Antibiotic-free hydrogel for nursing staphylococcus aureus infection as well as preparation method and application of antibiotic-free hydrogel

Abstract

The invention discloses antibiotic-free hydrogel for nursing staphylococcus aureus infection as well as a preparation method and application of the antibiotic-free hydrogel. The hydrogel is prepared from metal ion compounds such as zinc ions, copper ions and iron ion compounds, a natural high polymer material and an antioxidant as raw materials, and has the advantages of efficient antibacterial property, high safety, long-acting slow release and the like. An in-vitro antibacterial experiment shows that the sterilizing rate of the metal ion compound on staphylococcus aureus can reach 90%-99.9%; animal experiments prove that the hydrogel can effectively relieve inflammatory reaction of infected parts, and skin irritation experiments show that the product is free of irritation and anaphylactic reaction. The invention provides a safe and effective nonreactive solution for nursing of staphylococcus aureus infection.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine, and in particular relates to an antibiotic-free hydrogel for treating Staphylococcus aureus infection, and a preparation method and application thereof. Background Art

[0002] Staphylococcus aureus is an extremely common and highly harmful pathogen. It is widely distributed in the natural environment and in areas of the human body, such as the skin and nasal cavity, and can cause a variety of serious infectious diseases. Common skin and soft tissue infections include furuncles, carbuncles, and cellulitis, which cause patients to experience localized symptoms such as redness, swelling, heat, and pain. Among lung infections, Staphylococcus aureus pneumonia is often more severe, especially for those with weakened immune systems, such as the elderly, children, and long-term hospitalizations, and can lead to serious consequences such as respiratory failure. In sepsis, Staphylococcus aureus invades the bloodstream and multiplies rapidly, releasing toxins and triggering a systemic inflammatory response, resulting in a high mortality rate.

[0003] Currently, clinical treatment for Staphylococcus aureus infections primarily relies on antibiotics. However, the long-term, widespread, and even overuse of antibiotics has led to a serious problem of bacterial resistance. Methicillin-resistant Staphylococcus aureus (MRSA), for example, is resistant to multiple commonly used antibiotics, significantly increasing the difficulty of clinical treatment, increasing treatment costs, prolonging hospital stays, and worsening prognosis. Furthermore, antibiotic treatment can also trigger numerous adverse reactions, such as intestinal dysbiosis, leading to digestive symptoms like diarrhea and abdominal pain, disrupting the normal microbial balance in the intestines. Some patients may also experience allergic reactions, ranging from mild rashes to severe anaphylactic shock, which pose a serious threat to their health and life. Therefore, the development of safe, effective, and antibiotic-free care products to combat Staphylococcus aureus infections is an extremely urgent clinical need and holds great market potential.

[0004] While some existing non-antibacterial products, such as common disinfectants, can kill bacteria to a certain extent, their effect is often short-lived and they are highly irritating to the skin, potentially leading to dryness and peeling. While antimicrobial dressings can form a protective barrier on the wound surface, their antimicrobial effectiveness is limited, making it difficult to effectively inhibit the continued growth and reproduction of Staphylococcus aureus. Furthermore, some dressings have poor air permeability, which can easily lead to localized moisture in the wound, favoring bacterial growth.

[0005] Therefore, it is necessary to design an antibacterial material with long-lasting sustained-release antibacterial properties, high activity against drug resistance, and mild and skin-friendly properties. Summary of the Invention

[0006] In order to overcome the shortcomings of the above-mentioned prior art, the purpose of the present invention is to provide an antibiotic-free hydrogel for the care of Staphylococcus aureus infection, as well as its preparation method and application. By introducing specific metal ion compounds into the hydrogel system and utilizing the inhibitory effect of metal ions on Staphylococcus aureus, an antibiotic-free and antibacterial effect is achieved, the slow release of antibacterial components is achieved, and the antibacterial effect is improved.

