Method for extracting collagen peptide from fish byproducts

Abstract

The invention discloses a method for extracting collagen peptide from fish byproducts, which comprises the following steps: putting minced fish bones into an EDTA (Ethylene Diamine Tetraacetic Acid) solution for decalcification treatment to obtain decalcified minced fish bones; putting the minced fish skin into an ethanol solution for degreasing treatment to obtain degreased minced fish skin; putting the minced fish scales into an organic acid solution for deashing treatment to obtain deashed minced fish scales; mixing the decalcified minced fish bone, the degreased minced fish skin and the deashed minced fish scale, then adding a compound enzyme for enzymolysis treatment, and filtering to obtain an enzymolysis mixture; recovering the compound enzyme in the enzymolysis mixture to obtain enzymolysis liquid, then sequentially performing microfiltration, ultrafiltration and nanofiltration to obtain nanofiltration interception liquid, and then desalting and concentrating to obtain collagen peptide; according to the method, a pretreatment process combining microwave-assisted EDTA decalcification, ultrasonic-assisted ethanol degreasing and enzymolysis is adopted, and an immobilized enzyme membrane reactor and a multi-stage membrane separation technology are matched, so that efficient extraction and purification of the fish collagen peptide and resource utilization of byproducts are realized.

Description

Technical Field

[0001] This invention relates to the field of polypeptide extraction technology, and more specifically to a method for extracting collagen peptides from fish by-products. Background Technology

[0002] Fish scales, skin, and bones are the main byproducts of freshwater fish processing, accounting for more than 25% of the fish's fresh weight. In traditional processing, these byproducts are often processed into low-value animal feed, fertilizer, or even discarded, resulting in significant waste. In recent years, to achieve high-value utilization of freshwater fish byproducts, domestic and international scholars have discovered that these byproducts contain large amounts of fish collagen. For example, collagen accounts for approximately 70-80% of the total protein in fish scales, about 80% in fish skin, and a high 20-30% in fish bones. Collagen peptides, as an important bioactive substance, have wide applications in food, medicine, and cosmetics. However, current technologies for extracting collagen peptides from freshwater fish byproducts have many shortcomings:

[0003] 1. Traditional acid-base method: Hydrochloric acid or sodium hydroxide is used to dissolve collagen. Although the operation is simple, the decalcification efficiency is low (usually only 60-70%), and a large amount of saline wastewater is generated, which causes serious environmental pollution. The collagen yield is also low (about 8-10%).

[0004] 2. Enzymatic hydrolysis: Although the extraction rate is high (up to 99%), the enzyme cost is high, the reaction time is long (usually 6-8 hours), and the wastewater treatment after enzymatic hydrolysis is complicated, making it difficult to achieve industrial application.

[0005] 3. Supercritical fluid extraction: This method uses a supercritical fluid (such as CO2) as the extractant. Although low-temperature operation can preserve the activity of collagen, it requires large equipment investment and high energy consumption, making it difficult to apply on a large scale.

[0006] 4. Three-phase system separation: Collagen is separated by forming a three-phase system using salt and solvent, but the recovery rate of enzymes immobilized in the middle liquid layer is low (usually less than 70%), and the utilization value of the lower reaction medium is limited.

[0007] 5. Multi-stage membrane separation technology: The microfiltration-ultrafiltration-nanofiltration cascade system can purify collagen, but it suffers from severe membrane fouling, requires frequent cleaning, and has high maintenance costs.

[0008] 6. Insufficient utilization of by-product resources: Calcium carbonate and enzymatic hydrolysis residue after decalcification are not fully utilized and are only used as low-value feed raw materials, resulting in resource waste; wastewater treatment is complicated and it is difficult to achieve zero discharge.

[0009] To address the aforementioned issues, several improved technologies have emerged in recent years, such as ultrasound-assisted enzymatic hydrolysis, membrane separation technology, and subcritical water extraction. However, these technologies still have limitations in the extraction of collagen peptides from freshwater fish by-products, particularly lacking efficient extraction methods for collagen from fish bones and comprehensive utilization of extraction residues.

[0010] In summary, existing technologies for preparing collagen peptides from freshwater fish processing by-products suffer from problems such as low raw material utilization, low extraction efficiency, and insufficient product quality and resource utilization. Summary of the Invention

[0011] The purpose of this invention is to provide a method for extracting collagen peptides from fish by-products. This method employs a pretreatment process that combines microwave-assisted EDTA decalcification, ultrasound-assisted ethanol defatting, and enzymatic hydrolysis, along with an immobilized enzyme membrane reactor and multi-stage membrane separation technology, to achieve efficient extraction, purification, and resource utilization of fish collagen peptides and by-products.

