Preparation and characterization of Chitosan and PLGA‐based scaffolds for tissue engineering applications

Three dimensional (3D) biodegradable porous scaffolds play a crucial role in bone tissue repair. In this study, four types of 3D polymer/hydroxyapatite (HAp) composite scaffolds were prepared by freeze drying technique in order to mimic the organic/inorganic nature of the bone. Chitosan (CH) and poly(lactic acid‐co‐glycolic acid) (PLGA) were used as the polymeric part and HAp as the inorganic component. Properties of the resultant scaffolds, such as morphology, porosity, degradation, water uptake, mechanical and thermal stabilities were examined. 3D scaffolds having interconnected macroporous structure and 77–89% porosity were produced. The pore diameters were in the range of 6 and 200 µm. PLGA and HAp containing scaffolds had the highest compressive modulus. PLGA maintained the strength by decreasing water uptake but increased the degradation rate. Scaffolds seeded with SaOs‐2 osteoblast cells showed that all scaffolds were capable of encouraging cell adhesion and proliferation. The presence of HAp particles caused an increase in cell number on CH‐HAp scaffolds compared to CH scaffolds, while cell number decreased when PLGA was incorporated in the structure. CH‐PLGA scaffolds showed highest cell number on days 7 and 14 compared to others. Based on the properties such as interconnected porosity, high mechanical strength, and in vitro cell proliferation, blend scaffolds have the potential to be applied in hard tissue treatments. POLYM. COMPOS., 36:1917–1930, 2015. © 2014 Society of Plastics Engineers     

Stabilizing effect of carbodiimide and dehydrothermal treatment crosslinking on the properties of collagen/hydroxyapatite scaffolds

This paper reports the effect of the combined technique of dehydrothermal treatment (DHT) and a mixture of 1‐ethyl‐3(3‐dimethylaminopropyl) carbodiimide (EDC) and N‐hydroxysuccinimide (NHS) crosslinking on the physicochemical properties of collagen/hydroxyapatite materials. Collagen and collagen/hydroxyapatite porous scaffolds containing different amounts of collagen and hydroxyapatite were prepared with use of the freeze‐drying technique. All samples were capable of absorbing a large quantity of phosphate buffered saline. Samples crosslinked by DHT+EDC/NHS presented higher resistance to collagenase degradation (with slightly reduced degradation in DHT+EDC/NHS crosslinked scaffolds prepared from 2% collagen solution), whereas DHT scaffolds exhibited faster degradation. Mechanical testing results suggested that scaffolds crosslinked by DHT+EDC/NHS treatment have an improved compressive modulus compared with EDC/NHS crosslinking. The qualitative analysis of colour intensity resulting from the CellTiter 96 Aqueous One Solution Cell Proliferation Assay (MTS) led to the conclusion that all samples, regardless of the crosslinking method, were well tolerated by cells. However, DHT and EDC/NHS crosslinked scaffolds seem to support better cell viability, in contrast to DHT+EDC/NHS crosslinked scaffolds that support cell differentiation instead. DHT+EDC/NHS crosslinked scaffolds markedly increase the specific alkaline phosphatase activity of cells, which may be of benefit in bone tissue engineering. © 2017 Society of Chemical Industry     

Preparation,characterization and mechanical properties of controlled porous gelatin/hydroxyapatite nanocomposite through layer solvent casting combined with freeze-drying and lamination techniques

In this present study, to mimic the mineral and organic component of natural bone, hydroxyapatite (HA) and gelatin (GEL) nanocomposite was prepared via layer solvent casting combined with freeze-drying and lamination techniques. Glutaraldehyde (GA) was used as cross-linking agent. The synthesized nanocrystalline hydroxyapatite and nanocomposite samples were characterized by the commonly used bulk techniques. The results showed that GEL/HA nanocomposite were porous with 3-dimension interconnected microstructure, pore sizes were 100 μm to 1 mm, porosity were 75% to 93% and HA particles are dispersed evenly among gelatin fibers. It was also found that increasing initial GEL concentration and HA content enhance the elastic modulus (E) and reduce toughness and affect pore size and morphology. Finally, the stress–strain behavior in compression was very similar to natural spongy bone where the compressive modulus obtained was about 180 MPa.     

