemical enzyme coupling. Based on the advantage of mechanochemistry in promoting polymer reactions, the method activates protein peptide bonds through mechanochemical treatment, promotes protein hydrolysis while preventing secondary damage to the produced amino acids, and overcomes the problem of poor degradation ability of kitchen waste proteins.

[0007] In order to achieve the above-mentioned purpose, the present invention provides the following technical solutions:

[0008] A method for degrading kitchen waste protein by mechanochemical enzyme coupling comprises the following steps: mixing a kitchen waste stock solution with a solid phase reagent and placing the mixture in a polytetrafluoroethylene ball mill; adding stainless steel beads; setting the ball mill speed to 100-500 rpm for a grinding reaction; and performing a grinding reaction for 6-12 cycles. After the ball milling reaction is completed, the residue is filtered out to obtain a kitchen waste protein degradation solution.

[0009] In some embodiments, the mass ratio of the kitchen waste liquid to the solid phase reagent is 100:(5-12).

[0010] In some embodiments, the solid-phase reagent includes an alkaline reagent and an enzyme reagent, wherein the alkaline reagent includes at least one of sodium bicarbonate, sodium carbonate, potassium carbonate, sodium hydroxide, potassium hydroxide, and urea; and the enzyme reagent includes at least one of alkaline protease, neutral protease, and papain. The alkaline protease is derived from Bacillus subtilis (Uniprot ID: P00782); the neutral protease is derived from Bacillus subtilis (Uniprot ID: P39899); and the papain is derived from papaya (Uniprot ID: P00784).

[0011] In some embodiments, the mass ratio of the alkaline reagent to the enzyme reagent is 10:(0.1-3).

[0012] In some embodiments, the stainless steel bead has a diameter of 14 mm.

[0013] In some embodiments, the grinding cycle is 6-12 cycles, specifically: grinding for 30 minutes as one cycle, resting for 5 minutes, and grinding for a total of 6-12 cycles.

[0014] In some embodiments, the ball mill rotates at 400 rpm.

[0015] In some embodiments, the grinding cycle is 10 cycles.

[0016] Compared with the prior art, the method of the present invention for degrading kitchen waste protein by mechanochemical enzyme coupling has the following beneficial effects:

[0017] (1) The use of mechanical ball milling reaction and enzyme combination to degrade kitchen waste has simple operation, short process route and short degradation time.

[0018] (2) Compared with traditional methods, mechanical ball milling to assist in the degradation of kitchen waste protein can greatly reduce the amount of chemical reagents used, lower production costs, and alleviate environmental pollution.

[0019] (3) The degradation process takes place at room temperature, the conditions are mild and the rate is much higher than that of simple enzymatic hydrolysis.

[0020] (4) The loss of amino acids obtained is small, and the product is highly effective as a fertilizer.

[0021] In summary, the present invention has the advantages of a short process route, low reagent usage, simple operation, low cost, low pollution, high protein degradation rate, large amount of amino acids obtained, and good degradation efficiency. It is a highly efficient protein degradation technology. It can also be used as a fertilizer to better promote plant growth, and is a technology with good prospects for promotion and application. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1Effect of solid phase reagent addition amount on the degradation rate of kitchen waste protein.

[0023] Figure 2 The effect of ball mill speed on the degradation rate of kitchen waste protein.

[0024] Figure 3 The effect of ball milling cycle on the degradation rate of kitchen waste protein.

[0025] Figure 4 This is the free amino acid analysis chart of kitchen waste protein degradation products.

[0026] Figure 5 These are TEM images of food waste before and after degradation.

[0027] Figure 6 It is the effect of the degradation liquid on the plant height and biological quality of Arabidopsis thaliana. DETAILED DESCRIPTION

[0028] The present invention is described in detail below with reference to specific embodiments.

