Method for extracting collagen peptide based on multi-wavelength illumination assistance

Abstract

The invention discloses a method for extracting collagen peptide based on multi-wavelength illumination assistance, and belongs to the technical field of collagen peptide preparation. The method comprises the following steps: degreasing and crushing bone raw materials, mixing the crushed bone raw materials with water to prepare a suspension, and carrying out alkali extraction and acid precipitation to obtain a protein solution; a compound enzyme is added into the protein solution, multi-wavelength illumination auxiliary treatment of blue light of 430-470 nm and / or red light of 620-680 nm is synchronously applied in the enzymolysis process, the illumination intensity is 10-100 mW / cm < 2 >, and the time is 30-120 min; and filtering, concentrating and drying the enzymatic hydrolysate to obtain a collagen peptide product. According to the method, through the synergistic effect of multiband illumination and enzymolysis, the extraction efficiency of collagen peptide and the yield of small molecule peptide are remarkably improved, the biological activity of the product is effectively protected, and the method has the advantages of being mild in condition, capable of saving energy and reducing consumption, environmentally friendly, efficient and the like.

Description

Technical Field

[0001] This invention belongs to the field of collagen peptide preparation technology, specifically relating to a method for extracting collagen peptides based on multi-wavelength light assisted extraction. Background Technology

[0002] Collagen peptides, as small molecular weight active fragments obtained from the controlled degradation of collagen, are increasingly in demand in functional foods, high-end skincare products, medical dressings, and nutritional supplements due to their excellent biocompatibility, high absorbability, and various physiological functions (such as moisturizing, anti-oxidation, promoting skin health, and joint repair). Their application value is highly dependent on the efficiency and yield of the extraction technology, as well as the degree of protection of the product's bioactivity.

[0003] Currently, the mainstream method for extracting collagen peptides industrially is enzymatic hydrolysis. This method uses proteases (such as trypsin, papain, and neutral proteases) to specifically hydrolyze the peptide bonds of collagen under mild conditions. Compared to traditional acid or alkaline methods, it better avoids drastic damage to the helical structure, reduces the use of harmful chemicals, and thus yields peptides with a more concentrated molecular weight distribution and more complete functionality. However, conventional enzymatic hydrolysis technology still has several significant bottlenecks. First, the enzymatic hydrolysis process is time-consuming, usually requiring several hours or even tens of hours, and production efficiency needs to be improved. Second, to increase the yield, it is often necessary to increase the amount of enzyme used, resulting in high production costs. Third, the reaction efficiency of enzymes is limited by the probability of contact between the substrate and the enzyme and the mass transfer efficiency; enzyme molecules cannot fully penetrate into the interior of collagen fibers, and the limited number of sites of action leads to incomplete extraction.

[0004] To overcome these limitations, researchers have attempted to introduce physical field-assisted techniques to enhance the enzymatic hydrolysis process, such as ultrasound, microwaves, and high-pressure homogenization. Ultrasonic cavitation can disrupt collagen structure, increasing the contact area between the enzyme and substrate; microwaves accelerate molecular motion through in vivo heating. However, these techniques often involve strong thermal effects or shear forces, potentially leading to localized overheating, enzyme inactivation, or changes in peptide structure, affecting the activity of the final product. Furthermore, they are energy-intensive and require complex equipment.

[0005] Existing research has shown that appropriate light exposure can influence the spatial conformation of enzymes through photochemical effects, moderately enhancing the binding ability of their active sites to substrates. Simultaneously, light energy may weaken secondary bonds such as hydrogen bonds between collagen molecules, making their structure looser and easier for enzyme molecules to approach and cleave. However, current research largely focuses on single-wavelength light sources (such as ultraviolet light or single visible light), which have limited modes of action, limited synergistic effects, and may pose a risk of photodegradation, failing to simultaneously achieve efficient cell disruption and enzyme activity regulation. Summary of the Invention

[0006] To address the problems of insufficient extraction efficiency, easy loss of peptide activity, and high cost in existing collagen peptide extraction technologies, there is an urgent need to develop a green new method that can synergistically utilize the characteristics of different wavelengths of light energy to extract collagen peptides, significantly improving extraction efficiency, reducing enzyme dosage, and maximizing the protection of collagen peptide bioactivity. The purpose of this invention is to provide a method for extracting collagen peptides based on multi-wavelength light-assisted extraction. By applying specific wavelengths of light (including blue and red light) during the enzymatic hydrolysis of bone tissue, the efficiency of the enzymatic hydrolysis reaction and the characteristics of the products are controlled, achieving targeted regulation of collagen breakage sites and enrichment of oligopeptides, effectively improving peptide yield, small molecule peptide content, and functional activity.

