CRYSTAL GROWTH & DESIGN
Selective Synthesis of Various Nanoscale Morphologies of Hydroxyapatite via an Intermediate phase
2008 VOL. 8, NO. 3 1055–1059
Hiroyuki Ito, Yuya Oaki, and Hiroaki Imai* Department of Applied Chemistry, Faculty of Science and Technology, Keio UniVersity, 3-14-1 Hiyoshi, Kohoku, Yokohama 223-8522, Japan ReceiVed May 15, 2007; ReVised Manuscript ReceiVed NoVember 15, 2007
ABSTRACT: Dicalcium phosphate (DCP) was found to be a suitable precursor for nanoscopically controlled hydroxyapatite (HAp) crystals. Nanoscale needles, fibers, and sheets of HAp were selectively prepared through the hydrolysis of a solid precursor crystal of DCP in an alkali solution by varying the pH and ion concentrations. An oriented array of bundled nanoneedles of HAp elongated in the c axis was obtained under a highly basic condition at pH 11–13. The ordered architecture originated from the spatially periodic nucleation of HAp seeds on the DCP surface through topotactic solid–solid transformation. Long HAp fibers were observed under a relatively mild basic condition at pH 9–10. The fibrous morphology evolved from the nanoneedles produced by the solid–solid transformation with the elongation of the c-axis through a dissolution-precipitation route. Flaky HAp nanosheets consisting of a parallel assembly of nanoneedles were observed with an excess amount of phosphate ions under mild basic conditions. The presence of phosphate ions suppressed the solid–solid transformation and promoted the formation of a two-dimensional structure with the dissolution-precipitation process. Introduction Hydroxyapatite (HAp, Ca5(PO4)3OH) is attracting much attention as a material for artificial bones,1 scaffolds for tissue engineering,2,3 and chromatographic packing4 because of its high bioactivity and particular adsorbability for various ions and organic molecules. The performance of HAp in these applications is influenced by the nanoscale morphology and crystallinity. Fibrous and needle-like morphologies with a high specific surface area are advantageous for adsorption and ion exchange, while high mechanical strength due to a rigid structure is required for other applications. Moreover, the chemical and biological properties are known to depend on the crystal faces of HAp.5–7 Therefore, techniques for the preparation of HAp with a controlled architecture would be highly important for the development in biologically applicable inorganic materials. Wet chemical routes using aqueous solution systems have been applied for the morphological control of HAp crystals. Nanofibers of HAp were prepared in reverse micelles under hydrothermal conditions.8 Needle-like HAp crystals were produced by precipitation with the direct reaction of calcium hydroxide and phosphoric acid with organic molecules, such as collagen.9 Various fibrous morphologies were also prepared through the hydrolysis of precursor crystals, such as dicalcium phosphate (DCP,monetite,CaHPO4),10,11 R-tricalciumphosphate(R-TCP),12,13 and octacalcium phosphate (OCP).14 On the other hand, sheetlike and flaky HAp crystals were formed on the surface of glass15,16 and titanium dioxide17 in simulated body fluid (SBF). In addition, calcium phosphate cement was hydrolyzed and transformed into flaky HAp by soaking in SBF18 and calf serum.19 However, the controllability of the nanoscale morphologies of HAp crystals by these methods was insufficient; in addition, the formation mechanism of the nanoscale fibrils and sheets has not been clarified. The elucidation of the detailed mechanism would provide great advantages for the synthesis of biomaterials as well as a better understanding of the mineralization process of HAp in the body of vertebrates. * Corresponding author: E-mail:
[email protected]. Telephone: +8145-566-1556. Fax: +81-45-566-1551.
