Influence of silver content on the tribomechanical behavior on Ag-TiCN bioactive coatings

1 Influence of silver content on the tribomechanical behaviour on AgTiCN bioactive coatings J.C. Sánchez-López1, M.D. Abad1, I. Carvalho2, R. Escobar Galindo3, N. Benito4, S. Ribeiro2, M. Henriques5, A. Cavaleiro6, S.Carvalho2 1Instituto de Ciencia de Materiales de Sevilla (CSIC-US), Avda. Americo Vespucio 49, 41092 Sevilla, Spain 2Universidade do Minho, Dept. Física, Campus de Azurém, 4800-058 Guimarães, Portugal 3 Instituto de Ciencia de Materiales de Madrid (ICMM-CSIC), Cantoblanco, 28049, Madrid, Spain 4 Departamento de Física Aplicada (CXII), Universidad Autónoma de Madrid, Cantoblanco, 28049, Madrid, Spain 5 IBB-Institute for Biotechnology and Bioengineering Centre for Biological Engineering Universidade do Minho Campus de Gualtar, 4700-057, Portugal 6 SEG-CEMUC Mechanical Engineering Department, University of Coimbra, 3030788 Coimbra, Portugal Abstract Surface modification of bulk materials used in biomedical applications has become an important prerequisite for better biocompatibility. In particular, to overcome the particle generation, low-wear coatings based on carbon (nitrogen) and containing antimicrobial elements such as silver are promising candidates. Thus, the present work explores the potentialities of silver-containing carbonitride-based (Ag-TiCN) thin films prepared by direct current unbalanced reactive magnetron sputtering. The silver content in the coatings was varied from 0 to 26.7 at.% by changing the targets and the fraction of C2H2 and N2 in the gas mixture with Ar. The obtained Ag-TiCN based coatings were characterized in terms of composition and microstructure. Mechanical and tribological properties of the films were studied by nanoindentation and reciprocating pin-on disk *Manuscript changes highlighted Click here to view linked References 2 testing in a fetal bovinum serum solution, respectively. Raman, SEM and energy dispersive X-ray (EDX) analysis was carried out in the contact region after tribological tests to obtain information about the friction mechanism. The cytotoxicity of the coatings was assessed by in vitro tests using fibroblast cells. The coatings comprised a mixture of TiCxN1-x, Ag and a-C(N)x phases whose relative proportion varied depending on the Ag/Ti ratio. The mechanical, tribological and cytotoxicity were correlated with the chemical and phase composition. When the Ag/Ti ratios were below 0.20 (Ag contents < 6.3 at.%) the films resulted harder (18 GPa) with higher wear resistance (10-6 mm3/Nm), showing similar friction coefficient (0.3) and good biocompatibility. Keywords: Biocompatibility, silver, nanocomposite, amorphous carbon phases, tribology, fetal bovine serum, wear mechanism, cytotoxicity. 3 1. Introduction Application of thin films in the biomedical engineering field represents an attractive challenge due to the multiple situations where they may improve or even functionalize a certain part of the human body. Although the use of hip implants is continuously increasing, implant failure is a huge problem for both the patient and governmental agencies, once it involves repeated surgeries and consequently considerable economical resources, as well as patients’ death. This failure can be attributed to excessive wear and wear debris and also to microbial infection, which promotes the short durability [1]. For these reasons the investigation of new biomaterials is required to obtain good mechanical, tribological and biological properties that allow the development of better prosthesis. To overcome the problem of particle generation, the use of a very low wear coating material as diamond-like carbon (DLC), transition-metal carbides (MeCx), nitrides (MeNx), or others protective thin films, have been proposed [2-5]. In recent years, the research has been directed towards the use of antibacterial materials that could reduce the failure of medical devices provoked by the infections. Many of these studies focus on the use of silver, which is known to be an antimicrobial element [5-8]. When silver is transformed in ions and enters the environment, the multiplication of bacteria may be stopped because it is believed that Ag+ absorbs the