Bletilla striata polysaccharide gel patch for wound repair and preparation method of bletilla striata polysaccharide gel patch

Abstract

The invention discloses a bletilla striata polysaccharide gel patch for wound repair and a preparation method of the bletilla striata polysaccharide gel patch, and relates to the field of high polymer materials. When the bletilla striata polysaccharide gel patch for wound repair is prepared, graphene oxide sequentially reacts with 3-methylaminopropylamine and 4-(N-maleimidomethyl) cyclohexane-1-carboxylic acid succinimide ester, and modified graphene oxide is prepared; the preparation method comprises the following steps: extracting bletilla striata polysaccharide from bletilla striata, and reacting the bletilla striata polysaccharide with maleic anhydride and N-methyldopamine in sequence to prepare modified bletilla striata polysaccharide; and reacting the modified bletilla striata polysaccharide, the modified graphene oxide and 1, 5-dibromopentane, mixing with deionized water, coating, and preparing a film, so as to obtain the bletilla striata polysaccharide gel patch for wound repair. The bletilla striata polysaccharide gel patch for wound repair, prepared by the invention, has antibacterial, high-adhesion and antioxidant capacities.

Description

Technical Field

[0001] The present invention relates to the field of polymer materials, and specifically to a Bletilla striata polysaccharide gel patch for wound repair and a preparation method thereof. Background Art

[0002] A gel is a three-dimensional network polymer that forms a porous polymer network. This special structure endows the gel material with a series of unique physical and chemical properties, such as viscoelasticity, water absorption and retention, and high biocompatibility. These characteristics have led to the rapid development of wound dressings based on hydrogel materials, and various wound dressings for different wound types have been developed in the past few decades.

[0003] When the skin is traumatized, the wound is very fragile, and bacteria may invade the wound and cause infection, which may lead to symptoms such as wound swelling, pain, and suppuration, bringing negative impacts to human health. Therefore, this application introduces a Bletilla striata polysaccharide gel patch for wound repair with antibacterial effects and a preparation method thereof. Summary of the Invention

[0004] The purpose of the present invention is to provide a Bletilla striata polysaccharide gel patch for wound repair and a preparation method thereof to solve the problems existing in the prior art.

[0005] A Bletilla striata polysaccharide gel patch for wound repair is prepared by reacting modified Bletilla striata polysaccharide, modified graphene oxide with 1,5-dibromopentane, mixing with deionized water, coating, and film-forming.

[0006] The modified Bletilla striata polysaccharide is prepared by extracting Bletilla striata polysaccharide from Bletilla striata and then reacting Bletilla striata polysaccharide with maleic anhydride and N-methyldopamine in sequence.

[0007] The modified graphene oxide is prepared by reacting graphene oxide with 3-methylaminopropylamine and 4-(N-maleimidomethyl)cyclohexane-1-carboxylic acid N-hydroxysuccinimide ester in sequence.

[0008] A preparation method of a Bletilla striata polysaccharide gel patch for wound repair mainly includes the following preparation steps:

[0009] (1) Mix pre-modified graphene oxide, 4-(N-maleimidomethyl)cyclohexane-1-carboxylic acid N-hydroxysuccinimide ester and chloroform according to a mass ratio of 1:0.18 - 0.22:20 - 30, stir at 45 - 55 °C, 200 - 300 r / min under argon protection for 4.5 - 5.5 h, filter, wash with chloroform 3 - 5 times, and vacuum dry at -10 - 0 °C for 22 - 26 h to obtain modified graphene oxide.

