Acta Biomaterialia 7 (2011) 2769–2781

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Acta Biomaterialia

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Review

Nanoscale hydroxyapatite particles for bone tissue engineering

Hongjian Zhou, Jaebeom Lee ⇑Department of Nanomedical Engineering, College of Nanoscience and Nanotechnology, Pusan National University, Miryang 627-706, Republic of Korea

a r t i c l e i n f o a b s t r a c t

Article history:Received 7 December 2010Received in revised form 11 March 2011Accepted 16 March 2011Available online 1 April 2011

Keywords:BiomaterialsBioceramicsNanoscale particlesHydroxyapatiteBone tissue engineering

1742-7061/$ - see front matter � 2011 Acta Materialdoi:10.1016/j.actbio.2011.03.019

⇑ Corresponding author.E-mail address: jaebeom@pusan.ac.kr (J. Lee).

Hydroxyapatite (HAp) exhibits excellent biocompatibility with soft tissues such as skin, muscle andgums, making it an ideal candidate for orthopedic and dental implants or components of implants. Syn-thetic HAp has been widely used in repair of hard tissues, and common uses include bone repair, boneaugmentation, as well as coating of implants or acting as fillers in bone or teeth. However, the lowmechanical strength of normal HAp ceramics generally restricts its use to low load-bearing applications.Recent advancements in nanoscience and nanotechnology have reignited investigation of nanoscale HApformation in order to clearly define the small-scale properties of HAp. It has been suggested that nano-HAp may be an ideal biomaterial due to its good biocompatibility and bone integration ability. HAp bio-medical material development has benefited significantly from advancements in nanotechnology. Thisfeature article looks afresh at nano-HAp particles, highlighting the importance of size, crystal morphologycontrol, and composites with other inorganic particles for biomedical material development.

� 2011 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

1. Introduction

Bone is a natural organic–inorganic ceramic composite consist-ing of collagen fibrils containing embedded, well-arrayed, nano-crystalline, rod-like inorganic materials 25–50 nm in length [1–3]. Structural order in bone occurs at several hierarchical levelsand reflects the materials and mechanical properties of its compo-nents (Fig. 1). Hydroxyapatite (HAp) is chemically similar to theinorganic component of bone matrix – a very complex tissue withgeneral formula Ca10(OH)2(PO4)6. The close chemical similarity ofHAp to natural bone has led to extensive research efforts to usesynthetic HAp as a bone substitute and/or replacement in biomed-ical applications [4,5].

Tissue engineering is intensively researching solutions thathave the potential to reduce the complications related to currenttreatment methods. Tissue engineering can be defined as an inter-disciplinary field that applies the principles of engineering and lifesciences to develop biological substitutes that restore, maintain orimprove tissue function [6]. This concept involves three main strat-egies: the use of isolated cells or cell substitutes to replace limitedfunctions of the tissue; utilization of tissue-inducing substancessuch as growth factors; and scaffolds to direct tissue development.An ideal scaffold for bone tissue engineering is a matrix that acts asa temporary substrate allowing cell growth and tissue develop-ment. This occurs initially in vitro and eventually in vivo. The scaf-fold should be able to mimic the structure and biological functionof the native extracellular matrix (ECM) in terms of both chemical

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composition and physical structure. Scaffolds used for tissue engi-neering applications should also be biocompatible; able to provideappropriate mechanical support; exhibit favorable surface proper-ties such as promoting adhesion, proliferation and differentiationof cells; and provide an environment in which cells can maintaintheir phenotypes.

Recently, HAp has been used for a variety of biomedical applica-tions, including matrices for drug release control and bone tissueengineering materials [8,9]. Since HAp has chemical similarity tothe inorganic component of bone matrix, synthetic HAp exhibitsstrong affinity to host hard tissues. Chemical bonding with the hosttissue offers HAp a greater advantage in clinical applications com-pared to most other bone substitutes such as allografts or metallicimplants [10]. The main advantages of synthetic HAp are its biocom-patibility, slow biodegradability in situ, and good osteoconductiveand osteoinductive capabilities [1,11]. A study by Taniguchi et al.showed that sintered HAp exhibits excellent biocompatibility withsoft tissues such as skin, muscle and gums. Such capabilities havemade HAp an ideal candidate for orthopedic and dental implantsor components of implants. Synthetic HAp has been widely usedto repair hard tissues. Common uses include bone repair, bone aug-mentation, as well as coating of implants or acting as fillers in boneor teeth [12–18]. However, the low mechanical strength of normalHAp ceramics restricts its use mainly to low load-bearing applica-tions. Recent advances in nanoscience and nanotechnology havereignited interest in the formation of nanosized HAp and the studyof its properties on the nanoscale.

Nanocrystalline HAp powders exhibit improved sinterabilityand enhanced densification due to greater surface area, whichmay improve fracture toughness, as well as other mechanical

ll rights reserved.

Fig. 1. The hierarchical structure of bone at its various length scales. The microstructure of cortical bone consists of osteons with Haversian canals and lamellae, and at thenanoscale, the structural units are collagen fibers composed of bundles of mineralized collagen fibrils. Copyright Elsevier and reproduced with permission [7].

2770 H. Zhou, J. Lee / Acta Biomaterialia 7 (2011) 2769–2781

properties [11]. Moreover, nano-HAp, compared to coarser crystals,is expected to have better bioactivity [19]. Thus, nano-HAp parti-cles can be utilized for engineered tissue implants with improvedbiocompatibility over other implants. Nanotechnology has the po-tential to significantly benefit development of HAp biomedicalmaterials. To our knowledge, several reviews of nanocrystallinecalcium orthophosphates have been published in recent years.For example, Dorozhkin et al. [20,21] reviewed the current stateof technology and recent developments of various nanosized andnanocrystalline calcium orthophosphates, involved in synthesisand characterization as well as biomedical and clinical applica-tions. Moseke et al. [22] reviewed the synthesis and properties oftetracalcium phosphate (TTCP) in biomaterial applications suchas cements, sintered ceramics and coatings on implant metals;Johnson et al. [18] reviewed the compression, flexural and tensileproperties of calcium phosphate (CaP) and CaP–polymer compos-ites for applications in bone replacement and repair; Tran et al.[23] summarized studies that have demonstrated enhancedin vitro and in vivo osteoblast functions (e.g. adhesion, prolifera-tion, synthesis of bone-related proteins and deposition of cal-cium-containing mineral) on nanostructured metals, ceramics,polymers, and composites. After reviewing these feature articlesto avoid any redundancy, we focus on calcium orthophosphate,and characterize its properties in the condition of nano-HAp withdifferent morphologies and porous structures-materials that offergreat promise as bone substitutes and/or replacements in biomed-ical applications. Moreover, we summarize how composites of HApand other inorganic nanomaterials can enhance the bioactivity andbiocompatibility of HAp – an area that has become the focus of re-cent research. The remainder of this feature article is organizedinto five sections. In the Section 2, the synthesis of morphologicallydifferent nano-HAps is introduced. Section 3 discusses the fabrica-tion of the porous structure of nano-HAp. Section 4 reviews thebio-orthopedic properties of nanoscale HAp for application in bonetissue engineering. Section 5 introduces composites of HAp andother inorganic nanomaterials for enhancing the bioactivity andbiocompatibility of HAp. Finally, in Section 6, we provide a sum-mary and our own perspectives on this active area of research.

