CN109350762B

CN109350762B - Medical dressing applied to chronic wound surface and preparation method thereof

Info

Publication number: CN109350762B
Application number: CN201811407593.8A
Authority: CN
Inventors: 朱翠兰; 尚丽霞; 陶秀梅; 陈鹏
Original assignee: Beijing Nuokangda Pharmaceutical Technology Co ltd
Current assignee: Beijing Nuokangda Pharmaceutical Technology Co ltd
Priority date: 2018-11-23
Filing date: 2018-11-23
Publication date: 2021-07-23
Anticipated expiration: 2038-11-23
Also published as: CN109350762A

Abstract

The invention belongs to the field of medical dressings, and relates to a medical dressing applied to chronic wounds, which comprises bacterial cellulose, honey, chitosan and derivatives thereof, wherein the honey and the chitosan are prepared into an antibacterial agent to be compounded on the bacterial cellulose membrane in a physical compounding way, or the honey and/or the chitosan are added into a culture medium in a bacterial cellulose membrane film forming process in a biological compounding process, and then the medical dressing is prepared in a physical compounding way, 3 components have good biocompatibility, the healing of wounds is promoted through mutual synergistic action, the formation of scars and the like is avoided, and the medical dressing has a wide application prospect.

Description

Medical dressing applied to chronic wound surface and preparation method thereof

Technical Field

The invention belongs to the field of medical dressings, and particularly relates to a medical dressing applied to chronic wounds and a preparation method thereof.

Background

The chronic wound refers to a wound which can not heal through a normal healing process under the action of various internal and external factors, enters a pathological inflammatory reaction state to cause the wound to be durable and difficult to heal and can heal by means of external force. Chronic wounds are often complicated with bacterial infections, particularly penicillin-resistant staphylococcus aureus infections, which make wound infections difficult to control, and chronic wound treatment becomes difficult due to factors such as necrotic tissues, poor microcirculation, changes in local growth factors and the like existing on the local part of the wound. In addition to treating primary diseases, control of wound infection and removal of necrotic tissue are important steps in the treatment of chronic wounds. When the antibiotic is applied to the whole body, the medicine concentration at the local part of the wound often cannot reach the effective range for preventing and treating infection, and in addition, the medicine dosage can be increased, and the side effect is increased. The use of a suitable topical antimicrobial dressing can substantially reduce wound infection while promoting overall wound healing.

At present, various dressings loaded with antibacterial properties are developed, and the dressings can be mainly classified into types of fibers, hydrogels, sponges, films and the like according to the forms of carriers. The fiber type antibacterial dressing can prevent bacterial infection and provide gas exchange, but the fiber type antibacterial dressing cannot keep a large amount of liquid and is easy to adhere to a wound to cause pain in dressing change; the hydrogel antibacterial dressing has good combination with the wound surface, easy replacement, good water absorption, but poor mechanical property; the sponge type antibacterial dressing has good absorptivity and water vapor permeability, but allows tissues to grow into pores, so that secondary wound is easily caused during dressing change, and scraps are easily left on the wound surface; the film type antibacterial dressing is transparent and can keep the wound moist, but has poor absorption performance.

On the other hand, in the selection of antibacterial agents, antibiotics tend to induce drug resistance in the body, and the development of novel antibacterial agents has been a focus in recent years. The bacterial cellulose is a cellulose material with a three-dimensional nano-net structure generated by microbial fermentation, and has a plurality of unique properties, such as extremely strong water binding capacity, high polymerization degree and crystallinity, high wet tensile strength, good biocompatibility and the like; the manuka honey is a carbohydrate complex with extremely complex components, contains various antibacterial substances, and has the advantages of promoting wound healing, promoting lymphocyte proliferation, improving immunity, promoting endothelial progenitor cell mobilization, accelerating wound neovascularization and the like; the chitosan and the derivatives thereof have good biocompatibility and in-vivo biodegradability, also have the effects of resisting bacteria, diminishing inflammation, stopping bleeding, reducing wound exudation, promoting wound tissue regeneration, repair, healing and the like, and have the advantages of flexibility, water absorption, air permeability and the like.

