Photo-cured xyloglucan hydrogel as well as preparation method and application thereof

Abstract

The invention relates to the field of biomedical polymer materials, in particular to light-cured xyloglucan hydrogel as well as a preparation method and application thereof. Methacrylic anhydride and xyloglucan react to obtain methallyl esterified xyloglucan, carbon-carbon double bonds are successfully modified on xyloglucan, so that the xyloglucan has photosensitivity, then free radical polymerization is initiated by ultraviolet light to form the photocured xyloglucan hydrogel, and the mechanical property and the stability of the xyloglucan hydrogel are improved. The photocuring xyloglucan pregel solution prepared by the method has good injectability, can be directly delivered to a target site, and is beneficial to filling irregular-shaped and deep wounds. The light-cured xyloglucan pre-gel solution prepared by the invention can generate light-cured xyloglucan hydrogel microspheres, and the porous microsphere structure imitating the extracellular matrix is suitable for drug loading, cell culture scaffolds and tissue engineering construction, so that the application scene of the xyloglucan hydrogel is effectively expanded.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of biomedical polymer materials, and particularly relates to a photocured xyloglucan hydrogel as well as a preparation method and application thereof. BACKGROUND

[0002] The information disclosed in this Background section is only for the purpose of increasing an understanding of the general background of the application and does not necessarily constitute an acknowledgement or any form of suggestion that this information forms part of the prior art already known to a person of ordinary skill in the art.

[0003] Hydrogel is a kind of polymer material with three-dimensional network structure, which has good biocompatibility, biodegradability and adjustable mechanical properties, and shows great potential in the field of biomedical materials, and is widely used in tissue repair, drug delivery and wound care. Traditional hydrogel dressings are mostly based on covalent crosslinking network, which is difficult to control the mechanical properties, and the formed hydrogel is difficult to completely match the irregular wound, which brings inconvenience to clinical operation.

[0004] Xyloglucan (XG) is derived from the primary cell wall of monocotyledonous plant seeds, and is a natural biomass-based material with good biocompatibility and biodegradability, which is an ideal tissue defect repair material. In particular, xyloglucan is a high molecular weight neutral branched polysaccharide, and its cellulose skeleton has galactose-substituted side chains. This unique branched structure not only makes it have high viscosity, but also plays a role in promoting cell aggregation and tissue regeneration in the wound healing process. Studies have shown that when xyloglucan is applied to local and mucosal wound dressings, it exhibits anti-inflammatory and immunomodulatory properties, and is a very potential and widely used tissue engineering material.

[0005] However, the current xyloglucan-based hydrogel is limited by the insufficient mechanical properties of the crosslinking network and the introduction of exogenous crosslinking agents, which restricts its application potential in tissue repair. SUMMARY

[0006] In order to overcome the above problems, the present application provides a photocured xyloglucan hydrogel as well as a preparation method and application thereof.

[0007] In order to achieve the above technical purposes, the present application adopts the following technical solutions: In a first aspect of the present application, a photocured xyloglucan hydrogel is provided, which is obtained by photocuring of methacrylated xyloglucan mixed with a photoinitiator.

[0008] In one or more embodiments, the average molecular weight of the unit sugar ring in the methacrylated xyloglucan is 155.0-184.3 g / mol, the degree of substitution is 9.1-53.3%, and the content of carbon-carbon double bond is 0.5-3.4 mmol / g.

[0009] In one or more embodiments, the photoinitiator comprises lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP), 2-hydroxy-2-methyl-1-[4-(2-hydroxyethoxy)phenyl]-1-propanone (Irgacure 2959).

[0010] In one or more embodiments, the mass ratio of the methacrylated xyloglucan to the photoinitiator is (0.6-5):(0.1-2).

[0011] In a second aspect of the present application, a preparation method of the photocured xyloglucan hydrogel is provided, and the preparation method comprises the following steps: (1) reacting methacrylic anhydride with xyloglucan to obtain methacrylated xyloglucan; (2) dissolving the methacrylated xyloglucan in water, then adding a photoinitiator, and mixing uniformly to obtain a photocured xyloglucan pre-gel solution; (3) curing the photocured xyloglucan pre-gel solution by ultraviolet irradiation to obtain the photocured xyloglucan hydrogel.

[0012] In one or more embodiments, the preparation method of the xyloglucan comprises: dispersing tamarind powder in deionized water, continuously stirring at a limited temperature, then separating the suspension, collecting the supernatant, and freeze-drying to obtain the xyloglucan.

[0013] Preferably, the mass ratio of the tamarind powder to the deionized water is 1:(90-110), preferably 1:100.

