A method for constructing a dynamic boronic ester cross-linked anti-allergic hydrogel loaded with low molecular weight fucoidan and application to atopic dermatitis

Abstract

The application discloses a kind of low molecular weight fucoidan loaded dynamic borate ester crosslinking antiallergic hydrogel construction method and application to resist atopic dermatitis, belong to medical biomaterial field, to solve the poor performance of existing fucoidan hydrogel carrier, drug release effect is not good.The present application extracts and purifies natural fucoidan with Sargassum fusiforme as raw material, and obtains low molecular weight fucoidan by mild degradation using hydrogen peroxide-vitamin C system;3-carboxyphenylboronic acid is grafted to gelatin by N-hydroxysuccinimide and 1-ethyl-(3-dimethylaminopropyl) carbonyldiimidazole hydrochloride mediated reaction, and then polyvinyl alcohol is obtained by dynamic borate ester crosslinking to obtain crosslinked hydrogel.The hydrogel obtained by the present application has good self-repairing and biocompatibility, can long-acting release active ingredients, and the hydrogel is applied to resist atopic dermatitis.

Description

Technical Field

[0001] This invention belongs to the field of medical biomaterials and relates to a method for constructing a dynamic borate ester crosslinked anti-allergic hydrogel loaded with low molecular weight fucoidan and its application in treating atopic dermatitis. Background Technology

[0002] Food allergies have become an increasingly serious global public health problem. Among the symptoms caused by food allergies, skin allergies are the most common, with atopic dermatitis (AD) being a typical example. AD is a chronic, relapsing allergic skin disease with a continuously rising global prevalence, severely impacting patients' quality of life. Currently, routine clinical prevention and control mainly rely on oral antihistamines or topical corticosteroids. While these can relieve symptoms, long-term use can lead to problems such as skin atrophy, pigmentation, and systemic side effects. Natural polysaccharides have become a research hotspot for AD prevention and control due to their good biocompatibility and anti-allergic activity. However, high molecular weight polysaccharides have poor transdermal absorption and low bioavailability, limiting their effectiveness in local application. Therefore, developing low molecular weight polysaccharide-based hydrogel systems with controllable degradation, efficient penetration, and intelligent responsive drug release is of great significance for improving the safety and effectiveness of AD prevention and control. However, research on constructing local delivery systems based on low molecular weight fucoidan is still relatively limited, and studies on using low molecular weight fucoidan to construct dynamically responsive hydrogels to intervene in allergic skin inflammation are scarce. Summary of the Invention

[0003] This invention provides a method for constructing a dynamic borate ester crosslinked anti-allergic hydrogel loaded with low molecular weight fucoidan and its application in treating atopic dermatitis, in order to solve the problems of poor performance and unsatisfactory drug sustained-release effect of existing fucoidan hydrogel carriers.

[0004] 1. A method for constructing a dynamically borate ester crosslinked anti-allergic hydrogel loaded with low molecular weight fucoidan, comprising the following steps: S1. Wash and dry the Sargassum fusiforme, pulverize it and sieve it. Place it in a petroleum ether Soxhlet extraction apparatus and heat it under reflux to defatt it. After defatting, air dry it to obtain defatted Sargassum fusiforme powder. S2. After thoroughly mixing defatted Sargassum powder with citric acid solution, extracting in a water bath, the resulting extract was centrifuged, precipitated with ethanol, removed from organic reagents, purified by dialysis and freeze-dried to obtain fucoidan; S3. Prepare an aqueous solution of fucoidan, add a compound degradation agent composed of hydrogen peroxide and vitamin C to the fucoidan solution to carry out the degradation reaction; after the reaction is completed, adjust the pH of the system to neutral, purify by dialysis, freeze dry, and obtain low molecular weight fucoidan; S4. At a certain temperature, gelatin is dissolved in morpholine ethanesulfonic acid buffer, and 3-carboxyphenylboronic acid, N-hydroxysuccinimide, and 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride are dissolved together in dimethyl sulfoxide for carboxyl activation treatment to obtain an activated mixed solution. After the activated mixed solution is fully reacted with the gelatin solution, it is purified by dialysis and freeze-dried to obtain phenylboronic acid grafted modified gelatin. S5. Low molecular weight fucoidan is dispersed in pure water to prepare a polysaccharide aqueous solution, and then a polyvinyl alcohol mixed solution is prepared with the polysaccharide aqueous solution. Subsequently, it is mixed with a phenylboronic acid grafted modified gelatin solution and then doped and composited through dynamic borate ester crosslinking to obtain a dynamic borate ester crosslinked hydrogel loaded with low molecular weight fucoidan.

[0005] Furthermore, in S1, the drying temperature is 60°C, the powder is passed through a 65-mesh sieve, and the heating and reflux degreasing time is 6 hours.

[0006] Furthermore, S2 specifically includes: Using a 1% (w / w) citric acid solution as the extraction solvent, defatted Sargassum fusiforme powder was mixed with the citric acid solution at a mass-to-volume ratio of 1:50 and extracted in an 80℃ water bath for 3 hours. After cooling, the extract was collected by centrifugation at 4000 rpm. The extraction was repeated once, and all extracts were combined. The combined extract was concentrated to one-quarter of its original volume by rotary evaporation. 95% ethanol was added to the concentrated extract to achieve a final ethanol concentration of 80%. The extract was allowed to stand for 12 hours for alcohol precipitation, and then centrifuged at 4000 rpm to obtain the precipitate. The precipitate was washed three times with 95% ethanol to obtain the alcohol precipitate. The alcohol precipitate was then reacted with boiling water... Dissolve the sample at a mass-to-volume ratio of 1:20, heat in an 80°C water bath for 1 hour, and then centrifuge to collect the supernatant. Mix the supernatant with Sevage reagent (obtained from chloroform and n-butanol at a volume ratio of 4:1) at a volume ratio of 4:1, shake for 20-30 minutes, centrifuge at 4000 rpm, and collect the supernatant for deproteinization. Repeat the deproteinization process of mixing the supernatant and Sevage reagent, shaking, and centrifuging three times. After deproteinization, the supernatant is rotary evaporated at 30 rpm and 50°C for 60 minutes to remove organic reagents. Dialyze the supernatant in a dialysis bag for 48 hours and then freeze-dry to obtain fucoidan.