[0007] In order to achieve the above object, the present invention adopts the following technical solutions: One of the purposes of the present invention is: The present invention provides an antibiotic-free hydrogel for treating Staphylococcus aureus infection. The antibiotic-free hydrogel is prepared using metal ion compounds, natural polymer materials and antioxidants as raw materials, wherein the concentration of the metal ions is 0.5 mM to 5 mM.

[0008] Preferably, the metal ion compound is selected from at least one of a zinc ion compound, a copper ion compound, and an iron ion compound, and the concentration of the metal ion is 1 mM to 3 mM.

[0009] Further preferably, the zinc ion compound is selected from zinc sulfate, zinc chloride or zinc acetate.

[0010] Preferably, the natural polymer material is selected from carrageenan, xanthan gum, sodium alginate, carbomer, hyaluronic acid or collagen.

[0011] Preferably, the antioxidant is selected from ascorbic acid or tocopherol.

[0012] The second purpose of the present invention is: The present invention provides a method for preparing an antibiotic-free hydrogel for treating Staphylococcus aureus infection, comprising the following steps: S1, dissolving a metal ion compound and an antioxidant in water to prepare a mixed solution; S2, heating the mixed solution, and adding natural polymer materials to the mixed solution during the heating process, stirring until a stable colloid is formed, thereby preparing an anti-inflammatory hydrogel for treating Staphylococcus aureus infection.

[0013] Preferably, in step S1, the molar ratio of the metal ion compound to the antioxidant is (1-2):1.

[0014] Preferably, in step S2, the heating temperature is 40-60°C, and the amount of the natural polymer material added is 2%-5% of the mass of the mixed solution.

[0015] Preferably, in step S2, the stirring speed is 200-500 r / min, and the stirring time is 30-60 min.

[0016] The third purpose of the present invention is: The present invention proposes the application of the above-mentioned antibiotic-free hydrogel for treating Staphylococcus aureus infection, which is mainly used for treating skin and soft tissue infection or wound infection.

[0017] Compared with the prior art, the present invention has the following beneficial effects: The invention discloses a non-antibiotic hydrogel for nursing Staphylococcus aureus infection and its preparation method and application, a specific metal ion compound is introduced into the hydrogel system, the inhibitory effect of metal ions on Staphylococcus aureus is utilized, the effect of non-antibiotic is achieved, the slow release of antibacterial components is achieved, the antibacterial effect is improved, and the growth and reproduction of Staphylococcus aureus can be efficiently inhibited. Its antibacterial effect can be applied to a variety of common infection types infected by Staphylococcus aureus, including but not limited to skin and soft tissue infections, such as infections after various traumas, furuncles, carbuncles, etc.; and traumatic infections, such as local or systemic infections caused by Staphylococcus aureus such as surgical incision infections and burn wound infections. These infections are relatively common clinically and seriously affect the health and quality of life of patients. The hydrogel product of the present invention can be targeted and effectively nursed for these infections. DETAILED DESCRIPTION

[0018] In order to enable those skilled in the art to better understand the present invention, the technical solution of the present invention will be clearly and completely described below in conjunction with the embodiments of the present invention. Obviously, the embodiments described are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts should fall within the scope of protection of the present invention.

[0019] As a novel biomaterial, hydrogels possess a unique three-dimensional network structure that absorbs and retains large amounts of water, exhibiting excellent moisture retention, which is crucial for maintaining the skin's normal physiological state and the wound healing environment. Furthermore, hydrogels exhibit excellent biocompatibility, reducing irritation to the skin and wounds, and lowering the likelihood of adverse reactions such as allergies. They also exhibit excellent sustained-release properties, slowly releasing loaded antimicrobial ingredients, prolonging their duration of action and enhancing their antimicrobial efficacy. Therefore, the use of hydrogels as carriers for the development of antibiotic-free products for the care of Staphylococcus aureus infections has significant research value and practical application potential.

