Induction Plasma Sprayed Biological-like Apatite Coatings for Biomedical Applications

Max Loszach, François Gitzhofer

CREPE, Department of Chemical and Biotechnological Engineering,

Université de Sherbrooke

Sherbrooke, Québec, CANADA

Induction plasma sprayed coatings were developed based on a sol-gel suspension, including different ionic substitutions such as K, Mg, F, Cl and Na in order to enhance the biological compatibility. Calcium phosphates coatings are well-known for their use as bones substitutes considering their excellent biocompatibility and bioactivity [1-2]. Although the crystallographic structure of the synthetic hydroxyapatite (HAP) is similar to natural bone, biological apatites are always non-stoechiometric and presents some incorporated elements on trace level in its lattice [3]. Several studies [4-7] have shown that these elements, even in trace levels, play an important role on the biological process as well during implantation as during the life-time of the bone.

A suspension based on nitrates and ammonium [8] was prepared by a sol-gel way including the different ionic substitutions at different concentration rate. The coating was then realized using induction plasma spray under in Ar/O2 gas mixture. The deposited coatings were characterized using X-ray diffraction (XRD) and scanning electron microscope (SEM). The concentration of the different substitution as well as the Ca/P ratio was analysed using neutron activation.

The reactivity of the coatings was also studied by an immersion for different time (1, 3, 7, 14 and 28 days) in a simulated body fluid (SBF) based on Kokubo method [9]. Samples were characterized after immersion using SEM and XRD and Ion Chromatography/Mass Spectrometry (IC/MS). Analysis were performed on the SBF in order to see the dissolution rate as a function of the different ions added to the HA suspension.

Key words: Inductive plasma, sol-gel suspension, biomaterials, biological-like apatite

1 Introduction Hydroxyapatite [HA: Ca10 (PO4)6(OH)2] [10] is a bio-ceramic material which belongs to the calcium phosphate family which is well-known for its use in biological applications. Having similarity with the crystallographic structure of natural bone, it is often applied clinically as a coating on an inert metallic implant such as Ti-6Al-4V. Despite of the structural and crystallographic similarities with synthetic Hydroxyapatite (HAP), natural apatites always presents a nonstoichiometric nature incorporating several elements on trace level in its lattice [11, 12]. It is generally accepted that HAP crystallizes in the hexagonal space group P63/m with 2 formula units Ca5(PO4)3(OH) per unit cell [13, 14]. Geometrically, it is an hexagonal stack of isolated PO4

3- tetrahedrous creating two kinds of tunnels parallel to the c-axis. The first kind of tunnel is filled with Ca+ ions which forms CaO9- polyhedral while the second is lined by oxygen and other Ca ions is occupied by OH- anions [10, 13]. The presence of such tunnels gives appatites structure ion exchange properties. Although these elements are at trace level, several studies have shown their importance on the different level of biological process upon implantation [15, 16]. Numerous studies have been trying to incorporate elements such as Mg, K, Na, Cl, F… in order to obtain a mono or pluri-substitued HAP using conventional sintering approach. The aim of the present work is to produce an HAP coating with combined substituted element present in biological apatite on a titanium-based substrate using induction plasma spraying of a suspension and to characterize it.

2 Experimental procedures

2-1 Preparation The suspension injected in the induction plasma system was prepared using the Tanahashi’s method [17] and a protocol developed by Kannan et al. [18]. Calcium nitrate tetrahydrate [Ca(NO3)2.4H2O, Sigma Aldrich], diammonium hydrogen phosphate [(NH4)2HPO4, Fisher], sodium nitrate [NaNO3, Sigma Aldrich], magnesium nitrate hexahydrate [Mg(NO3)2.6H2O, Sigma Aldrich], potassium nitrate [KNO3, Sigma Aldrich], ammonium chloride [NH4Cl, Sigma Aldrich], and ammonium fluoride [NH4F, Sigma Aldrich] were used as precursors for the synthesis. A mixture containing nitrates of Ca, Na, K and Mg was stirred at a rate of 2000 rpm, while a mixture of (NH4)2HPO4, NH4F and NH4Cl was slowly added at a rate of 40 mL/min. The concentration of the different precursors can be found on table 1. After this step, the pH value was around 4. It was then increased to a value of 10 by adding an ammonium hydroxide (NH4OH) solution. After the equilibration of the pH value, the mixture was heated at 90°C for 2h under a constant stirring of 2000 rpm. The precipitated suspension was then settled down for 24h in order to maturate the precipitate. After 24h, the suspension was centrifuged for 10 min at 2000 rpm in order to eliminate the last trace of liquid and obtain a concentrate paste. It was then mixed with deionised water in order to obtain the suspension.