[0007] To achieve the above objectives, the technical solution of the present invention is as follows:

[0008] In a first aspect, the present invention discloses a method for extracting collagen peptides based on multi-wavelength light assisted extraction, the method comprising the following steps:

[0009] (1) After degreasing, crushing and sieving the bone raw materials, deionized water is added to prepare a suspension;

[0010] (2) Step (1) The suspension was subjected to alkaline extraction and acid precipitation to obtain a protein solution;

[0011] (3) Add the complex enzyme to the protein solution in step (2) and simultaneously irradiate with multi-wavelength light to assist enzymatic hydrolysis to obtain the enzymatic hydrolysate;

[0012] (4) The enzymatic hydrolysate from step (3) was filtered, concentrated and dried to obtain collagen peptides;

[0013] The multi-wavelength illumination is selected from blue light with wavelengths of 430-470nm and / or red light with wavelengths of 620-680nm.

[0014] Specifically, the conditions for the multi-wavelength illumination include: an illumination intensity of 10-100 mW / cm². 2 The illumination time is 30-120 minutes.

[0015] Preferably, the conditions for the multi-wavelength illumination include: an illumination intensity of 30-60 mW / cm². 2 The illumination time is 60-100 minutes.

[0016] Specifically, the dosage of the compound enzyme is 10,000-16,000 U / g, which means that 1g of defatted, pulverized and sieved bone raw material needs to be supplemented with 10,000-16,000 U of compound enzyme.

[0017] Specifically, the complex enzyme includes any one or more of neutral protease, trypsin, papain, and alkaline protease.

[0018] Specifically, the enzymatic hydrolysis temperature in step (3) is 40-45℃, and the enzymatic hydrolysis time is 2-5h.

[0019] Specifically, the bone material mentioned in step (1) is mixed with deionized water at a ratio of 1:10 to 1:15.

[0020] Specifically, the filtration in step (4) uses a filter membrane with a molecular cutoff range of 500-10000 Da.

[0021] Specifically, the pulverization method in step (1) includes mechanical pulverization or ball milling, and the screening is done through a 20-200 mesh sieve.

[0022] In some embodiments, the alkaline extraction and acid precipitation method refers to extracting with 0.8-2% sodium hydroxide solution for 1.5-2 hours, followed by adding 0.1% hydrochloric acid to adjust the pH to 3.5-4.0, thereby precipitating the protein.

[0023] Secondly, the present invention discloses a collagen peptide prepared by the aforementioned method.

[0024] This collagen peptide product has a high content of small molecule peptides and retains its functional activity completely, making it suitable for functional foods, medical aesthetics, nutritional interventions and other fields.

[0025] Existing studies have shown that both red and blue light can induce the upregulation of protease and antioxidant enzyme activities in fungi and plants, suggesting that light has the potential to universally regulate proteolysis. Meanwhile, red light photobiological regulation has been widely used to promote collagen and elastin synthesis, indicating that it may also promote the stability and breakage of collagen molecules. Therefore, it is reasonable to speculate that introducing blue-red dual-band light into the enzymatic hydrolysis of animal bone collagen can, on the one hand, enhance the catalytic conformational activity of enzymes through blue light, and on the other hand, promote the breakage of bone collagen molecular chains and the generation of small peptides through red light, thereby achieving a dual-channel synergistic effect, shortening reaction time, increasing the proportion of small peptides, and retaining more active structures.

