Fabrication of Electrospun Gelatin Composite Nanofibers and Its Relevant Application
Author:Deng Ling Li
Electrospinning has attracted more and more attention from various fields.Biopolymer nanofibers has been applied for bioactive and drug delivery,active package,tissue engeering et al.Gelatin,owing to its biocompatibility,biodegradability,and easy availability,is the most commonly used FDA approved biopolymer and has long been researched for electrospinning.The main limitation of gelatin nanofibers is the poor water-resistance.Cross-linking is necessary to prevent the rapid dissolution of gelatin nanofibers and to retain its three dimensional structure.Gelatin nanofibers can be crosslined by chemical and enzymatic methods.However,the toxicity of the cross-linkers should be taken into consideration before application for food contact or oral consumption.So,study of the non-toxic cross-linking method for gelatin nanofibers is crucial for application of gelatin nanofibers or other biopolymer nanofibers in food industry.Hence,we studied the effects of non-ionic Tween 80,anionic sodium dodecyl sulfonate(SDS)and cationic cetyltrimethyl ammonium bromide(CTAB)surfactants on the morphology of electrospun gelatin nanofibers,and on the release behavior,antioxidant activity and antimicrobial activity of encapsulated curcumin.Then we added a typical hydrophobic protein,zein,into the gelatin solution to compare the morphology,characteristics,and solvent resistance of gelatin,zein,and gelatin/zein films fabricated by electrospinning vs.solvent casting.We characterized the gelatin/zein nanofibers with various ratios of gelatin/zein.Moreover,gelatin/zein nanofibers cross-linked by glucose via Maillard reaction,which were further applied for controlled release and cranial bone regeneration.(1)Effects of surfactants on the formation of gelatin nanofibers for controlled release of curcuminThe effects of non-ionic Tween 80,anionic sodium dodecyl sulfonate(SDS),and cationic cetyltrimethyl ammonium bromide(CTAB)surfactants on the morphology of electrospun gelatin nanofibers and the release behavior,antioxidant and antimicrobial activities of the encapsulated curcumin were studied.Scanning electron micrographs showed that the addition of SDS significantly increased the nanofiber diameter.Fourier transform infrared and differential scanning calorimetry analysis indicated that gelatin and SDS intimately interacted via electrostatic and hydrophobic interactions.However,these interactions inhibited the release of curcumin from the nanofibers with SDS,while CTAB and Tween 80 both facilitated the release.SDS and Tween 80 showed protective effects on curcumin from the attack of 2,2-diphenyl-1-picryl-hydrazyl-hydrate(DPPH)radicals,and the increased release of curcumin from nanofibers with CTAB or Tween 80 resulted in a higher reducing power.The antimicrobial activity results suggested that the curcumin encapsulated gelatin nanofibers with CTAB exhibited effective inhibition against Staphylococcus aureus.(2)Characterization of gelatin/zein films fabricated by electrospinning vs.solvent castingThe gelatin,zein,and gelatin/zein films were fabricated by electrospinning and solvent casting,respectively.Scanning electron microscopy,differential scanning calorimetry,Fourier transform infrared spectroscopy and water contact angle measurements were performed to characterize the morphology,molecular interactions,thermal behavior,and surface hydrophilicity of the electrospun and casted films.Results showed that the zein particles prevented the gelatin chains from aggregating intra-molecularly in the casted films due to heterogeneous mixing,while strong hydrogen bonding was formed in the nanofibrous film as a result of homogeneous mixing.Interestingly,the gelatin/zein nanofibrous film had a hydrophobic surface(water contact angle of 118.0o),while the casted gelatin/zein film had a hydrophilic surface(water contact angle of 53.5o).The confocal laser scanning microscopy observations suggested that the zein particles were well dispersed in the gelatin network to maintain the nanofibrous structures after immersion in water or ethanol,leading to the improved solvent resistance.(3)Characterization of gelatin/zein nanofibers by hybrid electrospinningThe gelatin/zein nanofibers by hybrid electrospinning were fabricated.The morphology observations by field emission scanning electron microscopy showed that the diameter of gelatin/zein nanofibers was significantly increased with the increasing gelatin ratio.The results of Fourier transform infrared spectroscopy and differential scanning calorimetry indicated that gelatin and zein strongly interacted via hydrogen bonding.The homogeneous mixing resulted in the crystalline structure for the gelatin/zein nanofiber at a 1:1 weight ratio,which showed a hydrophobic surface(water contact angle of 118°).The mechanical tests showed that the gelatin/zein nanofibers with weight ratios of 1:3 and 1:2 gave rise to much higher elongation at break of 87.9% and 69.0%,indicating good deformability and flexibility.In contrast to the pure gelatin or zein nanofibers,the gelatin/zein fibers preserved the 3D porous structures after immersed in water or ethanol for 24 h.