Hydroxyapatite Films Research Articles

Physical-chemical characterization and biological assessment of simple and lithium-doped biological-derived hydroxyapatite thin films for a new generation of metallic implants

We report on the synthesis by PLD of simple and lithium-doped biological-origin hydroxyapatite (HA) films. The role of doping reagents (Li2CO3, Li3PO4) on the morphology, structure, chemical composition, bonding strength and cytocompatibility of the films was investigated.SEM investigations of the films evidenced a surface morphology consisting of particles with mean diameters of (5–7) µm. GIXRD analyses demonstrated that the synthesized structures consisted of HA phase only, with different degrees of crystallinity, mainly influenced by the doping reagent type. After only three days of immersion in simulated body fluid, FTIR spectra showed a remarkable growth of a biomimetic apatitic film, indicative of a high biomineralization capacity of the coatings. EDS analyses revealed a quasi-stoichiometric target-to-substrate transfer, the values inferred for the Ca/P ratio corresponding to a biological apatite. All synthesized structures displayed a hydrophilic behavior, suitable for attachment of osteoblast cells. In vitro cell viability tests showed that the presence of Li2CO3 and Li3PO4 as doping reagents promoted the hMSC growth on film surfaces.Taking into consideration these enhanced characteristics, corroborated with a low fabrication cost generated by sustainable resources, one should consider the lithium-doped biological-derived materials as promising prospective solutions for a next generation of coated implants with rapid osteointegration.

The a-axis oriented Hydroxyapatite film of epitaxially growth mechanism.

The a-axis oriented Hydroxyapatite film of epitaxially growth mechanism.

Effect of thickness on optical and microwave dielectric properties of Hydroxyapatite films deposited by RF magnetron sputtering

This study present findings on the structural, optical and dielectric properties of polycrystalline Hydroxyapatite [HAp, Ca10(PO4)6(OH)2] films, deposited using radio-frequency (RF) magnetron sputtering. The X ray diffraction (XRD) studies revealed that the unit cell volume and crystallite size of the films deposited on quartz substrates enhanced with an increase in film thickness. The Young's modulus (Ehkl) and the Poisson ratio (νhkl) of the thin films along different crystallographic directions have been calculated using the X-ray elastic constants. The Young's modulus of the films exhibited crystallographic direction dependence which suggests that the sputtered films are anisotropic. The dielectric constant εr and the loss tangent tanδ of the sputtered films were in the range 29–85 and 0.0028–0.0014 respectively at a frequency of 1 MHz. These values are by far the best for films deposited under a pure argon environment. The improvement in crystallinity and uniform grain size of the films are used as parameters to understand the variation in dielectric properties of the sputtered films. The best microwave dielectric properties were in the range εr = 75 - 65 and tanδ = 0.014–0.032 when measured at 5 and 10 GHz, employing split post dielectric resonator (SPDR) technique. The obtained results suggest that HAp sputtered films can be promising for optical limiting and applications in tunable microwave devices.

Application of fluoridated hydroxyapatite thin film coatings using KrF pulsed laser deposition.

Fluoridated hydroxyapatite (FHA) was investigated for application as an implant coating for titanium bone substitute materials in dental implants. A KrF pulsed excimer deposition technique was used for film preparation on a titanium plate. The compacts were ablated by laser irradiation at an energy density of 1 J/cm2 on an area 1×1 mm2 with the substrate at room temparature. Energydispersive spectrometric analysis of the FHA film revealed peaks of fluorine in addition to calcium and phosphorus. X-ray diffraction revealed the presence of crystalline FHA on the FHA film after a 10 h post annealing treatment at 450°C. The FHA film coating exhibited significant dissolution resistance to sodium phosphate buffer for up to 21 days, and favorable cell attachment of human mesenchymal stem cells compared with HA film. The results of this study suggest that FHA coatings are suitable for real-world implantation applications.

