Dynamic study of calcium phosphate formation on porous HA/TCP ceramics

Bone-like apatite formation on porous calcium phosphate ceramics was investigated in static simulated body fluid (SBF) and dynamic SBF at different flowing rates. The results of a 14-day immersion in static SBF showed that the formation of bone-like apatite occurred both on the surface and in the pores of the samples. When SBF flowed at the physiological flow rate in muscle (2 ml/100 ml⋅min), bone-like apatite could be detected only in internal surface of the pores of samples. The result that bone-like apatite formation could only be found in the pores when SBF flowed at physiological flow rate was consistent with that of porous calcium phosphate ceramics implanted in vivo: osteoinduction was only detected inside the pores of the porous calcium phosphate ceramics. This result implicates that the bone-like apatite may play an important role in the osteoinduction of Ca-P materials. The dynamic model used in this study may be better than usually used static immersion model in imitating the physiological condition of bone-like apatite formation. Dynamic SBF method is very useful to understand bone-like apatite formation in vivo and the mechanism of ectopic bone formation in calcium phosphate ceramics.     

Dynamic study of calcium phosphate formation on porous HA/TCP ceramics

Bone-like apatite formation on porous calcium phosphate ceramics was investigated in static simulated body fluid (SBF) and dynamic SBF at different flowing rates. The results of a 14-day immersion in static SBF showed that the formation of bone-like apatite occurred both on the surface and in the pores of the samples. When SBF flow at the physiological flow rate in muscle (2 ml/100 ml min1), bone-like apatite could be detected only in internal surface of the pores of samples. The result that bone-like apatite formation could only be found in the pores when SBF flown at physiological flow rate was consistent with that of porous calcium phosphate ceramics implanted in vivo: osteoinduction was only detected inside the pores of the porous calcium phosphate ceramics. This result implicates that the bone-like apatite may play an important role in the osteoinduction of Ca-P materials. The dynamic model used in this study may be better than usually used static immersion model in imitating the physiological condition of bone-like apatite formation. Dynamic SBF method is very useful to understand bone-like apatite formation in vivo and the mechanism of ectopic bone formation in calcium phosphate ceramics.     

In-vitro apatite formation on phosphorylated bamboo

Natural self-reinforced composite, bamboo, was surface modified by phosphorylation with urea–H3PO4 and NaOH–H3PO4 methods; then precalcification was performed by immersing samples in saturated Ca(OH)2 solution. After that, calcium phosphate can be formed on the surface of bamboo samples in calcification media: simulated body fluid (1.5 SBF) and accelerated calcification solution (ACS). Experimental results reveal that pre-calcification is an inevitable step for the formation of calcium phosphate. The calcium phosphate formed in 1.5 SBF was identified by thin-film X-ray diffraction as apatite which was not well crystallized. Compared with the urea–H3PO4 method, the NaOH–H3PO4 method has the advantages of quicker and continuous apatite formation and stronger adhesive between apatite and bamboo.     

Fabrication and biological characteristics of β-tricalcium phosphate porous ceramic scaffolds reinforced with calcium phosphate glass

The fabrication process, compressive strength and biocompatibility of porous β-tricalcium phosphate (β-TCP) ceramic scaffolds reinforced with 45P₂O₅–22CaO–25Na₂O–8MgO bioglass (β-TCP/BG) were investigated for their suitability as bone engineering materials. Porous β-TCP/BG scaffolds with macropore sizes of 200–500 μm were prepared by coating porous polyurethane template with β-TCP/BG slurry. The β-TCP/BG scaffolds showed interconnected porous structures and exhibited enhanced mechanical properties to those pure β-TCP scaffolds. In order to assess the effects of chemical composition of this bioglass on the behavior of osteoblasts cultured in vitro, porous scaffolds were immersed in simulated body fluid (SBF) for 2 weeks, and original specimens (without soaked in SBF) seeded with MC3T3-E1 were cultured for the same period. The ability of inducing apatite crystals in simulated body fluid and the attachment of osteoblasts were examined. Results suggest that apatite agglomerates are formed on the surface of the β-TCP/BG scaffolds and its Ca/P molar ratio is ~1.42. Controlling the crystallization from the β-TCP/BG matrix could influence the releasing speed of inorganic ions and further adjust the microenvironment of the solution around the β-TCP/BG, which could improve the interaction between osteoblasts and the scaffolds.     