proteins of the cell wall [9]. The antibacterial activity is dependent on the total amount of Ag+ ions which is responsible for the destruction of the bacteria. Conversely, if the quantity of silver released from the films is too high it can produce cytotoxicity [10]. Early works have studied the combination of hard phases (TiN or TiC), that provide hardness, with silver as soft metal to play the role of lubricant, to investigate their tribological performance [11,12]. For the use in artificial implants, good 4 biocompatibility is also required, in particular due to the known toxicity of the silver to the human body. In the last few years, the research efforts have been directed to assess the the biocompatibility of these multiphase nanocomposite materials with optimum tribological properties [5,6, 13-15]. We have investigated previously the deposition of TiC(O)N-based coatings by direct current (DC) magnetron sputtering gaining knowledge about the synthesis conditions that yielded a good compromise between tribological and hardness properties [16,17]. In the present study the addition of variable concentrations of Ag into TiCN films is explored with the goal of maintaining a good tribological performance without cytotoxicity effects. To achieve this purpose, the coatings are evaluated in terms of structure and composition and the tribological properties are studied in lubricated conditions using fetal bovinum serum (FBS) to simulate the biological conditions. The final mechanical and tribological performance is correlated with the evolution of the ratio of Ag/Ti inside the coatings as well as the effect of the Ag content in the cytotoxicity. 2. Experimental Details Ag-TiCN samples were deposited by reactive DC magnetron sputtering using an Alcatel SCM650 apparatus onto polished and ultrasonically cleaned 316L steels and single crystalline silicon (100) substrates. Two types of targets (pure Ti and mixed Ti/Ag) of dimensions (200100 mm2) were used in Ar+C2H2+N2 mixtures, with the substrates rotating at 70 mm over the target at a constant speed of 7 rpm. Argon flow was kept constant at 60 sccm while the reactive gases fluxes, C2H2 and N2, were changed in the ranges of 5-11 and 5-12 sccm, respectively, in order not to change significantly the C and N contents. This represents a variation of the working pressure of the deposition chamber between 0.31 to 0.47 Pa. The films were grown at a constant 5 temperature (300 ºC) and bias voltage (-70 V). Varying the density of current applied to each magnetron and the chemical composition of the mixed Ti/Ag target, a set of samples was prepared with Ag content varying from 0 to 26.7 at. %. Further details about the synthesis conditions can be found elsewhere [18]. The atomic composition of the deposited samples was measured by electron probe microanalysis (EPMA) using a Cameca SX 50 apparatus. Ball crater tests were used to obtain the film thickness. The structure and phase distribution of the coatings were determined by X-ray diffraction (XRD) using a conventional Philips PW 1710 diffractometer, operating with Cu Kα radiation, in a Bragg–Brentano configuration. Xray photoelectron spectroscopy (XPS) was measured using a hemispherical analyzer (SPECS EA-10 Plus) and Al K radiation as exciting source at a constant power of 300 W. The pass energy was 15 eV giving a constant resolution of 0.9 eV. The Ag 3d5/2 line at 367.9 was used to calibrate the binding energies. The samples were sputtercleaned in situ using a broad 3 keV Ar+ beam for 10 minutes. Hardness measurements were conducted using a MicroMaterials Nanotest system equipped with a Berkovich indenter applying a maximum load of 10 mN. Correction of the geometrical defects in the tip of the indenter, thermal drift of the equipment and uncertainty of the initial contact was done [19]. The residual stresses, , were obtained by the deflection method from the Stoney’s equation, using substrate curvature radii, both before and after coating deposition [20]. The tribological properties were evaluated by reciprocating friction tests using alumina-6-mm balls in diluted fetal bovine serum (FBS, 10% solution in water) in a CSM tribometer. The test parameters were set to 0.5 N of applied load; 3 