[0007] Furthermore, in step S3, the hydrogen peroxide is a 30% hydrogen peroxide solution, and the feed ratio of fucoidan solution, vitamin C, and 30% hydrogen peroxide solution is 1:2.64:1.70. The final concentration of the degradation system is 30 mmol / L, the reaction temperature is 50℃, the reaction time is 2 h, the solution for adjusting the pH of the system is NaOH solution, and the dialysis purification time is 48 h.

[0008] Furthermore, in S4, the temperature is 45°C and the pH of the morpholine ethanesulfonic acid buffer is 5.5.

[0009] Further, in S4, the molar ratio of 3-carboxyphenylboronic acid, N-hydroxysuccinimide, and 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride is 5:4:10; The activation treatment time was 2 hours, and the dialysis purification time was 72 hours.

[0010] Furthermore, S5 specifically includes: Prepare aqueous solutions of low molecular weight fucoidan with different mass fractions, wherein the mass fraction is 0.05% to 0.50%; A 10% polyvinyl alcohol mixed solution was prepared using low molecular weight fucoidan aqueous solutions of different mass fractions as solvents. The obtained polyvinyl alcohol mixed solution was mixed with a 5% phenylboronic acid grafted modified gelatin solution at a mass ratio of 1:1 and allowed to stand at room temperature to gel and form, thus obtaining dynamic borate crosslinked hydrogels with different drug loading capacities loaded with low molecular weight fucoidan.

[0011] Furthermore, the 10% polyvinyl alcohol mixed solution is prepared by adding polyvinyl alcohol to aqueous solutions of low molecular weight fucoidan with different mass fractions as solvents, followed by swelling at room temperature and heating in an oil bath at 95°C for 4–6 hours until completely dissolved.

[0012] Furthermore, the 5% phenylboronic acid grafted modified gelatin solution is prepared by dissolving phenylboronic acid grafted modified gelatin in a phosphate buffer solution with a pH of 7.2-7.4 and a concentration of 0.01 mol / L, adjusting the pH to 7.5-8.5 with sodium hydroxide solution, and stirring until completely dissolved.

[0013] This invention also provides an application of a dynamic borate ester crosslinked hydrogel loaded with low molecular weight fucoidan, obtained by a method for constructing such a hydrogel, in the treatment of atopic dermatitis.

[0014] Compared with the prior art, the present invention has the following beneficial effects: This invention uses Sargassum fusiforme fucoidan (SFF) as a raw material and obtains a low molecular weight degradation product (degraded Sargassum fusiforme fucoidan, DSFF) through a hydrogen peroxide-vitamin C degradation method. Phenylboronic acid-modified gelatin (Gel-BA) is further synthesized, and innovatively, it is cross-linked with polyvinyl alcohol (PVA) via dynamic borate ester to construct an anti-allergy hydrogel loaded with DSFF (DSFF@Gel-BA-PVA). The anti-allergy hydrogel aims to achieve local sustained release of DSFF, prolong the duration of action of low molecular weight fucoidan at the site of inflammation, and exert a synergistic intervention effect through the hydrogel's own moisturizing and barrier repair properties.

[0015] A hydrogel substrate was constructed by crosslinking 3-carboxyphenylboronic acid-grafted modified gelatin with polyvinyl alcohol via dynamic borate ester bonds. The dynamic and reversible bonds of the borate ester endow the hydrogel with excellent self-healing and mechanically adjustable properties. Low molecular weight fucoidan was in situ incorporated into the crosslinking network to achieve uniform loading and sustained release of active ingredients. The resulting composite hydrogel can effectively improve the condition of atopic dermatitis lesions and soothe skin inflammation. It also has good biocompatibility and skin adhesion. The preparation process is mild and controllable, and the raw materials are readily available, making it suitable for the large-scale development of topical anti-allergic skin preparations. Attached Figure Description

[0016] Figure 1 Fourier transform infrared (FT-IR) spectra of fucoidan (SFF) and low molecular weight fucoidan (DSFF); Figure 2 The UV-Vis absorption spectra of fucoidan (SFF) and low molecular weight fucoidan (DSFF) are shown. Figure 3 The molar percentage of each monosaccharide in fucoidan (SFF) and low molecular weight fucoidan (DSFF); Figure 4 The ¹H NMR spectra of 3-carboxyphenylboronic acid (BA), gelatin (Gel), and phenylboronic acid grafted gelatin (Gel-BA) are shown. Figure 5 FT-IR spectra of 3-carboxyphenylboronic acid (BA), gelatin, and phenylboronic acid grafted gelatin (Gel-BA); Figure 6 The storage modulus G' and loss modulus G'' of blank Gel-BA-PVA hydrogels with different mass ratios of Gel-BA to PVA (1~5 are 3:1, 2:1, 1:1, 1:2, 1:3 respectively) vary with angular frequency. Figure 7The storage modulus G' and loss modulus G'' of composite hydrogels with different DSFF loading amounts are shown as curves of change with angular frequency; (3: unloaded blank hydrogel, L, M, and H correspond to the 0.05%, 0.10%, and 0.50% DSFF addition groups, respectively). Figure 8 Statistical results of the color intensity of R, G, and B parameters of skin lesions in each group of mice; Figure 9 Statistical results of standardized H, S, and V values ​​in skin lesions of mice in each group; Figure 10 Results of skin thickness measurement at the lesion sites in each group of mice; Figure 11 The statistical results of body weight testing for each group of mice; Figure 12 The results of surface temperature measurements at the skin lesion sites of mice in each group; Figure 13 The results of spleen weight analysis for each group of mice are as follows; Figure 14 The results of spleen index statistics for each group of mice; Figure 15 This is a flowchart of the technology of the present invention. Detailed Implementation

[0017] The present invention will be further illustrated below with reference to embodiments. These embodiments are for illustrative purposes only and are not intended to limit the invention in any way. It should be understood that the described embodiments are merely some, not all, of the embodiments described in this application. All other embodiments obtained by those skilled in the art based on the embodiments in this application without inventive effort are within the scope of protection of this application.

[0018] Example 1 like Figure 15 As shown, this invention provides a method for constructing a dynamic borate ester crosslinked anti-allergic hydrogel loaded with low molecular weight fucoidan, comprising the following steps: After washing the purchased Sargassum fusiforme, dry it at 60℃, pulverize it using a grinder, and then pass it through a 65-mesh sieve. Take an appropriate amount of the sieved powder sample and treat it with petroleum ether in a Soxhlet extraction apparatus under reflux for 6 hours to remove lipid components from the sample. After defatting, defatted Sargassum fusiforme powder is obtained.

