Copper ion hydrogel as well as preparation method and application thereof

Abstract

The invention provides copper ion hydrogel as well as a preparation method and application thereof, and the copper ion hydrogel is prepared from the following raw materials in percentage by mass: 0.02%-0.1% of soluble copper salt, 0.6%-8% of xanthan gum, 0.1%-1% of sodium hyaluronate, 0.5%-9% of butanediol, 0.1%-10% of glycerol, 0.02%-1% of betaine, 0.8%-9% of panthenol, 0.05%-1% of allantoin and the balance of water. The copper ion hydrogel can promote collagen regeneration, accelerate skin barrier structure reconstruction and improve chronic skin injury, and can be used for treating dermatitis skin diseases of pets.

Description

Technical Field

[0001] This invention relates to the field of skin repair and biomaterials technology, specifically to a copper ion hydrogel, its preparation method, and its application. Background Technology

[0002] With the continuous changes in living environment, lifestyle, and external stimuli, the incidence of skin damage and chronic skin barrier dysfunction is increasing year by year. As an important defense barrier against external aggressors, the skin is highly susceptible to physical friction, environmental stimuli, microbial infections, and abnormal immune responses, leading to problems such as dryness, erythema, desquamation, itching, persistent inflammation, or slow repair. Especially in chronic skin diseases, damage to skin structure is often persistent and recurrent, making the recovery of the skin barrier even more complex.

[0003] During skin repair, the content and quality of collagen in the dermis are key factors in maintaining tissue stability, elasticity, and repair capabilities. When skin is damaged or in a state of chronic inflammation, collagen is prone to degradation, and its regeneration rate decreases significantly, leading to delayed tissue repair, difficulty in restoring barrier function, and recurrent skin damage. Whether caused by inflammation, dry skin conditions, barrier damage, or local microenvironmental disturbances, promoting collagen regeneration has become an important means to improve skin repair efficiency. Existing skin repair products mainly focus on anti-inflammation, moisturizing, disinfection, or promoting stratum corneum repair, but most lack the ability to promote dermal matrix reconstruction and cannot directly enhance collagen regeneration. While antibiotics or antifungal agents have some effect on infection control, long-term use may disrupt the skin's microecological balance and is not suitable for continuous skin repair programs. Furthermore, skincare preparations with plant extracts or traditional moisturizers as main ingredients, although gentle, usually have limited effects on promoting deep skin structure repair and are insufficient to meet the collagen renewal needs of chronic damage repair.

[0004] Furthermore, seborrheic dermatitis is one of the most common chronic relapsing skin diseases in companion animals, and its occurrence is closely related to multiple factors such as sebaceous gland secretion disorders, skin barrier damage, microbial community imbalance, and abnormal immune regulation. Among these factors, Malassezia furfur (… Malassezia spp. The abnormal proliferation of Malassezia furfur is widely considered a significant contributing factor to seborrheic dermatitis. This fungus proliferates rapidly in a sebum-dependent environment, causing typical symptoms such as itching, erythema, scaling, oiliness, and folliculitis, severely impacting the skin health and quality of life of pets.

[0005] Currently, the clinical treatment of seborrheic dermatitis in pets mainly relies on antibiotics and antifungal drugs. However, antibiotics have significant limitations in the management of pet skin diseases. Their use easily leads to a gradual increase in drug resistance in the skin microbiome, and long-term or repeated use may even induce persistent colonization of drug-resistant bacteria. Furthermore, antibiotics lack specificity against lipid-dependent fungi such as Malassezia furfur, making it difficult to effectively control the core pathogenic factors of seborrheic dermatitis. In addition, antibiotic preparations are generally unsuitable for daily skin care in pets; overuse can not only disrupt the normal skin microecology but may also further weaken the stability of the skin barrier. While topical preparations with traditional Chinese medicine as the main ingredient are relatively mild, their inhibitory effect on fungi such as Malassezia furfur is limited, often failing to meet the actual needs of antifungal treatment and daily care for seborrheic dermatitis.

[0006] Therefore, developing a topical material that can directly promote collagen regeneration, assist in the reconstruction of the skin barrier structure, and has good antifungal properties is of great significance for improving the skin's self-repair ability, shortening the damage recovery cycle, and improving seborrheic dermatitis caused by fungi. Summary of the Invention

[0007] To address the technical limitations of existing topical skin preparations in terms of collagen renewal, barrier reconstruction, and antifungal effects, this invention provides a copper ion hydrogel, its preparation method, and its applications. The copper ion hydrogel can promote collagen regeneration, accelerate skin barrier structure reconstruction, and improve chronic skin damage, and can also be used to treat pet dermatitis.