[0026] Compared with the prior art, the present invention has the following beneficial effects:

[0027] (1) Enzymatic hydrolysis is carried out with the assistance of multi-wavelength light (430-470nm blue light and / or 620-680nm red light). The light energy provides additional energy and may activate enzyme molecules, making their binding with the substrate more efficient, thereby significantly accelerating the hydrolysis process of collagen and improving the peptide yield (up to 80% or more) and extraction efficiency.

[0028] (2) By adopting a combination strategy of blue light and red light, while promoting enzymatic hydrolysis, the red light band may help reduce oxidative stress, protect the active structure of light-sensitive functional amino acids (such as hydroxyproline) and peptides, thereby enabling the final product to maintain higher antioxidant activity (such as DPPH scavenging rate) and other biological functions.

[0029] (3) Light, as a clean physical field aid, reduces the dependence on high-concentration chemical reagents. This method is carried out at a mild temperature (40-45℃), and the enzymatic hydrolysis time is shortened by light enhancement, which reduces energy consumption and enzyme dosage, conforms to the trend of green production, and helps to reduce industrial production costs.

[0030] (4) The light parameters (wavelength, intensity, time) and enzymatic hydrolysis conditions (temperature, time, enzyme type) of the present invention can be precisely controlled, which facilitates the optimization of the process to adapt to different raw materials and target products. It is easy to operate, has high stability, and has good repeatability and industrial scale-up potential. Attached Figure Description

[0031] Figure 1 The results show the DPPH free radical scavenging rate of collagen peptides at different concentrations. Detailed Implementation

[0032] The following embodiments are merely illustrative of the present invention and do not limit the scope of protection of the present invention in any way. For those skilled in the art, all equivalent implementations or modifications made without departing from the spirit of the present invention are within the scope of protection of the present invention.

[0033] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to embodiments.

[0034] Example 1

[0035] (1) Take fresh pig spine, remove fat and membrane, autoclave (121℃, 15min), then crush and pass through a 40-mesh sieve. Add deionized water at a material-to-liquid ratio of 1:10, add 1% NaOH to extract for 2h, then add 0.1% hydrochloric acid to adjust the pH to 3.8 to obtain a protein solution;

[0036] (2) Add neutral protease (10000 U / g) + trypsin (5000 U / g) to the protein solution, and simultaneously irradiate with alternating blue light (450 nm) + red light (650 nm) (40 mW / cm²). 2 (Switch every 10 minutes) 90 minutes, enzymatic hydrolysis at 45℃ for 3 hours to obtain enzymatic hydrolysate;

[0037] (3) Then, the enzyme was inactivated by heating at 90°C for 10 min, and the collagen peptides were obtained by filtration, concentration (50°C, -0.08MPa, 60 min), and drying (air inlet 180°C, air outlet 85°C).

[0038] Example 2

[0039] (1) Take fresh beef tibia, remove fat and membrane, autoclave (121℃, 15min), then crush and pass through a 40-mesh sieve. Add deionized water at a material-to-liquid ratio of 1:10, add 1% NaOH to extract for 2h, and then add 0.1% hydrochloric acid to adjust the pH to 3.8 to obtain a protein solution;

[0040] (2) Add neutral protease (8000 U / g) + papain (3000 U / g) to the protein solution, and simultaneously irradiate with alternating blue light (460 nm) + red light (660 nm) (35 mW / cm²). 2 (Switch every 10 minutes) 80 minutes, enzymatic hydrolysis at 42℃ for 4 hours to obtain enzymatic hydrolysate;

[0041] (3) Then, the enzyme was inactivated by heating at 90°C for 10 min, and the collagen peptides were obtained by filtration, concentration (50°C, -0.08MPa, 60 min), and drying (air inlet 180°C, air outlet 85°C).

[0042] Example 3

[0043] (1) Take fresh fish scales and bones, remove fat and membranes, autoclave (121℃, 15min), then pulverize and pass through a 40-mesh sieve. Add deionized water at a material-to-liquid ratio of 1:15, add 0.8% NaOH to extract for 1.5h, then add 0.1% hydrochloric acid to adjust the pH to 3.9 to obtain a protein solution;

[0044] (2) Add neutral protease (10000 U / g) to the protein solution, and simultaneously irradiate with blue light (450 nm) + red light (660 nm) continuously (30 mW / cm²). 2 The enzyme hydrolysate was obtained by enzymatic hydrolysis at 40℃ for 4 hours after 60 minutes.

