A process for achieving controlled sustained-release antibacterial biological patch with high drug loading

Abstract

The present invention relates to the field of medical materials, and more specifically, to a process for producing a high-drug-loaded antibacterial biological patch with controlled sustained release. Unlike conventional coating structures, the effective antibacterial components of the antibacterial patch of the present invention are distributed throughout the patch structure network, rather than being confined to a single layer. This not only ensures that the resulting patch possesses excellent mechanical properties and a structure that resists loosening upon contact with water, but also enhances process stability. Furthermore, the drug loading amount and loading speed can be quantitatively and controllably adjusted according to actual needs. This method can be applied to various surgical scenarios, such as open abdominal surgery, and is particularly suitable for immediate antibacterial use after surgical procedures. It not only facilitates anti-infection, but also guides tissue regeneration and degrades as tissue repair progresses.

Description

Technical Field

[0001] The present invention relates to the field of medical materials, and in particular to a process for realizing a high-drug-loaded antibacterial biological patch with controlled sustained release. Background Art

[0002] As an implantable medical device, patches face significant infection risks during use. Antimicrobial functionalization of patches has become a research focus, with methods typically including coating and metal ion deposition. Patent CN118001456A provides a novel method for preparing an anti-adhesion abdominal wall patch. This involves degreasing and sensitizing a polypropylene patch, then electroplating nanosilver in an ion sputtering apparatus to deposit antimicrobial silver ions on the patch. However, this method is generally not suitable for biological patches. Patent No. US8911765B2 discloses a biodegradable polymer covering for breast implants. The document describes immersing a synthetic polymer patch in a methanol / tetrahydrofuran organic solvent containing 0.0055g / mL of rifampicin and 0.0055g / mL of minocycline. After continuous immersion, the patch is oven-dried to obtain an antimicrobial patch. In this method, an adherent drug coating is formed on the polymer surface by immersion and oven drying. Due to the difference in the properties of the synthetic polymer patch and the biological patch, under the same conditions, the drug loading of the biological patch is much lower than that of the synthetic polymer patch. In clinical practice, the biological patch formed by this coating method usually shows a low drug loading, resulting in the inability to achieve antibacterial treatment. The problem of therapeutic effect; Patent No. CN118453962A provides a method for preparing a decellularized extracellular matrix patch with antibacterial function, wherein the decellularized small intestinal submucosal tissue is immersed in a NaClO solution for 12 hours to obtain an N-Cl antibacterial coating, which is freeze-dried and hot-pressed at 60°C to prepare an antibacterial patch; although this method superimposes the antibacterial coating, the drug loading capacity is still greatly limited, and the document does not record the chlorine loading capacity of the final product. In addition, the addition of a large amount of antibacterial chlorine is particularly cautious for medical devices, especially for patches of Class III implantable medical devices. It is clearly stipulated that chlorine, chlorine dioxide, hypochlorous acid, etc. may have adverse effects on the human body, and their addition requires strict approval. Summary of the Invention

[0003] To overcome the above problems, a non-coated patch with antimicrobial components distributed throughout the patch's internal network and surface is obtained. The present invention provides a process for achieving a high-drug-loaded antimicrobial biological patch with controlled sustained release. The process is as follows: a base layer, an intermediate layer, and a top layer are sequentially laid with a medium from bottom to top, and then laminated to obtain the patch.

[0004] The medium is selected from one or more of water, antibiotic solution X, and antibiotic Y solution.

[0005] The middle layer includes at least one strip insertion layer; the strip insertion layer is formed by arranging bio-based strips at intervals; and the intervals between the bio-based strips are 0.7-2.2 cm.

[0006] Furthermore, laying the base layer with the medium specifically comprises: wetting the substrate with the medium, laying the base layer on the substrate, flattening the base layer on the substrate, and then scraping off excess medium on the surface of the base layer with a scraper.

[0007] The purpose of wetting the substrate is to remove the air between the base layer and the substrate and to utilize the adhesion between the wetting medium and the base layer to make the base layer flatter on the substrate.

[0008] The material of the substrate is not limited. The substrate only serves to provide a horizontal operating surface for the base layer. It can be a commercially available glass substrate, steel plate, copper plate, etc.

[0009] Similar to laying the base layer with a medium, laying the middle layer with a medium is specifically to moisten the base layer with the medium, lay the middle layer on the base layer, flatten the middle layer on the base layer, and then use a scraper to scrape off excess medium on the surface of the middle layer.

[0010] Similar to laying the base layer with a medium, laying the top layer with a medium is to wet the top layer with the medium, lay the top layer on the middle layer, flatten the top layer on the middle layer, and then use a scraper to scrape off excess medium on the surface of the top layer.

[0011] Preferably, the strips are interleaved in layers, with each bio-based strip being arranged in parallel.

[0012] Preferably, the strip insertion layer is formed by bio-based strips arranged at intervals; the intervals between the bio-based strips are 0.7-2 cm.

[0013] Further preferably, the strip insertion layer is formed by bio-based strips arranged at intervals; the intervals between the bio-based strips are 0.9-1.5 cm.

[0014] More preferably, the strip insertion layer is formed by bio-based strips arranged at intervals; the intervals between the bio-based strips are 1-1.5 cm.

[0015] To achieve an antibacterial patch in which the antimicrobial components are distributed throughout the patch structure network, rather than just in a single layer, the inventors, through in-depth creative thinking, employed a medium (selected from one or more of water, antibiotic solution X, and antibiotic Y solution) to form the base, middle, and top layers, replacing the conventional lamination process (e.g., the simple stacking of dry film layers followed by pressing as described in patents CN105102009B or CN115054743B). This not only ensures that the resulting patch possesses excellent mechanical properties and a structure that resists loosening upon contact with water, but also effectively achieves the slow release of tetracycline from antibiotic solution X and antibiotic Y from antibiotic solution Y while maintaining a high drug loading, and further allows for the controlled and regulated release rates of tetracycline and antibiotic Y. In the present invention, the further controlled spacing of the bio-based strips makes this sustained-release effect particularly pronounced.

[0016] The angle between the length direction of the bio-based strip and the length direction of the base layer is 0-90°.

