Protein powder and preparation method thereof, hydrogel wound dressing and preparation method thereof

Abstract

The invention relates to the technical field of hydrogel, in particular to protein powder and a preparation method thereof, and hydrogel wound dressing and a preparation method thereof. The invention provides protein powder. The protein powder is prepared from the following components: giant salamander collagen freeze-dried powder, a natural eutectic solvent and water, the mass ratio of the giant salamander collagen freeze-dried powder to the natural eutectic solvent is 1: (45-55); the mass volume ratio of the giant salamander collagen freeze-dried powder to the water is 1g: (5-15) mL. The invention further provides the hydrogel wound dressing, the hydrogel wound dressing has good antibacterial performance and anti-inflammatory performance, has the effects of wound monitoring and drug controlled release, and can effectively improve the healing state of a wound.

Description

Technical Field

[0001] The invention relates to the technical field of hydrogels, and in particular to a protein powder and a preparation method thereof, a hydrogel wound dressing and a preparation method thereof. Background Art

[0002] Giant salamanders, known as "ginseng in water", not only contain high-quality protein and essential amino acids, but also contain a variety of unsaturated fatty acids, minerals and vitamins that are beneficial to the human body. In particular, the collagen in the skin of giant salamanders plays an important role in maintaining skin elasticity and luster, promoting wound healing and bone health, and is deeply favored by consumers.

[0003] Giant salamander skin collagen can keep human skin elastic, smooth and delicate, and has a strong skin care effect. At the same time, the ancient regenerative factors in it can induce specific cell proliferation and differentiation, accelerate wound healing, and repair damaged tissues and organs. There is no relevant research on the use of giant salamander skin collagen as a dressing in the prior art.

[0004] In addition, the traditional dressings currently used in medical treatment have high permeability, which can easily lead to wound dehydration, gauze adhesion, pain and bleeding during dressing changes, and increase the risk of infection. Therefore, it is necessary to provide a new wound dressing that can keep the wound moist, create a low-oxygen, slightly acidic environment that is conducive to healing, and relieve pain and avoid secondary injuries. Summary of the invention

[0005] The purpose of the present invention is to provide a protein powder and a preparation method thereof, a hydrogel wound dressing and a preparation method thereof.

[0006] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0007] The present invention provides a protein powder, which is prepared from the following components: giant salamander collagen freeze-dried powder, natural low eutectic solvent and water;

[0008] The mass ratio of giant salamander collagen freeze-dried powder to natural low eutectic solvent is 1:45-55;

[0009] The mass volume ratio of giant salamander collagen freeze-dried powder and water is 1g:5-15mL.

[0010] Preferably, the natural deep eutectic solvent comprises D-xylose and choline chloride;

[0011] The molar mass ratio of D-xylose to choline chloride is 0.5-1.5:0.5-1.5.

[0012] The present invention also provides a method for preparing the protein powder, comprising the following steps:

[0013] (1) mixing giant salamander collagen freeze-dried powder, a natural low eutectic solvent and water, and performing ultrasonic water bath to obtain a collagen glycosylation product;

[0014] (2) The collagen glycosylation product is dialyzed and dried to obtain protein powder.

[0015] Preferably, in step (1), the temperature of the ultrasonic water bath is 60-90° C., the power of the ultrasonic water bath is 1000-1500 W; and the time of the ultrasonic water bath is 50-90 min.

[0016] Preferably, in step (2), the dialysis time is 36 to 48 hours; the molecular weight of the dialysis bag used for dialysis is 250 to 350 Da;

[0017] The vacuum freeze drying includes precooling and low temperature drying; the precooling temperature is -70 to -90°C; the precooling time is 1 to 3 hours; the low temperature drying temperature is 10 to 30°C; the low temperature drying time is 40 to 56 hours;

[0018] The vacuum degree of the drying is 0-5 kPa.

[0019] The present invention also provides a hydrogel wound dressing, comprising the following components in parts by volume: 0.5 to 1.5 parts of a protein mixed solution, 1 to 3 parts of a sodium alginate solution and 3 to 7 parts of a chitosan solution;

[0020] The method for preparing the protein mixed solution comprises the following steps: redissolving the protein powder to a concentration of 0.005 to 0.015 g / mL to obtain a protein solution; mixing curcumin and the protein solution to obtain a protein mixed solution;

[0021] The protein powder is the protein powder described above;

[0022] The mass volume ratio of the curcumin and protein solution is 0.005-0.015 g:1 mL.