Patent application CN 103550817A discloses a bacterial cellulose/chitosan composite sponge dressing, which has the advantages that the effect of promoting wound healing is obviously improved through the composite action of bacterial cellulose and chitosan, but the sponge dressing is also added with an adhesive, a hot melt adhesive or a heat-sensitive adhesive to help the positioning of the dressing, and the existence of the adhesive and other substances has greater potential safety hazard on sensitive skin; for another example, patent application CN 103480027 a discloses a method for preparing a bacterial cellulose-chitosan fiber-based wet dressing, in which bacterial cellulose is dissolved in an organic solvent, and chitosan is compounded onto a bacterial cellulose membrane by electrostatic spinning to form a bacterial cellulose-chitosan composite wet dressing, but the above preparation method has a high cost; patent application CN 103463124A also discloses a bacterial cellulose chitosan composite gel, which is composed of chitosan, bacterial cellulose, an antibacterial agent and a thickening agent, wherein the additionally added antibacterial agent belongs to chemicals and is easy to generate the problem of bacterial drug resistance after long-term use.

Therefore, the researches on compounding and preparing the bacterial cellulose and the chitosan into the medical dressing are very many at present, but some chemical reagents are often added in the preparation process of the medical dressing, so that the application of the medical dressing is limited, and therefore, how to select proper natural raw materials to prepare the medical dressing with excellent antibacterial performance, biological performance and physical and chemical performance is a hotspot of the current researches.

Disclosure of Invention

Based on the defects of the prior art, the bacterial cellulose, the manuka honey, the chitosan and the derivatives thereof are selected as the raw materials of the medical dressing, no chemical auxiliary agent is added, the biocompatibility and the application range are wider, the manuka honey, the chitosan and the derivatives thereof are compounded on the bacterial cellulose membrane in a biological compounding and physical compounding mode in the preparation process, the preparation process is simple, the use of organic reagents is avoided, and the operation convenience and the safety are higher.

In order to achieve the purpose, the invention provides a medical dressing applied to chronic wounds, which comprises bacterial cellulose, honey, chitosan and derivatives thereof.

Further, the medical dressing comprises the following components in parts by weight: 5-10 parts of bacterial cellulose, 0.5-5 parts of honey, 0.5-5 parts of chitosan and derivatives thereof.

Furthermore, the medical dressing comprises the following components in parts by weight: 6-9 parts of bacterial cellulose, 1-3 parts of honey, 1-3 parts of chitosan and derivatives thereof.

Further, the bacterial cellulose is synthesized by one or more microorganisms selected from the group consisting of Acetobacter, Agrobacterium, Rhizobium, and Sarcina.

Further, the honey is manuka honey.

Furthermore, the chitosan and the derivatives thereof are selected from one or more of chitosan, carboxymethyl chitosan and chitosan quaternary ammonium salt.

The invention also provides a preparation method of the medical dressing for the chronic wound, which comprises the following steps:

1) preparation of bacterial cellulose membrane: inoculating the seed liquid into a fermentation culture medium for culture to obtain a bacterial cellulose membrane;

2) purification treatment of bacterial cellulose membrane: cleaning the bacterial cellulose membrane prepared in the step 1) with purified water, then putting the bacterial cellulose membrane into alkali liquor for passivation, and then cleaning the bacterial cellulose membrane with water for injection to obtain a purified bacterial cellulose membrane;

3) physical compounding: adding Mel, chitosan and its derivatives into water for injection, and filtering to obtain antibacterial solution; adding the purified bacterial cellulose membrane obtained in the step 2) into the filtered antibacterial agent solution for physical compounding;

4) drying the composite membrane: and drying the compounded film to obtain the composite membrane.

Further, the preparation method of the medical dressing for the chronic wound comprises the following steps:

1) preparation of bacterial cellulose membrane: inoculating the seed solution into a fermentation culture medium in an inoculation amount of 1-20%, and performing static culture at 28-35 ℃ for 5-14 days to obtain a bacterial cellulose membrane;

2) purification treatment of bacterial cellulose membrane: repeatedly cleaning the bacterial cellulose membrane obtained in the step 1) with purified water until the pH value of the bacterial cellulose membrane is more than or equal to 5.0; then putting the bacterial cellulose membrane into alkali liquor according to the ratio of the membrane volume to the alkali liquor volume of 1:2, heating to 80-110 ℃, stirring for 1-12h, taking out, washing the bacterial cellulose membrane with water for injection until the pH value is 5.0-7.0 and the content of bacterial endotoxin is less than or equal to 0.03EU/ml, and obtaining a purified bacterial cellulose membrane;

3) physical compounding: accurately weighing Manuka honey, chitosan and derivatives thereof, adding into water for injection, and filtering to obtain 0.01% -10% antibacterial agent solution; adding the bacterial cellulose membrane purified in the step 2) into an antibacterial agent solution, and stirring for 1-12 h;

4) drying the composite membrane: drying the membrane obtained in the step 3);

further, the preparation process of the seed liquid in the step 1) comprises the following steps:

firstly, rejuvenating strains: inoculating the freeze-dried thallus into a slant culture medium under aseptic conditions, culturing at 30 ℃ for 36h, and repeating for 2 times; wherein the slant culture medium comprises the following components: 10% of glucose, 1% of yeast powder, 2% of calcium carbonate and 1.8% of agar;

② strain expanding culture: picking strains cultured by the slant into a sterile culture solution, and performing shake culture at 30 ℃ and 150r/min for 24 h; wherein the culture solution comprises 25g/L of glucose, 3g/L of peptone and 5g/L of yeast extract.

