Frontiers of Materials Science (v.3, #2)

Editorial for Biomin-10 by Fu-zhai Cui; Lun Bai (103-103).

Overview of the amorphous precursor phase strategy in biomineralization by Steve Weiner; Julia Mahamid; Yael Politi; Yurong Ma; Lia Addadi (104-108).
It was assumed for a long time that organisms produce minerals directly from a saturated solution. A few exceptions were known, including the well documented mineralized teeth of the chiton. In 1997 it was demon-strated that sea urchin larvae form their calcitic spicules by first depositing a highly unstable mineral phase called amorphous calcium carbonate. This strategy has since been shown to be used by animals from other phyla and for both aragonite and calcite. Recent evidence shows that vertebrate bone mineral may also be formed via a precursor phase of amorphous calcium carbonate. This strategy thus appears to be widespread. The challenge now is to understand the mechanisms by which these unstable phases are initially formed, how they are temporarily stabilized and how they are destabilized and transform into a crystalline mature product.
Keywords: amorphous calcium carbonate; precursor phase; calcium carbonate

The ocean hosts inorganic raw materials to a magnitude, which surpasses the resources of these materials available on land. Those mineral resources include industrial minerals, metalliferous oxides, hydrothermal metalliferous sulfides, and dissolved minerals. Hence, a significant source of minerals for sustainable recovery in the future may be ocean waters. Among of those mineral resources, there are two kinds of very important minerals which are consolidated on the seabeds of ocean basins in polymetallic nodules and on the surface of seamounts in polymetallic crusts. Until now, the (bio)- chemical processes that result in the formation of metal deposits in the form of nodules or crusts are not understood. In the present review, we concentrate on the (potential) biogenic origin of nodule and crust formation.We studied polymetallic/ferromanganese nodules that had been collected from the Clarion-Clipperton Zone in the Eastern Pacific Ocean, by high-resolution scanning electron microscopy (HR-SEM) to search for microorganisms. The nodules are made up of small-sized micronodules, 100 to 450 μm in size, which are bound/glued together by an interstitial whitish material. In these micronodules, dense accumulations of microorganisms/bacteria can be visualized that display only two morphotypes: (i) round-shaped cocci and (ii) elongated rods. The microorganisms are decorated on their surfaces with S-layers, which are indicative for bacteria. Moreover, the data suggest that these S-layers are the crystallization seeds for the mineralization process. We conclude that the mineral material of the nodule has a biogenic origin and propose consequently the view that mineralization in nodules is caused by biologically controlled mineralization processes.In a second series of investigations, first evidence for a biogenic origin of ferromanganese crusts formation is given. Crusts were obtained from the Magellan seamounts and analyzed for their chemical composition using the EDX technique. Again, special emphasis had been put on the (potential) biogenic origin of the mineral deposition in these ferromanganese crusts. We could demonstrate by HR-SEM that, in those deposits, vast amounts of coccoliths (calcareous unicellular algae) exist. Surprisingly, the coccoliths are composed of Mn besides Ca and C, as analyzed by EDX. This result could be further substantiated by EDX mappings. We propose that initiation of crust formation involves the dissolution of calcite from the coccoliths, resulting in an oxidation of Mn2+ to Mn4+ and subsequent precipitation of Mn4+O2. Following this scheme, it can be assumed that crust formation may serve as an example for a biologically induced mineralization process.
Keywords: polymetallic nodules; ferromanganese crusts; bacteria; coccolithophores; microorganisms; biomineralization; biogenic materials

Abstract Biomineralization processes result in organic/inorganic hybrid materials with complex shapes, hierarchical structures, and superior material properties. Recent developments in biomineralization and biomaterials have demonstrated that calcium phosphate particles play an important role in the formation of hard tissues in nature. In this paper, current concepts in biomineralization, such as nano assembly, biomimetic shell structure, and their applications are introduced. It is confirmed experimentally that enamel- or bone-liked apatite can be achieved by oriented aggregations using nano calcium phosphates as starting materials. The assembly of calcium phosphate can be either promoted or inhibited by different biomolecules so that the kinetics can be regulated biologically. In this paper, the role of nano calcium phosphate in tissue repair is highlighted. Furthermore, a new, interesting result on biomimetic mineralization is introduced, which can offer an artificial shell for living cells via a biomimetic method.
Keywords: biomineralization; calcium phosphate; tissue repair