[0008] This invention is achieved through the following technical solution: In a first aspect, the present invention provides a copper ion hydrogel, wherein, by mass percentage, the raw materials for preparing the copper ion hydrogel include: 0.02%~0.1% soluble copper salt, 0.6%~8% xanthan gum, 0.1%~1% sodium hyaluronate, 0.5%~9% butylene glycol, 0.1%~10% glycerol, 0.02%~1% betaine, 0.8%~9% panthenol, and 0.05%~1% allantoin, with the balance being water.

[0009] Furthermore, by mass percentage, the raw materials for preparing the copper ion hydrogel include: 0.05%~0.08% soluble copper salt, 2%~4% xanthan gum, 0.1%~0.5% sodium hyaluronate, 3%~5% butylene glycol, 6%~10% glycerol, 0.5%~1% betaine, 1%~3% panthenol, and 0.05%~0.1% allantoin, with the balance being water.

[0010] Preferably, the soluble copper salt is copper sulfate, copper gluconate, copper citrate, or copper chloride.

[0011] Furthermore, the soluble copper salt is copper sulfate.

[0012] Secondly, the present invention provides a method for preparing the copper ion hydrogel, comprising: S1. Dissolve soluble copper salt in water to obtain a copper salt solution; S2. Add xanthan gum to the copper salt solution and mix well to obtain copper salt-xanthan gum hydrogel; S3. Add sodium hyaluronate, butylene glycol, glycerin, betaine, panthenol, and allantoin to the copper salt-xanthan gum hydrogel, and adjust the pH to 5.8-7.2 to obtain a copper ion hydrogel.

[0013] Preferably, step S2 specifically involves: heating the copper salt solution to 35-45°C, adding xanthan gum and mixing thoroughly to obtain a copper salt-xanthan gum hydrogel. Adding xanthan gum after heating the copper salt solution promotes more complete mixing between the xanthan gum and copper ions.

[0014] Thirdly, the present invention provides the application of the copper ion hydrogel in the preparation of topical formulations for skin repair.

[0015] Preferably, the topical preparation is a drug used to promote collagen regeneration, accelerate the reconstruction of the skin barrier structure, and improve chronic skin damage.

[0016] The topical preparation can promote collagen regeneration, accelerate the reconstruction of the skin barrier structure, and improve chronic skin damage. Specifically, within 7 to 14 days after application to the skin wound site, it can significantly promote the synthesis of type I collagen, increasing its expression level by 18.7% to 26.9%, decrease the expression level of type III collagen by 29.7% to 37.6%, and significantly promote the synthesis of type IV collagen, increasing its expression level by 29.8% to 32.6%. This marks a key shift in skin wounds from rapid filling to strength reconstruction and firm epidermal adhesion.

[0017] Fourthly, the present invention provides the application of the copper ion hydrogel in the preparation of topical formulations for treating pet dermatitis-like skin diseases.

[0018] Preferably, the dermatitis is a type of dermatitis characterized by inflammatory response and barrier damage, such as seborrheic dermatitis, eczematous dermatitis, or contact dermatitis.

[0019] Furthermore, the seborrheic dermatitis is a fungal dermatitis.

[0020] Furthermore, the fungus is Malassezia furfur.

[0021] The topical preparation can be used for skin care in cases of inflammatory response accompanied by impaired skin barrier function, which includes skin conditions characterized by sebum secretion imbalance, disordered stratum corneum structure, or chronic irritation, preferably including seborrheic dermatitis, eczematous skin condition, or skin barrier damage related to contact irritation.

[0022] The topical skin care preparation exhibits good biocompatibility and safety. HE staining sections show that the copper ion hydrogel has no significant tissue toxicity, induces a controllable inflammatory response, degrades rapidly, and causes minimal irritation to local tissues.

[0023] The topical skin care preparation can significantly heal seborrheic dermatitis lesions within 14 days of use, with a healing rate of over 88%. It can be used as an adjunct treatment during the acute phase of seborrheic dermatitis in pets and as a daily care during the remission phase, in order to control the pathogenic fungal load, relieve erythema and itching, and reduce the risk of recurrence.

