Composite antibacterial dressing, preparation method and application of composite antibacterial dressing in bone cultural relic protection material

Abstract

The invention discloses a composite antibacterial dressing, a preparation method and application of the composite antibacterial dressing in bone cultural relic protection materials, and relates to the technical field of bacteriostasis and the technical field of cultural relic protection.The composite antibacterial dressing comprises an elastomer layer and a hydrogel layer, and the hydrogel layer is attached to or embedded into the elastomer layer; the hydrogel layer is used for being attached to a cultural relic, and the hydrogel layer is hydrogel containing chlorhexidine digluconate; the elastomer layer is a polymer elastomer containing lipopeptide, a lipopeptide solution can inhibit the growth of pseudomonas, the composite antibacterial dressing can effectively inhibit the growth of aspergillus, trichoderma and pseudomonas in bone cultural relics, and meanwhile, the dressing can achieve the effects of preserving moisture and slowing down the water loss speed of the cultural relics, and is convenient to operate and carry. The invention also provides a preparation method of the composite antibacterial dressing, and the product obtained by the method has stable antibacterial and moisturizing effects.

Description

Technical Field

[0001] The invention relates to the technical fields of antibacterial treatment and cultural relic protection, and in particular to a composite antibacterial dressing and a preparation method thereof, and application of the composite antibacterial dressing in bone cultural relic protection materials. Background Art

[0002] Sanxingdui is an ancient cultural site located in Guanghan City, Sichuan Province. It has a history of thousands of years and covers an area of ​​about 12 square kilometers. It is the most important pre-Qin site discovered in the southwest region so far, and has important cultural significance and historical value. The archaeological excavation of Sanxingdui began as early as 1934, and has a history of more than 90 years. During this period, more than 40 archaeological excavations have been carried out, and 8 sacrificial pits have been discovered. A rich variety of historical relics such as jade, bronze, and gold have been unearthed, including a large number of whole ancient ivory and ivory products. According to statistics, more than 80 ancient ivory tusks were unearthed in the No. 1 and No. 2 sacrificial pits of Sanxingdui, 100 relatively complete ivory tusks were unearthed in the No. 3 pit, 47 ivory tusks were unearthed in the No. 4 pit, and about 160 ivory tusks have been found in the No. 7 pit, and the total data is estimated to be close to 200, which provides important archaeological clues for exploring the ancient Shu culture, ivory sacrificial traditions, and ancient ivory crafts. If these unearthed ancient ivory can be preserved intact, they will not only leave important cultural and historical marks for our country's long history, but also provide reference ideas for the protection of other cultural relics, and have important research and cultural value.

[0003] The ivory unearthed from the Sanxingdui site has experienced a sudden change from a stable underground environment with darkness, low oxygen, low temperature and high humidity to an environment with sudden changes in temperature and humidity, a sudden increase in oxygen content, increased light and an increase in harmful organisms, which has a great impact on the stability of the ivory. If there are no effective emergency protection measures, the moisture inside the ivory will be rapidly lost after being unearthed. At the same time, under the action of harmful microorganisms such as mold and oxygen, the internal structure of the ivory will be destroyed, the surface will crack and powder, and irreversible damage will occur. The most ideal protection method for ancient ivory is to reinforce it on site when it is just unearthed, but due to the limitations of the archaeological environment, the excavation site may not have the equipment required for curing, and on-site curing will also affect the progress of the site, so the unearthed ivory needs to be transported back to the room for cleaning and reinforcement. Therefore, it is necessary to develop an emergency protection method for ancient ivory, so that the ancient ivory can maintain its original humidity and water content as much as possible during the transportation process from unearthed to curing, so as to buy more time for the subsequent protection work.

[0004] Research reports that microbial corrosion of ancient ivory may occur through the following pathways: (1) Acidification: According to research, ancient ivory, fresh ivory, and natural apatite are all easily soluble in acid, and the carbon dioxide produced by microbial respiration and the organic acids secreted create an acidic environment, accelerating the corrosion of ancient ivory; (2) Mechanical destruction: The biggest difference between ancient ivory and fresh ivory is that the organic components that mainly act as a bonding phase in ivory materials have been eroded and destroyed, and the loose and porous structure provides space for the growth of microbial hyphae, further exacerbating the damage to the internal structure; (3) Other metabolic effects: During the growth process, microorganisms may absorb nutrients from cultural relics and produce complex biochemical ligands, which play an important role in mineral dissolution and inducing / controlling mineral formation. Therefore, when developing emergency ivory protection materials, the antibacterial properties of the materials should also be considered to reduce the corrosion of ancient ivory by microorganisms.