[0023] Preferably, the sodium alginate solution uses water as solvent and comprises the following components in final concentrations: 0.015-0.035 g / mL sodium alginate, 0.01-0.03 g / mL polyvinyl alcohol and 0.005-0.015 g / mL glycerol.

[0024] Preferably, the chitosan solution uses water as solvent and comprises the following components in final concentrations: 0.005-0.015 g / mL chitosan, 0.005-0.025 g / mL CaCl2 and 0.005-0.015 mL / mL acetic acid.

[0025] Preferably, the method for mixing the curcumin and the protein solution is a pH driven method.

[0026] The present invention also provides a method for preparing the hydrogel wound dressing, comprising the following steps:

[0027] 1) mixing the protein mixed solution and the sodium alginate solution to obtain a mixed solution;

[0028] 2) pouring the mixed solution into a mold to obtain a mold containing the mixed solution; immersing the mold containing the mixed solution in a chitosan solution to obtain a formed hydrogel;

[0029] 3) demolding and rinsing the formed hydrogel to obtain a hydrogel wound dressing;

[0030] In step 2), the soaking time is 20 to 40 minutes.

[0031] Beneficial effects of the present invention:

[0032] The invention uses natural low eutectic solvent and water to carry out ultrasound-assisted glycosylation modification on giant salamander collagen freeze-dried powder to prepare a protein powder.

[0033] Among them, giant salamander collagen freeze-dried powder has excellent biocompatibility and healing-promoting properties, and it has unique antimicrobial peptides with potential antibacterial properties, which can play an antibacterial role in the wound healing process.

[0034] The present invention also provides a hydrogel wound dressing, to which glycosylated protein (protein solution obtained by re-dissolving protein powder) is added, the mechanical properties of conventional ionic hydrogels are improved, and the reconstruction of epithelial cells in the process of wound healing is beneficial, and the specific antimicrobial peptides can play an antibacterial role in wound healing; curcumin with broad-spectrum antibacterial, antioxidant, anti-inflammatory properties and pH responsiveness is also introduced as an active substance; the glycosylated protein is combined with the sodium alginate chitosan gel network to effectively increase the loading amount of the active substance curcumin, thereby improving the wound healing rate; and the hydrogel prepared after the combination has a high water content, can keep the wound surface moist, reduce the wound temperature, and has a good sustained-release effect on curcumin.

[0035] The hydrogel wound dressing of the present invention has good antibacterial and anti-inflammatory properties, and has the functions of "wound surface monitoring" and "drug controlled release", and can effectively improve the healing state of the wound. BRIEF DESCRIPTION OF THE DRAWINGS

[0036] Figure 1 The infrared spectrum analysis results of Experimental Example 1;

[0037] Figure 2 The differential scanning calorimetry results in Experimental Example 1;

[0038] Figure 3Evaluation of wound healing degree in Experimental Example 2;

[0039] Figure 4 The results of immunofluorescence staining analysis of inflammatory factors (IL-6, TNF-α) in Experimental Example 2;

[0040] Figure 5 The results of immunofluorescence staining of CD31 and α-SMA in Experimental Example 2;

[0041] Figure 6 HE staining and Masson staining in Experimental Example 2;

[0042] Figure 7 The results of the antibacterial experiment in Experimental Example 3 are shown. DETAILED DESCRIPTION

[0043] The present invention provides a protein powder, which is prepared from the following components: giant salamander collagen freeze-dried powder, natural low eutectic solvent and water;

[0044] The mass ratio of giant salamander collagen freeze-dried powder to natural deep eutectic solvent is 1:45-55, preferably 1:48-52, and more preferably 1:50;

[0045] The mass volume ratio of giant salamander collagen freeze-dried powder and water is 1g:5-15mL, preferably 1g:8-12mL, and more preferably 1g:10mL.