[0020] Based on the above understanding, the present invention proposes an antibiotic-free hydrogel for treating Staphylococcus aureus infection, which is prepared using metal ion compounds, natural polymer materials and antioxidants as raw materials.

[0021] Among them, the selection of metal ion compounds is selected from at least one of zinc ion compounds, copper ion compounds, and iron ion compounds. These metal ions have varying degrees of inhibitory activity against Staphylococcus aureus, and zinc ion compounds are preferred because of their relatively high safety and good antibacterial effect. Common zinc ion compounds such as zinc sulfate can stably release zinc ions in aqueous solution, bind to certain key enzymes or proteins of Staphylococcus aureus, and interfere with the normal metabolism and growth process of bacteria; zinc chloride also has strong antibacterial ability, and its mechanism of action is similar to that of zinc sulfate, but it may show different antibacterial efficiencies under different environmental conditions; zinc acetate, due to the presence of acetate ions in its molecular structure, may synergize with zinc ions to exert antibacterial effects in some cases, and is relatively less irritating to the skin.

[0022] The hydrogel is composed of metal ion compounds, natural polymer materials, and antioxidants as raw materials to prepare the hydrogel. Among them, the natural polymer materials have good biocompatibility and degradability, providing a stable skeleton structure for the hydrogel. Common natural polymer materials include carrageenan, a polysaccharide extracted from red algae, which has unique gel properties and can form a stable gel system under mild conditions and is non-irritating to the skin; xanthan gum is a polysaccharide produced by microbial fermentation, and its aqueous solution has high viscosity and good stability, which can enhance the mechanical properties of the hydrogel; hyaluronic acid is widely present in human tissues and has excellent moisturizing properties and biological activity. It can promote cell proliferation and migration and is beneficial for wound healing; carbomer is a commonly used hydrogel matrix material that can quickly swell in water to form a gel and has good thickening and emulsifying properties; sodium alginate is a natural polysaccharide extracted from seaweed that can undergo cross-linking reactions with metal ions, further enhancing the stability of the hydrogel; collagen is an important component of human skin and connective tissue, has good biocompatibility and cell adhesion, and can provide a suitable microenvironment for cell growth and repair.

[0023] Antioxidants are added to improve the stability and bioactivity of the hydrogel and reduce the adverse effects of metal ions during oxidation. Common antioxidants include ascorbic acid (vitamin C), which has strong reducing properties and can scavenge free radicals in the system, preventing metal ion oxidation and hydrogel aging. Tocopherol (vitamin E) also has excellent antioxidant properties, protecting the bioactive components in the hydrogel from oxidative damage.

[0024] The present invention also provides a method for preparing the hydrogel, which specifically comprises the following steps: S1. Dissolve the metal ion compound and antioxidant in water to prepare a mixed solution, and heat it to 40-60°C. Within this temperature range, the metal ion compound and antioxidant can better dissolve and evenly disperse in the solution, which is also conducive to subsequent mixing with natural polymer materials. The molar concentration ratio of antioxidant to metal ion compound is 1:1-2:1. This ratio range has been verified by a large number of experiments and can fully exert the antibacterial activity of metal ions while ensuring the antioxidant effect. For example, when zinc sulfate is selected as the metal ion compound and ascorbic acid as the antioxidant, at this molar concentration ratio, the redox balance in the hydrogel system can be well maintained, thereby ensuring the stable performance of the hydrogel.

[0025] S2. Add the natural polymer material during the heating process and stir until a stable colloid is formed to obtain the non-resistance hydrogel. The amount of natural polymer material added is 2% to 5% of the mass of the mixed solution. If the amount added is too small, the strength and stability of the hydrogel may be insufficient; if the amount added is too large, the hydrogel may become too viscous, affecting its performance. During the stirring process, the stirring speed and time need to be controlled. Generally, the stirring speed is 200 to 500 rpm and the stirring time is 30 to 60 minutes to ensure that the natural polymer material can be fully dissolved and evenly mixed with the mixed solution to form a stable three-dimensional network structure.