[0014] Preferably, the limited temperature is 35-45 ℃, preferably 40 ℃.

[0015] Preferably, the stirring time is 20-30 h, preferably 24 h.

[0016] In one or more embodiments, the average molecular weight of the unit sugar ring in the xyloglucan is 149.1 g / mol, and the content of hydroxyl group is 13.4 mmol / g.

[0017] In one or more embodiments, in step (1), the method for obtaining the methacrylated xyloglucan by reacting methacrylic anhydride with xyloglucan comprises: The xylanoglucan is dissolved in water to obtain a xylanoglucan solution; methacrylic anhydride is added, and the pH is adjusted to be alkaline, followed by light-shielded reaction; after the reaction is completed, the precipitate is collected after alcohol precipitation; the precipitate is dispersed in water, and the supernatant is collected by centrifugation, and the methacrylated xylanoglucan is obtained by dialysis, filtration and freeze-drying.

[0018] Preferably, the concentration of the xylanoglucan aqueous solution is 0.1-5% (w / v).

[0019] Preferably, the volume ratio of the xylanoglucan solution to the methacrylic anhydride is 100:(1-10).

[0020] Preferably, the pH is adjusted to be 8-10.

[0021] Preferably, the light-shielded reaction is carried out at room temperature for 6-24 h.

[0022] Preferably, the dialysis is carried out in deionized water for 72-168 h by using a dialysis bag with a molecular weight cut-off of 8-14 kDa.

[0023] In one or more embodiments, in step (2), the mass ratio of the methacrylated xylanoglucan to the photoinitiator is (0.6-5):(0.1-2).

[0024] In one or more embodiments, the concentration of the methacrylated xylanoglucan dissolved in water is 0.6-5% (w / v).

[0025] In one or more embodiments, in step (3), the light-cured xylanoglucan pre-gel solution is irradiated under ultraviolet light with an intensity of 30-60 mW / cm 2 for 15-60 s to be cured into a gel, i.e. to obtain the light-cured xylanoglucan hydrogel.

[0026] In a third aspect, the application provides the use of the light-cured xylanoglucan hydrogel of the first aspect or prepared by the preparation method of the second aspect in the field of medical tissue repair and biomaterials.

[0027] The application has the following advantages: (1) The application successfully modifies carbon-carbon double bonds on xylanoglucan by reacting methacrylic anhydride with xylanoglucan, so that the xylanoglucan has photosensitivity, and then a light-cured xylanoglucan hydrogel is formed by free radical polymerization initiated by ultraviolet light, thereby improving the mechanical properties and stability of the xylanoglucan hydrogel.

[0028] (2) The light-cured xylanoglucan pre-gel solution prepared by the application has good injectability, and can be directly delivered to the target site, which is beneficial to filling irregular shapes and deep wounds.

[0029] (3) The photocured xyloglucan pre-gel solution prepared by the present application can generate photocured xyloglucan hydrogel microspheres, and such porous microsphere structure similar to extracellular matrix is suitable for drug loading, cell culture scaffold and tissue engineering construction, effectively expanding the application scenarios of xyloglucan hydrogel. BRIEF DESCRIPTION OF DRAWINGS

[0030] The drawings accompanying the specification of the present application form a part thereof, serve to provide further understanding of the application, and together with the description, explain the application. The present application should not be limited by the specific illustrative embodiments herein.

[0031] Figure 1 It is a product diagram of photocured xyloglucan hydrogel and a corresponding scanning electron microscope diagram; Figure 2 It is a schematic diagram of the structure of xyloglucan extracted in Example 1 and the reaction to generate methacrylated xyloglucan, wherein (a) is the chemical structure of xyloglucan extracted from tamarind gum, and (b) is a schematic diagram of esterification reaction; Figure 3 It is a schematic diagram of preparing xyloglucan (XG) and methacrylated xyloglucan (MXG) by ultraviolet irradiation in Example 2; Figure 4 It is a structure characterization diagram of methacrylated xyloglucan generated in Example 2, wherein (a) is an infrared spectrum, and (b) is a nuclear magnetic resonance hydrogen spectrum; Figure 5 It is a scanning electron microscope diagram of photocured xyloglucan hydrogel with different solid contents prepared in Example 3; Figure 6 It is a rheological data diagram of photocured xyloglucan hydrogel with different solid contents prepared in Example 4; Figure 7 It is a structure characterization diagram of methacrylated xyloglucan with different degrees of substitution generated in Example 5, wherein (a) is an infrared spectrum, and (b) is a nuclear magnetic resonance hydrogen spectrum; Figure 8 It is a scanning electron microscope diagram of photocured xyloglucan hydrogel with different degrees of substitution prepared in Example 5; Figure 9 It is a rheological data diagram of photocured xyloglucan hydrogel with different degrees of substitution prepared in Example 5; Figure 10 It is a MTT cytotoxicity data diagram of photocured xyloglucan hydrogel with different degrees of substitution prepared in Example 6; Figure 11 It is a schematic diagram of the process of preparing photocured xyloglucan hydrogel microspheres in Example 7, wherein (a) is a microsphere preparation observation diagram, and (b) is a microsphere morphology observation diagram; Figure 12Scanning electron micrograph of photocured xyloglucan hydrogel microspheres prepared in Example 7. DETAILED DESCRIPTION