[0005] There are few reports on emergency protection of unearthed ivory abroad. At first, China used isolation materials such as hemp paper or plastic film to temporarily protect unearthed ivory. However, since the materials are easy to mold and difficult to remove, and the objects may be displaced due to improper bonding, this method cannot protect the ivory well. The Chengdu Archaeological Institute currently uses silicone potting to temporarily protect unearthed ivory. The moisture of ivory is completely locked in the initial stage of silicone potting, so that the unearthed ivory is preserved in an environment close to the original burial environment for a period of time, which greatly reduces the water loss rate of ancient ivory and isolates the pollution of ancient ivory by harmful gases and microorganisms from the outside. The initial effect of potting is significant, but the storage device of ancient ivory is huge after potting, which will cause certain inconveniences for transportation and exhibition, and silicone is expensive and costly.

[0006] At present, emergency protection measures for ivory at home and abroad need to be further optimized to improve the above problems. Summary of the invention

[0007] The purpose of the present invention is to provide a composite antibacterial dressing and a preparation method. The obtained composite antibacterial dressing has a good inhibitory effect on Aspergillus, Trichoderma and Pseudomonas on bone cultural relics, and isolates the pollution of external harmful gases and microorganisms to ancient ivory to a certain extent. At the same time, the antibacterial dressing can effectively slow down the water loss rate of ancient ivory, which is beneficial to maintaining the appearance of bone cultural relics such as ancient ivory, and is also more convenient for transportation.

[0008] The present invention is achieved through the following technical scheme: a composite antibacterial dressing, comprising an elastomer layer and a hydrogel layer, wherein the hydrogel layer is adhered to or embedded in the elastomer layer, and the hydrogel layer is used to be adhered to the cultural relics, wherein the hydrogel layer is a hydrogel containing chlorhexidine digluconate; and the elastomer layer is a polymer elastomer containing lipopeptides, and the lipopeptide solution can inhibit the growth of Pseudomonas.

[0009] Furthermore, the concentration of chlorhexidine digluconate in the hydrogel layer is 0.5-4%; and the concentration of lipopeptide in the elastomer layer is 12.5-50 mg / mL.

[0010] Furthermore, the thickness of the hydrogel layer is 0.5-2 mm; the thickness of the elastomer layer is 0.1-1 mm.

[0011] The preparation method of the aforementioned composite antibacterial dressing comprises the following steps: S1. A 0.5-4% chlorhexidine digluconate solution was prepared, and then AM, SDS, MBA, glycerol, deionized water and a photoinitiator were placed in a container, and then the chlorhexidine digluconate solution was added to the container and stirred to obtain a 0.5~4 chlorhexidine digluconate hydrogel; S2. Inoculating and fermenting Bacillus amyloliquefaciens M73 on Landy medium, isolating lipopeptides that inhibit Pseudomonas, dissolving the lipopeptides in a methanol-chloroform solvent to obtain a lipopeptide solution, and immersing a silicone elastomer in the lipopeptide solution. After 3 to 10 minutes, an elastomer layer having a lipopeptide concentration of 12.5 to 50 mg / mL is obtained; S3. The hydrogel layer prepared in step S1 is then bonded to the elastomer layer prepared in step S2, or the hydrogel layer prepared in step S1 is embedded in the elastomer layer prepared in step S2 to obtain a composite antibacterial dressing.

[0012] Furthermore, in step S1, the method for preparing the hydrogel comprises the following steps: a1. Take raw materials AM 23.06%, SDS 0.03%, MBA 7.69%, glycerol 15.38%, deionized water 53.82% and photoinitiator 2959 0.02% by mass percentage, and then place the above raw materials in a container; b1. Use a stirrer to stir the material in the container. When the solid is completely dissolved, use a vacuum pump to extract the dissolved gas in the solution; c1 After the gas was exhausted, chlorhexidine digluconate mother solution was added thereto to obtain chlorhexidine digluconate hydrogel; d1. After stirring evenly, pour the solution into the mold and let it stand under the UV lamp for 0.4~1h. After it is formed, demould it for later use.

[0013] Furthermore, in step S2, the method for preparing the lipopeptide comprises the following steps: a2. Preparation of culture medium: Prepare Landy medium and sterilize; b2. Inoculation and fermentation: Inoculate 1×10 5CFU / mL of Bacillus amyloliquefaciens M73 culture was fermented in a shaker at 30°C and 2000 rpm for 3 days; c2. Lipopeptide extraction: After fermentation, centrifuge at 8000rpm for 5min, collect the supernatant, adjust the pH to 2 with hydrochloric acid, and let stand until the lipopeptide precipitates; d2. Centrifuge again, discard the supernatant, dry the precipitate to obtain lipopeptide powder, then dissolve the lipopeptide powder with methanol-chloroform solvent, filter and take the filtrate to obtain a lipopeptide complex solution.