[0045] (3) Then, the enzyme was inactivated by heating at 90°C for 10 min, and the collagen peptides were obtained by filtration, concentration (50°C, -0.08MPa, 60 min), and drying (air inlet 180°C, air outlet 85°C).

[0046] Example 4

[0047] (1) Take fresh chicken breastbone, remove fat and membrane, autoclave (121℃, 15min), then pulverize and pass through a 40-mesh sieve. Add deionized water at a material-to-liquid ratio of 1:15, add 1.2% NaOH to extract for 2h, then add 0.1% hydrochloric acid to adjust the pH to 3.6 to obtain a protein solution;

[0048] (2) Add alkaline protease (12000 U / g) + trypsin (4000 U / g) to the protein solution, and simultaneously irradiate with blue light (445 nm) + red light (680 nm) continuously (50 mW / cm²). 2 The enzyme hydrolysate was obtained by enzymatic hydrolysis at 47℃ for 3 hours after 75 minutes.

[0049] (3) Then, the enzyme was inactivated by heating at 90°C for 10 min, and the collagen peptides were obtained by filtration, concentration (50°C, -0.08MPa, 60 min), and drying (air inlet 180°C, air outlet 85°C).

[0050] Example 5

[0051] (1) Take fresh pig spine, remove fat and membrane, autoclave (121℃, 15min), then crush and pass through a 40-mesh sieve. Add deionized water at a material-to-liquid ratio of 1:10, add 1% NaOH to extract for 2h, then add 0.1% hydrochloric acid to adjust the pH to 3.8 to obtain a protein solution;

[0052] (2) Add neutral protease (10000 U / g) + trypsin (5000 U / g) to the protein solution, and simultaneously irradiate with blue light (450 nm) (40 mW / cm²). 2 The enzyme hydrolysate was obtained by enzymatic hydrolysis at 45℃ for 3 hours after 90 minutes.

[0053] (3) Then, the enzyme was inactivated by heating at 90°C for 10 min, and the collagen peptides were obtained by filtration, concentration (50°C, -0.08MPa, 60 min), and drying (air inlet 180°C, air outlet 85°C).

[0054] Example 6

[0055] (1) Take fresh pig spine, remove fat and membrane, autoclave (121℃, 15min), then crush and pass through a 40-mesh sieve. Add deionized water at a material-to-liquid ratio of 1:10, add 1% NaOH to extract for 2h, then add 0.1% hydrochloric acid to adjust the pH to 3.8 to obtain a protein solution;

[0056] (2) Add neutral protease (10000 U / g) + trypsin (5000 U / g) to the protein solution, and simultaneously irradiate with red light (650 nm) (40 mW / cm²). 2 The enzyme hydrolysate was obtained by enzymatic hydrolysis at 45℃ for 3 hours after 90 minutes.

[0057] (3) Then, the enzyme was inactivated by heating at 90°C for 10 min, and the collagen peptides were obtained by filtration, concentration (50°C, -0.08MPa, 60 min), and drying (air inlet 180°C, air outlet 85°C).

[0058] Comparative Example 1

[0059] Compared with Example 1, the difference is that only enzymatic hydrolysis was performed, without light-assisted hydrolysis.

[0060] (1) Take fresh pig spine, remove fat and membrane, autoclave (121℃, 15min), then crush and pass through a 40-mesh sieve. Add deionized water at a material-to-liquid ratio of 1:10, add 1% NaOH to extract for 2h, then add 0.1% hydrochloric acid to adjust the pH to 3.8 to obtain a protein solution;

[0061] (2) Add neutral protease (10000U / g) + trypsin (5000U / g) to the protein solution and hydrolyze at 45℃ for 3h to obtain the enzymatic hydrolysate;

[0062] (3) Then, the enzyme was inactivated by heating at 90°C for 10 min, and the collagen peptides were obtained by filtration, concentration (50°C, -0.08MPa, 60 min), and drying (air inlet 180°C, air outlet 85°C).