Relationship between apatite-forming ability and mechanical properties of bioactive PMMA-based bone cement modified with calcium salts and alkoxysilane

Polymethylmethacrylate (PMMA)-based bone cement is used for the fixation of artificial joints in orthopaedics. However, the fixation is liable to loosen in the body, because the cement does not bond to living bone. So-called bioactive ceramics bond directly to living bone through the apatite layer formed on their surfaces in the body. We previously revealed that modification using γ-methacryloxypropyltrimethoxysilane (MPS) and water-soluble calcium salts such as calcium acetate and calcium hydroxide was effective for providing the PMMA-based bone cement with apatite-forming ability in a simulated body fluid (SBF, Kokubo solution) that closely reproduces the body environment. However, the effect of the chemical reaction forming the apatite on the mechanical properties of the cements has not been clarified. The present work aimed to investigate this issue from the viewpoint of the interface structure between the apatite and the cement. The surface of the cement modified with calcium acetate and MPS was fully covered with newly formed apatite after soaking in Kokubo solution within 7 days, while half of the surface area of the cement modified with calcium hydroxide and MPS was covered with the apatite. The bending strength of the modified cements decreased after soaking in Kokubo solution. Porous structure was observed in the region about 50–100 μm in depth from the top surface because of release of the Ca²⁺ ions by both modified cements after soaking in Kokubo solution. The decrease in bending strength of the modified cements could be attributed to the formation of the pores. In addition, the pores on the top surfaces of the cements were filled with the newly formed apatite. The apatite formation would be effective not only for bioactivity but also for decreasing the reduction of mechanical strength.     

Formation mechanism of biomedical apatite coatings on porous titania layer

A titania containing calcium and phosphate with rough and porous structure was prepared by microarc oxidation. The in vitro bioactivity was examined by immersing the samples into the simulated body fluid (SBF). And the mechanism was also discussed. The results show that only 3 days of immersion in SBF, apatite was formed on the surface, and after 6 days, nearly all the surface covered by apatite. This indicates that the layer can induce the formation of apatite in simulated body fluid. It is analyzed that the key factors of the apatite formation are the hydrolysis of the CaTiO₃ and special structure.     

Apatite wollastonite–poly methyl methacrylate bio-composites

Bio-composites consisting of sol–gel processed apatite wollastonite (AW) glass ceramics and poly methyl methacrylate (PMMA) were prepared by hot compaction method. Density of the composites decreased with increase in PMMA content, while, biaxial flexural strength (BFS) was 21 MPa for 20 wt.% PMMA and beyond which it decreased. A correlation between phase compositions of AW glass ceramics with BFS was attempted from the XRD results. In vitro bioactivity of the composites in a simulated body fluid (SBF) showed the formation of spherical globules on the surface within 7 days of soaking as observed by environmental SEM. Thin film XRD and EDX measurement confirmed these globules to be bone like apatite with Ca/P ratio 1.53 and FTIR measurement showed the corresponding peaks for phosphates. Results indicated the bone bonding ability of the composites by forming a surface apatite (calcium phosphate) layer in SBF and the growth increased with increase in soaking durations. ICP measurement of the remaining SBF after 7, 14 and 21 days soaking of samples was found to be in good agreement with the EDX analysis results.     

Biphasic calcium phosphate coating on cobalt-base surgical alloy during investment casting

The biphasic calcium phosphate (BCP) yields higher bioactivity and efficiency than the Hydroxyapatite (HA) alone. The HA/β-TCP ratio significantly affects BCP bioactivity as well as the extent of BCP resorption. In this study, the BCP coating on ASTM F-75 cobalt base alloy during the investment casting process was investigated. For this purpose, molten metal was poured at 1,470°C into previously coated investment molds preheated to 750, 850, 950, 1,050°C in order to investigate the effect of mold preheating temperatures on coating phase transformations. For in vitro evaluation, samples were immersed in the simulated body fluid (SBF) at 37°C for 4 weeks and characterized by XRD, SEM, EDS, and optical microscopy. The weight percentages of HA and β-TCP of the specimens were calculated to find that the HA/β-TCP ratio significantly depended on the mold preheating temperature as it caused changes in the dissolution behavior of BCP coating and the bone-like apatite precipitation on coating during in vitro evaluation.     