[0010] (2) Mix the crude extract of Bletilla striata, deionized water, and Sevag reagent at a mass ratio of 1:3 - 5:1, stir at 200 - 300 r / min for 30 - 50 min, filter, take the filtrate, and vacuum dry the filtrate at -10 - 0 °C for 46 - 50 h to obtain Bletilla striata polysaccharide;

[0011] (3) Mix the pre-modified Bletilla striata polysaccharide, N-methyldopamine, and deionized water at a mass ratio of 1:0.18 - 0.22:14 - 16, stir at 45 - 55 °C, 200 - 300 r / min under argon protection for 7.5 - 8.5 h, filter, wash with deionized water 3 - 5 times, and vacuum dry at -10 - 0 °C for 22 - 26 h to obtain the modified Bletilla striata polysaccharide;

[0012] (4) Stir the gel mixture at 55 - 65 °C, 200 - 300 r / min for 20 - 40 min, filter, wash with deionized water 3 - 5 times, vacuum dry at -10 - 0 °C for 22 - 26 h, add deionized water, stir at 200 - 300 r / min for 22 - 24 h, uniformly coat on a polytetrafluoroethylene plate, place in a calcium chloride solution, let stand for 22 - 26 h, take out the membrane, and then let stand in deionized water for 10 - 14 h to obtain the gel patch.

[0013] As an optimization, the pre-modified graphene oxide in step (1) is prepared by mixing graphene oxide and ethanol at a mass ratio of 1:12 - 16, ultrasonically treating for 15 - 25 min, adding 3-methylaminopropylamine which is 0.18 - 0.22 times the mass of graphene oxide, continuing to ultrasonically treat for 15 - 25 min, heating to 75 - 85 °C, stirring at 200 - 300 r / min for 5.5 - 6.5 h, filtering, washing with ethanol 3 - 5 times, and vacuum drying at 75 - 85 °C for 22 - 26 h.

[0014] As an optimization, the graphene oxide is 2000-mesh graphene oxide.

[0015] As an optimization, the crude extract of Bletilla striata in step (2) is prepared by washing the dried slices of Bletilla striata with deionized water 3 - 5 times, placing in an oven at 20 - 30 °C, drying for 22 - 26 h, pulverizing to 70 - 90 mesh to obtain Bletilla striata powder; mixing the Bletilla striata powder and deionized water at a mass ratio of 1:9 - 11, stirring at 200 - 300 r / min for 22 - 26 h, filtering, taking the filtrate, vacuum drying at -10 - 0 °C for 22 - 26 h to obtain a concentrated solution, mixing the concentrated solution and ethanol at a mass ratio of 1:8 - 10, stirring at 2 - 6 °C, 200 - 300 r / min for 22 - 26 h, centrifuging at 7000 - 9000 r / min for 2 - 3 min, taking the precipitate, and vacuum drying at -10 - 0 °C for 22 - 26 h.

[0016] As an optimization, the Sevag reagent described in step (2) is prepared by uniformly mixing n-butanol and chloroform at a volume ratio of 1:4.

[0017] As an optimization, the pre-modified Bletilla striata polysaccharide described in step (3) is prepared by heating maleic anhydride to 60 - 64 °C, adding Bletilla striata polysaccharide at 0.15 - 0.17 times the mass of maleic anhydride, stirring at 105 - 115 °C and 200 - 300 r / min for 3.5 - 4.5 h, washing 3 - 5 times with deionized water, and vacuum drying at 105 - 115 °C for 22 - 26 h.

[0018] As an optimization, the gel mixture described in step (4) is prepared by mixing the modified Bletilla striata polysaccharide, modified graphene oxide, 1,5-dibromopentane, and ethanol at a mass ratio of 4 - 6:1:0.18 - 0.22:8 - 10.

[0019] As an optimization, the specific addition amount of the deionized water described in step (4) is 18.5 - 19.5 times the mass of the modified Bletilla striata polysaccharide.

[0020] As an optimization, the calcium chloride solution described in step (4) is a calcium chloride solution with a mass percentage of 2.4 - 2.6%.

