Fabrication of gelatin/chitosan nanofibrous scaffold: process optimization and empirical modeling

Electrospinning is a very useful technique for producing polymeric nanofibers by applying electrostatic forces. This study reports on the modeling and optimization of the electrospinning process of gelatin/chitosan, using response surface methodology. The individual and the interaction effects of the gelatin/chitosan blend ratio (50/50, 60/40 and 70/30), applied voltage (20, 25 and 30 kV) and feeding rate (0.2, 0.4 and 0.6 mL h^?1) on the mean fiber diameter and standard deviation of the fiber diameter were investigated on optimization section, using scanning electron microscopy. To fabricate the nanofibrous gelatin/chitosan blend, trifluoroacetic acid/dichloromethane was selected as the solvent system. The model obtained for the mean fiber diameter has a quadratic relationship with applied voltage and feeding rate. The interaction between applied voltage and flow rate were found significant but the interactions of blend ratio and flow rate and also blend ratio and applied voltage were negligible. A quadratic relationship was obtained for applied voltage and flow rate with standard deviation of the fiber diameter and there was no interaction between the parameters in the model. The optimum condition for electrospinning of gelatin/chitosan was also introduced using the model obtained in this study. Scanning electron micrographs of human dermal fibroblast cells on the nanofibrous structures show good attachment and proliferation on the fabricated scaffold surface. Electrospun gelatin/chitosan nanofibrous mats have great potential for use as a scaffold for skin tissue engineering. © 2014 Society of Chemical Industry     

Improvement of mechanical properties and in vitro bioactivity of freeze-dried gelatin/chitosan scaffolds by functionalized carbon nanotubes

Functionalized multiwall carbon nanotubes (f-MWCNTs) were used to reinforce the freeze-dried gelatin (G)/chitosan (Ch) scaffolds for bone graft substitution. Two types of G/Ch scaffolds at a ratio of 2:1 and 3:1 by weight incorporated with 0.025, 0.05, or 0.1 and 0.2 or 0.4?wt% f-MWCNT, respectively, were prepared by freeze drying, and their structure, morphology, and physicochemical and compressive mechanical properties were evaluated. The scaffolds exhibited porous structure with pore size of 80–300 and 120–140?µm for the reinforced scaffolds of G/Ch 2:1 and 3:1, respectively, and porosity 90–93% which slightly decreased with an increase in f-MWCNTs content for both types. Incorporation of f-MWCNTs led to 11- and 9.6-fold increase in modulus, with respect to their pure biopolymer blend scaffolds at a level of 0.05?wt% for G/Ch 2:1 and 0.2?wt% for G/Ch 3:1, respectively. The higher content of f-MWCNTs resulted in loss of mechanical properties due to agglomeration. The highest value of compressive strength and modulus was obtained for G/Ch 2:1 with 0.05?wt% f-MWCNT as 411?kPa and 18.7?MPa, respectively. Improvement of in vitro bioactivity as a result of f-MWCNTs incorporation was proved by formation of a bone-like apatite layer on the surface of scaffolds upon immersion in simulated body fluid. The findings indicate that the f-MWCNT-reinforced gelatin/chitosan scaffolds may be a suitable candidate for bone tissue engineering.     

Preparation and characterization of porous scaffold composite films by blending chitosan and gelatin solutions for skin tissue engineering

Biodegradable polymers have significant potential in biotechnology and bioengineering. However, for some applications, they are limited by their inferior mechanical properties and unsatisfactory compatibility with cells and tissues. In the present investigation blends of chitosan and gelatin with various compositions were produced as candidate materials for biomedical applications. Fourier transform infrared spectral analysis showed good compatibility between these two biodegradable polymers. The composite films showed improved tensile properties, highly porous structure, antimicrobial activities, low water dissolution, low water uptake and high buffer uptake compared to pure chitosan or gelatin films. These enhanced properties could be explained by the introduction of free ? OH, ? NH₂ and ? NHOCOCH₃ groups of the amorphous chitosan in the blends and a network structure through electrostatic interactions between the ammonium ions (? NH³⁺) of the chitosan and the carboxylate ions (? COO^?) of the gelatin. Scanning electron microscopy images of the blend composite films showed homogeneous and smooth surfaces which indicate good miscibility between gelatin and chitosan. The leafy morphologies of the scaffolds indicate a large and homogeneous porous structure, which would cause increased ion diffusion into the gel that could lead to an increase in stability in aqueous solution, buffer and temperature compared to the gelatin/chitosan system. In vivo testing was done in a Wistar rat (Rattus norvegicus) model and the healing efficiencies of the scaffolds containing various compositions of chitosan were measured. The healing efficiencies in Wistar rat of composites with gelatin to chitosan ratios of 10:3 and 10:4 were compared with that of a commercially available scaffold (Eco‐plast). It was observed that, after 5 days of application, the scaffold with a gelatin to chitosan ratio of 10:3 showed 100% healing in the Wistar rat; however, the commercial Eco‐plast showed only a little above 40% healing of the dissected rat wound. Copyright © 2012 Society of Chemical Industry     

