Insect peptide protein production method and use of products thereof

Abstract

The present application is a kind of insect peptide protein manufacturing method, comprising: freezing step, freezing insect larvae to make it lose activity;Grinding step, the frozen insect larvae are mixed with pure water, and the insect paste is ground;Enzymolysis step: the insect paste is mixed with protease, and the insect peptide protein with degree of hydrolysis of 70% to 80% is obtained by enzymolysis;Filtering step, remove impurities and oil in the insect paste after enzymolysis;Inactivation step, the protease in the insect paste after enzymolysis is inactivated by heating. The insect peptide protein has high nutrient utilization rate, the insect peptide protein after enzymolysis is rich in amino acids and small molecular proteins, is easy to be absorbed by animals and plants, and can be applied to various products.

Description

Technical Field

[0001] This invention relates to the field of protein technology, and more specifically to a method for manufacturing insect peptide proteins. In particular, it refers to a method for manufacturing insect peptide proteins that applies enzymatic hydrolysis technology to the field of insect protein decomposition, using low-temperature enzymatic hydrolysis to efficiently decompose insect proteins into peptides or amino acids, thereby improving the absorption and utilization rate of proteins. Background Technology

[0002] Prior art includes Taiwan Patent Publication No. 202428544, entitled "Amino Acid Fertilizer Made from Black Soldier Fly Larvae and Method for Manufacturing the Same," which discloses a method for manufacturing amino acid fertilizer using black soldier fly larvae. The method involves inactivating black soldier fly larvae in warm water at 45°C to 65°C; then, after ultrasonic pulverization and heating, the larvae are allowed to stand; finally, deionized water, molasses, and symbiotic bacteria are added to the larvae in a specific ratio, followed by fermentation to produce an amino acid. This amino acid is then mixed with kaolin to form an amino acid fertilizer. The amino acid fertilizer is produced by mixing 10% to 15% by weight of amino acid with 85% to 90% by weight of kaolin. This amino acid fertilizer contains at least the following components: nitrogen, phosphorus, potassium, calcium, magnesium, and organic matter, thereby providing nutrients needed by various plants to replace existing plant and animal proteins.

[0003] Another example is the Chinese Patent Publication No. CN117682911A, "Preparation Method and Application of Black Soldier Fly Body Amino Acid Liquid Fertilizer," which discloses a preparation method and application of black soldier fly body amino acid liquid fertilizer. This preparation method utilizes farmed black soldier fly larvae and employs an optimized enzymatic hydrolysis process. Through this enzymatic hydrolysis process, the total amino acid content of the black soldier fly larvae hydrolysate reaches 100.739 g / L, with a relatively balanced amino acid composition. Except for methionine and serine, which have lower contents, the contents of the other 14 amino acids are all greater than 4 g / L, and the calcium content is high, which is beneficial to plant growth and absorption. This preparation method has low raw material costs, shortens processing time, and produces significant product effects. The black soldier fly body amino acid liquid fertilizer prepared by this invention has a significant promoting effect on crop growth and can improve crop yield and quality.

[0004] The prior art has two drawbacks. First, the process of extracting insect peptide proteins using dried insects is energy-intensive, time-consuming, and requires the use of chemical agents (strong acids and strong alkalis) as buffer solutions. The process is not environmentally friendly and produces chemical residues, and the cost is also high. Second, the environmental parameters of the enzymatic hydrolysis process in the prior art can lead to a lower degree of hydrolysis of insect peptide proteins, which in turn leads to lower nutritional absorption efficiency, insufficient release of functional components, and potentially poor taste and solubility. Summary of the Invention

[0005] This invention proposes a method for manufacturing insect peptide proteins, comprising: The freezing step involves freezing the insect larvae to render them inactive; The grinding step involves mixing the frozen insect larvae with pure water and grinding them into an insect paste. Enzymatic hydrolysis step: This enzymatic hydrolysis step involves adding the insect slurry to a protease mixture and then hydrolyzing it to obtain insect peptide proteins with a degree of hydrolysis of 70% to 80%. The filtration step removes impurities and oils from the enzymatically hydrolyzed insect slurry. The inactivation step involves heating the enzymatically hydrolyzed insect slurry to inactivate the protease in the slurry.