[0026] The metal ion concentration range in the hydrogel is 0.5 mM to 5 mM, preferably 1 mM to 3 mM. Within this concentration range, the metal ions can significantly inhibit Staphylococcus aureus while not causing significant toxicity to human cells. In vitro antibacterial and cytotoxicity experiments have demonstrated that a metal ion concentration of 1 mM can achieve a bactericidal rate of over 90% against Staphylococcus aureus, with minimal effect on the activity of normal cells. When the concentration is increased to 3 mM, the bactericidal rate further increases to over 95%, while remaining within the safe concentration range.

[0027] The methods of the present invention are further illustrated below with reference to the examples. The following examples utilize conventional instruments and equipment in the art. Unless otherwise specified, the various raw materials and reagents used are conventional commercially available products, with specifications conventional in the art, or can be prepared or formulated by known methods or reagent instructions. Experimental procedures in the following examples, where specific conditions are not specified, generally follow conventional conditions or the conditions recommended by the manufacturer.

[0028] Example 1: Preparation of 1 mM zinc sulfate hyaluronic acid hydrogel Ingredients: Zinc sulfate, ascorbic acid, hyaluronic acid.

[0029] Preparation steps: S1. Dissolve zinc sulfate and ascorbic acid in sterile ultrapure water to prepare a mixed solution, which is then heated to 50°C. The zinc sulfate concentration is 1 mM, and the molar ratio of ascorbic acid to zinc sulfate is 1:1. During the dissolution process, stir using a magnetic stirrer at a speed of 300-500 rpm to ensure complete dissolution of the zinc sulfate and ascorbic acid.

[0030] S2. During the heating process, add hyaluronic acid powder at a concentration of 3% by weight of the mixed solution. Slowly and evenly sprinkle the powder into the mixed solution while stirring continuously for 30-60 minutes until a stable colloid is formed, yielding a 1 mM zinc sulfate hyaluronic acid hydrogel.

[0031] Example 2: Zinc chloride sodium alginate hydrogel The preparation process differed from that of the embodiment only in that zinc chloride of the same concentration was used instead of zinc sulfate, and an equal amount of sodium alginate was used instead of ascorbic acid.

[0032] Test Example 1: In vitro antibacterial experiment of metal ion compounds against Candida albicans Method: A single colony of Staphylococcus aureus was picked and inoculated into 5 mL of LB liquid medium. The suspension was cultured at 37°C and 180 rpm with shaking until the logarithmic growth phase. The concentration of the bacterial suspension was adjusted to 1×10 8 CFU / mL. Pour sterilized LB solid medium into a sterile culture dish. After the medium solidifies, use a pipette to draw 100 μL of the adjusted bacterial suspension and evenly spread it on the surface of the medium. Place it in a clean bench to air-dry for 5-10 minutes.

[0033] Using sterile tweezers, immerse 6 mm diameter sterile filter paper discs in solutions of metal ions of varying concentrations, a positive control antibiotic solution, and sterile deionized water. After soaking for 3–5 minutes, remove and drain any excess liquid. Place the filter paper discs evenly on the surface of the culture medium coated with the bacterial solution, placing four to five discs per culture dish, maintaining an appropriate distance between each disc. Incubate the culture dish upside down in a 37°C incubator for 18–24 hours. Observe and measure the diameter of the inhibition zone. Three parallel groups were set up for each experimental condition.

[0034] Results: Metal ion compounds at different concentrations exhibited antibacterial activity against Staphylococcus aureus, forming distinct zones of inhibition. The diameter of the inhibition zone gradually increased with increasing metal ion concentration. For silver nitrate at a concentration of 10 mg / mL, the inhibition zone diameter reached (22.5 ± 1.2) mm; for copper sulfate at the same concentration, the inhibition zone diameter was (18.3 ± 0.8) mm; and for zinc chloride at 10 mg / mL, the inhibition zone diameter was (15.6 ± 0.6) mm. The inhibition zone diameter for penicillin, the positive control, was (25.8 ± 1.5) mm, while no inhibition zone was observed in the negative control.

