HAp Nanocrystals Research Articles

Synthesis, characterization and luminescence properties of Eu3+-doped hydroxyapatite nanocrystal and the thermal treatment effects

In this work, we present the synthesis, characterization and the luminescence properties of Ca10(PO4)6(OH)2 (hydroxyapatite/HAp) nanocrystals doped with europium trivalent ions. The most important processes that lead to europium emissions in the visible region were identified. Eu:HAp nanopowder excited at 394nm (or 460nm) exhibits several emissions: (i) weak emissions at 579nm, 592nm and 616nm due to the 5D0→7F0, 5D0→7F1 and 5D0→7F2 transitions, respectively, with europium ion occupying site I in hydroxyapatite structure and (ii) strong emissions due to the 5D0→7F0 (574nm), 5D0→7F1 (602nm) and 5D0→7F2 (610–630nm) transitions, when Eu3+ is occupying site II. The emission spectrum and the time-resolved luminescence analysis showed that the HAp nanocrystals (nanopowder) thermally treated at temperature (T) between 500 and 800°C have a change in the initial Eu3+ site distribution of 100 % of Eu3+ at site I to a more stable one where the majority of europium ions are at site II: 30% remains at site I and 70% migrates to site II. In addition, an enhancement of the Eu3+ emission intensity is observed due to the increasing crystallite size. A time-resolved luminescence investigation using a short pulse laser excitation at 460nm was employed to measure the luminescence decays and to determine the most important mechanisms involved in the deexcitation process of 5D0 excited state of Eu3+, where it is seen a fast (2.9μs) energy transfer from Eu3+- site I (donor) to Eu3+- site II (acceptor) in the thermally treated nanopowders with T>500°C. The initial presence of 100% of Eu3+ at site I in the synthesized nanocrystals is gradually modified by the thermal treatments with temperatures above 500°C by thermal activation of Ca2+ vacancy (the charge compensator) diffusion through the HAp lattice, which propitiates the Ca2+- vacancies and Eu3+ ions to exchange positions in the lattice. By this thermal activated mechanism, Eu3+ ion migrates through the lattice until get the final distribution of 30% at site I and 70% at site II. As a result, the complete description of the Eu3+ (5D0) decay and the energy transfer process from Eu3+ (site I)→Eu3+ (site II) were proposed.

Control of one‐dimensional hydroxyapatite nanocrystals at mild conditions: organic additive effects

One-dimensional hydroxyapatite (HAp, Ca10(PO4)6(OH)2) nanocrystals have been successfully synthesised via the simple precipitation-hydrolysis process on mild solution conditions. At the first precipitation step, the precursor samples of monetite phase CaHPO4 were prepared with three kinds of organic additives. The results of X-ray diffraction (XRD) patterns show that the structure and crystallinity of CaHPO4 prepared with the different organic additives are different, which indicates that the organic additives exert influence on the crystal growth of CaHPO4. During the second process, the HAp nanocrystals with high aspect ratio grew up under a high pH condition. The XRD patterns show that the preference of every nanocrystal growth is the (002) plane. Transmission electron microscopy micrographs indicate that the HAp nanocrystals were affected by the reaction temperature and the pH values of the solution. The organic additives influenced the aspect ratio of HAp nanocrystals, and Fourier transform infrared spectra show that the organic additives were completely dissolved in the aqueous system. This indicates that the organic additives affected the formation of nucleation and growth of HAp nanocrystals, and the influence gradually decreased with the extension of the hydrolysis time, until the organic additives were detached from the HAp crystals.