[0021] Compared with the prior art, the beneficial effects achieved by the present invention are as follows:

[0022] When preparing the Bletilla striata polysaccharide gel patch for wound repair, the present invention reacts graphene oxide with 3-methylaminopropylamine and 4-(N-maleimidomethyl)cyclohexane-1-carboxylic acid N-hydroxysuccinimide ester in sequence to obtain modified graphene oxide; extracts Bletilla striata polysaccharide from Bletilla striata, and then reacts Bletilla striata polysaccharide with maleic anhydride and N-methyldopamine in sequence to obtain modified Bletilla striata polysaccharide; reacts the modified Bletilla striata polysaccharide, modified graphene oxide with 1,5-dibromopentane, mixes with deionized water, coats, and makes a film to obtain the Bletilla striata polysaccharide gel patch for wound repair.

[0023] First, react graphene oxide with 3-methylaminopropylamine and 4-(N-maleimidomethyl)cyclohexane-1-carboxylic acid N-hydroxysuccinimide ester in sequence to obtain modified graphene oxide; introduce succinimide onto graphene oxide, and succinimide can covalently crosslink with the amine groups on the tissue surface. By reacting the succinimide ester with the amine groups on the tissue surface, stable chemical bonds are formed on the tissue surface, enhancing the adhesion strength and long-term stability of the material.

[0024] Second, extract Bletilla striata polysaccharide from Bletilla striata, and then react Bletilla striata polysaccharide with maleic anhydride and N-methyldopamine in sequence to obtain modified Bletilla striata polysaccharide; dopamine is introduced onto the surface of Bletilla striata polysaccharide, and dopamine can scavenge free radicals, playing an antioxidant effect, and further playing an anti-inflammatory effect at the wound site.

[0025] Finally, react the modified Bletilla striata polysaccharide, modified graphene oxide with 1,5-dibromopentane, mix with deionized water, coat, and form a film to obtain a Bletilla striata polysaccharide gel patch for wound repair; utilize the reaction between bromine group and tertiary amine to generate quaternary ammonium salt. The quaternary ammonium salt has hydrophilic and hydrophobic groups. The hydrophilic group enables it to dissolve in water, and the hydrophobic group can interact with the lipids in the bacterial cell membrane. When the quaternary ammonium salt contacts bacteria, it can destroy the envelope structure of the virus, expose the important components of the bacteria, and then lose their activity, achieving the antibacterial effect. Detailed implementation mode

[0026] Next, in combination with the embodiments of the present invention, the technical solutions in the embodiments of the present invention will be clearly and completely described. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative work shall fall within the protection scope of the present invention.

[0027] Example 1:

[0028] A preparation method of a Bletilla striata polysaccharide gel patch for wound repair mainly includes the following preparation steps:

[0029] (1) Mix graphene oxide and ethanol at a mass ratio of 1:12, ultrasonicate for 15 min, add 3-methylaminopropylamine at 0.18 times the mass of graphene oxide, continue ultrasonication for 15 min, heat up to 75 °C, stir at 200 r / min for 5.5 h, filter, wash with ethanol 3 times, and vacuum dry at 75 °C for 22 h to obtain pre-modified graphene oxide; mix the pre-modified graphene oxide, 4-(N-maleimidomethyl)cyclohexane-1-carboxylic acid succinimide ester and chloroform at a mass ratio of 1:0.18:20, stir at 45 °C, 200 r / min under argon protection for 4.5 h, filter, wash with chloroform 3 times, and vacuum dry at -10 °C for 22 h to obtain modified graphene oxide;

[0030] (2) Wash the dried Bletilla striata slices with deionized water three times, place them in an oven at 20 °C, dry for 22 h, crush to 70 mesh to obtain Bletilla striata powder; mix the Bletilla striata powder and deionized water at a mass ratio of 1:9, stir at 200 r / min for 22 h, filter, take the filtrate, vacuum dry at -10 °C for 22 h to obtain a concentrated solution, mix the concentrated solution and ethanol at a mass ratio of 1:8, stir at 2 °C and 200 r / min for 22 h, centrifuge at 7000 r / min for 2 min, take the precipitate, vacuum dry at -10 °C for 22 h to obtain the crude Bletilla striata extract; mix n-butanol and chloroform at a volume ratio of 1:4 to obtain Sevag reagent; mix the crude Bletilla striata extract, deionized water, and Sevag reagent at a mass ratio of 1:3:1, stir at 200 r / min for 30 min, filter, take the filtrate, and vacuum dry the filtrate at -10 °C for 46 h to obtain Bletilla striata polysaccharide;