[0024] Compared with the prior art, the present invention has the following beneficial effects: On one hand, this invention utilizes a three-dimensional network structure formed by the synergistic interaction of xanthan gum and sodium hyaluronate to load copper ions. Copper ions can electrostatically attract and weakly coordinate with the carboxyl groups in the side chains of xanthan gum molecules, forming ion associations between the xanthan gum molecular chains, thereby stabilizing and regulating the gel network. This cross-linking process not only improves the mechanical strength of the hydrogel, making it less prone to collapse or loss during skin repair, but also allows for the continuous release of active copper ions during the critical stages of tissue regeneration. The released copper ions can stimulate fibroblast activity and promote the regeneration of collagen and extracellular matrix, thereby accelerating the reconstruction of skin tissue structure. Simultaneously, copper ions also exhibit significant antifungal activity, capable of disrupting the cell membrane structure of Malassezia furfur, thus effectively inhibiting its growth and reproduction, and effectively suppressing the excessive proliferation of Malassezia furfur, a common cause of seborrheic dermatitis in dogs and cats. It avoids the drug resistance problems that may arise from traditional antibiotic preparations, demonstrating excellent anti-Malassezia furfur activity and exhibiting high safety and application stability. On the other hand, ingredients such as sodium hyaluronate, glycerin, butylene glycol, and betaine have excellent moisturizing and skin barrier repair effects, which can significantly increase skin hydration and improve common problems in seborrheic dermatitis such as barrier damage, dryness, and microecological imbalance. Panthenol and allantoin can promote skin tissue repair and healing, relieve inflammatory responses, and improve clinical symptoms such as itching, peeling, and oiliness. Furthermore, the film-forming properties of the hydrogel can effectively reduce the impact of external stimuli on wounds or damaged skin, enhance skin barrier function, and improve chronic skin lesions caused by long-term inflammation and barrier damage. Therefore, the copper ion hydrogel of this invention has antifungal, anti-inflammatory, moisturizing, and skin repair effects, while also possessing good biocompatibility and high stability. It can be widely used in various fields such as skin injury care, recovery from chronic skin lesions, postoperative tissue regeneration, and repair of damaged barrier function. It can also be used to improve seborrheic dermatitis, chronic skin inflammation, sebum secretion imbalance, and follicular microenvironment disorder caused by excessive growth of Malassezia furfur in pets, as well as the long-term management of sensitive skin, showing good application prospects and promotional value.

[0025] The preparation method of copper ion hydrogel of the present invention is simple, mild, requires no complicated operation or organic solvent, is safe and reliable, has low cost, and is easy to industrialize and promote. Attached Figure Description

[0026] To more clearly illustrate the technical solutions in the examples of the present invention or the prior art, the drawings used in the description of the examples or the prior art will be briefly introduced below. Obviously, the drawings described below are some examples of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0027] Figure 1This study compares the inhibitory effects of different copper sources on Malassezia furfur and measures the minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MFC).

[0028] Figure 2 A is a schematic diagram of the preparation process of copper ion hydrogel and a physical morphology display; A is a schematic diagram of the preparation process of copper ion hydrogel; B~D are physical photographs of the copper ion hydrogel, including the appearance of the copper ion hydrogel in a container (B), the shape retention under tilt (C), the moldable / writeable state on the surface of a petri dish, and a schematic diagram of extrusion by a syringe (D).

[0029] Figure 3 This is a scanning electron microscope (SEM) schematic diagram of the copper ion hydrogel of the present invention.

[0030] Figure 4 Cu, a copper ion hydrogel 2+ Characterization of release behavior, rheological properties, and antibacterial effect. A represents Cu in a copper ion hydrogel. 2+ A) In vitro release curve; B) Apparent viscosity changes of copper ion hydrogels in Examples 1 and 2 at different shear rates; C) Colony growth at different time points after copper ion hydrogel treatment (0~4 h); D) Statistical bar chart of colony survival rate on plates.

[0031] Figure 5 This is a schematic diagram of the subcutaneous injection experiment of the copper ion hydrogel of the present invention in guinea pigs; wherein, A is a subcutaneous anatomical diagram of the guinea pig abdomen, and B is an HE staining result of the skin tissue in contact with the copper ion hydrogel in the subcutaneous tissue of the guinea pig abdomen.

[0032] Figure 6 This invention demonstrates the therapeutic effect of copper ion hydrogel on seborrheic dermatitis caused by Malassezia furfur; wherein, A represents the process of skin lesion changes in guinea pigs with seborrheic dermatitis; and B represents the statistical results of the skin lesion area in guinea pigs.