Preparation of porous PLGA/HA/collagen scaffolds with supercritical CO2 and application in osteoblast cell culture

Biocompatible three-dimensional scaffolds for cell culturing may facilitate methods for the repair of damaged human tissues. A novel hybrid porous scaffold of poly(lactic-co-glycolic acid), hydroxyapatite and collagen was prepared using a supercritical CO₂ saturation technique. Expansion factors of scaffolds with different compositions were studied after supercritical CO₂ treatment to choose the optimal composition for three-dimensional culture. The supercritical CO₂ process conditions, such as saturation temperature, saturation time and saturation pressure were varied to evaluate their influence on pore structure. The results showed that the pore size and porosity of the scaffold could be controlled by manipulating these process conditions. The porous samples were characterized by environmental scanning electron microscopy, energy-dispersive X-ray spectroscope, Fourier transform infrared spectroscopy and X-ray diffractometry. Finally, MG-63 cells were successfully cultured on the porous scaffold as assessed by electron and confocal microscopy, confirming the biocompatibility of this new hybrid porous scaffold.     

Bone Regeneration in rat using a gelatin/bioactive glass nanocomposite scaffold along with endothelial cells (HUVECs)

In our previous study, a three‐dimensional gelatin/bioactive glass nanocomposite scaffold with a total porosity of about 85% and pore sizes ranging from 200 to 500 μm was prepared through layer solvent casting combined with lamination technique. The aim of this study was to evaluate in vitro biocompatibility and in vivo bone regeneration potential of these scaffolds with and without endothelial cells when implanted into a critical‐sized rat calvarial defect. MTT assay, SEM observation, and DAPI staining were used to evaluate cell viability and adhesion in macroporous scaffolds and results demonstrated that the scaffolds were biocompatible enough to support cell attachment and proliferation. To investigate the in vivo osteogenesis of the scaffold, blank scaffolds and endothelial/scaffold constructs were implanted in critical‐sized defects, whereas in control group defects were left untreated. Bone regeneration and vascularization were evaluated at 1, 4, and 12 weeks postsurgery by histological, immunohistochemical, and histomorphometric analysis. It was shown that both groups facilitated bone growth into the defect area but improved bone regeneration was seen with the incorporation of endothelial cells. The data showed that the porous Gel/BaG nanocomposite scaffolds could well support new bone formation, indicating that the proposed strategy is a promising alternative for tissue‐engineered bone defects.     

Bi-/multi-modal pore formation of PLGA/hydroxyapatite composite scaffolds by heterogeneous nucleation in supercritical CO2 foaming

Scaffolds with multimodal pore structure are essential to cells differentiation and proliferation in bone tissue engineering.Bi-/multi-modal porous PLGA/hydroxyapatite composite scaffolds were prepared by supercritical CO_2 foaming in which hydroxyapatite acted as heterogeneous nucleation agent.Bimodal porous scaffolds were prepared under certain conditions,i.e.hydroxyapatite addition of 5%,depressurization rate of 0.3 MPa·min~(-1),soaking temperature of 55℃,and pressure of 9 MPa.And scaffolds presented specific structure of small pores(122 μm±66 μm)in the cellular walls of large pores(552μm±127μm).Furthermore,multimodal porous PLGA scaffolds with micro-pores(37 μm±11 μm)were obtained at low soaking pressure of 7.5 MPa.The interconnected porosity of scaffolds ranged from(52.53±2.69)% to(83.08±2.42)%by adjusting depressurization rate,while compression modulus satisfied the requirement of bone tissue engineering.Solvent-free CO_2 foaming method is promising to fabricate bi-/multi-modal porous scaffolds in one step,and bioactive particles for osteogenesis could serve as nucleation agents.     

Preparation and characterization of gelatin/hydroxyapatite nanocomposite for bone tissue engineering

Homogeneous gelatin/hydroxyapatite (GEL/HA) nano‐composites were synthesized by a novel in situ precipitation method, and its corresponding characterizations, including composition, morphology, pore structure, thermal stability, mechanical strength, and biocompatibility, were carried out. High‐magnified scanning electron microscope (SEM) images indicated that nano‐HA with particle size ranging from 20 to 50 nm were uniformly distributed in GEL matrix and tightly integrated with organic phase. Wide angle X‐ray diffraction (XRD) analysis and transmission electron microscope (TEM) images showed that, during the process of mineralization, there existed preferred oriented growth of HA crystals along (002) and (211) crystal planes. Thermogravimetric analysis (TGA) and differential thermal analysis (DTA) indicated that, the thermal stability of GEL molecules enhanced by hybridizing with HA nanocrystals. Interconnected porous GEL/HA nanocomposites with pore size ranging between 50 and 350 μm were prepared by a freeze‐drying method. This pore size was adequate for bone tissue engineering (BTE) applications. In addition, in vitro MG63 osteoblast‐like cell culture illuminated that GEL/HA nanocomposites had excellent cytocompatibility and could promote proliferation of cells. These results suggested that GEL/HA nanocomposite might be an ideal bone substitute. POLYM. COMPOS., 38:1579–1590, 2017. © 2015 Society of Plastics Engineers     