[0006] Furthermore, the freezing step involves fasting the insect larvae for 5 to 24 hours, then freezing the larvae at 10°C to 20°C for 6 to 12 hours to render them inactive.

[0007] Furthermore, the grinding step involves mixing the insect larvae that have undergone the aforementioned freezing step with pure water in a weight ratio of 2:1 to 5:3, and grinding them into insect paste at a rotation speed of 2500 RPM to 3500 RPM.

[0008] Furthermore, the enzymatic hydrolysis step involves mixing the insect plasma with a protease and then ultrasonically agitating the mixture. The insect plasma and the protease are mixed in a weight ratio between 1:0.005 and 1:0.06. The ultrasonic agitation is performed at a temperature between 34°C and 44°C and a pH value between 3.0 and 9.0, with a frequency between 18kHz and 24kHz, and a duration between 1 minute and 30 minutes, to obtain the insect peptide protein with a degree of hydrolysis of 70% to 80%.

[0009] Furthermore, the filtration step involves removing solid impurities from the insect slurry after the above enzymatic hydrolysis step through vacuum filtration, and then allowing it to stand in an environment with a temperature between 5°C and 12°C for 5 to 30 minutes to remove the solidified grease on the surface of the insect slurry. The inactivation step involves heating the insect slurry after the filtration step in an environment with a temperature between 50°C and 60°C for 5 to 30 minutes to inactivate the protease in the insect slurry.

[0010] Furthermore, it includes an evaporation step, in which the enzymatically hydrolyzed insect plasma is heated in an environment at a temperature of 50°C to 60°C for 20 to 120 minutes to evaporate the water in the insect plasma and concentrate the insect plasma, thereby increasing the purity of the insect peptide protein by 25% to 80%.

[0011] Furthermore, the species of insect larvae include one or a combination of black soldier fly, mealworm, cricket, silkworm pupa, bee pupa, fly larva, earthworm, mealworm, superworm, mosquito larva, bloodworm and locust.

[0012] Furthermore, the types of proteases include one or a combination of pepsin, papain, bromelain, kiwifruit enzyme, peptidase, figase, and calpain.

[0013] The present invention also provides a use of the product manufactured according to the aforementioned method for manufacturing insect peptide protein, which is used as a nutritional supplement in the process of cultivating Chinese cabbage. The diluted insect peptide protein is used to cultivate Chinese cabbage to provide the amino acids and nutrients required for the growth of Chinese cabbage. The dilution ratio ranges from 500 times to 1000 times.

[0014] The following effects can be achieved based on the above technical features: 1. The present invention has a high nutritional utilization rate. Compared with conventional peptide proteins, the insect peptide proteins hydrolyzed by the present invention are rich in amino acids and small molecule proteins, which are more easily absorbed by animals and plants.

[0015] 2. This invention uses biotechnology for decomposition, reducing the impact of chemical drugs on the environment and minimizing damage to the Earth's ecosystem.

[0016] 3. This invention can be applied to a variety of products. Insect peptide proteins can be used in agricultural irrigation, livestock and aquaculture feed to improve crop quality and increase the absorption efficiency of plants and animals. The nutritional composition of the product can also be adjusted according to different insects and enzyme formulations.

[0017] 4. The insect larvae organic waste of the present invention has a high conversion rate and can be converted into nutrient-rich biomass and protein, requiring much less water, land and feed than traditional livestock (such as cattle, pigs, sheep and chickens).

[0018] 5. This invention overcomes the shortcomings of previous technologies in manufacturing dried insects, such as high power consumption, time consumption, and chemical residues, by freezing insect larvae. The pretreatment method of freezing insect larvae has the advantages of being environmentally friendly, safe, fast, and pollution-free.