Usage of polymer brushes as substrates of bone cells by Sabine A. Letsche; Annina M. Steinbach; Manuela Pluntke; Othmar Marti; Anita Ignatius; Dirk Volkmer (132-144).
Implant medical research and tissue engineering both target the design of novel biomaterials for the improvement of human health and clinical applications. In order to develop improved surface coatings for hard tissue (bone) replacement materials and implant devices, we are developing micropatterned coatings consisting of polymer brushes. These are used as organic templates for the mineralization of calcium phosphate in order to improve adhesion of bone cells. First, we give a short account of the current state-of-the-art in this particular field of biomaterial development, while in the second part the preliminary results of cell culture experiments are presented, in which the biocompatibility of polymer brushes are tested on human mesenchymal stem cells.
Keywords: polymer brush; ATRP; micropatterning; bone cell; cell adhesion

Hydroxyapatite-alginate biocomposite promotes bone mineralization in different length scales in vivo by F. L. De Paula; I. C. Barreto; M. H. Rocha-Leão; R. Borojevic; A. M. Rossi; F. P. Rosa; M. Farina (145-153).
Tissue engineering is a multidisciplinary research area that aims to develop new techniques and/or biomaterials for medical applications. The objective of the present study was to evaluate the osteogenic potential of a composite of hydroxyapatite and alginate in bone defects with critical sizes, surgically made in the calvaria region of rats. The rats (48 adult males), Rattus norvegicus Wistar, were divided into two groups: control (without composite implantation) and experimental (with composite implantation) and analyzed by optical microscopy at the biological time points 15, 45, 90 and 120 d, and transmission electron microscopy 120 d after the implantation of the biomaterial. It was observed that the biomaterial presented a high degree of fragmentation since the first experimental points studied, and that the fragments were surrounded by new bone after the duration of the project. These areas were studied by analytical transmission electron microscopy using an energy dispersive X-ray spectrometer. Three regions could be distinguished: (1) the biomaterial rich in hydroxyapatite; (2) a thin contiguous region containing phosphorus but without calcium; (3) a region of initial ossification containing mineralizing collagen fibrils with a calcium/phosphorus ratio smaller than the particles of the composite. The intermediate region (without calcium or containing very low amounts of calcium), which just surrounded the composite had not been described in the literature yet, and is probably associated specifically to the biocomposite used. The high performance of the biomaterial observed may be related to the fact that alginate molecules form highly anionic complexes and are capable of adsorbing important factors recognized by integrins from osteoblasts. Regions of fibrotic tissue were also observed mainly in the initial experimental points analyzed. However, it did not significantly influence the final result. In conclusion, the biomaterial presents a great potential for application as bone grafts in the clinical area.
Keywords: bone engineering; bone healing; hydroxyapatite; alginate; biocomposite; analytical microscopy

Biomimetic mineral coatings in dental and orthopaedic implantology by Yue-lian Liu; Klaas de Groot; Ernst B. Hunziker (154-162).
Biomimetic techniques are used to deposit coatings of calcium phosphate upon medical devices. The procedure is conducted under near-physiological, or “biomimetic”, conditions of temperature and pH primarily to improve their biocompatibility and biodegradability of the materials. The inorganic layers generated by biomimetic methods resemble bone mineral, and can be degraded within a biological milieu.The biomimetic coating technique involves the nucleation and growth of bone-like crystals upon a pretreated substrate by immersing this in a supersaturated solution of calcium phosphate under physiological conditions of temperature (37°C) and pH (7.4). The method, originally developed by Kokubo in 1990, has since undergone improvement and refinement by several groups of investigators.Biomimetic coatings are valuable in that they can serve as a vehicle for the slow and sustained release of osteogenic agents at the site of implantation. This attribute is rendered possible by the near-physiological conditions under which these coatings are prepared, which permits an incorporation of bioactive agents into the inorganic crystal latticework rather than their mere superficial adsorption onto preformed layers. In addition, the biomimetic coating technique can be applied to implants of an organic as well as of an inorganic nature and to those with irregular surface geometries, which is not possible using conventional methodologies.
Keywords: biomimetic; biomineralization; osteoinduction; osteogenic agent; tissue engineering