In situ synthesized chitosan–gelatin/ZnO nanocomposite scaffold with drug delivery properties: Higher antibacterial and lower cytotoxicity effects

In this work, chitosan–gelatin/zinc oxide nanocomposite hydrogel scaffolds (CS–GEL/nZnO) were prepared via in situ synthesis of ZnO nanoparticles (nZnO) to reach a scaffold with both inherent antibacterial and drug delivery properties. The prepared nanocomposite hydrogel scaffolds were characterized using scanning electron microscopy, transmission electron microscopy, atomic absorption spectrometer, Fourier transform infrared spectroscopy, and X-ray diffraction. In addition, swelling, biodegradation, antibacterial, cytocompatibility, and cell attachment of the scaffolds were evaluated. The results showed that the prepared scaffolds had high porosity with a pore size of 50–400 μm and nZnO were well distributed without any agglomeration on the CS–GEL matrix. In addition, the nanocomposite scaffolds showed enhanced swelling, biodegradation, and antibacterial properties. Moreover, the drug delivery studies using naproxen showed that nZnO could control naproxen release. Cytocompatibility of the samples was proved using normal human dermal fibroblast cells (HFF2). In comparison to the previous reports which nZnO were simply added to the matrix of the scaffold, in situ synthesis of nZnO was led to higher antibacterial and lower cytotoxicity effects as a result of well distribution of nZnO in this method. According to the findings, the in situ synthesized CS–GEL/nZnO is strongly recommended for biomedical applications especially skin tissue engineering. © 2019 Wiley Periodicals, Inc. J. Appl. Polym. Sci. 2019 , 136, 47590.     

Bacterial cellulose/collagen composite: Characterization and first evaluation of cytocompatibility

The novel bacterial cellulose (BC)/collagen composites were prepared by immersing wet BC pellicle excreted by Acetobacter xylinum in collagen solution followed by freeze‐drying process. The product looks like a foam structure. The morphology of BC/collagen composite was examined by scanning electron microscope (SEM) and compared with pristine BC. SEM images showed that collagen molecules was not only coated on the BC fibrils surface but also could penetrate inside BC and hydrogen bond interactions were formed between BC and collagen. The prepared BC/collagen composite was also characterized by Fourier transform infrared (FTIR) spectroscopy, X‐ray diffraction (XRD), thermogravimetric analysis (TGA), and mechanical test. With the incorporation of collagen in the BC, no changes happened in the crystal structure but the thermal stability was improved. Tensile test results indicate that the Young's Modulus and tensile strength have a big increase while the elongation at break has a slight decrease. The cytocompatibility of composite was preliminarily evaluated by cell adhesion studies. The tests were carried out using 3T3 fibroblast cells. The cells incubated with BC/collagen scaffolds for 48 h were capable of forming cell adhesion and proliferation. It showed much better cytocompatibility than pure BC. So, the prepared BC/collagen scaffolds are bioactive and may be suitable for cell adhesion/attachment suggesting that these scaffolds can be used for wound dressing or tissue‐engineering scaffolds. Therefore, these results suggest that these novel BC/collagen scaffolds may have the potential to be sued for some biomedical applications. © 2011 Wiley Periodicals, Inc. J Appl Polym Sci, 2011     

Injection‐molded porous hydroxyapatite/polyamide‐66 scaffold for bone repair and investigations on the experimental conditions