[0046] In the present invention, the method for preparing the giant salamander collagen freeze-dried powder comprises the following steps:

[0047] a. Soak the giant salamander skin in a 0.1 mol / mL NaCl solution for 20 to 28 h, and wash with distilled water to obtain the initially treated giant salamander skin;

[0048] b. The initially treated giant salamander skin and isopropanol (purity 99%) were mixed at a mass volume ratio of 1g: 8 to 12mL for 20 to 28h, and washed with distilled water to obtain the treated giant salamander skin;

[0049] c. The treated giant salamander skin was crushed using a beater and dried to obtain a giant salamander skin homogenate powder;

[0050] d. The giant salamander skin homogenate powder and the mixed solvent were mixed in a mass ratio of 1:18 to 22, heated, dialyzed, centrifuged, and the supernatant was taken; the supernatant was freeze-dried to obtain giant salamander collagen freeze-dried powder;

[0051] In step a, the soaking time is preferably 24 hours;

[0052] In step b, the mass volume ratio of the initially treated giant salamander skin and isopropanol is preferably 1 g:10 mL, and the mixing time is preferably 24 h;

[0053] In step c, the drying method is vacuum freeze drying, which includes precooling and low-temperature drying; the precooling temperature is -70 to -90°C, preferably -80°C; the precooling time is 1 to 3 hours, preferably 2 hours; the low-temperature drying temperature is 10 to 30°C, preferably 20°C; the low-temperature drying time is 40 to 56 hours, preferably 48 hours;

[0054] The vacuum degree of the drying is 0-5 kPa, preferably 2.5 kPa;

[0055] In step d, the mixed solvent comprises anhydrous oxalic acid and choline chloride; the molar mass ratio of the anhydrous oxalic acid to the choline chloride is 0.5-1.5:0.5-1.5, preferably 0.8-1.2:0.8-1.2, and more preferably 1:1;

[0056] The preparation method of the mixed solvent comprises the following steps: mixing anhydrous oxalic acid and choline chloride to obtain a mixed solvent; the mixing temperature is 70 to 90° C., preferably 80° C.;

[0057] The mass ratio of the giant salamander skin homogenate powder to the mixed solvent is preferably 1:20;

[0058] In step d, the heating method is water bath heating; the heating temperature is 50-70°C, preferably 60°C; the heating time is 4-6h, preferably 5h;

[0059] In step d, the dialysis time is 40 to 56 hours, preferably 48 hours;

[0060] In step d, the centrifugal speed is preferably 10000-14000 rpm, more preferably 12000 rpm; the centrifugal time is 5-15 min, more preferably 10 min;

[0061] The freeze-drying method is vacuum freeze drying, which includes precooling and low-temperature drying; the precooling temperature is -70 to -90°C, preferably -80°C; the precooling time is 1 to 3 hours, preferably 2 hours; the low-temperature drying temperature is 10 to 30°C, preferably 20°C; the low-temperature drying time is 40 to 56 hours, preferably 48 hours;

[0062] The vacuum degree of the drying is 0-5 kPa, preferably 2.5 kPa.

[0063] In the present invention, the natural deep eutectic solvent comprises D-xylose and choline chloride;

[0064] The molar mass ratio of D-xylose to choline chloride is 0.5-1.5:0.5-1.5, preferably 0.8-1.2:0.8-1.2, and more preferably 1:1;

[0065] The preparation method of the natural deep eutectic solvent comprises the following steps: mixing D-xylose and choline chloride to obtain a natural deep eutectic solvent;

[0066] The mixing speed is 15 to 25 rpm, preferably 18 to 22 rpm, and more preferably 20 rpm;

[0067] The mixing temperature is 85-95°C, preferably 88-92°C, and more preferably 90°C;

[0068] The mixing time is 2.5 to 3.5 hours, preferably 2.8 to 3.2 hours, and more preferably 3 hours.

[0069] The present invention also provides a method for preparing the protein powder, comprising the following steps:

[0070] (1) mixing giant salamander collagen freeze-dried powder, a natural low eutectic solvent and water, and performing ultrasonic water bath to obtain a collagen glycosylation product;

[0071] (2) The collagen glycosylation product is dialyzed and dried to obtain protein powder.

[0072] In the present invention, in step (1), the temperature of the ultrasonic water bath is 60-90°C, preferably 70-80°C, and more preferably 75°C; the power of the ultrasonic water bath is 1000-1500W, preferably 1200-1300W, and more preferably 1250W; the time of the ultrasonic water bath is 50-90min, preferably 60-80min, and more preferably 70min.