[0031] (3) Heat maleic anhydride to 60 °C, add Bletilla striata polysaccharide which is 0.15 times the mass of maleic anhydride, stir at 105 °C and 200 r / min for 3.5 h, wash with deionized water three times, and vacuum dry at 105 °C for 22 h to obtain pre-modified Bletilla striata polysaccharide; mix the pre-modified Bletilla striata polysaccharide, N-methyl dopamine, and deionized water at a mass ratio of 1:0.18:14, stir at 45 °C, 200 r / min under argon protection for 7.5 h, filter, wash with deionized water three times, and vacuum dry at -10 °C for 22 h to obtain modified Bletilla striata polysaccharide;

[0032] (4) Mix the modified Bletilla striata polysaccharide, modified graphene oxide, 1,5-dibromopentane, and ethanol at a mass ratio of 4:1:0.18:8, stir at 55 °C and 200 r / min for 20 min, filter, wash with deionized water three times, vacuum dry at -10 °C for 22 h, add deionized water which is 18.5 times the mass of the modified Bletilla striata polysaccharide, stir at 200 r / min for 22 h, evenly coat on a polytetrafluoroethylene plate, place in a calcium chloride solution with a mass percentage of 2.4%, let stand for 22 h, take out the membrane, and then let stand in deionized water for 10 h to obtain the gel patch.

[0033] Example 2:

[0034] A preparation method of a Bletilla striata polysaccharide gel patch for wound repair mainly includes the following preparation steps:

[0035] (1) Graphene oxide and ethanol were mixed at a mass ratio of 1:14, ultrasonicated for 20 min, 3-(methylamino)propylamine with 0.2 times the mass of graphene oxide was added, and ultrasonicated for another 20 min. Then the temperature was raised to 80 °C, and stirred at 250 r / min for 6 h. After filtration, it was washed 4 times with ethanol and vacuum dried at 80 °C for 24 h to obtain pre-modified graphene oxide; the pre-modified graphene oxide, 4-(N-maleimidomethyl)cyclohexane-1-carboxylic acid N-hydroxysuccinimide ester and chloroform were mixed at a mass ratio of 1:0.2:25, and stirred at 50 °C, 250 r / min under argon protection for 5 h. After filtration, it was washed 4 times with chloroform and vacuum dried at -5 °C for 24 h to obtain modified graphene oxide;

[0036] (2) The dried slices of Bletilla striata were washed 4 times with deionized water, placed in an oven at 25 °C and dried for 24 h, then crushed to 80 mesh to obtain Bletilla striata powder; the Bletilla striata powder and deionized water were mixed at a mass ratio of 1:10, stirred at 250 r / min for 24 h, filtered, and the filtrate was taken and vacuum dried at -5 °C for 24 h to obtain a concentrated solution. The concentrated solution and ethanol were mixed at a mass ratio of 1:9, stirred at 4 °C, 250 r / min for 24 h, centrifuged at 8000 r / min for 2.5 min, the precipitate was taken and vacuum dried at -5 °C for 24 h to obtain the crude extract of Bletilla striata; n-butanol and chloroform were mixed at a volume ratio of 1:4 to prepare Sevag reagent; the crude extract of Bletilla striata, deionized water and Sevag reagent were mixed at a mass ratio of 1:4:1, stirred at 250 r / min for 40 min, filtered, and the filtrate was taken and vacuum dried at -5 °C for 48 h to obtain Bletilla striata polysaccharide;