[0033] Figure 7 Histological observation and quantitative analysis of type I collagen expression in skin tissues of different treatment groups. A shows representative images of tissue sections from the control group, Example 1, and Example 2 at 7 and 14 days, with red boxes indicating sampling areas and green boxes indicating positive signal areas; B shows the statistical results of the proportion of positive signals in each group at the corresponding time points, where ns indicates no statistically significant difference, and * and ** indicate statistically significant differences (p<0.05 and p<0.01), respectively.

[0034] Figure 8Histological observation and quantitative analysis of type III collagen expression in skin tissues of different treatment groups. A shows representative images of tissue sections from the control group, Example 1, and Example 2 at 7 and 14 days, with red boxes indicating sampling areas and green boxes indicating positive signal areas; B shows the statistical results of the proportion of positive signals in each group at the corresponding time points. ns indicates no statistically significant difference, and * and ** indicate statistically significant differences (p<0.05 and p<0.01), respectively.

[0035] Figure 9 Histological observation and quantitative analysis of type IV collagen expression in skin tissues of different treatment groups. A shows representative tissue sections from the control group, Example 1, and Example 2 at 7 and 14 days, with red boxes indicating sampling areas and green boxes indicating positive signal areas; B shows the statistical results of the proportion of positive signals in each group at the corresponding time points. ns indicates no statistically significant difference, and * and ** indicate statistically significant differences (p<0.05 and p<0.01), respectively. Detailed Implementation

[0036] The present invention will be further illustrated below with reference to specific embodiments. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the invention. Furthermore, it should be understood that after reading the teachings of this invention, those skilled in the art can make various alterations or modifications to the invention, and these equivalent forms also fall within the scope defined by the appended claims.

[0037] The following examples use instruments and equipment conventional in the art. Experimental methods in the following examples, unless otherwise specified, are generally performed under conventional conditions or as recommended by the manufacturer. All raw materials used in the following examples are conventional commercially available products with specifications conventional in the art. In this specification and the following examples, unless otherwise specified, "%" represents a percentage by mass.

[0038] Example 1 A copper ion hydrogel, prepared from the following raw materials by weight percentage: 0.08% copper sulfate, 2% xanthan gum, 0.5% sodium hyaluronate, 3% butylene glycol, 6% glycerin, 1% betaine, 3% panthenol, 0.1% allantoin, with the balance being water. like Figure 1 As shown, the preparation method of the copper ion hydrogel includes the following steps: S1. Dissolve copper sulfate in sterile deionized water to obtain a copper sulfate solution; S2. Heat the copper sulfate solution to 40 degrees Celsius in a water bath, then add xanthan gum to the copper sulfate solution and stir at 60 degrees Celsius for 12 hours to obtain copper sulfate-xanthan gum hydrogel. S3. Add sodium hyaluronate, butylene glycol, glycerin, betaine, panthenol, and allantoin to the copper sulfate-xanthan gum hydrogel in sequence, adjust the pH to 6.0, and stir at 60℃ for 12 hours to obtain copper ion hydrogel.

[0039] Example 2 A copper ion hydrogel, prepared from the following raw materials by weight percentage: 0.05% copper sulfate, 4% xanthan gum, 0.1% sodium hyaluronate, 5% butylene glycol, 10% glycerin, 0.5% betaine, 1% panthenol, 0.05% allantoin, with the balance being water.

[0040] The preparation method of the copper ion hydrogel is as described in Example 1.

[0041] Example 3 A copper ion hydrogel, prepared from the following raw materials by weight percentage: 0.02% copper sulfate, 8% xanthan gum, 1% sodium hyaluronate, 0.5% butylene glycol, 0.1% glycerin, 0.2% betaine, 5% panthenol, 1% allantoin, with the balance being water.

[0042] The preparation method of the copper ion hydrogel is as described in Example 1.

[0043] Example 4 A copper ion hydrogel, prepared from the following raw materials by weight percentage: 0.03% copper sulfate, 4% xanthan gum, 0.4% sodium hyaluronate, 9% butylene glycol, 0.5% glycerin, 0.8% betaine, 9% panthenol, 0.3% allantoin, with the balance being water. The preparation method of the copper ion hydrogel is as described in Example 1.

[0044] Example 5 A copper ion hydrogel, prepared from the following raw materials by weight percentage: 0.1% copper sulfate, 0.6% xanthan gum, 0.7% sodium hyaluronate, 3.5% butylene glycol, 10% glycerin, 0.4% betaine, 0.8% panthenol, 0.2% allantoin, with the balance being water. The preparation method of the copper ion hydrogel is as described in Example 1.

[0045] The prepared copper ion hydrogel is characterized below.