[0014] Furthermore, in step c2, the lipopeptides are separated by high performance liquid chromatography and analyzed by electrospray mass spectrometry, and the high performance liquid chromatography conditions satisfy: Chromatographic column: Octadecylsilane bonded silica gel is used as filler; Column temperature: 25°C; Flow rate: 4.0ml / min; Injection volume: 1 μL; Detection wavelength: 214nm; Mobile phase A was acetonitrile, mobile phase B was 0.1% formic acid solution, gradient elution; The elution process is: 0~50min, mobile phase A is 2~95%, mobile phase B is 98~5%.

[0015] Mass spectrometer detection conditions: electrospray voltage 5000 V, capillary injection port voltage 60 V, capillary heating temperature 320 °C, acceleration voltage 6000 V, reflector voltage 1210 V, repulsion pulse frequency 5000 Hz, mass detection range 0.5-2.5 kDa.

[0016] Furthermore, in step d2, the methanol-chloroform solvent is a mixed solution of methanol:chloroform in a volume ratio of 1:1, and 12.5-50 mg of lipopeptide powder is dissolved in each milliliter of methanol-chloroform solvent.

[0017] Furthermore, in step S2, the preparation method of the silicone elastomer is: mix the main agent and the curing agent in the SYLGARD 184 silicone elastomer kit in a ratio of 10-20:1, add 0.5-2% TMSPMA of the total volume, mix well and then cure in an oven at 60°C to obtain the silicone elastomer.

[0018] The composite antibacterial dressing as described above or the composite antibacterial dressing obtained by the above-mentioned preparation method can be used as a protective material for inhibiting Aspergillus, Trichoderma and Pseudomonas in bone cultural relics.

[0019] Compared with the prior art, the present invention has the following advantages and beneficial effects: 1. In this solution, the composite antibacterial dressing solves the problem that there is no good protection measure for ancient ivory and bone cultural relics when they are unearthed. At present, silicone potting is used to temporarily protect the unearthed ivory when such cultural relics are unearthed, which makes transportation and exhibition inconvenient, and the materials involved in the process are expensive and costly, which is not conducive to the development of archaeological work.

[0020] 2. In this plan, the advantages of hydrogels and elastomers in the existing technology are combined and applied to the protection of cultural relics. At the same time, the characteristics of this type of cultural relics are investigated and studied in depth. By combining technologies from multiple fields such as biology, chemistry, and materials, products suitable for the protection of bone cultural relics are developed, which promotes the development of the archaeological industry.

[0021] 3. In this plan, according to the characteristics of bone cultural relics, taking the ancient ivory of Sanxingdui as an example, a composite antibacterial dressing containing chlorhexidine and lipopeptide suitable for this type of cultural relics is prepared. Among them, the lipopeptide is obtained by fermentation and separation of Bacillus amyloliquefaciens M73, that is, the application of Bacillus amyloliquefaciens M73 is increased to ensure sufficient source of antibacterial raw materials. It is also a further development and application of products with independent intellectual property rights. The technology is more mature and the product use is less restricted. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 The results of HPLC analysis of Bacillus amyloliquefaciens M73 are shown.

[0023] Figure 2 This is the mass spectrum of the a2 fraction.

[0024] Figure 3 This is the secondary mass spectrum analysis diagram of the active substances in the a2 fraction.

[0025] Figure 4 is a schematic diagram of the hydrogel.

[0026] Figure 5 This is a graph showing the antibacterial effect of SDS-free hydrogel on several ivory-corrosive bacteria.

[0027] Figure 6 This is a diagram showing the antibacterial effect of a hydrogel compounded with 2% chlorhexidine digluconate.

[0028] Figure 7 This is a graph showing the antibacterial effect of the viscous formula hydrogel on several ivory-corrosive bacteria.

[0029] Figure 8 This is a graph showing the antibacterial effect of non-sticky formula hydrogel on several ivory-corrosive bacteria.

[0030] Fig. 9 is a schematic diagram of a blank elastomer.

[0031] Fig.10This is the antibacterial effect diagram of the blank elastomer.

[0032] Fig.11 This is a diagram showing the antibacterial effect of elastomer soaked in lipopeptide.

[0033] Fig.12 This is a diagram showing the antibacterial effect of the composite antibacterial dressing.

[0034] Fig.13 This is a comparison chart of the antibacterial effect of the composite antibacterial dressing on Trichoderma one month before and one month after. DETAILED DESCRIPTION

[0035] The present invention is further described in detail below in conjunction with the examples, but the embodiments of the present invention are not limited thereto. In the present invention, the equipment and reagents not specifically described are all conventional equipment and reagents, and can be directly purchased on the market.