[0035] Test Example 2: In vitro antibacterial experiment of metal ion compounds against Staphylococcus aureus Method: Take the frozen Staphylococcus aureus strain and spread it on LB agar plate to activate the bacteria. After culturing at 37℃ for 24 hours, pick a single colony and inoculate it into 5 mL LB liquid culture medium. Incubate at 37℃ with shaking for 18-24 hours and adjust the bacterial solution concentration to OD600=0.5.

[0036] The bacterial suspension was mixed with different concentrations of metal ion compounds (zinc sulfate, copper chloride, and iron sulfate) and incubated at 37°C for 2 hours. The total number of colonies was counted using the serial dilution drop plate method, and the bactericidal rate was calculated. The specific procedure was as follows: the mixed bacterial suspension was serially diluted 10-fold, and 0.1 mL of the appropriate dilution was dripped onto LB agar plates. The plates were evenly spread with a sterile spreader. After incubation at 37°C for 24 hours, the number of colonies on the plates was counted, and the bactericidal rate was calculated according to the formula: Bactericidal rate (%) = (number of colonies in the control group - number of colonies in the experimental group) / number of colonies in the control group × 100%. Table 1 Grading standard of in vitro antibacterial efficacy of metal ions

[0037] Results: Zinc sulfate reached a strong level of efficacy at a moderate concentration and achieved a sterilization level effect at a high concentration, which was significantly better than other metal ions (see Table 1), fully demonstrating the strong inhibitory effect of metal ion compounds on Staphylococcus aureus.

[0038] Test Example 3: Experiment on the treatment of skin infection in rats using a non-antibiotic hydrogel product Model establishment: A skin injury model was established on the back of SD rats. A 1 cm diameter punch was used to create a circular wound on the back of the rats, reaching the dermis. Staphylococcus aureus (1×10 8 CFU / mL), and 0.1 mL of bacterial solution was inoculated into each rat wound to establish the infection model. Grouped Dosing: Model rats were randomly divided into three groups, with five rats in each group. The control group received normal saline; experimental group 1 received the zinc sulfate hyaluronic acid hydrogel of Example 1; and experimental group 2 received the zinc chloride sodium alginate hydrogel of Example 2. Dosing was performed twice daily for seven consecutive days. During administration, an appropriate amount of hydrogel was evenly applied to the wound surface to a thickness of approximately 1-2 mm.

[0039] Table 2 Effects of different treatment groups on healing of infected skin wounds in rats

[0040] Results: The control group experienced significant redness and swelling at the infected site, with abundant purulent secretions. Wound healing was slow, and by day 7 the wound had not yet fully healed, with only approximately 30% of the wound area healed. By day 7, the wounds in experimental group 1 had healed to approximately 60%, and in experimental group 2, to approximately 55%. The effects of the different treatment groups on the healing of infected skin wounds in rats (see Table 2) further demonstrate the therapeutic efficacy of the hydrogel product for skin infections. Test Example 4: Skin irritation test Methods: Healthy rabbits were selected and the hydrogels of Example 1 and Example 2 were applied to the depilatory area on their backs. The skin was observed for irritation reactions such as erythema and edema within 48 hours. Depilatory cream was used to remove hair, and the depilatory area was approximately 5 cm x 5 cm. The hydrogel was applied evenly in a layer approximately 1 mm thick. Skin reactions were observed 24 and 48 hours after application.

[0041] Results and Evaluation: Each animal's skin irritation response was scored according to Table 2, and the average score was calculated. Irritation intensity was evaluated according to Table 3. The scoring results are shown in Table 4.

[0042] Table 3 Rating criteria for skin irritation reactions

[0043] Table 4 Skin irritation reaction results of some examples

[0044] Results: All experimental animals showed no abnormal reactions such as erythema and edema on the skin of the application site, and the scores were all 0 points, indicating that the non-anti-hydrogel product of the present invention has no skin irritation.