[0046] 1. Compare the inhibitory effects of different copper sources at different concentrations on the growth of Malassezia furfur. To compare the inhibitory effects of different copper sources on Malassezia furfur, a 96-well plate microdilution method was used to screen for copper sulfate (CuSO4), copper chloride (CuCl2), copper gluconate (Glu-Cu), and copper citrate at varying concentrations. Figure 1 In the table, A represents the treatment results of copper sulfate (CuSO4) and copper gluconate (Glu-Cu), and B represents the treatment results of copper chloride (CuCl2) and copper citrate (Cu3(C6H5O7)2·xH2O). The color changes within the pores reflect the metabolic activity and growth status of the microorganisms, with the blue boxes indicating representative concentration ranges showing significant inhibitory effects.

[0047] The results showed that the MICs (minimum inhibitory concentrations) of copper sulfate (CuSO4), copper chloride (CuCl2), copper gluconate (Glu-Cu), and copper citrate were 0.5 mM, 2 mM, 1 mM, and 4 mM, respectively. Further investigation was conducted using copper sulfate, which exhibited the best antibacterial effect, for plating to determine the minimum fungicidal concentration (MFC). The control group (CK) was untreated, and 1 / 2 MIC, 1 MIC, and 2 MIC corresponded to different copper ion concentrations and their respective colony growth. The results showed that the number of colonies decreased significantly with increasing CuSO4 concentration; when the concentration reached 1 mM, no visible colonies were observed on the plate, indicating that this concentration reached the MFC. Considering both liquid dilution and plating results, CuSO4 showed the best inhibitory and bactericidal effect against Malassezia furfur, with an MIC of 0.5 mM and an MFC of 1 mM.

[0048] 2. Characterization of the injection molding properties of copper ion hydrogels: The preparation process of the copper ion hydrogel in Example 1 is as follows: Figure 2 As shown. Figure 2 The specific morphology and flow state of the copper ion hydrogel in Example 1 at a 45° angle can be observed. Figure 2 (B and C), and then Example 1 was loaded into a 5 mL syringe with a needle and injected onto a culture dish, demonstrating that the resulting hydrogel has good transparency, morphological stability and extrudability and coatability.

[0049] 3. Microscopic morphological observation of copper ion hydrogels: The copper ion hydrogels from Examples 1 and 2 were placed in sterile petri dishes and pre-frozen at -80°C for 12 h. After pre-freezing, the samples were transferred to a vacuum freeze-dryer and dried continuously at -50°C and 10 Pa for 48 h. The freeze-dried samples were then rapidly fractured in liquid nitrogen, and cross-sectional samples were obtained using tweezers and adhered to conductive tape for electron microscopy. After gold sputtering, their microscopic morphological structure was observed using a scanning electron microscope.

[0050] Result: As Figure 3 The results show that the copper ion hydrogel exhibits a dense and uniform surface structure, with a more compact and slightly disordered pore structure. Some areas have smaller pore sizes, and the pore morphology is complex, exhibiting obvious layering characteristics. This wrinkled, rough-surfaced microstructure suggests that it may possess higher mechanical strength, endowing it with excellent morphological stability and effectively protecting wounds.

[0051] 4. Investigating the Cu in copper ion hydrogels 2+ release: Take 1 mL of the copper ion hydrogel from Example 1, place it in a dialysis bag, and immerse it in 100 mL of sterile ultrapure water. Vortex at a constant speed, and collect 1 mL of the immersion solution every 2 hours. Analyze the Cu in the immersion solution using ICP-AES. 2+ The amount.

[0052] Result: As Figure 4 As shown in Figure A, 0~4 h Cu 2+ The release rate is the fastest, reaching over 60% by the 4th hour, Cu 2+ It was rapidly released from the copper ion hydrogel. Subsequently, Cu 2+ The release curve shows a relatively slow and continuous trend. At 16 hours, Cu... 2+ Release rate exceeds 80%. Cu was observed. 2+ The release continues, indicating that the copper ion hydrogel can continuously release Cu. 2+ And it has the ability to continuously release Cu 2+ Its sustained-release properties.

[0053] 5. Analyze the rheological properties of copper ion hydrogels: The copper ion hydrogel samples of Examples 1 and 2 were measured using a rotational rheometer on a 25 mm diameter plate with a spacing of 1.5 mm and the test temperature was kept constant at 25 °C within the frequency range of 0 to 100 Hz.