[0019] The defatted Sargassum fusiforme powder was mixed with a 1% citric acid solution at a material-to-liquid ratio of 1:50 (w / v). After thorough mixing, the mixture was extracted in an 80°C water bath for 3 hours. After cooling, the extract was centrifuged (4000 rpm) to obtain a clear extract. The extraction was repeated once, and the extracts were combined. The extract was evaporated to one-quarter of its original volume using a rotary evaporator. 95% ethanol was added to the concentrated extract to achieve an ethanol concentration of 80%, and the mixture was allowed to stand for 12 hours. The precipitate was obtained by centrifugation (4000 rpm). The precipitate was washed three times with 95% ethanol to obtain an alcohol precipitate. The alcohol precipitate was dissolved in boiling water at a ratio of 1:20 (w / v), and the mixture was heated in an 80°C water bath for 1 hour. The supernatant was obtained by centrifugation (4000 rpm). The supernatant was mixed with Sevage reagent (obtained by mixing chloroform and n-butanol at a volume ratio of 4:1) at a volume ratio of 4:1, and shaken thoroughly for 20-30 min. The mixture was then centrifuged (4000 rpm) to obtain the supernatant. This deproteinization process of mixing the supernatant and Sevage reagent, shaking, and centrifuging was repeated three times. The supernatant was then subjected to rotary evaporation (30 rpm, 50°C, 60 min) to remove organic reagents. Subsequently, the mixture was transferred to a 3500D dialysis bag and dialyzed for 48 h. After lyophilization, fucoidan was obtained, denoted as SFF (Sargassum fusiforme fucoidan).

[0020] A 2 mg / mL fucoidan solution was prepared, and hydrogen peroxide-vitamin C (H2O2-Vc) was added as a degrading agent to achieve a final concentration of 30 mmol / L (i.e., 306 μL of 30% H2O2 solution and 528.36 mg of solid Vc were added to 100 mL of fucoidan solution, and the reaction was carried out at 50 °C for 2 h). The mixture was adjusted to neutral with NaOH solution to terminate the reaction. After centrifuging to remove insoluble matter, the supernatant was transferred to a 1000 D dialysis bag and dialyzed for 48 h. The resulting low molecular weight fucoidan was obtained by freeze-drying and designated as DSFF (degraded Sargassum fusiforme fucoidan).

[0021] At 45°C, 1 g of gelatin was dissolved in 100 mL of morpholine ethanesulfonic acid buffer (0.1 mol / L, pH 5.5). Subsequently, 2.5 mmol of 3-carboxyphenylboronic acid (BA), 2 mmol of N-hydroxysuccinimide, and 5 mmol of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride were dissolved in dimethyl sulfoxide. After activating the carboxyl groups for 2 h, these solutions were added dropwise to the gelatin solution, and the mixture was stirred at 45°C for 48 h. The reaction mixture was then transferred to a 14000D dialysis bag and dialyzed for 72 h. The resulting gelatin was freeze-dried to obtain phenylboronic acid-grafted modified gelatin, designated Gel-BA.

[0022] Weigh 1.00 g of Gel-BA and dissolve it in 20 mL of 0.01 mol / L phosphate-buffered saline (PBS) buffer with a pH of 7.2-7.4. Then, adjust the pH of the solution dropwise to 7.5-8.5 using 1 mol / L NaOH solution. Next, weigh 10.00 g of PVA and add it to 100 mL of ultrapure water. Allow it to swell at room temperature for 2 hours, then heat it in a 95°C oil bath for 4-6 hours until the solution is clear and free of particulate matter. Add 5% (w / w) Gel-BA solution and 10% (w / w) PVA solution to 6-well plates at mass ratios of 3:1, 2:1, 1:1, 1:2, and 1:3, respectively. Stir magnetically at 200 rpm to ensure thorough mixing. Gelation is considered complete when magnetic stirring stops due to the formation of a self-supporting network, thus successfully preparing a Gel-BA-PVA blank hydrogel. Comparative analysis revealed that a 1:1 mass ratio of Gel-BA to PVA resulted in the most uniform hydrogel pore structure. Therefore, a 1:1 mass ratio of Gel-BA to PVA was used to prepare DSFF@Gel-BA-PVA hydrogels. Low molecular weight fucoidan (DSFF) was loaded into the Gel-BA-PVA hydrogel system to prepare DSFF@Gel-BA-PVA hydrogels. Three DSFF loading amounts were set: 0.05%, 0.10%, and 0.50%. The specific procedures were as follows: DSFF powder was dissolved in ultrapure water to prepare 0.05%, 0.10%, and 0.50% DSFF solutions, respectively. Subsequently, 10.00g of PVA was weighed and added to 100mL of 0.05%, 0.10%, and 0.50% low molecular weight fucoidan aqueous solutions to obtain a 10% polyvinyl alcohol mixed solution. The 10% polyvinyl alcohol mixed solution (DSFF@PVA) and the 5% phenylboronic acid grafted modified gelatin solution (Gel-BA) were placed in a 6-well plate and mixed evenly at a 1:1 mass ratio. The mixture was allowed to stand at room temperature to gel and form. When the gelation phenomenon was observed, DSFF@Gel-BA-PVA was prepared.

[0023] Chemical composition determination Total sugar content was determined using the phenol-sulfuric acid method, with L-fucoside as the standard: 1.00 mL of working solution was mixed with 1.00 mL of 6% phenol solution, followed by the addition of 5.00 mL of concentrated sulfuric acid. After thorough mixing, the reaction mixture was incubated in an 85°C water bath for 15 min, followed by cooling in a cold water bath for 10 min. The absorbance was measured at 490 nm. Uronic acid content was determined using the sulfuric acid-carbazole method, with D-glucuronic acid as the standard: 1.00 mL of working solution was mixed with 6.00 mL of concentrated sulfuric acid (18.4 mol / L), and the mixture was heated in an 80°C water bath for 5 min, followed by cooling in a cold water bath for 10 min. Then, 200 μL of carbazole-ethanol solution was added, and the mixture was heated again at 80°C for 10 min, followed by cooling in a cold water bath for 10 min. The absorbance was measured at 530 nm. The sulfate group content was determined using the gelatin-barium chloride method, with potassium sulfate as the standard: 1.0 mL of a 2.0 mg / mL sample solution was mixed with 1.0 mL of a 2.0 mol / mL hydrochloric acid solution and hydrolyzed at 105 °C for 4 h. Subsequently, 400 μL of the working solution was mixed sequentially with 3.60 mL of 3% trichloroacetic acid solution and 1.00 mL of gelatin-barium chloride solution. After mixing, the reaction mixture was allowed to stand for 20 min, and the absorbance value at 360 nm was recorded as A1. The above steps were repeated, except that 1.00 mL of gelatin solution was replaced with 1.00 mL of gelatin-barium chloride solution, and the absorbance value at 360 nm was recorded as A2. A standard curve was plotted with sulfate concentration on the x-axis and absorbance (A1-A2) on the y-axis. Protein content was determined using the Coomassie Brilliant Blue method, with bovine serum albumin (BSA) as the standard: 100 μL of working solution was mixed with 1.00 mL of Coomassie Brilliant Blue G-250 reagent diluted at a ratio of 1:4. After mixing, the mixture was allowed to stand for 5 min, and the absorbance was measured at a wavelength of 595 nm. The contents of all chemical components were calculated based on the corresponding standard curves.