[0036] Example 1 In this embodiment, a composite antibacterial dressing including an elastomer layer and a hydrogel layer is taken as an example, and a preparation method of the composite antibacterial dressing is provided. The antibacterial effect of the product obtained by the method of the present invention and the antibacterial effect when used in bone cultural relics are examined to further illustrate this solution.

[0037] 1. Preparation of hydrogel layer 1.1. The main reagents used to prepare antibacterial hydrogels are shown in Table 1.

[0038] Table 1 Main reagents for preparing antibacterial hydrogels 1.2. Preparation of antibacterial hydrogel S1. According to the reagents in Table 1, weigh AM (7.5 g), SDS (0.01 g), MBA (2.5 g, 1 / 1000 mass fraction aqueous solution), glycerol (5 g), deionized water (17.5 g) and photoinitiator 2959 (0.0075 g) into a container; S2. Use a magnetic stirrer to stir the material in the container. When the solid is completely dissolved, use a vacuum pump to extract the dissolved gas in the solution; S3. When the gas is completely exhausted and no more bubbles are generated, the vacuum pump is turned off, and an appropriate amount of 20% (w / v) chlorhexidine digluconate mother solution is added thereto to obtain a chlorhexidine digluconate hydrogel; S4. After stirring evenly, pour the solution into the mold and let it stand under the UV lamp for about 0.5h. After it is formed, demould it for use.

[0039] 2. Preparation of the Elastomer Layer 2.1 Preparation of lipopeptides 2.1.1 Strain used for preparation of lipopeptides: Bacillus amyloliquefaciens M73.

[0040] 2.1.2 Preparation of main reagents for lipopeptides The main reagents used to prepare lipopeptides are shown in Table 2.

[0041] Table 2 Main reagents for preparing lipopeptides 2.1.3 Preparation of lipopeptides Step 1: Prepare Landy medium Weigh 20 g of glucose, 5 g of sodium L-glutamate, 5 g of tryptone, 1 g of yeast extract, 0.5 g of MgSO4·7H2O, 0.5 g of KCl, 0.5 g of K2HPO4, 0.15 mg of Fe2SO4·7H2O, 5 mg of MnSO4, and 0.16 mg of CuSO4·5H2O into a conical flask, completely dissolve them with 1000 mL of distilled water, adjust the pH to 7.3, and sterilize them by high-pressure steam at 121°C for 20 min.

[0042] Step 2: Inoculation and fermentation In the clean bench, 1×10 5 CFU / mL of Bacillus amyloliquefaciens M73 culture liquid was fermented in a shaker at 30°C and 2000 rpm for 3 days.

[0043] Step 3: Extraction of lipopeptides After fermentation, centrifuge at 8000 rpm for 5 min, collect the supernatant, adjust the pH to 2 with hydrochloric acid, and let stand overnight to wait for the lipopeptide to precipitate.

[0044] In this example, the active ingredients in the lipopeptide were further investigated.

[0045] The lipopeptides obtained by fermenting M73 with Bacillus amyloliquefaciens were detected and separated by high performance liquid chromatography (HPLC). The lipopeptides were separated by high performance liquid chromatography and analyzed by electrospray mass spectrometry. The high performance liquid chromatography conditions met the following requirements: Chromatographic column: Octadecylsilane bonded silica gel is used as filler; Column temperature: 25°C; Flow rate: 4.0ml / min; Injection volume: 1 μL; Detection wavelength: 214nm; Mobile phase A was acetonitrile, mobile phase B was 0.1% formic acid solution, gradient elution; The elution process is: 0~50min, mobile phase A is 2~95%, mobile phase B is 98~5%.

[0046] Mass spectrometer detection conditions: electrospray voltage 5000 V, capillary injection port voltage 60 V, capillary heating temperature 320 °C, acceleration voltage 6000 V, reflector voltage 1210 V, repulsion pulse frequency 5000 Hz, mass detection range 0.5-2.5 kDa.

[0047] A. The first preparation yielded three fractions, a, b, and c, of which fraction a had antibacterial activity; B. Fraction a1, a2 and a3 are obtained by separating and re-preparing fraction a, wherein fraction a2 has gray mold inhibitory activity, see Figure 1 The main active substances in the a2 fraction were further detected by electrospray ionization (ESI)-Q-TOF-MS. The MS / MS high-resolution mass spectrum of the a2 fraction gave a (M+H) ion peak at 1021.5153m / z, 1035.5315m / z, and 1049.5473m / z, see Figure 2 , they are p4, p5, and p6 respectively.