[0012] In order to achieve the above-mentioned objectives of the present invention, the following technical solution is adopted:

[0013] The first aspect of this invention provides a method for extracting collagen peptides from fish by-products, the method comprising the following steps:

[0014] (a) Remove impurities and clean the fish scales, fish skin and fish bones respectively, and then crush them to obtain fish scale paste, fish skin paste and fish bone paste;

[0015] (b) Under stirring and microwave conditions, fish bone porridge was placed in EDTA solution for decalcification to obtain decalcified fish bone porridge;

[0016] Under stirring and ultrasonic conditions, fish skin paste was placed in an ethanol solution for defatting treatment to obtain defatted fish skin paste;

[0017] Under stirring and microwave conditions, fish scale paste was placed in an organic acid solution for deashing treatment to obtain deashed fish scale paste;

[0018] (c) Mix the decalcified fish bone paste, defatted fish skin paste and deashed fish scale paste, add a compound enzyme for enzymatic hydrolysis, and filter to obtain an enzymatic hydrolysate mixture;

[0019] (d) The complex enzyme in the enzymatic hydrolysate is recovered to obtain the enzymatic hydrolysate, which is then subjected to microfiltration, ultrafiltration and nanofiltration in sequence to obtain the nanofiltration intercept liquid. Then, collagen peptides are obtained by desalting and concentration.

[0020] Preferably, in step (b), the concentration of the EDTA solution is 2%~5%, the pH value is 6.5~7.0, the mass ratio of fish bone slurry to EDTA solution is 1:(5~7), and the decalcification treatment time is 30~60 min.

[0021] Preferably, in step (b), the concentration of the ethanol solution is 75%~95%; the mass ratio of fish skin paste to fish scales is 1:(3~5); and the degreasing treatment time is 10~30 min.

[0022] Preferably, in step (b), the concentration of the organic acid solution is 2% to 5%; the mass ratio of fish scale paste to organic acid solution is 1:(15 to 20); the deashing treatment time is 10 to 30 minutes; and the organic acid includes citric acid.

[0023] Preferably, in step (c), the mass ratio of decalcified fish bone paste, degreased fish skin paste, and deashed fish scale paste is 1:(1~2):1.

[0024] Preferably, in step (c), the complex enzyme is composed of neutral protease and alkaline protease; the mass ratio of neutral protease to alkaline protease is (0.5~2):1; the amount of complex enzyme added is 2%~3% of the mixed mass of decalcified fish bone paste, degreased fish skin paste and deashed fish scale paste.

[0025] Preferably, in step (c), the enzymatic hydrolysis temperature is 40~50℃, the pH value is 7~7.5, and the time is 2~3h.

[0026] Preferably, in step (d), an immobilized enzyme membrane reactor is used to recover the complex enzyme from the enzymatic hydrolysis mixture; the immobilized membrane in the immobilized enzyme membrane reactor is a polyacrylamide gel; the flow rate of the enzymatic hydrolysis mixture is 1~2 L / min; and the temperature is 40~50℃.

[0027] Preferably, in step (d), the microfiltration membrane has a pore size of 0.22 μm, a microfiltration pressure of 0.2~0.4 MPa, and a temperature of 25~35℃; the ultrafiltration membrane has a pore size of 10~100 kDa, an ultrafiltration pressure of 0.25~0.5 MPa, a temperature of 25~35℃, and a pH value of 6.5~7; and the nanofiltration membrane has a pore size of 500 Da, a nanofiltration pressure of 0.5~1 MPa, a temperature of 35~45℃, and a pH value of 3~4.

[0028] Preferably, the method for extracting collagen peptides from fish by-products further includes step (e):

[0029] Calcium carbonate was recovered from the decalcification liquid obtained from the decalcification treatment; hydroxyapatite was recovered from the deashing liquid obtained from the deashing treatment; the enzymatic hydrolysis residue after enzymatic hydrolysis was mixed with plant protein, and the calcium-to-phosphorus ratio was adjusted to (1.5~2):1 to produce high-calcium protein feed.