Influence of morphology on the in vitro bioactivity of hydroxyapatite nanostructures prepared by precipitation method

It seems that the available studies about the effect of morphology on the bioactivity of pure hydroxyapatite (Ca10(PO4)6(OH)2, HAP) are yet insufficient. For this purpose, a novel morphology-controlled synthesis method to fabricate HAP nanostructures is employed by using different Schiff bases as chelating ligands. The formation mechanism of zero-dimensional (0-D) and one-dimensional (1-D) nanostructures of HAP is also considered. Based on the morphological studies of the final products, it is found that the formation of tetradentate and bidentate ligands leads to the preparation of 0-D and 1-D HAP nanocrystals, respectively. The in vitro bioactivity of the as-produced HAP nanostructures is evaluated by soaking the products in simulated body fluid (SBF). The analytical measurements indicate that the changes in the Ca and P concentrations of the 1-D HAP nanostructures are more than those of the 0-D HAP nanostructures, indicating good bioactivity of the nanorods and nanobundles. The formation of new apatite crystals on the surfaces of the HAP nanostructures soaked in the SBF solutions is proved by field emission scanning electron microscopy (FESEM) and diffuse reflectance Fourier transform infrared spectrometry (ATR-FTIR).

Fabrication of dispersible calcium phosphate nanocrystals via a modified Pechini method under non-stoichiometric conditions

Various techniques for preparing ceramic nanoparticles have been developed; however, most of them start from a preparation of precursor nanoparticles that are generally amorphous or in poorly crystallized phases. Thermal treatments used to obtain crystalline phases typically result in the sintering of the products into large polycrystals. In this study, we developed a process to fabricate dispersible hydroxyapatite (HAp; Ca10(PO4)6(OH)2) nanocrystals via a modified Pechini method, which is a sol–gel like solid-state synthesis method for the preparation of multicomponent oxides. We demonstrated that the HAp nanocrystals sintered into large polycrystals ranging in size from several tens to several hundreds of microns via a conventional Pechini method using the stoichiometric Ca/P molar ratio of 1.67. When the Ca/P molar ratio in the precursor was >1.67, a mixture of HAp nanocrystals and removable CaO matrix was obtained. The HAp nanocrystals were dispersed in aqueous media mostly in the form of nanoparticles when the amount of CaO matrix was sufficiently greater than the amount of HAp.

Strontium-Substituted Hydroxyapatite Nanocrystals Synthesized in Ultrasonic Field: Use for Protein Adsorption

Hydroxyapatite Ca10(PO4)6(OH)2(HAP) is widely applied in a biomaterial because of its excellent biocompatibility and bioactivity. However, all properties of HAP, especially the adsorption properties can be tailored by modifying the composition with ionic substitutions. Among many cations that can substitute for calcium in the structure of HAP, strontium is a kind of essential trace elements in human body. It has been known as one of the cationic substitute for calcium in the HAP lattice. The strontium-substituted hydroxyapatite (Sr-HAP) is formed when strontium is doped in HAP structure. The bioactivity, biocompatibility, and adsorption properties are improved when the original lattice of HAP is destroyed. In this study, we focus on synthesis of strontium-substituted HAP nanocrystals in ultrasonic field which is a mild and simple synthesis method. The Sr-HAP with different Sr contents was synthesized. The effects of reaction temperature, sintering temperature and reaction time on protein adsorption of Sr-HAP were studied. In addition, the crystalline phase, chemical compound, surface area and morphology were characterized by XRD, FTIR, TEM and BET. The results indicate that the original lattice of HAP was destroyed and the structure of HAP doped with strontium formed. Sr-HAP with smaller crystal size, larger specific surface area and homogeneous distribution was prepared. Especially, it has great adsorption to target protein, bovine serum albumin (BSA).

Hydroxyapatite nanocrystals: colloidal chemistry, assembly and their biological applications

In this review, recent advances in the tunable synthesis, ion doping, assembly and applications of monodisperse HAp nanocrystals are summarized.