[0024] The degradation treatment affected the main chemical composition of the fucoidan (SFF) and low molecular weight fucoidan (DSFF) prepared in Example 1. Compared with SFF, the total sugar content of DSFF was significantly reduced by about 15.01%, the uronic acid content was significantly reduced by about 11.71%, and the sulfate group content was reduced by about 5.29%, but the retention was better; the protein content was less than 1% in both cases, indicating that the sample purity was high.

[0025] Spectroscopic measurement Fourier transform infrared spectroscopy analysis was performed using the potassium bromide pellet method: the sample was mixed with potassium bromide powder dried to constant weight, thoroughly ground in an agate mortar, and then pressed into a transparent and uniform pellet. The Fourier transform infrared spectrometer was used at 4000-500 cm⁻¹. -1Infrared spectra were recorded within the specified range, with potassium bromide used as a blank control for background correction. Ultraviolet (UV) spectroscopy analysis: SFF and DSFF were dissolved in deionized water to prepare 0.1 mg / mL solutions. Samples were scanned using a UV spectrophotometer in the wavelength range of 200-400 nm, with deionized water used as a blank control. Absorption curves were recorded, and characteristic absorption peaks in the UV region were analyzed.

[0026] like Figure 1 Infrared spectral analysis showed that DSFF was at 3428 cm⁻¹ -1 (Hydroxy group), 2935cm -1 (CH), 1044cm -1 (Pyranose ring) and 824cm -1 The characteristic absorption peak at the (sulfate group) is basically consistent with that of SFF, confirming that the degradation treatment did not change the structure of the main functional groups of fucoidan. Figure 2 Ultraviolet spectroscopy analysis showed that SFF and DSFF had no absorption peaks at 280 nm and 400 nm, indicating that the content of protein and pigment impurities was extremely low; meanwhile, DSFF showed a characteristic absorption peak at 260 nm.

[0027] Monosaccharide composition determination Weigh 2.00 mg of sample and dissolve it in 2 mol / L trifluoroacetic acid solution, then hydrolyze at 105 °C for 4 h. Add methanol to the hydrolysate, and evaporate the mixture to dryness using a rotary evaporator (30 r / min, 50 °C). Repeat this step five times to completely remove residual trifluoroacetic acid. Redissolve the dried residue in an appropriate amount of ultrapure water to prepare a 10 μg / mL solution. After filtration through a 0.22 μm aqueous microporous membrane, transfer the solution to a 500 μL liquid chromatography vial. Nine monosaccharides were selected as standards, including mannose, rhamnose, glucuronic acid, galacturonic acid, glucose, galactose, xylose, arabinose, and fucose. Each standard was prepared as a 10 μg / mL solution, and then mixed to form a standard mixture. The mass of each monosaccharide was determined by absolute quantification, and the molar ratio was calculated based on the corresponding molecular weight. Fucose was the most abundant monosaccharide component in both SFF and DSFF. Figure 3 As shown, the degradation treatment led to a significant change in the monosaccharide composition: the molar percentage of fucose in DSFF increased from 43.17% to 46.71%, mannose from 9.57% to 15.58%, and glucuronic acid from 8.78% to 11.01%; while galactose, glucose, and xylose decreased to 16.79%, 4.92%, and 5.00%, respectively, and galacturonic acid was not detected. This remodeling stems from the selective retention of sulfated fucose-rich structural regions and the preferential cleavage of neutral sugar side chains during the degradation process, suggesting that DSFF may have superior activity in anti-allergy and immunomodulatory effects.

[0028] Molecular weight determination A laser light scattering detector and a differential refractive index detector connected in series were used: 10.00 mg of sample was weighed, dissolved in an appropriate amount of ultrapure water, and diluted to 1.00 mL to prepare a 10.0 mg / mL sample solution. After filtration through a 0.22 μm aqueous microporous membrane, the solution was analyzed by high performance size exclusion chromatography to determine its weight-average molecular weight (Mw), number-average molecular weight (Mn), and polydispersity index (PDI, Mw / Mn).

[0029] The degradation treatment reduced the weight-average molecular weight (Mw) of DSFF by approximately 69.81%, the number-average molecular weight (Mn) by approximately 12.78%, and the polydispersity index (PDI) by approximately 65.39%, resulting in a more uniform molecular weight distribution.

[0030] Particle size and zeta potential determination Dynamic light scattering (DLS) was used for determination: 5.00 mg of sample was weighed, dissolved in an appropriate amount of ultrapure water, and diluted to 10.00 mL to prepare a 0.5 mg / mL solution. After filtration through a 0.45 μm aqueous microporous membrane, the solution was sonicated for 15 min to ensure uniform dispersion. Particle size analysis was performed using a Malvern nanoparticle size analyzer: For particle size analysis, the sample solution was transferred to a dedicated cuvette, equilibrated at 25 °C for 2 min, and then measured three times repeatedly. For zeta potential determination, the sample solution was placed in a pleated capillary cell, equilibrated at 25 °C for 2 min, and then measured three times repeatedly under an applied voltage of 150 V.

[0031] Particle size analysis showed that the average particle size of DSFF decreased from 145.3 nm to 104.3 nm, a reduction of 28.22%, which helps improve water solubility and bioavailability. The absolute value of the Zeta potential of DSFF increased from 14.29 mV to 18.25 mV, an increase of 27.71%, and the surface negative charge density increased, indicating that it has better dispersion stability and colloidal stability in aqueous solution.