Hydroxyapatite/polyamide‐66 (HA/PA66) composite scaffolds were prepared using injection‐molding technique and also analyzed by means of scanning electron microscopy, X‐ray diffraction, differential scanning calorimetry, Fourier transform infrared spectroscopy, and mechanical testing. Compared with common methods to fabricate scaffolds, this process can fabricate composite scaffolds in a rapid and convenient manner by adjusting the experimental conditions of foaming agent and shot size. The interactions between PA66 and HA particles affect the crystallization behavior of PA66 and the pore structure of scaffolds. HA particles can increase the stiffness of composite scaffolds accompanied by the reduction of impact strength, pore size and porosity. The obtained 40 wt% HA/PA66 composite scaffolds with a pore size ranging from 100–500 μm and a porosity more than 65% can simultaneously meet the requirements of porous structure and mechanical performance. POLYM. ENG. SCI., 54:1003–1012, 2014. © 2013 Society of Plastics Engineers     

Fabrication of β‐tricalcium phosphate composite ceramic scaffolds based on spheres prepared by extrusion‐spheronization

In this study, β‐tricalcium phosphate/phosphate‐based glass (β‐TCP/PG) composite spheres were prepared by an extrusion‐spheronization method featuring high production and fine control of sphere size. Subsequently, fully interconnected β‐TCP composite ceramic sphere‐based (TCCS) scaffolds were fabricated by sintering the randomly packed β‐TCP/PG composite spheres. The results manifested that at least 20% microcrystalline cellulose (MCC) was required to obtain β‐TCP/PG composite spheres in good spherical shape. The prepared TCCS scaffolds showed hierarchical pore architecture, which consisted of interconnected macropores among the spheres, a hollow core in the sphere, plentiful medium‐sized pores in the sphere shell and micropores among the grains. The pore architecture and mechanical strength of the TCCS scaffolds could be tailored by adjusting the sintering temperature, sphere size, and amounts of PG and MCC in the β‐TCP/PG composite spheres. This work is believed to open up new paths for the design and fabrication of interconnected bioceramic scaffolds for application in bone regeneration.     

Preparation of chitosan‐nanohydroxyapatite composite scaffolds by a supercritical CO2 assisted process

In this study, chitosan‐nanohydroxyapatite composite scaffolds were prepared by a supercritical fluid assisted process. For this purpose, different amounts of nanohydroxyapatite particles, that is, 0.25, 0.50, and 1.00 wt% were added to chitosan (deacetylation degree: DD 75–85%) solution (2%, w/v, in acetic acid). The gels were then frozen at −20°C, treated in acetone and dried in a supercritical fluid extractor under a constant CO₂ flow of 15 g/min at 35°C and 200 bar for 5 h to obtain porous scaffolds. Scanning electron microscope views showed that the drying of gels under supercritical CO₂ lead to the formation of microporous scaffolds with a pore size distribution of 30–150 μm. Addition of nanohydroxyapatite particles did not significantly affect the pore size distribution. Attenuated Total Reflectance Fourier Transform Infrared Spectroscopy and X‐Ray diffraction analyses supported the successful incorporation of nanohydroxyapatite particles in the scaffold. An increase in water uptake and mechanical strength were observed in composite scaffolds. The results obtained from this study indicated that chitosan‐nanohydroxyapatite scaffolds prepared by using supercritical CO₂ shall be considered as a potential candidate for bone tissue engineering applications. POLYM. COMPOS., 2012. © 2012 Society of Plastics Engineers     

Scaffolds for Hard Tissue Engineering by Ionotropic Gelation of Alginate–Influence of Selected Preparation Parameters

Cylindrical constructs with parallel aligned pores were prepared by using ionotropic gelation of alginate/calcium phosphate hydroxyapatite (HAP) mixtures in regard to applications as scaffold for bone regeneration. The starting powder and stabilizing agents were characterized by measurement of electrosonic amplitude, particle size distribution, and specific surface. The shrinkage of the gels was investigated in dependence on the drying methods. The pore size relied on preparation conditions such as amount of HAP and concentrations of gelling agent or alginate sol. A wide field of pore sizes could be fabricated by varying the kind and concentration of additives. Micro computer tomography-investigations of freeze dried scaffolds demonstrated the pore progression over a length of 4 mm. The pore dimension and structure were adequate for cell seeding and blood capillary ingrowth. Biocompatibility was proven by in vitro experiments with human mesenchymal stem cells by fluorescence microscopy. A high stability of the wet gels was maintained under cell culture conditions for a period of 3 weeks.     