Fabrication and characterization of hydroxyapatite/collagen bone-like nanocomposite through a self-assembly method

AbstractBased on a self-assembly mechanism, a co-precipitation method was utilized to fabricate bone-like biomimetic nanocomposite with a simplified preparation approach and accessible materials to investigate in depth some characteristics of hydroxyapatite/collagen(HAp/Col) nanocomposite for the elucidation of performances in some respects. The as-prepared composite was characterized by X-ray diffraction, scanning electron microscopy, transmission electron microscopy, Fourier transformation infrared spectroscopy, and thermal analysis. The results show that HAp nanocrystals formed as preferentially oriented slender needles 50–100 nm in length on a felt-like Col matrix which is composed of large numbers of randomly oriented Col fibers and showed polycrystalline behavior. The as-prepared cellular composites are analogous in both composition and nanostructured architecture to native bone, longer aging time promotes the growth and purification of nano-HAp on Col, and characterization confirms that chemical interaction occurs and causes intimate bonding between HAp and Col.

Synthesis and characterization of hydroxyapatite nanocrystal and gelatin doped with Zn2+ and cross linked by glutaraldehyde

Different nanocrystals of hydroxyapatite-[HAp]-gelatin [GEL], HAp/GEL/Glutaraldehyde (GA), HAp/GEL/Zn and HAp/GEL/GA/Zn were prepared using the biomimetic process. The nanocrystals were characterized by Scanning Electron Microscopy (SEM), Fourier Transform Infrared Spectroscopy (FTIR) and X-ray diffraction analysis (XRD). The FT-IR spectra of HAp/GEL nanocrystal materials are similar to the spectrum of real bone. Nanocrystals show (002), (222), (211), and (112) planes, which confirm formation of hydroxyapatite phase. Lattice parameters from X-ray diffraction spectra showed that the metal cation was incorporated into the hydroxyapatite structure and has substituted for calcium in the HAp crystal structure and all of the doped HAp formulations were biphasic and crystalline. The hydroxyapatite/gelatin/zinc (HAp/GEL/Zn) composite sintered in 900°C demonstrated that the c-axes of HAp nanocrystals were regularly aligned along collagen fibers. Key words: Hydroxyapatite, Nanocomposite, Zinc.

Preparation and Characterization of Calcium Phosphate Crystals by Precursor Thermolysis Method

In this paper, a simple precursor thermolysis method is applied to prepare calcium phosphate crystals and macroporous ceramic. The phase composition of sample was determined by XRD and the morphology of sample was observed by TEM and SEM. The macroporous ceramic was observed by 3D digital microscope. Results show that 25-100 nm HAP nanocrystals were obtained at 500°C and HAP rod-like crystals with an equivalent diameter of about 0.50-1.6μm and a length of about 5-10μm were obtained at 700°C. In addition, about 100 nm β-TCP nanoparticles were also obtained at 900°C. The colloidal precursors can also be used to fabricate macroporous calcium phosphate ceramic. The size of pores is about 100-200μm. Pores are well opened and interconnected in three dimensions.

Nano/Microstructural Effect of Hydroxyapatite Nanocrystals on Hepatocyte Cell Aggregation and Adhesion

Hepatocyte cell aggregation and adhesion to HAp nanocrystals covered with SU-8 polymer micropatterns by nano/microfabrication techniques is demonstrated. The surface roughness and wettability of the HAp nanocrystals are significantly different from those of the SU-8 polymer. QCM-D and microscopic observation clearly reveal that the cells realize the surface properties to form aggregation and preferentially adhere to the HAp nanocrystals at 2 h after seeding, indicating the importance of the microstructures as well as the interfacial phenomena at a nanometer scale.

<I>In Situ</I> Biomimetic Synthesis and Characterization of Nano Hydroxyapatite in Gelatin Matrix

In this work, nano hydroxyapatite (nHAp) composites were synthesized via an in situ biomimetic process at room temperature by using gelatin as a template agent. Formation of hydroxyapatite nanoparticles and the effect of biopolymer concentration on the shape and morphology of the nHAp nanoparticles were confirmed and investigated by using X-ray diffraction (XRD), Fourier Transform Infrared spectroscopy (FT-IR), Scanning and Transmission Electron Microscopy (SEM and TEM). The results indicated that, the gelatin biopolymer as a template agent in the preparation batch has influences the size and morphology of the HAp nanocrystals precipitated in an aqueous solution of gelatin.