[0032] Thermogravimetric determination A 2.00 mg sample was placed in an alumina crucible and analyzed using a thermogravimetric analyzer under a nitrogen atmosphere at a flow rate of 20 mL / min. The temperature was programmed to increase from room temperature (25 °C) to 800 °C at a heating rate of 10 °C / min. Thermogravimetric (TG) curves (representing the relationship between mass loss and temperature) and derivative thermogravimetric (DTG) curves (representing the relationship between the rate of mass loss and temperature) were recorded in real time. The initial decomposition temperature, the temperature range of the main weight loss stage, and the percentage of weight loss were determined using the TG curves.

[0033] Thermogravimetric analysis of SFF and DSFF revealed that the main weight loss peak temperature of DSFF decreased from 242.5℃ to 218.5℃, a drop of about 24℃, indicating that its thermal stability was slightly lower than that of SFF. This was attributed to the weakening of chemical bond energy due to the shortening of the molecular chain. However, the char residues of the two at 800℃ were similar, at 33.18% and 30.65%, respectively, indicating that the final inorganic residue content was not significantly different and that both could maintain structural stability within the conventional processing and storage temperature range for biomaterials.

[0034] Bioactivity assay The anti-allergic activity was assessed using hyaluronidase inhibition rate as an indicator: 700 μL of acetate-sodium acetate solution, 100 μL of DSFF solution, and 100 μL of hyaluronidase solution were added sequentially, and the mixture was incubated at 37°C for 10 min. Then, 100 μL of sodium hyaluronate solution was added and thoroughly mixed, and the mixture was incubated at 37°C for another 15 min. The reaction was immediately terminated by adding CTAB solution. After mixing, the reaction mixture was allowed to stand at room temperature for 10 min, and the absorbance was measured at 400 nm. The inhibition rate was calculated using the following formula: ; In the formula, The absorbance of the buffer solution was used instead of the polysaccharide sample. The absorbance of the polysaccharide sample; The absorbance of the buffer solution instead of the enzyme solution; The absorbance of the buffer solution is used instead of the enzyme solution and polysaccharide sample.

[0035] Antioxidant activity was assessed by measuring DPPH free radical scavenging activity: 1 mL of working solution was mixed with 1 mL of 0.2 mmol / L DPPH-ethanol solution. After mixing, the mixture was reacted at room temperature in the dark for 30 min, and the absorbance at 517 nm was recorded. Repeat the above steps, but use 1 mL of ethanol solution instead of 1 mL of DPPH-ethanol solution, and record the absorbance at 517 nm. In addition, 1 mL of 0.2 mmol / L DPPH-ethanol solution was added to 1 mL of ultrapure water, and the absorbance at 517 nm was recorded. The DPPH free radical scavenging rate is calculated using the following formula: ; DSFF exhibited significantly enhanced in vitro physiological activity compared to SFF. Both SFF and DSFF showed a good dose-response relationship in hyaluronidase inhibition and DPPH radical scavenging, with DSFF showing significantly superior activity compared to SFF at the same concentrations. At a concentration of 1.0 mg / mL, DSFF achieved a hyaluronidase inhibition rate of 79.31% and a DPPH radical scavenging rate of 80.74%, approximately 1.36 times and 2.52 times that of SFF, respectively. This enhanced activity is attributed to the synergistic effect of multiple factors, including the exposure of active groups due to the reduced molecular weight, an increased proportion of key active monosaccharides, and an increased surface charge density.

[0036] Construction of dynamic borate crosslinked hydrogels Gel-BA structural characterization 10.00 mg of Gel-BA was dissolved in 0.5 mL of L2O, and then the solution was transferred to a 5 mm NMR tube. The proton resonance frequency was 400 MHz, the number of scans was 64, the relaxation delay was 2 seconds, and the temperature was 25 °C. Fourier transform infrared spectroscopy was used, and the specific operating procedures were the same as described above. XPS technology was used to analyze the elemental composition and chemical state of the sample surface: after grinding the sample into a uniform powder, an appropriate amount of sample was evenly coated onto the conductive adhesive surface on the sample holder. Before testing, the sample should be kept in a temperature better than 1 × 10⁻⁶ m² / m². -8 Samples were pretreated in a mbar vacuum environment to remove surface-adsorbed impurities and moisture. A monochromatic AlKα X-ray source with a beam size of 400 μm, an operating voltage of 15 kV, and an emission current of 10 mA was used. The scanning range covered 0–1350 eV with a step size of 1 eV, and a total of 3 scans were performed. High-resolution spectra were acquired for the characteristic peaks of C1s, N1s, O1s, and B1s, with a step size of 0.05 eV, for a total of 10 scans.

[0037] Gel-BA was systematically characterized using ¹H NMR, FTIR, XPS, and UV spectroscopy. Figure 4 As shown, ¹H NMR reveals characteristic proton peaks of the benzene ring in Gel-BA at δ 7.3–8.5 ppm; Figure 5 As shown, FTIR at 1333 cm⁻¹ -1 A stretching vibration peak of the BO bond was detected; XPS full-spectrum and fine-spectrum analysis confirmed the presence of element B and the chemical bonding state of the boric acid group.

[0038] Determination of the degree of substitution of Gel-BA phenylboronic acid The characteristic absorption peak of phenylboronic acid was identified. Standard solutions of different concentrations were prepared using phenylboronic acid as a standard, and a standard curve was established by fitting the relationship between absorbance and concentration at the characteristic absorption wavelength. Aqueous solutions of Gel and Gel-BA were precisely prepared, each with a concentration of 100 μg / mL. The absorbance of both solutions was measured at the same wavelength using a multi-mode microplate reader, and the content of BA in the solution was calculated based on the difference in absorbance between Gel-BA and Gel.

[0039] The degree of substitution was determined to be 37.09 ± 0.14 mg / g, indicating that the phenylboronic acid groups were successfully and appropriately grafted onto the gelatin molecular chain.

[0040] Morphological observation and state comparison of hydrogels The state of different precursor materials was observed using the inverted vial method. The gel-forming ability of hydrogels prepared with different mass ratios and hydrogels containing different concentrations of DSFF was further compared. The internal structure of the hydrogels was morphologically characterized using scanning electron microscopy (SEM). Gel-BA / PVA hydrogel samples prepared with different mass ratios were immersed in ultrapure water for 30 min to swell, then transferred to a -80℃ freezer overnight, and subsequently freeze-dried to constant weight. The freeze-dried hydrogels were immersed in liquid nitrogen and gently broken up. The broken surfaces were fixed to a sample substrate with the cross-section facing upwards, and gold sputtering was performed on the samples using an ion sputtering system (120 seconds, 15 mA) to improve conductivity. After SEM observation, images were acquired from different fields of view, with at least three independent regions examined for each sample.

