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Mineralized tissues are biological tissues that incorporate minerals into soft matrices. Typically these tissues form a protective shield or structural support. Bone, mollusc shells, deep sea sponge ''Euplectella'' species, radiolarians,
diatoms A diatom (Neo-Latin ''diatoma''), "a cutting through, a severance", from el, διάτομος, diátomos, "cut in half, divided equally" from el, διατέμνω, diatémno, "to cut in twain". is any member of a large group comprising sev ...
,
antler Antlers are extensions of an animal's skull found in members of the Cervidae (deer) family. Antlers are a single structure composed of bone, cartilage, fibrous tissue, skin, nerves, and blood vessels. They are generally found only on ...
bone,
tendon A tendon or sinew is a tough, high-tensile-strength band of dense fibrous connective tissue that connects muscle to bone. It is able to transmit the mechanical forces of muscle contraction to the skeletal system without sacrificing its ability ...
,
cartilage Cartilage is a resilient and smooth type of connective tissue. In tetrapods, it covers and protects the ends of long bones at the joints as articular cartilage, and is a structural component of many body parts including the rib cage, the neck ...
,
tooth enamel Tooth enamel is one of the four major tissues that make up the tooth in humans and many other animals, including some species of fish. It makes up the normally visible part of the tooth, covering the crown. The other major tissues are dentin, ...
and
dentin Dentin () (American English) or dentine ( or ) (British English) ( la, substantia eburnea) is a calcified tissue of the body and, along with enamel, cementum, and pulp, is one of the four major components of teeth. It is usually covered by e ...
are some examples of mineralized tissues. These tissues have been finely tuned to enhance their mechanical capabilities over millions of years of evolution. Thus, mineralized tissues have been the subject of many studies since there is a lot to learn from nature as seen from the growing field of
biomimetics Biomimetics or biomimicry is the emulation of the models, systems, and elements of nature for the purpose of solving complex human problems. The terms "biomimetics" and "biomimicry" are derived from grc, βίος (''bios''), life, and μίμησ ...
. The remarkable structural organization and engineering properties makes these tissues desirable candidates for duplication by artificial means. Mineralized tissues inspire miniaturization, adaptability and multifunctionality. While natural materials are made up of a limited number of components, a larger variety of material chemistries can be used to simulate the same properties in engineering applications. However, the success of biomimetics lies in fully grasping the performance and mechanics of these biological hard tissues before swapping the natural components with artificial materials for engineering design. Mineralized tissues combine stiffness, low weight, strength and toughness due to the presence of minerals (the inorganic part) in soft protein networks and tissues (the
organic Organic may refer to: * Organic, of or relating to an organism, a living entity * Organic, of or relating to an anatomical organ Chemistry * Organic matter, matter that has come from a once-living organism, is capable of decay or is the product ...
part). There are approximately 60 different minerals generated through biological processes, but the most common ones are calcium carbonate found in mollusk shells and
hydroxyapatite Hydroxyapatite, also called hydroxylapatite (HA), is a naturally occurring mineral form of calcium apatite with the formula Ca5(PO4)3(OH), but it is usually written Ca10(PO4)6(OH)2 to denote that the crystal unit cell comprises two entities. ...
present in teeth and bones. Although one might think that the mineral content of these tissues can make them fragile, studies have shown that mineralized tissues are 1,000 to 10,000 times tougher than the minerals they contain. The secret to this underlying strength is in the organized layering of the tissue. Due to this layering, loads and stresses are transferred throughout several length-scales, from macro to micro to nano, which results in the dissipation of energy within the arrangement. These scales or hierarchical structures are therefore able to distribute damage and resist cracking. Two types of biological tissues have been the target of extensive investigation, namely
nacre Nacre ( , ), also known as mother of pearl, is an organicinorganic composite material produced by some molluscs as an inner shell layer; it is also the material of which pearls are composed. It is strong, resilient, and iridescent. Nacre is ...
from mollusk shells and bone, which are both high performance natural composites. Many mechanical and imaging techniques such as nanoindentation and atomic force microscopy are used to characterize these tissues. Although the degree of efficiency of biological hard tissues are yet unmatched by any man-made ceramic composites, some promising new techniques to synthesize them are currently under development. Not all mineralized tissues develop through normal physiologic processes and are beneficial to the organism. For example, kidney stones contain mineralized tissues that are developed through pathologic processes. Hence,
biomineralization Biomineralization, also written biomineralisation, is the process by which living organisms produce minerals, often to harden or stiffen existing tissues. Such tissues are called mineralized tissues. It is an extremely widespread phenomenon; ...
is an important process to understand how these diseases occur.


