Properties Of Collagen Superfine Fiber In-situ Crosslinked By Citric Acid

Bovine tendon collagen is the main component of animal connective tissue,it's a kind of biological material with good biocompatibility,biodegradability and bioactivity.It has potential application in artificial skin,external dressing and other medical fields.The collagen electrospun fiber membrane has micro-nanometer fiber structure,which can mimic human cell interstitial structure,and has high cell compatibility.However,the electrospun collagen microfiber membrane is poor in water stability,swelling when contact with water or tissue fluid,thus losing the micro-nanofiber structure,affecting the cell proliferation and regeneration,greatly reducing its cell compatibility,this is also restricting its development in the field of biomedical.In order to improve the water stability of bovine tendon collagen static membrane in tissue fluid and to maintain the micro-nanometer fiber morphology of microfiber membrane,then to exert its structural advantage,we study the in-situ crosslinking bovine tendon collagen ultrafine fiber membrane by citric acid.The physical and chemical properties of collagen superfine fiber membrane were analyzed.Specifically,morphological changes before and after cross-linking of the ultra-fine fibers and after water treatment were characterized by SEM;Water solubility was used to characterize the water stability of the fiber;The cross-linking degree of collagen fibers was calculated under different cross-linking conditions,which was used to characterize the degree of cross-linking;The thermal properties of fiber membranes were characterized by DSC;FTIR was used to characterize the changes of the fiber structure and functional groups before and after the reaction.Finally,the biological properties of collagen fiber membranes were evaluated.In order to quantitatively evaluate the crosslinking effect of citric acid cross-linking,the cross-linking effect was evaluated,we also conducted a thermal cross-linking of collagen static film for comparison,illustrates the advantages of citric acid crosslinking.The contents of this paper include:Firstly,bovine tendon collagen was used as the raw material,polyoxyethylene(PEO)as the spinning aid,and the mixed solution of hexafluoroisopropanol(HFIP)and acetic acid(AA,50 vt.%)was used as the solvent to prepare different concentrations of collagen/PEO electrostatic spinning film.According to the observation of fiber morphology,the best electrospinning parameters corresponding to different concentration of spinning solution were obtained,and the morphology of electrospinning of collagen was observed under different concentration.The morphology of the fibers was analyzed by SEM,the mechanical properties were tested,spectral analysis and other performance evaluation and analysis.Secondly,the bovine tendon collagen film was thermally cross-linked for comparative analysis.The bovine tendon collagen electrospinning membrane was prepared by thermal cross-linking with 10% concentrated bovine tendon collagen.The properties of the bovine tendon collagen static-treated membrane and untreated bovine tendon collagen static-spun membrane were measured by infrared spectroscopy,DSC,mechanical strength test and water solubility test.The results showed that the thermo-crosslinked bovine tendon collagen film had a stronger thermal cross-linking time,smaller fiber diameter and stronger mechanical properties,but the water solubility did not improveBased on the properties of non-cross-linked and thermally cross-linked bovine tendon collagen films,we studied the spinnability and the optimum spinning process of the cross-linked bovine tendon collagen fibers in situ at different solution concentrations.Finally,in-situ crosslinking was carried out using a solution of 8% and 10% solution concentrations,the ratio of cross-linking agent citric acid was 5% and 10% of bovine tendon collagen,respectively,and the two solutions were spun,the obtained fiber membranes were subjected to infrared spectroscopy,DSC,water solubility test,mechanical property test and so on.The experimental results showed that with the increase of citric acid content,the diameter of superfine collagen fibers increased gradually.10% citric acid in situ crosslinked collagen fibers diameter of 523.7 ± 167 nm,immersed in PBS 48 h,the fiber morphology remained.The in-situ crosslinking of citric acid enhanced the mechanical properties of collagen-spun-film,and the addition did not affect the basic structure of collagen.The cross-linking degree of citric acid was 30.1%,compared with the degree of heat cross-linking have a greater advantage.The cross-linked collagen has a significantly lower water absorption and solubility than the un-crosslinked and thermally crosslinked samples.Finally,the biological properties of citric acid in-situ cross-linked collagen microfiber membrane were evaluated.The in vitro cytotoxicity test of 10% citric acid cross-linked collagen showed that there was no obvious bio-cytotoxicity of citric acid in situ crosslinked microfiber membrane.Animal acute toxicity test showed that citric acid in situ crosslinked collagen static film without biological acute toxic physiological symptoms.Hemolysis test showed that the addition of the crosslinking agent citric acid did not reduce the original hemolytic properties of the collagen.Therefore,citric acid in situ cross-linked collagen has excellent biological properties,the process is simple,low cost,in the field of biomedical feasibility.