Crosslinking of electrospun fibrous gelatin scaffolds for apatite mineralization

Fibrous gelatin scaffolds fabricated via electrospinning followed by crosslinking were used as substrates for apatite mineralization. Gelatin macromolecules were confined by their fibers and further restricted by the crosslinked structure while proper flexibility could be attained upon hydration. After 4 or 5 days of mineralization, partially carbonated hydroxyapatite was proved to deposit uniformly on the surface of the fibers. The property of the substrate, such as stiffness of the scaffolds and flexibility of macromolecules chain, was changed by different crosslinking ways. The influences of these properties on the formation of apatite were also investigated. Results showed that a relatively less rigid interface and more flexible chain acquired by glutaraldehyde solution crosslinking seemed to favor the nucleation of minerals and to reduce the size of the inorganic products. © 2010 Wiley Periodicals, Inc. J Appl Polym Sci, 2011     

Synergistic effects of crosslinking and chitosan molecular weight on the microstructure,molecular mobility,thermal and sorption properties of porous chitosan/gelatin/hyaluronic acid scaffolds

In this study, synergistic effects of crosslinking and chitosan molecular weight on the microstructure, molecular mobility, thermal, and sorption properties of porous chitosan/gelatin/hyaluronic acid hybrid foams are reported. Fourier transform infrared spectroscopy has been utilized to confirm the covalent attachment of hyaluronic acid to gelatin and chitosan, and covalent chemical crosslinking between gelatin and chitosan. Detailed image analysis of scanning electron microscopy images of the porous scaffold hydrids reveal that the pore size of the materials formulated using either low‐ or high‐molecular‐weight chitosan increases significantly upon crosslinking using ethyl(dimethylaminopropyl) carbodiimide/N‐Hydroxysuccinimide. These microstructural changes are even more pronounced for the crosslinked hybrid scaffolds formulated using low‐molecular‐weight chitosan, highlighting a synergistic effect between crosslinking and the use of low‐molecular‐weight chitosan. Results obtained using differential scanning calorimetry demonstrate a significant reduction in molecular mobility reduction in molecular mobility for crosslinked scaffolds formed using high‐molecular‐weight chitosan compared to non‐crosslinked hybrids and crosslinked hybrids formulated using low‐molecular‐weight chitosan. Correspondingly, dynamic vapor sorption evidenced significantly lower water vapor sorption for crosslinked scaffolds formulated using high‐molecular‐weight chitosan. © 2017 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2017 , 134, 44772.     

pH-sensitive chitosan/gelatin hybrid polymer network microspheres for delivery of cimetidine

pH responsive chitosan/gelatin hybrid polymer network (HPN) microspheres were prepared via the inverse emulsion crosslinking method. Cimetidine release from the microspheres was studied. The drug only delivers in acidic medium, while the release rate can be controlled by the HPN composition and the degree of deacetylation.     

Fabrication and characterization of chitosan/gelatin porous scaffolds with predefined internal microstructures