Evolution

The evolution of mineralized tissues has been puzzling for more than a century. It has been hypothesized that the first mechanism of animal tissue mineralization began either in the oral skeleton of
conodont Conodonts ( Greek ''kōnos'', " cone", + ''odont'', "tooth") are an extinct group of agnathan (jawless) vertebrates resembling eels, classified in the class Conodonta. For many years, they were known only from their tooth-like oral elements, whi ...
or the dermal skeleton of early agnathans. The dermal skeleton is just surface
dentin Dentin () (American English) or dentine ( or ) (British English) ( la, substantia eburnea) is a calcified tissue of the body and, along with enamel, cementum, and pulp, is one of the four major components of teeth. It is usually covered by e ...
and basal bone, which is sometimes overlaid by enameloid. It is thought that the dermal skeleton eventually became scales, which are homologous to teeth. Teeth were first seen in chondrichthyans and were made from all three components of the dermal skeleton, namely dentin, basal bone and enameloid. The mineralization mechanism of mammalian tissue was later elaborated in actinopterygians and sarcopterygians during bony fish evolution. It is expected that genetic analysis of agnathans will provide more insight into the evolution of mineralized tissues and clarify evidence from early fossil records.


Hierarchical structure

Hierarchical structures are distinct features seen throughout different length scales. To understand how the hierarchical structure of mineralized tissues contributes to their remarkable properties, those for nacre and bone are described below. Hierarchical structures are characteristic of biology and are seen in all structural materials in biology such as bone and nacre from seashells


Nacre

Nacre has several hierarchical structural levels.


The macroscale

Some mollusc shells protect themselves from predators by using a two layered system, one of which is nacre. Nacre constitutes the inner layer while the other, outer, layer is made from
calcite Calcite is a carbonate mineral and the most stable polymorph of calcium carbonate (CaCO3). It is a very common mineral, particularly as a component of limestone. Calcite defines hardness 3 on the Mohs scale of mineral hardness, based on scratc ...
. The latter is hard and thus prevents any penetration through the shell, but is subject to brittle failure. On the other hand, nacre is softer and can uphold inelastic deformations, which makes it tougher than the hard outer shell. The mineral found in nacre is aragonite, CaCO3, and it occupies 95% vol. Nacre is 3000 times tougher than aragonite and this has to do with the other component in nacre, the one that takes up 5% vol., which is the softer organic biopolymers. Furthermore, the nacreous layer also contains some strands of weaker material called growth lines that can deflect cracks.


The microscale

The Microscale can be imagined by a three-dimensional brick and mortar wall. The bricks would be 0.5 μm thick layers of microscopic aragonite polygonal tablets approximately 5-8 μm in diameter. What holds the bricks together are the mortars and in the case of nacre, it is the 20-30 nm organic material that plays this role. Even though these tablets are usually illustrated as flat sheets, different microscopy techniques have shown that they are wavy in nature with amplitudes as large as half of the tablet's thickness. This waviness plays an important role in the fracture of nacre as it will progressively lock the tablets when they are pulled apart and induce hardening.


The nanoscale

The 30 nm thick interface between the tablets that connects them together and the aragonite grains detected by scanning electron microscopy from which the tablets themselves are made of together represent another structural level. The organic material “gluing” the tablets together is made of proteins and
chitin Chitin ( C8 H13 O5 N)n ( ) is a long-chain polymer of ''N''-acetylglucosamine, an amide derivative of glucose. Chitin is probably the second most abundant polysaccharide in nature (behind only cellulose); an estimated 1 billion tons of chit ...
. To summarize, on the macroscale, the shell, its two layers (
nacre Nacre ( , ), also known as mother of pearl, is an organicinorganic composite material produced by some molluscs as an inner shell layer; it is also the material of which pearls are composed. It is strong, resilient, and iridescent. Nacre is ...
and
calcite Calcite is a carbonate mineral and the most stable polymorph of calcium carbonate (CaCO3). It is a very common mineral, particularly as a component of limestone. Calcite defines hardness 3 on the Mohs scale of mineral hardness, based on scratc ...
), and weaker strands inside nacre represent three hierarchical structures. On the microscale, the stacked tablet layers and the wavy interface between them are two other hierarchical structures. Lastly, on the nanoscale, the connecting organic material between the tablets as well as the grains from which they are made of is the final sixth hierarchical structure in nacre.