2. Synthesis of nanoscale HAp

HAp (Ca10(OH)2(PO4)6) nano- and microcrystals with multiformmorphologies (separated nanowires, nanorods, microspheres,

microflowers and microsheets) have been successfully synthesizedby many powder processing techniques, including sol–gel synthe-sis [24–28], solid state reactions [29], co-precipitation [30], hydro-thermal reactions [31], microemulsion syntheses [32] andmechanochemical synthesis [33].

2.1. HAp nanoparticles

From a practical application perspective, suitable nano- ormicromaterials with specific morphologies not only need to becapable of being synthesized in large quantities with a desiredcomposition, reproducible size and structure, but also of beingprepared and assembled using environmentally responsible tech-niques. Recently, environmentally friendly synthetic methodolo-gies, including molten-salt synthesis, hydrothermal processing,biomimic synthesis and template synthesis, have been imple-mented as viable techniques for the synthesis of a range of materi-als [34,35]. Nanosized HAp particles can be prepared by a variety oftechniques such as mechanochemical synthesis [36], combustionpreparation [37] and various wet chemistry techniques [38,39].Among the most reported precipitation processes, chemical agentssuch as citric acid [40,41], amino acids [42] and ethylenediamine-tetraacetic acid (EDTA) [43,44] have been used to mediate HApnucleation and crystal growth. These modifiers exert significantcontrol over crystal morphology due to affinity between the mod-ifying agent and the HAp crystals. However, there has been less fo-cus on the precipitation kinetics of nucleation and growth, whichare related to the degree of supersaturation, SHAp. This value canbe calculated as follows:

S ¼ IAPHAp

Ksp; ð1Þ

where IAPHAp is the ionic activity product expressed as:

IAPHAp ¼ ½Ca2þ�5½PO3�4 �

3½OH��c5Ca2þc3

PO3�4cOH� : ð2Þ

The brackets represent ion concentrations of the respective spe-cies and c values are the activity coefficients of the ions. Ksp is thesolubility product of HAp. With a higher degree of supersaturation,a greater driving force for precipitation, i.e. a faster precipitationrate, was expected with increasing IAPHAp [45].

Biological mineralization (or biomineralization) is the processof in vivo inorganic material formation. The new theory of ‘‘aggre-

H. Zhou, J. Lee / Acta Biomaterialia 7 (2011) 2769–2781 2771

gation-based crystal growth’’ [46] and the new concept of the‘‘mesocrystal’’ [47] highlight the roles of nanoparticles in biologicalcrystal engineering. Mimicking the formation of natural CaP, hardtissues contribute significantly to the biological function of engi-neered materials. Many advances have been made in biomaterialresearch with the rapid growth of nanotechnology. The study ofCaPs is a specific area in nanotechnology, which may be appliedreadily in the repair of hard and soft skeletal tissues [48–50]. Moll-azadeh et al. [51] prepared HAp crystals using an in situ biomi-metic process in the presence of polyvinyl alcohol (PVA). Theysystematically investigated the effects of polymer amount andmolecular weight on the physical properties of HAp crystals. Theresults indicated that the development (size and shape) of HApnanocrystals precipitated in an aqueous solution of PVA was inver-sely related to the polymer molecular weight (i.e. the smallestcrystallite size was observed with the highest PVA molecularweight). It is thought that HAp formation is initiated through theinteraction of Ca2+ ions with the negative side groups on the poly-mer surface. The larger number of reaction sites in the highermolecular weight PVA polymer led to a higher number of HAp nu-clei, and therefore a smaller crystallite size.

The hydrothermal method, a typical solution-based approach,has proven to be an effective and convenient process to preparevarious inorganic materials with diverse, controllable morpholo-gies and architectures [52–55]. The environmentally acceptableadvantages of this method include easily controllable reaction con-ditions, relatively large scale and high yield in terms of quantity ofthe desired products, and frequent use of water as the reactionmedium. Zhang et al. [56] developed a general strategy for the syn-thesis of HAp nano- and microstructures using water as a reactionmedium through a simple hydrothermal process. Several dominantmorphologies were achieved (nanorods, nanowires, microsheets,burr-like microspheres and microflowers). The pH value plays acrucial role in obtaining Ca10(OH)2(PO4)6 samples with variousmorphologies. The use of trisodium citrate also has an important

Fig. 2. Schematic for the formation and morphology evolution mechanism of Ca5(PO

influence on product shape. Possible formation mechanisms forCa10(OH)2(PO4)6 nano- and microcrystals with diverse morphologyare presented in Fig. 2. A strong blue emission peak at approxi-mately 428 nm was observed at room temperature, and the photo-luminescence (PL) intensity of this emission varied withCa10(OH)2(PO4)6 morphology over a range of pH values. CO��2 radi-cals in the HAp lattice interstitials may be responsible for self-acti-vated luminescence. These types of phosphors do not containmetal ions as activators and contain no toxic elements; thus, theyare considered environmentally friendly luminescent materials.The microsized Ca10(OH)2(PO4)6 sample prepared at pH 5.0 withstrong blue emission, spherical morphology, non-aggregation andhigh crystallinity can potentially be used as a new, efficient andenvironmentally friendly blue luminescent material.

Qiu et al. [57] prepared nanocrystalline HAp by precipitationwith the aid of ultrasonic atomization using Ca(NO3)2�4H2O and(NH4)2HPO4 as raw materials. The results showed that the synthe-sis method used in this study can effectively shorten reaction timewhile improving powder homogeneity compared to other pub-lished methods. It was also found that addition of a small amountof the surfactant glycine during precipitation synthesis can reduceHAp nanoparticle agglomeration. However, Poinern et al. [1] andLeGeros [11] developed a chemical route to synthesize HAp usingcalcium nitrate and potassium hydrogen phosphate as the mainraw materials. This hydrothermal method also used ultrasonic irra-diation followed by heat treatment to manufacture nanosized HAp.This study has shown that nano-HAp particles with a sphericalmorphology in the nanometer range (approximately 30 nm ± 5%)can be synthesized using a hydrothermal chemical precipitationmethod incorporating low-power sonic irradiation. The crystallin-ity and morphology of HAp depends on the Ca/P ratio, ultrasonicirradiation and temperature. An ultrasonic power of 50 W andtemperatures in the range of 400 �C are sufficient to produce thenano-HAp. This offer an economic route for the synthesis ofnano-HAp with a strong scale-up capability.