mm/s of linear speed over a track length of 3 mm and 3000 cycles. The specific film wear rate was estimated after dividing the worn volume by the applied load and the sliding distance. Scanning electron microscopy (SEM) and energy 6 dispersive X-ray analysis (EDX) of the friction contact region were recorded in a FEG Hitachi S5200 microscopes operating at 5 keV. Raman spectra measurements (2002000 cm-1) were carried out in a LabRAM Horiba Jobin Yvon spectrometer equipped with a CCD (charge-coupled device) detector and a He-Ne laser (532 nm) at 5 mW. Cytotoxicity tests were performed using fibroblasts 3T3 (CCL-163) obtained from American Type Cell Collection. Coated coupons (previously sterilized at 121ºC for 15 min) were inserted in six well plates and 3 ml of Dulbecco modified eagle medium DMEM (Gibco) were added to each well. The plates with the materials were then incubated with 5% CO2 at 37ºC for 7 days. Meanwhile, the cells were grown in DMEM containing 10% of FBS (Gibco) and 1% penicillinstreptomicin, PS (Gibco). The cells were allowed to grow until attaining 80% confluence and after detachment 500 μL of cell suspension with 1x105 cells/ml were added to each well of a 24 wells' plate. After 7 days of material contact with the medium, 500 μL were removed from each well and added to the plates with cells. The plates were incubated with 5% CO2 at 37ºC for 48h. After that time, all the medium was removed and a solution containing 100 μL of MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2Htetrazolium, inner salt (Promega CellTiter 96® AQueous Non-Radioactive Cell Proliferation Assay) and 1 ml of DMEM without phenol red was added to each well. After 1 h, the absorbance of the resulting solution was read at 490 nm. The percentage of fibroblasts death was determined by the ratio of the difference between cell growth in the absence of Ag-TiCN sample (control – 100%) and the growth in the presence of a sample over the control growth. The assays were performed at least three times and in triplicate. 3. Results and discussion 7 3.1 Coating chemistry and microstructure The chemical composition of the deposited coatings obtained by EPMA is shown in Table 1 together with the film thicknesses and the result obtained after evaluation of the tribo-mechanical properties. The carbon content do not vary significantly from 30 at.%. The N content decreases slightly from 33.9 to 21.8 at. %, the Ti content decreases from 36.9 to 17.9 at. % while the Ag content increases from 0 to 26.7 at %. Oxygen appears as contaminant in a range of 4-8 at.%, usually below 5 at.%. Since titanium and silver display an opposite trend, the Ag/Ti ratio will be used hereafter to label the samples and to discuss the changes observed in the structure and functional properties. The XRD patterns obtained for the different Ag/Ti ratios are shown in Fig. 1. In the first sample (non-containing Ag) exhibit three reflections at 36º, 42º and 61º corresponding to the typical (111), (200) and (220) peaks originated by a cubic lattice whose positions lie intermediate between those for bulk TiC and TiN phases. By considering that the incorporated silver cannot form carbide or nitride phases and the complete mutual solid solubility of the TiN-TiC system, the formation of a TiCyN1-y phase is foreseen. A contribution to the peak shift due to the compressive residual stress in the coatings can be also possible. With the incorporation of silver, the TiCyN1-y diffraction peaks get broader and lose intensity concomitantly with the development of the characteristic pattern of metallic silver. In particular, for Ag/Ti ratios above 0.40 the main identified peaks correspond to fcc-Ag. The grain size of TiCyN1-y and Ag phases was determined by Scherrer formula using the (111) peak. The calculated values vary from 15 down to 9 nm and from 5 up to 8 nm, for TiCyN1-y and Ag respectively, with the increase of Ag/Ti. Under these premises, taking into account the atomic compositions given in Table 1, an excess of C and N is noticed over the Ti content and this difference becomes more significant as the Ag content increases. This can be 8 understood by assuming the formation of amorphous carbon-based phases (a-C and aCNx) where C atoms are