[0054] Result: As Figure 4As shown in Figure B, under high strain, the copper ion hydrogel exhibits shear-thinning behavior, meaning that the viscosity of the copper ion hydrogel decreases with increasing shear rate. This implies that the copper ion hydrogel possesses suitable mechanical properties to withstand and adapt to the mechanical environment of human tissue, allowing it to be easily applied to uneven wound surfaces and quickly adapt to the shape of the wound, achieving comprehensive coverage of irregular wound areas.

[0055] 6. Investigate the antibacterial activity of copper ion hydrogels against Malassezia furfur: A bactericidal experiment was conducted using copper ion hydrogel from Example 1. Prepare 10... 7 CFU / mL of Malassezia furfur was inoculated onto a copper ion hydrogel. The mixture was centrifuged every 1 hour, and the collected cells were resuspended and spread on solid culture medium for fungal viability determination.

[0056] Result: As Figure 4 Results C and D showed that the bactericidal effect of copper ion hydrogel on Malassezia furfur was time-dependent, with the survival rate of Malassezia furfur gradually decreasing with increasing exposure time. After 3 hours of exposure, the survival rate of Malassezia furfur reached a stable level, showing a significant difference from other time-treated groups, with only a few colonies growing on the solid culture medium, indicating that most of the Malassezia furfur had been killed.

[0057] 7. Evaluate the biocompatibility of copper ion hydrogels: The biodegradability and biosafety of copper ion hydrogels were tested in guinea pigs. Guinea pigs were subcutaneously injected with 0.5 mL of copper ion hydrogels from Examples 1 and 2, respectively. Histological sections were excised and stained at 30 min, 1 day, and 3 days after treatment. Changes in local tissues of the guinea pigs after copper ion hydrogel injection were recorded by photography.

[0058] Result: As Figure 5As shown, a spherical protrusion appeared on the guinea pig's back immediately 30 minutes after injection. Dissection revealed a distinct viscous hydrogel within the subcutaneous mass, surrounded by fibrous tissue. In the early 30-minute stage, only a small number of neutrophils infiltrated around the two copper ion hydrogels. This was a physiological acute inflammation induced by the injection procedure, not due to severe toxicity of the copper ion hydrogel. In Example 2, the local vascular congestion was more pronounced after treatment with the copper ion hydrogel, indicating a slightly stronger early acute inflammation. On the first day after injection, the mass shrank, indicating the gradual degradation and absorption of the copper ion hydrogel material. Simultaneously, the infiltrating cells shifted to a predominantly macrophage-based structure with a small number of lymphocytes, indicating a transition from acute to chronic repair. This is a normal foreign body recognition response of the body to the implanted material; neither Example 1 nor Example 2 showed excessive inflammation with the copper ion hydrogels. On the third day after injection, no erythema or ulceration appeared at the injection site, and the guinea pig's subcutaneous tissue returned to normal. In Example 1, a small number of macrophages encapsulated the material around the injection site of the copper ion hydrogel, which accelerated its rapid degradation. In contrast, in Example 2, the injection site of the copper ion hydrogel showed less macrophage infiltration and a milder inflammatory response, but the copper ion hydrogel material retained its morphology more intact, and the degradation process was more gradual. Overall, the inflammatory responses induced by the two copper ion hydrogels in Examples 1 and 2 were within a controllable range, without serious tissue damage, cell degeneration, or tissue necrosis, demonstrating good biocompatibility.

[0059] 8. Evaluate the therapeutic effect of copper ion hydrogel on seborrheic dermatitis caused by Malassezia furfur: To verify the inhibitory effect of the hydrogel of this invention on skin lesions associated with Malassezia furfur, a model of increased Malassezia furfur load on the skin surface was established. Guinea pigs were anesthetized by intraperitoneal injection of a mixture of ketamine (50 mg / mL) and xylazine (8 mg / mL) in a 1:1 ratio, with a total volume of 0.1 mL. The hair on the back of each guinea pig was shaved with a pet-specific shaving tool, and any remaining hair was removed with depilatory cream. The back of the guinea pig was then wiped with cotton balls soaked in 75% alcohol. After drying, a 4 cm x 4 cm modeling area was marked with a ruler. The back of the guinea pig was rubbed with sandpaper until it bled but did not bleed. 200 L of Malassezia furfur suspension was evenly applied to the modeling area on the back of the guinea pig once a day for 7 consecutive days, and the model construction was monitored. After successful modeling, the modeling area of ​​the guinea pigs was treated once a day with the copper ion hydrogels of Examples 1 and 2, respectively, while the control group was treated with physiological saline, for 14 consecutive days. Photographs of the lesion sites were collected on day 1 (1 d), day 7 (7 d), and day 14 (14 d) to record the apparent changes, and the lesion areas were segmented, labeled, and their areas were statistically analyzed using image analysis.