NaOH treatment of vacuum-plasma-sprayed titanium on carbon fibre-reinforced poly(etheretherketone)

Carbon fibre-reinforced polyetheretherketone (CF-PEEK) substrates were coated with titanium by vacuum-plasma-spraying and chemically treated in 10 M sodium hydroxide (NaOH) solution. After NaOH treatment, the specimens were immersed in simulated body fluid (SBF) containing ions in concentrations similar to those of human blood plasma. Scanning electron microscopy, energy-dispersive X-ray analysis and diffuse reflectance Fourier transformed–infrared spectroscopy were used to analyse the NaOH-treated VPS-Ti surface and the calcium phosphate layer formed during immersion in SBF. It was observed that a carbonate-containing calcium phosphate layer was formed on the NaOH-treated VPS-Ti surface during immersion in SBF, whereas no calcium phosphate precipitation occurred on the untreated surfaces. It is therefore concluded that vacuum-plasma-spraying with titanium and subsequent chemical modification in 10 M NaOH solution at 60°C for 2 h is a suitable method for the preparation of bioactive coatings for bone ongrowth on CF-PEEK.     

磷酸钙陶瓷在不同动物的模拟体液中类骨磷灰石的形成研究

根据人、狗、猪、猴和兔五种动物体液的钙离子浓度和pH值的差异,配制了不同组分的模拟体液,将孔壁致密和有微孔的多孔磷酸钙陶瓷分别浸泡在这些模拟体液中,研究陶瓷孔隙表面类骨磷灰石的形成情况.结果表明:在模拟体液中浸泡14天后,孔壁致密的材料未见有类骨磷灰石层形成;有微孔的多孔磷酸钙陶瓷,材料孔壁表面(包括陶瓷表面较深孔隙)有类骨磷灰石层的形成,这与体内植入实验观察到的类骨磷灰石层形成和诱导成骨情况相似,可以推论类骨磷灰石层的形成的确是骨诱导的先决条件.随着钙离子浓度的增加,其孔壁表面类骨磷灰石层的形成也更为均匀,但类骨磷灰石生长快慢顺序与动物组织学观察到的骨诱导性高低的次序不完全一致.     

通过仿生法在硅橡胶表面制备磷灰石薄膜的研究

用CaCl2的乙醇溶液和K2HPO4溶液对硅橡胶进行预处理,将处理过的硅橡胶分别浸渍于模拟体液和钙磷饱和溶液中来制备磷灰石薄膜.利用薄膜X射线衍射、红外吸收光谱和扫描电子显微镜对形成的薄膜进行了表征.结果表明,分别在模拟体液中7d和在钙磷饱和溶液中5d后,硅橡胶表面形成了一层磷灰石薄膜;在模拟体液中的薄膜表面呈网状并分布有许多球状晶粒,在钙磷饱和溶液中的薄膜为结晶良好的片状晶体.     

XPS study of the process of apatite formation on bioactive Ti–6Al–4V alloy in simulated body fluid

Bioactive Ti–6Al–4V alloy, which spontaneously forms a bonelike apatite layer on its surface in the body and bonds to living bone through this apatite layer, can be prepared by producing an amorphous sodium titanate on its surface by NaOH and heat treatments. In this study, the process of apatite formation on the bioactive Ti–6Al–4V alloy was investigated in vitro, by analyzing its surface with X-ray photoelectron spectroscopy as a function of soaking time in a simulated body fluid (SBF). Thin-film X-ray diffractometry of the alloy surface and atomic emission spectroscopy of the fluid were also performed complementarily. It was found that immediately after immersion in the SBF, the alloy exchanged Na⁺ ions from the surface sodium titanate with H₃O⁺ ions in the fluid to form Ti-OH groups on its surface. The Ti-OH groups, immediately after their formation, incorporated the calcium ions in the fluid to form calcium titanate. The calcium titanate thereafter incorporated the phosphate ions in the fluid to form an amorphous calcium phosphate, which was later crystallized into bonelike apatite. This process of apatite formation on the alloy was the same as on the pure titanium metal, because the alloy formed the sodium titanate free of Al and V by the NaOH and heat treatments. The initial formation of the calcium titanate is proposed to be a consequence of the electrostatic interaction of negatively charged units of titania dissociated from the Ti-OH groups with the positively charged calcium ions in the fluid. The calcium titanate is postulated to gain a positive charge and interact with the negatively charged phosphate ions in the fluid to form amorphous calcium phosphate, which eventually stabilizes into crystalline apatite.     