[0017] Preferably, the angle between the length direction of the bio-based strip and the length direction of the base layer is 15-75°.

[0018] Further preferably, the angle between the length direction of the bio-based strip and the length direction of the base layer is 30-60°.

[0019] More preferably, the angle between the length direction of the bio-based strip and the length direction of the base layer is 35-45°.

[0020] On the basis of the interval of the bio-based strips being 0.7-2.2 cm, the angle between the length direction of the bio-based strips and the length direction of the substrate layer can be further adjusted. The angle between the length direction of the bio-based strips and the length direction of the substrate layer is 15-75°, which to a certain extent improves the problem that tetracycline is unstable in contact with water and is prone to diastereoisomerization, resulting in a decrease in the activity of tetracycline in the antibiotic solution X and a decrease in the therapeutic effect.

[0021] Preferably, the process for achieving a high drug-loaded antibacterial biological patch with controlled sustained release is as follows: a base layer, an intermediate layer and a top layer are laid in sequence with a medium from bottom to top, and then laminated and pressed at 35-55° C. to obtain the patch.

[0022] More preferably, the base layer, the middle layer and the top layer are laid in sequence from bottom to top with a medium, and then laminated and pressed at 39-50° C. to obtain the product.

[0023] More preferably, the base layer, the middle layer and the top layer are laid in sequence from bottom to top with a medium, and then laminated and pressed at 39-48° C. to obtain the product.

[0024] As a preferred embodiment, the process for achieving a high-drug-loaded antibacterial biopatch with controlled sustained release is as follows:

[0025] S1. Decellularizing matrix material A with water to obtain a basal layer;

[0026] S2. The bio-based strip is placed on the substrate to form a strip insert layer covering the substrate layer, ie, the intermediate layer of the controlled-release high-drug-loaded antibacterial biopatch;

[0027] S3. Laying the acellular matrix material A on the strip insert layer with water to form the top layer of the antibacterial biological patch with high drug loading and controlled sustained release;

[0028] S4. Lamination.

[0029] As a preferred embodiment, the process for achieving a high-drug-loaded antibacterial biopatch with controlled sustained release is as follows:

[0030] S1. Laying the decellularized matrix material A with water to obtain a base layer; the middle layer includes two strip insert layers (strip insert layer 1 and strip insert layer 2), and the middle layer is obtained by the following steps S2-S3;

[0031] S2. The bio-based strip is placed on the substrate to form a strip insert layer 1 covering the substrate layer;

[0032] S3. The bio-based strip is placed on the strip insert layer 1 to form a strip insert layer 2;

[0033] S4. Laying the acellular matrix material A on the middle layer with water to form the top layer of the antibacterial biological patch with high drug loading and controlled sustained release;

[0034] S5. Lamination.

[0035] As a preferred embodiment, the process for achieving a high-drug-loaded antibacterial biopatch with controlled sustained release is as follows:

[0036] S1. Laying the decellularized matrix material A with water to obtain a base layer; the middle layer is obtained by the following steps S2-S3;

[0037] S2. Use antibiotic solution X to lay acellular matrix material B on the base layer to form a sandwich 1 covering the base layer. Repeat this operation until a sandwich is formed. n ; n is a positive integer, and 1≤n≤18;

[0038] S3. Place the bio-based strips in the sandwich n On, to form a strip insertion layer 1, repeat this operation until a strip insertion layer is formed m ; m is a positive integer, and 1≤m≤18;

[0039] S4. Laying the acellular matrix material A on the strip insert layer with water to form the top layer of the antibacterial biological patch with high drug loading and controlled sustained release;

[0040] S5. Laminate using a laminator.

[0041] As a preferred embodiment, the process for achieving a high-drug-loaded antibacterial biopatch with controlled sustained release is as follows:

[0042] S1. Laying the acellular matrix material A with water to obtain a basal layer;

[0043] S2. Use antibiotic solution X to lay acellular matrix material B on the base layer to form a sandwich 1 covering the base layer. Repeat this operation until a sandwich is formed. n ; n is a positive integer, and 1≤n≤18;

[0044] S3. Place the bio-based strips in the sandwich n On, to form a strip insertion layer 1, repeat this operation until a strip insertion layer is formed m ; m is a positive integer, and 1≤m≤18;

[0045] Preferably, 1≤m≤7.

[0046] Interlayer 1 formed after steps S2-S3 n and stripe insertion layer m The combination of is a unit module; the unit module is repeatedly stacked p times to obtain the middle layer; p is a positive integer, and 1≤p≤18;

[0047] Preferably, m+n≤20.

[0048] Furthermore, m+n≤18.

[0049] Preferably, p is a positive integer, and 1≤p≤16.

[0050] Preferably, (m+n)×p≤35. That is, the total number of intermediate layers is controlled within 35 layers.

[0051] Furthermore, (m+n)×p≤22. That is, the total number of intermediate layers is controlled within 22 layers.

[0052] Furthermore, (m+n)×p≤10.

[0053] Furthermore, (m+n)×p≤9.

[0054] S4. Laying the acellular matrix material A on the middle layer with water to form the top layer of the antibacterial biological patch with high drug loading and controlled sustained release;

[0055] S5. Laminate using a laminator.

[0056] In the present invention, the base layer and the interlayer n and the top layer are called continuous plane layers; stripe insertion layer 1, stripe insertion layer 2 and stripe insertion layer m They are all called discontinuous surface layers.

[0057] In the technical solution of the present invention, it is necessary to ensure that both sides of the discontinuous surface layer are in contact with the continuous planar layer.

[0058] Preferably, 60% to 100% of each planar area of ​​each non-continuous facing layer is in contact with a continuous planar layer.