Bone

Like nacre and the other mineralized tissues,
bone A bone is a rigid organ that constitutes part of the skeleton in most vertebrate animals. Bones protect the various other organs of the body, produce red and white blood cells, store minerals, provide structure and support for the body, ...
has a hierarchical structure that is also formed by the self-assembly of smaller components. The mineral in bone (known as bone mineral) is
hydroxyapatite Hydroxyapatite, also called hydroxylapatite (HA), is a naturally occurring mineral form of calcium apatite with the formula Ca5(PO4)3(OH), but it is usually written Ca10(PO4)6(OH)2 to denote that the crystal unit cell comprises two entities. ...
with a lot of carbonate ions, while the organic portion is made mostly of
collagen Collagen () is the main structural protein in the extracellular matrix found in the body's various connective tissues. As the main component of connective tissue, it is the most abundant protein in mammals, making up from 25% to 35% of the whol ...
and some other proteins. The hierarchical structural of bone spans across to a three tiered hierarchy of the collagen molecule itself. Different sources report different numbers of hierarchical level in bone, which is a complex biological material. The types of mechanisms that operate at different structural length scales are yet to be properly defined. Five hierarchical structures of bone are presented below.


The macroscale

Compact bone and spongy bone are on a scale of several millimetres to 1 or more centimetres.


The microscale

There are two hierarchical structures on the microscale. The first, at a scale of 100 μm to 1 mm, is inside the compact bone where cylindrical units called osteons and small struts can be distinguished. The second hierarchical structure, the ultrastructure, at a scale of 5 to 10 μm, is the actual structure of the osteons and small struts.


The nanoscale

There are also two hierarchical structures on the nanoscale. The first being the structure inside the ultrastructure that are fibrils and extrafibrillar space, at a scale of several hundred nanometres. The second are the elementary components of mineralized tissues at a scale of tens of nanometres. The components are the mineral crystals of
hydroxyapatite Hydroxyapatite, also called hydroxylapatite (HA), is a naturally occurring mineral form of calcium apatite with the formula Ca5(PO4)3(OH), but it is usually written Ca10(PO4)6(OH)2 to denote that the crystal unit cell comprises two entities. ...
, cylindrical
collagen Collagen () is the main structural protein in the extracellular matrix found in the body's various connective tissues. As the main component of connective tissue, it is the most abundant protein in mammals, making up from 25% to 35% of the whol ...
molecules, organic molecules such as lipids and proteins, and finally water. The hierarchical structure common to all mineralized tissues is the key to their mechanical performance.


Mineral component

The mineral is the inorganic component of mineralized tissues. This constituent is what makes the tissues harder and stiffer.
Hydroxyapatite Hydroxyapatite, also called hydroxylapatite (HA), is a naturally occurring mineral form of calcium apatite with the formula Ca5(PO4)3(OH), but it is usually written Ca10(PO4)6(OH)2 to denote that the crystal unit cell comprises two entities. ...
, calcium carbonate,
silica Silicon dioxide, also known as silica, is an oxide of silicon with the chemical formula , most commonly found in nature as quartz and in various living organisms. In many parts of the world, silica is the major constituent of sand. Silica is ...
, calcium oxalate, whitlockite, and monosodium urate are examples of minerals found in biological tissues. In mollusc shells, these minerals are carried to the site of mineralization in vesicles within specialized cells. Although they are in an
amorphous In condensed matter physics and materials science, an amorphous solid (or non-crystalline solid, glassy solid) is a solid that lacks the long-range order that is characteristic of a crystal. Etymology The term comes from the Greek language, Gr ...
mineral phase while inside the
vesicles Vesicle may refer to: ; In cellular biology or chemistry * Vesicle (biology and chemistry), a supramolecular assembly of lipid molecules, like a cell membrane * Synaptic vesicle ; In human embryology * Vesicle (embryology), bulge-like features o ...
, the mineral destabilizes as it passes out of the cell and crystallizes. In bone, studies have shown that calcium phosphate nucleates within the hole area of the collagen fibrils and then grows in these zones until it occupies the maximum space.