Recently, our research group reported the production of nanotextured and nanofibrous HAp through the hydrolysis of DCP prepared in a gelatin gel containing phosphate ions.20 In previous works, the formation mechanism of the hierarchical architectures was still quite unclear, whereas the effect of the organic molecule was suggested to be important for the morphological evolution. The aim of the present work is the selective production of nanoscale morphologies of HAp and the clarification of the formation mechanism of HAp through the hydrolysis of the precursor crystal. Here, we found that DCP produced by drying dicalcium phosphate dihydrate (DCPD, brushite, CaHPO4 · H2O) at 60 °C was a suitable precursor for the preparation of various types of nanoscale HAp. Although the conversion of DCPD into DCP commonly requires heating at approximately 200 °C,21–23 the presence of nitrate or chloride ions on the surface promoted the dehydration. The final nanoscale morphologies of HAp obtained from the precursor DCP were influenced by the conditions for hydrolysis, including the pH and the concentration of calcium and phosphate ions. In this study, the formation mechanism of various types of nanostructured HAp is discussed in the context of its morphological evolution under various conditions. The preparation techniques for nanostructured HAp crystals would be applicable for biomedical and chemical applications; moreover, new findings on the formation of the nanostructures will be informative for the science and technology of wet-chemical materials processing. Experimental Section The preparation of DCP as a precursor crystal with a platy shape was performed using a conventional solution technique. A 50 dm3 solution of 1.1 M NH4H2PO4 (Kanto Chemical, 99.0%) was mixed with the same volume of a 2.7 M Ca(NO3)2 · 4H2O (Junsei Chemical, 98.0%) solution. The mixture was vigorously stirred for 30 min to give a white precipitate of DCPD. We obtained DCP by filtration and subsequent drying of the precipitate in air at a temperature between 60 and 250 °C for 24 h. Commercially available DCPD (Junsei Chemical, 98.0%) was used as a reference sample. A 40 dm3 suspension containing 0.4 g of the precursor DCP was stirred at ca. 70 °C for 3 h for the hydrolysis of the precursor crystals. The pH of the suspension was adjusted and kept at a specific value in
10.1021/cg070443f CCC: $40.75 2008 American Chemical Society Published on Web 02/09/2008
1056 Crystal Growth & Design, Vol. 8, No. 3, 2008
Figure 1. XRD patterns of various samples. (a) as-deposited, (b) dried at 60 °C, (c) heated at 250 °C.
Ito et al.
Figure 2. XRD patterns of HAp crystals obtained in various alkaline solutions. (a) pH 9.0, (b) pH 13.0, (c) pH 9.0 with 3.0 M (NH4)2HPO4.
the range of 9.0 and 13.0 by the addition of ammonium hydroxide or sodium hydroxide during the hydrolysis reaction. Aqueous solutions including Ca(NO3)2 · 4H2O (Junsei Chemical, 98.0%, 0.1 M) and (NH4)2HPO4 (Kanto Chemical, 99.0%, 3.0 M) were also used for the hydrolysis. The solid products were washed with purified water and dried at 60 °C for 24 h. The morphologies of the products were characterized using a fieldemission scanning electron microscope (FESEM, Hitachi S-4700) and a field-emission transmission electron microscope (FETEM, FEI TECNAI F20). The X-ray diffraction (XRD) patterns were recorded on a Rigaku RAD-C system with Cu KR radiation. Fourier-transform infrared (FTIR) spectrometry was performed with a BIO-RAD FTS60A.
Results and Discussion A white precipitate was immediately produced in a highly supersaturated solution containing Ca(NO3)2 and NH4H2PO4. According to the XRD patterns (Figure 1), the wet precipitate and dried powder were identified to be DCPD and DCP, respectively. This means that DCPD rapidly changed into DCP with dehydration at 60 °C as well as at 250 °C. In contrast, a high temperature above 200 °C was required for the conversion of commercially available DCPD powder into DCP. An absorption peak at 1380 cm-1 corresponding to nitrate ions was observed in the FTIR spectra for DCP crystals prepared in a solution of Ca(NO3)2 and NH4H2PO4 (Figure S1, Supporting Information). Dipping of the commercial DCPD powder in an HNO3 or NH4NO3 solution promoted the transformation into DCP at 60 °C. Thus, the presence of nitrate ions on the surface induced the dehydration of DCPD at a low temperature. In our previous work,20 gelatin was suggested to promote the dehydration of DCPD. However, we now propose nitrate ions originating from Ca(NO3)2 as the real promoter in that system. In addition, we also found that chloride ions had the same promoting effect on the dehydration of DCPD crystals. As shown in Figure 2, HAp crystals were obtained in various alkali solutions at pH 9.0–13.0 from DCP crystals dried at 60 °C. On the other hand, DCP powder heated at a high temperature above 250 °C was not reacted in the alkali solutions. Since an absorption peak at 720 cm-1 correspond-
Figure 3. FESEM images of DCP dried at 60 °C.