[0048] In general, each peak in the spectrum may be due to the presence of various types of fragments. Fraction a2 was decomposed into small molecules by MS / MS, and the obtained spectrum was mainly composed of b-type and y-type ions. Figure 3From the mass spectrometry data, the b-type ions of the 1021.5153m / z compound are b1 (340.2593m / z), b2 (503.3219m / z), b3 (617.3638m / z), and b4 (704.4381m / z), and the corresponding main y-type ion detected is y5 (188. 0710m / z), and the corresponding other main fragments detected are Ser-Glu (216.0806m / z), (D-Tyr) + (D-Asn) (278.1141m / z), Thr + C14β-NH2FA + Asn (441.3095m / z), The compounds with 1035.5315 m / z are b1 (354.2751 m / z), b2 (517.3219 m / z), b3 (631.3638 m / z), b4 (718.4381 m / z), and the corresponding other major fragments detected are Thr + C14β-NH2FA + Asn (455.3218 m / z) and (CH2)15 (198.2214 m / z). We found that the b ions of 1021.5154 and 1035.5303 differ by one CH2(14 m / z), and (CH2)14(184.2058 m / z) and (CH2)15(198.2214 m / z) differ by one CH2(14 m / z). From the mass spectrometry data obtained and combined with literature reports, it was concluded that the antifungal compound was bacteriocin L containing Cnβ-amino fatty acids (n=14,15,16).

[0049] Step 14: Prepare lipopeptide solution After complete precipitation, centrifuge at 10000g for 5 minutes and discard the supernatant. Place the precipitate in an oven, weigh it after drying, dissolve it in methanol: chloroform 1:1, filter it, and collect the filtrate for later use. If needed, 5-50 mg of lipopeptide powder can be dissolved in 1 mL of methanol-chloroform solvent.

[0050] 2.2 Preparation of elastomer 2.2.1 Reagents used in preparing elastomers The main reagents used to prepare the elastomers are shown in Table 3.

[0051] Table 3 Main reagents for preparing elastomers *SYLGARD 184 silicone elastomer kit includes base and curing agent 2.2.2 Preparation method of elastomer Mix the main agent and curing agent in the SYLGARD 184 silicone elastomer kit in a ratio of 10:1, add about 1% of the total volume of TMSPMA, mix well, and wait for curing in a 60°C oven.

[0052] 2.3 Preparation of the elastomer layer Immerse the elastomer cured in step 2.2.2 in a 1:1 methanol:chloroform solution of the lipopeptide obtained in step 2.1.3, take it out after about 5 minutes, wipe the surface with filter paper and set aside.

[0053] 3. Preparation of composite antibacterial dressing By utilizing the adhesive property of the hydrogel, the hydrogel layer prepared in step 1.2 is adhered to the elastomer layer prepared in step 2.3 to obtain a composite antibacterial dressing.

[0054] In the present invention, the elastomer layer and the hydrogel layer are preferably laminated, or the hydrogel layer is embedded in the elastomer layer to obtain a composite antibacterial dressing with a stable structure and antibacterial effect. When the composite antibacterial dressing is used as a material for protecting bone cultural relics, the hydrogel layer is laminated to the cultural relics. The hydrogel layer has good water absorption and water retention. Taking the ancient ivory cultural relics of Sanxingdui as an example, the hydrogel layer can continuously provide moisture to the ancient ivory cultural relics to achieve moisturizing and antibacterial effects; at the same time, the outer elastomer layer can reduce water volatilization, and the elastomer layer impregnated with lipopeptides also has an inhibitory effect on Pseudomonas.

[0055] In this solution, considering the water retention performance for cultural relics, antibacterial effect and other factors, as well as cost and other factors, the elastomer layer preferably has a thickness of 0.1-1 mm; the hydrogel layer preferably has a thickness of 0.5-2 mm.

[0056] 4. Further investigate the antibacterial effect of the composite antibacterial dressing.

[0057] In the previous study, the bacteria and fungi were isolated, purified and identified by culture omics. Combined with the high-throughput sequencing results, the bacteria and fungi with relatively high abundance in the Sanxingdui ancient ivory were found, and their acid and enzyme production were measured. Finally, the key corrosion bacteria were determined to be Pseudomonas The key corrosion fungi are Aspergillus , Mortierella , Penicillium and Trichoderma This project carried out antibacterial experiments on Aspergillus, Trichoderma and Pseudomonas isolated from these genus.

[0058] 4.1 Materials required for determination of antibacterial effect 4.1.1 Strains Aspergillus, Trichoderma and Pseudomonas isolated from ancient ivory from Sanxingdui, Sichuan.

[0059] 4.1.2 Main reagents for determination of antibacterial effect The main reagents needed to determine the antibacterial effect include potato dextrose agar medium, LB medium, 0.9% saline, etc.

[0060] 4.1.3 Main instruments for determining antibacterial effect The main instruments needed to measure the antibacterial effect include high-pressure steam sterilizer, oven, clean workbench, constant temperature incubator, etc.

