Synthesis and Characterization of Hybrid Fiber Scaffold for Medical Implants

This work synthesized and characterized chitosan-silica based hybrid fiber from cowry shell and rice husk respectively with the aim of studying the behavior of a hybrid polymer-silicate composite via the electrospinning technique to produce nanofibers from natural polymer and agro waste, from which scaffolds can be produced which can be used for culturing and wound healing application in the biomedical field. The samples obtained were subjected to the following analyses: Energy Dispersive X-ray Florescent (ED-XRF), Fourier Transform Infrared Spectroscopy (FTIR), X-Ray Diffraction (XRD), Scanning Electron Microscopy (SEM) and Thermogravimetic Analysis (TA). It was observed from the ED-XRF analysis that the major constituent of rice husk ash is silica (SiO2) with some associate metallic oxides. The peaks obtained from the FTIR analysis of the polymer confirm the characteristics features for the polysaccharide structure of chitosan while the characteristic and positions of the peaks observed in the extracted silica spectra showed similar position with that of the commercially available silica obtained from BDH chemicals UK. The Chitosan-silica hybrid has Si AOH and Si AO ASi with the respective bands within the regions of 3270– 3400cm-1 and 989–1300cm-1.The X-ray diffraction analysis indicates the presence of crystallite in polymers which usually results to improve mechanical properties, unique thermal behavior, and increased fatigue strength. These attributes make crystalline polymers desirable materials for biomedical applications. Also, amorphous components of the hybrid composite would present an improved biodegradation behavior of a biomedical material. It was observed from the morphological analysis that there was good interfacial interaction between the chitosan and the dispersion of the silica reinforcement material. The TA of the sample was carried out to investigate the effect of silica reinforcement on the thermal stability of the hybrid composite. It was discovered that the reinforcement of the silica matrix and its reaction with the polymer makes the thermal resistance of the hybrids to increase and hence, increase in thermal decomposition temperature.


Introduction
The utilization of biomaterials in humans has experienced a great success in surgery, dentistry, orthopedics and the likes. For instance, heart valves prostheses are fabricated from carbons, metals, elastomers, fabrics and natural valves and other tissues chemically pretreated to reduce their immunologic response and to enhance durability [1]. Also, the human hip joints are fabricated from titanium, specific high strength alloys, ceramics, composites and ultrahigh molecular weight polyethylene. Dental implants fabricated from titanium materials to form an artificial tooth on which a crown is affixed. One of the advantages of the titanium implants is the bonding with the bone of the jaw. However, this attachment has been more accurately described as a tight apposition of mechanical fit and not true bonding [2]. Wear, corrosion and mechanical properties of titanium have also been of concern [3]. Several other materials have been used to fabricate these implants. Some of these in- applications such as orthopedics, dental implants, drug delivery and others [4,5]. The application of chitin in the biomedical field started from the investigation of the reaction and chemical attributes of lysozyme; an enzyme in the human body fluids. It dissolves certain bacteria by cleaving to the chitinous material of the cell walls. It has been suggested that chitosan may be used to inhibit fibroplasias in wound healing and to stimulate tissue growth and differentiation in tissue culture [6].
Silica based biomaterials, such as melt-derived bioactive glasses and sol-gel glasses, have been used for a long time in bone healing applications because of their ability to form hydroxyl apatite and to stimulate stem cell proliferation and differentiation [7]. They have also been reported to support the differentiation of osteoclasts. For these reasons silica based biomaterials have been suggested to be used as bone filling materials or as part of tissue engineering scaffolds in bone repair. Chemically, the most simple silicate material is silicon dioxide, or silica (SiO2). Structurally, it is a three-dimensional network that is generated when every corner oxygen atom in each tetrahedron is shared by adjacent tetrahedral. Thus, the material is electrically neutral and all atoms have stable electronic structures [3]. Silica fiber like every other fiber possesses a great strength, hence it is envisaged that if blended with chitosan and a hybrid fiber is made of the composite, it will enhance the strength of the material [8].
It has been discovered that most medical problems cannot be corrected by either the natural healing or traditional surgical process [9]. In such cases, implants made from biomaterials can be successfully used to interface with living host tissue to give remedies [10]. Hence, this work attempt to synthesize, characterize and fabricate a hybrid, nanofiber composed of chitosan and bio-silicate glass for biomedical application by preparing an organic and inorganic hybrid cross linking network and electro spinning process to produce the nanofiber.