(4)Study on wettability,mechanical property,and biocompatibility of electrospun gelatin/zein nanofibers cross-linked by glucoseThe gelatin/zein nanofibers were electrospun and then cross-linked by glucose via Maillard reaction.The morphology observations showed that the average diameter of the cross-linked gelatin/zein nanofibers decreased with the increasing weight ratio of zein.FTIR results indicated that gelatin and zein molecules were homogenously dispersed within the fibers,and the Maillard reaction occurred between the proteins and glucose.The addition of zein affected the mobility of the gelatin chains,leading to the decreased denaturation temperature.The cross-linked fibers with a 1:1 gelatin/zein ratio showed the most hydrophobic surface(water contact angle of 134°),and comparable mechanical properties to the cross-linked gelatin nanofibers,while a higher ratio of zein(more than 50%)resulted in the decreased elastic modulus.The L929 cell cytotoxicity test suggested that the cross-linked gelatin/zein nanofibers exhibited good biocompatibility and non-cytotoxicity.(5)Encapsulation of allopurinol by glucose cross-linked gelatin/zein nanofibers: characterization and release behaviorVarious concentrations of allopurinol were encapsulated into glucose cross-linked gelatin/zein(1:1,w/w)nanofibers.Encapsulation of allopurinol had no negative influence on the production of smooth and uniform nanofibers.Fourier infrared spectroscopy(FTIR)and differential scanning calorimeter(DSC)revealed the hydrogen bonding interactions between allopurinol and proteins,and the homogenous dispersion of allopurinol at concentrations of 2.5%(A2.5),5%(A5),and 10%(A10),while phase separation happened with a concentration of 20%(A20)of allopurinol by recrystallization.The nanofiber wettability was significantly increased for the A2.5 and A5 fibers by the strong interactions among the components,but the wettability of the A10 and A20 fibers showed even lower wettability compared to the unencapsulated nanofibers due to the dispersion of excess allopurinol on the surface.The release experiments of allopurinol from nanofibers into phosphate buffer,simulated gastric fluid,and simulated intestinal fluid suggested the anomalous pattern for the burst release,followed by the Fickian diffusion for the sustained release.The allopurinol encapsulated nanofibers showed effective suppression against xanthine oxidase.The effective suppression against xanthine oxidase by allopurinol encapsulated nanofibers indicated its future potential for the treatment of gout vial oral administration or surgery implantation.(6)Reconstruction of rat cranial bone defects by cross-linked gelatin/zein nanofibrous scaffoldsThe glucose cross-linked gelatin/zein nanofibrous scaffolds using the Maillard reaction were fabricated.The purpose of this study was to evaluate the cross-linked scaffolds for bone regeneration in vitro and in vivo.The nanofibrous scaffolds exhibited fast mineralization in concentrated simulated body fluid,and dicalcium phosphate dehydrate(DCPD)and octacalcium phosphate(OCP)were formed.A CCK-8 test indicated that the scaffolds had no cytotoxic effects on MC3T3-E1 cells.Additionally,SEM and CLSM images revealed that all the scaffolds were biocompatible and showed excellent support for MC3T3-E1 cells.In osteogenesis characterizations,Alizarin Red staining experiments revealed the improved calcium deposits on the cross-linked scaffolds,while the pure gelatin scaffold showed the best effect.In a rat cranial bone regeneration experiment,the cross-linked scaffolds exerted a strong positive effect on cranial bone regeneration with increased new bone volume and connective tissue formation,but the incorporation of zein in the gelatin scaffolds did not favor the bone regeneration effect.In inflammatory cytokine analysis,the cross-linked gelatin group exhibited significantly higher levels of IL-1β,promoting the differentiation of osteoblasts,while higher levels of IFN-γ and lower levels of IL-6 restricted the differentiation of osteoclasts.The study fabricated gelatin/zein nanofiber by hybrid electrospinning and crosslinked nanofibers by Maillard reaction,which were extensively applied for controlled release and rat cranial bone reconstruction.