Preparation of a Nanoscaled Poly(vinyl alcohol)/Hydroxyapatite/DNA Complex Using High Hydrostatic Pressure Technology for In Vitro and In Vivo Gene Delivery

Our previous research showed that poly(vinyl alcohol) (PVA) nanoparticles incorporating DNA with hydrogen bonds obtained by high hydrostatic pressurization are able to deliver DNA without any significant cytotoxicity. To enhance transfection efficiency of PVA/DNA nanoparticles, we describe a novel method to prepare PVA/DNA nanoparticles encapsulating nanoscaled hydroxyapatites (HAps) prepared by high hydrostatic pressurization (980 MPa), which is designed to facilitate endosomal escape induced by dissolving HAps in an endosome. Scanning electron microscopic observation and dynamic light scattering measurement revealed that HAps were significantly encapsulated in PVA/HAp/DNA nanoparticles. The cytotoxicity, cellular uptake, and transgene expression of PVA/HAp/DNA nanoparticles were investigated using COS-7 cells. It was found that, in contrast to PVA/DNA nanoparticles, their internalization and transgene expression increased without cytotoxicity occurring. Furthermore, a similar level of transgene expression between plasmid DNA and PVA/HAp/DNA nanoparticles was achieved using in vivo hydrodynamic injection. Our results show a novel method of preparing PVA/DNA nanoparticles encapsulating HAp nano-crystals by using high hydrostatic pressure technology and the potential use of HAps as an enhancer of the transfection efficiency of PVA/DNA nanoparticles without significant cytotoxicity.

Fabrication, Characterization and Cell Cultures on a Novel Composite Chitosan-Nano-Hydroxyapatite Scaffold

This paper deals with the characterizations made during the development of a nano-HAp loaded chitosan scaffold, obtained by the freeze-drying technique combined with a novel in situ crystal growth method. The nano-composites were characterized by a highly porous and interconnected structure. The XRD patterns and calculated domain sizes of the HAp nano-crystals nucleated on the chitosan scaffolds are very similar to the ones recorded for deproteinated bone apatite. Both osteoblasts (MG63) and mesenchimal cells (hMSC) were showing good proliferation and adhesion onto the scaffolds. The presence of extensive filopodia and excellent spreading in and around the interconnected porous structure, indicated a strong cellular adhesion and growth. Moreover a good hMSC osteogenic differentiation has been verified. The observations related to well-developed structure morphology, physicochemical properties and high cytocompatibility suggest that the obtained chitosan-nHA porous scaffolds are potential candidate materials for bone regeneration.

PREPARATION AND CHARACTERIZATION OF MONOSDISPERSED METAL PHOSPHATE PARTICLES AND CHARACTERISTICS OF PROTEIN ADSORPTION ONTO VARIOUS APATITE PARTICLES

There exists a unique class of orthophosphates in which both the phosphorous and metal atoms are tetrahedrally coordinated by oxygen, therefore, to form metal phosphates. Various kinds of metal phosphate particles have many applications of uses in pigments, catalysis, adsorbents, and bioceramics. It is well known that calcium hydroxy apatite(Hap), a kind of calcium phosphate particles, also shows a high affinity for proteins and much attention has been focused on Hap and substituted Hap nanocrystals as sorbents. In this review, I will present (1) the preparation and characterization of monodispersed metal phosphate particles and (2) characteristics of protein adsorption onto various kinds of apatite particles based on our previous investigations.

Carboxymethylation of the fibrillar collagen with respect to formation of hydroxyapatite

Control over crystal growth by acidic matrix macromolecules is an important process in the formation of many mineralized tissues. Highly acidic macromolecules are postulated intermediates in tissue mineralization, because they sequester many calcium ions and occur in high concentrations at mineralizing foci in distantly related organisms. A prerequisite for biomineralization is the ability of cations like calcium to bind to proteins and to result in concert with appropriate anions like phosphates or carbonates in composite materials with bone-like properties. For this mineralization process the proteins have to be modified with respect to acidification. In this study we modified the protein collagen by carboxymethylation using glucuronic acid. Our experiments showed unambigously, that N(epsilon)-carboxymethyllysine is the major product of the in vitro nonenzymatic glycation reaction between glucuronic acid and collagen. We hypothesized that the function of biomimetically carboxymethylated collagen is to increase the local concentration of corresponding ions so that a critical nucleus of ions can be formed, leading to the formation of the mineral. Thus, the self-organization of HAP nanocrystals on and within collagen fibrils was intensified by carboxymethylation.