4)3OH samples with various morphologies based upon different pH values [56].

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Li et al. [58] studied the crystalline behaviors of HAp influencedby different forms of citrates and explained this behavior using theclassical theory of crystal nucleation and growth in solutions. Theyfound that the supersaturation value decreased, i.e. the speed ofcrystal nucleation decreased, with increasing citric acid monohy-drate (C6H8O7�H2O, CiA) concentration or with decreasing triso-dium citrate (Na3C6H5O7�2H2O, NaCit) concentration. Thepresence of CiA inhibited the fast-growing of (0 0 l) planes and in-creased the crystallinity of (h k 0) planes. This resulted in homoge-neously grown particles rather than the prismatic particlesobtained when synthesis is performed without CiA. By raisingthe synthesis temperature, finer and more homogenously grownparticles could be prepared in the presence of CiA. With increasedripening time, a higher crystallinity of HAp with lower amounts ofCaO phase was obtained. This could be explained by crystal growthin solutions, i.e. prolonged maturation time resulting in a higheramount of unreacted Ca(OH)2 that can be involved in the crystalgrowth of CaP powders.

Utilizing nanotechnology, calcium and phosphate can bemanipulated at the molecular level and assembled to producematerials with unique structural and functional properties. Thepreparation of CaP powders with a particular morphology, stoichi-ometry, crystallinity and crystal size distribution is important inbiomedicine and materials science. Various processes have beenemployed to prepare CaP powders, including co-precipitation[59], sol–gel process [60,61], spray pyrolysis [62], hydrothermalsynthesis [63,64], emulsion processing [65,66] and mechanochem-ical method [67]. Among these methods, the sol–gel method hasreceived the most attention because of its well-known inherentadvantages, namely homogeneous molecular mixing, low process-ing temperature, and ability to generate nanocrystalline powders,bulk amorphous monolithic solids and thin films [68]. Natarajanet al. [69] prepared nanosized HAp particles using the sol–gelmethod from the water-based solution of calcium and phosphorusprecursors. In that study, two calcium precursors, i.e. calcium ni-trate tetrahydrate and calcium acetate, were chosen. The influenceof aging period, pH, viscosity and sintering temperature on thecrystallinity and morphology of HAp particles were investigatedfor the two calcium precursors and a triethyl phosphate precursor.The morphology of nano-HAp when the phosphorous precursorwas used was dependent on the type of calcium precursor used.Gopi et al. [70] reported the synthesis and characterization ofnano-HAp powders by a novel ultrasonic coupled sol–gel synthe-sis. The resulting powders were sintered by conventional meansat different temperatures. These results show that nano-HAp pow-ders synthesized by ultrasonic coupled sol–gel synthesis showed aremarkable particle size reduction compared with the conven-tional sol–gel method, hence; these powders could be used as acoating material in biomedical applications.

With the increasing need to develop clean, non-toxic and envi-ronmentally friendly techniques, HAp powders have been ex-tracted using bioproducts such as corals, cuttlefish shells, naturalgypsum, natural calcite and bovine bone [71,72]. Chemical analysishas shown that these products, typically considered as biowaste,are rich sources of calcium in the form of carbonates and oxide.One such biowaste is chicken eggshells. Every day, a million tonnesof eggshells are generated as biowaste around the globe. Eggshellrepresents �11% of the total egg weight and is primarily composedof calcium carbonate (�94%), calcium phosphate (�1%) and organicmatter (�4%) [37]. Besides being economically cheap and plentifulin nature, eggshells have been shown to be biocompatible with thehuman body during implantation but not osteoconductive. Hence,converting these powders into HAp prior to implantation is advan-tageous. Sanosh et al. [73] reported a simple sol–gel precipitationtechnique to synthesize nano-HAp powders using CaO derivedfrom chicken eggshells. Their study shows that biowaste egg shells

can effectively be utilized for synthesizing pure nano-HAppowders.

Particle formation by sol–gel is a very complex process. It in-volves nucleation, growth, aggregation and agglomeration [74,75].Lee et al. [76] also synthesized highly sinterable, nanosized HAppowders using a wet chemical route with recycled eggshell andphosphoric acid as calcium and phosphorous sources, respectively.Raw eggshell was easily converted to CaO by the calcining process,and phosphoric acid was mixed with the calcined eggshell by a wetball-milling method. The observed phases of the powder synthesisprocess were dependent on the mixing ratio (wt.%) of the calcinedeggshell to phosphoric acid and the heating temperature. Theball-milled, nanosized HAp powder, which has an average particlesize of 70 nm, is fully densified at 1300 �C for 1 h. The Ca/P ratiofor the stoichiometric composition of HAp was controlled by adjust-ing the mixing ratio. In addition, HAp can be successfully producedfrom recycled eggshells, seashells and phosphoric acid. The phasesobtained depend on the ratio of calcined eggshells/seashells tophosphoric acid, the calcination temperature and the mechano-chemical activation method (ball milling or attrition milling) [77].Mostaghaci et al. [78] synthesized nanosized HAp using an Iranianstrain of Serratia. Results showed that the optimum powder pro-duction was achieved at approximately pH 8 and a temperatureof 37 �C. The powder particles were single crystals and ranged insize from 25 to 30 nm. Moreover, particle shapes and the sizes wererelatively uniform and exhibited lower agglomeration relative toconventional methods. This powder could be used in the regenera-tion of bone defects, fabrication of medical implants, and as a vectorfor pharmaceutical and biological materials such as genes.

2.2. HAp nanorods and nanoflakes

Bone is a composite, consisting of HAp nanorods embedded in acollagen matrix. In the biomineralization process of vertebratehard tissues, some specific molecules control the nucleation andgrowth of inorganic crystals (HAp), resulting in the formation ofhierarchical structure of teeth and bones with superior mechanicalproperties [79,80]. Thus, controlled syntheses of apatitic crystalswith various morphologies have been the focus of intensive re-search in order to understand more completely their biominerali-zation and utility in industrial and biomedical applications [81].Inspired by biomineralization, molecular manipulation of inor-ganic crystals with organic growth modifiers gradually developedinto a powerful tool for the design of novel tissue engineeringmaterials [82,83].