Further, the fermentation medium of step 1) is: glucose 25g/L, peptone 3g/L, yeast extract 5g/L, pH 4.0-6.0, and sterilizing at 121 deg.C for 20 min.

Furthermore, the alkali liquor in the step 2) is sodium hydroxide solution, and the concentration is 0.01-2.0 mol/L.

Preferably, the concentration of the alkali liquor is 0.5-1.0 mol/L.

Further, the filtration mode in the step 3) is selected from one of filter paper filtration and membrane filtration.

Further, the stirring manner in the step 3) is selected from one of ultrasonic oscillation, magnetic stirring and electric stirring.

Further, the compounding temperature in the step 3) is 10-80 ℃.

Preferably, the compounding temperature in the step 3) is 30-40 ℃.

Further, the concentration of the antibacterial agent solution prepared in the step 3) is 3-7%.

Further, the drying manner in the step 4) is selected from one of a supercritical drying and molding technique, a vacuum drying and molding technique, a freeze drying and molding technique and a forced air drying and molding technique.

The invention also provides another preparation method of the medical dressing for the chronic wound, which comprises the following steps:

1) biological compounding: inoculating the seed solution into a fermentation culture medium, adding honey and/or chitosan and derivatives thereof into the fermentation culture medium, and then culturing to obtain a composite bacterial cellulose membrane;

2) purification treatment of bacterial cellulose membrane: cleaning the composite bacterial cellulose membrane prepared in the step 1) with purified water, then putting the composite bacterial cellulose membrane into alkali liquor for passivation, and then cleaning the composite bacterial cellulose membrane with water for injection to obtain a purified composite bacterial cellulose membrane;

3) physical compounding: adding Mel and/or chitosan and its derivatives into water for injection, and filtering to obtain antibacterial solution; adding the purified composite bacterial cellulose membrane obtained in the step 2) into the filtered antibacterial agent solution for physical compounding;

1) biological compounding: inoculating the seed solution into a fermentation culture medium at an inoculation amount of 1-20%, adding part of Manuka honey and/or chitosan and derivatives thereof, and standing at 28-35 deg.C for 5-14 days to obtain a composite bacterial cellulose membrane;

2) purification treatment of bacterial cellulose membrane: repeatedly cleaning the composite bacterial cellulose membrane obtained in the step 1) with purified water until the pH value of the composite bacterial cellulose membrane is more than or equal to 5.0; then putting the composite bacterial cellulose membrane into alkali liquor according to the ratio of the membrane volume to the alkali liquor volume of 1:2, heating to 80-110 ℃, stirring for 1-12h, taking out, washing the composite bacterial cellulose membrane with water for injection until the pH value is 5.0-7.0 and the content of bacterial endotoxin is less than or equal to 0.03EU/ml, and obtaining a purified composite bacterial cellulose membrane;

3) physical compounding: adding the rest Manuka honey and/or chitosan and derivatives thereof into water for injection, and filtering to obtain 0.01% -10% antibacterial agent solution; adding the purified composite bacterial cellulose membrane obtained in the step 2) into an antibacterial agent solution, and stirring for 1-12 h;

4) drying the composite membrane: drying the membrane of step 3).

Further, the fermentation medium of step 1) is: glucose 25g/L, peptone 3g/L, yeast extract 5g/L, pH 5.0, and sterilizing at 121 deg.C for 20 min.

Preferably, the concentration of the alkali liquor is 0.5-1.0 mol/L.

Further, the compounding temperature in the step 3) is 10-80 ℃.

Preferably, the compounding temperature in the step 3) is 30-40 ℃.

The invention has the beneficial effects that:

(1) the bacterial cellulose adopted by the medical dressing is of a non-animal source, does not generate immunological rejection reaction, has excellent biocompatibility, does not cause any toxic or allergic side effect when being contacted with living tissues, does not adhere to tissues, reduces scar formation, has super-strong water absorption capacity, and can absorb wound exudate in time and promote wound healing; the bacterial cellulose also contains rich pores, is easy to grow fibroblast and promotes wound tissue repair;

the manuka honey, the chitosan and the derivatives thereof are all natural substances, are non-toxic and harmless, and have good biocompatibility and broad-spectrum antibacterial activity; therefore, the medical dressing prepared from the bacterial cellulose, the manuka honey, the chitosan and the derivatives thereof has good biocompatibility and safety;

the interaction of the three components can promote the wound healing and inhibit the action of microorganisms, thereby more effectively avoiding the formation of scars.