The majority of the mineral phase of the Lytechinus variegatus tooth is comprised of magnesium containing calcite crystal elements, collectively arranged so that they appear as a single crystal under polarized light, as well as under X-ray or electron irradiation. However, the crystal elements are small, and in spite of the common alignment of their crystal axes, are not the same size or shape in different parts of the tooth. The toughness of the tooth structure arises from the fact that it is a composite in which the crystals are coated with surface layers of organic matter that probably act to inhibit crack formation and elongation. In the growth region the organic components represent a greater part of the tooth structure. In the most heavily mineralized adoral region the primary plates fuse with inter-plate pillars. Using Scanning Electron Microscopy; TOF-SIMS mapping of the characteristic amino acids of the mineral related proteins; and isolation and characterization of the mineral-protected protein we report that the late-forming inter-plate pillars had more than a three-fold greater Mg content than the primary plates. Furthermore, the aspartic acid content of the mineralrelated protein was highest in the high Mg pillars whereas the mineral-protected protein of the primary plates was richer in glutamic acid content.These results suggest that the Asp-rich protein(s) is important for formation of the late developing inter-plate pillars that fuse the primary plates and increase the stiffness of the most mature tooth segment. Supported by NIDCR Grant DE R01-01374 to AV.
Keywords: sea urchin tooth; mineral-related proteins; high magnesium calcite; TOF-SIMS; SEM

Proliferation of osteoblast cells on nanotubes by F. Watari; T. Akasaka; Xiaoming Li; M. Uo; A. Yokoyama (169-173).
Carbon nanotubes (CNT) have a unique structure and feature. In the present study, cell proliferation was performed on the scaffolds of single-walled CNTs (SWCNT), multiwalled CNTs (MWCNT), and on graphite, one of the representative isomorphs of pure carbon, for the sake of comparison. Scanning electron microscopy observation of the growth of osteoblast-like cells (Saos2) cultured on CNTs showed the morphology fully developed for the whole direction, which is different from that extended to one direction on the usual scaffold. Numerous filopodia were grown from cell edge, extended far long and combined with the CNT meshwork. CNTs showed the affinity for collagen and proteins. Proliferated cell numbers are largest on SWCNTs, followed by MWCNTs, and are very low on graphite. This is in good agreement with the sequence in the results of the adsorbed amount of proteins and expression of alkaline phosphatase activity for these scaffolds. The adsorption of proteins would be one of the most influential factors to make a contrast difference in cell attachment and proliferation between graphite and CNTs, both of which are isomorphs of carbon and composed of similar graphene sheet crystal structure. In addition, the nanosize meshwork structure with large porosity is another property responsible for the excellent cell adhesion and growth on CNTs. CNTs could be the favorable materials for biomedical applications.
Keywords: carbon nanotube; scaffold; osteoblast; regeneration; nanomaterial