The molecule–template combination exerts significant controlover crystal morphology and has been discussed in many articles[84–87]. A variety of organic molecules such as hexadecyltrimeth-yl-ammonium bromide (CTAB), sodium dodecyl sulfate (SDS), ami-no acids, proteins and monosaccharides have been used tosynthesize HAp with fibrous- and flake-like morphologies [86,88–92]. Kalita et al. [93] synthesized bioactive HAp (Ca10(PO4)6(OH)2)ceramic powder in the lower end of the nanoregime using micro-wave radiation, which offers several advantages. The appliedmicrowave power of 600 W, pH of the suspension, mole ratio ofCa/P in the starting chemicals, and the chelating effect of EDTAserved as important factors in the synthesis of nanocrystallineHAp powder. Results confirmed a highly crystalline nanopowder(5–30 nm) with elemental composition of Ca and P in the HApphase and possessing mixed (elliptical and rod-shaped) morphol-ogy. Tari et al. [94] reported that using poly(sodium 4-styrene sul-fonate) (PSSS) as a nucleation- and growth-controlling agentresulted in the precipitation of well-crystallized HAp nanoparticlesthrough microemulsion. During PSSS mixing with a calcium precur-sor, rod-like micelles were formed, which control the morphologyand crystallization of nano-HAp. The investigations showed that

H. Zhou, J. Lee / Acta Biomaterialia 7 (2011) 2769–2781 2773

the obtained HAp nanorods have an average width and length ofabout 30 and 200 nm, respectively. Shanthi et al. [95] reported thatCTAB is used as the capturing material for successfully preparednano-HAp rods at ambient temperature and normal atmosphericpressure. A cytotoxicity test using a normal cell line indicated thebiocompatible range of the nanocrystalline HAp and its fairlynon-toxic nature.

Calcium phosphates comprise a large family of compounds withimportant biological applications-including osteologic implant forcoatings, grafts, scaffolds and bone cavity fillings, and vehicles fordrug, protein and gene delivery-due to their similarity with themineral constituents of human bones and teeth. The propertiesof calcium phosphate, including bioactivity, biocompatibility, solu-bility, mechanical properties and absorption, can be tailored overwide ranges by controlling particle composition, size, morphologyand assembly. For these reasons, it is of great importance to devel-op synthesis methods focused on the precise control of particlesize, morphology and chemical composition. Jiang et al. [96] pre-sented an approach using CaP that offered careful size and struc-tural control via a template-guided process. In their study, HApnanorods and well-aligned hybrid HAp composites were preparedvia a template-guided synthesis procedure (a schematic illustra-tion of the formation of HAp nanorods is shown in Fig. 3). First,phosphate ester was used as a structure-directing agent for prepar-ing HAp nanorods. After hydrothermal treatment, CaP nanorodswith well-controlled particle size and porosity can be obtained.Second, carboxymethyl cellulose (CMC) molecules were found tobe effective in controlling particle size and the subsequent align-ment of HAp. HAp composites with well-organized microstruc-tures were prepared using CMC. CMC has carboxyl groups thatcan attract Ca2+ ions, thereby guiding the growth of HAp grains.These proposed processes can also be applied to the preparationof CaP materials with well-controlled microstructures using othersimilar templates for a wide range of applications.

HAp is the basic component of natural bone. In the bone forma-tion process, HAp mineralization is controlled by collagen, which isa special protein containing an ionic group that can interact withHAp, and a dispersive group, which can stabilize HAp in the phys-iological environment. However, dental structures are also com-posed of HAp, which is needle-like and forms under the controlof proteins. Nearly the entire biological mineralization process isa crystallization process controlled by organic components.

Fig. 3. Schematic illustration of the formation of HAp nanorods

Recently, considerable attention has been paid to the biomimeticmineralization process because this process may lead to the fabri-cation of novel materials that cannot be produced by conventionalmethods [97–101]. Zhang et al. [102] reported the mineralizationof nano-HAp on self-assembled collagen; the nanocompositeformed has a similar structure and biocompatibility to naturalbone. Tang et al. [103] reported the fabrication of nano-HAp underthe control of amino acids and found that both bone mimetic plate-like and dental-mimetic rod-like nano-HAp can be produced underthe control of different amino acids. This method makes it possibleto control nano-HAp mineralization by purely synthetic polymersrather than natural biomacromolecules. Since the synthetic poly-mer has a relatively low cost and can be more easily designed,nano-HAp synthesized by this method has many potentialapplications. Yao et al. [104] employed double-hydrophilic blockcopolymer (DHBC) poly-(vinylpyrrolidone)-b-poly(vinylpyrroli-done-alt-maleicanhydride)-b-poly-(vinylpyrrolidone) (PVP-b-P(NVP-alt-MAn)-b-PVP) to synthesize a biomimetic template forHAp nanocrystal synthesis. Needle-like HAp nanocrystals can beformed in the presence of PVP108–P(NVP-MAn)28–PVP108, as shownin Fig. 4. Compared to the HAp nanocrystals formed in the presenceof poly-(vinylpyrrolidone) (PVP) homopolymer, those formed withDHBC are more stable and do not precipitate in water after prepa-ration. The crystallization process and the morphology of the finalnano-HAp crystals can be controlled by adjusting the DHBC molec-ular structure. Cho et al. [105] prepared nanosized HAp powderswith high crystallinity and appropriate stoichiometry by a high-temperature flame spray pyrolysis process from spray solutionscontaining PEG. The mean sizes of HAp powders obtained fromPEG spray solutions were changed from several tens to severalhundred of nanometers according to the PEG concentrations inthe spray solutions at a post-treatment temperature of 800 �C.HAp powders post-treated at a low temperature of 400 �C had a fi-ber-like morphology. On the other hand, post-treated HAp pow-ders at temperatures of 600 and 1000 �C had a rod-likemorphology with a low aspect ratio and spherical-like morphology,respectively. The mean sizes of HAp powders post-treated at tem-peratures of 600 and 1000 �C were 32 and 213 nm, respectively.

The application of nanostructured HAp with different morpholo-gies for bone tissue engineering has been introduced above. How-ever, several other applications of nano-HAp are also in progress.Surface modifications of HAp nanoparticles have been performed

. Copyright Elsevier and reproduced with permission [97].

Fig. 4. TEM images of HAp nanocrystals templated by PVP108–P(NVP-MAn)28–PVP108 after the 1 day (a), 8 day (b) and 13 day (c) preparations. Copyright ACS and reproducedwith permission [104].