(2) The medical dressing only consists of the bacterial cellulose, the honey, the chitosan and the derivatives thereof, and other chemical reagents are not added additionally, so that the drug resistance of bacteria is avoided, the probability of discomfort of sensitive people to the chemical reagents is reduced, and the safety and the application range of the product are greatly improved.

(3) The preparation process of the medical dressing adopts a mode of physical compounding or combination of biological compounding and physical compounding, the preparation process is safe and simple, the manufacturing cost is low, and the medical dressing is suitable for large-scale production.

(4) The product of the invention is convenient for clinical use, can be well adhered to wounds, has good water permeability, high transparency and good durability, can relieve wound pain and accelerate wound healing, does not cause secondary wound in the dressing change process, and is beneficial to wound repair.

Detailed Description

EXAMPLE 1 medical dressing and preparation thereof

Medical dressing: 6 parts of bacterial cellulose, 2 parts of Manuka honey and 2 parts of chitosan

The preparation method comprises the following steps:

1) preparation of bacterial cellulose membrane: inoculating the seed solution into a fermentation culture medium according to the inoculation amount of 10%, and standing and culturing for 10 days at 30 ℃ to obtain a bacterial cellulose membrane;

2) purification treatment of bacterial cellulose membrane: repeatedly cleaning the bacterial cellulose membrane obtained in the step 1) with purified water until the pH value of the bacterial cellulose membrane is more than or equal to 5.0; then putting the bacterial cellulose membrane into NaOH solution with the concentration of 1.0mol/L according to the ratio of the membrane volume to the alkali liquor volume of 1:2, heating to 100 ℃, stirring for 8h, taking out, washing the bacterial cellulose membrane with water for injection until the pH value is 5.0-7.0 and the content of bacterial endotoxin is less than or equal to 0.03EU/ml, and obtaining the purified bacterial cellulose membrane;

3) physical compounding: accurately weighing Manuka honey and chitosan, adding into water for injection, and filtering with 0.22 μm filter membrane to obtain 8% antibacterial solution; adding the bacterial cellulose membrane purified in the step 2) into an antibacterial agent solution, and magnetically stirring for 8 hours at the compounding temperature of 25 ℃;

4) drying the composite membrane: drying the film in the step 3) by adopting a vacuum drying and forming technology;

5) and 4) drying the product, attaching a forming film PET to two sides, flattening by using a die, cutting to a size of 15.0cm x 15.0cm, carrying out inner wrapping and outer wrapping, conveying to a sterilization mechanism for cobalt-60 ray sterilization, wherein the sterilization dose is 25kGy, sterilizing and packaging to obtain a final product.

Wherein the preparation process of the seed liquid in the step 1) comprises the following steps:

Wherein the freeze-dried thallus is acetobacter xylinum, which is preserved in China general microbiological culture collection center, and the strain number is CGMCC 1.1812.

Wherein the fermentation medium in the step 1) is 25g/L of glucose, 3g/L of peptone and 5g/L of yeast extract, the pH value is 4.0-5.0, and the fermentation medium is sterilized at 121 ℃ for 20 min.

EXAMPLE 2 medical dressings and their preparation

Medical dressing: 7 parts of bacterial cellulose, 2 parts of manuka honey and 1 part of carboxymethyl chitosan

The preparation method comprises the following steps:

1) preparation of bacterial cellulose membrane: inoculating the seed solution into a fermentation culture medium by 8 percent of inoculation amount, and standing and culturing for 10 days at 28 ℃ to obtain a bacterial cellulose membrane;

2) purification treatment of bacterial cellulose membrane: repeatedly cleaning the bacterial cellulose membrane obtained in the step 1) with purified water until the pH value of the bacterial cellulose membrane is more than or equal to 5.0; then putting the bacterial cellulose membrane into NaOH solution with the concentration of 0.5mol/L according to the ratio of the membrane volume to the alkali liquor volume of 1:2, heating to 80 ℃, stirring for 4h, taking out, washing the bacterial cellulose membrane with water for injection until the pH value is 5.0-7.0 and the content of bacterial endotoxin is less than or equal to 0.03EU/ml, and obtaining the purified bacterial cellulose membrane;