[0073] In the present invention, in step (2), the dialysis time is 36 to 48 hours, preferably 40 to 44 hours, and more preferably 42 hours; the molecular weight of the dialysis bag used for dialysis is 250 to 350 Da, preferably 280 to 320 Da, and more preferably 300 Da; the vacuum freeze-drying includes precooling and low-temperature drying; the precooling temperature is -70 to -90 ° C, preferably -80 ° C; the precooling time is 1 to 3 hours, preferably 2 hours; the low-temperature drying temperature is 10 to 30 ° C, preferably 20 ° C; the low-temperature drying time is 40 to 56 hours, preferably 48 hours;

[0074] The vacuum degree of the drying is 0-5 kPa, preferably 2.5 kPa.

[0075] The present invention also provides a hydrogel wound dressing, comprising the following components in parts by volume:

[0076] 0.5 to 1.5 parts of protein mixed solution, preferably 0.8 to 1.2 parts, more preferably 1 part;

[0077] 1 to 3 parts of sodium alginate solution, preferably 1.5 to 2.5 parts, more preferably 2 parts;

[0078] 3 to 7 parts of chitosan solution, preferably 4 to 6 parts, more preferably 5 parts;

[0079] The method for preparing the protein mixed solution comprises the following steps: redissolving the protein powder to a concentration of 0.005 to 0.015 g / mL to obtain a protein solution; mixing curcumin and the protein solution to obtain a protein mixed solution;

[0080] The protein powder is the protein powder described above;

[0081] The mass volume ratio of the curcumin and the protein solution is 0.005-0.015 g:1 mL, preferably 0.008-0.012 g:1 mL, and more preferably 0.01 g:1 mL.

[0082] In the present invention, the concentration of the protein powder after redissolution is preferably 0.008 to 0.012 g / mL, and more preferably 0.01 g / mL.

[0083] In the present invention, the sodium alginate solution uses water as solvent and includes the following components with final concentrations:

[0084] Sodium alginate 0.015-0.035 g / mL, preferably 0.02-0.03 g / mL, more preferably 0.025 g / mL;

[0085] Polyvinyl alcohol 0.01-0.03 g / mL, preferably 0.015-0.025 g / mL, more preferably 0.02 g / mL;

[0086] The content of glycerol is 0.005 to 0.015 g / mL, preferably 0.008 to 0.012 g / mL, and more preferably 0.01 g / mL.

[0087] In the present invention, the chitosan solution uses water as solvent and includes the following components with final concentrations:

[0088] Chitosan 0.005-0.015 g / mL, preferably 0.008-0.012 g / mL, more preferably 0.01 g / mL;

[0089] CaCl2 0.005-0.025 g / mL, preferably 0.01-0.02 g / mL; more preferably 0.015 g / mL;

[0090] The acetic acid concentration is 0.005 to 0.015 mL / mL, preferably 0.008 to 0.012 mL / mL, and more preferably 0.01 mL / mL.

[0091] In the present invention, the method for mixing curcumin and protein solution is a pH-driven method, and the specific steps are: adjusting the pH of the protein solution to 12 using a 2mol / mL NaOH solution, adding curcumin, and stirring to obtain a mixed solution; and adjusting the pH of the mixed solution to 7 using a 2mol / mL HCl solution to obtain a protein mixed solution.

[0092] The present invention also provides a method for preparing the hydrogel wound dressing, comprising the following steps:

[0093] 1) mixing the protein mixed solution and the sodium alginate solution to obtain a mixed solution;

[0094] 2) pouring the mixed solution into a mold to obtain a mold containing the mixed solution; immersing the mold containing the mixed solution in a chitosan solution to obtain a formed hydrogel;

[0095] 3) demolding and rinsing the formed hydrogel to obtain a hydrogel wound dressing;

[0096] In step 2), the soaking time is 20 to 40 minutes, preferably 25 to 35 minutes, and more preferably 30 minutes.