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in order to modulate their colloid stability, prevent dissolution in thecase of low pH, prevent inflammation, serve as an intermediate layerto allow strong bond formation between HAp–polymer matrices,and potentially enhance its bioactivity or improve its conjugationability with special functional groups [106–108]. HAp nanoparticleshave also served as non-viral carriers for drug delivery and genetherapy because of their established biocompatibility, ease of han-dling and well-known adsorption affinity [109–112]. Furthermore,HAp nanoparticles can be stably loaded with radioisotopes [111].After loading with genes or drugs by adsorption, HAp nanoparticlesprovide a protective environment that shields them from degrada-tion while providing a convenient pathway for cell membrane pen-etration and the controlled release of the genes/drugs [112]. Theresearch results indicate the potential of nano-HAp in gene deliveryand as drug carriers [112,113]. Readers who are interested in learn-ing more may refer to several elegant and more comprehensive re-views, e.g. those by Dorozhkin [20,21,114].

3. Porous structure of nanoscale HAp

HAp ceramics have been widely used as artificial bone substi-tutes because of their high biocompatibility, bioaffinity and osteo-conductibility. However, induction of bone growth into HAp blocksis unsatisfactory, because it is very slowly replaced by host boneafter implantation. For this reason, porous bodies and granules ofHAp ceramics have been developed and have been widely usedin clinical settings. However, due to the closed structure of conven-tional porous HAp, which has non-uniform pore geometry and lowinterpore connections, it is very difficult for implant pores to be-come completely filled with newly formed host bone [115]. How-ever, porous HAp ceramics with highly interconnecting structureshave been developed, and osteoconduction can occur deep insidesuch ceramics. Wang et al. [116] systematically studied the effectsof electrical polarization of porous HAp ceramics using two typesof cylindrical porous HAp ceramics with high and low interporeconnections (HAp-H and HAp-L, respectively) on bone ingrowth.Electrical polarization was effective in enhancing bone ingrowththrough all the pores of HAp-H implants; however, this advantagewas not apparent in the HAp-L implants. This suggested that en-hanced bone ingrowth into HAp porous bodies due to electricalpolarization may be a co-operative interaction between the osteo-conductivity of HAp porous bodies and the enhanced osteogeniccell activity induced by large charges stored on pore surfaces.

The size and density of interpore connections, as well as thoseof pores, are important factors for osteoconduction into the centralarea of porous HAp. Diverse characteristics, such as pore size, poreshape, pore interconnectivity and total porosity of the scaffold, areconsidered important factors for successful tissue regeneration[117,118]. Microwave heating has also been applied to fabricateinterconnective porous structured bodies by foaming as-synthe-

sized, calcium-deficient HAp (Ca-deficient HAp) precipitate con-taining H2O2. The porous bodies were sintered by a microwaveprocess with activated carbon as the embedding material to pre-pare nano- and submicron-structured ceramics. The study resultssuggest that porous carbonated biphasic CaP ceramics with a nano-structure promote osteoblast adhesion, proliferation and differen-tiation [119]. In conclusion, porous carbonated biphasic calciumphosphate (BCP) ceramics with a nanostructure are simple andquick to prepare using microwaves. Compared to those producedby conventional sintering, carbonated BCP ceramics may be betterbone graft materials.

Since synthesized HAp is very brittle, it cannot be used for load-bearing bone replacements. Hence, implant materials composed ofhard and soft phases (composite materials) are used for total bonereplacement [120]. Composites of HAp with polymers such as poly-methyl methacrylate, poly(3-hydroxybutyrate-co-3-hydroxyval-eate) and polyacrylic acid show improved mechanical properties,as well as good biocompatibility and bioactivity [121,122]. Degrad-able and non-degradable polymers are used in controlled drugdelivery, scaffolds for tissue engineering, wound dressing, cosmeticskin masks and protective clothing. Imai, Furuichi and co-workers[122,123] reported hierarchical laminated architecture and porousstructures after calcinations of the polymer composite (poly(-acrylic acid)) at 700 �C. Polyacrylamide hydrogel is a biomaterialand non-degradable water-based polymer used as tissue filler.Joshy et al. [124] investigated mineralization of HAp in a UV-poly-merized acrylamide gel matrix by varying precursor concentrationand pH (pH 8–10). During polymerization, diammonium hydrogenphosphate ions were implanted in the gel matrix and subsequentlyimmersed in calcium nitrate solution. Thin, laminated, macropo-rous structures, embedded with nanospheres and ribbons of HApwere mineralized. The HAp was found to be oriented along thec-axis, which could lead to preferential binding of acidic proteinson its surface. The laminated structures displayed a resorbable nat-ure, whereas flake-like structures obtained at higher concentra-tions were found to be bioactive. This composite could be analternative to the use of silicone gel to avoid the long-term riskof fibrosis and migration when implanted. Muthutantri et al.[125] developed a novel fabrication technique, a combination ofslurry dipping and electrospraying, to produce HAp foams as po-tential matrices for bone tissue engineering applications. Slurry-dipped and electrosprayed scaffolds for different time intervalswere compared with foams prepared by the individual methodsof dipping and electrospraying. Significant differences in strutcrack distribution and strut thickness and porosity were observedon sintered foams prepared under various conditions. All sinteredstructures had average porosities in the range of 84–94% and desir-able pore interconnections, whereas the combined method pro-duced foams of uniform pore distribution, thicker struts andimproved mechanical properties.

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A new direct rapid prototyping process called low-temperaturedeposition manufacturing (LDM) was proposed to fabricate scaf-folds [126]. This process integrated extrusion/jetting and phaseseparation and can therefore fabricate scaffolds with hierarchicalporous structures, creating an ideal environment for new tissuegrowth. Scanning electron microscopy (SEM) images of fabricatedscaffold structures with different polymer matrices are shown inFig. 5. The interconnected computer-designed macropores allowcells in new tissues to grow throughout the scaffold. Moreover,the parameter-controlled micropores allow nutritional compo-nents in, and metabolic wastes out. The macrocellular morphology,microcellular morphology, porosity and mechanical properties ofpoly(a-hydroxy acid)-tricalcium phosphate (TCP) composite scaf-folds prepared by the proposed method were investigated. Thesehighly controllable scaffolds may play an important role in tissueengineering. LDM could also be combined with multinozzle depo-sition or cell deposition to accurately control materials or cellspoint-by-point. Sinha et al. [127] report a novel method of produc-ing HAp–PVA microspheres suitable for biomedical application.Spray drying is a well-established industrial process that producesfine ceramic powders. Integrating this with a method akin to bio-mineralization provides a direct route to produce HAp micro-spheres with highly controlled morphological features.