[0059] The arrangement of both sides of the discontinuous surface layer in contact with the continuous planar layer not only improves interlayer bonding, preventing the final pressed product from loosening, but also helps increase the drug loading of tetracycline in antibiotic solution X and antibiotic Y in antibiotic solution Y, and further promotes the distribution of tetracycline in antibiotic solution X and antibiotic Y in antibiotic solution Y throughout the final patch structure network (rather than the distribution trend of the drug being distributed only in a single layer). In addition, the inventors unexpectedly discovered that the contact between both sides of the discontinuous surface layer and the continuous planar layer unexpectedly reduces the drug loss of tetracycline and antibiotic Y caused by subsequent lamination, reducing the drug loss to as low as 15% by weight, and controlling the drug loss within the range of 15-50% by weight, thereby significantly reducing drug waste and lowering production costs. The inventors speculate that this may be because the basal layer, middle layer and top layer of the decellularized matrix material retain the complete triple helix structure of the collagen fibers. In addition to being able to partially deposit in the porous pores of each layer, some drug crystal nuclei can also be embedded in the spiral grooves on the triple helix chains. The setting of the discontinuous surface layer of the specific strip insertion layer has a more significant effect on this deposition adsorption and embedding adsorption, accompanied by the unique toughness between the fibers formed between the structures at this time, thereby increasing the binding force.

[0060] Acellular matrix material A and acellular matrix material B were purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd. and were prepared from the submucosal layer or full layer of hollow organs (intestine, stomach, bladder) of mammals (human, pig, cattle, fetal bovine, horse, sheep, dog) according to the perfusion-pressure differential method disclosed in patent CN106075583B.

[0061] Preferably, in step S1 and step S6, the decellularized matrix material A is derived from the porcine bladder submucosa and is purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.; that is, the decellularized matrix material A is prepared from the porcine bladder submucosa according to the perfusion-pressure differential method disclosed in patent CN106075583B.

[0062] Preferably, in steps S2-S5, the decellularized matrix material B is derived from the porcine small intestinal submucosa and is purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.; that is, the decellularized matrix material B is prepared from the porcine small intestinal submucosa according to the perfusion-pressure differential method disclosed in patent CN106075583B.

[0063] As a preferred embodiment, the antibiotic liquid X is a suspension of a tetracycline metal chelate, which is obtained by adjusting the pH of an aqueous solution of a tetracycline / metal ion mixture.

[0064] As a preferred embodiment, the pH is 7.2-8.1.

[0065] Preferably, the pH is 7.2-7.9.

[0066] More preferably, the pH is 7.2-7.7.

[0067] More preferably, the pH is 7.4.

[0068] As a preferred embodiment, in the aqueous solution of the tetracycline / metal ion mixture, the molar ratio of metal ions to tetracycline is (1.2-1.9):1 or (3-6.5):1.

[0069] Preferably, in the aqueous solution of the tetracycline / metal ion mixture, the molar ratio of metal ions to tetracycline is (1.5-1.8):1 or (3.2-4.3):1.

[0070] Preferably, the tetracycline is selected from any one of minocycline, minocycline hydrochloride, doxycycline hydrochloride and chlortetracycline.

[0071] More preferably, the tetracycline is minocycline hydrochloride.

[0072] Preferably, the metal ions are selected from one or more of calcium ions, magnesium ions, aluminum ions, and zinc ions.

[0073] More preferably, the metal ion is a calcium ion.

[0074] As a preferred embodiment, the preparation method of the bio-based strip is: laying the acellular matrix material C with the antibiotic Y solution, repeating this operation 1-10 times, laminating, and cutting into rectangular strips with a width of 1-20 mm.

[0075] Acellular matrix material C was purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd. and was prepared from the submucosal layer or full layer of hollow organs (intestine, stomach, bladder) of mammals (human, pig, cattle, fetal bovine, horse, sheep, dog) according to the perfusion-pressure differential method disclosed in patent CN106075583B.

[0076] Preferably, the acellular matrix material C is purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd. and is prepared from the submucosal layer of the intestine and stomach of mammals (human, pig, cattle, fetal bovine, horse, sheep, dog) according to the perfusion-pressure difference method disclosed in patent CN106075583B.

[0077] Further preferably, the preparation method of the bio-based strip is: laying the decellularized matrix material C with the antibiotic Y solution to flatten the decellularized matrix material C, repeating this operation 3-7 times, laminating using a laminator, and cutting into rectangular strips with a width of 1-20 mm.

[0078] More preferably, the preparation method of the bio-based strip is as follows: laying the decellularized matrix material C with the antibiotic Y solution, flattening the decellularized matrix material C on a horizontal plane, repeating this operation 4-5 times, laminating using a laminator, and cutting into rectangular strips with a width of 1-20 mm.

[0079] Preferably, the width of the rectangular strip is 1-15 mm.

[0080] More preferably, the width of the rectangular strip is 1-8 mm.

[0081] Further preferably, the width of the rectangular strip is 2-8 mm.

[0082] For example, the width of the rectangular strip is 2 mm, 3 mm, 4 mm, 5 mm, 6 mm, 7 mm, 8 mm, etc.

[0083] In the present invention, the number of bio-based strips is not particularly limited, and they can be laid at intervals of 0.7-2.2 cm.

[0084] As a preferred embodiment, the antibiotic Y is a rifamycin antibiotic.

[0085] Preferably, the rifamycin antibiotic is selected from one or more of rifampicin, rifapentine, rifamycin sodium, and rifabutin.

[0086] More preferably, the rifamycin antibiotic is rifampicin.

[0087] As a preferred embodiment, the preparation method of the antibiotic Y solution is: adding antibiotic Y to 85wt%-95wt% ethanol aqueous solution to make the concentration of antibiotic Y in the antibiotic Y solution be 0.01-0.07g / mL.

[0088] Preferably, the preparation method of the antibiotic Y solution is: adding antibiotic Y to a 90wt%-95wt% ethanol aqueous solution to make the concentration of antibiotic Y in the antibiotic Y solution be 0.01-0.07g / mL.

[0089] Further preferably, the preparation method of the antibiotic Y solution is: adding antibiotic Y to a 90 wt % ethanol aqueous solution so that the concentration of antibiotic Y in the antibiotic Y solution is 0.01-0.07 g / mL.

[0090] Preferably, the concentration of antibiotic Y in the antibiotic Y solution is 0.01-0.04 g / mL.