[0032] It should be noted that the following detailed description is exemplary in nature and is intended to provide further description of the application. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs.

[0033] It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of example embodiments in accordance with the present application. As used herein, the singular forms "a", "an" and "the" are intended to include the plural forms as well, unless the context clearly indicates otherwise. It will be further understood that the terms "comprises" and / or "comprising," when used in this specification, specify the presence of stated features, steps, operations, elements, and / or components, but do not preclude the presence or addition of one or more other features, steps, operations, elements, components, and / or groups thereof.

[0034] In recent years, hydrogel materials with three-dimensional cross-linked network structure and excellent hydrophilic properties have attracted extensive attention in the biomedical field. Studies have shown that polysaccharide compounds have been recognized as the most promising medical hydrogel building blocks due to their inherent excellent biocompatibility, controllable physicochemical properties, and significant cost advantage. The molecular characteristics possessed by this natural polymer are highly consistent with clinical needs, making it an ideal material for developing new synthetic medical hydrogels.

[0035] Xyloglucan (XG) is derived from the primary cell wall of monocotyledonous plant seeds and is a high molecular weight neutral branched polysaccharide with a cellulose backbone bearing galactose-substituted side chains. It is characterized by about 75% of the glucose units in the cellulose backbone being substituted with (1-6)-a-D-xylose branches, and some of the xylose residues being further substituted with (1-2)-b-D-galactose. The unique branched structure of xyloglucan not only makes it highly viscous in water, but also plays a role in promoting cell aggregation and tissue regeneration during the wound healing process.

[0036] In particular, xyloglucan contains a large number of hydroxyl groups, which can undergo esterification with methacrylic anhydride under certain conditions, allowing the hydroxyl groups in the sugar chain to react with the anhydride to form methacrylated xyloglucan (MXG).

[0037] In order for those skilled in the art to have a clearer understanding of the technical solutions of the present application, the technical solutions of the present application will be described in detail below in conjunction with specific examples.

[0038] Example 1 Xyloglucan (XG) was extracted from tamarind powder by water immersion method: 10 g of tamarind powder was added into 1 L of deionized water, and stirred at 40 °C for 24 h; the obtained suspension was centrifuged at 8000 rpm for 5 min to remove insoluble substances, and the supernatant was collected, and the supernatant was freeze-dried to obtain xyloglucan raw material; wherein the temperature of the centrifugation process is controlled at 4 °C; wherein the freeze-drying is preceded by pre-freezing at -20 °C overnight; The tamarind powder was from Tokyo Chemical Industry Co., Ltd.; the specific reagent name was tamarind gum (TSP).

[0039] The average molecular weight of the xyloglucan unit sugar ring prepared in this example was 149.1 g / mol, and the hydroxyl content was 13.4 mmol / g.

[0040] Example 2 Preparation of photocured xyloglucan hydrogel: 2 g of xyloglucan (XG) was weighed and dissolved in 100 mL of deionized water to prepare a xyloglucan solution with a concentration of 2% (w / v); under continuous stirring, 2 mL of methacrylic anhydride (MA) was added dropwise to the xyloglucan solution at a constant rate of 0.1 mL / min using a microsyringe pump; then, 2.5 mol / L sodium hydroxide solution was added dropwise to the xyloglucan solution to control the pH of the reaction system to 8; immediately transferred into a sealed photochemical reactor, and reacted at room temperature in the dark for 24 h to complete the chemical modification; after the reaction was terminated, 2 times the volume of anhydrous ethanol was added dropwise to the mixed system to induce precipitation of the product, and centrifuged at 12000 rpm and 20 °C for 15 min; the precipitate was collected, dispersed in 100 mL of ultrapure water, and ultrasonically assisted; centrifuged at 12000 rpm and 20 °C for 30 min to remove insoluble impurities; the supernatant was collected, filtered through a 0.22 μm filter membrane, and then separated by dialysis bag with a molecular weight cutoff of 8-14 kDa to retain methacrylated xyloglucan with a molecular weight of 200-300 kDa; residual reactants (sodium hydroxide, methacrylic anhydride, etc., molecular weight <1000 Da) were removed by 168 h continuous dialysis (dialysis solution was replaced every 8 h); the final product solution was pre-frozen (-20 °C, 12 h) and then freeze-dried in a freeze-drier for 72 h to obtain white sponge-like methacrylated xyloglucan (MXG).

