Easily stripped puffball spore dressing as well as preparation method and application thereof

Abstract

The invention relates to a preparation method and application of a lasiosphaera seu calvatia spore dressing easy to peel, and belongs to the field of biological medicine. According to the dressing, the puffball spore powder and the medical proof fabric are directly combined, no excipient needs to be added, the preparation process is simple, the cost is low, and the dosage is small. The puffball spore dressing prepared by the method has excellent easy-to-peel property, can effectively prevent wound exudate from being adhered to the dressing, and relieves the pain of a patient during dressing change; meanwhile, the hemostatic effect is remarkable, and bleeding can be quickly stopped. In addition, the puffball spores, which are in direct contact with the wound to exert effects, of the dressing are bio-based materials, are good in biocompatibility and can be safely used clinically. The invention further provides a specific preparation method and application of the dressing.

Description

Technical Field

[0001] The invention belongs to the field of biomedicine, and in particular relates to an easily peelable puffball spore dressing, a preparation method and an application thereof. Background Art

[0002] Trauma is a common and frequently occurring disease in clinical practice. At present, traffic accidents, natural disasters, war conflicts, medical trauma, etc. still occur from time to time. [1-2] Wound dressings are an important part of trauma care. Ideal wound dressings should have good biocompatibility, excellent hemostatic properties, and non-adhesive properties. Traditional wound dressings, such as gauze and cotton pads, have the disadvantages of easy adhesion to wounds and poor hemostatic effects. Currently, non-adhesive dressings are mainly constructed by combining hydrophobic layers to create Janus structures or by loading hydrophobic materials onto the surface of the dressing substrate.

[0003] Puffball spores are a natural biomaterial with the advantages of good biocompatibility and significant hemostatic effect. It has a history of medicinal use in my country for nearly two thousand years. [3] Puffball is a fungus of the family Calvataceae, and there are many varieties, such as the big puffball Calvatia gigantea (Batschex Pers.) Lloyd, the peeling puffball Lasiosphaerafenzlii Reich. or the purple puffball Calvatialacina (Mont. et Berk.) Lloyd, Bovistella sp., and Calvatia sp. The spores of different varieties of puffball have different microscopic morphologies, which affect their hydrophilic and hydrophobic properties.

[0004] Ancient books and modern clinical records show that puffball spores are often used as medicine. [4-5] However, there are many limitations to using powdered puffball spores directly for hemostasis, such as easy scattering. It has been reported that puffball spore dressings usually require excipients such as gelatin and chitosan to improve their formability and mechanical properties. The addition of these excipients not only increases the complexity of the preparation process, but also may affect the hydrophilicity and hydrophobicity of the puffball spore dressing, thereby affecting the peeling force from the wound. Therefore, it is of great practical value to develop an easy-to-peel puffball spore dressing that is simple to prepare and effective in hemostasis. Summary of the invention

[0005] In order to solve the above technical problems, the present invention provides the following technical solutions:

[0006] A first aspect of the present invention provides an easily peelable puffball spore dressing, which is composed of puffball spores and medical adhesive tape, and the puffball spores are evenly coated on the medical adhesive tape.

[0007] Furthermore, in the puffball spore dressing, the source of the puffball spores includes one or more of Bovistellasp., Calvatia sp., and Lasiosphaera sp.

[0008] Furthermore, in the puffball spore dressing, the amount of puffball spores applied is 3.38 to 20.25 g / m 2 , preferably 6.76~16.88g / m 2 .

[0009] A second aspect of the present invention provides a method for preparing any of the above-mentioned puffball spore dressings, comprising the following steps:

[0010] (1) According to the above ratio, puffball spores are attached to the sticky side of medical adhesive tape;

[0011] (2) After the spores are firmly attached, the prepared dressing is cut into the required size and sealed and packaged;

[0012] (3) Sterilize and keep for later use.

[0013] Furthermore, the specific steps of step (1) include: using a spray gun to evenly spray the puffball spores on the sticky side of the medical tape; or using a screen to evenly sprinkle the puffball spores on the sticky side of the medical tape; or immersing the medical tape in a spore suspension to attach the puffball spores to the medical tape.

