Particle Accretion Mechanism Underlies Biological Crystal Growth from an Amorphous Precursor Phase

Many biogenic minerals are composed of aggregated particles at the nanoscale. These minerals usually form through the transformation of amorphous precursors into single crystals inside a privileged space controlled by the organism. Here, in vitro experiments aimed at understanding the factors responsible for producing such single crystals with aggregated particle texture are presented. Crystallization is achieved by a two‐step reaction in which amorphous calcium carbonate (ACC) is first precipitated and then transformed into calcite in small volumes of water and in the presence of additives. The additives used are gel‐forming molecules, phosphate ions, and the organic extract from sea urchin embryonic spicules ‐ all are present in various biogenic crystals that grow via the transformation of ACC. Remarkably, this procedure yields faceted single‐crystals of calcite that maintain the nanoparticle texture. The crystals grow predominantly by the accretion of ACC nanoparticles, which subsequently crystallize. Gels and phosphate ions stabilize ACC via a different mechanism than sea urchin spicule macromolecules. It is concluded that the unique nanoparticle texture of biogenic minerals results from formation pathways that may differ from one another, but given the appropriate precursor and micro‐environment, share a common particle accretion mechanism.     

Amorphous Calcium Carbonate is Stabilized in Confinement

Biominerals typically form within localized volumes, affording organisms great control over the mineralization process. The influence of such confinement on crystallization is studied here by precipitating CaCO₃ within the confines of an annular wedge, formed around the contact point of two crossed half‐cylinders. The cylinders are functionalized with self‐assembled monolayers of mercaptohexadecanoic acid on gold. This configuration enables a systematic study of the effects of confinement since the surface separation increases continuously from zero at the contact point to macroscopic (mm) separations. While oriented rhombohedral calcite crystals form at large (>10 µm) separations, particles with irregular morphologies and partial crystallinity are observed as the surface separation approaches the dimensions of the unconfined crystals (5–10 µm). Further increase in the confinement has a significant effect on the crystallization process with flattened amorphous CaCO₃ (ACC) particles being formed at micrometer separations. These ACC particles show remarkable stability when maintained within the wedge but rapidly crystallize on separation of the cylinders. A comparison of bulk and surface free‐energy terms shows that ACC cannot be thermodynamically stable at these large separations, and the stability is attributed to kinetic factors. This study therefore shows that the environment in which minerals form can have a significant effect on their stability and demonstrates that ACC can be stabilized with respect to the crystalline polymorphs of CaCO₃ by confinement alone. That ACC was stabilized at such large (micrometer) separations is striking, and demonstrates the versatility of this strategy, and its potential value in biological systems.     

The Effect of Additives on Amorphous Calcium Carbonate (ACC): Janus Behavior in Solution and the Solid State

Amorphous calcium carbonate (ACC) is an important intermediate in the formation of crystalline CaCO₃ biominerals, where its crystallization is controlled using soluble additives. However, although this transformation often occurs in the solid state, experiments mainly focus on the effect of additives on ACC crystallization in solution. This paper addresses this issue and compares the crystallization, in solution and in the solid state, of ACC precipitated in the presence of a range of additives. Surprisingly, these results show that some additives exhibit a Janus behavior, where they retard crystallization in solution, yet accelerate it in the solid state. This is observed for all of the larger molecules examined, while the small molecules retard crystallization in both solution and the solid state.     

Crystal Growth of Calcium Carbonate in Hydrogels as a Model of Biomineralization

In recent years, the prevalence of hydrogel‐like organic matrices in biomineralization has gained attention as a route to synthesizing a diverse range of crystalline structures. Here, examples of hydrogels in biological, as well as synthetic, bio‐inspired systems are discussed. Particular attention is given to understanding the physical versus chemical effects of a broad range of hydrogel matrices and their role in directing polymorph selectivity and morphological control in the calcium carbonate system. Finally, recent data regarding hydrogel‐matrix incorporation into the growing crystals is discussed and a mechanism for the formation of these single‐crystal composite materials is presented.     