[0041] Weigh 10 mg of the methacrylated xyloglucan (MXG) prepared above, and completely dissolve it in 1 mL of deionized water after ultrasonic treatment in a water bath at room temperature for 30 min to prepare a 1% (w / v) MXG base solution. Subsequently, add 5 mg of photoinitiator phenyl-2,4,6-trimethylbenzoyl lithium phosphine (LAP) to the homogenized solution and vortex for 5 min until completely dissolved. Finally, filter through a 0.22 μm sterile filter membrane to obtain a photocurable xyloglucan pregel solution, which is stored at 4 °C in the dark for later use.

[0042] The prepared xylo-glucan pregel solution was uniformly injected into a polytetrafluoroethylene gelling mold or skin defect using a sterile syringe, and a 405 nm ultraviolet light source (30 mW / cm²) was used. 2 Vertical irradiation for 15 s induces in-situ photocrosslinking, resulting in a photocured xylo-glucan hydrogel with a complete structure and excellent mechanical properties.

[0043] Figure 1 The image and corresponding scanning electron microscope (SEM) image of the photocurable xylo-glucan hydrogel product prepared in this embodiment are shown below. Figure 1 As shown, the photocurable xyloglucan hydrogel prepared in this embodiment exhibits good structural plasticity, and precise three-dimensional shaping (such as self-supporting structures like the irregular letters "SDU") can be achieved based on photocuring in situ gelation. Scanning electron microscopy characterization shows that the material presents a three-dimensional porous network structure with a uniform pore size distribution and an average pore size of approximately 100 μm, providing an ideal channel for nutrient transport and metabolic waste removal in the tissue regeneration microenvironment. The photocurable xyloglucan hydrogel prepared in this embodiment exhibits macroscopically controllable configuration and microscopic network characteristics, making the xyloglucan hydrogel system a promising candidate for applications in wound healing, tissue repair, and minimally invasive filling.

[0044] Figure 2 The structural formula of xyloglucan extracted from tamarind fruit and the basic structure of xyloglucan esterified by the reaction are shown. Observation Figure 2 (a) Schematic diagram of xylo-glucan structure. The main chain is glucan, and the side chains are composed of xylose and galactose-xylose substituents. It contains a large number of hydroxyl groups, which can serve as active sites for esterification reactions with acid anhydrides. Observation Figure 2 In (b), the hydroxyl groups in xyloglucan are used to esterify with methacrylic anhydride, which can introduce photosensitive carbon-carbon double bonds into xyloglucan to prepare methacrylic esterified xyloglucan.

[0045] To further verify the photocuring properties of the xyloglucan hydrogel, a pregel solution was prepared by combining xyloglucan methacrylated xyloglucan (MXG) and MXG. This solution was injected into sample vials and irradiated under a 405 nm UV lamp for 30 seconds to observe gel formation. Figure 3The preparation method of the wood glucan (XG) pre-gel solution is the same as that of the light-cured wood glucan pre-gel solution, except that the methacrylated wood glucan (MXG) is replaced by the wood glucan (XG).

[0046] As can be seen from Figure 3 , the XG group does not form a hydrogel after UV irradiation and still presents the flow characteristics of a liquid, while the MXG group forms a hydrogel in situ after UV irradiation, and the sample bottle can maintain the original shape when it is tilted or inverted, indicating the successful preparation of the light-cured wood glucan hydrogel.

[0047] Figure 4 In (a) of Figure 4 , it can be seen from the infrared spectra of the wood glucan (XG) and the methacrylated wood glucan (MXG) that a new peak appears at 1720 cm -1 , which is the characteristic peak of the stretching vibration of the ester carbonyl (C=O); the appearance of this peak indicates that the methacrylate has been successfully modified to the wood glucan molecule, introducing a photosensitive double bond into the wood glucan.

[0048] Figure 4 In (b) of Figure 4 , it can be seen from the nuclear magnetic resonance hydrogen spectrum of the wood glucan (XG) and the methacrylated wood glucan (MXG) that the peaks appearing at 5.66 ppm and 6.08 ppm belong to the olefin protons of the modified methacrylate, and the peak at 1.83 ppm belongs to the methyl protons of the methacrylate, which further confirms the structure of the MXG.