Tooth enamel and enameloid in actinopterygian fish by I. Sasagawa; M. Ishiyama; H. Yokosuka; M. Mikami; T. Uchida (174-182).
The morphological features of tooth enamel and enameloid in actinopterygian fish are reviewed to provide basic data concerning the biomineralization of teeth in lower vertebrates. Enameloid, which covers the tooth surface, is a unique well-mineralized tissue and usually has the same functions as mammalian tooth enamel. However, the development of enameloid is different from that of the enamel produced by dental epithelial cells. Enameloid is made by a combination of odontoblasts and dental epithelial cells. An organic matrix that contains collagen is provided by odontoblasts, and then dental epithelial cells dissolve the degenerate matrix and supply inorganic ions during advanced crystal growth in enameloid. It is likely that enameloid is a good model for studying the growth of well-mineralized hard tissues in vertebrates. Some actinopterygian fish possess a collar enamel layer that is situated at the surface of the tooth shaft, indicating that the origin of tooth enamel is found in fish. Collar enamel is thought to be a precursor of mammalian enamel, although it is thin and not well mineralized in comparison with enameloid. In Lepisosteus and Polypterus, both of which are living actinopterygians, both enameloid and enamel are found in the same tooth. Therefore, they are suitable materials for examining the developmental processes of enameloid and enamel and the relationship among them.
Keywords: actinopterygian; enamel; enameloid; fish; tooth

CAP-1 is a cuticle peptide isolated from the acid-insoluble fraction of the exoskeleton of the crayfish Procambarus clarkii. CAP-1 is an acidic peptide and comprises 78 amino acid residues. The C-terminal part is especially highly acidic by a phosphoserine and an Asprepeat, which is thought to be responsible for the calcification inhibitory activity in vitro. To examine the significance of the Asp-repeat and to get information on structure-activity relationship, various small related peptides with different sequences were synthesized and tested for the inhibitory activity. The results showed that 1) the activity depends not on the Asp-containing sequence but on the total number of Asp residues, 2) peptide conformation does not affect the activity, and 3) Asp is more effective in inhibitory activity than Glu. These characteristics seem to be consistent with the fact that acidic matrix proteins identified so far from various biominerals have almost no sequence similarity, leading to the idea that the molecular evolution of matrix proteins and peptides in biominerals might be intrinsically different from that of enzymes, hormones and other important functional proteins possibly due to the difference in the mode of interaction between proteins and inorganic compounds, or between proteins and organic compounds.
Keywords: CAP-1; peptide; Procambarus clarkii ; inhibitory activity; biomineral

Tentative application of computed tomography to study of otoliths and their responses to environment variations by Li-na Yan; Sheng-rong Li; Jun-yan Luo; Feng-qin Du; Guo-lin Ma; Yu-li Wang; Wu Wang (187-193).
As the carrier of biomineral aragonite, fish otoliths memorize various messages of environment throughout the fish’s life. In the past three decades, quite a few achievements have been made in the studies of fish otoliths, but no advances in research using medical instruments have been reported. The authors tentatively applied X-ray computed tomography (CT) to the studies of the internal structure of wild carp otoliths, with the CT value determined by variations in the sample density and element composition. The wild carps were collected, respectively, from the Baiyangdian Shallow Lake in Hebei Province and Miyun Reservoir in the Beijing metropolis, whose water environments are quite different. The former has suffered serious pollution and eutrophication, whereas the latter has nearly experienced no pollution. The primary result indicates that differences exists in CT values of otoliths for the carps from the above-mentioned waters. With in-depth studies, it is possible that X-ray computed tomography could serve as a useful tool in the study of fish otoliths, and the CT values can be taken as typomorphic parameters to distinguish the waters with different degrees of pollution.
Keywords: carp otoliths; X-ray computed tomography; water environment; Baiyangdian Shallow Lake; Miyun Reservoir

Polypyrrole/multiwalled carbon nanotube (MWNT) composite films were electrochemically deposited in the presence of an ionic surfactant, sodium dodecyl sulfate (SDS), acting as both supporting electrolyte and dispersant. The effects of the surfactant and the MWNT concentrations on the structure of the resulting composite films were investigated. The electrochemical behavior of the resulting polypyrrole/MWNT composite film was investigated as well by cyclic voltammogram. The effect of the additional alternating electric field applied during the constant direct potential electrochemical deposition on the morphology and electrochemical behavior of the resulting composite film was also investigated in this study.
Keywords: carbon nanotubes; conducting polymers; electrochemical polymerization; electrochemical capacitors; nanocomposites; surfactant