[0030] Compared with the prior art, the beneficial effects of the present invention include at least the following:

[0031] This invention combines microwave-assisted EDTA decalcification, ultrasound-assisted ethanol degreasing, and microwave-assisted organic acid deashing, increasing the collagen peptide extraction rate from less than 10% in traditional acid-base methods to 85%-90%, and shortening the processing time from 24 hours to less than 1 hour, thus improving extraction efficiency. Furthermore, the immobilized enzyme membrane reactor technology reduces enzyme usage by 70%; ethanol and organic acid degreasing / deashing can be recycled and reused; membrane system energy consumption is reduced by more than 60%, and overall costs are reduced by more than 40%, resulting in significant economic benefits. Additionally, low-temperature operation avoids high-temperature oxidation, maintaining the integrity of collagen peptide molecules and increasing purity from 82%-85% in traditional methods to over 95%, meeting food-grade standards.

[0032] In this invention, EDTA and organic acid wastewater are efficiently recovered through bio-adsorption, ethanol can be recycled, and the wastewater concentrate meets national standards after treatment, achieving green production with zero emissions. At the same time, all by-products are utilized as resources. Calcium carbonate in the decalcification liquid can be used for the preparation of nano-calcium carbonate, hydroxyapatite in the deliming liquid can be used for the preparation of biomaterials, enzymatic hydrolysis residue is used for feed, and wastewater concentrate is used for fertilizer. The resource utilization rate is increased from less than 30% to more than 95%, achieving a closed-loop resource system with zero waste.

[0033] In summary, this invention provides a continuous process for the efficient extraction of fish collagen peptides from freshwater fish processing by-products (fish scales, fish skin, fish bones). Through a pretreatment process combining microwave-assisted EDTA decalcification, ultrasound-assisted ethanol degreasing, microwave-assisted organic acid deashing, and enzymatic hydrolysis, coupled with an immobilized enzyme membrane reactor and multi-stage membrane separation technology, the efficient extraction, purification, and resource utilization of fish collagen peptides and by-products are achieved. This technology features high operability, low cost, and high resource utilization. It can extract high-purity fish collagen peptides (purity above 95%) from fish scales, skin, and bones. Simultaneously, it can recover calcium carbonate from decalcification liquid to prepare nanoscale materials (particle size within 100nm), recover hydroxyapatite from deliming liquid to prepare biomaterials (particle size within 500nm), prepare high-calcium protein feed from enzymatic hydrolysis residue (protein content above 70%, calcium content 1.5-2.0%), use wastewater concentrate as a functional feed additive (total amino acid content above 40%), and prepare organic fertilizer (nitrogen, phosphorus, and potassium content ≥5%) through composting, achieving a zero-waste resource closed loop. Detailed Implementation

[0034] The embodiments of the technical solution of the present invention will be described in detail below with reference to the examples. The following embodiments are only used to illustrate the technical solution of the present invention more clearly, and are therefore only examples, and should not be used to limit the scope of protection of the present invention.

[0035] It should be noted that, unless otherwise stated, the technical or scientific terms used in this application should have the ordinary meaning as understood by one of ordinary skill in the art to which this invention pertains.

[0036] The first aspect of this invention provides a method for extracting collagen peptides from fish by-products, the method comprising the following steps:

[0037] (a) Remove impurities and clean the fish scales, fish skin and fish bones respectively, and then crush them to obtain fish scale paste, fish skin paste and fish bone paste;

[0038] (b) Under stirring and microwave conditions, fish bone porridge was placed in EDTA solution for decalcification to obtain decalcified fish bone porridge;

[0039] Under stirring and ultrasonic conditions, fish skin paste was placed in an ethanol solution for defatting treatment to obtain defatted fish skin paste;

[0040] Under stirring and microwave conditions, fish scale paste was placed in an organic acid solution for deashing treatment to obtain deashed fish scale paste;

[0041] (c) Mix the decalcified fish bone paste, defatted fish skin paste and deashed fish scale paste, add a compound enzyme for enzymatic hydrolysis, and filter to obtain an enzymatic hydrolysate mixture;

[0042] (d) The complex enzyme in the enzymatic hydrolysate is recovered to obtain the enzymatic hydrolysate, which is then subjected to microfiltration, ultrafiltration and nanofiltration in sequence to obtain the nanofiltration intercept liquid. Then, collagen peptides are obtained by desalting and concentration.

[0043] In one embodiment, in step (a), the particle size of the pulverized material is 2-3 mm.

