1. Introduction to the structure and metabolism of dental tissue
2. Substitution of apatite structures
3. Demineralization – tooth decay
Introduction to the structure and metabolism of dental tissue
Teeth are phylogenetically old structures composed of hard tissues. Their function includes grasping, cutting and grinding the consumed food. Human teeth are known to be heterodontous – meaning that they have different shapes and functional specializations.
Tooth – dens (gr. odus, gen. odontos) is made of several parts:
1) Crown – corona dentis
2) Neck – collum dentis
3) Root – radix dentis
Crown sticks out of gum and is covered by a layer of enamel (enamelum). The enamel is made of cells called ameloblasts and is the hardest tissue found in the human body. The inner structure consists of columns called enamel prisms. Dentin (dentinum) represents the main substance of teeth. Though it contains high percentage of inorganic material making it harder than bone, it is still elastic. Dentin is created by cells known as odontoblasts. Cementum is a part of the tooth complex, covering the neck and the root of teeth. It is actually a fibrous bone poor in bone cells (cementocytes and cementoblasts) that is connected to the surface of dentin via collagen fibers.
Chemical composition of human teeth
From the chemical point of view, the components of teeth can be divided into organic and inorganic. The content of organic substances varies during the development of tooth. Immature enamel consists of 50 % of organic material and 50 % of mineral components. Compared to it, in the adult enamel, the mineral components make up as much as 96 % of the material; 0.5 % is organic material and the rest (approximately 3.5 %) is watery substance. Dentin contains more organic material (20-25 %), mineral substances make up around 60-65 % of its content.
Among the low molecular weight organic substances found in teeth are organic acids, lipid components and monosaccharides. The most abundant organic acids are citric, lactic and pyruvic acid. They are the products of teeth tissue cells metabolism. Citrate, the product of the citrate cycle, can be found in teeth tissue in complexes with calcium ions. Lactate and pyruvate are created in the glycolysis pathway. At the end of the teeth development, the organic substances become a part of the mineral structures of teeth. Inorganic substances are represented mostly by compounds of calcium, phosphorous, carbonates, magnesium, fluorine and sodium. These are the basic building blocks of the apatite structures.
Human enamel is a unique mineralized complex consisting of hydroxyapatite (92-94 %), water (2-3 %), carbonate (2 %), trace elements (sodium, magnesium, potassium, chloride, zinc; 1 %), fluoride (0.01-0.05 %), proteins and lipids (composition of enamel depends on many factors – diet, age and pathological states of hard teeth tissues. So far, at least 40 trace elements have been identified in the human enamel. Some get into the oral cavity during stomatological procedures, other (like strontium) can act as indicators of pollution in the environment. Furthermore, different places on the same teeth show differences in the enamel composition caused by fluctuations in the concentrations of individual elements. From the surface towards the dentin-enamel boundary, the concentrations of fluorine, iron, tin, chlorine, calcium, carbonates, magnesium and sodium decreases.
Varying ratios of calcium and phosphate give rise to small apatite crystals, they are however not always stoichiometric compounds according to the hydroxyapatite formula (see Table I). Due to lack of calcium, phosphates and hydroxyl ions and presence of carbonates and bicarbonate, non-stoichiometric apatite crystals emerge. Fluorinated structures have more stable matrix than hydroxyapatite. Mineral component can incorporate carbonates as well, such apatites are, however, more prone to tooth decay compared to hydroxyapatite (see subchapter concerning substitutions).
Calcium-phosphate components found in enamel during demineralization and remineralisation.
|Hydroxyapatite (enamel, dentin, cementum): Ca/P = 1,61-1,64||(Ca, Mg, Na)10(PO4, HPO4, CO3)6 (OH)2|
|Hydroxyapatite (generally, including bone): Ca/P = 1,67||(Ca)10(PO4)6(OH)2|
|Fluor-hydroxyapatite: Ca/P = 1,67||(Ca)10(PO4)6(F, OH)2|
|Fluor-apatite: Ca/P = 1,67||(Ca)10(PO4)6F2|
|Carbonate apatite: Ca/P = 1,67||(Ca)10(PO4)6CO3|
|Carbonate hydroxyapatite: Ca/P = 1,7-2,4||(Ca, X)10(PO4, CO3)6(OH)2|
|Calcium deficient hydroxyapatite: Ca/P = less then 1,67||(Ca, X)10(HPO4, CO3)6(OH)2|
|Dicalcium phosphate dihydrate: Ca/P = 1,1||CaHPO4.2H2O|
|Tricalcium phosphate: Ca/P = 1,5||(Ca, Mg)3(PO4)2|
|Octacalcium phosphate pentahydrate: Ca/P = 1,3||Ca8H2(PO4)6.5H2O|
|Amorphous calcium phosphate: Ca/P = 1,1-3||(CaC)x(PO4,Y)y|
X = calcium can be substituted by Mg, Na, Zn, Sr and others
Y = substitution for PO4
Water found in enamel has two forms. First is water bound in crystals in form of their hydration shell and second is free water mostly fixed to organic substances. Free water can evaporate when heated up. Water can be taken up by enamel from moisture environment, which can explain some physical phenomena during the formation and prevention of tooth decay. Enamel works as an imaginary molecular sieve, through which ions together with water phase can enter or leak.