Electrocatalytic properties of hydroxyapatite thin films electrodeposited on stainless steel substrates

In this work, we have investigated the structural, microstructural, and electrocatalytic properties of hydroxyapatite (HAp) thin films. The HAp films were electrodeposited on stainless-steel (SS) substrates by chronopotentiometry mode from an electrolytic solution. The HAp films were characterized by X-ray diffraction (XRD), scanning electron microscopy (SEM) and X-ray energy dispersion spectroscopy (EDS). The electrodeposition and electrochemical processes of the hydroxyapatite (HAp) phase were studied by cyclic voltammetry. This HAp/SS system acting as an anode was used for the first time to electrodegrade Rhodamine B (RhB) in aqueous solutions. To follow the degradation kinetics, we used UV-visible spectroscopy. Several parameters such as the current density and the initial concentration of electrolytic solution were determined to optimize the electrodegradation of RhB. The decrease of RhB concentration followed pseudo-first order kinetics law. The rate of RhB degradation in presence of HAp/SS electrode can reach interesting high performance, depending on the electrocatalysis experimental conditions.

Characterization of Hydroxyapatite (HA) Sputtering Targets by APS Methods

Radio frequency (RF) sputtering is a potential medical device coating technology that is commercializable; however, a suitable commercialized target for sputtering the hydroxyapatite (HA) coating onto titanium medical devices is more important. Therefore, this study used three HA targets in conducting sputtering experiments for HA films, which were manufactured in a laboratory by using three different processes: cold pressing and sintering (CPS), hot isostatic pressing (HIP), and atmospheric plasma spraying (APS). Subsequently, the sputtering performance of each type of target and the properties of the HA films were assessed to develop an appropriate process for modifying the surfaces of medical devices. The experimental results showed that the APS target, with a density of approximately 2.83 g/cm3, was suitable for use in HA sputtering. Additionally, the APS target could withstand a high discharge power over 300 W, whereas the CPS target could nearly endure a power below 70 W. The APS target, with Ca/P ratio of 2.401, consisted of a combination of HA, α-tricalcium phosphate (α-TCP), β-TCP, and tetracalcium phosphate phases (TTCP). In addition to being able to perform at a high sputtering power of more than 300 W, the APS target achieved a higher deposition rate than did the CPS target. This study shows that the processing technology used for the APS target is a potential method for applying HA sputtering for the surface modification of artificial aggregates.

Adhesive Evaluation by Brushing Tests for Hydroxyapatite Films Fabricated on Dentins Using a Water Mist Assisted Er:YAG Laser Deposition Method

Hydroxyapatite (HAp) is one of the main structural components in tooth. Therefore, the HAp is an ideal material to repair tooth substances. However, it has not yet been realized that tooth enamel and dentin are artificially repaired using the HAp in operative dentistry. We have proposed and developed a tooth restoration technique with intraoral laser ablation using an Er:YAG (Erbium: Yttrium-Aluminum-Garnet) laser in order to fabricate HAp films on tooth surfaces in the atmosphere at room temperature. Although HAp thin films were formed on dentin surfaces using the Er:YAG laser deposition method, it has still been unclear how is the mechanical durability of the HAp film on tooth substances. In the present study, we conducted brushing tests to evaluate the durability of adhesion between the HAp film and dentin. To form HAp layers on dentin surfaces, an ablation phenomenon was induced by the Er:YAG laser irradiated to the target of α-tricalcium phosphate (α-TCP) under humid conditions. The deposited layers were hydrolyzed by dripping artificial saliva on the surface at 37 degrees Celsius for 24, 48, 72, 120, and 168 hours to create HAp films. The films hydrolyzed for more than 48 hours were attached on the dentin surface after the brushing tests. This result indicates that the α-TCP layer deposited on the dentin surface changes into a crystallized HAp material by the hydrolyzation for more than 48 hours. The crystallization of the film possibly contributes to the high adhesive durability of the HAp film deposited on the dentin.