A Mesocrystal‐Like Morphology Formed by Classical Polymer‐Mediated Crystal Growth

Growth by oriented assembly of nanoparticles is a widely reported phenomenon for many crystal systems. While often deduced through morphological analyses, direct evidence for this assembly behavior is limited and, in the calcium carbonate (CaCO₃) system, has recently been disputed. However, in the absence of a particle‐based pathway, the mechanism responsible for the creation of the striking morphologies that appear to consist of subparticles is unclear. Therefore, in situ atomic force microscopy is used to investigate the growth of calcite crystals in solutions containing a polymer additive known for its ability to generate crystal morphologies associated with mesocrystal formation. It is shown that classical growth processes that begin with impurity pinning of atomic steps, leading to stabilization of new step directions, creation of pseudo‐facets, and extreme surface roughening, can produce a microscale morphology previously attributed to nonclassical processes of crystal growth by particle assembly.     

The Intrinsic Ability of Silk Fibroin to Direct the Formation of Diverse Aragonite Aggregates

As an analogue of the main protein contained in naturally formed nacre, reconstituted silk fibroin (SF) from the Bombyx mori silkworm silk shows a strong preference for the formation of the aragonite form of CaCO₃ crystals and allows fine control over their size and morphology. The aragonite phase could be generated via two different routes: direct growth or dissolution and recrystallization, depending on the concentration of Ca²⁺ and SF. Generally, lower concentrations of Ca²⁺ and SF favor the formation of aragonite needles and their aggregates, of which the lattice structure of the precursor is similar to that of the organic matrix in natural shell. Higher concentrations lead to the formation of aragonite aggregates via a dissolution and recrystallization process through intermediates of lens‐like vaterite. Molecular modeling shows that the β‐strand conformers of silk fibroin molecules has an excellent match with the ionic spacing in the aragonite (010) plane, which can promote growth along the (001) long axis of aragonite crystals. This synergy between silk fibroin and the aragonite phase may help our understanding of the function of organic matrices involved in the biomineralization process, and facilitate the fabrication of synthetic materials with the potential for high performance mechanical properties.     

Colloidal Stabilization of Calcium Carbonate Prenucleation Clusters with Silica

Calcium carbonate precipitation proceeds via a complex multistage scenario involving neutral ion clusters as precursors and amorphous phases as intermediates, which finally transform to crystals. Although the existence of stable clusters in solution prior to nucleation has been demonstrated, the molecular mechanisms by which they precipitate are still obscure. Here, direct insight into the processes that drive the transformation of individual clusters into amorphous nanoparticles is provided by progressive colloidal stabilization of different transient states in silica‐containing environments. Nucleation of calcium carbonate in the presence of silica can only take place via cluster aggregation at low pH values. At higher pH, prenucleation clusters become colloidally stabilized and cannot aggregate. Nucleation through structural reorganization within the clusters is not observed under these conditions, indicating that this pathway is blocked by kinetic and/or thermodynamic means. The degree of stabilization against nucleation is found to be sufficient to allow for a dramatic enrichment of solutions with prenucleation clusters and enable their isolation into the dry state. This approach renders direct analyses of the clusters by conventional techniques possible and is thus likely to facilitate deeper insight into the chemistry and structure of these elusive species in the future.     