[0091] Generally speaking, due to the inherent characteristics of synthetic polymer substrates, such as slow degradation, the drug release rate from drug-loaded biological substrates is typically much faster than that from drug-loaded synthetic polymer substrates, provided all other conditions are the same, including the substrate type, the drug loading and the release environment. However, if the drug is released too quickly, it will not exert a sustained antibacterial effect and may not meet the needs of clinical antibacterial therapy. This necessitates controlling the drug release rate from drug-loaded biological substrates. This is particularly true for antibiotic Y, a rifamycin antibiotic (specifically rifampicin) in the present invention, which is highly hydrophilic. Once applied to the patch, it dissolves rapidly in water and is released, further accelerating the drug release rate. Therefore, controlling the drug release rate from drug-loaded biological substrates is crucial. The inventors of the present invention have overcome the technical resistance to the aforementioned rapid release of drugs from the biological matrix by ensuring that the intermediate layer includes at least one strip insertion layer (i.e., the antimicrobial biological patch includes at least one discontinuous surface layer) and that the spacing and angle of the bio-based strips are within specific ranges. This overcoming is particularly significant when the antibiotic Y is a rifamycin antibiotic and the antibiotic solution X is a suspension of a tetracycline metal chelate. Furthermore, by controlling the number of strip insertion layers, the width of the strip insertion layers, and the width range of the bio-based strips, the process stability of the sustained-release effect can be enhanced and the sustained-release rate can be adjusted according to actual needs.

[0092] The antibacterial patch obtained by the process of producing an antibacterial biological patch with controlled sustained release and high drug loading according to the present invention can be provided with uniform small holes with a hole diameter of 0.1-3.0 mm and a distance between two adjacent holes of 4-20 mm.

[0093] Beneficial Effects: The high infection rate after surgical procedures typically occurs within the first three days. The process of this invention achieves high drug loading and controlled sustained release in antimicrobial biopatch. Unlike conventional coating structures, the effective antimicrobial components of the antimicrobial patch are distributed throughout the patch structure, rather than confined to a single layer. This process not only ensures the resulting patch possesses excellent mechanical properties and a structure that resists loosening upon contact with water, but also enhances process stability. Furthermore, the drug loading amount and loading rate can be quantitatively and controllably adjusted according to actual needs. This allows for application in a variety of surgical scenarios, including open abdominal surgery, and is particularly suitable for immediate antimicrobial use after surgical procedures. Not only does it aid in infection prevention, but the biopatch also guides tissue regeneration and degrades as tissue repair progresses. BRIEF DESCRIPTION OF THE DRAWINGS

[0094] Figure 1 This is a schematic structural diagram of the antibacterial biological patch with high drug loading and controlled sustained release described in Example 2; Figure 1 In the figure, 1 is the base layer, 2 is the strip insert layer 1, 3 is the strip insert layer 2, 4 is the middle layer, and 5 is the top layer. (The layers are staggered in the figure only to clearly show the structure of each layer. In the actual product, the edges of each quadrilateral are aligned.)

[0095] Figure 2 This is a schematic structural diagram of the antibacterial biological patch with high drug loading and controlled sustained release described in Example 3; Figure 2 Middle, 8-intermediate layer', 6-strip insertion layer 1’ , 7-strip insertion layer 2’ (The staggered layers in the diagram are only for the purpose of clearly showing the structure of each layer. In the actual product, the edges of each quadrilateral are aligned.)

[0096] Figure 3 (a) is a schematic cross-sectional view of the antibacterial biological patch with controlled sustained release and high drug loading as described in Example 4; in the figure, 9 is the middle layer;

[0097] Figure 3 (b) is a schematic cross-sectional view of the antibacterial biological patch with controlled sustained release and high drug loading as described in Example 5; in the figure, 11 is a unit module, 10 is an intermediate layer;

[0098] Figure 4 This is a schematic diagram of the structure of the high-drug-loaded, controlled-release antibacterial biopatch described in Comparative Example 4. (The staggered layers in the diagram are only for the purpose of clearly illustrating each layer; in the actual product, the edges of each quadrilateral are aligned.)

[0099] Figure 5 This is a SEM scanning electron microscope image of the antibacterial biological patch obtained in Example 4, wherein: Figure 5 (a) is a scanning electron microscope image of the upper surface of the patch. Figure 5(b) is a scanning electron microscope image of the lower surface of the patch. Figure 5 (c) and Figure 5 (d) is a scanning electron microscope image of a cross section on the patch. DETAILED DESCRIPTION

[0100] All materials used in the examples are commercially available.

[0101] Example 1

[0102] This example provides a process for achieving controlled sustained release of a high-drug-loaded antibacterial biopatch. The process is as follows:

[0103] S1. Use 6 ml of water to lay the acellular matrix material A (derived from the submucosa of porcine bladder, purchased from Zhuoran Medical Technology (Suzhou) Co., Ltd.) so that the basal layer 实施例1 Flattened on a horizontal plane to obtain the base layer; the middle layer 实施例1 Obtained by the following steps S2-S5; the intermediate layer 实施例1 The strip insertion layer comprises a strip insertion layer; the strip insertion layer is formed by bio-based strips arranged at intervals;

[0104] S2. Place the bio-based strips on the substrate 实施例1 The angle between the length direction of the bio-based strip and the length direction of the base layer is 45 degrees, the interval between the bio-based strips is 1.5 cm, and each bio-based strip is arranged in parallel to form a strip insertion layer covering the base layer, that is, the middle layer of the antibacterial biological patch with controlled sustained release and high drug loading is obtained. 实施例1 .

[0105] The preparation method of the bio-based strip is as follows: 6 mL of antibiotic Y solution is used to lay the decellularized matrix material C (derived from the submucosa of porcine small intestine, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.), flatten the decellularized matrix material C, repeat this operation 4 times, laminate it using a laminator, and cut it into rectangular strips with a width of 3 mm.

[0106] The antibiotic Y is a rifamycin antibiotic, and the rifamycin antibiotic is rifampicin.

[0107] The preparation method of the antibiotic Y solution is as follows: adding the antibiotic Y to a 90 wt % ethanol aqueous solution so that the concentration of the antibiotic Y in the antibiotic Y solution is 0.01 g / mL.