The fabrication process for a novel three-dimensional (3D) chitosan/gelatin scaffold with predefined multilevel internal architectures and highly porous structures is presented combining solid freeform fabrication (SFF), microreplication and lyophilization techniques. The computer model of the scaffold is designed with biological data such as branching angle in liver vascular cast incorporated. Stereolithography (SL), known as a SFF technique, is utilized to build the resin mould, based on which poly-dimethylsilicone (PDMS) mould is produced by microreplication. The chitosan/gelatin hybrid solution is then cast onto the PDMS mould for pre-freeze and the monolayer porous structures with organized internal morphology are produced upon lyophilization. The 3D scaffold can be constructed via stacking these monolayer structures. The properties of porous structure, such as porosity, pore size and micromorphology as well as wall thickness, were investigated. Scanning electron microscopy (SEM) demonstrated that the scaffold possesses multilevel organized internal morphologies including vascular systems (portal vein, artery and hepatic vein) and parenchymal component (hepatocyte chamber). These organized structures enable orderly arrangement of hepatocyte and hepatic nonparenchymal cells and co-culture in the same 3D scaffold to guide liver regeneration in a controlled manner. Cell culture experiment in vitro showed that hepatocytes perform better in the well-defined chitosan/gelatin scaffold than in porous scaffold. This approach makes it flexible to investigate the relationship between internal scaffold microstructure and hepatocyte behavior in vitro. It also provides a new way to fabricate complex 3D scaffold using various natural biomaterials for vital organ engineering.     

A study on the fabrication of porous chitosan/gelatin network scaffold for tissue engineering

A novel porous chitosan/gelatin scaffold for tissue engineering was prepared via polyelectrolyte complex formation, freeze drying and post‐crosslinking with glutaraldehyde. The porosity and mean pore diameters could be controlled within 30∼100 µm by varying the original water content and the freezing conditions. Dipping the scaffolds in poly(lactic acid) provided good mechanical properties making it a promising candidate towards tissue engineering. © 2000 Society of Chemical Industry     

Preparation and characterization of novel chitosan/zeolite scaffolds for bone tissue engineering applications

In this study, chitosan-based novel scaffolds containing zeolite A were fabricated by freeze-drying technique. The nanocomposite scaffolds were prepared from chitosan and zeolite A nanocrystals with different amounts (0.5, 1.0, and 2.0%) in aqueous media. The zeolite A nanocrystals and nanocomposite scaffolds were characterized by using FTIR, X-ray powder diffraction, scanning electron microscope, and thermogravimetric analysis. The scaffolds were seeded with bone marrow-derived human mesenchymal stem cell line (UE7T-13), and cell attachment, viability, and cytotoxicity assays were performed. In vitro cytotoxicity of scaffolds toward human mesenchymal stem cell line was evaluated through the evaluation of cell viability and cell attachment assays.     

Novel pH,ion sensitive polyampholyte gels based on carboxymethyl chitosan and gelatin

Natural pH and ion sensitive polyampholyte gels were successfully prepared by blending carboxymethyl chitosan (CM‐chitosan) and gelatin using glutaraldehyde as crosslinking agent. Their swelling behaviour under the influence of pH and ionic strength of the solution has been studied, and molecular interaction and, morphology of the gels were investigated by infrared spectroscopy and scanning electron microscopy. At the isoelectric point (IEP) the blend gels shrunk most, and when pH deviated from the IEP they behaved as polycations or polyanions. A decrease in swelling degree (D_s) with rising NaCl concentration up to 0.15 M was observed for all gels, while addition of Ca²⁺ made them shrink due to formation of Ca²⁺ crosslinked bridges. The blended gels showed increasing sensitivity to pH and ionic strength as the weight fraction of CM‐chitosan increased. Two essential components seem to regulate the swelling behaviour of blended gel: the first is related to osmotic pressure difference caused by the redistribution of mobile ions; the second is related to the possible formation of interactions corresponding to hydrogen bonding and hydrophobic interactions. © 2003 Society of Chemical Industry     

Properties of polyelectrolyte complex films of chitosan and gelatin

A series of chitosan–gelatin complexes was prepared by varying the ratio of constituents. Differential scanning calorimetry was used to determine the amount of the different states of water. The interaction between chitosan and gelatin was checked by IR and X-ray analysis and was related to mechanical strength. The results indicate that the water take-up of a chitosan–gelatin complex is depressed by strong interactions within networks. Chitosan can improve the tensile strength of complex films, and even with high water content these can keep appropriate tensile strength and higher elongation. © 1999 Society of Chemical Industry     

MTGase反胶束纯化及其催化合成壳聚糖胶原改性物的展望

综述了微生物转谷氨酰胺酶（MTGase）的性质、用途及反胶束萃取纯化效果,展望了用MTGase催化合成等日化用新功能性原料的可行性及其前景。     

Hemocompatible polyurethane/gelatin-heparin nanofibrous scaffolds formed by a bi-layer electrospinning technique as potential artificial blood vessels