[0049] Example 3 10 mg, 20 mg and 30 mg of methacrylated wood glucan (MXG) were weighed, respectively, and completely dissolved in 1 mL of deionized water by room temperature water bath ultrasonic treatment for 30 min to prepare MXG base solutions with concentrations of 1%, 2% and 3% (w / v); then, 5 mg of the light initiator lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) was added to the homogeneous solution and vortexed for 5 min to completely dissolve, and finally filtered through a 0.22 μm sterile filter membrane to obtain a light-cured wood glucan pre-gel solution, which was stored in the dark at 4 °C for use.

[0050] The above-prepared cured wood glucan pre-gel solution was uniformly injected into a polytetrafluoroethylene gel mold using a sterile syringe, and a 405 nm ultraviolet light source (30 mW / cm 2 ) was vertically irradiated for 15 s to initiate in situ photocrosslinking, obtaining light-cured wood glucan hydrogels with solid contents of 1%, 2% and 3%.

[0051] After freeze-drying, the photocured Xyloglucan hydrogel was pasted onto the surface of conductive glue, and after 15 s gold spraying treatment, scanning electron microscope observation was carried out.

[0052] Figure 5 The scanning electron microscope images of photocured Xyloglucan hydrogels with different solid contents were prepared for this example. Figure 5 As can be seen from

[0053] Example 4 To illustrate the dependence of the mechanical properties of photocured Xyloglucan hydrogels on their solid content, rheological analysis was performed on MXG pre-gel solutions with different solid contents (0.5%, 1%, 2%, 3% (w / v)). 500 μL of sample was placed between the parallel plates of the rheometer, and the modulus change was monitored in time sweep mode (strain 1%, frequency 10 rad / s, total time 180 s). Photocuring was performed using a 405 nm light source (30 mW / cm 2 ) : first in the dark state (0-60 s), then with the light source turned on for 60 s (60-120 s), and finally after the light was turned off (120-180 s). The rheological data of photocured Xyloglucan hydrogels with different solid contents are shown in Figure 6 .

[0054] As shown in Figure 6 , before irradiation, the pre-gel of all concentrations showed typical viscoelastic fluid characteristics (i.e. storage modulus G' ≤ loss modulus G''). After the start of irradiation (60-120 s interval), all samples rapidly completed the photocuring transition (G' > G'') within 15 s, indicating the successful formation of photocured hydrogels. After curing (120-180 s), even in the last 60 s of testing after stopping the light, the G' and G'' of the hydrogels remained stable, indicating that they had good stability. In addition, with increasing solid content, the equilibrium G' of the MXG hydrogel after curing showed a significant enhancement trend, combined with the scanning electron microscope (SEM) observation results (pore size decreased with increasing solid content), which can be attributed to the fact that higher solid content promotes the construction of more dense hydrogel network by MXG, thereby endowing it with more excellent mechanical properties.

[0055] In combination with the experimental results in Examples 2-4, it can be seen that the photocured MXG hydrogel has excellent light responsiveness, plasticity and mechanical properties. Its wood glucomannan photocuring ability realizes in-situ filling of irregular defects and forms a uniform three-dimensional network hydrogel, thereby widening its application prospects in the fields of wound repair, tissue regeneration and minimally invasive filling.

[0056] Example 5 After confirming the excellent in-situ gelation performance of the photocured wood glucomannan hydrogel, the key parameter of methacrylate modification, i.e., the degree of substitution, was then systematically screened, with the specific process as follows: Take 2 g of wood glucomannan (XG) and dissolve it in 100 mL of deionized water to prepare a 2% (w / v) wood glucomannan solution; under continuous stirring, 0.5, 1.0, 2.0, 3.0 or 5.0 mL of methacrylic anhydride (MA) was added dropwise into five 2% (w / v) wood glucomannan solutions using a microsyringe pump (0.1 mL / min constant speed); then, 2.5 mol / L sodium hydroxide solution was added dropwise to each solution to adjust the pH of the reaction system to 8.0; immediately, the reaction solution was transferred into a sealed photochemical reactor and reacted at room temperature in the dark for 24 h to complete the methacrylate modification; after the reaction was terminated, 2 volumes of anhydrous ethanol were added to the mixture to precipitate the product, which was collected by high-speed refrigerated centrifuge (12000 rpm, 20 °C) for 15 min; the precipitate was dispersed in 100 mL of ultrapure water with ultrasonic assistance; the insoluble impurities were removed by centrifugation at 12000 rpm and 20 °C for 30 min; the supernatant was filtered through a 0.22 μm filter membrane, followed by continuous dialysis for 168 h (dialysis solution was replaced every 8 h); the final product solution was pre-frozen (-20 °C, 12 h) and then transferred into a freeze dryer for 72 h to obtain five groups of white sponge-like methacrylated wood glucomannan. According to the amount of MA added, they were labeled as 0.5 mL-MXG1, 1.0 mL-MXG2, 2.0 mL-MXG3, 3.0 mL-MXG4 and 5.0 mL-MXG5, respectively.

