Transparent Aggregates of Nanocrystalline Hydroxyapatite Anders C. S. Jensen, Casper J. S. Ibsen, Duncan Sutherland, and Henrik Birkedal* Department of Chemistry and iNANO, Aarhus University, DK-8000 Aarhus C, Denmark * S Supporting Information ABSTRACT: Assemblies of nanoparticles into transparent aggre- gates have solicited strong research interest in the form of both crystalline or amorphous aggregates of nanoparticles. In the present work, we make short-range ordered several millimeter-sized transparent aggregates of citrate modified calcium phosphate nanoparticles and discuss the mechanism of their formation. Microparticles of hydroxyapatite (HAP) nanocrystals and amor- phous calcium phosphate (ACP) were synthesized with citrate as a growth and assembly modifier. Millimeter-sized transparent aggregates of these microparticles were made with 0 to 7.5% citrate/Ca 2+ . The degree of crystallinity, i.e., the ratio between nanocrystalline HAP and ACP in the microparticles, was determined by Rietveld refinement of powder X-ray diffraction data with an internal standard. It was found to decrease with increasing citrate concentration. Citrate also reduced the nanocrystallite size at low citrate concentrations. Above ∼3% added citrate, the crystallite size did not reduce further. Transparent aggregates were obtained by drying a suspension of particles. The aggregates lacked long-range order and in many cases featured spiral fractures partially propagating through the aggregates. The assembly mechanisms were studied by in situ video imaging, polarized optical microscopy, transmission electron microscopy, and confocal microscopy. The transparent aggregates consisted of polydisperse microparticles. The transparent aggregates form due to evaporation, but sedimentation leads to vertical size segregation with larger microparticles preferentially located at the bottom of the sample. ■ INTRODUCTION In nature, highly ordered and homogeneous assemblies of nanoparticles are widely used to make versatile materials. 1-5 These find use in structural support, 5 sensing, 6 and even in reproductive organs. 7 One biomineral that is famous for its optical properties is mother of pearl or nacre. It shows remarkable iridescent colors as a result of diffraction from layers of aragonite platelets with a periodicity of ∼500 nm, thereby reflecting visible light. In order to create this mesostructure, the platelets must be homogeneous and ordered. 2,4,8 By the example of nacre, it is clear that any transparent aggregates of nanoparticles must avoid any ordered structure in the range of visible light. Several examples of these materials have already been made. However, many of these materials are thin films made from inorganic/organic composites. 9-13 They can also be made from Ag nanoribbons 14 or by denaturized cellulose. 15 Although there are numerous examples of trans- parent thin films, there are only a handful of transparent bulk materials 16-19 made from either TiO 2 or CaCO 3 in combination with polymers. They are typically densely packed disordered, homogeneous structures. In the thin films, high aspect ratio building blocks are common with a tendency for the materials to display local ordering of particles. Transparent bulk materials have thus far only been reported for aggregates of isotropic particles of <5 nm size. Oaki et al. found no local order observed in these bulk materials by cross polarized microscopy. 17 In the current study, we obtain transparent aggregates of citrate stabilized amorphous calcium phosphate (ACP)/nano- crystalline hydroxyapatite (HAP) particles. Citrate is an important component in bone where it is tighly associated with bone apatite nanocrystals. 30 It is also associated with a host of other apatitic biomineralized tissues. 31 In synthetic systems, citrate has been shown to strongly impact apatite crystallization. At near neutral pH, it stabilizes ACP 32 and reduced apatite crystallite size, 33 an effect also observed at elevated tempar- atures. 34,35 Herein we have synthesized HAP at high pH following the work of Ibsen et al. 36,37 but using citrate as a growth modifier. The synthesis yielded transparent aggregates with a structure lacking long-range order. We shed light on the internal structure of the aggregates and on the mechanism that governs the assembly of anisotropic nanoparticle assemblies. ■ EXPERIMENTAL SECTION Microparticle Synthesis. All samples were made from 10 mL of 0.6 M CaCl 2 ·2H 2 O (>99% Sigma-Aldrich) that was temperated under stirring at 25 °C for 5 min. A mixture of 10 mL of 0.36 M NaH 2 PO 4 · H 2 O (>99% Sigma-Aldrich) and 0.84 M NaOH (>98% Sigma- Aldrich) and 10 mL 0-0.045 M Na 3 (citrate)·2H 2 O (>99% Sigma- Aldrich) was added. The reaction was kept at 25 °C for 24 h using a water bath controlled by a Julabo ED/F 12 (Julabo GmbH, Seelbach, Germany) and a MIXdrive 15 2Mag magnetic stirrer (2mag AG, Mü nchen, Germany). The pH started at a value of about 12.5 and fell during the reaction as phosphate and hydroxy ions were consumed during formation of calcium phosphates. All samples were purified by Received: July 18, 2014 Revised: October 10, 2014 Article pubs.acs.org/crystal © XXXX American Chemical Society A dx.doi.org/10.1021/cg501080c | Cryst. Growth Des. XXXX, XXX, XXX-XXX