The Research On Preparation And Application Of Keratin Compound Gel

Keratin is a fibrous structural protein with a large amount of cystine.It mainly exists in hair,wool,feathers,nails,and horns of mammals,reptiles and birds.Therefore,the source of keratin is abundant,and has good biocompatibility and degradability.In addition,the keratin peptide chain also has cell binding motifs,such as: glutamic acid-aspartic acid-serine(EDS),leucine Acid-aspartate-valine(LDV)and arginine-glycine-aspartate(RGD).These specific amino acid composition can promote cell proliferation,adhesion and transfer,etc.Moreover,keratin also has the function of rapid hemostasis and coagμlation,and can be well combined with the tissues.It can also promote the expression of type IV and VII collagen and stimμlate the migration of human keratinocytes.Keratin has been used as a biomaterial in the form of 2D and 3D scaffolds and hydrogels.However,keratin has poor processing and mechanical properties and is brittle.Usually,flexible polymer materials are used as cross-linking agents and plasticizers to improve these shortcomings of keratin,making it suitable for biomedical applications.Therefore,in this paper,firstly,by using gelatin and chitosan modified by gallic acid,the graft polymer obtained mimics the properties of mussels.Therefore,it can be chemically and physically cross-linked with keratin to prepare keratin composite hydrogel.and then chemically and physically cross-linked with keratin to prepare a keratin composite hydrogel.In addition,in order to improve the application of the keratin composite hydrogel,curcumin nanoparticles are loaded into the keratin composite hydrogel to improve the anti-inflammatory and antioxidant properties of the composite hydrogel.The main specific research contents are as follows:1.Extract keratin from experimental rabbit hair by reduction method;and use Fourier infrared spectroscopy(FT-IR)scanning,elementary binary chromatography(SD),sodium dodecyl s μlfate-polyacrylamide gel Electrophoresis(SDS-PAGE),amino acid analysis and Ellman reagents were used to characterize the extracted keratin.The resμlts showed that: 1)FT-IR detection of keratin amide A band and amide I,II,III bands were: 3295 cm-1,1650 cm-1,1536cm-1,1238cm-1.2)SD detects the secondary structure of keratin.The resμlts show that 90% of keratin isα-helix,and there are also a small amount of β-sheet and γ-keratin.3)The molecμlar weight of extracted keratin is mainly 45-60 KDa,which is mainly α-helix,of course,there are also a small amount of β keratin,the molec μ lar weight of which is 15-20 KDa,and the res μ lts are consistent with the SD scan resμlts of keratin.4)Pre-column derivatization was used to detect the composition of keratin amino acids using AQC as the derivatizing agent.The proportion of active amino acids in the entire keratin system was: arginine 11.61%,glycine 10.15%,aspartic acid1.81% Leucine 11.33%,valine 3.07%,and cysteine 2.7%.5)The content of sμlfhydryl in keratin detected by Ellman reagent is: 0.628 mmol / g.2.Preparation and Characterization of Gallic Acid Functionalized Gelatin(GA-g Gel)and Chitosan(GA-g-CS)Grafted onto chitosan and gelatin to obtain three gallic acid grafted chitosans(GA-g-CS-I,GA-g-CS-II,GA-g-CS-III)and three gallates Acid grafted gelatin(GA-g-Gel-I,GA-g-Gel-II,GA-g-Gel-III).Potential linear titration was used to determine the grafting rate of gallic acid in chitosan.The grafting rates of GA-g-CS-I,GA-g-CS-II,and GA-g-CS-III were 9.69± 0.56,%,39.60 ±0.34%,46.50 ±0.13% respectively.Folin phenol reagent calcμlates the total phenol content of the functionalized polymer,GA-g-CS-I,GA-g-CS-II,GA-g-CS-III are: 35.6±2mg/g,111.3 ± 1 mg / g,121 ± 3mg / g,GA-g-Gel-I,GA-g-Gel-II,GA-g-Gel-III are: 9.7 ± 4.1mg / g,39.6 ± 3.3mg / g,46.5 ± 2.9mg /g.According to the data,the carbodiimide method has the highest grafting rate,followed by the free radical method in an inert environment,and the lowest free radical method in an atmospheric environment.3.Preparation of keratin composite hydrogel.The obtained graft polymer with high grafting rate(prepared by carbodiimide method)was cross-linked with different amounts of keratin under the condition of oxidant sodium periodate.The keratin complex hydrogel is prepared by adding 10μl of 1% sodium periodate.And the gel time of the composite gel was measured by the tube inversion method.The gel time of the 15% keratin solution exceeds 24 hours.After adding functionalized gelatin and chitosan,the gel time is greatly reduced.The shortest gel time is 4.7min and 6.5 min.4.The gel properties of the composite hydrogels prepared with different proportions of keratin,functionalized chitosan and gelatin were detected by texture analyzer.The optimal specific gravity of the composite hydrogel is 2: 1(functional gelatin: keratin(w / w)),named GA-g-Gel-K hydrogel.The gel time is: 4 minutes and 54 seconds.The gel properties of this hydrogel are cohesion: 0.70 ± 0.03,elasticity: 0.37 ± 0.024,hardness: 1.7 ± 0.2N,adhesiveness: 0.60 ± 0.011 m J Compressive strength: 2.92 ± 0.13 k Pa.The best specific gravity of functionalized chitosan: keratin is 1: 7(W / W),named GA-g-CS-K hydrogel.The gel time is: 3 minutes and 30 seconds,the corresponding gel properties are cohesion: 0.83 ± 0.02,elasticity: 0.59 ± 0.017,hardness: 1.82 ±0.4N,adhesiveness: 0.60 ± 0.03 m J,compression Strength: 3.2 ± 0.03 k Pa.The porosity of the composite hydrogel at the optimal ratio was detected by ethanol replacement method.The porosity of keratin,GA-g-Gel-K hydrogel and GA-g-CS-K hydrogel is about 90.8 ± 0.9%,81.2 ± 0.5% and83.6 ± 0.7%.SEM analysis shows that the two kinds of composite gel networks have a uniform porous structure and the connection between the pores is relatively strong,which improves the problem that the keratin gel network easily collapses.The pores of the G-g-CS-K gel network are slightly larger than those of the G-g-Gel-K gel,indicating that the keratin ratio in the G-g-CS-K gel is relatively large,making the pore size of the hydrogel larger.These two hydrogels meet the performance indicators of wound adjuvant gels.5.Preparation of curcumin nanoparticles.The curcumin water-soluble sustained-release preparation is formed by chitosan ion gelation to form chitosan nanoparticles encaps μ lated curcumin(CUR-CSNPs),its particle size is 445.0 ± 16.8nm(PI = 0.306),zeta potential is 32 ± 1.4m V.Loaded into keratin complex hydrogel,can effectively release drugs for 6 days or more.6.The biocompatibility test of keratin complex hydrogel,the biocompatibility of keratin complex hydrogel by MTT method.The results showed that it had good biocompatibility.