[0057] Take 10 mg of the above-prepared methacrylated wood glucomannan (MXG i ) sample and treat it with ultrasonic waves in a room temperature water bath for 30 min to completely dissolve it in 1 mL of deionized water to obtain a 1% (w / v) MXG i solution. Then, 5 mg of a photoinitiator LAP was added to each MXG i solution and vortexed for 5 min until completely dissolved; finally, each solution was filtered through a 0.22 μm sterile filter membrane to obtain a photocured wood glucomannan (MXG iThe pregel solution was stored at 4 °C protected from light until use.

[0058] The photocurable xylo-glucan pregel solution prepared above was uniformly injected into a polytetrafluoroethylene gelling mold using a sterile syringe, and then a 405 nm ultraviolet light source (light intensity: 30 mW / cm²) was used. 2 The injection area was vertically irradiated for 15 seconds to induce in-situ photocrosslinking, ultimately yielding various methacrylated xyloglucans (MXG) with complete three-dimensional network structures and good mechanical properties. i Hydrogel.

[0059] To verify different methacrylated xyloglucan (MXG) i Successful preparation of the sample and investigation of the relationship between its degree of substitution DS (MA) and the amount of methacrylic anhydride (MA) added were conducted using Fourier transform infrared spectroscopy (FT-IR) and proton nuclear magnetic resonance spectroscopy (NMR). 1 Chemical modifications were characterized by 1H NMR, and the degree of substitution of each sample was calculated based on the NMR data. Figure 7 The following are structural characterization diagrams of methacrylic esterified xyloglucan with different degrees of substitution generated in the embodiments of the present invention, wherein (a) is the infrared spectrum and (b) is the nuclear magnetic resonance hydrogen spectrum.

[0060] like Figure 7 As shown in (a), compared to the infrared spectrum of xyloglucan (XG), MXG... i All samples were at 1720 cm. -1 The characteristic peak of stretching vibration of ester carbonyl (C=O) appears at the position, and the area ratio of this peak increases with the increase of the amount of methacrylic anhydride (MA) added after normalization. Figure 7 (b) Proton NMR spectrum 1 ¹H NMR analysis showed that, compared with XG, MXG i The characteristic proton peaks (=CH2) of methacrylate olefins appeared at 5.66 ppm and 6.08 ppm, while the methyl proton peak (-CH3) appeared at 1.83 ppm. After normalization, the peak area ratios of both peaks showed the same increasing trend. (The text then abruptly shifts to a different topic: combining FT-IR and...) 1 ¹H NMR characterization results confirm that with the increase of MA addition in the esterification reaction, MXG i The degree of substitution (DS) was significantly increased. The integral ratio of olefin protons to xyloglucan backbone protons was determined by quantitative analysis of MXG. i The degree of substitution ranges from 9.1% to 53.3%, as shown in Table 1 below: Table 1. Different samples and corresponding degrees of substitution

[0061] The photo-cured xyloglucan (MXG i ) hydrogel prepared in this example was freeze-dried, fixed on the surface of conductive glue, and treated with 15 s gold spraying to improve the surface conductivity. The microstructure was observed by scanning electron microscopy (SEM). Figure 8 The scanning electron microscopy images of photo-cured xyloglucan hydrogels with different degrees of substitution prepared in this example are shown.

[0062] As shown in Figure 8 , the photo-cured xyloglucan (MXG i ) hydrogels with different degrees of substitution (DS) all showed a three-dimensional network porous structure. With the increase of the degree of substitution, the internal network density of the hydrogel was significantly enhanced, and the pore size gradually decreased. This phenomenon indicated that a higher degree of substitution on the MXG molecular chain could introduce more photo-polymerizable olefin groups, effectively promoting the cross-linking reaction during UV curing, and thus forming a more dense gel network structure.

[0063] To clarify the relationship between the mechanical properties of photo-cured xyloglucan (MXGi) hydrogels and the degree of substitution (DS), the photo-curing kinetics and modulus evolution were analyzed by rheological method (the same method as in Example 4). As shown in Figure 9 , the changes of storage modulus (G') and loss modulus (G'') of MXGi hydrogels with different degrees of substitution in time scanning mode (strain 1%, frequency 10 rad / s) were as follows: after 60 s of dark state equilibrium, the photo-curing was triggered by 405 nm light source (30 mW / cm 2 ) for 60 s, and then the behavior after curing was monitored for 60 s.