[0108] S3. Use 6mL of water in the middle layer 实施例1 The acellular matrix material A (derived from the submucosa of the porcine bladder, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.) was laid on the top layer and flattened to form the top layer of the antibacterial biological patch with high drug loading and controlled sustained release. 实施例1 ;

[0109] S4.Laminate at 45℃ for 12h to obtain the product.

[0110] Example 2

[0111] This example provides a process for achieving controlled sustained release of a high-drug-loaded antibacterial biopatch. The process is as follows:

[0112] S1. 6 ml of water was used to lay down acellular matrix material A (derived from porcine bladder submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.), flattening the basal layer to obtain basal layer 1. Intermediate layer 4 was obtained by the following steps S2-S5. Intermediate layer 4 comprised two strip inserts, obtained by the following steps S2-S3. The strip inserts were formed by spaced bio-based strips.

[0113] S2. Place the bio-based strips on the base layer, with the angle between the length direction of the bio-based strips and the length direction of the base layer being 45°, the interval between the bio-based strips being 1.5 cm, and each bio-based strip being arranged in parallel to form a strip insertion layer 1 2 covering the base layer.

[0114] The preparation method of the bio-based strip is as follows: 6 mL of antibiotic Y solution is used to lay the decellularized matrix material C (derived from the submucosa of porcine small intestine, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.), flatten the decellularized matrix material C, repeat this operation 4 times, laminate it using a laminator, and cut it into rectangular strips with a width of 3 mm.

[0115] The antibiotic Y is a rifamycin antibiotic, and the rifamycin antibiotic is rifampicin.

[0116] The preparation method of the antibiotic Y solution is as follows: adding the antibiotic Y to a 90 wt % ethanol aqueous solution so that the concentration of the antibiotic Y in the antibiotic Y solution is 0.01 g / mL.

[0117] S3. Continue placing the bio-based strips on the strip insert layer 1, the angle between the length direction of the bio-based strips and the length direction of the substrate is 45°, the interval between the bio-based strips is 1.5 cm, and each bio-based strip is arranged in parallel to form strip insert layers 2 3;

[0118] The angle between the length direction of the bio-based strips of strip insertion layer 1 and the length direction of the bio-based strips of strip insertion layer 2 is 90°.

[0119] S4. Lay the acellular matrix material A (derived from porcine bladder submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.) on the middle layer 4 with 6 mL of water and flatten it to form the top layer 5 of the antibacterial biopatch with controlled sustained release and high drug loading;

[0120] S5. Lamination at 45℃ for 12h is completed. The structure of the obtained antibacterial biopatch with high drug loading and controlled sustained release is as follows: Figure 1 .

[0121] Example 3

[0122] This example provides a process for achieving a controlled sustained release antibacterial biopatch with a high drug loading. The difference from Example 2 is that the middle layer '8 includes two strip insertion layers. 1’ 6. The length direction of the bio-based strips and the strip insertion layer 2’ The angle between the length directions of the bio-based strips of 7 is 0°; the strips are inserted into the layer 2’ The bio-based strip of 7 falls exactly on the strip insertion layer 1’ The structure of the obtained antibacterial biopatch with high drug loading and controlled sustained release is as follows: Figure 2 .

[0123] Example 4

[0124] This example provides a process for achieving controlled sustained release of a high-drug-loaded antibacterial biopatch. The process is as follows:

[0125] S1. Lay the acellular matrix material A (derived from the submucosa of porcine bladder, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.) with 6 mL of water to flatten the basal layer to obtain the basal layer; the middle layer '' 9 is obtained by the following steps S2-S3;

[0126] S2. Use 6 mL of antibiotic solution X to lay acellular matrix material B (derived from porcine small intestinal submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.) on the basal layer and flatten it to form interlayer 1 covering the basal layer. Repeat this operation until interlayer 2 is formed; n = 2;

[0127] The antibiotic liquid X is a suspension of a tetracycline metal chelate, which is obtained by adjusting the pH of an aqueous solution of a tetracycline / metal ion mixture to 7.4 with an aqueous sodium hydroxide solution.

[0128] In the aqueous solution of the tetracycline / metal ion mixture, the molar ratio of the metal ion to the tetracycline is 1.7:1.

[0129] The tetracycline is minocycline hydrochloride, and the metal ion is calcium ion.

[0130] The preparation method of antibiotic solution X is as follows: 0.1 g of minocycline hydrochloride is added to 10 mL of water, mixed, and then a 0.0045 mol / mL calcium chloride aqueous solution is added to adjust the molar ratio of calcium ions to minocycline hydrochloride in the antibiotic solution X to 1.7:1.

[0131] S3. The bio-based strips are placed on the interlayer 2, with the angle between the length direction of the bio-based strips and the length direction of the substrate being 45°, and the interval between the bio-based strips being 1.5 cm; to form a strip insertion layer covering the interlayer 2; in the strip insertion layer, each bio-based strip is arranged in parallel;

[0132] The preparation method of the bio-based strip is as follows: 6 mL of antibiotic Y solution is used to lay the decellularized matrix material C (derived from the submucosa of porcine small intestine, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.), flatten the decellularized matrix material C, repeat this operation 4 times, laminate it using a laminator, and cut it into rectangular strips with a width of 3 mm.

[0133] The antibiotic Y is a rifamycin antibiotic, and the rifamycin antibiotic is rifampicin.

[0134] The preparation method of the antibiotic Y solution is as follows: adding the antibiotic Y to a 90 wt % ethanol aqueous solution so that the concentration of the antibiotic Y in the antibiotic Y solution is 0.01 g / mL.

[0135] S4. Using 6 mL of water, acellular matrix material A (derived from porcine bladder submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.) was laid on the middle layer '' 9 and flattened to form the top layer of the antibacterial biological patch with controlled sustained release and high drug loading;

[0136] S5. Use a laminator to laminate at 45°C for 12 hours.

[0137] The cross-sectional structure of the obtained antibacterial biological patch with high drug loading and controlled sustained release is as follows: Figure 3 (a).