XPS study of the process of apatite formation on bioactive Ti—6Al—4V alloy in simulated body fluid

Bioactive Ti—6Al—4V alloy, which spontaneously forms a bonelike apatite layer on its surface in the body and bonds to living bone through this apatite layer, can be prepared by producing an amorphous sodium titanate on its surface by NaOH and heat treatments. In this study, the process of apatite formation on the bioactive Ti—6Al—4V alloy was investigated in vitro, by analyzing its surface with X-ray photoelectron spectroscopy as a function of soaking time in a simulated body fluid 4SBF). Thin-film X-ray diffractometry of the alloy surface and atomic emission spectroscopy of the fluid were also performed complementarily. It was found that immediately after immersion in the SBF,the alloy exchanged Na¹ ions from the surface sodium titanate with H₃O¹ ions in the fluid to form Ti-OH groups on its surface. The Ti-OH groups, immediately after their formation,incorporated the calcium ions in the fluid to form calcium titanate. The calcium titanate thereafter incorporated the phosphate ions in the fluid to form an amorphous calcium phosphate, which was later crystallized into bonelike apatite. This process of apatite formation on the alloy was the same as on the pure titanium metal, because the alloy formed the sodium titanate free of Al and V by the NaOH and heat treatments. The initial formation of the calcium titanate is proposed to be a consequence of the electrostatic interaction of negatively charged units of titania dissociated from the Ti-OH groups with the positively charged calcium ions in the fluid. The calcium titanate is postulated to gain a positive charge and interact with the negatively charged phosphate ions in the fluid to form amorphous calcium phosphate, which eventually stabilizes into crystalline apatite.     

In–vitro calcium phosphate growth over functionalized cotton fibers

Biomimetic growth of calcium phosphate compound on cotton sheets treated with tetraethoxy silane and soaked in simulated body fluid solution was studied using scanning electron microscopy (SEM), energy dispersive X-ray analysis (EDAX), micro-Fourier transform infrared spectroscopy (FTIR) and X-ray diffractometry (XRD). Micro-FTIR and EDAX results show that silicon was coupled to the cotton fiber when cotton was treated with tetra-ethoxy silane (TEOS) at 125°C for 1 h. Calcium phosphate nucleation started to occur on the surface of TEOS-treated cotton fibers upon immersion in 1.5×SBF (simulated body fluid solution) within 3 days and after 20 days, all the fiber surfaces were found covered with a thick and porous coating of calcium phosphate. The Ca and P determined by inductively coupled plasma spectroscopy (ICP) analysis revealed that the Ca/P ratio as well as the amount of calcium phosphate coating depends on the soaking time in SBF solution. © 1999 Kluwer Academic Publishers     

多孔Nano-dHA/PLA/BCP复合支架的制备及性能研究

为改善常规的多孔聚乳酸/双相钙磷陶瓷(PLA/BCP)支架表面亲水性不佳及降解时呈酸性等不足,采用马弗炉烧结制备的BCP多孔支架浸入纳米缺钙羟基磷灰石/聚乳酸(nano-dHA/PLA)混悬液后,真空干燥得到多孔纳米缺钙羟基磷灰石/聚乳酸/双相钙磷陶瓷(nano-dHA/PLA/BCP)复合支架,利用万能测试机测试支架抗压强度,阿基米德法测定支架孔隙率,扫描电子显微镜(SEM)观察支架表面形貌,并对其保水率和体外降解过程中pH值的变化情况等进行了研究. 结果表明：多孔nano dHA/PLA/BCP复合支架表面粗糙,保水率和强度均有较大提高,在磷酸盐缓冲液（PBS）浸泡过程中pH值下降较慢,在模拟体液（SBF）中浸泡1个月后发现有较多的类骨磷灰石形成.     

In vitro apatite formation on organic–inorganic hybrids in the CaO–SiO<Subscript>2</Subscript>–PO<Subscript>5/2</Subscript>–poly(tetramethylene oxide) system

The osteoconduction potential of artificial materials is usually evaluated in vitro by apatite formation in a simulated body fluid (SBF) proposed by Kokubo and his colleagues. This paper reports the compositional dependence of apatite formation on organic–inorganic hybrids in the CaO–SiO₂–PO_5/2–poly(tetramethylene oxide) system, initiated from tetraethoxysilane (TEOS), triethyl phosphate (OP(OEt)₃), calcium chloride (CaCl₂) and poly(tetramethylene oxide)(PTMO) modified with alkoxysilane. Formation of an apatite layer was observed on the surface of the organic–inorganic hybrids with molar ratios of TEOS/OP(OEt)₃ ranging from 100/0 to 20/80. The rate of apatite formation remarkably decreased when the hybrids were synthesized with TEOS/OP(OEt)₃ ratios of 40/60 or less. Hybrids without TEOS showed no apatite formation in SBF for up to 14 days. Addition of small amounts of OP(OEt)₃ to TEOS in the hybrids led to the high dissolution of calcium and silicate, while addition of large amounts of OP(OEt)₃ decreased the dissolution of calcium and silicate ions and resulted in reduced apatite formation regardless of the dissolution of phosphate ions from the hybrids.     