[0097] The technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0098] Example 1

[0099] Preparation of giant salamander collagen freeze-dried powder:

[0100] a. 100 g of giant salamander skin was soaked in 0.1 mol / mLNaCl solution for 24 h and washed with distilled water to obtain the initial treatment of giant salamander skin;

[0101] b. The initially treated giant salamander skin and isopropanol (purity 99%) were mixed at a mass volume ratio of 1g:10mL for 24h, and washed with distilled water to obtain the treated giant salamander skin;

[0102] c. The treated giant salamander skin was crushed using a beater, the vacuum degree was set to 2.5 kPa, precooled at -70 ℃ for 2 h, and then low-temperature dried at 20 ℃ for 42 h to obtain a giant salamander skin homogenate powder;

[0103] d. Mix the giant salamander skin homogenate powder and the mixed solvent (anhydrous oxalic acid and choline chloride mixed at a molar mass ratio of 1:1 at 80°C) in a mass ratio of 1:20, heat in a water bath at 60°C for 5 hours, dialyze for 48 hours, centrifuge at 12000rpm for 10 minutes, and take the supernatant; evacuate to 0kPa, precool the supernatant at -80°C for 2 hours, and then low-temperature dry at 20°C for 48 hours to obtain giant salamander collagen freeze-dried powder.

[0104] Preparation of natural deep eutectic solvent: D-xylose and choline chloride were mixed at a molar mass ratio of 1:1 at a speed of 20 rpm and a temperature of 90° C. for 3 h to obtain a natural deep eutectic solvent.

[0105] Preparation of protein powder:

[0106] (1) 1 g of giant salamander collagen freeze-dried powder, 50 g of natural low eutectic solvent and 10 mL of water were mixed, and ultrasonicated in a water bath at a power of 1250 W and a temperature of 75° C. for 70 min to obtain a collagen glycosylation product;

[0107] (2) The collagen glycosylation product was dialyzed for 42 hours, wherein the molecular weight of the dialysis bag was 300Da; the evacuation was performed to 0 kPa, the dialyzed material was precooled at -80°C for 2 hours, and then low-temperature dried at 20°C for 48 hours to obtain protein powder.

[0108] Preparation of a hydrogel wound dressing:

[0109] 1) Dissolving 1 g of protein powder to a concentration of 0.01 g / mL to obtain a protein solution; adjusting the pH of 100 mL of the protein solution to 12 using a 2 mol / mL NaOH solution, adding 1 g of curcumin, and stirring to obtain a mixed solution; adjusting the pH of the mixed solution to 7 using a 2 mol / mL HCl solution to obtain a protein mixed solution;

[0110] 2) mixing 100 mL of the protein mixed solution and 200 mL of a sodium alginate solution (using water as a solvent, comprising the following components at the final concentrations: 0.025 g / mL sodium alginate, 0.02 g / mL polyvinyl alcohol, and 0.01 g / mL glycerol) to obtain a mixed solution;

[0111] 3) pouring the mixed solution into a mold to obtain a mold containing the mixed solution; immersing the mold containing the mixed solution in 500 mL of a chitosan solution (using water as a solvent, comprising the following components at the final concentrations: 0.01 g / mL chitosan, 0.015 g / mL CaCl2, and 0.01 mL / mL acetic acid) to obtain a formed hydrogel;

[0112] 4) demolding the formed hydrogel and rinsing it with distilled water to obtain a hydrogel wound dressing.

[0113] Example 2

[0114] Preparation of giant salamander collagen freeze-dried powder and natural deep eutectic solvent is the same as in Example 1;

[0115] Preparation of protein powder:

[0116] (1) 1 g of giant salamander collagen freeze-dried powder, 55 g of natural low eutectic solvent and 5 mL of water were mixed, and ultrasonicated in a water bath at a power of 1500 W and a temperature of 90° C. for 50 min to obtain a collagen glycosylation product;

[0117] (2) The collagen glycosylation product was dialyzed for 48 hours, wherein the molecular weight of the dialysis bag was 250Da; the vacuum was evacuated to 2.5 kPa, the dialyzed material was precooled at -70°C for 3 hours, and then low-temperature dried at 30°C for 56 hours to obtain protein powder.