3) physical compounding: accurately weighing Manuka honey and carboxymethyl chitosan, adding into water for injection, and filtering with 0.22 μm filter membrane to obtain 10% antibacterial solution; adding the bacterial cellulose membrane purified in the step 2) into an antibacterial agent solution, and magnetically stirring for 6 hours at the compounding temperature of 40 ℃;

4) drying the composite membrane: drying the film in the step 3) by adopting a forced air drying and forming technology;

EXAMPLE 3 medical dressing and preparation thereof

Medical dressing: 7 parts of bacterial cellulose, 1 part of manuka honey and 2 parts of chitosan quaternary ammonium salt

The preparation method comprises the following steps:

1) preparation of bacterial cellulose membrane: inoculating the seed solution into a fermentation culture medium in an inoculation amount of 12%, and standing and culturing at 35 ℃ for 10 days to obtain a bacterial cellulose membrane;

2) purification treatment of bacterial cellulose membrane: repeatedly cleaning the bacterial cellulose membrane obtained in the step 1) with purified water until the pH value of the bacterial cellulose membrane is more than or equal to 5.0; then putting the bacterial cellulose membrane into NaOH solution with the concentration of 0.2mol/L according to the ratio of the membrane volume to the alkali liquor volume of 1:2, heating to 90 ℃, stirring for 6h, taking out, washing the bacterial cellulose membrane with water for injection until the pH value is 5.0-7.0 and the content of bacterial endotoxin is less than or equal to 0.03EU/ml, and obtaining the purified bacterial cellulose membrane;

3) physical compounding: accurately weighing Manuka honey and chitosan quaternary ammonium salt, adding into water for injection, and filtering with 0.22 μm filter membrane to obtain 0.01% antibacterial solution; adding the bacterial cellulose membrane purified in the step 2) into an antibacterial agent solution, and magnetically stirring for 4 hours at the compounding temperature of 60 ℃;

4) drying the composite membrane: drying the membrane in the step 3) by adopting a freeze drying molding technology;

EXAMPLE 4 medical dressing and preparation thereof

The preparation method comprises the following steps:

1) biological compounding: inoculating the seed solution into a fermentation culture medium in an inoculation amount of 20%, adding manuka honey into the fermentation culture medium in an amount of 0.5%, and standing and culturing at 30 ℃ for 10 days to obtain a composite bacterial cellulose membrane;

3) physical compounding: accurately weighing chitosan, adding into water for injection, and filtering with 0.22 μm filter membrane to obtain 8% antibacterial solution; adding the bacterial cellulose membrane purified in the step 2) into an antibacterial agent solution, and magnetically stirring for 8 hours at the compounding temperature of 25 ℃;

EXAMPLE 5 medical dressing and preparation thereof

The preparation method comprises the following steps:

1) biological compounding: inoculating the seed solution into a fermentation culture medium by 8 percent of inoculation amount, adding 0.5 percent of carboxymethyl chitosan into the fermentation culture medium, and standing and culturing for 10 days at 28 ℃ to obtain a composite bacterial cellulose membrane;

2) purification treatment of bacterial cellulose membrane: repeatedly cleaning the bacterial cellulose membrane obtained in the step 1) with purified water until the pH value of the bacterial cellulose membrane is more than or equal to 5.0; then putting the bacterial cellulose membrane into NaOH solution with the concentration of 0.5mol/L according to the ratio of the membrane volume to the alkali liquor volume of 1:2, heating to 80 ℃, stirring for 6h, taking out, washing the bacterial cellulose membrane with water for injection until the pH value is 5.0-7.0 and the content of bacterial endotoxin is less than or equal to 0.03EU/ml, and obtaining the purified bacterial cellulose membrane;

3) physical compounding: accurately weighing Manuka honey, adding into water for injection, and filtering with 0.22 μm filter membrane to obtain 10% antibacterial solution; adding the bacterial cellulose membrane purified in the step 2) into an antibacterial agent solution, and magnetically stirring for 6 hours at the compounding temperature of 40 ℃;

EXAMPLE 6 medical dressing and preparation thereof

The preparation method comprises the following steps:

1) biological compounding: inoculating the seed solution into a fermentation culture medium in an inoculation amount of 12%, adding half of Manuka honey and chitosan quaternary ammonium salt into the fermentation culture medium, wherein the addition amount is 0.5%, and standing and culturing at 32 ℃ for 10 days to obtain a composite bacterial cellulose membrane;

2) purification treatment of bacterial cellulose membrane: repeatedly cleaning the bacterial cellulose membrane obtained in the step 1) with purified water until the pH value of the bacterial cellulose membrane is more than or equal to 5.0; then putting the bacterial cellulose membrane into NaOH solution with the concentration of 0.2mol/L according to the ratio of the membrane volume to the alkali liquor volume of 1:2, heating to 90 ℃, stirring for 4h, taking out, washing the bacterial cellulose membrane with water for injection until the pH value is 5.0-7.0 and the content of bacterial endotoxin is less than or equal to 0.03EU/ml, and obtaining the purified bacterial cellulose membrane;