[0064] As shown in Figure 9 , compared with unmodified xyloglucan (XG), all MXG i hydrogels with different degrees of substitution (DS) showed photo-induced curing characteristics, and could maintain the curing modulus after the light was stopped. With the increase of DS, the equilibrium storage modulus (G') of curing was significantly improved. Combined with the phenomenon of decreasing pore size observed by SEM with the increase of DS, this trend could be attributed to the fact that at the same concentration, a higher DS introduced more photo-polymerizable olefin groups, promoting MXG to form a densified cross-linked network, thereby significantly improving its mechanical properties.

[0065] Example 6 To evaluate the biocompatibility of photo-cured xyloglucan (MXG i ) hydrogels, MTT method was used to detect their cytotoxicity. Mouse fibroblasts (L929) were co-cultured with different degrees of substitution MXG i hydrogel extracts, the absorbance (OD) was measured and the cell survival rate was calculated. Figure 10 The MTT cytotoxicity data graph of photo-cured xyloglucan hydrogels with different degrees of substitution prepared in this example is shown.

[0066] As shown in Figure 10 comparison with the blank control group, the unmodified xylan group (XG) showed significant cell proliferation behavior, which was attributed to the mucus-like side chains of XG forming a cell extracellular matrix-like adhesion microenvironment, promoting cell proliferation. Low-substitution MXG (MXG 1~3 ) completely retains this property, showing comparable cell compatibility and proliferation-promoting ability to XG. While high-substitution MXG (MXG 4~5 ) due to the excessive introduction of methacrylate groups, which destroys the original molecular side chain structure, leading to a decrease in cell compatibility with increasing substitution degree, and ultimately inhibiting cell expansion.

[0067] As can be seen from the experimental data in Examples 5 and 6, by adjusting the amount of methacrylic anhydride in the single-step esterification reaction, methacrylated xylan with a substitution degree of 9.1% to 53.3% can be synthesized, and a photocured xylan hydrogel with excellent mechanical modulus and good biocompatibility can be constructed. Among them, MXG3 maintains high mechanical modulus while showing excellent cell compatibility, significantly expanding the practical application potential of xylan hydrogel materials.

[0068] Example 7 Preparation of photocured xylan hydrogel microspheres: 20 mg of methacrylated xylan (MXG3) sample was accurately weighed and treated with room temperature water bath ultrasonic for 30 min to completely dissolve in 2 mL deionized water, to prepare 1% (w / v) MXG3 aqueous solution. 10 mg of photoinitiator LAP was added to the solution and vortexed for 5 min until completely dissolved. The obtained solution was filtered through a 0.22 μm sterile microporous filter membrane to obtain a photocured xylan pre-gel solution, which was stored at 4 °C in the dark as an aqueous phase for standby.

[0069] The aqueous phase used the above-mentioned photocured xylan pre-gel solution, and the oil phase was a volatile droplet generating oil (purchased from Shenzhen Yixin Life Science Co., Ltd., product number: YX-HFY-231215050). The two-phase solutions were injected into the microfluidic chip at a flow rate ratio of 1:5 (oil phase flow rate: aqueous phase flow rate) by microsyringe pumps. Based on the interfacial tension and shear force of the two phases, the aqueous phase formed monodisperse droplets in the oil phase and flowed out from the outlet. The droplets were collected in the volatile droplet generating oil and irradiated with 30 mW / cm 2 UV light for 30 s to crosslink and cure, generating photocured xylan hydrogel microspheres. Figure 11 The flowchart of photocured xylan hydrogel microspheres is shown in (a) and (b).

[0070] Observation Figure 11It can be seen from (a) that the monodisperse microsphere droplet array is formed in the microfluidic chip based on two-phase shearing action. After 30 mW / cm 2 After the UV irradiation and the completion of the photocuring crosslinking, the hydrogel microspheres with uniform size are obtained. The observation of the microspheres under the optical microscope shows that the microspheres have a monodisperse particle size and an intact spherical structure. Figure 11 It can be seen from (b) that the microspheres after curing have a monodisperse particle size and an intact spherical structure under the optical microscope.

[0071] The photocured xylomannan hydrogel microspheres prepared in the embodiment are freeze-dried, adhered to a conductive glue sample table and gold-sprayed for 15 s to enhance the surface conductivity, and then characterized by a scanning electron microscope (SEM) to observe the microstructure. Figure 12 The scanning electron microscope (SEM) image of the photocured xylomannan hydrogel microspheres prepared in the embodiment is shown in the following figure.