Table 1.Concentrations of the different precursors

Precursor Molarity Weight(g) in 1000 mL

Ca(NO3)2.4H2O 1,0 472,2 (NH4)2HPO4 0,6 158,5 KNO3 0,0017 0,35 NaNO3 0,057 9,64 Mg(NO3)2.6H2O 0,025 13,03 NH4Cl 0,005 0,55 NH4F 0,001 0,1 Ca/P ratio 1,67

2-2 Plasma process The system used to generate the plasma is the PL-50 supplied by Tekna Plasma Systems Inc (sherbrooke, Quebec, Canada) connected to a 3 MHz LEPEL HF power generator. The system including the deposition chamber is described in figure 1. The sample holder was passing under the plasma with a speed of 50 cm/s and a spraying distance of 16 cm. In the meantime, a suspension was axially injected by a peristaltic pump with a feed rate of 10 mL/min directly into the plasma chamber through the probe. The process parameters are summarized in table 2.

Figure 1 : RF plasma torch PL-50 by Tekna Plasma

Systems Inc [Boulos M.I, 1992]

Table 2 : process parameters during coating depositions

Plasma Power 30 kW

Central gas (Ar) 23 slpm Sheath gas (Oxygen) 63 slpm

Atomizing gas (Argon) 12 slpm Working pressure 100 mmTorr

2-3 Characterization Phase compositions were determined using X-ray diffraction (XRD) on a Philips X’pert Pro MPD X-ray diffractometer (Eindhoven, Netherlands). Microstructures of the coating were observed on a Hitachi VPSEM S3000N Scanning Electron Microscope (Tokyo, Japan) by secondary electron microscopy (SE) and backscattered electron microscopy (BSE). Elemental analysis for the presence of all elements were performed using neutron activation. Rietveld analysis have been performed using the MAUD software [19].

3 Results and Discussion The XRD pattern of the coating is presented in figure 2. Both hydroxyapatite and β-TCP phases are present on the pattern. The concentrations of the different substituted elements are not significantly high enough to affect the XRD pattern. It is also interesting to notice the low intensity of the CaO peak which corresponds to a low concentration of this phase which is normally a problem in HA plasma deposited coatings since CaO is cytotoxic for the different bones cells [20]. Quantitative analysis on the phase concentration using the Rietveld method shows a ratio close to 80/20 respectively for HA and β-TCP which at the same time is beneficial for the stability (HA) and for the resorbability (β-TCP) [21, 22].

 

Figure 2 XRD pattern for the s-HAP Coating

The elemental analyses of the s-HAP coating are presented in table 3. The characterization has been performed on three different series of coatings, in order to confirm the reproducibility of both of the suspension and deposition protocols. The results confirm the presence of all different substituted elements in the coating structure. The concentration of each element is very close to the biological apatite found in bones structure [2]. Although the existing scientific reports [24, 25] have well studied the distribution of the single or dual elements substitution in the apatite structure, there are no studies concerning multi-substitutions in the synthetic apatite. However, several studies have documented the importance of the influence of each of these elements, although they are only existing in trace levels in biological apatite. Magnesium, already well-known in substituted synthetic apatite, plays an important role in the calcification process, as well on the mineral metabolism [26]; sodium intercedes on bone metabolism and osteoporosis [27]; chlorine plays a significant role in the development of an acidic environment on the surface of the bone which activates osteoclastes in bone resorption process [28]; potassium has an active part in mineralization and biochemical processes [29]; fluorine is already well-known for its ability to stabilize the apatite structure which plays an

important role in dental caries prevention [30].

Table 3 Neutron activation results for the s-HAP coating

Sample 1 Sample 2 Sample 3

Ca (%wt) 36,8 39,6 39,9

K (ppm) 264 418 391

Mg (ppm) 5192 6122 5350

Na (ppm) 6272 8606 8696

F (ppm) 173 237 97

Cl (ppm) 793 1864 1165

P (%) 18,3 19,1 19,5

Element concentration (%wt) Ca P Na Mg K Cl F Bone 24,5 11,5 0,7 0,55 0,03 0,1 0,02 s-HAP

38,8 19 0,86 0,55 0,04 0,15 0,015

The SEM micrographs of the microstructure are presented in figure 3 and 4. The coating presents a high open microporosity around 10 µm, which results from the long deposition distance as well as the from high concentration of the suspension in comparison with the low power used. It presents a cauliflower microstructure with the presence of a nano-structured organisation showing an important surface rugosity. Although we have been able to reproduce the bone composition using the SPS technology, there is still some engineering to be done on the coating microstructure as there should be the presence of micropores in the range of 10 µm to allow for biological fluids circulation for bone cells in-growth and larger pores in the range of 100 -500 µm to allow for cell development. [31, 32]

Figure 3 SEM micrographs of the microstructure of the

coating (x250 and x5,0k)

Figure 4 SEM micrograph of the cross section of the

coating (x700 and x1,0k)

4 Conclusions To conclude, a bi-phasic mixture of β-TCP/HA, enable to enhance the resorbability of the material in order to favour the bones remodelling process has been developed by inductive plasma using a SPS method. The coatings possess a microstructure enables to enhance the fluid circulation trough the material, which present an important open microporosity around 10 µm. The elemental analysis performed by neutron activation shows that the elements substituted inside the s-HAP structure are at concentrations really close from natural bone.

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