Arrangement of hydroxyapatite crystals
Enamel is acellular tissue composed mostly of hydroxyapatite crystals. Crystals of apatite, up to 1 mm long, 50 nm wide and 25 nm thick, run from dentin to the surface of enamel. Enamel prisms are arranged in bundles containing approximately 1000 crystals. Cross-sectional profile of prisms differs in shape (circular or keyhole-shaped). Hydroxyapatite crystals are primarily arranged (according to their long axes (c-)) in parallel with long axes of prisms. On the periphery of each prism, crystals rather divert from their original orientation. Thus on a borderline between prisms intercrystalic spaces arise, which mediate the diffusion in the tissue. This is an important feature in the creation of the tooth decay. Interprismatic crystals can exists as independent structures, it is however quite difficult to distinguish the ends of the adjacent prisms. Density profile of crystals and prisms in the enamel depends on the content of mineral compounds and is not uniform. Generally, the density decreases in the direction from surface towards dentin. On the contrary the porosity and amount of water substances and organic material increases in the same direction. The knowledge of hydroxyapatite structure is important, because it makes it possible to determine how the hard tooth tissue would behave, should it undergo a process of demineralization – dissolution.
The mineral component of enamel is basically a substituted calcium hydroxyapatite. The stoichiometric structure of hydroxyapatite can be easily imagined by a consideration of the arrangement of ions around the central hydroxyl column, which extends in the c-axis direction through the long axes of the crystals. In the plane of the diagram, the hydroxyl ion is enclosed in the triangle of calcium ions (Ca II.). This triangle is surrounded on one side by a triangle of phosphate ions shifted by 60 degrees. The triangles are surrounded by a hexagon made of calcium ions (Ca I.). The whole crystalline structure can be shown as a row of hexagonal prisms stacked one on the other, each rotated 60° compared to the previous one.
Enamel apatite and naturally all other mineralized tissues can show profound differences in their composition. These include missing ions – mostly calcium and hydroxyl, less often carbonate, fluoride, sodium or magnesium. Changes and substitutions have considerable influence over the behavior of apatite, mainly with respect to its solubility at low pH.
Composition of dentin and cementum
Dentin consists mainly of inorganic substances (70 %). Other components include organic substances (20 %) and water (10 %). Inorganic constituent is made of calcium hydroxyapatite crystals. Crystals are smaller than the ones found in enamel and they are located on and between the collagen fibers.
The main inorganic component of dentin is hydrated calcium phosphate or calcium hydroxyapatite. Its structure and composition is close to the one of apatite group minerals found in earth’s crust. Due to a small size of crystals (50 x 25 x 4 nm), a determination of an exact composition and crystalline structure of this biomineral is quite difficult. It mostly resembles the structure of hydroxyapatite, however it often contains elements and molecules other than calcium and phosphates.
Cementum is made of inorganic materials (65 %), organic substances (23 %) and water (12 %). The main inorganic component is hydroxyapatite. Its crystals are thin and arranged similarly to a fibular bone. Organic components are represented mostly by collagen of type I.
Substitution of apatite structures
Apatite, in comparison to other minerals, exhibits higher flexibility and is very prone to ion exchanges. Substitution changes the composition of mineral and often has decisive influence on its characteristics like solubility, hardness, fragility, tension, temperature stability etc.. Some ion exchanges in the crystalline structure of hydroxyapatite significantly affect the size of the crystals and solubility. Limited number of elements found in human body restricts the amount of substitutions in apatite, which is less than for apatite occurring in nature.
Structure of apatite group of minerals has four crystallographic positions:
1) Tetrahedral position for six P5+ ions, every ion makes 4 bonds with oxygen
2) Ca I. position for 4 Ca2+ ions
3) Ca II. position for 6 other Ca2+ ions (arranged in a way that they create a channel along the c- axis)
4) Position in channel, which is occupied by up to 2 monovalent anions (mostly OH–, F–, rarely Cl– ions)
Within the structure, ions or ion groups can substitute to different positions.
Inclusion of ions to the enamel apatite
The position of OH– group in the structure of hydroxyapatite, (Ca)10(PO4)6(OH)2,can be partially of fully occupied by fluorine. In this way crystals of fluorohydroxyapatite and fluorapatite are formed. Other elements from the halide group (like chlorine, bromine or iodine) can be incorporated to the structure; there is however no commonly occurring mineral with fully substituted OH– group by any of these atoms.
Fluorine forms insoluble salts with more elements. They can either be simple with Ca, Mg, Fe or complex salts with Ca and P. Bones and teeth contain resistant crystals of fluorapatite. Fluorine is of crucial significance for the growth of teeth. It inhibits bacterial growth and modulates metabolism in enamel. It is therefore used in prophylactic teeth supplements.