[0041] A dynamic borate ester crosslinked hydrogel system was constructed. The effects of different mass ratios of Gel-BA to PVA (3:1, 2:1, 1:1, 1:2, 1:3) and different DSFF loadings (0.05%, 0.10%, 0.50%) on hydrogel formation ability and microstructure were investigated by vial inversion method and scanning electron microscopy. It was found that unmodified Gel-PVA could not form a gel when mixed with gel-BA, PVA, PVA@DSFF, Gel-BA / PVA, and Gel-BA / PVA@DSFF. However, stable gels could be formed at different Gel-BA to PVA ratios (3:1, 2:1, 1:1, 1:2, 1:3 by mass), with the hydrogel exhibiting the most uniform pore structure at a Gel-BA to PVA mass ratio of 1:1. When DSFF solutions with different DSFF loadings (0.05%, 0.10%, 0.50%) were used instead of pure water to prepare hydrogels with a mass ratio of 1:1, it was found that the system had good DSFF carrying capacity and the hydrogel pore structure was relatively stable.

[0042] Rheological behavior testing of hydrogels Dynamic frequency scanning was used to study the viscoelastic response behavior of hydrogel samples at different angular frequencies. The hydrogel samples were placed between parallel plates of a rheometer, with the temperature set at 25℃, the strain amplitude fixed at 1%, and the frequency scanning range at 0.1-100Hz. Within this frequency range, the changes in storage modulus (G′) and loss modulus (G″) with angular frequency were recorded.

[0043] like Figure 6 and Figure 7 Frequency scanning results showed that all groups of hydrogels exhibited typical viscoelastic behavior, with liquid-like properties in the low-frequency region and solid-like properties in the high-frequency region. With the increase of Gel-BA ratio, the cross-linking frequency of G′ and G″ shifted to lower frequencies, indicating an increase in cross-linking density; DSFF loading shifted the cross-linking frequency to higher frequencies, suggesting that the network structure tended to be looser, but all groups maintained an intact gel state.

[0044] In vitro drug release studies of hydrogels Weigh 0.50 g of DSFF@Gel-BA-PVA hydrogel and place it in 20 mL of PBS (pH 5.5, 6.8, and 7.4) to ensure complete immersion. Then transfer the hydrogel to a 37°C constant-temperature shaker and shake at 200 rpm in the dark. At predetermined time points (0.5, 1, 2, 4, 6, 8, 12, and 24 h), 1 mL of release medium was collected and immediately replaced with an equal volume of fresh PBS at the same temperature and pH. The absorbance of the collected release medium was measured at the maximum absorption wavelength of DSFF using a multi-mode microplate reader. The cumulative release at each time point was calculated based on the DSFF standard curve, and a release curve was plotted. Triple-sample experiments were performed for each pH condition, and the results are expressed as averages.

[0045] The in vitro drug release behavior and kinetic characteristics of the hydrogel were investigated. The drug release process exhibited a typical two-stage release pattern: rapid initial release (0-4 h) and slow, sustained release in the later stage (4-24 h). The cumulative release amounts after 24 h were 61.13%, 58.85%, and 47.04% at pH 5.5, 6.8, and 7.4, respectively, showing a trend of accelerated release in a weakly acidic environment. The kinetic model fitting results showed that the first-order kinetic model had the highest goodness of fit (R²>0.94). The release index n of the Korsmeyer-Peppas model was less than 0.45, indicating that the drug release behavior was mainly controlled by the Fick diffusion mechanism, and may also be accompanied by local dynamic rearrangement of the cross-linked network.

[0046] Example 2 A dynamic borate ester crosslinked anti-allergic hydrogel loaded with low molecular weight fucoidan is used in the treatment of atopic dermatitis.

[0047] The effect of hydrogels on the prevention and control of atopic dermatitis in mice Experimental mice: 6-8 week old SPF grade BALB / c female mice, weighing 18-22g, were used in this experiment. The housing environment was strictly controlled: temperature 25±2℃, relative humidity 50±5%, 12h light / dark cycle, and mice had free access to food and water.

[0048] Experimental grouping: After 1 week of acclimatization, mice were randomly divided into 8 groups of 6 mice each using a random number table. The groups were: blank control group (Ctrl), model control group (Mod), blank hydrogel group (Gel-B), low-dose DSFF hydrogel group (Gel-L), medium-dose DSFF hydrogel group (Gel-M), high-dose DSFF hydrogel group (Gel-H), desonide group (Drug-1), and mometasone furoate group (Drug-2). Drug-1 and Drug-2 served as positive control groups. Experimental protocol: One day prior to the experiment, hair removal cream was used to remove hair from a 2cm × 2cm area on the back of mice. During the sensitization phase, on days 1 and 3, 200 μL of a 1.0% 2,4-dinitrochlorobenzene (DNCB) solution was evenly applied to the shaving area on the back of the mice once daily; on days 5 and 7, 200 μL of a 0.5% DNCB solution was applied once daily. The blank control group received an equal volume of the solvent. By repeatedly applying DNCB solution, the mouse delayed-type skin sensitization model was established by simulating repeated antigen stimulation. The intervention phase was conducted from day 7 to day 13, for a total of 7 times. In this study, the Ctrl and Mod groups did not receive local drug treatment; the Gel groups (including Gel-B, Gel-L, Gel-M, and Gel-H groups) received topical application of 0.5g of Gel-BA-PVA hydrogel, measuring 2cm×2cm, either as a blank or containing different concentrations (0.05%, 0.10%, and 0.50%) of low molecular weight fucoidan (DSFF); and the positive control groups received topical application of 0.5g of cream containing 0.05% desonide or 0.10% mometasone furoate. Both the hydrogel and the drug adhered tightly to the hair-removed area on the mouse's back and remained in effect for 24 hours. To ensure the stability of the hydrogel at the application site, breathable medical nonwoven fabric was used for auxiliary fixation. On day 14, the mice were weighed using a scale and their body temperature was measured with a thermometer; the data were recorded in detail. Subsequently, the skin tissue and spleen of the mice were removed and weighed, and the results were accurately recorded for subsequent analysis.