[0044] In one embodiment, in step (b), the stirring speed is 300-500 rpm; the microwave power during the decalcification process is 300-500 W; the ultrasonic power during the degreasing process is 300-500 W; and the microwave power during the deashing process is 200-400 W.

[0045] In one embodiment, in step (b), the concentration of the EDTA solution is 2% to 5%, the pH value is 6.5 to 7.0, the mass ratio of fish bone slurry to EDTA solution is 1:(5 to 7), and the decalcification treatment time is 30 to 60 min.

[0046] In one embodiment, in step (b), the concentration of the ethanol solution is 75%~95%; the mass ratio of fish skin paste to fish scales is 1:(3~5); and the degreasing treatment time is 10~30 min.

[0047] In one embodiment, in step (b), the concentration of the organic acid solution is 2% to 5%; the mass ratio of fish scale paste to organic acid solution is 1:(15 to 20); the deashing treatment time is 10 to 30 minutes; and the organic acid includes citric acid.

[0048] In one embodiment, in step (c), the mass ratio of decalcified fish bone paste, degreased fish skin paste, and deashed fish scale paste is 1:(1~2):1.

[0049] Preferably, in step (c), the complex enzyme is composed of neutral protease and alkaline protease; the mass ratio of neutral protease to alkaline protease is (0.5~2):1; the amount of complex enzyme added is 2%~3% of the mixed mass of decalcified fish bone paste, degreased fish skin paste and deashed fish scale paste.

[0050] In one embodiment, in step (c), the enzymatic hydrolysis treatment temperature is 40~50℃, the pH value is 7~7.5, and the time is 2~3h.

[0051] In one embodiment, in step (c), ultrasonic treatment is performed during the enzymatic hydrolysis process, with an ultrasonic frequency of 20-40 kHz and an ultrasonic density of 100-400 W / m². 3 The ultrasonic treatment time is 10-20 minutes.

[0052] In one embodiment, in step (d), ultrasonic treatment is performed during the ultrafiltration process, with an ultrasonic frequency of 20-40 kHz and an ultrasonic density of 100-400 W / m³. 3 .

[0053] In one embodiment, in step (d), a composite enzyme is recovered from the enzymatic hydrolysis mixture using an immobilized enzyme membrane reactor; the immobilized membrane in the immobilized enzyme membrane reactor is a polyacrylamide gel; the flow rate of the enzymatic hydrolysis mixture is 1-2 L / min; the temperature is 40-50°C; and the immobilized enzyme density is 10-20 mg / cm³. 3 .

[0054] In one embodiment, in step (d), the microfiltration membrane has a pore size of 0.22 μm, a microfiltration pressure of 0.2~0.4 MPa, and a temperature of 25~35℃; the ultrafiltration membrane has a pore size of 10~100 kDa, an ultrafiltration pressure of 0.25~0.5 MPa, a temperature of 25~35℃, and a pH value of 6.5~7; the nanofiltration membrane has a pore size of 500 Da, a nanofiltration pressure of 0.5~1 MPa, a temperature of 35~45℃, and a pH value of 3~4.

[0055] In one embodiment, the method for extracting collagen peptides from fish by-products further includes step (e):

[0056] Calcium carbonate was recovered from the decalcification liquid obtained from the decalcification treatment; hydroxyapatite was recovered from the deashing liquid obtained from the deashing treatment; the enzymatic hydrolysis residue after enzymatic hydrolysis was mixed with plant protein, and the calcium-to-phosphorus ratio was adjusted to (1.5~2):1 to produce high-calcium protein feed.

[0057] In one embodiment, calcium carbonate is recovered by a reverse microemulsion method and produced into calcium carbonate with a particle size of less than 100 nm; hydroxyapatite is recovered by a sol-gel method and produced into hydroxyapatite with a particle size of no more than 500 nm.

[0058] In one embodiment, the plant protein includes soybean meal and / or rapeseed meal; the calcium content in the high-calcium protein feed is 1.5% to 2%; and the protein content is not less than 70%.

[0059] In one embodiment, the fish bone, fish scale, and fish skin processing equipment in step (b) can be linked with the enzymatic hydrolysis reactor in step (c) through a pipeline, and the immobilized enzyme membrane reactor in step (d) can be linked with the multi-stage membrane separation system through a pipeline, so that the steps can achieve continuous production and reduce pollution and energy consumption in intermediate steps.