[0061] 4.2 Determination of antibacterial effect 4.2.1 Equipment preparation Prepare consumables such as EP tubes, coating rods, and pipette tips, and prepare potato dextrose agar (PDA) medium, LB medium, and 0.9% saline, and sterilize with high-pressure steam at 121°C for 20 min. After sterilization, pour the medium into plates in a clean bench, and dry the remaining consumables for later use.

[0062] 4.2.2 Quantification of tested strains Wash the test bacteria in the culture dish with physiological saline to obtain bacterial liquid. The spore-forming fungi need to be filtered with filter paper to remove the hyphae to obtain a spore suspension. Use a hemocytometer to measure the concentration of each bacterial liquid. The spore concentration of spore-forming fungi is measured, and the bacterial concentration of bacteria is measured. Fungi that do not produce spores do not need to be counted. Dilute each bacterial liquid with physiological saline to a concentration of 3×10 7 CFU (colony forming units) / mL, no dilution for non-spore-forming fungi.

[0063] 4.2.3 Determination of antibacterial effect This experiment uses the method of measuring the size of the inhibition zone to evaluate the antibacterial effect of the antibacterial material. The diluted bacterial solution is evenly spread on the culture medium at a rate of 100 μL per dish, where fungi are inoculated in PDA culture medium and bacteria are inoculated in LB culture medium. The material to be tested is cut into small square pieces with a side length of about 0.5 cm and placed on the culture medium coated with the bacterial solution. Three pieces are placed on each dish as three repeated experiments. After completion, the fungi are placed in a 27°C incubator for 3 days, and the bacteria are placed in a 37°C incubator for 2 days. After the culture is completed, the size of the inhibition zone is measured.

[0064] 4.3 Antibacterial experiment 4.3.1 Study on the antibacterial effect of hydrogel alone The hydrogel stock solution prepared according to the above method is light milky white after adding chlorhexidine digluconate mother solution, and is converted into a transparent gel block after UV curing. Figure 4 As shown, it is soft, tough, stretchable, and viscoelastic. It is not easy to break after repeated physical pulling. It has a certain resilience against deformation. It can easily adhere to the surface of various objects and is not easy to fall off. It has the ability to absorb and retain a large amount of water, and will swell to a certain extent after absorbing water.

[0065] In this experiment, the hydrogel stock solution was divided into 5 groups, and 4%, 2%, 1%, 0.5%, and 0% (blank group) of chlorhexidine digluconate stock solution were added to investigate their antibacterial effects. The results are shown in Table 4.

[0066] Table 4 As shown in Table 4, in the CK (blank) group without antibacterial agent, the hydrogel had no obvious antibacterial effect on the strains used. The antibacterial test results are shown in Figure 5 . The hydrogel of chlorhexidine digluconate mother solution has a better antibacterial effect on bacteria than on fungi. 4% and 2% chlorhexidine gluconate have excellent antibacterial effects on fungi and bacteria, and the antibacterial effects are similar, both greater than 1.8 cm and maintained at about 2 cm. Compared with 2% chlorhexidine gluconate, the antibacterial effects of 1% and 0.5% chlorhexidine gluconate on bacteria and fungi are less than 1.8 cm, and the antibacterial effect of 0.5% chlorhexidine gluconate on fungi is only maintained at about 1 cm. Therefore, it is preferred to use chlorhexidine gluconate with a concentration of 2% to obtain a hydrogel layer and prepare an antibacterial excipient with a lipopeptide elastomer layer.

[0067] The antibacterial activity of the hydrogel compounded with 2% chlorhexidine digluconate was determined by the cross measurement method. The antibacterial activity was evaluated by the diameter of the inhibition zone. The sizes of the inhibition zones formed by the material on the plates coated with Aspergillus, Trichoderma, and Pseudomonas were approximately 18.92 mm, 21.02 mm, and 23.81 mm, respectively, all showing high sensitivity. Figure 6 .

[0068] By comparing the blank control with the experimental group, it can be found that the antibacterial effect of the hydrogel mainly comes from chlorhexidine digluconate. According to the above preparation method, the hydrogel formula used contains SDS. SDS, as an anionic surfactant, can interfere with the normal function of bacterial cell membranes, thus having certain antibacterial properties. In order to further investigate whether the addition of SDS will increase the antibacterial effect of the system, this experiment added an experimental group of hydrogels after removing SDS, and found that there was no significant difference in the antibacterial effect of the two. It can be seen that the main role of SDS in the hydrogel is to enhance the interface effect, and it has little effect on the antibacterial effect.