[0049] Observation and quantitative color analysis of mouse skin condition During the experiment, changes in the appearance of the mouse's back skin were recorded using a mobile phone camera. All photos were taken with the same mobile phone, maintaining consistent background, lighting conditions, and focal length. Image analysis was performed using both RGB and HSV modes: Specifically, the mouse back skin photos were imported into ImageJ software, and the exposed back skin area was selected as the region of interest (ROI), while avoiding interference from edge hairs and the background. After completing white balance correction for all images, the built-in color analysis tool was used to extract red (R), green (G), and blue (B) values, as well as hue (H), saturation (S), and value (V) from the ROI.

[0050] NCB successfully induced typical specific dermatitis symptoms in mice, manifested as skin wrinkling, infiltration, and erythema. After intervention with DSFF@Gel-BA-PVA hydrogel, the appearance of mouse skin was significantly improved. On day 14, the skin in the gel group returned to smoothness without obvious infiltration, while the drug group showed side effects such as skin thinning and pigmentation.

[0051] Mouse skin score The skin scoring criteria primarily involve four dimensions: erythema, edema, desquamation, and erosion. This scoring system categorizes symptoms in each dimension into four levels, corresponding to 0, 1, 2, and 3 points respectively. During the assessment, mice were fixed on a platform, and skin parameters were carefully observed and accurately recorded under natural light. Each mouse's total score was calculated by summing the scores for erythema, edema, desquamation, and erosion, with a total score ranging from 0 to 12 points (0 representing the mildest and 12 representing the most severe).

[0052] like Figure 8 and Figure 9 As shown, RGB and HSV color quantitative analysis indicated that DSFF@Gel-BA-PVA hydrogel effectively relieved skin erythema and promoted color normalization, while drugs, although able to inhibit erythema, were accompanied by decreased brightness and pigmentation. Meanwhile, the four-dimensional and three-dimensional skin allergy scores showed that the Gel-H group significantly reduced skin scores by day 11, comparable to the Drug group; by day 14, the Gel group's scores further decreased and were significantly lower than the Drug group, which showed a 42.11%-46.24% increase in scores.

[0053] H&E staining of mouse skin sections Skin tissue was collected from the dorsal region of mice and fixed with 4% paraformaldehyde for 24 hours at room temperature. After dehydration with a gradient of ethanol and clearing with xylene, the tissue was embedded in paraffin. The embedded tissue block was serially sectioned into 4 μm thick sections using a rotary microtome. The sections were fixed onto glass slides and baked in a 60°C oven for 2 hours. The sections were then dewaxed with xylene, rehydrated with a gradient of ethanol, stained with hematoxylin, and rinsed with tap water to remove blue staining. Eosin staining was followed by rinsing again. The sections were then dehydrated with a gradient of ethanol, cleared with xylene, and finally mounted with neutral resin. After the mounting medium dried, the sections were observed and images were taken under an optical microscope. Three non-overlapping random fields of view were selected for observation of each section.

[0054] H&E staining confirmed that the Gel-M and Gel-H groups could effectively improve epidermal thickening, spongy edema and vacuolar degeneration, and restore the integrity of the epidermal structure, while the Drug group was accompanied by a tendency of epidermal atrophy.

[0055] Measurement of mouse skin thickness Skin tissue from the subject's back area was removed using sterile surgical scissors, ensuring complete preservation of the epidermis, dermis, and subcutaneous tissue. The tissue surface was rinsed with PBS to remove residual blood and debris, and then trimmed from both sides towards the center to a size of 1.5cm × 1.5cm. Excess moisture was blotted with filter paper, and the tissue was weighed. The thickness was estimated based on the weight of the skin per unit area.

[0056] Regarding skin thickness and histopathology, such as Figure 10 As shown, the skin thickness in the Mod group increased by 43.31% compared to the Ctrl group, while the Gel group showed a dose-dependent decrease with the highest recovery rate of 58.13%. Furthermore, there was no overcorrection phenomenon observed in the Drug group (skin thickness decreased by 33.80%-48.08%).

[0057] Measurement of mouse body weight and body temperature On day 14 of the experiment, the weight of each mouse was measured using a scale, and the body temperature was measured using an electronic thermometer. All data were recorded. Weight measurement: Each mouse was placed in a fixed container and allowed to settle before being weighed on a scale, with the weight recorded to the nearest 0.1 gram. Body temperature measurement: An electronic thermometer was used. The probe was lubricated with a small amount of petroleum jelly and gently inserted into the mouse's rectum approximately 1 cm deep. The reading was recorded after stabilization, accurate to 0.1°C. All measurements were performed gently and quickly to minimize stress on the animals.

[0058] like Figure 11 and Figure 12 As shown, the gel group had a mild effect on mouse body weight and could effectively restore hypothermia caused by allergies; the drug group had a significant decrease in body weight of 21.55%-26.77%, and body temperature was negatively correlated with body weight, suggesting a risk of metabolic disorders.

[0059] Determination of mouse spleen and spleen index After obtaining skin tissue, the mouse peritoneum was quickly opened, and the spleen was carefully separated and removed intact. The spleen was rinsed with pre-cooled PBS to remove residual blood and connective tissue, and the surface moisture was blotted dry with filter paper. The spleen was then weighed using an electronic analytical balance to an accuracy of 0.1 mg, and the weight of each mouse's spleen was recorded. Gentle handling was used to prevent mechanical damage to the spleen and to ensure accurate and reliable weighing results. The spleen index was calculated using the following formula: ; like Figure 13 and Figure 14 As shown, spleen index analysis revealed that the Gel group significantly reduced the elevated spleen index in model mice to near-normal levels, while the Drug group showed excessive suppression, suggesting a risk of impaired immune function.

[0060] Of course, the above description is not intended to limit the present invention, and the present invention is not limited to the examples given above. Any changes, modifications, additions or substitutions made by those skilled in the art within the scope of the present invention should also fall within the protection scope of the present invention.