[0029] Protein content of food waste solution was determined using the BCA assay kit. Using bovine serum albumin as the standard, 2g of bovine serum albumin was accurately weighed and diluted to 100mL to create a 20g / L protein standard stock solution. 10, 6, 4, 2, and 1ml of the standard solution were then added to 100mL to create a series of standard solutions. Fifty parts of BCA reagent A were mixed with one part of BCA reagent B (a ratio of Reagent A to Reagent B = 50:1) to produce a clear green working solution. 25µL of each diluted protein standard was added to a microplate, and 200µL of the working solution was added to each well. The solution was shaken for 30 seconds to mix thoroughly. The solution was incubated at 37°C for 30 minutes and then cooled to room temperature. The absorbance of the sample was measured at 562nm. The same sample was measured in triplicate. The blank-corrected average absorbance of the standard at 562nm was plotted against its concentration (g / L) to generate a protein content standard curve. Take 10 mL of the sample to be tested, make up to 100 mL, measure according to the above method, and calculate the protein concentration of the sample according to the absorbance.

[0030] Determination of free amino acid content in degradation solution: The amino acid analyzer method and principle for free amino acid determination are as follows: First, the treated sample is centrifuged (8000g, 10 minutes) and then filtered through a 0.22μm filter. The sample is then separated using a cation exchange column (Na+-type). This process utilizes amino acids' structural differences, acidity, polarity, and molecular size to separate them on the cation exchange column. The separated individual amino acid components then react with ninhydrin to form purple or yellow compounds. Derivatization agents include o-phthalaldehyde (OPA), dinitrofluorobenzene (FDNB), 9-fluorenylmethylchloroformate (FMOC), and phenyl isothiocyanate (PITC). The resulting compounds are then photometrically measured at specific wavelengths. Proline is detected at 440nm, and other amino acids at 570nm. The content of each amino acid is calculated using an internal standard method, and the sum is used to obtain the total free amino acid content. The final results are usually analyzed and processed by the amino acid analyzer's built-in software (EZChrom Elite).

[0031] Protein content determination and degradation rate determination method:

[0032] Degradation rate (%g / g) = (protein content of degradation solution - total amino acid content of degradation solution) / (protein content before degradation - total amino acid content before degradation) Example 1

[0033] A method for mechanochemical-enzymatically degrading food waste protein comprises the following steps: adding 100 mL of food waste stock solution, 10 g of sodium carbonate, 1 g of alkaline protease, and six 14 mm diameter stainless steel beads to a 250 mL polytetrafluoroethylene ball mill. The mixture is milled in the mill at 400 rpm for 30 minutes per cycle, followed by a 5-minute rest period, for a total of 10 cycles. After ball milling, the milled material is centrifuged, and the protein and amino acid contents of the degraded solution are measured. The protein degradation rate in this example was 94.32%, and the amino acid content reached 9.82 g / L. Example 2

[0034] A method for mechanochemical-enzymatically degrading food waste protein comprises the following steps: adding 100 mL of food waste stock solution, 10 g of sodium bicarbonate, 1 g of alkaline protease, and six 14 mm diameter stainless steel beads to a 250 mL polytetrafluoroethylene ball mill. The mixture is milled in the mill at 400 rpm for 30 minutes per cycle, followed by a 5-minute rest period, for a total of 10 cycles. After ball milling, the milled material is centrifuged, and the protein and amino acid contents of the degraded solution are measured. The protein degradation rate in this example was 85.51%, and the amino acid content reached 8.36 g / L. Example 3

[0035] A method for mechanochemical-enzymatic degradation of kitchen waste protein comprises the following steps: adding 100 mL of kitchen waste stock solution, 10 g of sodium hydroxide, 1 g of alkaline protease, and six 14 mm diameter stainless steel beads to a 250 mL polytetrafluoroethylene ball mill. The mixture is milled in the mill at 400 rpm for 30 minutes per cycle, followed by a 5-minute rest period, for a total of 10 cycles. After ball milling, the milled material is centrifuged, and the protein and amino acid contents of the degraded solution are measured. The protein degradation rate in this example was 88.31%, and the amino acid content reached 7.99 g / L. Example 4

[0036] A method for mechanochemical-enzymatic degradation of kitchen waste protein comprises the following steps: adding 100 mL of kitchen waste stock solution, 10 g of potassium carbonate, 1 g of alkaline protease, and six 14 mm diameter stainless steel beads to a 250 mL polytetrafluoroethylene ball mill. The mixture is milled in the mill at 400 rpm for 30 minutes per cycle, followed by a 5-minute rest period, for a total of 10 cycles. After ball milling, the milled material is centrifuged, and the protein and amino acid contents of the degraded solution are measured. The protein degradation rate in this example was 95.53%, and the amino acid content reached 11.34 g / L. Example 5