Citrate Effects on Amorphous Calcium Carbonate (ACC) Structure,Stability, and Crystallization

Understanding the role of citrate in the crystallization kinetics of amorphous calcium carbonate (ACC) is essential to explain the formation mechanisms, stabilities, surface properties, and morphologies of CaCO₃ biominerals. It also contributes to deeper insight into fluid–mineral interactions, both in nature and for industrial processes. In this study, ACC formation and its crystallization are monitored in real time as a function of citrate (CIT) concentration in solution. Additionally, synchrotron radiation pair distribution function analyses combined with solid‐state, spectroscopic, and microscopic techniques are used to determine the effect of CIT on ACC structure, composition, and size. Results show an increase in ACC lifetime coupled with an increase in CIT uptake by ACC and slight changes in ACC atomic structure with an increase in CIT concentration. ACC does not form at concentrations ≥ 75% CIT/Ca and vaterite is absent in all cases where CIT is present. These findings can be explained by CIT binding with Ca ions, thereby forming Ca–CIT complexes in solution and decreasing ACC and calcite saturation levels. The formation of CIT‐bearing ACC with calcitic structure and the absence of vaterite formation suggest that these solution complexes form a calcite‐type atomic arrangement while CIT probably also acts as a growth inhibitor.     

Screening the Incorporation of Amino Acids into an Inorganic Crystalline Host: the Case of Calcite

Organisms have the ability to produce structures with superior characteristics as in the course of biomineralization. One of the most intriguing characteristics of biominerals is the existence of intracrystalline macromolecules. Despite several studies over the last two decades and efforts to mimic the incoporation of macromolecules synthetically, a fundamental understanding of the mechanism of incorporation is as yet lacking. For example, which of the common 20 amino acids are really responsible for the interaction with the mineral phase? Here a reductionist approach, based on high‐resolution synchrotron powder diffraction and analytical chemistry, is utilized to screen all of these amino acids in terms of their incorporation into calcite. We showed that the important factors are amino‐acid charge, size, rigidity and the relative pKa of the carboxyl and amino functional groups. It is also demonstrated that cysteine, surprisingly, interacts very strongly with the mineral phase and therefore, like acidic amino acids, becomes richly incorporated. The insights gained from this study shed new light on the incorporation of organic molecules into an inorganic host in general, and in particular on the biomineralization process.     

Silk Fibroin‐Regulated Crystallization of Calcium Carbonate

Mollusk shell is one of the best studied of all calcium carbonate biominerals. Its silk‐like binder‐matrix protein plays a pivotal role during the formation of aragonite crystals in the nacre sheets. Here, we provide novel experimental insights into the interaction of mineral and protein compounds using a model system of reconstituted Bombyx mori silk fibroin solutions serving as templates for the crystallization of calcium carbonate (CaCO₃). We observed that the inherent (self‐assembling) aggregation process of silk fibroin molecules affected both the morphology and crystallographic polymorph of CaCO₃ aggregates. This combination fostered the growth of a novel, rice‐grain‐shaped protein/mineral hybrid with a hollow structure with an aragonite polymorph formed after ripening. Our observations suggest new hypotheses about the role of silk‐like protein in the natural biomineralization process, but it may also serve to shed light on the formation process of those ‘ersatz’ hybrids regulated by artificially selected structural proteins.     

Additives Control the Stability of Amorphous Calcium Carbonate via Two Different Mechanisms: Surface Adsorption versus Bulk Incorporation

The mechanisms by which organisms control the stability of amorphous calcium carbonate (ACC) are yet not fully understood. Previous studies have shown that the intrinsic properties of ACC and its environment are critical in determining ACC stability. Here, the question, what is the effect of bulk incorporation versus surface adsorption of additives on the stability of synthetic ACC, is addressed. Using a wide range of in situ characterization techniques, it is shown that surface adsorption of poly(Aspartic acid) (pAsp) has a much larger stabilization effect than bulk incorporation of pAsp and only 1.5% pAsp could dramatically increase the crystallization temperature from 141 to 350 °C. On the contrary, surface adsorption of PO₄^3? ions and OH^? ions does not effectively stabilize ACC. However, bulk incorporation of these ions could significantly improve the ACC stability. It is concluded that the stabilization mechanism of pAsp is entirely different from that of PO₄^3? and OH^? ions: while pAsp is effectively inhibiting calcite nucleation at the surface of ACC particle, the latter acts to modify the ion mobility and delay crystal propagation. Thus, new insights on controlling the stability and crystallization processes of metastable amorphous materials are provided.     