4. Bio-orthopedic properties of nanoscale HAp

Bone substitutes are required to repair segmental defectscaused by the removal of infected tissue or bone tumors. The mostdesirable form of bone substitutes, in such cases, is autologousbone. However, autografts are not always available and may resultin morbidity at the donor site. An allograft is preferred in somecases, but the possible immune response and disease (i.e. humanimmunodeficiency virus (HIV) or hepatitis B) transmission are det-rimental to the recipient [128]. Bone graft substitutes have at-tracted much attention because of their advantages over bothautografts and allografts [129]. In the case of a bone, an optimizedbiomaterial should be as biomimetic as possible, i.e. it should con-sist of poorly crystalline, carbonate-substituted apatite with suffi-cient mechanical properties [130]. HAp artificial bones are nowwidely used in clinical practice and show satisfying repair functionin a series of studies [131–133]. Nonetheless, there are some weak-nesses, such as weak intensity and slow degradation. To strengthenHAp, researchers have carried out omnidirectional investigationsand the challenge in HAp development is whether biological prop-erties can match mechanical properties [134].

Recently, our group reported HAp coating on scratched areas ofa human tooth and HAp disks by the immersion method in a HApcolloidal solution (620 lm average diameter dispersed in deion-ized water) [135]. The surface morphologies of the scratched area

Fig. 5. SEM micrographs of scaffolds fabricated with different polyme

after immersion for 1–3 months were investigated and showedthat the damaged surfaces were remarkably recovered. Themechanical property and chemical stability of the HAp coating lay-ers on both specimens were then determined via the Vickers hard-ness test and concentration measurement of extracted Ca2+ ions,respectively, after strong acidic treatment. The cellular behaviorof mouse calvaria-derived pre-osteoblastic cells (MC3T3-E1) wasalso examined on the HAp layers regenerated on micro-scratchedHAp disks for the purpose of their potential applications on maxil-lofacial bone conservation and reconstruction for prosthetic den-tistry, and artificial disk preparation of a vertebral column. Theseresults of HAp coating on the scratched areas of the tooth suggestthat this technique could be suitable for the development of long-term prevention of micro-cleavage and tooth health supporters toreduce discoloration, and for other maxillofacial and orthopedicapplications. Another interesting result published by our groupconcerned the regeneration of a micro-scratched tooth enamellayer by nanoscale HAp solution [136]. Nanoscale HAp powderswith a mean particle size of 200 nm were used to regenerate theenamel layers of damaged teeth. An artificially scratched toothwas immersed in a nanoscale HAp powder suspension in deionizedwater (70 wt.% HAp) at 37 �C for a period of 1–3 months. After3 months, the scratched surface was finally inlaid with HAp andthe roughness increased from 2.80 to 5.51. Moreover, the hardnessof the newly generated HAp layer on the crown was similar to thatof the innate layer. Ca2+ and PO3�

4 ions from the HAp powders dis-solved in deionized water were precipitated on the tooth to pro-duce cement pastes on the enamel surface due to its high degreeof recrystallization, resulting in a hard, newly regenerated HAplayer on the enamel layer. This nanoscale HAp powder solutionmight be used to heal decayed teeth as well as to develop tooth-whitening materials.

In addition, our group reported the clinical evaluation of SB-1™as a synthetic bone in sinus bone grafting. The SB-1™ used consistsof synthetic HAp with 500–1400 lm particle size and has the samechemical composition as the inorganic part of human bones. Rab-bits with damaged bones were treated surgically with SB-1™ andthe progress of bone recovery was monitored using X-ray tech-niques. The result showed that the SB-1™ was well integrated withthe surrounding host bone and promoted induction of new bone,leading to bonding with newly formed bone and recovery of dam-aged bone tissue. In particular, it was found through an in vivostudy with rabbit that SB-1™ was superior in terms of bondingwith host bones as well as induction and integration with newbone compared to the allografts and bone grafts with low HApcontents.

Zhu et al. [128] evaluated the osteoconductive properties ofnano-HAp material and its potential application as artificial bonein repairing bone defects, and attempted to analyze the scientific

r matrices. Copyright ACS and reproduced with permission [126].

2776 H. Zhou, J. Lee / Acta Biomaterialia 7 (2011) 2769–2781

basis of these properties. Their animal model of bone defects wasbased on the bilateral radius of 39 New Zealand white rabbits,which were randomly divided into an experimental group (bonedefect repaired with nano-HAp artificial bone), a control group(bone defect repaired with HAp artificial bone) and a blank group(defect left empty). The experimental group was found to stimulatea callus bonier than the control and blank groups. These differencesin bone conduction are statistically significant (P < 0.05). Therefore,nano-HAp artificial bone can potentially be used for bone defecttreatment. Osteosarcoma is a primary malignant bone tumor, mostprevalent in children and adolescents, and is usually highly aggres-sive and eventually lethal. Despite multimodal therapies, there isno effective approach to treat this malignant disease. Shi et al.[137] observed the biological response of osteosarcoma cells totwo kinds of nano-Hap: nano-HAp-S and nano-HAp-L. These nano-spheres have the same crystallinity (phase) and morphology butdiffer in size. Cells treated with both forms of nano-HAp wereinhibited and mainly led to apoptotic cell death. The caspase-9-dependent intrinsic apoptotic pathway plays a role in this. Inter-estingly, the suppression and apoptosis of osteosarcoma cells weredirectly related to the size of nanoparticles, and the larger-sizednano-HAp was more effective than the smaller particles. In the col-lagen matrix, tens to hundreds of these nano-blocks combine intoself-assembled biomaterials that have remarkable physical andchemical features such as unique mechanical strength, insensitiv-ity to growth/dissolution and flexible structures [138,139]. Thus,features of smaller HAp nanoparticles may more closely resemblefeatures of HAp during biomineralization than features of the lar-ger HAp particles that are conventionally used. Therefore, nano-HAp may promote osteoblast adhesion, proliferation and synthesisof alkaline phosphatase and lead to rapid repair of hard tissue in-jury [140,141].

Nano-HAp has been reported to be a better candidate in bio-medical applications. Cai et al. [142] prepared nano-HAp, typically

Fig. 6. Bone tissue cross-section of N-HA scaffold under phase contrast (a) 3 and (c) 12 w(b,d) representing birefringence of collagen strands. 200� original magnification. S, scareproduced with permission [144].