[0037] A method for mechanochemical-enzymatic degradation of kitchen waste protein comprises the following steps: adding 100 mL of kitchen waste stock solution, 10 g of potassium hydroxide, 1 g of alkaline protease, and six 14 mm diameter stainless steel beads to a 250 mL polytetrafluoroethylene ball mill. The mixture is milled in the mill at 400 rpm for 30 minutes per cycle, followed by a 5-minute rest period, for a total of 10 cycles. After ball milling, the milled material is centrifuged, and the protein and amino acid contents of the degraded solution are measured. The protein degradation rate in this example was 74.19%, and the amino acid content reached 7.30 g / L. Example 6

[0038] A method for degrading kitchen waste protein by mechanochemical-enzymatic coupling comprises the following steps: adding 100 mL of kitchen waste stock solution, 10 g of potassium carbonate, 1 g of neutral protease, and six 14 mm diameter stainless steel beads to a 250 mL polytetrafluoroethylene ball mill. The mixture is milled in the mill at 400 rpm for a 30-minute cycle, followed by a 5-minute rest period, for a total of 10 cycles. After ball milling, the milled material is centrifuged, and the protein and amino acid contents of the degraded solution are measured. The protein degradation rate in this example was 72.37%, and the amino acid content reached 7.38 g / L. Example 7

[0039] A method for mechanochemical-enzymatic degradation of kitchen waste protein comprises the following steps: adding 100 mL of kitchen waste stock solution, 10 g of potassium carbonate, 1 g of papain, and six 14 mm diameter stainless steel beads to a 250 mL polytetrafluoroethylene ball mill. The mixture is milled in the mill at 400 rpm for a reaction, with a 30-minute cycle followed by a 5-minute rest period, for a total of 10 cycles. After ball milling, the milled material is centrifuged, and the protein and amino acid contents of the degraded solution are measured. The protein degradation rate in this example was 73.52%, and the amino acid content reached 7.79 g / L. Example 8

[0040] A method for mechanochemical-enzymatically degrading kitchen waste protein comprises the following steps: adding 100 mL of kitchen waste stock solution, 10 g of potassium carbonate, 0.5 g of alkaline protease, and six 14 mm diameter stainless steel beads to a 250 mL polytetrafluoroethylene ball mill. The mixture was milled at 300 rpm for 30 minutes per cycle, followed by a 5-minute rest period, for a total of 10 milling cycles. After ball milling, the milled material was centrifuged, and the protein and amino acid contents of the degraded solution were determined. The protein degradation rate in this example was 91.72%, and the amino acid content reached 9.56 g / L. Example 9

[0041] A method for mechanochemical-enzymatic degradation of kitchen waste protein comprises the following steps: adding 100 mL of kitchen waste stock solution, 10 g of potassium carbonate, 0.2 g of alkaline protease, and six 14 mm diameter stainless steel beads to a 250 mL polytetrafluoroethylene ball mill. The mixture is milled at 200 rpm for 30 minutes per cycle, followed by a 5-minute rest period, for a total of 10 cycles. After ball milling, the milled material is centrifuged, and the protein and amino acid contents of the degraded solution are measured. The protein degradation rate in this example was 92.45%, and the amino acid content reached 9.22 g / L. Example 10

[0042] A method for mechanochemical-enzymatic degradation of kitchen waste protein comprises the following steps: adding 100 mL of kitchen waste stock solution, 10 g of potassium carbonate, 2 g of alkaline protease, and six 14 mm diameter stainless steel beads to a 250 mL polytetrafluoroethylene ball mill. The mixture is milled at 500 rpm for 30 minutes per cycle, followed by a 5-minute rest period, for a total of 10 milling cycles. After ball milling, the milled material is centrifuged, and the protein and amino acid contents of the degraded solution are measured. The protein degradation rate of the degraded food waste in this example was 93.09%, and the amino acid content reached 10.08 g / L.

