Preparation And Properties Of Artemisinin-Loaded Silk Fibroin/Gelatin Composite Hydrogel

The unique three-dimensional structure and excellent physicochemical properties such as water retention and swelling of the hydrogel and the high efficiency of the drug make drug-loaded hydrogel widely used in biomedical fields.Research on the application of drug-loaded hydrogels in wound healing and tumor treatment has been a research hotspot.In the application of wound healing,the high water content of the drug-loaded hydrogel provides a wet healing environment and physical barrier for the wound,which is conducive to the recovery of wounds.The three-dimensional structure can promote the adsorption and growth of cells,and drugs can havesome specific effects in wound healing,such as resisting pathogenic microorganisms,inhibiting excessive inflammatory responses,and promoting the secretion of related cytokines.In terms of tumor treatment,hydrogel can be used as a carrier to transport drugs to the tumor site,and then control the drug release according to the set speed and concentration to achieve the purpose of maintaining effective drug concentration in the tumor site.This treatment scheme can reduce the side effects of oral administration of the drug and increase the bioavailability of the drug.Hydrogels can be made from natural or synthetic polymers.Hydrogels based on silk fibroin and gelatin,which are natural protein polymers,have good biocompatibility and have been used in wound healing and tumor treatment research.However,single-molecule?gelatin or silk fibroin?crosslinked network hydrogel is hard to satisfy the requirements of practical applications due to its high brittleness,rapid degradation,and poor stability at body temperature.Therefore,a composite hydrogel of silk fibroin and gelatin is considered to avoid these problems caused by a single hydrogel.Artemisinin is a sesquiterpene lactone compound with a variety of pharmacological activities,and has effects on antibacterial,anti-inflammatory and anti-tumor properties.Artemisinin has poor water solubility.How to load artemisinin into a hydrogel and achieve the effect of controlled drug release are problems to be solved.In this study,a cosolvent?polyethylene glycol 4000?was used to dissolve artemisinin to obtain a high-concentration artemisinin solution.The biological performance of the high-concentration artemisinin solution was tested and analyzed.The preparation of artemisinin-loaded silk fibroin/gelatin composite hydrogel is completed by the combination of high concentration artemisinin solution,silk fibroin and gelatin and cross-linking of genipin.The physicochemical properties and biological properties of prepared composite hydrogels were tested.The findings are as follows:1.Preparation of artemisinin solution and its biological performance testArtemisinin/polyethylene glycol 4000 solution was obtained by physical blending of artemisinin/ethanol solution and polyethylene glycol 4000 aqueous solution and natural volatilization.The solubility of artemisinin can reach 0.982±0.0889 mg/mL,which was 12 times higher than that of pure water.The prepared artemisinin solution had obvious inhibitory effect on bacteria and fungi and the effect on gram-negative bacteria was better.The effect of artemisinin solution on cells was dose-dependent,and experiments showed that artemisinin at a concentration of 0.031 mg/mL-0.063 mg/mL had no significant toxicity to normal cells.The IC₅₀ of the three types of hepatoma cells BEL-7404,SMMC7721,and HepG2 were 0.0016mg/mL,0.0084 mg/mL,and 0.0541 mg/mL,indicating that artemisinin had a good inhibitory effect on the three types of liver cancer cells.0.031 mg/mL artemisinin solution can inhibit lipopolysaccharide-induced NO production in mouse macrophages,showing anti-inflammatory properties.2.Preparation and performance evaluation of artemisinin-loaded silk fibroin/gelatin composite hydrogelThe prepared artemisinin solution was mixed with silk fibroin and gelatin to prepare artemisinin-loaded silk fibroin/gelatin composite hydrogel under the cross-linking by genipin.The preparation scheme of the composite hydrogel was optimized and the physical,chemical and biological properties were studied.The drug-loaded composite hydrogel had a stable three-dimensional porous structure with an average pore diameter of 83.25?m,and had good mechanical properties with compressive strength is 0.44 MPa and the tensile strength is 1.42 MPa.The addition of polyethylene glycol-assisted artemisinin improved the thermal stability of the composite hydrogel,but the swelling performance was reduced to 2.79 g/g.The release of artemisinin was pH-responsive.The release conditions within 24 hours fit well with the first-order release model,and the correlation coefficients is 0.9926?pH3.4?,0.9975?pH4.5?,0.9995?pH6.0?,0.9789?pH7.4?,indicating drug diffusion caused by swelling;the release of artemisinin within 24-120h was more in line with the zero-order kinetic equation and the correlation coefficients is 0.9505?pH3.4?,0.9124?pH4.5?,0.9587?pH6.0?,0.9755?pH7.4?.It is directly related to the erosion of the composite hydrogel.The artemisinin-loaded composite hydrogel exhibited good anti-inflammatory properties and could be applied in the study of wound healing.In addition,artemisinin-loaded composite hydrogel can inhibit the growth of human liver cancer cells HepG2 and SMMC7721 for a long time through the continuous release of artemisinin,indicating that the hydrogel had application value in cancer treatment.3.Wound healing test of artemisinin-loaded silk fibroin/gelatin composite hydrogelA full-thickness skin resection model of SD rats was established to evaluate the role of artemisinin-loaded silk fibroin/gelatin composite hydrogel in wound healing.In the process of wound healing,the hydrogel can absorb the exudate from the wound and provide a more favorable environment for wound healing.The artemisinin-loaded silk fibroin/gelatin composite hydrogel can reduce the infiltration of inflammatory cells,shorten the inflammation time,and accelerate wound recovery by slow-release artemisinin.And the wound healing degree?99.51%?was significantly greater than that of the control group?silk fibroin/gelatin composite hydrogel group,98.39%?and the blank control group?96.03%?when it was applied to the wound for 18 days.In addition,under the action of artemisinin-loaded silk fibroin/gelatin composite hydrogel,collagen fibers are arranged in an orderly manner during the recovery process of wound tissues,and cells,growth factors and epidermis are generated faster.Finally,it formed a complete skin structure,including epidermis,cuticle,hair follicles and sebaceous glands.All the test results showed that the artemisinin-loaded silk fibroin/gelatin composite hydrogel can accelerate the repair of damaged tissue and promote the formation of new tissue epidermis.4.Antitumor test of artemisinin-loaded silk fibroin/gelatin composite hydrogelThe role of artemisinin-loaded silk fibroin/gelatin composite hydrogel in the treatment of cancer was evaluated by establishing a balb/c-nude tumor model of HepG2 cells in nude mice.Artemisinin sustained-released by artemisinin-loaded silk fibroin/gelatin composite hydrogel can continue to act on tumor sites,inhibit tumor cell proliferation,and promote tumor cell apoptosis and tumor tissue necrosis.After 15 days of drug-loaded composite hydrogel treatment,the tumor volume growth rate was 47.62%,much lower than the control group?563.04%?and the blank group?494.42%?.The test results showed that the artemisinin-loaded silk fibroin/gelatin composite hydrogel can inhibit tumor growth.In general,the artemisinin-loaded silk fibroin/gelatin composite hydrogel prepared in this study has a stable internal structure and good comprehensive performance,has a significant role in wound healing and tumor treatment,and has a good application prospect in the field of biomedicine.