[0019] 6. This invention involves grinding insect larvae with pure water in a specific ratio, then mixing the ground insect pulp with protease in a weight ratio between 1:0.005 and 1:0.06. Enzymatic hydrolysis is carried out under controlled conditions of 34°C to 44°C and pH value between 3.0 and 9.0. The enzymatic hydrolysis process is performed using ultrasound at a frequency between 18kHz and 24kHz for 1 to 30 minutes. According to experiments, the degree of hydrolysis of the obtained insect peptide protein can be as high as 70% to 80%. Attached Figure Description

[0020] Figure 1 This is a flowchart of an embodiment of the present invention.

[0021] Explanation of icon numbers: S1 Freezing step; S2 grinding step; S3 enzymatic hydrolysis step; S4 filtration steps; S5 deactivation step; S6 Evaporation Step. Detailed Implementation

[0022] To further explain the technical solution of the present invention, the present invention will be described in detail below through specific embodiments.

[0023] Please refer to the following first. Figure 1 The process of this embodiment of the invention is described, including freezing step S1, grinding step S2, enzymatic hydrolysis step S3, filtration step S4, inactivation step S5 and evaporation step S6.

[0024] First, freezing step S1 involves freezing the insect larvae that have been raised to the collection stage, such as 4th to 5th instar black soldier fly larvae, at -20°C for 12 hours after fasting for 24 hours, so that the black soldier fly larvae lose their activity and are then cleaned.

[0025] The next step is grinding step S2, which involves mixing frozen black soldier fly larvae with pure water at a weight ratio of 2:1 to dilute the insect paste, and then grinding it with a high-speed mixer at a speed of 2500 RPM to 3500 RPM to form a homogeneous insect paste.

[0026] The next step, S3, involves mixing insect plasma with protease at a weight ratio of 1:0.01. In S3, the mixture is placed at pH 7.0 and 40°C, and shaken at 20 kHz for 20 minutes to obtain insect peptide proteins.

[0027] The next step is filtration step S4, which uses vacuum filtration to remove solid impurities from the insect paste. After filtration, the paste is left to stand at 10°C for 20 minutes to facilitate the scraping off of condensed grease from the surface of the insect paste.

[0028] The next step is inactivation step S5, which involves heating the filtered insect slurry to 60°C and continuing to heat for 10 minutes to completely inactivate the protease and obtain stable insect peptide proteins.

[0029] Insect peptides can be applied in agriculture, animal husbandry, and aquaculture. In agriculture, diluted insect peptides can be used for irrigation or foliar spraying of plants to provide the necessary amino acids and nutrients, thereby improving crop yield and quality. Specific benefits include promoting cell division and growth, particularly in root and lateral bud development; helping to delay leaf senescence and maintain green foliage; and promoting lateral branching, increasing the number of branches and thus increasing crop yield. For example, insect peptides can be used as a nutritional supplement in the cultivation of bok choy. This is achieved by diluting the insect peptides with deionized water (500-1000 times) to provide the necessary amino acids and nutrients for bok choy growth. In animal husbandry, diluted insect peptide protein can be added to drinking water for livestock. In aquaculture, it can be used as a feed additive or processed into powder or granules to provide a protein source. Specifically, it contains cellulose, minerals, vitamins, and amino acids. Regarding cellulose, insect peptide protein generally has a low fiber content, approximately 5% to 10%, making it digestible for animals. Regarding minerals, it contains important minerals such as nitrogen, potassium, calcium, zinc, phosphorus, iron, magnesium, manganese, copper, and molybdenum, all essential for the health of plants and animals. Regarding vitamins, it contains various vitamins, including B vitamins, which help strengthen the immune system and promote growth. As for amino acids, insect peptide protein... Providing a complete amino acid composition, including all essential amino acids, insect larvae are particularly important for animal nutrition. They contain relatively high levels of micronutrients such as nitrogen, phosphorus, potassium, magnesium, minerals (calcium, zinc, iron, molybdenum, copper, and manganese), and vitamins (A, K, D, E, C, B1, B2, and B6), offering a variety of nutritional benefits. These components enable insect peptide proteins to provide another new protein source in plant and animal industries, especially in fisheries, agriculture, and livestock farming. They can also be further applied in the cosmetic, medical, and food industries. For example, insect peptide proteins can be used in whitening and moisturizing lotion formulas with the following ingredients: 100ml to 150ml rose water, 5ml to 20ml insect peptide protein, 0.5% to 2% hyaluronic acid, and 1ml to 5ml simple emulsifier.