Materials and Method
Silica separation from rice husk ash Rice husks were obtained from Owode, Ogun state, Nigeria and incinerated at 650 ˚C for a period of 4 hours to form rice husk ash.
The ash was mixed with the solution of 1 molar concentration of sodium hydroxide (NaOH) solution. The result was sintered at 850˚C for 3 hours to form sodium silicate (NaSiO3)

Synthesis of chitosan
The cowry shell was washed thoroughly to get rid of impurities such as sand, dust, dirt and insect larva before sun drying. The subsequent product was pulverized to produce a powder of particle size of 250 microns which was subjected to the following chemical hydrolysis processes to form chitosan [11,12].
Deproteinization: 80g of the powder was weighed into a conical flask and 100ml of 4% NaOH was added. NaOH was prepared by weighing 4g of NaOH pellet and dissolved and made up in 100ml to obtain 4% NaOH. The mixture was boiled and stirred at 100 ºC for 2hrs in a water bath. It was filtered and washed with deionized water. Red litmus paper was used to check if the base was completely washed away. After the washing, the mixture was filtered and the residue was scraped, gently into the Petri dishes, followed by oven drying at 110 ºC for 3hours. The weight of the sample was 62g after deproteinization.
Demineralization: 1M HCl was prepared by measuring 55ml of Conc. HCl and mixed with 1liter of distilled water in a cylindrical flask of 1000cm 3. 5% of the prepared solution was measured (30ml) and mixed with the deproteinized sample. The mixture was boiled and stirred for 60 minutes at 100 ºC in a water bath. Subsequent washing was done with distilled water followed by filtration.
The mixture was examined with blue litmus to check the acidity of the mixture. The residue (chitin) obtained from above was scarped into the Petri dish and dried in the oven at 100 ºC for 2hrs.

Deacetylation:
The isolated chitin was soaked in 510ml of 50% NaOH (weighing 50g of NaOH pellets and dissolved in 100ml of distilled water), boiled at 100 ºC for 2hrs in water bath and cooled for 30mins at room temperature. The mixture was placed on a magnetic stirrer at 30 ºC for 4hrs, filtered, washed and examined with red litmus to check if the base was completely washed away. The mixture was filtered to retain the solid matter which is chitosan.
The chitosan was oven dried at 90 ºC for 24hrs.

Preparation of Silica-Polymer (chitosan-Polyetheleneoxide) Solutions
Polymer solutions were prepared with distilled water and glacial acetic acid. The concentration of the polymer solutions was prepared in wt/wt%.
The polymer solutions were stirred over night to allow homogeneity of the solutions.
iii. Silica solution was prepared by weighing 1.5g of the syn-

Am J Biomed Sci & Res
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Discussion
The extracted chitosan (polymer) solution on electropinning did not form fibers; but drops and beads were formed with low degree of fiber deposition even at higher viscosity. This is in agreement with reports from other literatures that showed that electrospinning of chitosan in aqueous solvent was unsuccessful unless another electrospinning-inducing polymer [13] such as PEO is introduced into the solution. On addition of PEO, an increase in polymer viscosity was observed which led to the formation of nanofiber deposited on aluminum foil and glass surface.

ED-XRF analysis
The product of the incinerated process of rice husk was white rice husk ash whose chemical composition was investigated by ED-XRF analysis (Table 3). It can be observed from Table 3 that the major constituent of rice husk ash is silica (SiO2) which occurs with some other metallic oxides like alumina, ferric oxide, titanium oxide, calcium oxide, magnesium oxide, zinc oxide, manganese (II) oxide, phosphorus pent oxide and others [14]. After the extraction of silica from rice husk ash, the major chemical and functional groups were identified from the FTIR spectra as  [17]. The adsorption peak at 1442cm -1 is similar to in-plane OH bending. The peak observed at 1075cm -1 is related to Si AO ASi stretch [17] whereas the peak at 780cm -1 is associated to the overlapping of Si AC stretch and NH2wag. The peak at 459cm -1 is assigned to O-Si-bending [18]. The large intensity of the peak at 1080cm -1 might be due to the overlapping of the Si AO ASi and the AC AO AC A of glycosidic linkage [17,19]. Chitosan-silica hybrid has Si AOH and Si AO ASi with the respective bands within the regions of 3270-3400cm -1 and 989-1300cm -1 [16].

XRD results analysis
The diffraction pattern from amorphous materials (including many polymers) is devoid of the sharp peaks characteristic of crystals and consists of broad features or halos. The X-ray diffraction pattern of chitosan twelve peaks were identified at 2θ with the peaks showing an hydrated partially crystalline material with amorphous background identified between 55º to 70º at 2θ. This result suggests that the extracted chitosan is a semi crystalline polymeric material.
The XRD spectra of 2θº diffraction between 30º and 40º showed three specific peaks at 31.27º, 32.98˚ and 37.89˚. These features of extracted silica spectrum suggested the characteristic of amorphous micro porousilica [18]. The XRD result is presented in Figure   6.    groups [22]. Chitosan has low rate of weight loss in the axis ranging between 175 ˚C and 290 ˚C because of the decomposition of low molecular weight species [22]. The rate of thermal decomposition is much around 180 ˚C and 465 ˚C due to the complex dehydration of the saccharide rings, depolymerization, and decomposition of the acetylated and deacetylated units of the polymer [23]. Therefore, the incorporation of silica and its reaction with the polymer makes the thermal resistance of the hybrids to increase and hence, increase in thermal decomposition temperature [24,25].