20 ± 5, 40 ± 10 and 80 ± 12 nm in diameter, and studied their ef-fects on proliferation of two bone-related cells: bone marrow mes-enchymal stem cells (MSCs) and osteosarcoma cells (U2OS). Cellculture experiments showed improved cytophilicity of the nano-phase mineral relative to conventional HAp. Greater cell viabilityand proliferation of MSCs were measured for nano-HAp, remark-ably so for the 20 nm particles. Interestingly, the growth of osteo-sarcoma cells was inhibited by nano-Hap, and 20 nm particleswere the best retardant. Nano-HAp has been suggested to exhibitfavorable cell proliferation to optimize biological functionality,for which particle size is believed to play a key role. Thesein vitro findings are of great significance for understanding thecytophilicity and biological activity of nanoparticles during bio-mineralization. Li et al. [143] investigated the effects on highlymalignant melanoma cells of nano-HAp particles with differentmorphologies. Three types of HAp particles with different mor-phologies were synthesized and co-cultured with highly malignantmelanoma cells using phosphate-buffered saline (PBS) as a control.A precipitation method with or without citric acid addition as asurfactant was used to produce rod-like nano- and micron-sizedHAp particles, respectively, and a novel oil-in-water emulsionmethod was employed to prepare ellipse-like nano-HAp particles.Experiment results indicated that the particle nanoscale effect,rather than particle morphology of HAp, was more effective ininhibiting highly malignant melanoma cell proliferation. Applefordet al. [144] found that the bone formation and angioconductive po-tential of HAp scaffolds closely matched to trabecular bone in a ca-nine segmental defect after 3 and 12 weeks of implantation.Histomorphometric comparisons were made between naturallyforming trabecular bone (control) and defects implanted with scaf-folds fabricated with micro-size HAp (M-HAp) and nanosized HAp(N-HAp) ceramic surfaces. As shown in Fig. 6, no significant differ-ences were identified between the two HAp scaffolds; however,significant bone in-growth was observed after 12 weeks with

eeks post-surgery with corresponding cross-polarized light micrographs shown inffold; M, mineralized bone; C, collagen birefringence; V, vessel. Copyright ACS and

H. Zhou, J. Lee / Acta Biomaterialia 7 (2011) 2769–2781 2777

43.9 ± 4.1% and 50.4 ± 8.8% of the cross-sectional area filled withmineralized bone in M-HAp and N-HAp scaffolds, respectively. Thisstudy showed the potential of trabecular bone modeled, highlyporous and interconnected HAp scaffolds for regenerativeorthopedics.

5. Composites of HAp and inorganic nanomaterials

The low fracture toughness and poor wear resistance of HAp canbe improved by adding second-phase reinforcement. Carbon nano-tubes (CNTs) have already shown their potential as effective rein-forcements for HAp and other ceramics [145–148] to improvefracture toughness. Reports are available on the processing ofHAp–CNT composite coatings for orthopedic implants throughplasma spraying [149–152], laser surface alloying [153,154], elec-trophoretic deposition [155,156] and aerosol deposition [157]. Inaddition to conventional sintering [158,159] and hot isostaticpressing [160,161], spark plasma sintering (SPS) [161–163] hasalso been employed to fabricate free-standing HAp–CNT compos-ites. Omori et al. [161] reported consolidation of multiwalled(MW) CNTs by SPS followed by dip-coating with HAp and a secondround of SPS consolidation of the coated preform. They observed aconsolidated coating of HAp on CNTs without crack formation. An-other study on SPS of HAp–CNT composites used nano-HAp pow-ders and CNTs, mixed by ball milling, as the starting material.Sarkar et al. [162] found the fracture toughness of HAp–CNT com-posite to be as high as 1.27 MPa m0.5, with 2.5 vol.% CNT addition,which was a 30% increase over HAp. Xu et al. [163] performedextensive mechanical stirring to homogeneously mix CNTs withspray-dried HAp powder to prepare feedstock for SPS processing.The Young’s modulus has been reported to be 131 GPa with2 vol.% CNT addition, although no direct comparison with HAphas been reported. The biocompatibility of CNT-reinforced HApcomposite has also been studied by Xu et al. This group determinedthe beneficial effect of CNT on osteoblast cell proliferation. Lahiriet al. [164] investigated CNT-reinforced HAp composite synthe-sized using SPS. Quantitative microstructural analysis suggeststhat CNTs play a role in grain boundary pinning and are responsiblefor improved densification and retention of nanostructurethroughout the thickness of the sintered pellet. HAp crystal formsa coherent interface with CNTs, resulting in a strong interfacialbond. The uniform distribution of 4 wt.% CNTs in the HAp matrix,good interfacial bonding and fine HAp grain size help to improvefracture toughness by 92% and elastic modulus by 25% comparedto a HAp matrix without CNT.

Xiao et al. [165] introduced a simple and effective approach tofunctionalize MWCNTs by in situ deposition of HAp to improvehydrophilicity and biocompatibility. The scheme of biomineraliza-

Fig. 7. Scheme of biomineralization mechanism of the preparation of HAp–P

tion mechanism of HAp–PEG–MWCNT preparation is shown inFig. 7. First, two types of pre-functionalized MWCNTs were pre-pared: acid-oxidized MWCNTs, and MWCNTs covalently modifiedby PEG. The influence of acid-oxidation time, pre-phosphorylationand PEGylation of MWCNTs on in situ growth of HAp was furtherinvestigated in simulated body fluid (SBF) with ionic concentra-tions of 2, 5 and 10 times, respectively, at 37 �C for 24 h. The resultsshowed that these factors have positive effects on HAp crystalgrowth, with PEGylation of MWCNTs playing a key role duringdeposition. Finally, the methyl thiazolyl tetrazolium (MU) assaywas performed to evaluate cytotoxicity, which showed that PEGy-lated MWCNTs wrapped by HAp crystals have the bestbiocompatibility.

Inspired by self-assembly of nano-HAp association with the67 nm periodic microstructure of collagen, Liao et al. [166] usedMWCNTs with an approximately 40 nm bamboo periodic micro-structure as a template for nano-HAp deposition to form a nano-HAp–MWCNT composite. Spindle-shaped units consisting of anassembly of near-parallel, fibril-like nano-HAp polycrystals wereformed and oriented at a specific angle to the long axis of CNTs.Spindle-shaped units detached from the MWCNT template are ableto maintain the ordered parallel structure of the nano-HAp poly-crystal fibril. Tang et al. [167] found that biomimetic synthesis ofHAp on SDS functionalized MWCNTs by using an alternate soakingprocess (ASP) in Ca/P solutions. The results showed that nano-HApcrystals were formed on SDS functionalized MWCNTs, and miner-alized MWCNTs remained in a dispersed state. HAp–MWCNTnanohybrids, combining the osteconductive properties of HApand the excellent mechanical properties of MWCNTs, will providea promising material for bone tissue engineering.