In this paper, a scaffold, which mimics the morphology and mechanical properties of a native blood vessel is reported. The scaffold was prepared by sequential bi-layer electrospinning on a rotating mandrel-type collector. The tubular scaffolds (inner diameter 4 mm, length 3 cm) are composed of a polyurethane (PU) fibrous outer-layer and a gelatin-heparin fibrous inner-layer. They were fabricated by electrospinning technology, which enables control of the composition, structure, and mechanical properties of the scaffolds. The microstructure, fiber morphology and mechanical properties of the scaffolds were examined by means of scanning electron microscopy (SEM) and tensile tests. The PU/gelatin-heparin tubular scaffolds have a porous structure. The scaffolds achieved a breaking strength (3.7±0.13 MPa) and an elongation at break (110±8%) that are appropriate for artificial blood vessels. When the scaffolds were immersed in water for 1 h, the breaking strength decreased slightly to 2.2±0.3 MPa, but the elongation at break increased to 145±21%. In platelet adhesion tests the gelatin-heparin fibrous scaffolds showed a significant suppression of platelet adhesion. Heparin was released from the scaffolds at a fairly uniform rate during the period of 2^nd day to 9^th day. The scaffolds are expected to mimic the complex matrix structure of native arteries, and to have good biocompatibility as an artificial blood vessel owing to the heparin release.     

Characterization of chitosan/citrate and chitosan/acetate films and applications for wound healing

In this work, we aimed to develop a scaffold of chitosan (CS) with a porous sponge structure for an artificial skin. The scaffolds were prepared from both CS/citric and CS/acetic solutions. In addition, the cast films were also prepared from the same solutions to compare some of their properties. They were characterized using WAXD, FTIR, DSC, tensile measurements, and SEM observation. It was found that CS/acetate had low crystallinity but CS/citrate was in an amorphous state, resulting in a large ductility with rubbery softness. Despite the different morphologies of CS/citrate and CS/acetate scaffolds, both scaffolds exhibited the wound healing effect available for tissue engineering. © 2008 Wiley Periodicals, Inc. J Appl Polym Sci, 2008     

3D mesoporous bioactive glass/silk/chitosan scaffolds and their compatibility with human adipose-derived stromal cells

Human adipose-derived stem/stromal cells (hASCs) have been popularly studied as cell-based therapy in the field of regenerative medicine due to their ability to differentiate into several cell types. In this study, in order to improve the mechanical strength and bioactivity of scaffolds for bone tissue engineering, three types of mesoporous bioactive glasses with different shapes and compositions were dispersed in the silk fibroin/chitosan (SF/CS)-based scaffolds, which were fabricated with a combination of freezing and lyophilization. The characteristic and physical properties of these composite scaffolds were evaluated. The biocompatibility was also assessed through hASCs in vitro tests. Both Alamar Blue® and Live/Dead assay® revealed that the spherical mesoporous bioactive glass doped scaffolds enhanced cell viability and proliferation. Furthermore, the addition of spherical mesoporous bioactive glass into SF/CS scaffolds encouraged hASC osteogenic differentiation as well. These results suggested that this composite scaffold can be applicable material for bone regeneration.     

Collagen‐based scaffolds reinforced by chitosan fibres for bone tissue engineering

In this paper, porous bone scaffolds reinforced with chitosan fibres were prepared. The porosity and pore size of the reinforced scaffolds were both satisfactory. The reinforced scaffolds resembled natural bone in both components and crystal size. Only if the length of the fibres was no shorter than the critical length, could the fibres reinforce the material. We have proposed an empirical formula to calculate the critical length of the fibres for the porous materials and determined the modifying factor (F_l) for the porous bone scaffold investigated in this work. Along with the increase of the fibres' volume content, the compressive strength of the scaffold also increased. We have proposed a further empirical formula for calculating the compressive strength of the porous reinforced materials and determined the modifying factor (F_σ) for the porous reinforced bone scaffold examined in these studies. Along with the degradation in vitro, the decrease in strength of the reinforced scaffold was less than that of the unreinforced scaffold. The growth rate of osteoblast cells on the reinforced scaffold was higher than that on the unreinforced scaffold. These results suggest that the reinforced scaffold may be a promising candidate matrix for repairing large bone defects. Copyright © 2005 Society of Chemical Industry     