[0043] Example 11 Optimal mass ratio of solid phase reagent to kitchen waste solution

[0044] A method for degrading kitchen waste protein by mechanical chemical enzyme coupling, comprising the following steps: adding 100 mL of kitchen waste stock solution to a 250 mL polytetrafluoroethylene ball mill, controlling the addition amount of solid phase reagent (potassium carbonate: alkaline protease = 10:1) to 5, 6, 7, 8, 9, 10, 11, and 12 g, respectively, and 6 stainless steel beads with a diameter of 14 mm, grinding the reaction at 400 rpm, with a cycle of 30 minutes, a rest of 5 minutes, and 10 cycles of grinding. After ball milling, the ball milled material is centrifuged and the protein content and amino acid content of the degradation solution are determined. Figure 1 As shown in the figure, the test results show that if the amount of solid phase reagent added is too little, the protein cannot be fully oxidized and degraded, and the degradation effect is poor; if the amount of solid phase reagent added is too much, the alkalinity of the system will be too strong, which will further destroy the activity of the enzyme and also cause the loss of amino acids. Therefore, the preferred mass ratio of solid phase reagent to kitchen waste liquid is 1:10.

[0045] Example 12 Optimal Ball Mill Speed

[0046] A method for degrading kitchen waste protein by mechanochemical enzyme coupling comprises the following steps: adding 100 mL of kitchen waste stock solution, 10 g of potassium carbonate, 1 g of alkaline protease, and 6 stainless steel beads with a diameter of 14 mm to a 250 mL polytetrafluoroethylene ball mill, mixing them evenly, placing the ball mill into a planetary ball mill, controlling the speed of the ball mill to 100, 150, 200, 250, 300, 350, 400, 450, and 500 rpm for 10 cycles, centrifuging the milled material after ball milling, and measuring the protein content and amino acid content of the degradation solution. Figure 2 As shown in the figure, the test results show that if the rotation speed is too low, the grinding and dispersion are insufficient, resulting in poor degradation effect; if the grinding rate is too high, the heat release of the system increases, causing partial inactivation of the enzyme and reduced degradation efficiency, so the preferred rotation speed is 400 rpm.

[0047] Example 13 Optimization of ball milling cycle

[0048] A method for degrading kitchen waste protein by mechanochemical enzyme coupling comprises the following steps: adding 100 mL of kitchen waste stock solution, 10 g of potassium carbonate, 1 g of alkaline protease, and 6 stainless steel beads with a diameter of 14 mm to a 250 mL polytetrafluoroethylene ball mill, mixing them evenly, placing the ball mill into a planetary ball mill, controlling the ball mill speed to 400 rpm, and performing 6, 7, 8, 9, 10, 11, and 12 cycles respectively; after ball milling, centrifuging the milled material, and measuring the protein content and amino acid content of the degradation solution. Figure 3 As shown in FIG, the test results show that when the ball milling time is short, the materials are not fully mixed and reacted in the ball mill, and the degradation rate is low; while when the ball milling time is too long, the degradation rate of the product does not increase, so the preferred ball milling time is 10 cycles.

[0049] Example 14 Liquid Phase Enzymatic Hydrolysis of Kitchen Waste

[0050] A method for degrading kitchen waste protein by mechanochemical-enzymatic coupling includes the following steps: adding 100 mL of kitchen waste stock solution, 10 g of potassium carbonate, and 1 g of alkaline protease to a 250 mL beaker, maintaining the enzymatic hydrolysis temperature at 35°C for 5 hours, centrifuging the solution after completion, and measuring the protein and amino acid content. Results showed a 42.15% protein degradation rate and a 4.59 g / L amino acid content. These results demonstrate that enzymatic hydrolysis cannot be fully effective without mechanochemical assistance.

[0051] Example 15 Analysis of amino acid types in protein degradation from kitchen waste

[0052] A method for mechanochemical enzyme coupling degradation of kitchen waste protein comprises the following steps: adding 100 mL of kitchen waste stock solution, 10 g of potassium carbonate, 1 g of alkaline protease, and 6 stainless steel beads with a diameter of 14 mm to a 250 mL polytetrafluoroethylene ball mill, mixing them evenly, placing the ball mill into a planetary ball mill, controlling the speed of the ball mill to 400 rpm for 10 cycles, centrifuging the milled material after ball milling, and using an amino acid analyzer to determine the amino acid content. Under optimal conditions, the protein degradation solution prepared has an amino acid content of 11.34 g / L. Its composition and distribution are shown in Tables 1 and Figure 4 .