[0118] Preparation of a hydrogel wound dressing:

[0119] 1) Dissolve 0.015 g of protein powder to a concentration of 0.015 g / mL to obtain a protein solution; adjust the pH of 100 mL of the protein solution to 12 using a 2 mol / mL NaOH solution, add 1.5 g of curcumin, and stir to obtain a mixed solution; adjust the pH of the mixed solution to 7 using a 2 mol / mL HCl solution to obtain a protein mixed solution;

[0120] 2) mixing 100 mL of the protein mixed solution and 150 mL of a sodium alginate solution (using water as a solvent, comprising the following components at the final concentrations: 0.015 g / mL sodium alginate, 0.01 g / mL polyvinyl alcohol, and 0.015 g / mL glycerol) to obtain a mixed solution;

[0121] 3) pouring the mixed solution into a mold to obtain a mold containing the mixed solution; immersing the mold containing the mixed solution in 600 mL of chitosan solution (using water as a solvent, including the following components with final concentrations: 0.015 g / mL chitosan, 0.005 g / mL CaCl2, and 0.015 mL / mL acetic acid) to obtain a formed hydrogel;

[0122] 4) demolding the formed hydrogel and rinsing it with distilled water to obtain a hydrogel wound dressing.

[0123] Example 3

[0124] Preparation of giant salamander collagen freeze-dried powder and natural deep eutectic solvent is the same as in Example 1;

[0125] Preparation of protein powder:

[0126] (1) 1 g of giant salamander collagen freeze-dried powder, 45 g of natural low eutectic solvent and 15 mL of water were mixed, and ultrasonicated in a water bath at a power of 1100 W and a temperature of 60° C. for 90 min to obtain a collagen glycosylation product;

[0127] (2) The collagen glycosylation product was dialyzed for 36 hours, wherein the molecular weight of the dialysis bag was 350Da; the vacuum was drawn to 5 kPa, and the dialyzed material was precooled at -90°C for 1.5 hours, and then low-temperature dried at 15°C for 40 hours to obtain protein powder.

[0128] Preparation of a hydrogel wound dressing:

[0129] 1) Dissolving 0.005 g of protein powder to a concentration of 0.005 g / mL to obtain a protein solution; adjusting the pH of 100 mL of the protein solution to 12 using a 2 mol / mL NaOH solution, adding 0.05 g of curcumin, and stirring to obtain a mixed solution; adjusting the pH of the mixed solution to 7 using a 2 mol / mL HCl solution to obtain a protein mixed solution;

[0130] 2) mixing 100 mL of the protein mixed solution and 300 mL of a sodium alginate solution (using water as a solvent, comprising the following components at the final concentrations: 0.035 g / mL sodium alginate, 0.03 g / mL polyvinyl alcohol, and 0.005 g / mL glycerol) to obtain a mixed solution;

[0131] 3) pouring the mixed solution into a mold to obtain a mold containing the mixed solution; immersing the mold containing the mixed solution in 400 mL of a chitosan solution (using water as a solvent, comprising the following components at the final concentrations: 0.005 g / mL chitosan, 0.025 g / mL CaCl2, and 0.005 mL / mL acetic acid) to obtain a formed hydrogel;

[0132] 4) demolding the formed hydrogel and rinsing it with distilled water to obtain a hydrogel wound dressing.

[0133] Comparative Example 1

[0134] Preparation of giant salamander collagen freeze-dried powder is the same as in Example 1;

[0135] A hydrogel wound dressing:

[0136] 1) re-dissolving 1 g of giant salamander collagen freeze-dried powder to a concentration of 0.01 g / mL to obtain a giant salamander collagen solution;

[0137] 2) Mixing 100 mL of giant salamander collagen solution and 200 mL of sodium alginate solution (same as in Example 1) to obtain a mixed solution;

[0138] 3) pouring the mixed solution into a mold to obtain a mold containing the mixed solution; soaking the mold containing the mixed solution in 500 mL of chitosan solution (same as in Example 1) for 30 min to obtain a formed hydrogel;

[0139] 4) demolding the formed hydrogel and rinsing it with distilled water to obtain a hydrogel wound dressing.

[0140] Comparative Example 2

[0141] Preparation of giant salamander collagen freeze-dried powder and protein powder is the same as in Example 1;

[0142] A hydrogel wound dressing:

[0143] 1) Dissolve 1 g of protein powder to a concentration of 0.01 g / mL to obtain a protein solution;

[0144] 2) Mixing 100 mL of the protein solution and 200 mL of the sodium alginate solution (same as in Example 1) to obtain a mixed solution;

[0145] 3) pouring the mixed solution into a mold to obtain a mold containing the mixed solution; soaking the mold containing the mixed solution in 500 mL of chitosan solution for 30 minutes to obtain a formed hydrogel;

[0146] 4) demolding the formed hydrogel and rinsing it with distilled water to obtain a hydrogel wound dressing.