[0069] In this scheme, the hydrogel prepared according to the above preparation method has good toughness and viscosity, and can be well adhered to the elastomer. However, if you want to further increase the viscosity of the hydrogel on this basis, you can add 1.14 g of acrylic acid to the formula, and the obtained hydrogel has better viscosity. However, the antibacterial effect on Aspergillus is weakened. One of the reasons is that the release of chlorhexidine digluconate is hindered to a certain extent, resulting in a decrease in antibacterial efficacy, but it still has a good antibacterial effect on several other strains of tested bacteria. The results are shown in Figure 7. If you want to reduce the viscosity of the hydrogel, you can replace the crosslinker MBA with TMSPMA on the basis of adding 1.14 g of acrylic acid, increase the amount of water to 20 mL, and increase the amount of photoinitiator to 0.01 g. The hydrogel prepared in this way is almost non-sticky and has a good antibacterial effect. The results are shown in Figure 8.

[0070] 4.3.2 Study on the antibacterial effect of lipopeptide elastomer alone The lipopeptide was prepared according to the above method. After drying, the lipopeptide was a brown solid. The solution after dissolving the solid in a methanol:chloroform 1:1 solution was brown, and the color of the solution gradually deepened with the increase of the lipopeptide concentration. Elastomers were prepared according to the preparation method of 2.2.2. The elastomers were divided into 4 groups, and lipopeptides of different concentrations as shown in Table 5 were immersed in the elastomers to examine the antibacterial effect of the lipopeptides. The blank group was a blank elastomer without the addition of the lipopeptide solution ( Fig. 9 ), which is a transparent elastic film with weaker stretchability than hydrogel.

[0071] Table 5 As shown in Table 5, the elastomer without antibacterial agent was used as blank control, and the elastomer obtained by compounding lipopeptide solution was used as experimental group. The results showed that the blank elastomer had no antibacterial effect. Fig.10 The elastomer of the compound lipopeptide solution has a certain antibacterial effect on Aspergillus, Trichoderma and Pseudomonas, see Fig.11 The antibacterial activity of the elastomer obtained from the composite lipopeptide solution was determined by the cross measurement method. The antibacterial activity was evaluated by the diameter of the inhibition zone. The diameter of the inhibition zone formed by the 50 mg / mL lipopeptide concentration material on the plate coated with Aspergillus, Trichoderma and Pseudomonas was approximately 15.03 mm, 16.31 mm, and 12.68 mm.

[0072] 4.3.3 Antibacterial effect of composite antibacterial dressing According to the contents of Items 4.3.1 and 4.3.2, we further selected a hydrogel containing 2% chlorhexidine digluconate and an elastomer containing 50 mg / mL lipopeptide solution to initially obtain a composite antibacterial dressing, and further investigated the performance of the composite antibacterial dressing.

[0073] The multilayer structure combines the characteristics of hydrogel and elastomer, that is, it has the water retention capacity of hydrogel, and the surface elastomer can also prevent the water in the hydrogel from diffusing into the air to prolong its moisturizing effect. The antibacterial effect of the double-layer structure (elastic side facing down) was measured.

[0074] The antibacterial activity of the composite material was determined, and its antibacterial activity was evaluated by the diameter of the inhibition zone. The sizes of the inhibition zones formed by the composite material on the plates coated with Aspergillus, Trichoderma, and Pseudomonas were approximately 8.52 mm, 15.08 mm, and 21.97 mm, respectively, showing medium to high sensitivity. The inhibition zone for Aspergillus was not obvious, but no colony grew on the contact part ( Fig.12 ), after the material was stored at room temperature for 1 month, the sizes of the inhibition zones formed by the material on the plates coated with Aspergillus, Trichoderma, and Pseudomonas were measured to be approximately 4.13 mm, 10.62 mm, and 11.02 mm, respectively. It can be seen that the antibacterial effect on Aspergillus was significantly reduced, and only the antibacterial effect on the contact surface could be achieved without obvious inhibition zones. However, it still showed medium and high sensitivity inhibition effects on several other ivory-corrosive bacteria, as shown in Figure 2. Fig.13 , Fig.13 This is a comparison of the antibacterial effect of the gel on Trichoderma before (left) and after (right) one month of storage.

[0075] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. Any simple modification or equivalent change made to the above embodiment based on the technical essence of the present invention shall fall within the protection scope of the present invention.

Claims

1. A composite antibacterial dressing, characterized in that: It includes an elastic body layer and a hydrogel layer. The hydrogel layer is attached to or embedded in the elastic body layer. The hydrogel layer is used to be attached to the cultural relic. The hydrogel layer is a hydrogel containing chlorhexidine digluconate; The elastic body layer is a polymer elastic body containing lipopeptide, and the lipopeptide solution can inhibit the growth of pseudomonas.

2. A composite antibacterial dressing according to claim 1, characterized in that: The concentration of chlorhexidine digluconate in the hydrogel layer is 0.5-4%; the concentration of lipopeptide in the elastomer layer is 12.5-50 mg / mL.