[0063] Comparative Example 2

[0064] The difference compared to Example 1 is that only light exposure was used, without enzymatic hydrolysis.

[0065] (1) Take fresh pig spine, remove fat and membrane, autoclave (121℃, 15min), then crush and pass through a 40-mesh sieve. Add deionized water at a material-to-liquid ratio of 1:10, add 1% NaOH to extract for 2h, then add 0.1% hydrochloric acid to adjust the pH to 3.8 to obtain a protein solution;

[0066] (2) Irradiate the protein solution with alternating blue light (450nm) and red light (650nm) (40mW / cm²). 2 (Switch every 10 minutes) 90 minutes;

[0067] (3) Collagen peptides were obtained by filtration, concentration (50℃, -0.08MPa, 60min), and drying (inlet air 180℃, outlet air 85℃).

[0068] Comparative Example 3

[0069] The difference compared to Example 1 is that the enzymatic hydrolysis process is assisted by ultrasound.

[0070] (1) Take fresh pig spine, remove fat and membrane, autoclave (121℃, 15min), then crush and pass through a 40-mesh sieve. Add deionized water at a material-to-liquid ratio of 1:10, add 1% NaOH to extract for 2h, then add 0.1% hydrochloric acid to adjust the pH to 3.8 to obtain a protein solution;

[0071] (2) Add neutral protease (10000U / g) + trypsin (5000U / g) to the protein solution, and simultaneously use ultrasound assistance (200W, 5s on / 5s off) for 20min, and then enzymatically hydrolyze at 45℃ for 3h to obtain the enzymatic hydrolysate.

[0072] (3) Then, the enzyme was inactivated by heating at 90°C for 10 min, and the collagen peptides were obtained by filtration, concentration (50°C, -0.08MPa, 60 min), and drying (air inlet 180°C, air outlet 85°C).

[0073] Comparative Example 4

[0074] Compared with Example 1, the difference is that blue light (450nm) + green light (530nm) are used as auxiliary light.

[0075] (1) Take fresh pig spine, remove fat and membrane, autoclave (121℃, 15min), then crush and pass through a 40-mesh sieve. Add deionized water at a material-to-liquid ratio of 1:10, add 1% NaOH to extract for 2h, then add 0.1% hydrochloric acid to adjust the pH to 3.8 to obtain a protein solution;

[0076] (2) Add neutral protease (10000 U / g) + trypsin (5000 U / g) to the protein solution, and simultaneously irradiate with alternating blue light (450 nm) + green light (530 nm) (40 mW / cm²). 2 (Switch every 10 minutes) 90 minutes, enzymatic hydrolysis at 45℃ for 3 hours to obtain enzymatic hydrolysate;

[0077] (3) Then, the enzyme was inactivated by heating at 90°C for 10 min, and the collagen peptides were obtained by filtration, concentration (50°C, -0.08MPa, 60 min), and drying (air inlet 180°C, air outlet 85°C).

[0078] Example of effect

[0079] The total peptide content, small molecule peptide ratio, hydroxyproline content, and antioxidant properties of the collagen peptides prepared in the above examples and comparative examples were tested.

[0080] Total peptide content was determined using the biuret method.

[0081] (1) Preparation of standard curve: Prepare a series of peptide standard solutions with concentrations (e.g., 0, 1, 2, 3, 4, 5 mg / mL) precisely. Take 6 test tubes and add 1.0 mL of each concentration of standard solution to each tube.

[0082] (2) Dilute the sample to be tested appropriately so that its estimated concentration falls within the range of the standard curve. Take 1.0 mL of the diluted sample solution into a test tube. Add 4.0 mL of biuret reagent to all standard tubes and sample tubes, and vortex thoroughly to mix. Let the mixture stand at room temperature (20-25℃) for 30 minutes. Afterward, use a spectrophotometer at a wavelength of 540 nm, zeroing the instrument with a blank tube (standard tube with a concentration of 0), and measure the absorbance value (OD value) of each tube.