The role of lower hybrid resonance and helicon waves excitations in a magnetized plasma for coating production of complex crystalline structures as hydroxyapatite

The non-equilibrium electro-thermodynamic properties of magnetized plasma produced by a modified radio-frequency magnetron sputtering system were investigated in detail. The plasma was analyzed using Hall, Langmuir, and Faraday probes and optical emission spectroscopy, and numerical simulations were conducted to determine the electric and magnetic field profiles. It is found that the occurrence of lower hybrid resonance and excitation of helicon waves are satisfied locally along the plasma profile (Z). This unusual effect in magnetron sputtering systems plays an important role in the increase of the electrons and ions temperature improving the quality of the films produced. Films of complex structure as hydroxyapatite that have biomedical applications were produced along the Z to confirm our plasma results. The films were characterized by performing grazing incidence X-ray diffraction using synchrotron radiation, X-ray photoelectron spectroscopy and atomic force microscopy. The microstructure analysis of hydroxyapatite films produced at different Z revealed that films with high crystallinities and with ideal stoichiometries (Ca/P = 1.67 ± 0.05) were produced within a magnetic cusp region (26 mm < Z < 32 mm). Based upon the results from plasma diagnostics and films characterization, we proposed a model of plasma of this system. The delivered energy by ions bombardment for films formation was determined and reached values around 8 eVs−1Å−2 and plasma temperature of T ≈ 10 eV inside the magnetic cusp region. Finally, this work opens the possibility to produce coatings of other complex compounds by the fine-tuning of resonant waves and their high delivered energies in vacuum magnetized plasmas.

Preparation of bio-polyurethane using castor oil and antibacterial hybrid films thereof with silver-doped hydroxyapatite

The hybrid films of bio-polyurethane (Bio-PU) and silver-doped hydroxyapatite (HA-Ag) for breathable and antibacterial textile applications. The Bio-PU was synthesized using a mixture of castor oil based polyol with petroleum based poly(ethylene glycol). Silver doping to hydroxyapatite(HA) was carried out through ion exchange mechanism between calcium ion in HA and silver ion (Ag+) in aqueous solution of AgNO3. The concentration of Ag+ was controlled to 100-300 ppm. The existence of silver in HA was proved using SEM-EDS while the silver doping amount was estimated by measuring residual concentration of Ag+ after doping using ICP-OES. It was found that the hybrid films exhibited excellent antibacterial activity against bacteria of S. aureus and K. pneumonia by showing 99.9 % reduction of bacteria.

Effect of laser fluence and ambient gas pressure on surface morphology and chemical composition of hydroxyapatite thin films deposited using pulsed laser deposition

The dependence of the surface morphology and chemical composition of hydroxyapatite (HA) thin films on the laser fluence and ambient gas pressure during their formation by pulsed laser deposition was studied as the first step to investigate the effect of physical and chemical interactions between the ablated chemical species and ambient gas molecules on HA film formation. It was found that a higher fluence could decrease the number of large protrusions on the surface of HA thin films. However, too high a fluence caused a phosphorus deficiency from the stoichiometric value, particularly in the case of lower ambient gas pressure. It was also found that for lower fluences, the atomic species among the ablated chemical species were easily scattered by collision processes with ambient gas molecules. This was caused by the lower velocity of the ablated chemical species and higher ambient gas pressure, which induced a shorter mean free path. In addition, these collision processes played an important role in the adsorption, migration, and re-evaporation of the ablated chemical species on the substrate via chemical reactions.

Characteristics of RF-Sputtered Thin Films of Calcium Phosphate on Titanium Dental Implants