[0147] Comparative Example 3

[0148] Preparation of giant salamander collagen freeze-dried powder is the same as in Example 1;

[0149] A hydrogel wound dressing:

[0150] 1) Dissolving 1 g of giant salamander collagen freeze-dried powder to a concentration of 0.01 g / mL to obtain a giant salamander collagen solution; adjusting the pH of 100 mL of the giant salamander collagen solution to 12 using a 2 mol / mL NaOH solution, adding 1 g of curcumin, and stirring to obtain a mixed solution; adjusting the pH of the mixed solution to 7 using a 2 mol / mL HCl solution to obtain a protein mixed solution;

[0151] 2) Mixing 100 mL of the protein mixed solution and 200 mL of the sodium alginate solution (same as in Example 1) to obtain a mixed solution;

[0152] 3) pouring the mixed solution into a mold to obtain a mold containing the mixed solution; soaking the mold containing the mixed solution in 500 mL of chitosan solution (same as in Example 1) for 30 min to obtain a formed hydrogel;

[0153] 4) demolding the formed hydrogel and rinsing it with distilled water to obtain a hydrogel wound dressing.

[0154] Comparative Example 4

[0155] Curcumin powder.

[0156] Experimental Example 1 Thermal Stability

[0157] The thermal stability of the hydrogel wound dressings described in Example 1 and Comparative Examples 1 to 3 was tested by infrared spectra analysis and differential scanning calorimetry (DSC) (reference: Novel chitosan-based hydrogels as promising wound dressing materials with advanced properties.), and Comparative Example 4 was used as a control. The results are shown in FIG. Figures 1-2 shown.

[0158] from Figure 1 It can be seen that the characteristic peak of curcumin disappears and is completely wrapped;

[0159] from Figure 2 It can be seen that the thermal denaturation temperature of protein powder is higher than that of lyophilized collagen powder in the range of 90-130°C; compared with comparative examples 1 and 2, the decomposition stages of comparative examples 3 and example 1 are delayed, and in addition, the weight loss rate of example 1 is the lowest and the residual mass is the highest. Therefore, the hydrogel wound dressing described in example 1 is more stable than the hydrogel wound dressings described in comparative examples 1-3.

[0160] Experimental Example 2

[0161] Two full-thickness wounds with a diameter of 10 mm were removed from the backs of 18 SD rats (sourced from Jiangsu Qingyuanyi Biotechnology Co., Ltd.) using a puncher to obtain a rat model with full-thickness loss; the rat models were evenly divided into 6 groups, namely, the embodiment group, the comparative example group 1, the comparative example group 2, the comparative example group 3, the negative control group, and the positive control group; the rats in the embodiment group were smeared with the hydrogel wound dressing of embodiment 1, and the hydrogel wound dressing was changed every two days, and 1 g was applied each time; the comparative examples 1 to 3 groups were smeared with the hydrogel wound dressings described in comparative examples 1 to 3 respectively; the positive control was smeared with medical hydrogel dressings (Hais Hainuo hydrocolloid dressing type artificial skin medical sterile mole wound wound), and the smearing method was the same as that of the embodiment group, and the rats in the negative control group were bandaged with gauze; the wound healing degree of the rats' wound skin was observed on the 8th, 12th, and 16th days after administration, and the relative area of ​​the wound was calculated. The results are as follows Figure 3 As shown; and the wound skin was subjected to immunofluorescence staining analysis of inflammatory factors (IL-6, TNF-α), CD31, α-SMA immunofluorescence staining, HE staining, and Masson staining. The results are shown Figures 3 to 6 As shown;

[0162] The calculation formula for the relative wound area is as follows:

[0163]

[0164] from Figures 3 to 6 It can be seen that the hydrogel wound dressing described in Example 1 has a high wound healing rate compared with the hydrogel wound dressings described in Comparative Examples 1 to 3 and the negative control and the positive control, and can effectively reduce the level of inflammatory factors, promote the formation of new capillaries and granulation tissue, and promote epidermal tissue remodeling, thereby accelerating wound healing.