3) physical compounding: adding the rest Manuka honey and chitosan quaternary ammonium salt into water for injection, and filtering with 0.22 μm filter membrane to obtain 0.01% antibacterial solution; adding the bacterial cellulose membrane purified in the step 2) into an antibacterial agent solution, and magnetically stirring for 4 hours at the compounding temperature of 60 ℃;

Comparative example 1 medical dressing free of chitosan and its derivatives and preparation thereof

Medical dressing: 6 parts of bacterial cellulose and 2 parts of Manuka honey

The preparation method is the same as the example 1, and specifically comprises the following steps:

3) physical compounding: accurately weighing Manuka honey, adding into water for injection, and filtering with 0.22 μm filter membrane to obtain 8% antibacterial solution; adding the bacterial cellulose membrane purified in the step 2) into an antibacterial agent solution, and magnetically stirring for 8 hours at the compounding temperature of 25 ℃;

Comparative example 2 medical dressing without honey and preparation thereof

Medical dressing: 6 parts of bacterial cellulose and 2 parts of chitosan

3) physical compounding: adding chitosan and its derivatives into water for injection, and filtering with 0.22 μm filter membrane to obtain 8% antibacterial solution; adding the bacterial cellulose membrane purified in the step 2) into an antibacterial agent solution, and magnetically stirring for 8 hours at the compounding temperature of 25 ℃;

Experimental example 1 testing of absorption Properties of different medical dressings

Test objects: examples 1-6, comparative examples 1-2.

The test principle is as follows: according to standard YY/T0471.1-2004 contact wound dressing test method part 1: liquid absorbency, water absorption experiments were performed.

The specific test process is as follows:

cutting the above dressing into 10 pieces of 5cm × 5cm size;

preparing a test solution A: 8.298g of sodium chloride and 0.368g of calcium chloride dihydrate powder are respectively weighed, and the volume is determined by using a 1L volumetric flask.

The samples of 5cm × 5cm are placed in a culture dish, and then the test solution preheated to 37 ± 1 ℃) is added, the adding amount is about 40 times of the sample. Transferring into drying oven, and maintaining at 37 + -1 deg.C for 30 min; the sample was held in one corner with forceps, suspended for 30 seconds and weighed.

The other 9 samples were repeated and the average value was taken to obtain the absorption capacity of the sample, with higher absorption capacity indicating better absorption of the sample.

Table 1 medical dressing absorption multiple results

Item	Absorption multiple
		Example 1	20.1
Example 2	18.9
		Example 3	19.8
Example 4	20.5
		Example 5	21.3
Example 6	19.5
		Comparative example 1	15.6
Comparative example 2	1.5

The data in table 1 show that the dressing provided by the invention has good absorption performance, and is beneficial to helping the affected part to recover as soon as possible, wherein if any one of honey or chitosan and derivatives thereof is lacked, the absorption performance of the dressing is obviously reduced.

Experimental example 2 cytotoxicity test of different medical dressings

Test objects: examples 1-6, comparative examples 1-2, and a commercially available silver ion antimicrobial dressing (yubang antimicrobial medical dressing 5.5 x 7.5cm self-adhesive).

The test principle is as follows: cytotoxicity experiments were carried out according to the standard "GB 16886.5-2017 medical device biology evaluation part 5 in vitro cytotoxicity tests".

TABLE 2 criterion for cytotoxicity

Survival Rate of viable cells (%)	Means of
		≥70	Has no cytotoxicity
<70	Potential cytotoxicity

TABLE 3 cytotoxicity of antibacterial materials

As can be seen from the cytotoxicity detection results, compared with the commercially available silver ion antibacterial dressing, the medical dressing has the advantages of better cell proliferation effect, better biocompatibility, smaller probability of causing adverse reactions clinically and safer clinical use of the product.

Experimental example 3 tests on bacteriostatic properties of different medical dressings

Test objects: the antibacterial repairing materials prepared in the examples 1-6 and the comparative examples 1-2 are silver ion antibacterial dressings sold on the market.

The test principle is as follows: evaluation of the antibacterial Properties of textiles according to the Standard GB/T20944.1-2007 part 1: the antibacterial performance test was carried out by the agar plate diffusion method.