[0072] Figure 12 The characterization results show that the freeze-dried photocured xylomannan hydrogel microspheres have a spherical morphology with a particle size of about 500 μm and form a three-dimensional network structure with a pore size of about 50 μm. Such porous microsphere structure simulating the extracellular matrix is suitable for drug loading, cell culture scaffolds and tissue engineering construction, effectively expanding the application scenarios of xylomannan hydrogels.

[0073] The above only describes the preferred embodiments of the present application and is not used to limit the present application. For those skilled in the art, the present application can have various modifications and changes. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A photocurable xylo-glucan hydrogel, characterized in that, It is obtained by photocuring a mixture of methacrylated xylodecanol and a photoinitiator.

2. The photocurable xylo-glucan hydrogel as described in claim 1, characterized in that, The average molecular weight of the unit sugar ring in methacrylated xylodecanol is 155.0~184.3 g / mol, the degree of substitution is 9.1~53.3%, and the carbon-carbon double bond content is 0.5~3.4 mmol / g.

3. The photocurable xylo-glucan hydrogel as described in claim 1, characterized in that, The photoinitiator includes lithium phenyl-2,4,6-trimethylbenzoylphosphine and 2-hydroxy-2-methyl-1-[4-(2-hydroxyethoxy)phenyl]-1-propanone; Alternatively, the mass ratio of methacrylated xyloglucan to photoinitiator is (0.6~5):(0.1~2).

4. The method for preparing the photocurable xylo-glucan hydrogel according to any one of claims 1 to 3, characterized in that, Includes the following steps: (1) Methacrylic anhydride reacts with xyloglucan to obtain methacrylic esterified xyloglucan; (2) Dissolve methacrylated xyloglucan in water, then add a photoinitiator and mix well to obtain a photocurable xyloglucan pregel solution; (3) The xyloglucan pregel solution was cured into a gel by ultraviolet light irradiation to obtain a photocurable xyloglucan hydrogel.

5. The preparation method according to claim 4, characterized in that, The method for preparing the xyloglucan includes: Tamarind powder was dispersed in deionized water and stirred continuously at a limited temperature. The suspension was then separated, the supernatant was collected, and the supernatant was freeze-dried to obtain xyloglucan. Preferably, the mass ratio of tamarind powder to deionized water is 1:(90~110), more preferably 1:100; Preferably, the temperature is 35~45℃, more preferably 40℃; Preferably, the stirring time is 20-30 hours, more preferably 24 hours; Alternatively, the average molecular weight of the unit sugar ring of xyloglucan is 149.1 g / mol, and the hydroxyl content is 13.4 mmol / g.

6. The preparation method according to claim 4, characterized in that, In step (1), the method for reacting methacrylic anhydride with xyloglucan to obtain methacrylic esterified xyloglucan includes: Xyloglucan was dissolved in water to obtain a xyloglucan solution; methacrylic anhydride was added and the pH was adjusted to alkaline, followed by a reaction in the dark; after the reaction was completed, the precipitate was collected after alcohol precipitation; the precipitate was dispersed in water, the supernatant was collected by centrifugation, and the supernatant was obtained by dialysis, filtration, and freeze-drying.

7. The preparation method according to claim 6, characterized in that, The concentration of xyloglucan aqueous solution is 0.1~5% (w / v); Alternatively, the volume ratio of xyloglucan solution to methacrylic anhydride is 100:(1~10). Alternatively, adjust the pH to 8-10; Alternatively, the reaction conditions are: 6-24 h at room temperature in the dark; Alternatively, the dialysis conditions are as follows: use a dialysis bag with a molecular weight cutoff of 8-14 kDa and dialyze in deionized water for 72-168 h.

8. The preparation method according to claim 4, characterized in that, In step (2), the mass ratio of methacrylated xylodecanol to photoinitiator is (0.6~5):(0.1~2); Alternatively, the concentration of methacrylated xyloglucan dissolved in water is 0.6-5% (w / v).

9. The preparation method according to claim 4, characterized in that, In step (3), the light-cured xylo-glucan pregel solution is placed in a light source with an intensity of 30~60 mW / cm². 2 The xyloglucan hydrogel is obtained by irradiating it with ultraviolet light for 15-60 seconds to cure it into a gel.

10. The application of the photocurable xylo-glucan hydrogel according to any one of claims 1 to 3 or the photocurable xylo-glucan hydrogel prepared by the preparation method according to any one of claims 4 to 9 in the field of medical tissue repair and biomaterials.

Images