Effects of formaldehyde solution and nanoparticles on mechanical properties and biodegradation of gelatin/nano β-TCP scaffolds

In this study, gelatin/beta tricalcium phosphate (β-TCP) nanocomposite scaffolds were prepared by solvent casting method. The cross-linking method was carried out by adding formaldehyde to gelatin. The microparticles of sodium chloride were used as porogen agent. Characterization of nano β-TCP was performed using XRD, FTIR, and SEM. Results showed that the size of the particles is about 100 nm with spherical morphology. In addition, the scaffold characterization was carried out using FTIR and SEM techniques. Observations showed a porous texture with pore size between 100 and 400 μm. The biodegradability and bioactivity evaluations of the scaffolds were done by immersing them in a simulated body fluid solution for different time periods. The biodegradability studies demonstrated a reduction in the degradation rate of gelatin/β-TCP nanocomposite scaffolds due to the presence of β-TCP nanoparticles. The obtained results of bioactivity tests confirmed the formation of apatite layer on the surface of the scaffolds. Furthermore, the effects of porosity, cross-linking agent, and β-TCP nanoparticles on the bending and compressive properties of the composite scaffolds were examined. According to the mechanical examinations of the scaffolds, the best bending and compressive properties occurred in the presence of 10 and 20 wt% of β-TCP nanoparticles, respectively. The appropriate mechanical properties and biodegradation rate for tissue engineering applications obtained at 1 g of the formaldehyde solution.     

Development of Collagen/Demineralized Bone Powder Scaffolds and Periosteum-Derived Cells for Bone Tissue Engineering Application

The aim of this study was to investigate physical and biological properties of collagen (COL) and demineralized bone powder (DBP) scaffolds for bone tissue engineering. DBP was prepared and divided into three groups, based on various particle sizes: 75–125 μm, 125–250 μm, and 250–500 μm. DBP was homogeneously mixed with type I collagen and three-dimensional scaffolds were constructed, applying chemical crosslinking and lyophilization. Upon culture with human periosteum-derived cells (PD cells), osteogenic differentiation of PD cells was investigated using alkaline phosphatase (ALP) activity and calcium assay kits. The physical properties of the COL/DBP scaffolds were obviously different from COL scaffolds, irrespective of the size of DBP. In addition, PD cells cultured with COL scaffolds showed significantly higher cell adhesion and proliferation than those with COL/DBP scaffolds. In contrast, COL/DBP scaffolds exhibited greater osteoinductive potential than COL scaffolds. The PD cells with COL/DBP scaffolds possessed higher ALP activity than those with COL scaffolds. PD cells cultured with COL/DBP scaffolds with 250–500 μm particle size yielded the maximum calcium deposition. In conclusion, PD cells cultured on the scaffolds could exhibit osteoinductive potential. The composite scaffold of COL/DBP with 250–500 μm particle size could be considered a potential bone tissue engineering implant.     

Polymer template fabrication of porous hydroxyapatite scaffolds with interconnected spherical pores

Porous hydroxyapatite (HA) scaffolds with interconnected spherical pores were fabricated by slip casting using a polymer template. Templates were produced using polymer beads, NaCl, and adhesive (N100). Effects of the preparation process on the pore structures and mechanical properties of the porous HA scaffolds were investigated. Pore interconnectivity was improved by adding NaCl particles with appropriate diameters to the polymer template. The size of the adhesive area could be controlled by adjusting the concentration of N100. The pore size could be controlled between 200 ± 42 and 400 ± 81 μm, and the porosity between 50.2 and 73.1%, by changing the size of the polymer beads and the volume of the NaCl particles. The compressive strength decreased as the porosity or pore size increased.     

Bone‐like apatite‐coated chitosan scaffolds: Characterization and osteoblastic activity

In this study, porous scaffolds were prepared from chitosan (2% w/v in acetic acid and deacetylation degree: DD > 85%) by freeze‐drying method, and freshly lyophilized scaffolds were stabilized with ethanol solutions. Bone‐like apatite formation on chitosan scaffolds was achieved by immersing the scaffolds into a novel concentrated simulated body fluid (10× SBF‐like solution) for different periods, i.e., 6 and 24 h. Scanning electron microscope views showed that the 6‐h treatment in 10× SBF‐like solution led to the formation of calcium phosphate nucleation sites on chitosan scaffolds, whereas the apatite particles showed characteristic cauliflower‐like morphology at the end of 24‐h treatment. X‐ray diffraction results supported the fact that mineral phase was made of hydroxyapatite. Osteogenic activities of untreated and SBF‐treated chitosan scaffolds were examined by preosteoblastic MC3T3 cell culture studies. The mitochondrial activity test showed that apatite‐coated scaffolds stimulated cell proliferation compared with uncoated scaffolds. Alkaline phosphatase and osteocalcine levels indicated that the differentiation of the cells on all scaffolds increased significantly from 15th day of culture to the 21th day of culture, especially for the cells on 24‐h SBF‐treated scaffolds. The results of this study indicated that 10× SBF‐like solution‐treated chitosan scaffolds may be evaluated for bone tissue engineering. POLYM. COMPOS., 31:1418–1426, 2010. © 2009 Society of Plastics Engineers     