[0060] In one embodiment, the wastewater concentrate generated during the production process contains amino acids and small peptides after nanofiltration membrane treatment, with a total amino acid content of over 40%, which can be used as a functional feed additive. The nitrogen, phosphorus, and potassium content of the wastewater concentrate is ≥5%, and the heavy metal content meets national standards. After composting treatment, it can be used as organic fertilizer.

[0061] The technical solution of the present invention will be further described in detail below through specific embodiments.

[0062] Example 1

[0063] This embodiment describes a method for extracting collagen peptides from fish by-products (carp). The method for extracting collagen peptides from fish by-products includes the following steps:

[0064] (a) After cleaning the carp scales and skin, mince them into a paste with a particle size of 2-3 mm. After removing impurities from the carp bones, mince them into a paste with a particle size of 2-3 mm to obtain fish scale paste, fish skin paste and fish bone paste.

[0065] (b) Fish bone paste was mixed with 3% EDTA solution at a ratio of 1:6. The microwave power was 400W and the microwave time was 45 minutes. At the same time, the stirring device was turned on and the stirring speed was 450rpm / min. Then, the mixture was filtered to obtain decalcified fish bone paste.

[0066] Fish skin paste was mixed with 85% ethanol solution at a ratio of 1:4, ultrasonicated at 400W for 20 minutes, while simultaneously turning on the stirring device at 350rpm / min; then filtered to obtain defatted fish skin paste.

[0067] Fish scale paste was mixed with a 3% citric acid solution at a ratio of 1:18. The mixture was microwaved at 300W for 20 minutes while the stirring device was turned on at a speed of 400rpm / min. Then, the mixture was filtered to obtain de-ashed fish scale paste.

[0068] (c) Decalcified fish bone paste, defatted fish skin paste and de-ashed fish scale paste were mixed in a mass ratio of 1:1:1. A compound enzyme of neutral protease and alkaline protease (1:1.5) was added, with the enzyme dosage being 2.5% of the mass of the mixed paste. The pH was adjusted to 7.3, the enzymatic hydrolysis temperature was 45℃, and the enzymatic hydrolysis time was 2.5 hours. At the same time, during the enzymatic hydrolysis process, an ultrasonic auxiliary device was turned on with a power of 400W for 15 minutes. After the enzymatic hydrolysis was completed, an enzymatic hydrolysate was obtained.

[0069] (d) Enzyme recovery via immobilized enzyme membrane reactor: The enzymatic hydrolysate is pumped into an immobilized enzyme membrane reactor (polyacrylamide gel-immobilized composite enzyme) for continuous processing to recover the enzyme used for hydrolysis. The flow rate of the hydrolysate is 1.5 L / min, and the temperature is 45 °C. The remaining hydrolysate after the composite enzyme recovery is first pumped into a microfiltration membrane (0.22 μm) treatment device at a pressure of 0.3 M. The process involves applying pressure at 30℃ and a pressure of 0.35 MPa to remove large particulate impurities. The filtrate is then pumped into an ultrafiltration membrane device with a pore size of 50 kDa, a pressure of 0.35 MPa, a temperature of 28℃, and a pH of 6.8 to retain collagen peptides of the target molecular weight. Simultaneously, ultrasound is activated to enhance membrane separation; the specific parameters are an ultrasound frequency of 30 kHz and an ultrasound power density of 300 W / m³. The filtrate is then pumped into a nanofiltration membrane with a pore size of 500 Da, a pressure of 0.7 MPa, a temperature of 40℃, and a pH of 3.5 to desalinate and concentrate collagen peptides with a molecular weight greater than 500 Da and less than 50 kDa, obtaining high-purity collagen peptides (purity up to 99.1%).

[0070] (e) Resource utilization of residues:

[0071] Calcium carbonate in the decalcification solution is recovered by adjusting the pH to neutral or alkaline and then precipitated. Nano-sized calcium carbonate is prepared using a reverse microemulsion method.

[0072] Hydroxyapatite in the deashing solution was recovered by adjusting the pH to neutral and then precipitated, and biological bone repair materials were prepared using the sol-gel method.

[0073] After dehydration of the enzymatic hydrolysis residue, it is mixed with soybean meal at a mass ratio of 1:1, and the calcium-to-phosphorus ratio is adjusted to 1.8:1 to prepare a high-calcium protein feed.