[0030] It is worth mentioning that the insect peptide protein produced by the insect larvae of the present invention after enzymatic hydrolysis step S3 has a degree of hydrolysis of up to 70% to 80% compared with the insect peptide protein obtained by conventional enzymatic hydrolysis process with environmental parameters.

[0031] In addition, conventional extraction methods are quite power-consuming, time-consuming, costly, and require the use of chemical reagents (strong acids and strong bases) as buffer solutions. The process is not environmentally friendly and produces chemical residues. The freezing step S1 can improve the aforementioned disadvantages of power consumption, time consumption, and high cost.

[0032] The manufacturing process described in this case is energy-saving, environmentally friendly, safe, rapid, and pollution-free. It yields insect peptide proteins with high purity (i.e., high concentration). A variety of combined proteases are used as biocatalysts to synthesize protein hydrolysates with high degrees of hydrolysis, ranging from 52.4% to 68.6%. The optimal conditions for hydrolysis are an enzyme concentration of 0.5% to 6%, a pH of 3.0 to 9.0, and a hydrolysis time of 1 to 30 minutes, resulting in a DH of 52.4% to 68.6%. The hydrolysates of the insect peptide proteins contain a large amount of hydrophobic essential amino acids, particularly glutamic acid (14.04%), aspartic acid (9.90%), and glycine (7.72%).

[0033] In addition to protein, insects also contain minerals, vitamins, fiber, and healthy fatty acids such as omega-6 and omega-3. Furthermore, insect products selectively promote the growth of beneficial bacteria in the gut microbiota. In short, insects are rich in protein, essential amino acids, healthy fats, fiber, vitamins, and minerals, providing the nutrients the human body needs.

[0034] Therefore, this invention has the advantages of high nutrient utilization, green environmental protection, and diversified products. The enzymatically hydrolyzed insect peptide protein is rich in amino acids and small molecule proteins, which are easily absorbed by plants and animals. At the same time, the use of biotechnology for decomposition reduces the impact of chemical drugs on the environment. The nutritional composition of the product can also be adjusted according to different insects and enzyme formulas to enhance product diversity. The conversion rate of insect organic waste is very high, which can be converted into nutrient-rich biomass and is also very efficient in converting it into protein. This requires much less water, land, and feed than traditional livestock (such as cattle, pigs, sheep, and chickens). In addition, insect farming produces less greenhouse gas emissions and less waste, making it a more sustainable choice for protein production.

[0035] Furthermore, in the evaporation step S6, the deactivated insect plasma can be heated at 55°C for 20 minutes to promote water evaporation, deactivate active enzymes, and stabilize them, thereby increasing the purity of the insect peptide protein. Preferably, the purity of the insect peptide protein can be increased by 25%-80%, to 1.25-1.8 times the original purity.

[0036] Using medium- and low-temperature enzymatic hydrolysis of enzymes and a wide pH range can create optimal conditions for protease activity. The short hydrolysis time and low enzyme concentration can obtain a large number of small peptide molecules. The specificity of the protease will affect the amino acid residues of the peptides, and the hydrolysis products have unique nutritional and functional properties.

[0037] It is worth mentioning that there are various combinations of protease addition, including single, multiple and continuous protease addition, protease with dual endopeptidase and exopeptidase activity, which can promote hydrolysis and generate peptides with significant biological activity, multi-enzyme addition and simultaneous addition of two or more enzymes.