[0014] Furthermore, the sterilization method includes ethylene oxide sterilization or irradiation sterilization.

[0015] Furthermore, in the puffball spore dressing, the dressing can absorb exudate to protect the wound.

[0016] Furthermore, in the puffball spore dressing, the dressing can quickly stop bleeding and protect the wound.

[0017] Beneficial Effects

[0018] The present invention provides a preparation method and application of an easily peelable puffball spore dressing. The effective effects include: (1) the dressing is made by directly combining puffball spore powder with medical adhesive tape without adding any excipients, and the preparation process is simple, the dosage is small, and the cost is low; (2) the puffball spore dressing prepared by the method of the present invention has excellent easy peelability, can effectively prevent wound exudate from adhering to the dressing, and reduce the pain of patients during dressing changes; (3) it has a significant hemostatic effect and can stop bleeding quickly; (4) the puffball spores that the dressing directly contacts with the wound to exert its effect are bio-based materials, have good biocompatibility, and can be safely used in clinical practice. BRIEF DESCRIPTION OF THE DRAWINGS

[0019] Figure 1 Shown are the results of the hemostatic efficacy of puffball spore dressing;

[0020] Figure 2 Shown are safety results for the puffball spore dressing. DETAILED DESCRIPTION

[0021] The present invention will be further illustrated below. It should be noted that the examples illustrate some preparation or use methods, however, it should be understood that these examples do not limit the present invention. The scope of protection of the present invention shall be subject to the contents recorded in the attached claims.

[0022] In the following examples, the 1 / 10,000 balance is BSA4202SA from Sartorius, Germany. The multifunctional centrifuge is CAX-371 produced by Tomy, Japan. The microplate reader is Synergy H1 produced by Berton Instruments, USA. The Spark full-wavelength microplate reader is produced by Tecan Biotech, Switzerland. The liquid transfer gun is produced by Shanghai Aibende Technology Co., Ltd. The tensile meter is produced by Wenzhou Weidu Electronics (load 1000mN / division value 0.01mN).

[0023] The materials used are GIBCO TM DMEM culture medium (GIBCO, USA); Hyclone TM Fetal bovine serum (Cytiva, USA); 0.25% trypsin, penicillin-streptomycin double antibody (Shanghai Yishen Biotechnology Co., Ltd.); CCK-8 test kit (Dojin Chemical Research Institute, Japan); anhydrous ethanol (Xilong Chemical Co., Ltd.); citrate anticoagulated sheep blood (Guangzhou Hongquan Biotechnology Co., Ltd.); medical tape (Hainuo Beishiwei Medical Products Qingdao Co., Ltd.); cotton fiber dressing (Shenzhen Wenjian Medical Company); QuikClot Combat Gauze dressing (Chinook Medical Gear, USA); hydrophilic dressing (3M Company, USA); Shutai (Vick Group, France); Speed ​​Sleep New (Dunhua Shengda Animal Pharmaceutical Co., Ltd.); the puffball spores are commercially available.

[0024] All experiments shown in the examples were designed for at least three parallel experiments per group, and the peeling force experimental data were expressed as mean values ​​(Mean); the experimental data at the cell level and animal level were expressed as mean ± standard deviation (Mean ± SD). SPSS26.0 was used to analyze the experimental data and GraphPad Prism 8 was used for graphing, and one-way analysis of variance was used for inter-group comparison. Statistical differences were indicated as follows: *p<0.05, **p<0.01, ***p<0.001, ns represents no significant difference.

[0025] Example 1 Preparation of puffball spore dressing using a sieve method

[0026] (1) Place a piece of 5*10 cm medical tape on the laboratory table;

[0027] (2) Take 0.1 g of puffball spores and place them on a 300-mesh sieve, and spread them evenly on the medical tape through the sieve to ensure uniform distribution;

[0028] (3) Gently press the spores to allow them to bond to the medical tape;

[0029] (4) Cut the dressing into appropriate sizes of 2*5 cm and seal it;

[0030] (5) Sterilize the dressing by irradiation for 2 hours.