Kinetics of in vitro drug release from chitosan/gelatin hybrid membranes

In vitro studies of controlled release from chitosan/gelatin hybrid membranes were carried out using drugs of different molecular weight. It was found that release of urea, 5-fluorouracil (5-Fu), sodium benzoate, sodium salicylate, sodium mandelate, and sulfacetamide sodium followed zero-order kinetics after a short time lag. Variation of the diffusion coefficient, permeation coefficient, and degree of hydration with crosslinking and varying weight percent of gelatin in membrane matrices were studied in detail by using 5-Fu as a model drug. The diffusion coefficient and permeation coefficient of 5-Fu are dependent on the degree of hydration of the swollen membrane. The transport process of drug molecules in the hydrogel membrane is presumed to be predominantly of the pore mechanism. © 1998 John Wiley & Sons, Inc. J Appl Polym Sci 68: 1751–1758, 1998     

载5-氟尿嘧啶壳聚糖/明胶微粒的制备及药物释放性能

壳聚糖及明胶是生物相容性良好的高分子药物载体,制备载5-氟尿嘧啶壳聚糖/明胶微粒,并进行体外释药研究。以石蜡油为外相,壳聚糖/明胶为内相,用乳化交联法制备微粒,以吸附药量为指标,采用正交设计实验优化获得最佳制备条件,用红外光谱、SEM对最佳条件下制备的微粒进行表征。结果表明壳聚糖/明胶微粒的最佳制备条件如下:水油比1:7,壳聚糖/明胶浓度比1:3,乳化剂100.7mmol/L,乳化5min,乳化温度60℃,交联剂戊二醛用量5.5mmol/L,交联时间1h。在此条件下,载药微粒的载药量为34.93%,包封率为38.36%。红外光谱图表明壳聚糖/明胶微粒已负载5-氟尿嘧啶,SEM表明微粒成球状,表面较光滑。模拟胃肠释放表明,微粒具有一定的缓释性能。采用乳化交联法制备载5-氟尿嘧啶壳聚糖/明胶微粒方法简单,重现性好,且其体外释放实验显示出明显的缓释作用。     

鱼鳞明胶/壳聚糖/薰衣草精油抗菌复合膜制备及性能

以鱼鳞明胶(SG)和壳聚糖(CS)为原料,薰衣草精油(LEO)为抗菌剂,通过共混法制备抗菌复合膜,通过FTIR、XRD、SEM和热重分析对复合膜的形貌和结构进行表征及对力学、光学、阻隔、抑菌等性能测试分析。结果表明,LEO与SG/CS基膜较好复合,与SG/CS基膜相比,当LEO添加量为1%时,复合膜具有良好的力学强度(26.36 MPa),断裂伸长率提升了12.96%；透明度有所下降,透光率为70.50%；耐水性能和水蒸气阻隔性能显著增强,含水率、水溶性和水蒸汽透过率分别下降了22.48%、30.49%和26.03%；DPPH自由基清除率提高了247.88%,抗氧化性显著提高；同时对大肠杆菌和金黄色葡萄球菌具有良好的抑菌作用。     

Preparation and characterization of a novel thermosensitive hydrogel based on chitosan and gelatin blends

In this study, a novel injectable in situ gelling thermosensitive hydrogel system based on chitosan and gelatin blends was designed and investigated. The addition of gelatin provides the correct buffering and other physicochemical conditions including control of hydrophobic interactions and hydrogen bonding, which are necessary to retain chitosan in solution at neutral pH near 4°C and furthermore to allow gel formation upon heating to body temperature. The chitosan/gelatin hydrogels were studied by FTIR, swelling, and rheological analysis. The rheological analysis evidenced the endothermic gelation of chitosan/gelatin solutions, which indicated their gelation temperatures and reflected the effect of gelatin concentration on the thermosensitive properties of gels. The morphology of this system was examined with laser scanning confocal microscopy and scanning electron microscopy. The images indicated that the gels were quite heterogeneous and porous. The investigation of these gels as vehicles for delivering bovine serum albumin as a model drug of protein showed that the system could sustain the release of the protein drug. These results show that chitosan/gelatin solutions can form gels rapidly at body temperature and have promising perspective for their use in local and sustained delivery of protein drug. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009