[0037] (3) Maleic anhydride was heated to 62 °C, and Bletilla striata polysaccharide with 0.16 times the mass of maleic anhydride was added, and stirred at 110 °C, 250 r / min for 4 h, washed 4 times with deionized water and vacuum dried at 110 °C for 24 h to obtain pre-modified Bletilla striata polysaccharide; the pre-modified Bletilla striata polysaccharide, N-methyldopamine and deionized water were mixed at a mass ratio of 1:0.2:15, stirred at 50 °C, 250 r / min under argon protection for 8 h, filtered, washed 4 times with deionized water and vacuum dried at -5 °C for 24 h to obtain modified Bletilla striata polysaccharide;

[0038] (4) The modified Bletilla striata polysaccharide, modified graphene oxide, 1,5-dibromopentane and ethanol were mixed at a mass ratio of 5:1:0.2:9, stirred at 60 °C, 250 r / min for 30 min, filtered, washed 4 times with deionized water and vacuum dried at -5 °C for 24 h. Then, deionized water with 19 times the mass of the modified Bletilla striata polysaccharide was added, stirred at 250 r / min for 23 h, evenly coated on a polytetrafluoroethylene plate, placed in a calcium chloride solution with a mass percentage of 2.5%, left standing for 24 h, the membrane was taken out and then left standing in deionized water for 12 h to obtain the gel patch.

[0039] Example 3:

[0040] A preparation method of a Bletilla striata polysaccharide gel patch for wound repair mainly includes the following preparation steps:

[0041] (1) Mix graphene oxide and ethanol at a mass ratio of 1:16, ultrasonicate for 25 min, add 3-methylaminopropylamine which is 0.22 times the mass of graphene oxide, continue ultrasonication for 25 min, raise the temperature to 85 °C, stir at 300 r / min for 6.5 h, filter, wash with ethanol 5 times, and vacuum dry at 85 °C for 26 h to obtain pre-modified graphene oxide; Mix the pre-modified graphene oxide, 4-(N-maleimidomethyl)cyclohexane-1-carboxylic acid N-hydroxysuccinimide ester and chloroform at a mass ratio of 1:0.22:30, stir at 55 °C, 300 r / min under argon protection for 5.5 h, filter, wash with chloroform 5 times, and vacuum dry at 0 °C for 26 h to obtain modified graphene oxide;

[0042] (2) Wash the dried Bletilla striata slices with deionized water 5 times, place them in an oven at 30 °C, dry for 26 h, and pulverize to 90 mesh to obtain Bletilla striata powder; Mix the Bletilla striata powder and deionized water at a mass ratio of 1:11, stir at 300 r / min for 26 h, filter, take the filtrate, vacuum dry at 0 °C for 26 h to obtain a concentrated solution, mix the concentrated solution and ethanol at a mass ratio of 1:10, stir at 6 °C, 300 r / min for 26 h, centrifuge at 9000 r / min for 3 min, take the precipitate, and vacuum dry at 0 °C for 26 h to obtain a crude Bletilla striata extract; Mix n-butanol and chloroform at a volume ratio of 1:4 to obtain Sevag reagent; Mix the crude Bletilla striata extract, deionized water, and Sevag reagent at a mass ratio of 1:5:1, stir at 300 r / min for 50 min, filter, take the filtrate, and vacuum dry the filtrate at 0 °C for 50 h to obtain Bletilla striata polysaccharide;

[0043] (3) Heat maleic anhydride to 64 °C, add Bletilla striata polysaccharide which is 0.17 times the mass of maleic anhydride, stir at 115 °C, 300 r / min for 4.5 h, wash with deionized water 5 times, and vacuum dry at 115 °C for 26 h to obtain pre-modified Bletilla striata polysaccharide; Mix the pre-modified Bletilla striata polysaccharide, N-methyldopamine and deionized water at a mass ratio of 1:0.22:16, stir at 55 °C, 300 r / min under argon protection for 8.5 h, filter, wash with deionized water 5 times, and vacuum dry at 0 °C for 26 h to obtain modified Bletilla striata polysaccharide;