[0038] To improve hydrolysis efficiency and increase the diversity of generated insect bioactive peptides, producing a wider range of peptides with different bioactivities, enzymes are added to the protein solution in a specific sequence in a continuous enzyme addition system. Compared to methods using a single enzyme, this increases the degree of hydrolysis and peptide yield. The results of the analysis of insect peptide proteins using high-performance liquid chromatography (HPLC) are shown in the table below:

[0039] Based on the above description of the embodiments, one can fully understand the operation, use and effects of the present invention. However, the above embodiments are only preferred embodiments of the present invention and should not be construed as limiting the scope of the present invention. Simple equivalent changes and modifications made in accordance with the scope of the patent application and the description of the invention are all within the scope of the present invention.

Claims

1. A method for manufacturing insect peptide proteins, characterized in that, Include: The freezing step involves freezing the insect larvae to render them inactive; The grinding step involves mixing the frozen insect larvae with pure water and grinding them into an insect paste. Enzymatic hydrolysis step: This enzymatic hydrolysis step involves adding the insect slurry to a protease mixture and then hydrolyzing it to obtain insect peptide proteins with a degree of hydrolysis of 70% to 80%. The filtration step removes impurities and oils from the enzymatically hydrolyzed insect slurry. The inactivation step involves heating the enzymatically hydrolyzed insect slurry to inactivate the protease in the slurry.

2. The method for manufacturing insect peptide proteins according to claim 1, characterized in that, The freezing step involves fasting the insect larvae for 5 to 24 hours, then freezing them at 10°C to 20°C for 6 to 12 hours to render them inactive.

3. The method for manufacturing insect peptide proteins according to claim 1, characterized in that, The grinding step involves mixing the insect larvae, which have undergone the aforementioned freezing step, with pure water in a weight ratio of 2:1 to 5:3, and grinding them into insect paste at a speed of 2500 RPM to 3500 RPM.

4. The method for manufacturing insect peptide proteins according to claim 1, characterized in that, The enzymatic hydrolysis step involves mixing the insect plasma with a protease and then ultrasonically agitating the mixture. The insect plasma and the protease are mixed in a weight ratio between 1:0.005 and 1:0.

06. The ultrasonic agitation is performed at a temperature between 34°C and 44°C and a pH value between 3.0 and 9.0, at a frequency between 18kHz and 24kHz, for a duration between 1 minute and 30 minutes, to obtain the insect peptide protein with a degree of hydrolysis of 70% to 80%.

5. The method for manufacturing insect peptide proteins according to claim 1, characterized in that, The filtration step involves removing solid impurities from the insect slurry after the enzymatic hydrolysis step through vacuum filtration, and then allowing it to stand in an environment with a temperature between 5°C and 12°C for 5 to 30 minutes to remove the solidified grease from the top layer of the insect slurry. The inactivation step involves heating the insect slurry after the filtration step in an environment with a temperature between 50°C and 60°C for 5 to 30 minutes to inactivate the protease in the insect slurry.

6. The method for manufacturing insect peptide proteins according to claim 1, characterized in that, The process also includes an evaporation step, in which the enzymatically hydrolyzed insect plasma is heated in an environment at a temperature of 50°C to 60°C for 20 to 120 minutes to evaporate the water in the insect plasma and concentrate it, thereby increasing the purity of the insect peptide protein by 25% to 80%.

7. The method for manufacturing insect peptide proteins according to claim 1, characterized in that, The species of insect larvae include one or a combination of black soldier fly, mealworm, cricket, silkworm pupa, bee pupa, fly larva, earthworm, mealworm, superworm, mosquito larva, bloodworm and locust.

8. The method for manufacturing insect peptide proteins according to claim 1, characterized in that, The types of proteases include pepsin, papain, bromelain, kiwifruit enzyme, peptidase, figase, calpain, or a combination thereof.

9. Use of a product manufactured by the method for manufacturing insect peptide proteins according to any one of claims 1 to 8, characterized in that, It is a nutritional supplement used in the cultivation of Chinese cabbage. The diluted insect peptide protein is used to cultivate Chinese cabbage, providing the amino acids and nutrients required for the growth of Chinese cabbage. The dilution ratio ranges from 500 to 1000 times.

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