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Formation of calcium phosphates on TiO2 nanotubes in the presence of albumine: insight in formation of multifunctional nanocomposites

Among largest health issues in modern society increased frequency of hard tissue chronical diseases takes special place [1]. These diseases could be a consequence of physical trauma, birth defects, other diseases (cancer), etc. In response to this problem, World health organization has appointed years 2000-2010 a Bone and Joint Decade [2]. Often the only treatment of such diseases is implantation with the aim to regenerate damaged or diseased tissue. However, only in the USA, approximately 10% of implants fail prematurely [3]. In addition, due to the continuous population ageing, many patients are outliving their implants [4]. These problems clearly urge the need to develop new, innovative implant materials. The solution of such problems is sought in development of multifunctional materials, which in addition to replacing missing tissue or enabling its regeneration, as well as having improved mechanical properties, will act as local drug delivery system [4-6]. New hard tissue regeneration biomaterials emerging in recent years are composite materials based on calcium phosphates (CaPs) and different inorganic nanomaterials as they offer possible ways of preparing multifunctional materials because inorganic nanomaterials embedded within CaP can increase their mechanical strength [7] and can be used as drug delivery systems, like TiO2 nanotubes (TiNTs). Nowadays it is possible to produce arrays of TiNTs with precisely controlled diameter, length, wall thickness and phase [8]. In vitro cells response to nanotubular surface is better compared to not treated surfaces and can be modulated by changing the diameter of TiNTs [9]. TiNTs also offer a possibility to covalently immobilize bioactive molecules, and in that way act as drug delivery system [10]. Modifying TiNTs arrays with CaP can be achieved by different techniques, biomimetic, electrochemical and magnetron sputtering being mostly used [11]. However, to the best of our knowledge TiNTs are not used as fiber reinforcements of CaP ceramics and cements. In this research we have investigated the impact of TiO2 nanotubes (TiNT) on properties of formed CaP solid phase at conditions closed to physiological in the presence of bovine serum albumine (BSA). BSA is the most abudant protein in human plasma and is one of first macromolecules that will interact with synthesised nanocomposites in human body. It is one of three soluble proteins which are immediately adsorbed on the surface of the implant, influencing its in vivo performance [12]. However, its role in mineralization of titanium implants is still controversial [13]. All experiment systems were preformed in a thermostated double-walled glass vessel with a 50 mL capacity at 25 °C. Precipitation systems were not additionaly stirred. Control precipitation system (CS) was prepared by fast mixing of equal volumes (20 mL) of equimolar CaCl2 and Na2HPO4 reactant solutions (c = 4 mmol dm—3), so-called anionic and cationic solution. The pH of anionic solution was adjusted to 7.4 with HCl. Precipitation systems containing TiNT and/or BSA were prepared by adding TiNT suspension or BSA solution to the anionic reactant solution. Before mixing the reactant solutions the pH was readjusted when necessary. The progress of reaction was followed by measuring the pH of the precipitation systems. Based on pH curves induction time (tind), the time passed from initiation of precipitation process untill the begining of amorphous CaP (ACP) transforms into more stable crystalline phase was calculated. Prepared precipitates were filtered after 60 minutes of reaction time. The influence of TiNT and BSA on structure and morphology of precipitated CaP phase was investigated by Fourier – Transform Infrared spectroscopy (FTIR), Raman spectroscopy, Poxwder Diffaction (XRD), Scanning Electron microscopy (SEM), Transmission Electron Microscopy (TEM). Representative pH vs time curves are shown in Fig. 1. Curves obtained in all investigated systems showed typical sigmoidal shape reflecting three stages of the precipitation process: i) initial slight pH decrease associated with the formation of ACP, during which the changes in pH and calcium concentrations are small or absent, ii) an abrupt decrease in pH associated with the secondary precipitation of crystalline phase upon ACP, iii) final slight pH change associated with solution-mediated growth and phase transformation [14, 15]. Induction times were obtained from intersection of the tangents drawn on the second and third section of pH vs. time curve (Table 1.). In the presence of TiNTs a significant reduction of induction time was observed, indicating promotion of ACP transformation, which was not concentration dependent in the investigate concentration region. Contrary to TiNTs, in the presence of BSA transformation is inhibited and the influence is concentration dependent. In the precipitatuion systems containing both TiNTs and BSA the transformation was faster at higher TiNTs concentrations. PXRD patterns and FTIR spectra showed that presence of either TiNTs or BSA, or both of them does not influence the composition of the formed precipitate. In all systems calcium deficient hydroxyapatite (CaDHA, Ca10−x(HPO4)x(PO4)6−x(OH)2−x, 0 < x < 2) was formed. CaDHA is considered to be a promising material for manufacturing biomaterials for hard tissue regeneration as biological apatite, the main inorganic part of animal hard tissues, is in fact ion-substituted CaDHA [16]. Althought TiNTs do not influence the morphology, SEM investigation have shown that CaDHA grows on TiNT surface in linear layout. The obtained results point to a biomimetic preparation route of multifunctional CaP based biomaterials.

Calcium Phosphates ; TiO2 nanoparticles ; Bovine Serum Albumine

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