Organic component

The organic part of mineralized tissues is made of proteins. In bone for example, the organic layer is the protein collagen. The degree of mineral in mineralized tissues varies and the organic component occupies a smaller volume as tissue
hardness In materials science, hardness (antonym: softness) is a measure of the resistance to localized plastic deformation induced by either mechanical indentation or abrasion. In general, different materials differ in their hardness; for example hard ...
increases. However, without this organic portion, the biological material would be brittle and break easily. Hence, the organic component of mineralized tissues increases their
toughness In materials science and metallurgy, toughness is the ability of a material to absorb energy and plastically deform without fracturing.nucleation or inhibition of hydroxyapatite formation. For example, the organic component in nacre is known to restrict the growth of aragonite. Some of the regulatory proteins in mineralized tissues are osteonectin, osteopontin, osteocalcin, bone sialoprotein and dentin phosphophoryn. In nacre, the organic component is porous, which allows the formation of mineral bridges responsible for the growth and order of the nacreous tablets.


Formation of minerals

Understanding the formation of biological tissues is inevitable in order to properly reconstruct them artificially. Even if questions remain in some aspects and the mechanism of mineralization of many mineralized tissues need yet to be determined, there are some ideas about those of mollusc shell, bone and sea urchin.


Mollusk shell

The main structural elements involved in the mollusk shell formation process are: a
hydrophobic In chemistry, hydrophobicity is the physical property of a molecule that is seemingly repelled from a mass of water (known as a hydrophobe). In contrast, hydrophiles are attracted to water. Hydrophobic molecules tend to be nonpolar and, ...
silk gel, aspartic acid rich protein, and a
chitin Chitin ( C8 H13 O5 N)n ( ) is a long-chain polymer of ''N''-acetylglucosamine, an amide derivative of glucose. Chitin is probably the second most abundant polysaccharide in nature (behind only cellulose); an estimated 1 billion tons of chit ...
support. The silk gel is part of the protein portion and is mainly composed of glycine and alanine. It is not an ordered structure. The acidic proteins play a role in the configuration of the sheets. The
chitin Chitin ( C8 H13 O5 N)n ( ) is a long-chain polymer of ''N''-acetylglucosamine, an amide derivative of glucose. Chitin is probably the second most abundant polysaccharide in nature (behind only cellulose); an estimated 1 billion tons of chit ...
is highly ordered and is the framework of the matrix. The main elements of the overall are: # The silk gel fills the matrix to be mineralized before the mineralization takes place. # The highly ordered
chitin Chitin ( C8 H13 O5 N)n ( ) is a long-chain polymer of ''N''-acetylglucosamine, an amide derivative of glucose. Chitin is probably the second most abundant polysaccharide in nature (behind only cellulose); an estimated 1 billion tons of chit ...
determines the orientation of the crystals. # The components of the matrix are spatially distinguishable. # Amorphous calcium carbonate is the first form of the mineral. # Once nucleation begins on the matrix, the calcium carbonate turns into crystals. # While crystals grow, some of the acidic proteins get trapped within them.


Bone

In bone, mineralization starts from a
heterogeneous Homogeneity and heterogeneity are concepts often used in the sciences and statistics relating to the uniformity of a substance or organism. A material or image that is homogeneous is uniform in composition or character (i.e. color, shape, siz ...
solution having calcium and phosphate ions. The mineral nucleates, inside the hole area of the collagen fibrils, as thin layers of calcium phosphate, which then grow to occupy the maximum space available there. The mechanisms of mineral deposition within the organic portion of the bone are still under investigation. Three possible suggestions are that nucleation is either due to the precipitation of calcium phosphate solution, caused by the removal of biological inhibitors or occurs because of the interaction of calcium-binding proteins.


Sea urchin embryo

The sea urchin embryo has been used extensively in developmental biology studies. The larvae form a sophisticated endoskeleton that is made of two
spicules Spicules are any of various small needle-like anatomical structures occurring in organisms Spicule may also refer to: *Spicule (sponge), small skeletal elements of sea sponges *Spicule (nematode), reproductive structures found in male nematodes ( ...
. Each of the spicules is a single crystal of mineral
calcite Calcite is a carbonate mineral and the most stable polymorph of calcium carbonate (CaCO3). It is a very common mineral, particularly as a component of limestone. Calcite defines hardness 3 on the Mohs scale of mineral hardness, based on scratc ...
. The latter is a result of the transformation of amorphous CaCO3 to a more stable form. Therefore, there are two mineral phases in larval spicule formation.