The Coral Protein CARP3 Acts from a Disordered Mineral Surface Film to Divert Aragonite Crystallization in Favor of Mg‐Calcite

Stony corals construct their aragonite skeleton by calcium carbonate precipitation, in a process recently suggested to be biologically controlled. Amorphous calcium carbonate and small amounts of calcite are also reported recently, however, their functional role is unknown. Coral acid‐rich proteins (CARPs) are extracted from the coral skeleton and are shown to be active in calcium carbonate precipitation in vitro. However, individual function of these proteins in coral mineralization is not known. Here, the regulatory activity of the aspartate‐rich CARP3 protein is examined. The whole protein and two peptides representing its acidic domain and its variable domain are used in CaCO₃ precipitation reactions from Mg‐rich solutions. The biomolecules alter crystallization pathways, promoting Mg‐calcite in place of aragonite, with the acidic peptide capable of eradicating aragonite formation. The activity of CARP3 and its representative peptides is exerted from disordered CaCO₃ mineral phases, coating the crystals formed, as shown by 2D ¹H–¹³C heteronuclear correlation nuclear magnetic resonance (NMR) measurements, localizing organic protons in atomic proximity to disordered carbonate carbons. The structures of the protein and individual domains as derived from NMR measurements and folding calculations and their amino acid compositions are discussed in the context of their observed activity and its implication to mineralization in hard corals.     

Time‐Resolved Evolution of Short‐ and Long‐Range Order During the Transformation of Amorphous Calcium Carbonate to Calcite in the Sea Urchin Embryo

Use of amorphous precursors is a widespread strategy in biomineralization. In sea urchin embryos, controlled transformation of amorphous calcium carbonate (ACC) to calcite results in smoothly curving and branching single crystals. However, the mechanism of the disorder‐to‐order transformation remains poorly understood. Here, the use of strontium as a probe in X‐ray absorption spectroscopy (XAS) greatly facilitates investigation of the evolution of order. In pulse‐chase experiments, embryos incorporate Sr²⁺ from Sr‐enriched seawater into small volumes of the growing endoskeleton. During the chase, the Sr‐labeled mineral matures under physiological conditions. Based on Sr K‐edge spectra of cryo‐frozen whole embryos, it is proposed that the transformation occurs in three stages. The initially deposited calcium carbonate has short‐range order resembling synthetic hydrated ACC. Within 3 h, the short‐range order of calcite is established. Between 3 h and 24 h, the short‐range order does not change, while long‐range order increases. These results refute the notion that organisms imprint the local order of the final crystal on ACC. Furthermore, it is proposed that the intermediate is more similar to disordered calcite than to anhydrous ACC. Pulse‐chase experiments in conjunction with heavy element labeling have great potential to improve understanding of phase transformations during biomineralization.     

银杏中种皮形态建成及结构特征

由于银杏中种皮的发育及结构与早期种仁形成、后期种核贮藏特性密切相关,本文利用半薄切片和扫描电镜观察了银杏中种皮的分化、发育过程及结构特征。结果表明,银杏授粉后不久珠被分化并形成细胞大小、形态明显差异的3个区,其中中区的5～6层细胞体积小,细胞核大,细胞排列紧密,这几层细胞即将发育形成中种皮;中种皮发育早期,细胞数目增殖速度较快,细胞质染色深,细胞核大并位于细胞的中央;授粉后70d细胞数目不再增加,此时细胞发生径向扩张,随着次生壁的形成,中种皮细胞壁开始加厚;授粉后100d中种皮细胞壁迅速增厚,细胞径向扩张加剧,细胞中央形成完整大液泡,细胞核被挤向靠近细胞壁的部位;之后细胞中液泡破裂,细胞质降解,细胞核解体消失,形成次生壁强烈增厚的多层结构;中种皮发育成熟后由大量管胞组成,根据管胞分布位置的不同可分为螺纹管胞和孔纹管胞2种,管胞侧壁均分布有大量纹孔。     