connected to C or N atoms preferentially by sp2 bonds, as observed in previous publications [21,22]. In order to obtain further information about these carbon-containing phases, XPS analysis was carried out on the C 1s and N 1s photoelectron spectra of Ag-TiCN coatings. Fig. 2a shows the C1s spectra for samples with Ag/Ti atomic ratios of 0, 0.20, 0.40 and 1.49. The spectra can be deconvoluted into four components at 282.0 (C-Ti), 283.2 (C-TiO), 285.1 (C-C) and 286.9 eV (C-N) bonds. The first conclusion that can be drawn from the analysis is the predominance of an amorphous carbon phase originated by the C2H2 precursor although a certain contribution from adventitious carbon cannot be discarded. The relative contribution of the carbide components (C-Ti and C-TiO) continuously decreases as the silver content increases, especially for the highest Ag/Ti ratios (0.40, 1.49), where the contribution of the C-C and C-N peaks is significantly higher. This results in agreement with the conclusions obtained previously attending to the changes denoted in chemical compositions and XRD data. In the N1s spectra of Fig. 2b, two contributions were found at 397.0 eV and 399.2 eV ascribed to N-Ti and N-C bonds, respectively. The intensity of the N-C component becomes significant when the Ag/Ti increases up to 0.40 and 1.49 (in agreement with the results for C1s spectra). By using the data obtained from the deconvolution of the N1s and the elemental chemical composition measured by EPMA it was possible to estimate the phase composition as a function of the Ag/Ti ratio. The obtained results are plotted in Fig. 3. The amorphous CNx is slightly increasing while the main changes are noticed in the crystalline phases where the increment of Ag correlates with a diminution of the TiCyN1-y phase. This multiphase structure will certainly have an influence on the functional properties of the coatings (tribological, mechanical and biological) as we revise in the next sections. 9 Figure 4 shows the cross-sectional SEM micrographs of fractured Ag-TiCN samples with Ag/Ti ratios of 0, 0.20, 0.40 and 1.49. The film morphology displays a typical columnar-like microstructure although finer columns and porosity is observed as the Ag/Ti atomic ratio increases. This denser microstructure may be attributed to the segregation of the immiscible Ag and a-C(N) phases leading to renucleation sites that disrupts the columnar morphology. Similar behavior has been observed previously in Ag-CrN nanocomposites [23]. At the highest Ag/Ti ratio (Fig. 4d), Ag clusters are segregated to the column boundaries as observed before [14] which can be identified as brighter spots on the surface. 3.2. Mechanical properties: Hardness and residual stress The hardness and internal stress values are presented in Fig. 5 as a function of the Ag/Ti ratio. The hardness varied in a range from 8 to 18 GPa, much lower than the typical values reported for pure TiCN coatings, ranging from 30 to 36 GPa [24-26]. These overall lower values should be certainly related with microstructural and chemical composition aspects. Thus, low energetic conditions of the deposition (low negative substrate bias and low deposition temperature) can lead to columnar and open morphologies. The phase composition of these nanocomposites is also affecting the dependence of the mechanical properties as it influences the balance between hard and soft phases and their distribution. Thus, according to the achieved accuracy increasing Ag content up to 6.3 at.% does not almost affect the hardness; however, further increase is accompanied with a significant hardness decrease. A similar behaviour of soft metals has been reported in the past in other systems, such as ZrN/Cu [27], ZrN/Ni [28], or others including silver, Ag/TiN [5] and Ag/TiC [29]. However, it is also likely that ioninduced defects in the films during the deposition or more compact film microstructure 16 [18] N. K. Manninen, R. Escobar Galindo, N. Benito, N.M. Figueiredo, A. Cavaleiro, S.Carvalho, “Ag-Ti(C,N)-based coatings for biomedical applications: influence of silver content on the structural properties”, Journal of Physics D: Applied Physics 44 (2011) 375501. [19] J.M. Antunes, A. Cavaleiro, L.F. Menezes, M.I. Simões, J.V. Fernandes, Surf. Coat. Technol. 