The Tersoff-potential based MD (molecular dynamics) method was used to simulate the radial compression of one (10,0) BN nanotube, and its compressive properties was compared with those of one (10,0) carbon nanotube. The semi-empirical PM3 QC (Quantum chemistry) method was adopted to calculate the electronic structures of the compressed BN-tube, and the effect of the radial compression on the electronic structures of the BN-tube was discussed. It is shown that (i) BN-tube has comparable radial compressive stiffness to carbontube, but lower energy-absorbing, load-support and deformation-support capabilities, and (ii) with the increase of compressive strain, the HOMO energy of the BN-tube increases, the LUMO energy and the LUMO-HOMO energy-gap decrease, and its chemical activity and conductance increase.
Keywords: BN nanotube; radial compression; compressive properties; electronic structures

The simplified one-dimensional dislocation equation for mixed dislocations is derived briefly from the two-dimensional modified Peierls-Nabarro equation taking into account the discreteness effect of crystals. The collinear dissociated core structure of 〈111〉 {110} superdislocations in the novel B2 structure YAg and YCu are investigated with the simplified equation. Both the core width and the dissociated width are increasing with the increases in the dislocation angle of superdislocations. The dissociated width determined by continuum elastic theory is inaccurate for the high antiphase boundary energy but is recovered for the low antiphase boundary energy. The Peierls stress of the dissociated dislocation is replaced by that of superpartials. The results show that both the unstable stacking fault energy and the core width are crucial for the Peierls stress in the case of a narrow core structure. However, the core width becomes the main factor in controlling the Peierls stress in the case of a wide core.
Keywords: core structure; dissociation width; variational method; Peierls stress

Effects of process parameters on arc shape and penetration in twin-wire indirect arc welding by Shun-shan Zhang; Mei-qing Cao; Dong-ting Wu; Zeng-da Zou (212-217).
In this study, the effects of variable parameters on arc shape and depth of penetration in twin-wire indirect arc gas shielded welding were investigated. The variation of arc shape caused by changes of the parameters was recorded by a high-speed camera, and the depths of penetration of specimen were measured after bead welding by an optical microscope. Experiments indicated that proper parameters give birth to a concentrated and compressed welding arc, which would increase the depth of penetration as the incensement of the arc force. Several principal parameters including the distance of twin wires intersecting point to base metal, the included angle, and the content of shielding gas were determined. The arc turned more concentrated and the depth of penetration increased obviously as the welding current increased, the arc turned brighter while unobvious change of penetration occurred as the arc voltage increased, and the deepest penetration was obtained when the welding speed was 10.5 mm/s.
Keywords: indirect arc; twin-wire welding; penetration; arc shape

Fracture behavior of HPHT synthetic diamond with micrometers metallic inclusions by He-sheng Li; Yong-xin Qi; Yuan-pei Zhang; Mu-sen Li (218-223).
The fracture behavior of the diamond single crystals with metallic inclusions was investigated in the present paper. Single diamond crystals with metallic inclusions were formed by a special process with high pressure and high temperature (HPHT). The inclusions trapped in the diamond were characterized mainly to be metallic carbide of (Fe,Ni)23C6 or Fe3C and solid solution of γ-(Fe,Ni) by transmission electronic microscopy (TEM). The grain size of the inclusions is about micrometers. The fracture characteristics of the diamond single crystals, after compression and heating, were investigated by optical microscopy (OM) and scanning electron microscopy (SEM). The fracture sections of the compressed and heated diamonds were found to be parallel to the (111) plane. The interface of the inclusions and diamond is deduced to be the key factor and the original region of the fracture formation. Mechanisms of the fracture behavior of the HPHT synthesized diamonds are discussed.
Keywords: synthetic diamond; metallic inclusions; fracture; microstructure