Claims

1. A method for constructing a dynamically borate ester-crosslinked anti-allergic hydrogel loaded with low molecular weight fucoidan, characterized in that, Includes the following steps: S1. Wash and dry the Sargassum fusiforme, pulverize it and sieve it. Place it in a petroleum ether Soxhlet extraction apparatus and heat it under reflux to defatt it. After defatting, air dry it to obtain defatted Sargassum fusiforme powder. S2. After thoroughly mixing defatted Sargassum powder with citric acid solution, extracting in a water bath, the resulting extract was centrifuged, precipitated with ethanol, removed from organic reagents, purified by dialysis and freeze-dried to obtain fucoidan; S3. Prepare an aqueous solution of fucoidan, add a compound degradation agent composed of hydrogen peroxide and vitamin C to the fucoidan solution to carry out the degradation reaction; after the reaction is completed, adjust the pH of the system to neutral, purify by dialysis, freeze dry, and obtain low molecular weight fucoidan; S4. At a certain temperature, gelatin is dissolved in morpholine ethanesulfonic acid buffer, and 3-carboxyphenylboronic acid, N-hydroxysuccinimide, and 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride are dissolved together in dimethyl sulfoxide for carboxyl activation treatment to obtain an activated mixed solution. After the activated mixed solution is fully reacted with the gelatin solution, it is purified by dialysis and freeze-dried to obtain phenylboronic acid grafted modified gelatin. S5. Low molecular weight fucoidan is dispersed in pure water to prepare a polysaccharide aqueous solution, and then a polyvinyl alcohol mixed solution is prepared with the polysaccharide aqueous solution. Subsequently, it is mixed with a phenylboronic acid grafted modified gelatin solution and then doped and composited through dynamic borate ester crosslinking to obtain a dynamic borate ester crosslinked hydrogel loaded with low molecular weight fucoidan.

2. The method for constructing a dynamic borate ester crosslinked anti-allergy hydrogel loaded with low molecular weight fucoidan according to claim 1, characterized in that, In S1, the drying temperature is 60℃, the powder is passed through a 65-mesh sieve, and the heating and reflux degreasing time is 6 hours.

3. The method for constructing a dynamic borate ester crosslinked anti-allergy hydrogel loaded with low molecular weight fucoidan according to claim 1, characterized in that, S2 specifically includes: Using a 1% (w / w) citric acid solution as the extraction solvent, defatted Sargassum fusiforme powder was mixed with the citric acid solution at a mass-to-volume ratio of 1:50 and extracted in an 80℃ water bath for 3 hours. After cooling, the extract was collected by centrifugation at 4000 rpm. The extraction was repeated once, and all extracts were combined. The combined extract was concentrated to one-quarter of its original volume by rotary evaporation. 95% ethanol was added to the concentrated extract to achieve a final ethanol concentration of 80%. The extract was allowed to stand for 12 hours for alcohol precipitation, and then centrifuged at 4000 rpm to obtain the precipitate. The precipitate was washed three times with 95% ethanol to obtain the alcohol precipitate. The alcohol precipitate was then reacted with boiling water... Dissolve the sample at a mass-to-volume ratio of 1:20, heat in an 80°C water bath for 1 hour, and then centrifuge to collect the supernatant. Mix the supernatant with Sevage reagent (obtained from chloroform and n-butanol at a volume ratio of 4:1) at a volume ratio of 4:1, shake for 20-30 minutes, centrifuge at 4000 rpm, and collect the supernatant for deproteinization. Repeat the deproteinization process of mixing the supernatant and Sevage reagent, shaking, and centrifuging three times. After deproteinization, the supernatant is rotary evaporated at 30 rpm and 50°C for 60 minutes to remove organic reagents. Dialyze the supernatant in a dialysis bag for 48 hours and then freeze-dry to obtain fucoidan.

4. The method for constructing a dynamic borate ester crosslinked anti-allergy hydrogel loaded with low molecular weight fucoidan according to claim 1, characterized in that, In step S3, the hydrogen peroxide is a 30% hydrogen peroxide solution, and the feed ratio of fucoidan solution, vitamin C, and 30% hydrogen peroxide solution is 1:2.64:1.

70. The final concentration of the degradation system is 30 mmol / L, the reaction temperature is 50℃, the reaction time is 2 h, the pH of the system is adjusted by NaOH solution, and the dialysis purification time is 48 h.

5. The method for constructing a dynamic borate ester crosslinked anti-allergic hydrogel loaded with low molecular weight fucoidan according to claim 1, characterized in that, In S4, the temperature is 45°C and the pH of the morpholine ethanesulfonic acid buffer is 5.

5.

6. The method for constructing a dynamic borate ester crosslinked anti-allergic hydrogel loaded with low molecular weight fucoidan according to claim 1, characterized in that, In S4, the molar ratio of 3-carboxyphenylboronic acid, N-hydroxysuccinimide, and 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride is 5:4:

10. The activation treatment time was 2 hours, and the dialysis purification time was 72 hours.

7. The method for constructing a dynamic borate ester crosslinked anti-allergy hydrogel loaded with low molecular weight fucoidan according to claim 1, characterized in that, S5 specifically includes: Prepare aqueous solutions of low molecular weight fucoidan with different mass fractions, wherein the mass fraction is 0.05% to 0.50%; A 10% polyvinyl alcohol mixed solution was prepared using low molecular weight fucoidan aqueous solutions of different mass fractions as solvents. The obtained polyvinyl alcohol mixed solution was mixed with a 5% phenylboronic acid grafted modified gelatin solution at a mass ratio of 1:1 and allowed to stand at room temperature to gel and form, thus obtaining dynamic borate crosslinked hydrogels with different drug loading capacities loaded with low molecular weight fucoidan.

8. The method for constructing a dynamic borate ester crosslinked anti-allergy hydrogel loaded with low molecular weight fucoidan according to claim 7, characterized in that, The 10% polyvinyl alcohol mixed solution was prepared by adding polyvinyl alcohol to low molecular weight fucoidan aqueous solutions of different mass fractions as solvents, followed by swelling at room temperature and heating in an oil bath at 95°C for 4–6 hours until completely dissolved.

9. The method for constructing a dynamic borate ester crosslinked anti-allergic hydrogel loaded with low molecular weight fucoidan according to claim 7, characterized in that, The 5% phenylboronic acid grafted modified gelatin solution is prepared by dissolving phenylboronic acid grafted modified gelatin in a phosphate buffer solution with a pH of 7.2-7.4 and a concentration of 0.01 mol / L, adjusting the pH to 7.5-8.5 with sodium hydroxide solution, and stirring until completely dissolved.

10. The application of the dynamic borate ester crosslinked hydrogel loaded with low molecular weight fucoidan obtained by any of the construction methods described in claims 1-9 in the treatment of atopic dermatitis.

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