Hydroxyapatite (HA) coatings on titanium have been investigated for many years, and have demonstrated advantageous biocompatibility in dental implants. Animal experiments have demonstrated that the biological response to plasma-sprayed HA-coated implants shows disadvantages in terms of adherence, thickness uniformity, and long-term osseointegration effects. Determining how to resolve the degradation problem of HA in the body by improving osseointegration and stability in alveolar bones has become an increasingly researched topic. The present study investigated the film characteristics and dissolution properties of calcium phosphate (CaP) coatings obtained by radio-frequency (RF) sputtering of a self-developed atmospheric plasma spray (APS) HA target. The experimental parameters varied, including RF power (60–250 W), sputtering time (15–120 min), and substrate roughness (0.4–4 μm). Analyses were conducted using synchrotron X-ray diffraction, Fourier transform infrared spectroscopy (FTIR), white light interferometry, and scanning electron microscopy combined with energy-dispersive X-ray spectroscopy (EDS). EDS analysis showed that the Ca/P ratio increased as the discharge power was increased. The analysis results also showed that a lower surface roughness resulted in higher crystallinity, because a larger surface-free energy was attained during sputtering. In-plane texturing has been proven when HA films are sputtered onto substrates of varying roughness, within appropriate deposition parameters. FTIR analysis revealed the presence of the principal PO43- bonds in the deposited calcium phosphate films. The CaP films induced calcium phosphate precipitation when immersed in simulated body fluid (SBF), suggesting that, based on in vitro bioactive behavior, the proposed combined surface modification can be used in dental implants.

Sol gel-derived hydroxyapatite films over porous calcium polyphosphate substrates for improved tissue engineering of osteochondral-like constructs.

Articular cartilage interfaces with bone through a zone of calcified cartilage. This study describes a method to generate an "osteochondral-like" implant that mimics this organization using isolated deep zone cartilage cells and a sol-gel hydroxyapatite coated bone substitute material composed of calcium polyphosphate (CPP). Developing a layer of calcified cartilage at the interface should contribute to enhancing the success of this "osteochondral-like" construct following implantation to repair cartilage defects.

Effect of ablation laser pulse repetition rate on the surface protrusion density of hydroxyapatite thin films deposited using pulsed laser deposition

The effect of the pulse repetition rate of the ablation laser on surface protrusion density was investigated for hydroxyapatite thin films deposited using pulsed laser deposition. Atomic force microscope surface images showed a marked decrease in surface protrusion density at lower pulse repetition rate, while protrusion-free areas exhibited similar morphologies independent of the pulse repetition rate. These results indicate that the surface protrusions were not formed or enlarged from surface migration of atomic species on the surface, which formed the smooth surface of the protrusion-free area. The postulated origin of the protrusions was large particles or droplets produced directly from the target irradiated with the ablation laser. These results suggest that the optimum pulse repetition rate for protrusion-free hydroxyapatite thin film surfaces is ≤3Hz.

Fabrication of oriented hydroxyapatite film by RF magnetron sputtering

Hydroxyapatite (HAp) is compatible with bone tissue and is used mainly as a bone prosthetic material, especially as the coating of implants. Oriented HAp film is expected to be a high-quality epitaxial scaffold of the neonatal bone. To fabricate highly oriented HAp thin films via the conventional plasma process, we deposited the HAp film on a Ti coated silica glass substrate using RF magnetron sputtering in low substrate temperature conditions. The X-ray diffraction pattern of the film sample consisted of an intense (002) peak, corresponding to the highly oriented HAp. The (002) peak in XRD diagrams can be attributed either to the monoclinic phase or the hexagonal phase. Pole figure analysis showed that the (002) plane grew parallel to the surface of the substrate, without inclination. Transmission Electron Microscope analysis also showed the fabrication of aligned HAp crystallites. The selected area diffraction patterns indicated the existence of monoclinic phase. The existence of hexagonal phase could not be judged. These results indicate the uniaxial films fabricated by this technique enable to be the epitaxial scaffold of the neonatal bone. This scaffold can be expected to promote connection with the surrounding bone tissue and recovery of the dynamic characteristics of the bone.