[0074] The wastewater generated in each process is treated by nanofiltration membrane to become a concentrated liquid containing amino acids and small peptides, with a total amino acid content of up to 40%. It can be used as a functional feed additive or mixed with plant waste for composting.

[0075] Example 2

[0076] This embodiment describes a method for extracting collagen peptides from fish by-products (grass carp). The method for extracting collagen peptides from fish by-products includes the following steps:

[0077] (a) After cleaning the grass carp scales and skin, mince them into a paste with a particle size of 2-3 mm. After removing impurities from the grass carp bones, mince them into a paste with a particle size of 2-3 mm. This yields fish scale paste, fish skin paste, and fish bone paste.

[0078] (b) Fish bone paste was mixed with 4% EDTA solution at a ratio of 1:5.5, microwaved at 450W for 35 minutes, with the stirring device turned on at 500rpm. Then, the mixture was filtered to obtain decalcified fish bone paste.

[0079] Fish skin paste was mixed with 90% ethanol solution at a ratio of 1:4.5, ultrasonicated for 18 minutes at a power of 450W, while simultaneously turning on the stirring device at a speed of 380 rpm; then filtered to obtain defatted fish skin paste.

[0080] Fish scale paste was mixed with a 4% citric acid solution at a ratio of 1:16. The mixture was microwaved at 350W for 15 minutes while the stirring device was turned on at 400rpm. Then, the mixture was filtered to obtain de-ashed fish scale paste.

[0081] (c) Decalcified fish bone paste, defatted fish skin paste, and de-ashed fish scale paste were mixed at a mass ratio of 1:1.5:1. A compound enzyme of neutral protease and alkaline protease (2:1) was added, with the enzyme dosage being 3% of the mass of the mixed paste. The pH was adjusted to 7.3, the enzymatic hydrolysis temperature was 48℃, and the enzymatic hydrolysis time was 3 hours. During the enzymatic hydrolysis process, an ultrasonic auxiliary device was turned on with a power of 450W for 12 minutes. After the enzymatic hydrolysis was completed, an enzymatic hydrolysate was obtained.

[0082] (d) Enzyme recovery via immobilized enzyme membrane reactor: The enzyme hydrolysate mixture is pumped into an immobilized enzyme membrane reactor (polyacrylamide gel-immobilized composite enzyme) for continuous processing to recover the enzyme used for hydrolysis. The flow rate of the hydrolysate is 1.8 L / min, and the temperature is 48℃. The remaining hydrolysate after the composite enzyme recovery is first pumped into a microfiltration membrane (0.22 μm) treatment device at a pressure of 0.35 MPa and a temperature of 32℃ to remove large particulate impurities. The filtrate is then pumped into an ultrafiltration membrane device with a pore size of 60 kDa, a pressure of 0.4 MPa, a temperature of 30℃, and a pH of 7 to retain collagen peptides of the target molecular weight. Simultaneously, ultrasound is activated during this process to enhance membrane separation. The specific parameters are: ultrasonic frequency 40 kHz and ultrasonic power density 400 W / m³. The filtrate is then pumped into a nanofiltration membrane for treatment with a pore size of 500 Da, a pressure of 0.8 MPa, a temperature of 42℃, and a pH of 3.8. This is suitable for molecular weights greater than 500 Da. Collagen peptides smaller than 60 kDa are desalted and concentrated to obtain high-purity collagen peptides (purity up to 98.5%).

[0083] (e) Resource utilization of residues:

[0084] Calcium carbonate in the decalcification solution is recovered by adjusting the pH to neutral or alkaline and then precipitated. Nano-sized calcium carbonate is prepared using a reverse microemulsion method.

[0085] Hydroxyapatite in the deashing solution was recovered by adjusting the pH to neutral and then precipitated, and biological bone repair materials were prepared using the sol-gel method.

[0086] After dehydration of the enzymatic hydrolysis residue, it is mixed with rapeseed meal at a mass ratio of 1:1.2, and the calcium-to-phosphorus ratio is adjusted to 1.7:1 to prepare a high-calcium protein feed.

[0087] The wastewater generated in each process is treated by nanofiltration membrane to become a concentrated liquid containing amino acids and small peptides, with a total amino acid content of up to 40%. It can be used as a functional feed additive or mixed with plant waste for composting.

[0088] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features therein. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention, and they should all be covered within the scope of the claims and specification of the present invention.