[0060] Figure 6The results showed that in the control group, the erythema and scratching / erosion-like changes of the skin lesions worsened over time, with significant erythema and expansion of the lesion area still visible on day 14. All groups in this invention showed improvement in the appearance of the skin lesions. In Group 1, the erythema and lesion severity were reduced compared to the control group, although some traces remained on day 14. Group 2 showed the most significant improvement; the erythema area significantly decreased on day 7, and the lesions basically returned to a normal appearance or only left slight traces on day 14. Quantitative analysis of wound area: On day 0, there was no significant difference in wound area among the three groups, all ranging from 7 to 8 cm². 2 To ensure consistency in the initial degree of injury, the wound area in the control group increased to 9.27 cm² on day 7. 2 This indicates that the infection led to the progression of the injury. In Case 1, the wound area decreased to approximately 4.55 cm². 2 In Example 2, the height decreased to approximately 2.83 cm. 2 The wound area in both groups was significantly smaller than that in the blank control group, and the repair effect in Group 2 was even better. On day 14, the wound area in the blank control group reached 10.98 cm². 2 The injury continued to worsen. In Case 1, the wound area decreased to 0.827 cm². 2 In Example 2, the lesion area was almost completely healed (approximately 0 mm), and both Example 1 and Example 2 were significantly smaller than the blank control group. The healing rate of Example 1 reached 88.19%, and the healing rate of Example 2 reached 100%. Furthermore, the quantitative representation of the lesion area / distribution based on image segmentation showed that the lesion area was the largest in the blank control group, while the lesion areas in Example 1 and Example 2 were significantly reduced, with the smallest in Example 2. This indicates that the copper ion hydrogel of the present invention can effectively reduce the degree and extent of Malassezia furfur-related lesions on the skin surface under external care conditions, and has good application value.

[0061] 9. Analyze the effect of copper ion hydrogel on skin collagen content: Tissue samples were collected from skin lesions in guinea pigs of each treatment group on days 1 (d), 7 (d), and 14 (d). The tissue sections were immunohistochemically stained with specific antibodies against type I, type III, and type IV collagen to differentiate the expression and distribution of different collagen types. After staining, the sections were dehydrated, cleared, and mounted with neutral resin. Images were then observed and acquired under an optical microscope. ImageJ was used to calculate the percentage of blue collagen area in each field of view relative to the total dermal area, serving as a semi-quantitative indicator of collagen content for inter-group statistical comparison analysis.

[0062] Result: As Figure 7 As shown, compared with the saline control group, copper ion hydrogel can significantly regulate and accelerate the deposition of structural collagen in wounds.

[0063] (1) Type I collagen like Figure 7 As shown, on day 7, the collagen content in the control group was 55.6%, while there was no significant difference between Example 1 (58.6%) and the control group. Example 2 (70.2%) showed a highly significant difference compared to Example 1. By day 14, collagen levels in all three groups had significantly increased. The collagen content in the control group was 65.3%, an increase of 9.7%. Example 1 (85.5%) and Example 2 (88.9%) were significantly higher than the control group, with no significant difference between the groups. These results indicate that the copper ion hydrogels in Examples 1 and 2 can significantly promote the synthesis of type I collagen, increasing its expression level by 18.7%–26.9%, and the effect continues to increase over time.

[0064] (2) Type III collagen like Figure 8 As shown, on day 7, the positive rate in the control group was 85.2%, while Example 1 (95.2%) showed a highly significant difference from the control group, and Example 2 (88.5%) showed no significant difference from the control group, but there was a highly significant difference between Example 1 and Example 2 (*P<0.001). By day 14, the positive rates in all three groups had decreased significantly, with the control group at 68.8%, a decrease of 16.4%. Example 1 (65.5%) showed no significant difference from the control group, while Example 2 (50.9%) was significantly lower than the control group. This dynamic change indicates that type III collagen is actively synthesized in the early stages of inflammation, but decreases in the later stages as inflammation subsides. The collagen expression level decreased by 29.7%–37.6% after treatment with copper ion hydrogel. This suggests that type III collagen forms a rapid but weak scaffold in the early stages of healing. In the later stages, as wound healing transitions to the mature stage, type I collagen is synthesized in large quantities and gradually replaces type III collagen, becoming the main component, and tissue strength increases.