Balani et al. [151] synthesized HAp-reinforced with sub-micrometer Al2O3 and CNTs as a coating on a Ti–6Al–4 V substratevia plasma spraying. The addition of Al2O3 and CNTs to HAp showsimprovements in the hardness and elastic modulus by 65% and50%, respectively, compared to HAp. Consequently, HAp–Al2O3–CNT coatings have been nano-scratched to determine their wearperformance. Reinforcement of HAp by Al2O3 decreases wear vol-ume by more than 13 times, whereas HAp–Al2O3–CNT coatingshows an even further decreased wear volume of 5 times com-pared to that of HAp–Al2O3 coating. Ajeesh et al. [168] preparedsintered iron oxide–HAp nanocomposite ceramics from powdersproduced through a co-precipitation process. The phase purityand bioactivity of the composites were analyzed as a function ofpercentage of iron oxide in the composite. In all the prepared com-posites, HAp retains the same phase identity and high X-ray opac-ity as a composition containing 40 wt.% iron oxide. Increased cellviability and cell adhesion showed that the prepared composite of-fers considerable potential for bone tissue engineering

EG–MWCNTs. Copyright Elsevier and reproduced with permission [165].

2778 H. Zhou, J. Lee / Acta Biomaterialia 7 (2011) 2769–2781

applications. Jiang et al. [169] designed and synthesized nano-HApcrystals carrying incorporated Fe ions by a co-precipitation meth-od. The results showed that lattice substitution or incorporationoccurs between Fe ions and nano-HAp, which is quite differentfrom the mechanical mixture of Fe3O4 nanoparticles and nano-HAp crystals. The nano-Fe–HAp with a synthetic molar ratio of1:2 (calculated by Fe3O4:HAp) exhibited a needle-like crystal shapeand relatively good crystallinity. However, when the molar ratiowas increased to 1:1, some Fe3O4 nanoparticles were separatedfrom the nano-Fe–HAp crystals. Magnetization measurementsshowed that nano-Fe–HAp crystals were in a superparamagneticstate, proving that the existence of Fe ions in nano-HAp crystalscontributed to the magnetic properties of nano-Fe–HAp crystals.As one component of biomedical composites, nano-Fe–HAp hasthe potential to promote bone regeneration.

Bioactive nano-titania ceramics with excellent biomechanicalcompatibility and bioactivity were prepared by using HAp additiveas a grain growth inhibitor. The mechanical properties of nano-titania ceramics were analogous to those of human bone. Theamount of HAp additive played an important role in determiningthe grain/particle size of nano-titania ceramics, which had a greateffect on osteoblast proliferation in cell culture experiments. Cellculture experiments also showed that the bioactive HAp additiveitself also significantly affected the cytocompatibility of nano-tita-nia ceramics. These results indicated that the content of HAp addi-tive not only had an effect on the bioactivity of nano-titaniaceramics due to the bioactivity of the additive itself, but also hadan effect both on the biomechanical compatibility and bioactivityof nano-titania ceramics by adjusting the grain/particle size ofthe ceramics. The HAp and ZrO2 nanopowders were prepared bychemical reactions and alcohol–aqueous solution heating, respec-tively. The nanosized HAp-ZrO2 powders with a homogeneous dis-tribution could then be synthesized by ball-milling. HAp-ZrO2

bioceramics with small grain sizes could be obtained by usinghot-press sintering technology [170]. The in vitro biocompatibilityof these bioceramics was studied, and the results showed thatthere was no reaction between HAp and ZrO2 powders-whichmay be attributed to the very short sintering time of the hot-presssintering- and that HAp-ZrO2 bioceramics possess non-toxic andnon-allergenic properties.

Biomaterials science has found many ways to enhance differentproperties of synthetic HAp by introducing various ionic substitu-tions into the apatite structure [171]. One of the most interestingsubstitutions is the incorporation of Si, which considerably im-proves the bioactivity of synthetic apatite [172]. The role of Si inbioactive processes involved in new bone tissue formation is wellknown [173]. Carlisle first reported the presence Si traces duringbone mineralization at early stages of calcification, and this hasproven the influence of an Si-deficient diet in diseases such as oste-oporosis [174]. Therefore, Si-substituted HAp (Si–HA) was chosenas an appropriate candidate for the preparation of an improvedCaP material and was subsequently produced by chemical synthe-sis [175,176]. Solla et al. [177] chemically synthesized Si–HA andpresented it as a new material with enhanced bioactivity. Si–HApfilms were deposited by pulsed laser deposition (PLD), using tar-gets composed of HAp mixtures with different Si-containingsources such as SiO2 and diatomaceous earth. Analysis revealedthat Si is successfully incorporated into the HAp structure, as wellas traces of other elements such as Na, Fe or K.

6. Summary and perspective

Nanophase HAp bioceramics have gained importance in the bio-medical field due to their superior biological and biomechanicalproperties. Development of HAp biomedical materials will benefit

from advances in nanotechnology. Several methods for synthesiz-ing HAp on the nanoscale have evolved in the past few decades.Due to the chemical similarity between HAp and mineralized boneof human tissue, synthetic HAp exhibits a strong affinity to hosthard tissues. A significant amount of research in this area is ex-pected to be focused on the nanoscale for enhanced applicationsas resorbable scaffolds that can be replaced by endogenous hardtissues over time. In the future, the ability to functionalize surfaceswith different molecules of varying natures and dimensions basedon their means of attachment to cells, as well as the potential tonanostructure the surface physically, chemically and topographi-cally, will enable selective targeting within biological systems toshow specificity towards individual proteins and peptides. Under-standing the influence of nano-HAp particle size, crystal morphol-ogy controls and interfaces with cells from a higher level isessential for the future development of nanotechnology and bio-technology. Such an interdisciplinary approach is very compli-cated, and the effective collaboration of scientists from differentdisciplines is necessary.

Acknowledgments

This research was supported by the International Research andDevelopment Program of the National Research Foundation of Kor-ea (NRF) funded by the Ministry of Education, Science and Technol-ogy (MEST) of Korea, and National Fisheries Research andDevelopment Institute (Grant Nos. K20091003000, FY2009,20100434961-00).

Appendix A. Figures with essential colour discrimination

Certain figures in this article, particularly Figures 4, 6, 7, are dif-ficult to interpret in black and white. The full colour images can befound in the on-line version, at doi:10.1016/j.actbio.2011.03.019.

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