[0138] Example 5

[0139] This example provides a process for achieving controlled sustained release of a high-drug-loaded antibacterial biopatch. The process is as follows:

[0140] S1. Lay the acellular matrix material A (derived from the submucosa of porcine bladder, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.) with 6 mL of water to flatten the basal layer to obtain the basal layer;

[0141] S2. Use 6 mL of antibiotic solution X to lay acellular matrix material B (derived from porcine small intestinal submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.) on the basal layer and flatten it to form interlayer 1 covering the basal layer. Repeat this operation until interlayer 2 is formed; n = 2;

[0142] The antibiotic liquid X is a suspension of a tetracycline metal chelate, which is obtained by adjusting the pH of an aqueous solution of a tetracycline / metal ion mixture to 7.6 with an aqueous sodium hydroxide solution.

[0143] In the aqueous solution of the tetracycline / metal ion mixture, the molar ratio of the metal ion to the tetracycline is 1.6:1.

[0144] The tetracycline is minocycline hydrochloride, and the metal ion is calcium ion.

[0145] The preparation method of antibiotic solution X is as follows: 0.1 g of minocycline hydrochloride is added to 10 mL of water, mixed, and then a 0.0045 mol / mL calcium chloride aqueous solution is added to adjust the molar ratio of calcium ions to minocycline hydrochloride in the antibiotic solution X to 1.6:1.

[0146] S3. Place the bio-based strips on interlayer 2, with the angle between the length direction of the bio-based strips and the length direction of the base layer being 45°, and the interval between the bio-based strips being 1.5 cm; to form a strip insertion layer covering interlayer 2; in the strip insertion layer, each bio-based strip is arranged in parallel; m = 1.

[0147] The preparation method of the bio-based strip is as follows: 7 mL of antibiotic Y solution is used to lay the decellularized matrix material C (derived from the submucosa of porcine small intestine, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.), flatten the decellularized matrix material C, repeat this operation 4 times, laminate it using a laminator, and cut it into rectangular strips with a width of 5 mm.

[0148] The antibiotic Y is a rifamycin antibiotic, and the rifamycin antibiotic is rifampicin.

[0149] The preparation method of the antibiotic Y solution is as follows: adding the antibiotic Y to a 95 wt % ethanol aqueous solution so that the concentration of the antibiotic Y in the antibiotic Y solution is 0.04 g / mL.

[0150] The combination of interlayer 1 - interlayer 2 and strip insertion layer 1 formed after steps S2 - S3 is a unit module 11 ; after the unit module 11 is repeatedly stacked three times, an intermediate layer ''' 10 is obtained; p=3.

[0151] S4. Using 6 mL of water, lay acellular matrix material A (derived from porcine bladder submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.) on the middle layer and flatten it to form the top layer of the antibacterial biopatch with controlled sustained release and high drug loading;

[0152] S5. Use a laminator to laminate at 43°C for 12 hours.

[0153] The cross-sectional structure of the obtained antibacterial biological patch with high drug loading and controlled sustained release is as follows: Figure 3 (b).

[0154] Comparative Example 1

[0155] This example provides a process for achieving controlled sustained release of a high-drug-loaded antibacterial biopatch. The process is as follows:

[0156] One layer of decellularized matrix material A (derived from porcine bladder submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.), four layers of decellularized matrix material B (derived from porcine small intestine submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.), and one layer of decellularized matrix material A (derived from porcine bladder submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.) were stacked in order from bottom to top, laminated at 40°C for 24 hours, removed, immersed in antibiotic solution X for 1 hour, removed, immersed in antibiotic solution Y for 1 hour, removed, and repeated the above immersion operation twice (equivalent to the laminated material being immersed in antibiotic solution X 3 times and antibiotic Y solution 3 times), with an interval of 1 hour between each immersion, and finally dried at room temperature for 24 hours, and placed in a 40°C oven for another 24 hours.

[0157] Wherein, antibiotic solution X and antibiotic solution Y were prepared according to the method of Example 4.

[0158] Comparative Example 2

[0159] This example provides a process for achieving controlled sustained release of a high-drug-loaded antibacterial biopatch. The process is as follows:

[0160] One layer of decellularized matrix material A (derived from porcine bladder submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.), four layers of decellularized matrix material B (derived from porcine small intestine submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.), and one layer of decellularized matrix material A (derived from porcine bladder submucosa, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.) were immersed in antibiotic solution X for 2 hours, taken out, and further immersed in antibiotic Y solution for 2 hours, taken out, and freeze-dried separately. The six layers of treated materials were stacked from bottom to top in the above order, laminated at 80°C for 18 hours using a laminator, and then taken out.

[0161] Wherein, antibiotic solution X and antibiotic solution Y were prepared according to the method of Example 4.

[0162] Comparative Example 3

[0163] This example provides a process for achieving a high-drug-loaded antibacterial biopatch with controlled sustained release. Unlike Comparative Example 2, the six layers of treated materials are stacked from bottom to top in the aforementioned order, laminated at 35°C for 24 hours, and then taken out.

[0164] Comparative Example 4

[0165] This example provides a process for achieving a controlled sustained-release antibacterial biopatch with a high drug loading. The difference from Example 2 is that the angle between the length direction of the bio-based strips of the strip insertion layer 1 and the length direction of the bio-based strips of the strip insertion layer 2 is 0°, and the bio-based strips of the strip insertion layer 1 and the strip insertion layer 2 are aligned and stacked. Figure 4 .

[0166] The base layer and the top layer are both called continuous planar layers; the strip insert layer 1 and the strip insert layer 2 are both called discontinuous surface layers. In this example, one side of the discontinuous surface layers, strip insert layer 1 and strip insert layer 2, is not in contact with the continuous planar layer.

[0167] Comparative Example 5

[0168] This example provides a process for achieving a high-drug-loaded antibacterial biopatch with controlled sustained release. The difference from Example 4 is that:

[0169] In steps S2 and S3, the antibiotic solution X is a suspension of a tetracycline metal chelate, which is obtained by adjusting the pH of an aqueous solution of a tetracycline / metal ion mixture to 7.3 with an aqueous sodium hydroxide solution.