Claims

1. A method of extracting collagen peptides from fish by-products, characterized by, The method for extracting collagen peptides from fish by-products comprises the following steps: (a) removing impurities and washing fish scales, fish skin and fish bones respectively, and then crushing them to obtain fish scale paste, fish skin paste and fish bone paste; (b) under stirring and microwave conditions, the fish bone paste is placed in an EDTA solution for decalcification treatment to obtain decalcified fish bone paste; under stirring and ultrasonic conditions, the fish skin paste is placed in an ethanol solution for defatting treatment to obtain defatted fish skin paste; under stirring and microwave conditions, the fish scale paste is placed in an organic acid solution for decalcification treatment to obtain decalcified fish scale paste; (c) mixing the decalcified fish bone paste, the defatted fish skin paste and the decalcified fish scale paste, and then adding a compound enzyme for enzymatic treatment and filtration to obtain an enzymatic mixture; (d) recovering the compound enzyme in the enzymatic mixture to obtain an enzymatic solution, and then sequentially performing microfiltration, ultrafiltration and nanofiltration to obtain a nanofiltration flow-through liquid, and then performing desalting and concentration to obtain collagen peptides.

2. The method of claim 1, wherein the collagen peptide is extracted from fish by-products, characterized by, In step (b), the concentration of the EDTA solution is 2-5%, the pH value is 6.5-7.0, the mass ratio of the fish bone paste to the EDTA solution is 1:(5-7), and the decalcification treatment time is 30-60 min.

3. The method of claim 1, wherein the collagen peptide is extracted from fish by-products, characterized by, In step (b), the concentration of the ethanol solution is 75-95%, the mass ratio of the fish skin paste to the fish scale is 1:(3-5), and the defatting treatment time is 10-30 min.

4. The method of claim 1, wherein the collagen peptide is extracted from fish by-products. In step (b), the concentration of the organic acid solution is 2-5%, the mass ratio of the fish scale paste to the organic acid solution is 1:(15-20), the decalcification treatment time is 10-30 min, and the organic acid includes citric acid.

5. The method of claim 1, wherein the collagen peptide is extracted from fish by-products. In step (c), the mass ratio of the decalcified fish bone paste, the defatted fish skin paste and the decalcified fish scale paste is 1:(1-2):

1.

6. The method of claim 1, wherein the collagen peptide is extracted from fish by-products. In step (c), the compound enzyme is composed of neutral protease and alkaline protease, the mass ratio of the neutral protease to the alkaline protease is (0.5-2):1, and the addition amount of the compound enzyme is 2-3% of the mixed mass of the decalcified fish bone paste, the defatted fish skin paste and the decalcified fish scale paste.

7. The method of claim 1, wherein the collagen peptide is extracted from fish by-products. In step (c), the enzymatic treatment temperature is 40-50°C, the pH value is 7-7.5, and the time is 2-3 h.

8. The method of claim 1, wherein the collagen peptide is extracted from fish by-products. In step (d), the compound enzyme in the enzymatic mixture is recovered by using an immobilized enzyme membrane reactor, the immobilized membrane in the immobilized enzyme membrane reactor is polyacrylamide gel, the flow rate of the enzymatic mixture is 1-2 L / min, and the temperature is 40-50°C.

9. The method of claim 1, wherein the collagen peptide is extracted from fish by-products, characterized by, In step (d), the microfiltration adopts a microfiltration membrane with a pore size of 0.22 μm, a microfiltration pressure of 0.2-0.4 MPa and a temperature of 25-35°C, the ultrafiltration adopts an ultrafiltration membrane with a pore size of 10-100 KDa, an ultrafiltration pressure of 0.25-0.5 MPa, a temperature of 25-35°C and a pH value of 6.5-7, and the nanofiltration adopts a nanofiltration membrane with a pore size of 500 Da, a nanofiltration pressure of 0.5-1 MPa, a temperature of 35-45°C and a pH value of 3-4.

10. The method of claim 1, wherein the collagen peptide is extracted from fish by-products. The method for extracting collagen peptides from fish by-products further comprises step (e): The calcium carbonate in the decalcification liquid obtained through decalcification treatment is recovered; the hydroxyapatite in the deashing liquid obtained through deashing treatment is recovered; the enzymolysis residue after enzymolysis treatment is mixed with plant protein, and the calcium-phosphorus ratio is adjusted to (1.5-2):1 to prepare high-calcium protein feed.