[0053] The results in Table 1 show that the free amino acid composition of the degradation liquid is complete, containing 18 natural amino acids, among which the content distribution of various amino acids is relatively balanced, which is consistent with the characteristics of kitchen waste food sources and can more comprehensively supplement amino acid nutrition for plants.

[0054] Table 1: Protein degradation products from food waste

[0055]

[0056] Microscopic morphology of the solution before and after kitchen waste degradation

[0057] Transmission electron microscopy was used to observe the aggregation of protein microparticles in the solution before and after degradation. The specific method was as follows: 1 mg of the sample to be tested was added to 3 mL of deionized water and mixed thoroughly. A small amount of the prepared liquid was dropped onto a copper sheet coated with a carbon film using a capillary tube. The excess solution was then wiped off with filter paper. After the sample solution was dried in air, the morphology was observed and photographed using a transmission electron microscope at a detection voltage of 100 kV. Figure 5As shown in the figure, morphological observation revealed that a large number of large and irregular protein particles were present in the food waste solution before degradation. After degradation, the number of particles in the solution decreased significantly, and the particles became smaller and rounder, indicating that mechanical force and enzymes synergistically converted most of the suspended insoluble protein into soluble amino acids and proteins, fully demonstrating the role of mechanical force.

[0058] Study on the effect of kitchen waste protein degradation liquid on promoting the growth of Arabidopsis thaliana

[0059] Preparation of chemical degradation liquid for food waste: According to a reference, 100 ml of food waste was added with 20 g of sodium hydroxide to make the alkali content above 20%, and the solution was boiled under reflux for 12 hours. After the solution was neutralized with acetic acid to a pH of ~7, the protein degradation rate was determined to be 52.37% after filtration, and the amino acid content was only 5.92 g / L.

[0060] Arabidopsis thaliana at the same growth stage were divided into three groups: experimental group, control group and blank group, with 10 plants in each group. The experimental group was fertilized with the degradation solution of Example 10, diluted 1000 times, the control group was fertilized with chemical degradation solution, diluted 1000 times, and the blank group was not fertilized. Fertilization was applied once a day for 3 consecutive months, and the average height and biomass were calculated after the end of the treatment. Figure 6 As shown, the test results show that when the method is used to degrade fertilizer, the plant height of Arabidopsis thaliana is significantly higher than that of the control group and the blank group, and the biomass content is increased. The experimental results show that the protein degradation liquid prepared by this method contains organic nitrogen elements that have a better effect in promoting plant growth.

[0061] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit the scope of protection of the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solutions of the present invention may be modified or replaced by equivalents without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A method for degrading kitchen waste protein by mechanical chemical enzyme coupling, characterized in that: The method comprises the following steps: mixing a raw kitchen waste solution with a solid-phase reagent and placing the mixture in a polytetrafluoroethylene ball mill; adding stainless steel beads; setting the ball mill speed to 100-500 rpm for a grinding reaction; and performing a grinding cycle of 6-12 cycles. After the ball milling reaction is completed, filtering and removing the residue to obtain a kitchen waste protein degradation solution; the solid-phase reagent comprises an alkaline reagent and an enzyme reagent, wherein the alkaline reagent comprises at least one of sodium bicarbonate, sodium carbonate, potassium carbonate, sodium hydroxide, potassium hydroxide, and urea; the enzyme reagent comprises at least one of alkaline protease, neutral protease, and papain; the alkaline protease is derived from Bacillus subtilis, Uniprot ID: P00782; the neutral protease is derived from Bacillus subtilis, Uniprot ID: P00782 ID: P39899; the papaya source of papain, Uniprot ID: P00784; the mass ratio of the kitchen waste stock solution to the solid phase reagent is 100:(5-12); the mass ratio of the alkaline reagent to the enzyme reagent is 10:(0.1-3); the grinding cycle is 6-12 cycles, specifically: grinding for 30 minutes as one cycle, resting for 5 minutes, and grinding for a total of 6-12 cycles.

2. The method for degrading kitchen waste protein by mechanochemical enzyme coupling according to claim 1, characterized in that: The diameter of the stainless steel beads is 14 mm.

3. The method for degrading kitchen waste protein by mechanochemical enzyme coupling according to claim 1, characterized in that: The ball mill rotates at 400 rpm.

4. The method for degrading kitchen waste protein by mechanochemical enzyme coupling according to claim 1, characterized in that: The grinding cycle is 10 cycles.

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