[0065] (3) Type IV collagen like Figure 9 As shown, on day 7, the positive rate in the control group was 32.6%, while there was no significant difference between Example 1 (48.3%) and the control group. Example 2 (62.6%) showed a highly significant difference compared to both the control group and Example 1. By day 14, the positive rates in all three groups had increased significantly, with the control group at 38.6%, and Example 1 (78.1%) and Example 2 (95.2%) being significantly higher than the control group. The results clearly indicate that copper ion hydrogel can significantly promote the synthesis of type IV collagen, increasing its expression level by 29.8%–32.6%. This increased expression level indicates good epidermal regeneration and basement membrane reconstruction.

[0066] Both copper ion hydrogel interventions in Examples 1 and 2 promoted collagen synthesis to varying degrees. The copper ion hydrogel in Example 2 showed superior long-term enhancement of type I and IV collagen, while the copper ion hydrogel in Example 1 was more prominent in early enhancement of type III collagen. The continuous increase in type I and IV collagen over time reflected the ongoing repair of skin tissue; type III collagen initially increased and then decreased, closely related to the dynamic changes during the inflammatory phase. From the perspective of the temporal and spatial dynamics of collagen subtypes, during the proliferative phase (day 7), the copper ion hydrogel showed faster and richer deposition of type III collagen (early, fine collagen), providing a rapid scaffold for granulation tissue. Figure 8 During the remodeling phase (day 14), copper ion hydrogels significantly promoted the synthesis and transformation of type I collagen (mature, strong collagen). Figure 7 ), and simultaneously enhanced the continuous expression of type IV collagen (a component of the basement membrane). Figure 9 This marks a crucial shift in wound healing from rapid filling to strength restoration and firm epidermal adhesion.

[0067] In summary, copper ion hydrogels directly promote collagen synthesis, transforming a temporary scaffold dominated by type III collagen into a mature structure dominated by type I collagen and a high-quality transformation of the intact basement membrane, thereby accelerating wound healing.

[0068] The above content is only for illustrating the technical concept of the present invention and should not be construed as limiting the scope of protection of the present invention. Any modifications made to the technical solution based on the technical concept proposed in this invention shall fall within the scope of protection of this invention.

Claims

1. A copper ion hydrogel, characterized in that, By mass percentage, the raw materials for its preparation include: 0.02%~0.1% soluble copper salt, 0.6%~8% xanthan gum, 0.1%~1% sodium hyaluronate, 0.5%~9% butylene glycol, 0.1%~10% glycerin, 0.02%~1% betaine, 0.8%~9% panthenol, and 0.05%~1% allantoin, with the balance being water.

2. The copper ion hydrogel according to claim 1, characterized in that, The raw materials for its preparation, by mass percentage, include: 0.05%~0.08% soluble copper salt, 2%~4% xanthan gum, 0.1%~0.5% sodium hyaluronate, 3%~5% butylene glycol, 6%~10% glycerin, 0.5%~1% betaine, 1%~3% panthenol, and 0.05%~0.1% allantoin, with the balance being water.

3. The copper ion hydrogel according to claim 1, characterized in that, The soluble copper salt is copper sulfate, copper gluconate, copper chloride, or copper citrate.

4. The method for preparing the copper ion hydrogel according to any one of claims 1 to 3, characterized in that, include: S1. Dissolve soluble copper salt in water to obtain a copper salt solution; S2. Add xanthan gum to the copper salt solution and mix well to obtain copper salt-xanthan gum hydrogel; S3. Add sodium hyaluronate, butylene glycol, glycerin, betaine, panthenol, and allantoin to the copper salt-xanthan gum hydrogel, and adjust the pH value to 5.8~7.2 to obtain copper ion hydrogel.

5. The use of the copper ion hydrogel according to any one of claims 1 to 4 in the preparation of topical formulations for skin repair.

6. The application according to claim 5, characterized in that, The topical preparation is a drug used to promote collagen regeneration.

7. The use of the copper ion hydrogel according to any one of claims 1 to 4 in the preparation of a topical formulation for treating pet dermatitis-like skin diseases.

8. The application according to claim 7, characterized in that, The dermatitis-related skin diseases mentioned are seborrheic dermatitis, eczematous dermatitis, or contact dermatitis.

9. The application according to claim 8, characterized in that, The seborrheic dermatitis mentioned is a type of dermatitis caused by fungi.

10. The application according to claim 9, characterized in that, The fungus is Malassezia furfur.

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