[0031] Example 2 Preparation of puffball spore dressing using spray gun method

[0032] (1) Place a piece of 5*10 cm medical tape on the laboratory table;

[0033] (2) Take 0.1 g of puffball spores and put them into a spray gun, and spray the spores evenly on the medical tape;

[0034] (3) Cut the dressing into 2*5cm size and seal it;

[0035] (4) Sterilize the dressing with ethylene oxide for 12 hours.

[0036] Example 3 Preparation of puffball spore dressing by soaking method

[0037] (1) Mix 1 gram of puffball spores with 100 ml of anhydrous ethanol to prepare a spore suspension;

[0038] (2) Immerse a piece of 5*10 cm medical tape in the spore suspension for 1 minute;

[0039] (3) Take out the dressing and dry it on a drying plate at 30°C for 1 hour;

[0040] (4) Cut the dressing into 2*5 cm size and seal it;

[0041] (5) Sterilize the dressing with ethylene oxide for 12 hours.

[0042] Example 4 Evaluation of the peeling force of puffball spore dressing

[0043] 4.1 Experimental methods

[0044] 3M hydrophilic dressing was used as the matrix material for blood coagulation, and Quikclot combat gauze was used as a control. The base material (100 mm wide and 100 mm long) was glued to a stainless steel substrate; sodium citrate anticoagulated sheep blood was mixed with 0.2 M CaCl 2The solution was dropped onto the substrate at a volume ratio of 10:1 and soaked with blood. Immediately place each coating amount of 20 mm wide and 50 mm long dressing on the matrix material to allow a clot to form between the hemostatic sample and the matrix material. First, coagulation was performed at 37°C for 1 hour; the coagulated sample was exposed to the hot air of a hair dryer for 30 minutes to accelerate the coagulation of the clot. After that, the sample was peeled off from one side with a tensile gauge to measure the peeling force. The maximum peeling force was divided by the width of the sample, and each group was tested three times in parallel.

[0045] 4.2 Experimental Results

[0046] By comparing the peel strength of the puffball spore dressing prepared by the sieve method with the commercially available dressing, the experiment found that the coating amount was not less than 6.76 g / m 2 The dressing can cover the surface of the dressing with spores, thereby effectively reducing the peeling strength and playing the role of easy peeling. The coating amount is 13.50g / m 2 It can save the amount of medicine while ensuring "easy to peel".

[0047] Table 1. Peeling force of puffball spore dressings with different coating amounts

[0048]

[0049] Example 5 Evaluation of the hemostatic efficacy of puffball spore dressing

[0050] 5.1 Experimental Methods

[0051] The rat tail amputation model was used to evaluate the hemostatic effect of puffball spore dressing. The SD rats were weighed and anesthetized with a compound anesthetic (Sutane: Sumianxin: saline = 1:1:8, 0.1 mL / 100 g). The rats were fixed with their backs facing up and cut short at 4 cm from the tip of the tail. The material was immediately applied to stop bleeding and photographed for record. After hemostasis, the hemostasis time was recorded.

[0052] 5.2 Experimental Results

[0053] The rat tail-cut experiment was used to evaluate the hemostatic efficacy of puffball spore dressings. The experiment found that puffball spore dressings can effectively promote rapid hemostasis of vascular injury, prevent excessive blood loss, avoid death caused by massive bleeding, and help the body recover faster. We used the commercially available first-line hemostatic agent QuikClot as a positive drug and cotton fiber dressings as a control. The powder group represented by puffball spores B.sp.spore and the puffball spore dressing B.sp.spore-dressing (coating amount of 13.50g / m 2 The results showed that B.sp.spore and B.sp.spore-dressing (coating amount of 13.50 g / m 2) can effectively accelerate the hemostasis process and is comparable to the existing first-line hemostatic drug QuikClot in terms of shortening the hemostasis time (see Figure 1 ).