[0044] (4) Mix the modified Bletilla striata polysaccharide, modified graphene oxide, 1,5-dibromopentane, and ethanol in a mass ratio of 6:1:0.22:10, stir at 65 °C and 300 r / min for 40 min, filter, wash 5 times with deionized water, vacuum dry at 0 °C for 26 h, add deionized water 19.5 times the mass of the modified Bletilla striata polysaccharide, stir at 300 r / min for 24 h, evenly coat on a polytetrafluoroethylene plate, place in a calcium chloride solution with a mass percentage of 2.6%, let stand for 26 h, take out the membrane, and then let stand in deionized water for 14 h to obtain the gel patch.

[0045] Comparative Example 1:

[0046] The preparation method of the Bletilla striata polysaccharide gel patch for wound repair in Comparative Example 1 is only different from that in Example 2 in that graphene oxide is not modified. The remaining steps are the same as those in Example 2.

[0047] Comparative Example 2:

[0048] The preparation method of the Bletilla striata polysaccharide gel patch for wound repair in Comparative Example 2 is only different from that in Example 2 in that Bletilla striata polysaccharide is not modified. The remaining steps are the same as those in Example 2.

[0049] Comparative Example 3:

[0050] The preparation method of the Bletilla striata polysaccharide gel patch for wound repair in Comparative Example 3 is different from that in Example 2 in step (4). Modify step (4) as follows: Mix the modified Bletilla striata polysaccharide and modified graphene oxide in a mass ratio of 5:1, add deionized water 19 times the mass of the modified Bletilla striata polysaccharide, stir at 250 r / min for 23 h, evenly coat on a polytetrafluoroethylene plate, place in a calcium chloride solution with a mass percentage of 2.5%, let stand for 24 h, take out the membrane, and then let stand in deionized water for 12 h to obtain the gel patch. The remaining steps are the same as those in Example 2.

[0051] Test Example 1:

[0052] Adhesion test

[0053] Test method: Refer to ASTM F2256 for the standard 180-degree peel test. First, attach the gel patches prepared in each example and comparative example to the surface of pig skin with a size of 6 cm × 2.5 cm × 0.1 cm, apply a pressure of 1 kPa with a universal testing machine and press continuously for 5 s, let stand for 5 min, use a universal testing machine (2.5 kN force sensor, Zwick / Roell Z2.5), measure the tensile force at a constant peeling speed of 50 mm / min, record the maximum adhesion force (Fmax), and calculate the adhesion tensile strength, where the adhesion tensile strength = 2Fmax / 2.5. The results are shown in Table 1.

[0054] Table 1

[0055]

[0056]

[0057] It can be found from the comparison of the experimental data in Table 1 that the Bletilla striata polysaccharide gel patch for wound repair prepared by the present invention has good adhesiveness.

[0058] It can be found from the comparison of the experimental data of Examples 1, 2, and 3 and Comparative Example 1 in Table 1 that the adhesion tensile strengths of Examples 1, 2, and 3 are large. The difference between Comparative Example 1 and the examples is that succinimide is not introduced onto graphene oxide. Succinimide can covalently crosslink with the amino groups on the tissue surface. By reacting the succinimide ester with the amino groups on the tissue surface, stable chemical bonds can be formed on the tissue surface, enhancing the adhesion strength and long-term stability of the material.

[0059] Test Example 2:

[0060] Antibacterial property test

[0061] The Bletilla striata polysaccharide gel patches for wound repair prepared in each example and comparative example were crushed to 30 mesh, 1 g was taken, and the test was carried out with reference to the standard of GB / T31402-2015. The selected bacterial strains were Staphylococcus aureus and Escherichia coli respectively. The results are shown in Table 2.

[0062] Table 2

[0063]

[0064]

[0065] It can be found from the comparison of the experimental data in Table 2 that the Bletilla striata polysaccharide gel patch for wound repair prepared by the present invention has good antibacterial ability.