[0083] A standard curve was plotted with the concentration of the standard on the x-axis and the absorbance value on the y-axis. The OD value of the sample tube was substituted into the regression equation of the standard curve to calculate the total peptide concentration in the sample dilution. This concentration was then multiplied by the dilution factor to obtain the total peptide content of the original sample.

[0084] Small molecule peptide percentage detection:

[0085] The trichloroacetic acid (TCA) precipitation method is used because large protein and peptide molecules can be precipitated by a certain concentration of TCA, while small peptide molecules are soluble in TCA solution. After centrifugation, the contents of the supernatant (small peptide molecules) and the precipitate (large protein molecules) are measured separately, allowing the calculation of the proportion of small peptide molecules.

[0086] (1) Pipette a certain volume (V0, e.g., 2.0 mL) of peptide solution (the total peptide content C0 must be known) into a centrifuge tube, add an equal volume of 15% TCA solution (2.0 mL), so that the final TCA concentration is 7.5%. Vortex to mix, and let it stand in a refrigerator at 4°C for 1 hour to precipitate. Centrifuge at 10,000 rpm for 15 minutes at 4°C, completely dissolve the precipitate with a small amount of 0.5 M NaOH solution, and make up to a certain volume (V1, e.g., 2.0 mL).

[0087] (2) The concentrations of small molecule peptides in the supernatant were determined using the biuret method (C0). small ) and the concentration of macromolecular proteins in the dissolved precipitate (C large )

[0088] Small molecule peptide percentage (%) = [C small ×(V0+V TCA [(C0×V0)]×100%; where V TCA For the volume of TCA added, V0 is usually equal to V. TCA .

[0089] Hydroxyproline content was determined by acid hydrolysis-colorimetric method.

[0090] (1) Weigh an appropriate amount of sample (containing approximately 1-10 mg of hydroxyproline) into a hydrolysis tube, add 10 mL of 6M HCl, seal the tube under vacuum or fill it with nitrogen and then tighten the cap. Hydrolyze the solution in an oven at 110°C for 16-24 hours. After cooling, filter and transfer the hydrolysate, adjust the pH to neutral with NaOH solution, and bring the volume to a certain level (V) with distilled water.

[0091] (2) Take an appropriate amount of diluted hydrolysate or standard solution into a test tube, add chloramine-T solution, and oxidize at room temperature for 5 minutes. Then add DMAB colorimetric reagent, mix well, and heat in a 60°C water bath for 20 minutes to ensure complete colorimetric reaction. Remove and cool to room temperature.

[0092] (3) Use a spectrophotometer at a wavelength of 560 nm, zero the instrument with a blank reagent, and measure the absorbance value.

[0093] First, plot the hydroxyproline standard curve. Calculate the hydroxyproline concentration in the test solution based on the sample absorbance. Then, multiply the concentration by the dilution factor and the total volume, and divide by the sample mass to obtain the hydroxyproline content in the sample.

[0094] Antioxidant performance testing: DPPH free radical scavenging capacity determination.

[0095] Prepare a series of solutions of different concentrations using distilled water or buffer solution. Take 2.0 mL of each solution and add 2.0 mL of DPPH ethanol working solution, then vortex to mix. Incubate at room temperature in the dark for 30 minutes. Measure the absorbance (A) of the reaction solution at 517 nm, using anhydrous ethanol as a blank. 样本组 Simultaneously, a control group (with 2.0 mL of water or anhydrous ethanol and 2.0 mL of DPPH ethanol working solution added) and a blank group (with the absorbance of 2.0 mL of sample solution + 2.0 mL of anhydrous ethanol) were set up, and A was measured. 对照组 and A 空白组 ;

[0096] DPPH free radical scavenging rate (%) = [1-(A 样本组 -A 空白组 ) / A 对照组 ×100%.