3. The composite antibacterial dressing according to claim 1, characterized in that: The thickness of the hydrogel layer is 0.5-2 mm; the thickness of the elastomer layer is 0.1-1 mm.

4. The method for preparing the composite antibacterial dressing according to claim 2, characterized in that: The following steps are involved: S1. A 0.5-4% chlorhexidine digluconate solution is prepared, AM, SDS, MBA, glycerol, deionized water and a photoinitiator are placed in a container, and the chlorhexidine digluconate solution is added to the container and stirred to obtain a 0.5~4% mass volume concentration of chlorhexidine digluconate hydrogel; S2. Inoculating and fermenting Bacillus amyloliquefaciens M73 on Landy medium, isolating lipopeptides that inhibit Pseudomonas, dissolving the lipopeptides in a methanol-chloroform solvent to obtain a lipopeptide solution, and immersing a silicone elastomer in the lipopeptide solution. After 3 to 10 minutes, an elastomer layer having a lipopeptide concentration of 12.5 to 50 mg / mL is obtained; S3. The hydrogel layer prepared in step S1 is then bonded to the elastomer layer prepared in step S2, or the hydrogel layer prepared in step S1 is embedded in the elastomer layer prepared in step S2 to obtain a composite antibacterial dressing.

5. The method for preparing the composite antibacterial dressing according to claim 4, characterized in that: In step S1, the method for preparing the hydrogel comprises the following steps: a1. Take raw materials AM 23.06%, SDS 0.03%, MBA 7.69%, glycerol 15.38%, deionized water 53.82% and photoinitiator 2959 0.02% by mass percentage, and then place the above raw materials in a container; b1. Use a stirrer to stir the material in the container. When the solid is completely dissolved, use a vacuum pump to extract the dissolved gas in the solution; c1 After the gas was exhausted, chlorhexidine digluconate mother solution was added thereto to obtain chlorhexidine digluconate hydrogel; d1. After stirring evenly, pour the solution into the mold and let it stand under the UV lamp for 0.4~1h. After it is formed, demould it for later use.

6. The method for preparing the composite antibacterial dressing according to claim 4, characterized in that: In step S2, the method for preparing lipopeptide comprises the following steps: a2. Preparation of culture medium: Prepare Landy medium and sterilize; b2. Inoculation and fermentation: Inoculate 1×10 5 CFU / mL of Bacillus amyloliquefaciens M73 culture was fermented in a shaker at 30°C and 2000 rpm for 3 days; c2. Lipopeptide extraction: After fermentation, centrifuge at 8000rpm for 5min, collect the supernatant, adjust the pH to 2 with hydrochloric acid, and let stand until the lipopeptide precipitates; d2. Centrifuge again, discard the supernatant, dry the precipitate to obtain lipopeptide powder, then dissolve the lipopeptide powder with methanol-chloroform solvent, filter and take the filtrate to obtain a lipopeptide complex solution.

7. The method for preparing the composite antibacterial dressing according to claim 6, characterized in that: In step c2, the lipopeptides are separated by high performance liquid chromatography and analyzed by electrospray mass spectrometry, and the high performance liquid chromatography conditions meet the following requirements: Chromatographic column: Octadecylsilane bonded silica gel is used as filler; Column temperature: 25°C; Flow rate: 4.0ml / min; Injection volume: 1 μL; Detection wavelength: 214nm; Mobile phase A was acetonitrile, mobile phase B was 0.1% formic acid solution, gradient elution; The elution process is: 0-50min, mobile phase A is 2-95%, mobile phase B is 98-5%; Mass spectrometer detection conditions: electrospray voltage 5000 V, capillary injection port voltage 60 V, capillary heating temperature 320 °C, acceleration voltage 6000 V, reflector voltage 1210 V, repulsion pulse frequency 5000 Hz, mass detection range 0.5-2.5 kDa.

8. The method for preparing the composite antibacterial dressing according to claim 6, characterized in that: In step d2, the methanol-chloroform solvent is a mixed solution of methanol:chloroform in a volume ratio of 1:1, and 12.5-50 mg of lipopeptide powder is dissolved in each milliliter of methanol-chloroform solvent.

9. The method for preparing the composite antibacterial dressing according to claim 4, characterized in that: In step S2, the preparation method of the silicone elastomer is as follows: the main agent and the curing agent in the SYLGARD 184 silicone elastomer kit are mixed in a ratio of 10-20:1, 0.5-2% of the total volume of TMSPMA is added, the mixture is mixed evenly, and then cured in an oven at 60° C. to obtain the silicone elastomer.

10. Use of the composite antibacterial dressing according to any one of claims 1 to 9 as a protective material for inhibiting Aspergillus, Trichoderma and Pseudomonas in bone cultural relics.

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