[0066] It can be found from the comparison of the experimental data of Examples 1, 2, and 3 and Comparative Example 3 in Table 2 that the bacteriostatic rates of Examples 1, 2, and 3 are high. The difference between Comparative Example 3 and the examples is that the reaction of bromine group with tertiary amine to generate quaternary ammonium salt is not utilized. Quaternary ammonium salt has hydrophilic and hydrophobic groups. The hydrophilic group enables it to dissolve in water, and the hydrophobic group can interact with the lipids in the bacterial cell membrane. When the quaternary ammonium salt contacts the bacteria, it can destroy the envelope structure of the virus, expose the important components of the bacteria, and then lose their activity, achieving the antibacterial effect.

[0067] Test Example 3:

[0068] Antioxidant test:

[0069] Test method: The MTT cell proliferation and cytotoxicity detection kit was used to detect the cell viability after different treatments. After culturing AML-12 in a 96-well plate until the cell density reached 80%, the gels prepared in each example and comparative example were crushed to 100 meshes, and a culture medium with a gel concentration of 500 μg / mL and a hydrogen peroxide concentration of 0.5 mmol / mL was prepared for culturing for 24 h. After the cell culture was completed, the cell viability of each group was detected. The results are shown in Table 3.

[0070] Table 3

[0071]

[0072]

[0073] It can be found from the comparison of the experimental data in Table 3 that the Bletilla striata polysaccharide gel prepared by the present invention for wound repair has good antioxidant ability.

[0074] It can be found from the comparison of the experimental data of Examples 1, 2, 3 and Comparative Example 2 in Table 3 that the expression of CD86 in Examples 1, 2, and 3 is small. The difference between Comparative Example 2 and the examples is that dopamine is not grafted onto the surface of Bletilla striata polysaccharide. Dopamine can scavenge free radicals and play an antioxidant effect.

[0075] The specific embodiments described above further elaborate on the purpose, technical solutions, and beneficial effects of the present invention. It should be understood that the above are only specific embodiments of the present invention and are not used to limit the protection scope of the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A method for preparing a Bletilla striata polysaccharide gel patch for wound repair, characterized in that: The bletilla striata polysaccharide gel patch for wound repair is prepared by reacting modified bletilla striata polysaccharide, modified graphene oxide and 1,5-dibromopentane, mixing with deionized water, coating and forming a film; The modified bletilla striata polysaccharide is prepared by extracting bletilla striata polysaccharide from bletilla striata, and then reacting the bletilla striata polysaccharide with maleic anhydride and N-methyl dopamine in sequence; The modified graphene oxide is prepared by reacting graphene oxide with 3-methylaminopropylamine and 4-(N-maleimidomethyl)cyclohexane-1-carboxylic acid succinimide ester in sequence.

2. A method for preparing a Bletilla striata polysaccharide gel patch for wound repair, characterized in that: The preparation method of the bletilla striata polysaccharide gel patch for wound repair mainly comprises the following preparation steps: (1) Pre-modified graphene oxide, 4-(N-maleimidomethyl)cyclohexane-1-carboxylic acid succinimide ester and chloroform are mixed in a mass ratio of 1:0.18-0.22:20-30, stirred at 45-55°C, 200-300 r / min, under argon protection for 4.5-5.5 hours, filtered, washed with chloroform for 3-5 times, and vacuum dried at -10-0°C for 22-26 hours to obtain modified graphene oxide; (2) Mix the crude extract of Bletilla striata, deionized water and Sevag reagent in a mass ratio of 1:3 to 5:1, stir at 200 to 300 r / min for 30 to 50 min, filter, take the filtrate, and vacuum dry the filtrate at -10 to 0°C for 46 to 50 h to obtain Bletilla striata polysaccharide; (3) pre-modified bletilla striata polysaccharide, N-methyl dopamine and deionized water are mixed in a mass ratio of 1:0.18-0.22:14-16, stirred for 7.5-8.5 h at 45-55° C., 200-300 r / min, under argon protection, filtered, washed with deionized water for 3-5 times, and vacuum dried at -10-0° C. for 22-26 h to obtain modified bletilla striata polysaccharide; (4) The gel mixture is stirred at 55-65°C and 200-300 r / min for 20-40 min, filtered, washed with deionized water for 3-5 times, vacuum dried at -10-0°C for 22-26 h, added with deionized water, stirred at 200-300 r / min for 22-24 h, evenly coated on a polytetrafluoroethylene plate, placed in a calcium chloride solution, allowed to stand for 22-26 h, the membrane removed, and allowed to stand in deionized water for 10-14 h to obtain a gel patch.