Biomimetic hydroxyapatite nanocrystals in composites with C60 and Au-DNA nanoparticles: IR-spectral study

Abstract Due to growing interest towards development so-called “smart biomaterials”—mainly composites with targeted biological action consisting of inorganic nanoparticles and organic compounds-the primary tasks in this field include synthesis of proper nanoparticles and studying chemical interaction between them and organic constituents. Exactly such kind of research is the object of this study - synthesis of composites in the system HAP:DNA:Au:C 60 :H 2 O including components of this system—composites HAP:DNA:Au:H 2 O, DNA:Au:H 2 O, HAP:C 60 :H 2 O, HAP:BSF, C 60 :H 2 O and studying chemical interaction in this system by comparison of IR spectra. All materials in this system are typical ingredients of smart biomaterials. The HAP nanocrystals were synthesized by chemical synthesis in a biomimetic route, all other materials were acquired on the market. We chose the simplest method of the composite synthesis—by mixing at room temperature. Comparative study of IR spectra of these composites revealed the considerable distortion, shifting or full disappearance of some absorption bands of the components of the composites. The largest alterations were found in the range of deformation vibrations and they are different for different modes of vibrations. In the range of valence vibrations the components’ absorption bands are mostly preserved. These features are the ample evidence of chemical interaction between these components. Identification of the absorption bands that were changed most of all indicates the chemical bonds which directly participate in the chemical interaction. For example loss of the DNA absorption band at wave number 1704 cm −1 can serve as indication of new chemical bond with this part of the DNA molecule, namely C O bond in thymine. The spectra alteration and their dependence vs. components concentration in the system HAP–DNA reveal that the HAP nanoparticles supposably are localized due to interaction with DNA molecules and that results in significant alteration of the spectra of DNA deformation vibrations. When DNA concentration increases the surplus of DNA molecules cannot interact with HAP and their spectrum is not changed. Thus there is something like threshold effect for the spectra's dependence vs. HAP concentration. The obtained results proved high efficiency of IR spectroscopy to study such kind of composite materials but unambiguous identification of every absorption band in the composites’ spectra demands special experiments coupled with thorough calculations.

Repair of large osteochondral defects in rabbits using porous hydroxyapatite/collagen (HAp/Col) and fibroblast growth factor‐2 (FGF‐2)

Articular cartilage has a limited capacity for self-renewal. This article reports the development of a porous hydroxyapatite/collagen (HAp/Col) scaffold as a bone void filler and a vehicle for drug administration. The scaffold consists of HAp nanocrystals and type I atelocollagen. The purpose of this study was to investigate the efficacy of porous HAp/Col impregnated with FGF-2 to repair large osteochondral defects in a rabbit model. Ninety-six cylindrical osteochondral defects 5 mm in diameter and 5 mm in depth were created in the femoral trochlear groove of the right knee. Animals were assigned to one of four treatment groups: porous HAp/Col impregnated with 50 microl of FGF-2 at a concentration of 10 or 100 microg/ml (FGF10 or FGF100 group); porous HAp/Col with 50 microl of PBS (HAp/Col group); and no implantation (defect group). The defect areas were examined grossly and histologically. Subchondral bone regeneration was quantified 3, 6, 12, and 24 weeks after surgery. Abundant bone formation was observed in the HAp/Col implanted groups as compared to the defect group. The FGF10 group displayed not only the most abundant bone regeneration but also the most satisfactory cartilage regeneration, with cartilage presenting a hyaline-like appearance. These findings suggest that porous HAp/Col with FGF-2 augments the cartilage repair process.