[0170] In the aqueous solution of the tetracycline / metal ion mixture, the molar ratio of the metal ion to the tetracycline is 1.8:1.

[0171] The tetracycline is minocycline hydrochloride, and the metal ion is calcium ion.

[0172] The preparation method of antibiotic solution X is as follows: 0.1 g of minocycline hydrochloride is added to 10 mL of water, mixed, and then a 0.0045 mol / mL calcium chloride aqueous solution is added to adjust the molar ratio of calcium ions to minocycline hydrochloride in the antibiotic solution X to 1.8:1.

[0173] The preparation method of the bio-based strip is as follows: 6 mL of antibiotic Y solution is used to lay the decellularized matrix material C (derived from the submucosa of porcine small intestine, purchased from Zhuoruan Medical Technology (Suzhou) Co., Ltd.), flatten the decellularized matrix material C, repeat this operation 4 times, laminate it using a laminator, and cut it into rectangular strips with a width of 10 mm.

[0174] The antibiotic Y is a rifamycin antibiotic, and the rifamycin antibiotic is rifampicin.

[0175] The preparation method of the antibiotic Y solution is as follows: adding the antibiotic Y to a 94 wt % ethanol aqueous solution so that the concentration of the antibiotic Y in the antibiotic Y solution is 0.07 g / mL.

[0176] Furthermore, in step S3 , the bio-based strips of the strip insertion layer have no gaps.

[0177] Comparative Example 6

[0178] This example provides a process for achieving a high-drug-loaded antibacterial biopatch with controlled sustained release. The difference from Example 5 is that in step S2, n=8, and the combination of interlayer 1-interlayer 8 and strip insertion layer 1 formed after steps S2-S3 is a unit module; the unit module is repeatedly stacked 4 times to obtain an intermediate layer. 对比例6 ; p = 4. That is, the middle layer 对比例6 The total number of floors is controlled to be greater than 35.

[0179] Test results

[0180] 1. Morphological characterization: The antibacterial biological patch with high drug loading and controlled sustained release obtained by the process of Example 4 was characterized by SEM scanning electron microscopy. The results are as follows: Figure 5 ,from Figure 5 It can be seen that the antimicrobial drug crystals are distributed throughout the internal network and surface of the patch structure.

[0181] 2. Mechanical properties test: The antibacterial biological patches obtained in Examples 1-5 and Comparative Examples 1-6 were completely immersed in PBS buffer and placed at a temperature of 22±2°C for 6 h. After that, they were taken out and the tensile strength was tested according to GB / T328.8-2007 (stretched at a steady speed of 100 mm / min until fracture, and the maximum tensile force was recorded). 0729.2-2009 peel strength test, each test was carried out 5 times in parallel, and the average value was taken. The results are shown in Table 1; according to the same process, 9 parallel experiments were carried out on Examples 4-5 and Comparative Example 6 respectively, and the average deviations of the tensile strength and peel strength of the ten experiments were calculated respectively (calculation of average deviation: the sum of the deviations (absolute values) of the individual measured values ​​from the average value, divided by the number of measurements). The smaller the average deviation, the more stable the sample process. The results show that the average deviation of the sample in Comparative Example 6 is much larger than the average deviation of Examples 4-5, indicating that the process stability of Comparative Example 6 is poor; the average deviations of Examples 4 and 5 are close, indicating that for this solution, the increase in the number of layers does not have a negative impact on process stability.

[0182] Table 1

[0183] Example Tensile strength (N / cm) Peel strength (N / cm) Example 1 18 1.04 Example 2 21 0.95 Example 3 20 0.95 Example 4 25 0.89 Example 5 27 0.73 Comparative Example 1 8 0.27 Comparative Example 2 5 0.18 Comparative Example 3 10 0.21 Comparative Example 4 15 0.53 Comparative Example 5 20 0.78 Comparative Example 6 29 0.46

[0184] The tensile strength of the antibacterial patches obtained in Examples 1 to 5 were all greater than 10 N / cm, and the peel strength was greater than 0.7 N / cm, indicating that the patches in Examples 1 to 5 had good mechanical properties and a structure that was not easily loosened when exposed to water. Among them, the tensile properties of the patch obtained in Example 5 increased due to the increase in the number of layers, and the increase in the number of layers also resulted in a slight decrease in the interlayer bonding strength (reduced peel strength).

[0185] In Comparative Examples 1-3, no medium was used for laying. Instead, each layer of material was impregnated to load the drug. In Comparative Example 1, the entire pressed material was impregnated. However, theoretically, this method only loads the drug on the surface of the material, which is equivalent to a layer of drug surface coating. It is difficult to achieve a high drug loading of the material. At the same time, it can be seen that the mechanical properties of Comparative Example 1 are very poor. In addition, during the experiments of Comparative Examples 2 and 3, the inventors also observed that the thickness of the edges of the resulting products was significantly greater than the thickness of the middle of the product. This may be due to the shrinkage or curling of the material after impregnation, especially the obvious curling of the edges of the material after impregnation, resulting in a significant uneven thickness of the overall material after the lamination process. This obvious uneven thickness phenomenon was not observed in Examples 1-5, Comparative Examples 1, and Comparative Examples 4-6.

[0186] Compared with Example 2, the bio-based strips of Strip Insertion Layer 1 and Strip Insertion Layer 2 of Comparative Example 4 are aligned and stacked, and it can be seen that their mechanical properties are significantly reduced. Not only the peel strength (bonding force between reaction layers) is reduced by 44%, but the tensile strength is also reduced by 29%.

[0187] Compared with Example 4, the bio-based strips in the strip insertion layer of Comparative Example 5 have no gaps, and it can be seen that their mechanical properties are similar.

[0188] Compared with Example 5, the number of intermediate layers in Comparative Example 6 is significantly increased, the increase in tensile strength is smaller (increase by 7%), and the peel strength (reflecting the interlayer bonding strength) is significantly decreased (decreased by 37%).