Organic-inorganic interface

The mineral-protein interface with its underlying adhesion forces is involved in the toughening properties of mineralized tissues. The interaction in the organic-inorganic interface is important to understand these toughening properties. At the interface, a very large force (>6-5 nN) is needed to pull the protein molecules away from the aragonite mineral in nacre, despite the fact that the molecular interactions are non-bonded. Some studies perform a finite element model analysis to investigate the behaviour of the interface. A model has shown that during tension, the back stress that is induced during the
plastic Plastics are a wide range of synthetic or semi-synthetic materials that use polymers as a main ingredient. Their plasticity makes it possible for plastics to be moulded, extruded or pressed into solid objects of various shapes. This adapta ...
stretch of the material plays a big role in the hardening of the mineralized tissue. As well, the nanoscale asperities that is on the tablet surfaces provide resistance to interlamellar sliding and so strengthen the material. A surface
topology In mathematics, topology (from the Greek words , and ) is concerned with the properties of a geometric object that are preserved under continuous deformations, such as stretching, twisting, crumpling, and bending; that is, without closing ...
study has shown that progressive tablet locking and hardening, which are needed for spreading large deformations over large volumes, occurred because of the waviness of the tablets.


Diseased mineralized tissues

In
vertebrates Vertebrates () comprise all animal taxa within the subphylum Vertebrata () (chordates with backbones), including all mammals, birds, reptiles, amphibians, and fish. Vertebrates represent the overwhelming majority of the phylum Chordata, with ...
, mineralized tissues not only develop through normal physiological processes, but can also be involved in pathological processes. Some diseased areas that include the appearance of mineralized tissues include atherosclerotic plaques, tumoral calcinosis, juvenile
dermatomyositis Dermatomyositis (DM) is a long-term inflammatory disorder which affects skin and the muscles. Its symptoms are generally a skin rash and worsening muscle weakness over time. These may occur suddenly or develop over months. Other symptoms may inc ...
,
kidney The kidneys are two reddish-brown bean-shaped organs found in vertebrates. They are located on the left and right in the retroperitoneal space, and in adult humans are about in length. They receive blood from the paired renal arteries; blo ...
and salivary stones. All physiologic deposits contain the mineral
hydroxyapatite Hydroxyapatite, also called hydroxylapatite (HA), is a naturally occurring mineral form of calcium apatite with the formula Ca5(PO4)3(OH), but it is usually written Ca10(PO4)6(OH)2 to denote that the crystal unit cell comprises two entities. ...
or one analogous to it. Imaging techniques such as
infrared spectroscopy Infrared spectroscopy (IR spectroscopy or vibrational spectroscopy) is the measurement of the interaction of infrared radiation with matter by absorption, emission, or reflection. It is used to study and identify chemical substances or functi ...
are used to provide information on the type of mineral phase and changes in mineral and matrix composition involved in the disease. Also, clastic cells are cells that cause mineralized tissue resorption. If there is an unbalance of clastic cell, this will disrupt resorptive activity and cause diseases. One of the studies involving mineralized tissues in dentistry is on the mineral phase of
dentin Dentin () (American English) or dentine ( or ) (British English) ( la, substantia eburnea) is a calcified tissue of the body and, along with enamel, cementum, and pulp, is one of the four major components of teeth. It is usually covered by e ...
in order to understand its alteration with aging. These alterations lead to “transparent” dentin, which is also called sclerotic. It was shown that a ‘‘dissolution and reprecipitation’’ mechanism reigns the formation of transparent dentin. The causes and cures of these conditions can possibly be found from further studies on the role of the mineralized tissues involved.


Bioinspired materials

The attractive properties of mineralized tissues like nacre and bone have led to the creation of a large number of biomimetic materials. Although improvements can be made, there are several techniques used to mimic these tissues. Some of the current techniques are described here for nacre imitation.