3. The method for preparing a Bletilla striata polysaccharide gel patch for wound repair according to claim 2, characterized in that: The pre-modified graphene oxide in step (1) is prepared by mixing graphene oxide and ethanol in a mass ratio of 1:12-16, ultrasonicating for 15-25 minutes, adding 3-methylaminopropylamine in an amount of 0.18-0.22 times the mass of graphene oxide, and continuing ultrasonicating for 15-25 minutes, heating to 75-85°C, stirring at 200-300 r / min for 5.5-6.5 hours, filtering, washing with ethanol for 3-5 times, and vacuum drying at 75-85°C for 22-26 hours.

4. The method for preparing a Bletilla striata polysaccharide gel patch for wound repair according to claim 3, characterized in that: The graphene oxide is 2000 mesh graphene oxide.

5. The method for preparing a Bletilla striata polysaccharide gel patch for wound repair according to claim 2, characterized in that: The crude extract of Bletilla striata in step (2) is prepared by washing dried Bletilla striata slices with deionized water for 3 to 5 times, placing the dried slices in an oven at 20 to 30° C., drying for 22 to 26 hours, and crushing the dried slices to 70 to 90 meshes to obtain Bletilla striata powder; mixing the Bletilla striata powder and deionized water in a mass ratio of 1:9 to 11, stirring at 200 to 300 r / min for 22 to 26 hours, filtering, taking the filtrate, and vacuum drying at -10 to 0° C. for 22 to 26 hours to obtain a concentrated solution; mixing the concentrated solution with ethanol in a mass ratio of 1:8 to 10, stirring at 200 to 300 r / min for 22 to 26 hours at 2 to 6° C., centrifuging at 7000 to 9000 r / min for 2 to 3 minutes, taking the precipitate, and vacuum drying at -10 to 0° C. for 22 to 26 hours to obtain the obtained solution.

6. The method for preparing a Bletilla striata polysaccharide gel patch for wound repair according to claim 2, characterized in that: The Sevag reagent in step (2) is prepared by uniformly mixing n-butanol and chloroform in a volume ratio of 1:

4.

7. The method for preparing a Bletilla striata polysaccharide gel patch for wound repair according to claim 2, characterized in that: The pre-modified Bletilla striata polysaccharide in step (3) is prepared by heating maleic anhydride to 60-64°C, adding Bletilla striata polysaccharide in an amount of 0.15-0.17 times the mass of maleic anhydride, stirring at 105-115°C and 200-300 r / min for 3.5-4.5 hours, washing with deionized water for 3-5 times, and vacuum drying at 105-115°C for 22-26 hours.

8. The method for preparing a Bletilla striata polysaccharide gel patch for wound repair according to claim 2, characterized in that: The gel mixture in step (4) is prepared by mixing modified bletilla striata polysaccharide, modified graphene oxide, 1,5-dibromopentane and ethanol in a mass ratio of 4-6:1:0.18-0.22:8-10.

9. The method for preparing a Bletilla striata polysaccharide gel patch for wound repair according to claim 2, characterized in that: The specific amount of deionized water added in step (4) is 18.5 to 19.5 times the mass of the modified Bletilla striata polysaccharide.

10. The method for preparing a Bletilla striata polysaccharide gel patch for wound repair according to claim 2, characterized in that: The calcium chloride solution in step (4) is a calcium chloride solution with a mass percentage of 2.4-2.6%.

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