The specific test process is as follows:

1. the principle is as follows: two layers of agar culture medium are injected into the plate, the lower layer is a sterile culture medium, and the upper layer is an inoculation culture medium. And placing the sample on two layers of culture media, and qualitatively evaluating the antibacterial performance of the sample according to the bacterial reproduction degree of the contact part of the culture media and the sample after culturing for a certain time.

2. Test bacteria

Gram-positive bacteria, staphylococcus aureus, and gram-negative bacteria, escherichia coli, were used.

3. Preparation of test bacterial solution

3.1 using inoculating loop to fetch the preserved bacteria, inoculating the preserved bacteria on an agar culture medium plate by a line drawing method, and culturing for 24 hours at 37 +/-2 ℃.

3.2 taking 20ml of the nutrient broth, putting the nutrient broth into a 100ml triangular flask, inoculating a typical bacterial colony on a 3.1 plate by using an inoculating ring, and culturing under the conditions that: the temperature is 37 +/-2 ℃, and the vibration frequency is 110min^-1And the time is 18h-24 h.

3.3 diluting the nutrient broth with distilled water 20 times, adjusting the concentration of the cultured broth to 1X 10⁸CFU/ml-5×10⁸CFU/ml, as test bacterial liquid. The concentration of the bacterial liquid is measured by a spectrophotometer or an appropriate method.

4. Preparation of the samples

4.1 test specimens

Representative samples were selected from the samples of examples 1-6, 4 (2 pieces on the front side and 2 pieces on the back side) round samples for each bacteria test, with a diameter of 25mm + -5 mm. The samples should not be sterilized.

4.2 control sample

Comparative examples 1-2 were control samples, the same size as the test samples.

5. Step (ii) of

5.1 preparation of the lower sterile medium. 10.0mL of agar medium was poured into a sterile plate and allowed to set.

5.2 preparing the upper layer of the inoculation medium. 150.0mL of agar medium with the temperature of 45 +/-2 ℃ is put into a beaker, and 1mL of test bacterium liquid is added. The beaker was shaken to distribute the bacteria evenly and 5.0mL of each plate in 5.1 was poured and allowed to clot.

5.3 Place the sample and control separately in the center of the dish with sterile forceps and press evenly on the agar medium until there is good contact between the sample and the agar medium.

5.4 immediately after placing the sample on the agar medium, the sample is placed in an incubator at 37 ℃. + -. 2 ℃ for 18h-24h, ensuring that the sample and the agar medium remain in contact throughout the incubation period.

6. Calculation and evaluation of results

6.1 at least 3 spots were measured per sample, and the width of the zone of inhibition of the sample was calculated as follows.

H＝(D-d)/2

In the formula:

h-width of the bacteriostatic strip, and the unit is millimeter (mm);

d-average of the outer diameter of the zone, in millimeters (mm)

d-sample diameter in millimeters (mm).

6.2 after the zone of inhibition was measured, the sample was removed from the agar medium with tweezers and the contact area under the sample was examined microscopically for bacterial growth.

6.3 the antibacterial effect of each sample was evaluated according to Table 1, based on the presence or absence of bacterial growth and the condition of the zone of inhibition.

TABLE 4 evaluation of antibacterial Effect

6.4 when all the samples satisfy the requirement of "good effect" in Table 1, the sample is considered to have antibacterial effect.

TABLE 5 antimicrobial Effect of different medical dressings

According to the antibacterial detection result, compared with the commercially available silver ion antibacterial dressing, the antibacterial repair material disclosed by the invention has the same antibacterial effect, but compared with the comparative example 1, the antibacterial effect is obviously improved.

Experimental example 4 content test of effective component per unit area of different medical dressings

Test objects: examples 1-6, comparative examples 1-2.

The test principle is as follows: detecting the content of sucrose and maltose according to the detection method of sucrose and maltose in honey of the first part of the edition of Chinese pharmacopoeia 2015, and calculating the content of honey according to conversion. And respectively measuring the nitrogen contents of the chitosan, the carboxymethyl chitosan, the chitosan quaternary ammonium salt, the pure BC film and the composite material by using an element analyzer, and then calculating the contents of the chitosan, the carboxymethyl chitosan and the chitosan quaternary ammonium salt in the composite film by using a formula.

The specific test process is as follows:

(1) sucrose and maltose: the content was calculated by measuring with high performance liquid chromatography according to the following method.

Chromatographic conditions and system applicability test: using Prevail carbonate ES as chromatographic column and acetonitrile-water (75:25) as mobile phase; and detecting by a differential refractive detector. The number of theoretical plates should not be less than 2000 calculated by fructose peak.