Preparation and characterization of magnetite/hydroxyapatite/chitosan nanocomposite by in situ compositing method

Chitosan is an important kind of biomaterial that is widely used in medical applications. One of the key concerns about its use is the preparation of composites used for bone engineering. Aim of this study concerns the preparation of three‐dimensional nanocomposites having potential use in bone repair and regeneration. The magnetite/hydroxyapatite/chitosan nanocomposites were prepared via in situ compositing method by preparing precursor solutions and molds with chitosan membrane. These nanocomposites were characterized by chemical, spectroscopic, magnetic, and morphological methods. X‐ray diffraction analysis results demonstrate the formation of magnetite and hydroxyapatite in the chitosan matrix. FTIR analysis indicates that inorganic nanoparticles were chemically bound to the amino and hydroxyl groups in CS molecules. From TG/DTA data, it can be concluded that during preparation raw materials were almost perfectly incorporated into the nanocomposites, and the decrease in decomposition temperatures indicates the formation of chemical bonds between inorganic nanoparticles and chitosan molecules. TEM results show that the maximum size of inorganic particles in the magnetite/hydroxyapatite/chitosan nanocomposites was under 50 nm, and these particles were dispersed homogeneously in the chitosan matrix. From the magnetic measurement, it could be concluded that the nanocomposites were superparamagnetic, which is also the peculiarity of nanomagnetites. © 2008 Wiley Periodicals, Inc. J Appl Polym Sci, 2008     

可注射nHA/PU复合多孔骨修复支架制备及表征

兼具良好孔隙率和原位任意塑形固化的可注射复合多孔骨修复材料在临床不规则骨缺损的治疗方面显示出巨大的优势。本研究通过优化双组分设计,以水为发泡剂制备可注射纳米羟基磷灰石/聚氨酯（nHA/PU）复合多孔支架。利用扫描电子显微镜（SEM）、傅里叶变换红外光谱（FTIR）、X射线衍射（XRD）、力学测试及Gillmore针测试等手段对制备的支架进行结构形貌、化学组成、力学性能和固化时间表征。结果表明,本研究制备的可注射nHA/PU复合多孔支架孔隙率高、孔隙贯通性好,孔径分布在100~700μm,适宜细胞和组织向孔内生长;添加20% nHA显著提高了PU支架的力学强度,但降低了支架的孔隙率;可注射支架在8h固化,适宜临床操作。本研究制备的可注射nHA/PU复合多孔支架在不规则骨缺损修复领域具有较大的应用潜力。     

Glutaraldehyde crosslinked gelatin/hydroxyapatite nanocomposite scaffold,engineered via compound techniques

In this study, a scaffold was designed to be used in bone tissue repair and the effect of glutaraldehyde (GA) concentration as crosslinking agent was investigated. To mimic the mineral and organic component of natural bone, hydroxyapatite (HAp) and gelatin (GEL) were used as the main components of this composite. Nanopowders of HAp were synthesized and also used together with GEL to engineer a three‐dimensional nanocomposite scaffold. The results show that GEL/HAp nanocomposite is porous with three‐dimensional interconnected structure, pore sizes ranging from 300 to 500 μm, and about 85% porosity. In addition, increasing GA concentration provokes the enhancement of compressive strength until 1 w/v% GA solution followed by a reduction to 2.5%, whereas it causes work fracture to decrease. It was concluded that optimum concentration for crosslinking GEL matrix for this purpose is 1 w/v% GA solution. A specific combination of commonly used techniques applied to engineer a scaffold with almost ideal properties intended for the bone tissue engineering is introduced. In addition, scaffolds that are prepared via this compound process has the potential to be used in the solid free form applications and so being formed in any dimension and geometry relevant to the defect size and shape. POLYM. COMPOS., 31:2112–2120, 2010. © 2010 Society of Plastics Engineers