149 (2002) 27. [20] G. Stoney, Proc. Roy. Soc. London A 82 (1909) 172. [21] D. Martínez-Martínez, C. López-Cartes, A. Fernández, J.C. Sánchez-López, Surf. Coat. Technol. 203 (2008) 756. [22] D. Martínez-Martínez, C. López-Cartes, A. Justo, A. Fernández, J. C. SánchezLópez, Solid State Sciences 11(2009) 660. [23] C.P. Mulligan, T.A. Blanchet, D. Gall,, Surf. Coat. Technol. 203 (2008) 584. [24] S.J. Bull, D.G. Bhat, M.H. Staia, Surf. Coat. Technol. 163 –164 (2003) 499. [25] A.P. Serro, C. Completo, R. Colaço, F. dos Santos, C. Lobato da Silva, J.M.S. Cabral, H. Araújo, E. Pires, B. Saramago, Surf. Coat. Technol. 203 (2009) 3701. [26] E.J. Bienk, H. Reitz, N.J. Mikkelsen, Surf. Coat. Technol. 76 - 77 (1995) 475. [27] J. Musil, Surf. Coat. Technol. 125 (2000) 322. [28] J. Šůna, J. Musil, V. Ondok, J.G. Han, Surf. Coat. Technol. 200 (2006) 6293. [29] J.L. Endrino, J.J. Nainaparampil, E. Krzanowki, Scripta Mater. 47 (2002) 613. [30] E. Silva, M. Rebelo de Figueiredo, R. Franz, R. Escobar Galindo, C. Palacio, A. Espinosa, S. Calderon V. , C. Mitterer, S. Carvalho, Surf. Coat. Technol. 205 (2010) 2134. [31] J.M. Lackner, W. Waldhauser, R. Ebner, R.J. Bakker, T. Schöberl, B. Major, Thin Solid Films 468 (2004) 125. [32] C.P. Constable, J. Yarwood, W.-D. Münz, Surf. Coat. Technol. 116–119 (1999) 155–159. [33] J.C. Sánchez-López, A. Fernández, in: A. Erdemir, C. Donnet (Eds.), Tribology of Diamond-Like Carbon Films: Fundamentals and Applications, Springer, New York, 2008, p. 311. [34] K. Holmberg, A. Matthews, Coat. Tribology, Tribology series, 28 Ed. D. Dowson, Elsevier, 1994. 17 [35] J.C. Sánchez-López, D. Martínez-Martínez, M.D. Abad, A. Fernández, Surf. Coat. Technol. 204, 947-954 (2009). [36] J. L. Endrino, J.C. Sánchez-López, R. Escobar Galindo, D. Horwat, A. Anders, Journal of Physics: Conference Series 252 (2010) 012005. [37] G. Malich, B. Markovic, C. Winder, Toxicology 124 (1997) 179. [38] P. V. AshaRani, G. L.K. Mun, M. P. Hande, S. Valiyaveettil, ACS Nano. 3 (2009) 279. [39] K. Kawata, M. Osawa, S. Okabe, Environ. Sci. Technol. 43 (2009) 6046. 18 Figure Captions Fig. 1. XRD patterns of the Ag-TiCN coatings deposited by DC reactive magnetron sputtering, with different Ag/Ti atomic ratios. Fig. 2. C1s (a) and N 1s (b) photoelectron spectra for 4 representative coatings with Ag/Ti ratios of 0, 0.20, 0.40 and 1.49. Fig. 3. Phase composition of the samples prepared with Ag/Ti ratios of 0, 0.20, 0.40 and 1.49. Fig. 4. Cross-sectional SEM micrographs for samples with atomic ratios Ag/Ti: a) 0; b) 0.20; c) 0.40 and d) 1.49. Fig. 5. Hardness and residual stress values of Ag-TiCN coatings vs. the Ag/Ti atomic ratio. Fig. 6. Variation of the hardness and wear rate properties as a function of the ratio of soft (Ag+CNx)/hard (TiCyN1-y) phases. Fig. 7. Friction coefficient (a) and wear rate (b) values of Ag-TiCN as a function of the Ag/Ti atomic ratio. 19 Fig. 8. Optical micrograph of the alumina ball after the friction test onto the Ag-TiCN coating with 10.8 at. % of Ag (a) and corresponding Raman spectra (b) and EDX (c) analysis. Fig. 9. Optical micrograph of the wear track after the friction test of the Ag-TiCN coating with 10.8 at. % of Ag (a) and corresponding Raman spectra (b). Fig. 10. The rate of inhibition of cellular as a function of the Ag/Ti atomic ratio, after 7days incubation. Table 1. Chemical composition and tribomechanical properties of the deposited Ag-TiCN coatings. Sample Chemical composition at. % Thickness m H GPa  GPa  K mm3/Nm Ag/Ti Ag Ti C N 0 0 36.9 29.2 33.9 2.9 17.0 -0.5 0.29 7.1E-06 0.06 2.0 35.1 29.9 33.0 2.0 17.8 -1.0 0.31 1.9E-05 0.15 4.6 31.1 32.4 31.9 2.4 18.2 -1.4 0.25 5.3E-06 0.20 6.3 31.6 30.5 31.6 1.4 18.0 -1.7 0.26 5.2E-06 0.27 8.0 29.2 29.5 33.3 1.5 16.5 -1.8 0.27 2.8E-05 0.40 10.8 26.9 31.2 31.1 3.0 15.0 -2.5 0.27 3.8E-05 0.77 19.8 25.7 30.6 23.9 2.9 10.0 -1.6 0.28 4.8E-05 1.49 26.7 17.9 33.6 21.8 3.2 8.0 -1.3 0.25 8.7E-05 Table 1 R1 Figure 1 Click here to download high resolution image Figure 2a Click here to download high resolution image Figure 2b Click here to download high resolution image Figure 3 Click here to download high resolution image Figure 4 Click here to download high resolution image