[0054] Example 6 Safety Evaluation of Puffball Spore Dressing

[0055] 6.1 Experimental methods

[0056] Cytotoxicity evaluation: Mouse fibroblast cell line L929 was used to evaluate the cytotoxicity of the dressings. This cell line is commonly used for cytotoxicity analysis of wound dressings. The cells were cultured in high-glucose DMEM supplemented with 10% FBS and 1% double antibody under 5% CO 2 , cultured at 37°C. CCK-8 counts of cell viability: L929 cells (5×10 3 / mL) were inoculated into a 96-well plate (100 μL / well), and 6-8 hours later, B. sp. spore-dressing (coating amount was 13.50 g / m 2 ) filtrate treatment. After 48 h, 10 μL CCK-8 solution was added to each well and incubated at 37 °C for 2 h. Finally, the absorbance was measured at 450 nm using an enzyme reader.

[0057] 6.2 Experimental Results

[0058] The crushed puffball spore dressing (coating amount of 13.50 g / m 2 ) was immersed in the culture medium overnight and filtered to obtain 100% filtrate. The cell survival rate was tested by CCK-8 method to investigate whether the filtrate had toxic effects on the mouse fibroblast cell line L929. The experimental results showed that the B.sp. spore-dressing filtrate had no significant cytotoxic effect on L929 cells (see Figure 2 ).

[0059] References:

[0060] [1]MBDowling,A.Chaturvedi,ICMacIntire,V.Javvaji,J.Gustin,SRRaghavan,TMScalea,M.Narayan,Injury-international Journal of The Care ofThe Injured,2016,47,2105-2109.

[0061] [2] JFKelly, AERitenour, DFMcLaughlin, KABAgg, ANApodaca, CTMallak, L.Pearse, MMLawnick, HRChampion, JBHolcomb, Journal of Trauma-InjuryInfection and Critical Care, 2008, 64, S21-S26.

[0062] [3] Chinese Pharmacopoeia Committee, Pharmacopoeia of the People’s Republic of China: 2020 Edition, Part 1, Beijing 2020.

[0063] [4] Bei Minmin, Journal of Traditional Chinese Medicine and External Therapy, 2000, 9, 47.

[0064] [5] Meng Longjiang, Chinese Journal of Traditional Chinese Medicine Information, 2001, 8, 70.

Claims

1. An easily peelable puffball spore dressing, characterized in that: The medical adhesive tape consists of puffball spores and medical adhesive tape, and the puffball spores are evenly coated on the medical adhesive tape.

2. The puffball spore dressing according to claim 1, characterized in that The sources of the puffball spores include one or more of Bovistella sp., Calvatia sp., and Lasiosphaera sp.

3. The puffball spore dressing according to claim 1, characterized in that The puffball spore coating amount is 3.38-20.25 g / m 2 , preferably 6.76~16.88g / m 2 .

4. A method for preparing the puffball spore dressing according to any one of claims 1 to 3, characterized in that: The following steps are involved: (1) According to the above ratio, puffball spores are attached to the sticky side of medical adhesive tape; (2) After the spores are firmly attached, the prepared dressing is cut into the required size and sealed and packaged; (3) Sterilize and keep for later use.

5. The method for preparing puffball spore dressing according to claim 4, characterized in that: The specific steps of step (1) include: using a spray gun to evenly spray the puffball spores on the sticky side of the medical tape; or using a screen to evenly sprinkle the puffball spores on the sticky side of the medical tape; or immersing the medical tape in a spore suspension to attach the puffball spores to the medical tape.

6. The method for preparing the puffball spore dressing according to claim 4, characterized in that: The sterilization method includes ethylene oxide sterilization or irradiation sterilization.

7. The puffball spore dressing according to any one of claims 1 to 3, characterized in that The dressing can absorb exudate and protect the wound.

8. The puffball spore dressing according to any one of claims 1 to 3, characterized in that The dressing can quickly stop bleeding and protect the wound.

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