ing to a stretching vibration mode of P-O-P bonds was detected in FTIR spectra for DCP treated at 250 °C (Figure S1, Supporting Information), the condensation of HPO42occurred on the surface of DCP. It has been reported that the crystal growth of HAp was inhibited by condensed phosphate.24–26 Therefore, the presence of condensed phosphate on the surface was deduced to prevent the transformation of precursor crystal DCP into HAp. In consequence, the low-temperature preparation of DCP from DCPD attached with nitrate ions is essential for the production of HAp in alkali solutions. As shown in Figure 3, DCP crystals produced from DCPD were platy-shaped, with a width of 5–50 µm and a thickness of 0.5–2.0 µm. A smooth surface was observed on the solid plates of DCP. Interestingly, various types of nanostructured HAp were formed from the solid crystals of DCP, as shown in Figures 4, 5, and 6, although the macroscopic morphology was hardly changed with the transformation. The nanometric morphology of HAp crystals was fundamentally influenced by varying the pH value. The resultant form obtained at pH 11.0–13.0 was an oriented array of bundled needles with a diameter of 150–200 nm (Figure 4a,b). The bundled needles were composed of ca. 10 nm-wide fibrous units elongated in the c direction (Figure 4c,d). Macroscopically, the c axis of the HAp needles was arranged parallel to the top face of the
Synthesis of Various Morphologies of Hydroxyapatite
Figure 4. FESEM images (a-c) and FETEM images (d) of HAp hydrolyzed for 3 h in an NaOH solution at pH 13.
Figure 5. FESEM images (a-c) and FETEM images (d) of HAp hydrolyzed for 3 h in an NH4OH solution at pH 9.
platy form. Long HAp fibers elongated along the c axis were obtained at pH 9.0–10.0 (Figure 5). Since the diffraction peaks in Figure 2a were relatively sharpened, the crystallinity of the long fibers was higher that that of the bundled needles. In this case, however, the orientation of the fibers was not directed in a particular direction. The nanoscale morphology was found to vary with the addition of specific ions into the alkaline solution. In the presence of 0.1 M calcium ions in an NH4OH solution at pH 10.0, we obtained needle-like HAp instead of long fibers. As shown in Figure 6, HAp nanosheets were produced at pH 9.0 in the presence of a sufficient amount of phosphate ions with the addition of 3.0 M (NH4)2HPO4. According to the X-ray diffraction peaks in Figure 2c, the crystallinity of
Crystal Growth & Design, Vol. 8, No. 3, 2008 1057
Figure 6. FESEM images (a-c) and FETEM images (d) of HAp hydrolyzed for 3 h in an NH4OH solution at pH 9 with 3.0 M of (NH4)2HPO4.
the nanosheets was lower than that of other products obtained in the absence of ionic additives. The split edges and alignment of fibrous units shown in Figure 6c,d suggest that the nanosheets were constructed with the parallel assembly of HAp nanoneedles elongated in the c direction. The presence of spots corresponding to (00l) and (h00) planes in an electron diffraction pattern (Figure S2, Supporting Information) indicates that the HAp needles were aligned in the [120] direction in the sheet. In our previous study,20 we observed oriented nanoscale units in the DCP crystals produced from DCPD containing gelatin molecules. The morphological evolution with the dehydration was tentatively ascribed to the presence of the organic molecules. In that case, a nanostructured HAp was certainly obtained through the transformation of the nanostructured DCP. In the present work, however, various types of nanostructured HAp were also found to be produced from the solid plates of DCP without any organic additives. The formation mechanism of the nanoarchitectures from a solid plate is discussed from the structural evolution of HAp at the early stages of crystal growth. At a very early stage of the formation of HAp needles, spindle-shaped seeds of HAp were immediately formed on the surface of the precursor DCP under a high pH condition (Figure 7a–c). The ridge of the spindle seeds grew up into an oriented array of bundled needles. The c axis of HAp, which was parallel to the surface of the plate, was inherited from the crystallographic direction of the precursor. This indicates that the topotactic transition, in which the crystal lattice of the product phase shows one or more crystallographically equivalent and orientational relationships to the crystal lattice of the parent phase, occurred with the transformation from DCP to HAp. This solid-state reaction was rapidly induced under a highly basic condition, as shown in eq 1. Since the spatially periodic nucleation of HAp would occur in the nanoscale on the DCP surface, the well-arranged HAp nanoneedles were formed through the solid–solid phase transition. Thus, we could show that the nanostructure of HAp was directly formed on the smooth surface of a solid DCP crystal.
1058 Crystal Growth & Design, Vol. 8, No. 3, 2008
Ito et al.
Figure 7. FESEM images of the growth behavior of various nanostructures of HAp at the initial stages. The reaction periods were (a) 1, (b) 5, and (c) 30 s in an NaOH solution at pH 13.0, (d) 20, (e) 30, and (f) 40 min in an NH4OH solution at pH 10.0, and (g) 5, (h) 10, and (i) 30 min in an NH4OH solution at pH 9.0 containing 3.0 M (NH4)2HPO4. Scale bar ) 500 nm.