Growth of a TiNb adhesion interlayer for bioactive coatings

Surface bioactivity has been under intensive study with reference to its use in medical implants. Our study is focused on coatings prepared from an electroactive material which can support bone cell adhesion. Until now, hydroxyapatite films have usually been utilized as a chemically-active surface agent. However, electrically-active films could set a new direction in hard tissue replacement. As a base for these films, it is necessary to prepare an intermediate film, which can serve as a suitable barrier against the possible diffusion of some allergens and toxic elements from the substrate. The intermediate film also improves the adaptation of the mechanical properties of the basic material to an electroactive film. The aim of our work was to select an implantable and biocompatible material for this intermediate film that is suitable for coating several widely-used materials, to check the possibility of preparing an electroactive film for use on a material of this type, and to characterize the structure and several mechanical properties of this intermediate film. TiNb was selected as the material for the intermediate film, because of its excellent chemical and mechanical properties. TiNb coatings were deposited by magnetron sputtering on various substrates, namely Ti, Ti6Al4V, stainless steel, and bulk TiNb (as standard), and important properties of the layers, e.g. surface morphology and surface roughness, crystalline structure, etc., were characterized by several methods (SEM, EBSD, X-ray diffraction, nanoindentation and roughness measurement). It was found that the structure and the mechanical properties of the TiNb layer depended significantly on the type of substrate. TiNb was then used as a substrate for depositing a ferroelectrically active material, e.g., BaTiO3, and the adhesion, viability and proliferation of human osteoblast-like Saos-2 cells on this system were studied. We found that the electroactive BaTiO3 film was not only non-cytotoxic (i.e. it did not affect the cell viability). It also enhanced the growth of Saos-2 cells in comparison with pure TiNb and with standard tissue culture polystyrene wells, and also in comparison with BaTiO3 films deposited on Ti, i.e. a material clinically used for implantation into the bone.

Influence of Heating Conditions for Formation of a Thin Apatite Film on Zirconia Using a Molecular Precursor Method

The influence of heating conditions, heating temperature, and heating time on the formation of a thin carbonate-containing hydroxyapatite (CA) film onto partially stabilized zirconia using a molecular precursor method was evaluated. The molecular precursor solution was prepared from a mixture of calcium-ethylenediaminetetraacetic acid complex and phosphate compounds at Ca/P ratio of 1.67. After the application of molecular precursor solution onto zirconia, four different heating conditions—namely, 600 °C-2 h, 800 °C-2 h, 1000 °C-2 h, and 600 °C-4 h—were applied. No distinct difference of surface appearance of CA coating was observed between 600 and 800 °C-2 h. Fusion of apatite crystals was observed at 1000 °C-2 h. Surface roughness of CA film at 1000 °C-2 h was significantly higher than those under other heating conditions. Heating at 800 °C produced a significantly more hydrophilic surface and higher degree of crystallization. No significant differences were recognized in the critical load at the first crack in the coating among the four samples by scratch tests. After 30 days’ immersion in phosphate buffered saline, the four different CA coating films were still present. Simulated body fluid immersion experiments were performed as in vitro biocompatibility tests. After 48 h immersion, the CA film at 800 °C-2 h showed a greater amount of spherical crystal precipitation. It was suggested that properties of CA coating on partially stabilized zirconia using a molecular precursor method were influenced by the heating temperature and time.

Adhesion profile and differentiation capacity of human adipose tissue derived mesenchymal stem cells grown on metal ion (Zn, Ag and Cu) doped hydroxyapatite nano-coated surfaces

Accelerated Mesenchymal Stem Cells (MSCs) condensation and robust MSC-matrix and MSC-MSC interactions on nano-surfaces may provide critical factors contributing to such events, likely through the orchestrated signal cascades and cellular events modulated by the extracellular matrix. In this study, human adipose tissue derived mesenchymal stem cells (hMSC)', were grown on metal ion (Zn, Ag and Cu) doped hydroxyapatite (HAP) nano-coated surfaces. These metal ions are known to have different chemical and surface properties; therefore we investigated their respective contributions to cell viability, cellular behavior, osteogenic differentiation capacity and substrate-cell interaction. Nano-powders were produced using a wet chemical process. Air spray deposition was used to accumulate the metal ion doped HAP films on a glass substrate. Cell viability was determined by MTT, LDH and DNA quantitation methods Osteogenic differentiation capacity of hMSCs was analyzed with Alizarin Red Staining and Alkaline Phosphatase Specific Activity. Adhesion of the hMSCs and the effect of cell adhesion on biomaterial biocompatibility were explored through cell adhesion assay, immunofluorescence staining for vinculin and f-actin cytoskeleton components, SEM and microarray including 84 known extracellular matrix proteins and cell adhesion pathway genes, since, adhesion is the first step for good biocompability. The results demonstrate that the viability and osteogenic differentiation of the hMSCs (in growth media without osteogenic stimulation) and cell adhesion capability are higher on nanocoated surfaces that include Zn, Ag and/or Cu metal ions than commercial HAP. These results reveal that Zn, Ag and Cu metal ions contribute to the biocompatibility of exogenous material.