[0097] As shown in Table 1, the indicators of Example 1 (e.g., total peptide 86.2%, DPPH scavenging rate 78.4%) are significantly better than those of Comparative Example 1 (enzyme only, no light) and Comparative Example 2 (light only, no enzyme), demonstrating a significant synergistic effect between multi-wavelength light irradiation (blue light + red light) and enzymatic hydrolysis. While Example 5 (blue light only + enzyme) is better than Comparative Example 1, it is inferior to Example 1 using both blue and red light. This indicates that, in addition to blue light (presumably its main function is to stimulate enzyme activity and promote hydrolysis), introducing red light (presumably its function is to reduce oxidative stress and protect active structures) is crucial for comprehensively improving product quality (especially antioxidant activity). While the results of Comparative Example 3 (ultrasound-assisted + enzyme) are better than those of traditional enzymatic hydrolysis (Comparative Example 1), they are all lower than those of Example 1 of this invention (light-assisted + enzyme). This indicates that the multi-wavelength light-assisted technology used in this invention is superior to common ultrasound-assisted technology in promoting enzymatic hydrolysis efficiency and protecting product functionality, and has greater application potential. Examples 1-4 also demonstrate that the present invention provides a highly versatile technology that can be adapted to different production needs, rather than a specific solution limited to certain conditions.

[0098] Table 1. Results of total peptide content, small molecule peptide ratio, hydroxyproline content, and antioxidant properties of the collagen peptides of this invention.

[0099]

[0100] Depend on Figure 1 It is evident that the product of Example 1 of this invention exhibits the best antioxidant activity, directly demonstrating the significant advantages and synergistic effect of the "blue light + red light" multi-wavelength combination strategy in enhancing the functional activity of the product. The antioxidant activity of the product is concentration-dependent; the DPPH scavenging rate increases with increasing collagen peptide concentration. Regardless of whether used at low or high doses, the collagen peptides prepared by this invention demonstrate stronger antioxidant capacity, and their functional advantages are inherent and universal.

[0101] In summary, the multi-wavelength light irradiation (blue light + red light) assisted enzymatic hydrolysis technology provided by this invention is an innovative method that is highly efficient, green, and can significantly improve the yield and quality of collagen peptides. Blue and red light play different but complementary roles, synergistically promoting enzymatic hydrolysis efficiency and protecting the bioactivity of the products, with effects far superior to single technologies or other physical assistance methods. This technical solution is mature, reliable, and easy to control, exhibiting good stability and repeatability under different raw materials and process parameters, and possesses great potential for industrial application.

Claims

1. A method for extracting collagen peptides based on multi-wavelength light assisted extraction, characterized in that, The method includes the following steps: (1) After degreasing, crushing and sieving the bone raw materials, deionized water is added to prepare a suspension; (2) Step (1) The suspension was subjected to alkaline extraction and acid precipitation to obtain a protein solution; (3) Add the complex enzyme to the protein solution in step (2) and simultaneously irradiate with multi-wavelength light to assist enzymatic hydrolysis to obtain the enzymatic hydrolysate; (4) The enzymatic hydrolysate from step (3) was filtered, concentrated and dried to obtain collagen peptides; The multi-wavelength illumination is selected from blue light with wavelengths of 430-470nm and / or red light with wavelengths of 620-680nm.

2. The method according to claim 1, characterized in that, The conditions for the multi-wavelength illumination include: illumination intensity of 10-100 mW / cm². 2 The illumination time is 30-120 minutes.

3. The method according to claim 2, characterized in that, The conditions for the multi-wavelength illumination include: an illumination intensity of 30-60 mW / cm². 2 The illumination time is 60-100 minutes.

4. The method according to claim 1, characterized in that, The dosage of the complex enzyme is 10,000-16,000 U / g.

5. The method according to claim 4, characterized in that, The complex enzyme includes any one or more of neutral protease, trypsin, papain, and alkaline protease.

6. The method according to claim 1, characterized in that, The enzymatic hydrolysis temperature in step (3) is 40-45℃, and the enzymatic hydrolysis time is 2-5h.

7. The method according to claim 1, characterized in that, The bone material described in step (1) is mixed with deionized water at a ratio of 1:10 to 1:

15.

8. The method according to claim 1, characterized in that, The filtration in step (4) uses a filter membrane with a molecular cutoff range of 500-10000 Da.

9. The method according to claim 1, characterized in that, The pulverization method described in step (1) includes mechanical pulverization or ball milling, and the screening is performed through a 20-200 mesh sieve.

10. A collagen peptide prepared by the method according to any one of claims 1-9.

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