[0045] In summary, in terms of high-efficiency antibacterial properties, the well-designed in vitro antibacterial experiment of the coating plate method was used to verify that about 10 6After incubating a suspension of S. aureus (CFU / mL) with hydrogels of different formulations for 12 hours, subsequent culture and counting revealed that the hydrogels containing specific metal ion compounds had a bactericidal rate of 90% to 99.9%. For example, the bactericidal rate of 1 mM zinc sulfate hydrogel reached 92%, increasing to 98% at 3 mM. The antibacterial mechanism is that metal ions disrupt bacterial cell membrane integrity and metabolic processes. Safety was verified by skin irritation tests (healthy rabbits, compared with saline, with no erythema or edema observed after 48 hours) and MTT cytotoxicity tests (the hydrogel extract did not affect cell viability), demonstrating its non-irritating, non-toxic, and excellent biocompatibility. The long-term sustained-release performance relies on a three-dimensional network structure. Using dialysis bags to simulate the in vivo environment, studies found that metal ions were released stably and continuously for 7 to 10 days, ensuring long-lasting antibacterial properties. The hydrogel can be formulated into a variety of dosage forms, including solution, colloid, and hydrogel dressings, suitable for both clinical and home use, meeting the care needs of S. aureus infections in different scenarios, including skin, soft tissue, and trauma.

[0046] The above content is only for explaining the technical idea of the present invention and cannot be used to limit the protection scope of the present invention. Any changes made on the basis of the technical solution in accordance with the technical idea proposed by the present invention shall fall within the protection scope of the claims of the present invention.

Claims

1. A non-antibiotic hydrogel for the care of Staphylococcus aureus infection, characterized in that The anti-oxidant hydrogel is prepared from metal ion compounds, natural polymer materials and antioxidants as raw materials, and the concentration of the metal ions is 0.5mM~5mM.

2. The non-antibiotic hydrogel for treating Staphylococcus aureus infection according to claim 1, characterized in that The metal ion compound is selected from at least one of a zinc ion compound, a copper ion compound, and an iron ion compound, and the concentration of the metal ion is 1 mM to 3 mM.

3. The non-antibiotic hydrogel for treating Staphylococcus aureus infection according to claim 2, characterized in that The zinc ion compound is selected from zinc sulfate, zinc chloride or zinc acetate.

4. The non-antibiotic hydrogel for treating Staphylococcus aureus infection according to claim 1, characterized in that The natural polymer material is selected from carrageenan, xanthan gum, sodium alginate, carbomer, hyaluronic acid or collagen.

5. The antibiotic-free hydrogel for treating Staphylococcus aureus infection according to claim 1, wherein The antioxidant is selected from ascorbic acid or tocopherol.

6. A method for preparing an anti-inflammatory hydrogel for treating Staphylococcus aureus infection, characterized in that: The following steps are involved: S1, dissolving a metal ion compound and an antioxidant in water to prepare a mixed solution; S2, heating the mixed solution, and adding natural polymer materials to the mixed solution during the heating process, stirring until a stable colloid is formed, thereby preparing an anti-inflammatory hydrogel for treating Staphylococcus aureus infection.

7. The preparation method according to claim 6, characterized in that In step S1, the molar ratio of the metal ion compound to the antioxidant is (1-2):

1.

8. The preparation method according to claim 6, characterized in that In step S2, the heating temperature is 40-60° C., and the amount of the natural polymer material added is 2%-5% of the mass of the mixed solution.

9. The preparation method according to claim 6, wherein In step S2, the stirring speed is 200-500 r / min, and the stirring time is 30-60 min.

10. An application of an anti-acid hydrogel for the treatment of Staphylococcus aureus infection, characterized in that: Used to treat skin and soft tissue infections or wound infections.

Images