[0189] 3. Drug loading and process drug loss test: The antibacterial patches obtained in Example 2, Examples 4-5, and Comparative Examples 1-5 were cut into 5 cm × 5 cm samples and respectively placed in methanol for extraction. h The extract was sampled and tested by liquid chromatography until the extract became colorless; sampling and testing were stopped; the drug loading was calculated according to the standard curve (the standards of rifampicin and minocycline hydrochloride were dissolved in methanol, diluted into a series of standard solutions of 1-500 μg / mL, and the standard curve was obtained by liquid chromatography injection testing), and the drug loss after the patch preparation process was calculated according to the amount of raw materials added in the process. The results are shown in Table 2; in the liquid chromatogram, under the same conditions, minocycline hydrochloride and the diastereomers of minocycline hydrochloride had different retention times and peak positions. Based on the proportion of peak area, the proportion of the diastereomers of minocycline hydrochloride in the mixture of minocycline hydrochloride and the diastereomers of minocycline hydrochloride in Examples 4-5 and Comparative Example 5 was calculated. The results are shown in Table 3; 5 parallel experiments were performed for each test and the average value was taken.

[0190] Table 2

[0191]

[0192] Table 3

[0193] Example Proportion of diastereomers of minocycline hydrochloride (%) Example 4 2.03 Example 5 2.14 Comparative Example 5 3.85

[0194] 4. In Vivo Drug Release Rate: Twenty-four rats were anesthetized with gaseous anesthesia, their abdominal hair shaved, and a 5 cm midline incision made to fully expose the abdominal cavity. The antibacterial patches obtained in Example 4 and Comparative Examples 1-5 were cut into 2 cm × 2 cm samples and placed on each side of the rat's abdominal cavity. The patches were secured with sutures and the wounds closed after implantation. Samples were collected at 4 hours, 1 day, 2 days, and 3 days after surgery. Following euthanasia by CO2 inhalation, the samples were removed and extracted with PBS buffer for 24 hours. The drug loading, W, was calculated based on a drug standard curve (rifampicin and minocycline hydrochloride standards were dissolved in PBS buffer, diluted to a series of standard solutions ranging from 1 to 500 µg / mL, and then injected and tested by liquid chromatography). In vivo release rate = (drug loading of each sample in Performance Test 3 - W) / drug loading of each sample in Performance Test 3 × 100%. The results are shown in Table 4:

[0195] Table 4

[0196]

[0197] The “ / ” in Table 4 indicates that the drug content on the sample patch is basically undetectable, which is equivalent to indicating that the drug loading amount of the corresponding drug on the sample patch is 0 at this time.

Claims

1. A process for achieving controlled sustained release of a high-drug-loaded antibacterial biological patch, characterized in that: The process is as follows: S1. Laying the decellularized matrix material A with water to obtain a base layer; the middle layer is obtained by the following steps S2-S3; S2. Use antibiotic solution X to lay acellular matrix material B on the base layer to form a sandwich 1 covering the base layer. Repeat this operation until a sandwich is formed. n ; n is a positive integer, and 1≤n≤18; S3. Place the bio-based strips in the sandwich n On, to form a strip insertion layer 1, repeat this operation until a strip insertion layer is formed m ; m is a positive integer, and 1≤m≤18; S4. Laying the acellular matrix material A on the strip insert layer with water to form the top layer of the antibacterial biological patch with high drug loading and controlled sustained release; S5. Laminate using a laminator to obtain; Alternatively, the process for producing a high-drug-loaded antibacterial biopatch with controlled sustained release is as follows: S1. Laying the acellular matrix material A with water to obtain a basal layer; S2. Use antibiotic solution X to lay acellular matrix material B on the base layer to form a sandwich 1 covering the base layer. Repeat this operation until a sandwich is formed. n ; n is a positive integer, and 1≤n≤18; S3. Place the bio-based strips in the sandwich n On, to form a strip insertion layer 1, repeat this operation until a strip insertion layer is formed m ; m is a positive integer, and 1≤m≤18; Interlayer 1 formed after steps S2-S3 n and stripe insertion layer m The combination of is a unit module; the unit module is repeatedly stacked p times to obtain the middle layer; p is a positive integer, and 1≤p≤18; S4. Laying the acellular matrix material A on the middle layer with water to form the top layer of the antibacterial biological patch with high drug loading and controlled sustained release; S5. Laminate using a laminator to obtain; Wherein, the antibiotic liquid X is a suspension of tetracycline metal chelate; the antibiotic Y is a rifamycin antibiotic, and the rifamycin antibiotic is rifampicin; The strip insertion layer is obtained by arranging the bio-based strips at intervals; the intervals between the bio-based strips are 0.7-2.2 cm, and the angle between the length direction of the bio-based strips and the length direction of the base layer is 15-75 degrees; The bio-based strip is prepared by laying the acellular matrix material C with an antibiotic Y solution, repeating this operation 1-10 times, laminating, and cutting into rectangular strips with a width of 1-20 mm. The total number of intermediate layers is controlled within 35 layers; The base layer, interlayer n and top layer are all called continuous plane layers, the strip insertion layer is called discontinuous surface layer, and 60%-100% of each plane area of ​​each discontinuous surface layer is in contact with the continuous plane layer.

2. The process for realizing controlled sustained release of a high drug loading antibacterial biological patch according to claim 1, characterized in that: The tetracycline metal chelate suspension is obtained by adjusting the pH condition of the aqueous solution of the tetracycline / metal ion mixture.

3. The process for realizing controlled sustained release of a high drug loading antibacterial biological patch according to claim 2, characterized in that: The pH is 7.2-8.

1.

4. The process for realizing controlled sustained release of a high drug loading antibacterial biological patch according to claim 2, characterized in that: In the aqueous solution of the tetracycline / metal ion mixture, the molar ratio of the metal ion to the tetracycline is (1.2-1.9):1 or (3-6.5):

1.

5. The process for realizing controlled sustained release of a high drug loading antibacterial biological patch according to claim 4, characterized in that: The preparation method of the antibiotic Y solution is as follows: adding antibiotic Y to 85wt%-95wt% ethanol aqueous solution to make the concentration of antibiotic Y in the antibiotic Y solution be 0.01-0.07g / mL.

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