Ultrastructural evaluation of in vitro mineralized calcium phosphate phase on surface phosphorylated poly(hydroxy ethyl methacrylate-co-methyl methacrylate)

The in vitro functionality of surface phosphorylated poly(hydroxy ethyl methacrylate-co-methyl methacrylate), poly(HEMA-co-MMA) to induce bioinspired mineralization of calcium phosphate phase is evaluated. The primary nucleation of calcium phosphate on the surface phosphorylated copolymer occurs within 3 days of immersion when immersed in 1.5× simulated body fluid and the degree of mineralization is proportional to the hydroxy ethyl methacrylate content in the copolymer. The calcium phosphate phase is identified as hydroxyapatite by X-Ray diffraction analysis. The transmission electron microscopic evaluation combined with selected area diffraction pattern and energy dispersive analysis exemplified that the primary nuclei of amorphous calcium phosphate transforms to crystalline needle like calcium rich apatite, within a period of 3 days immersion in simulated body fluid. The atomic force microscopic results corroborate the c-axis growth of the crystals within 3 days immersion in SBF.     

Hydrolysis of monetite/chitosan composites in α-MEM and SBF solutions

There are two objectives of this work. The first objective is to study the hydrolysis behavior of monetite cements formed in the presence and absence of the chitosan in cell culture media (α-MEM) and simulated body fluid (SBF) solutions at 37°C. During hydrolysis, monetite transformed to carbonated apatite. Therefore, the second objective is to examine how addition of chitosan affects on the formation of carbonated apatite phases. The changes in the phase structure of monetite after hydrolysis reactions were characterized using XRD, FTIR and SEM. Pure monetite and monetite/chitosan composite were soaked in α-MEM and SBF solution for 4 and 7 days. In α-MEM solution, the monetite particles started to transform into carbonated apatite with a slow rate. However, in SBF, the rate of monetite transformation to carbonated apatite was more rapid. The presence of the chitosan had no significant effect on the precipitation of carbonated apatite on the monetite particles.     

Regulation of bone-related genes expression by bone-like apatite in MC3T3-E1 cells

Bone-like apatite on HA/TCP ceramics sintered at 1,100 °C (HT1) and 1,200 °C (HT2) could be obtained via immersing substrates into simulated body fluid (SBF) for 3 days. When MC3T3-E1 preosteoblastic cells cultured on the surface of the bone-like apatite for 3 days, SEM observations revealed cell membrane features with secreted crystals very similar to in vivo bone formation during intramembranous ossification with a direct bone apposition on the ceramics. According to semi-quantitative RT-PCR method, mRNA expressions of osteocalcin (marker of late-stage differentiation) and type 1 collagen were increased in cultures with HT1S and HT2S when compared to HT1 and HT2 after cultured for 6 days. The results indicated that bone-like apatite had the ability to support the growth of osteoblast-like cells in vitro and to promote osteoblast differentiation by stimulating the expression of major phenotypic markers. Taken together, our findings will be helpful in understanding the mechanism of osteoinductivity of calcium phosphate ceramics and in constructing more appropriate biomimetic substrate.     

Bone-like apatite coating on Mg-PSZ/Al2O3 composites using bioactive systems

A biomimetic method was used to promote bioactivity on zirconia/alumina composites. The composites were composed of 80 vol% Mg-PSZ and 20 vol% Al₂O₃. Samples of these bioinert materials were immersed in simulated body fluid (SBF) for 7 days on either a bed of wollastonite ceramics or bioactive glass. After those 7 days, the samples were immersed in a more concentrated solution (1.4 SBF) for 14 days. Experiments were also performed without using a bioactive system during the first stage of immersion. A bone-like apatite layer was formed on the surface of all the materials tested, using wollastonite the bioactive layer was thicker and its morphology was close to that observed on the existing bioactive systems. A thinner apatite layer consisting of small agglomerates was obtained using bioactive glass. The thickness of the ceramic layers was within the range of 15 to 30 μm.