Comparative physical, chemical and biological assessment of simple and titanium-doped ovine dentine-derived hydroxyapatite coatings fabricated by pulsed laser deposition

We report on the synthesis by Pulsed Laser Deposition of simple and Ti doped hydroxyapatite thin films of biological (ovine dentine) origin. Detailed physical, chemical, mechanical and biological investigations were performed. Morphological examination of films showed a surface composed of spheroidal particulates, of micronic size. Compositional analyses pointed to the presence of typical natural doping elements of bone, along with a slight non-stoichiometry of the deposited films. Structural investigations proved the monophasic hydroxyapatite nature of both simple and Ti doped films. Ti doping of biological hydroxyapatite induced an overall downgrade of the films crystallinity together with an increase of the films roughness. It is to be emphasized that bonding strength values measured at film/Ti substrate interface were superior to the minimum value imposed by International Standards regulating the load-bearing implant coatings. In vitro tests on Ti doped structures, compared to simple ones, revealed excellent biocompatibility in human mesenchymal stem cell cultures, a higher proliferation rate and a good cytocompatibility. The obtained results aim to elucidate the overall positive role of Ti doping on the hydroxyapatite films performance, and demonstrate the possibility to use this novel type of coatings as feasible materials for future implantology applications.

Plasma-assisted deposition of bone apatite-like thin films from natural apatite

In orthopedics and dentistry, novel approaches for fabricating biomimetic and mechanically robust bioactive coatings are highly desirable in order to truly improve the clinical results of coated implants compared to uncoated ones. In this paper, a biological-like apatite coating is deposited for the first time by plasma-assisted deposition of a natural apatite source. Specifically, we deposited bone apatite-like (BAL) thin films from bone apatite targets by pulsed electron deposition (PED). Morphology, composition, structure and mechanical properties of as-deposited and annealed BAL and stoichiometric hydroxyapatite (HA) films were investigated. While as-deposited BAL and HA films were poorly crystalline at room temperature, they crystallized to an extent very close to that of natural apatite when annealed at 400°C. In addition, FTIR analysis pointed out that BAL films closely resembled the composition of the starting natural apatite target. Finally, nanoindentation tests indicated that BAL films with high mechanical properties could be deposited by PED.

In vivo study on the biocompatibility of chitosan-hydroxyapatite film depending on degree of deacetylation.

Chitosan, produced from chitin, is one of the polymers with promising applications in various fields. However, despite diverse research studies conducted on its biocompatibility, its uses are still limited. The main reason is the degree of deacetylation (DOD), which represents the proportion of deacetylated units in the polymer and is directly correlated with its biocompatibility property. In this article, the in vivo biocompatibility of three chitosan-hydroxyapatite composite films composed of chitosan with different DOD values was investigated by traditional biological protocols and novel optical spectroscopic analyses. The DOD of the chitosan obtained from three different manufacturers was estimated and calculated by Raman spectroscopy, Fourier transform infrared spectroscopy, and proton nuclear magnetic resonance spectroscopy. The chitosan with the higher DOD induced a higher incidence of inflammation in skin cells. The amino group density, biodegradability, and crystallinity of chitosan are the three possible factors that need to be considered when determining the biocompatibility of the films for in vivo application, as they led to complicated biological results, resulting in either better or worse inflammation even when using chitosan products with the same DOD. This basic study on the relationship between the DOD and inflammation is valuable for the development of further chitosan-based researches. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1637-1645, 2017.