Large scale model materials

The large scale model of materials is based on the fact that crack deflection is an important toughening mechanism of nacre. This deflection happens because of the weak interfaces between the aragonite tiles. Systems on the macroscopic scales are used to imitate these week interfaces with layered composite ceramic tablets that are held together by weak interface “glue”. Hence, these large scale models can overcome the brittleness of ceramics. Since other mechanisms like tablet locking and damage spreading also play a role in the toughness of nacre, other models assemblies inspired by the waviness of microstructure of nacre have also been devised on the large scale.


Ice templation

Ice Templation is a new method that uses the physics of ice formation to develop a layered-hybrid material. In this system, ceramic particles in a concentrated suspension are frozen using carefully controlled freezing kinetics. As a result, a homogeneous, porous scaffold can be made, which is then filled with a second organic or inorganic phase to build dense layered composites.


Layer-by-layer deposition

Layer-by-layer deposition is a technique that as suggested by its name consists of a layer-by-layer assembly to make multilayered composites like nacre. Some examples of efforts in this direction include alternating layers of hard and soft components of TiN/Pt with an
ion beam An ion beam is a type of charged particle beam consisting of ions. Ion beams have many uses in electronics manufacturing (principally ion implantation) and other industries. A variety of ion beam sources exists, some derived from the mercu ...
system. The composites made by this sequential deposition technique do not have a segmented layered microstructure. Thus, sequential adsorption has been proposed to overcome this limitation and consists of repeatedly adsorbing electrolytes and rinsing the tablets, which results in multilayers.


Thin film deposition: microfabricated structures

Thin film deposition focuses on reproducing the cross-lamellar microstructure of conch instead of mimicking the layered structure of nacre using micro-electro mechanical systems (MEMS). Among mollusk shells, the
conch Conch () is a common name of a number of different medium-to-large-sized sea snails. Conch shells typically have a high spire and a noticeable siphonal canal (in other words, the shell comes to a noticeable point at both ends). In North Am ...
shell has the highest degree of structural organization. The mineral aragonite and organic matrix are replaced by
polysilicon Polycrystalline silicon, or multicrystalline silicon, also called polysilicon, poly-Si, or mc-Si, is a high purity, polycrystalline form of silicon, used as a raw material by the solar photovoltaic and electronics industry. Polysilicon is produce ...
and photoresist. The MEMS technology repeatedly deposits a thin silicon film. The interfaces are etched by reactive ion etching and then filled with photoresist. There are three films deposited consecutively. Although the MEMS technology is expensive and more time-consuming, there is a high degree of control over the morphology and large numbers of specimens can be made.


Self-assembly

The method of self-assembly tries to reproduce not only the properties, but also the processing of
bioceramics Bioceramics and bioglasses are ceramic materials that are biocompatible. Bioceramics are an important subset of biomaterials.J. F. Shackelford (editor)(1999) ''MSF bioceramics applications of ceramic and glass materials in medicine'' Bioceramics ...
. In this process, raw materials readily available in nature are used to achieve stringent control of nucleation and growth. This nucleation occurs on a synthetic surface with some success. The technique occurs at low temperature and in an aqueous environment. Self-assembling films form templates that effect the nucleation of ceramic phases. The downside with this technique is its inability to form a segmented layered microstructure. Segmentation is an important property of nacre used for crack deflection of the ceramic phase without fracturing it. As a consequence, this technique does not mimic microstructural characteristics of nacre beyond the layered organic/inorganic layered structure and requires further investigation.


The future

The various studies have increased progress towards understanding mineralized tissues. However, it is still unclear which micro/nanostructural features are essential to the material performance of these tissues. Also constitutive laws along various loading paths of the materials are currently unavailable. For nacre, the role of some nanograins and mineral bridges requires further studies to be fully defined. Successful biomimicking of mollusk shells will depend will on gaining further knowledge of all these factors, especially the selection of influential materials in the performance of mineralized tissues. Also the final technique used for artificial reproduction must be both cost effective and scalable industrially.


See also

* Shell growth in estuaries *
Biomineralization Biomineralization, also written biomineralisation, is the process by which living organisms produce minerals, often to harden or stiffen existing tissues. Such tissues are called mineralized tissues. It is an extremely widespread phenomenon; ...


References


Bibliography

* * * * * * * * * * * * * {{DEFAULTSORT:Mineralized Tissues Bone products Pedology Physiology Biomineralization