[0165] Experimental Example 3

[0166] Escherichia coli and Staphylococcus aureus were spread on a plate, and a hole with a diameter of 10 mm was punched in the middle of the plate using a hole puncher. The hydrogel wound dressings described in Example 1 and Comparative Examples 1 to 3 were placed in the hole, and PBS buffer was used as a blank control group. The cells were cultured for 12 hours, and the size of the inhibition ring was observed. The results were as follows: Figure 7 shown.

[0167] from Figure 7 It can be seen that among the hydrogel wound dressings described in Example 1 and Comparative Examples 1 to 3, the antibacterial effect of Example 1 is more significant.

[0168] It can be seen from the above embodiments that the present invention provides a protein powder and a preparation method thereof, a hydrogel wound dressing and a preparation method thereof. The hydrogel wound dressing of the present invention has good antibacterial and anti-inflammatory properties, and has the functions of "wound surface monitoring" and "drug controlled release", which can effectively improve the healing state of the wound.

[0169] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. A protein powder, characterized in that The invention is prepared from the following components: giant salamander collagen freeze-dried powder, natural low eutectic solvent and water; The mass ratio of giant salamander collagen freeze-dried powder to natural low eutectic solvent is 1:45-55; The mass volume ratio of giant salamander collagen freeze-dried powder and water is 1g:5-15mL.

2. The protein powder according to claim 1, characterized in that The natural deep eutectic solvent comprises D-xylose and choline chloride; The molar mass ratio of D-xylose to choline chloride is 0.5-1.5:0.5-1.

5.

3. The method for preparing the protein powder according to claim 1 or 2, characterized in that, The steps include: (1) mixing giant salamander collagen freeze-dried powder, a natural low eutectic solvent and water, and performing ultrasonic water bath to obtain a collagen glycosylation product; (2) The collagen glycosylation product is dialyzed and dried to obtain protein powder.

4. The preparation method according to claim 3, characterized in that: In step (1), the temperature of the ultrasonic water bath is 60-90° C., the power of the ultrasonic water bath is 1000-1500 W, and the time of the ultrasonic water bath is 50-90 min.

5. The preparation method according to claim 4, characterized in that: In step (2), the dialysis time is 36 to 48 hours; the molecular weight of the dialysis bag used for dialysis is 250 to 350 Da; The drying is vacuum freeze drying; the vacuum freeze drying includes precooling and low temperature drying; the precooling temperature is -70 to -90°C; the precooling time is 1 to 3 hours; the low temperature drying temperature is 10 to 30°C; the low temperature drying time is 40 to 56 hours; The vacuum degree of the drying is 0-5 kPa.

6. A hydrogel wound dressing, characterized in that: The method comprises the following components in parts by volume: 0.5 to 1.5 parts of a protein mixed solution, 1 to 3 parts of a sodium alginate solution and 3 to 7 parts of a chitosan solution; The method for preparing the protein mixed solution comprises the following steps: redissolving the protein powder to a concentration of 0.005 to 0.015 g / mL to obtain a protein solution; mixing curcumin and the protein solution to obtain a protein mixed solution; The protein powder is the protein powder according to claim 1 or 2; The mass volume ratio of the curcumin and protein solution is 0.005-0.015 g:1 mL.

7. The hydrogel wound dressing according to claim 6, characterized in that The sodium alginate solution uses water as a solvent and includes the following components with final concentrations: 0.015-0.035 g / mL sodium alginate, 0.01-0.03 g / mL polyvinyl alcohol and 0.005-0.015 g / mL glycerol.

8. The hydrogel wound dressing according to claim 7, characterized in that The chitosan solution uses water as solvent and includes the following components with final concentrations: 0.005-0.015 g / mL chitosan, 0.005-0.025 g / mL CaCl2 and 0.005-0.015 mL / mL acetic acid.

9. The hydrogel wound dressing according to claim 8, characterized in that The method for mixing the curcumin and the protein solution is a pH driven method.

10. The method for preparing the hydrogel wound dressing according to any one of claims 6 to 9, characterized in that: The steps include: 1) mixing the protein mixed solution and the sodium alginate solution to obtain a mixed solution; 2) pouring the mixed solution into a mold to obtain a mold containing the mixed solution; immersing the mold containing the mixed solution in a chitosan solution to obtain a formed hydrogel; 3) demolding and rinsing the formed hydrogel to obtain a hydrogel wound dressing; In step 2), the soaking time is 20 to 40 minutes.

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