Preparation of a standard curve: precisely weighing 1.0g of fructose reference substance and 0.8g of grape reference substance respectively, placing into a conical flask with a plug, precisely adding 20ml of 40% acetonitrile, dissolving, and shaking to obtain fructose and glucose reference substance stock solution. And accurately weighing 0.2g of sucrose reference substance and 0.2g of maltose reference substance, placing the weighed substances into a conical flask with a plug, accurately adding 10ml of 40% acetonitrile, dissolving, and uniformly shaking to obtain the reference substance stock solution of sucrose and maltose. Precisely measuring fructose and glucose reference substance stock solutions and sucrose and maltose reference substance stock solutions respectively, and adding 40% acetonitrile to prepare fructose, glucose, sucrose and maltose mixed reference substance solutions with different concentrations. The amount of stock solution and the dilution volume in each concentration solution formulation are shown in the table below.

Precisely sucking 15 μ l of each mixed reference solution, injecting into liquid chromatograph, and measuring respectively. And (4) taking the concentration of the reference substance as a horizontal coordinate and the peak area value as a vertical coordinate, drawing a standard curve, and calculating a regression equation.

Preparation of a test solution: weighing about lg of the product, accurately weighing, placing in a conical flask with a plug, accurately adding 20ml of 40% acetonitrile, dissolving, shaking uniformly, filtering, and taking the subsequent filtrate to obtain the final product.

The determination method comprises the following steps: precisely measuring the sample solution 15ul, injecting into a liquid chromatograph, measuring, and calculating the content according to a standard curve method.

(2) Measuring the contents of chitosan, carboxymethyl chitosan and chitosan quaternary ammonium salt: the nitrogen content in chitosan, carboxymethyl chitosan, chitosan quaternary ammonium salt, pure BC film and composite material is respectively tested by using a German Elmentar variao EL type III element analyzer, and is calculated by the following formula:

chitosan (carboxymethyl chitosan, chitosan quaternary ammonium salt) (%) - (W)_C-W_BC)/Wch×100

W_CIs the nitrogen content in the composite material, W_BCIs the nitrogen content in the pure BC film, Wch is chitosan, carboxymethyl chitosan or chitosan quaternary ammonium saltThe nitrogen content of (1).

The results of the tests are given in the following table

TABLE 6 content of effective component per unit area of different medical dressings

Note: represents that this substance was not detected.

The data in the table show that the content of the functional components of the dressing prepared by combining biological compounding with physical mode is obviously higher than the content of the functional components of simple physical compounding.

The above detailed description is specific to one possible embodiment of the present invention, and the embodiment is not intended to limit the scope of the present invention, and all equivalent implementations or modifications without departing from the scope of the present invention should be included in the technical scope of the present invention.

Claims

1. A medical dressing applied to chronic wounds comprises bacterial cellulose, honey, chitosan and derivatives thereof, and is characterized in that the medical dressing comprises, by weight, 5-10 parts of bacterial cellulose, 0.5-5 parts of honey, 0.5-5 parts of chitosan and derivatives thereof;

the honey is Manuka honey;

the chitosan and the derivatives thereof are selected from: one or more of chitosan, carboxymethyl chitosan and chitosan quaternary ammonium salt.

2. The medical dressing applied to the chronic wound surface of claim 1, wherein the medical dressing is composed of, by weight, 6-9 parts of bacterial cellulose, 1-3 parts of honey, 1-3 parts of chitosan and derivatives thereof.

3. A method of preparing a medical dressing for application to a chronic wound surface as claimed in any one of claims 1 to 2,

the method comprises the following steps:

4. A method of manufacturing a medical dressing for chronic wounds according to claim 3, comprising the steps of:

3) physical compounding: adding manuka honey, chitosan and derivatives thereof into water for injection, and filtering to obtain an antibacterial agent solution with the concentration of 0.01-10%; adding the bacterial cellulose membrane purified in the step 2) into an antibacterial agent solution, and stirring for 1-12 h;

4) drying the composite membrane: drying the membrane of step 3).

5. A method of manufacturing a medical dressing for chronic wounds as claimed in any one of claims 1 to 4, comprising the steps of:

3) physical compounding: adding the rest Mel and/or chitosan and its derivatives into water for injection, and filtering to obtain antibacterial solution; adding the purified composite bacterial cellulose membrane obtained in the step 2) into the filtered antibacterial agent solution for physical compounding;

6. A method of manufacturing a medical dressing for a chronic wound according to claim 5, comprising the steps of:

1) biological compounding: inoculating the seed solution into fermentation culture medium at an inoculation amount of 1-20%, adding Manuka honey and/or chitosan and its derivatives, and standing at 28-35 deg.C for 5-14 days to obtain composite bacterial cellulose membrane;

4) drying the composite membrane: drying the membrane of step 3).