Structural analysis of hydroxyapatite coating on magnetite nanoparticles using energy filter imaging and electron tomography

Magnetic nanoparticle (MNP) composites with a magnetite (Fe(3)O(4)) core and a hydroxyapatite (HAp, Ca(10)(PO(4))(6)(OH)(2)) coating were prepared using a precipitation method and a subsequent hydrothermal treatment. The hydrothermal treatment diminished the lepidocrocite layer on the magnetite, enhanced the crystal growth of HAp and dissolved the MNPs. The divalent iron ions dissolved into solvent were not substituted for the HAp lattice. The three-dimensional (3D) nanostructure, the crystal morphology of HAp covered with the MNPs and the interfacial nanostructure of magnetite/HAp were analyzed using an energy-filter transmission electron microscopy (EF-TEM) and visualized by computer tomography in transmission electron microscopy (TEM). EF-TEM and 3D reconstruction images using a tilted series of high-angle annular dark-field images showed that the needlelike HAp nanocrystals covered with a magnetite core and the crystal growth of HAp attached to the magnetite surface was inhibited as a result of the lower density of the nucleation site of the lepidocrocite layer. The dissolution of iron ion from MNPs and the interfacial interaction of HAp and magnetite could cause the needlelike morphology of HAp nanocrystals.

Synthesis of positively charged calcium hydroxyapatite nano-crystals and their adsorption behavior of proteins

Positively charged Hap nano-crystals were prepared by using β-alanine and clarified the adsorption affinity of these surface amide functionalized Hap nano-crystals to proteins. Colloidal surface amide functionalized Hap nano-crystals were prepared by wet method in the presence of various amounts of β-alanine by changing molar ratio of β-alanine/Ca (β/Ca ratio) in the solution. The rod-like nano-crystals were lengthened with addition of β-alanine though their width did not vary; carboxyl groups of β-alanine are strongly coordinated to Ca 2+ ions exposed on ac and/or bc faces to inhibit particle growth to a- and/or b-axis directions and enhance the particle growth along to the c-axis. No difference can be recognized on the crystal structure among the synthesized Hap nano-crystals by XRD measurements. However, the large difference was recognized by TG-DTA and FTIR measurements. Those measurements revealed that β-alanine is incorporated on the Hap nano-crystal surface up to the β/Ca ratio of 1.0, though they are absent in the nano-crystals synthesized at β/Ca ratio ≥ 2.0. The zeta potential (zp) of β-alanine-Hap nano-crystals prepared at β/Ca = 0.4 and 1.0 of those incorporating β-alanine exhibited positive charge at pH ≤ 5.9. The saturated amounts of adsorbed BSA for the positively charged β-alanine-Hap nano-crystals were increased 2.3–2.4-fold by their electrostatic attraction force between positively charged β-alanine-Hap nano-crystals and negatively charged BSA molecules. We were able to control the adsorption affinity of Hap nano-crystal by changing their surface charge.

Fabrication of platy apatite nanocrystals loaded with TiO 2 nanoparticles by two-step emulsion method and their photocatalytic activity

Nanometer-sized TiO 2 island structure on the platy hydroxyapatite nanocrystals (HAp) has been accomplished by two-step emulsion process. At the first step, platy HAp nanocrystals, of which size was in the range of 70–200 nm after heat-treatment at 1078 K for 1 h, were prepared using the W/O emulsion system. Before the second step, HAp nanocrystals were immersed in NaH 2PO 4 solution for the formation of hydroxyl group on their surface. In the following, titanium tetraisopropoxide reacted with the hydroxyl group of HAp surface to form TiO 2 nanoparticles on the surface of HAp nanocrystals, which were dispersed in the micrometer-sized methanol droplets of polyethylene cetylether–cyclohexane mixture (methanol/oil emulsion). The resulting hydroxyapatite nanocrystals loaded with TiO 2 nanoparticles showed the high photocatalytic activity comparing to the commercial TiO 2 catalyst.