Solid–solid transformation 5CaHPO4 + 6OH- f Ca5(PO4)3OH + 2PO34 + 5H2O (1) Solid–liquid–solid transformation CaHPO4 + OH- f Ca2+ + PO34 + H2O
(2a)
5Ca2+ + 3PO34 + OH f Ca5(PO4)3OH
(2b)
The flaky HAp nanosheets were produced from small grains formed on the DCP surface in an NH4OH solution containing phosphate ions (Figure 7g,h). The orientation of the grown nanosheets was not correlated to that of the precursor, in contrast to the ordered array of the nanoneedles (Figure 7i). This suggests that the nanosheets were not produced through the solid-state transformation of DCP. The presence of a sufficient amount of phosphate ions would inhibit the direct transformation from DCP into HAp because the preferential dissolution of phosphoric acid from DCP (eq 1) was suppressed. In this case, the formation of HAp could be dominated by a solid–liquid–solid route including the total dissolution of the precursor DCP and subsequent crystal growth of HAp from the solution, as expressed by eq 2. Initially, many tiny seeds were formed on the DCP surface (Figure 7g,h), and flower-like sheets were then constructed on the seeds through the solid–liquid–solid transformation (Figure 7i). Since a similar sheet-like morphology of HAp was observed in SBF, the presence of an excess amount of phosphate ions is deduced to cause the two-dimensional growth. A specific adsorption of
phosphate ions on a crystal face, such as (110), could lead to the formation of the flaky morphology. However, further investigation is required to clarify the detailed formation mechanism of the specific morphology. As shown in Figure 7d,e, the initial stage of the formation of long HAp fibers was almost the same as that of the bundled needles, while the reaction was quite slower than that under the highly acidic condition. This means that the long fibers were produced by the crystal growth of small seeds formed through the solid–solid transformation. However, the ordered arrangement of the bundled needles deformed with the growth of the long fibers with the reaction time (Figure 7f). Finally, the randomly arranged long fibers evolved from the fibrous seeds with the dissolution of DCP (Figure 5a). The formation of a variety of nanostructured HAp is ascribed to the balance of the topotactic solid–solid phase transition and the solid–liquid–solid process, including dissolution–crystallization in solution. Under a highly basic condition, an ordered array of HAp nanoneedles is formed because the topotactic phase transition rapidly occurs with the dissolution of phosphoric acid. On the other hand, HAp sheets were produced with the dissolution–crystallization process with the inhibition of the preferential dissolution of phosphoric acid. Under a moderately basic condition, long HAp fibers grew with the dissolution–crystallization process from small fibrous seeds produced through the topotactic transformation of the precursor crystals. The formation of highly crystalline fibrous HAp is assisted by the dissolution of the precursor crystal and Ostwald ripening, including the dissolution of the products.
Synthesis of Various Morphologies of Hydroxyapatite
When the total dissolution of the precursor crystal (eq 2a) was suppressed by the addition of an excess amount of calcium ions, bundled needles were formed through the topotactic phase transition with preferential dissolution of phosphate ions. Conclusions Nanoscale needles, fibers, and sheets of hydroxyapatite (HAp) were selectively prepared by the hydrolysis of dicalcium phosphate (DCP) in alkali solutions by varying the pH and ion concentrations. The selective preparation is associated with the reactivity of the precursor DCP under various basic conditions. The formation of a variety of nanostructured HAp is attributed to the balance of the topotactic solid–solid phase transition and dissolution–crystallization processes. Powdery R-tricalcium phosphate is another candidate for a precursor of the nanoscale HAp.12,13 In that case, however, it is difficult to control the macroscopic form of the resultant HAp because the precursor crystal is commonly prepared by a conventional dry method. On the other hand, the hierarchical architecture of including the macroscopic morphology and the nanostructure could be controlled by a biomimetic wet-chemical route. Therefore, the total wet-chemical technique of the various types of nanostructured HAp described in the present study could have a wide variety of biomedical and chemical applications such as substitution of bones and adsorption of biomolecules. Moreover, the detailed mechanism of the formation of the nanostructured HAp is highly informative for understanding the biomineralization process in vertebrates because the transformation of a precursor crystal has been suggested to be associated with the formation of bones and teeth.27,28 Supporting Information Available: FTIR spectra of DCP, FFT image of HAp nanosheet. This material is available free of charge via the Internet at http://pubs.acs.org.
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CG070443F