Porous Single Crystals of Calcite from Colloidal Crystal Templates: ACC Is Not Required for Nanoscale Templating

This article investigates the formation of nanostructured single crystals of calcite using direct, ion‐by‐ion precipitation methods and shows that single crystals with complex morphologies and curved surfaces can readily be formed using this technique. Calcite crystals with inverse opal and direct opal structures are prepared using templates of colloidal crystals and polystyrene reverse opals, respectively, and excellent replication of the template structures are achieved, including the formation of 200‐nm spheres of calcite in the direct opal structure. These highly porous crystals also display extremely regular, crystalline gross morphologies. The methodology is extremely versatile and challenges the preconception that nanostructured crystals cannot be prepared by simple diffusion of reagents into the template due to blocking of the channels. The results are also discussed in light of alternative templating methods using amorphous calcium carbonate (ACC) as a precursor phase and provide insight into the role of ACC in biological calcification processes.     

氟化钙(CaF2)晶体研发进展

简述了氟化钙(CaF2)晶体的发展历史．总结其光学特性，介绍了氟化钙晶体的生长新工艺技术和发展。综述了氟化钙晶体在现代信息技术中的应用，特别对氟化钙晶体在紫外波段激光或光刻系统中的应用和发展状况作了重点说明，同时进一步指出了目前存在的问题，最后对氟化钙晶体的发展前景作了展望。     

Stabilization of Amorphous Calcium Carbonate with Nanofibrillar Biopolymers

Calcium carbonate is the most abundant biomineral that is biogenically formed with a vast array of nano and microscale features. Among the less stable polymorphs present in mineralized organisms, the most soluble, amorphous calcium carbonate (ACC), formed in chitin exoskeletons of some crustacea, is of particular interest since aqueous stability of isolated ACC is limited to a few hours in the absence of polyanions or magnesium. Here the influence of a selection of biopolymer gels on the mineralization of calcium carbonate is investigated. Mineralization is achieved in all biopolymers tested, but is particularly abundant in collagen hydrogels, in which a significant proportion of the calcium carbonate (≈18%) is found to be amorphous. In dense collagen gels, this amorphous fraction does not crystallize for up to six weeks in deionized water at room temperature. The reason why collagen in particular should stabilize this phase remains obscure, although the results suggest that the fiber diameter, fiber spacing, and the amphoteric nature of collagen fibers are important. Upon immersion in phosphate containing solutions, the calcium carbonate present within the collagen hydrogels is readily converted to carbonated hydroxyapatite, enabling the formation of a stiff bone‐like composite containing 78 wt% mineral, essentially equivalent to cortical bone.     

Optical Properties and Growth Aspects of Silver Nanoprisms Produced by a Highly Reproducible and Rapid Synthesis at Room Temperature

A rapid and readily reproducible seed‐based method for the production of high quality silver nanoprisms in high yield is presented. The edge‐length and the position of the main plasmon resonance of the nanoprisms can be readily controlled through adjustment of reaction conditions. From UV‐vis spectra of solutions of the nanoprisms, the inhomogeneously broadened line width of the in‐plane dipole plasmon resonance is measured and trends in the extent of plasmon damping as a function of plasmon resonance energy and nanoprism size have been elucidated. In addition, an in‐depth analysis of the lamellar defect structure of silver nanoprisms is provided that confirms that the defects can lead to a transformation of the crystal structure in the vicinity of the defects. These defects can combine give rise to lamellar regions, thicker than 1 nm, that extend across the crystal, where the silver atoms are arranged in a continuous hexagonal‐close‐packed (hcp) structure. This hcp structure has a periodicity of 2.50 Å, thus explaining the 2.50 Å lattice fringes that are commonly observed in 〈111〉 oriented flat‐lying nanoprisms. A new understanding of the mechanisms behind anisotropic growth in silver nanoprisms is presented.