Fluorine is resorbed in gastrointestinal tract, with significant proportion being resorbed in stomach and smallintestine. More than 90 % of ingested fluorine is resorbed. It is assumed that the absorption takes place via a passivediffusion and is inversely related to the proton gradient (pH). Therefore factors stimulating the secretion of gastric juices increase the absorption. Stomach resorbs hydrofluoric acid, whose amount is small in the small intestine due to a higher pH. On the contrary, concentration and a gradient of fluoride ions in the small intestine are high. Soluble fluorides like sodium fluoride or tin (II) fluoride are components of many types of toothpastes and are easily absorbed. Fast absorption leads to an increase in fluoride concentration in blood plasma, where they exist in the form of fluoride ions. Eliminationoffluorideions from the bloodstream takes place via two mechanisms – they get removed through urine or are taken up by calcified tissues, mostly teeth.
Incorporation of fluorides into the hydroxyapatite structure happens in two ways:
1) Fluoride ion fills in the gap after a missing OH– group in c-axis of crystal column
2) Fluoride displaces OH– ion from its position
High charge density of fluoride ions and symmetry leads to much closer arrangement of fluorides in the triangle near the Ca II position. This causes a decrease in energy barrier and effective stabilizationof crystalline structure. The result is more resistant, less soluble apatite (or fluorapatite) structure. Migration of fluorine within the structure plays an important role in the prevention and prophylaxis of tooth decay.
Distribution of fluoride in enamel is not homogenous – it concentrates only on surface and markedly decreases in abundance in the direction towards dentin. Surface possesses high concentrations of fluoride ions, probably related to their uptake from blood plasma during the time before tooth eruption. This process concentrates fluoride ions on the enamel surface and promotes the formation of a more stable form of fluorhydroxyapatite (compared to hydroxyapatite), which uptakes the fluoride ions more effectively and limits their passage to deeper layers of hard teeth tissues. Fluorohydroxyapatite or fluorapatite are not the only fluoridated forms found on the surface of enamel. Among the others are unspecified form of fluoride-phosphate complexes, calcium fluoride and hydrogenated calcium fluoride bound to complexes.
Other chemical substance commonly found in apatite is carbonate (CO32-). Its ions have different charge and molecular size than the dominant PO43- group. Carbonate is a trigonal planar ion, 0.2 nm in diameter, which gets into the crystalline structure only with difficulties. The mechanism of incorporation of carbonate to the apatite lattice is not fully understood. In principle, the carbonate ions are able to substitute the apatite structure either on position of OH– group, (Ca)10(PO4)6CO3 (the so called A-type substitution) or on PO43- group position, (Ca)10(PO4, CO3)6(OH)2 (the B-type substitution). Which of these substitutions would occur depends on pCO2 (CO2 partial pressure) during the growth of the crystal. Concentration increases in the direction from enamel (2 %) to dentin (4-6 %).
A general assumption is that the CO32- ions preferably substitute the PO43- group. The B-type substitution cause changes in various physical characteristics of hydroxyapatite – shortening of a- axis length, overall reduction is size of the crystal, extension of c- axis, decrease in thermal stability, increase in strain, solubility and optical birefringence. The increase in the solubility of apatites saturated with carbonate is caused by lower strength of Ca-CO32- bond, compared to Ca-PO43- bond of carbonate-free apatite. B-type apatite enriched with carbonate (similarly to HPO42- enriched apatite) can contain different amounts of carbonates and phosphates. Negative ion deficit, caused by the change of PO43- for either CO32- or HPO42- can be resolved by a loss of positive ion – or by the exclusion of Ca2+ from the crystal matrix. As a consequence of balancing the charge in carbonate-rich-apatite, the concentration of OH– in the matrix can be limited. The above-mentioned substitution results in the formation of less resistant apatite structures – the so-called “soft forms”, which are more soluble in acidic environment and thus more prone to various pathological changes.
A number of bivalent cations have the ability to substitute different Ca2+ positions in the crystalline matrix. Positions of cations can also be occupied by rare metal elements – their ions can substitute the apatite depending on their charge, ion radius and coordination number. The following general formula of apatite summarizes the substitutions:
(Ca, Mg, Na, Sr, Se, Zn, Pb,…)10(PO4, CO3, HPO4)6(OH, F)2.
Calcium can be replaced to some extent by magnesium, although this is thought to be limited, possibly to about 0.3 %. Magnesium is thought to be located on crystal surfaces or in separate, more vulnerable acid-soluble, phases as will be described below. The density charge of magnesium has destabilizing effect on the apatite structure, similar to the effect of carbonate substitution. Carbonate and magnesium have synergic effect in their incorporation into the hydroxyapatite matrix. They both contribute to its sensitivity towards acidic environment. Magnesium with carbonate are both concentrated on the surface or on the borderline between the prism and can be recrystallized during the development of hard teeth tissues: CaMg(CO3)2 and Ca9Mg(PO4)6(HPO4).
Tooth decay (caries) is among the most abundant pathological changes of hard teeth tissues. Tooth decay is a multicausal and multi-conditional disease. It forms on places, where optimal conditions arise. Participating factors are acids dissolving the mineral components of teeth by neutralization of negatively charged opposing ions in apatite. Acids come from food or are formed by microorganisms found on the teeth surfaces (under the protective layer of dextran). During the anaerobic catabolism of sugars by these microorganisms, mainly lactic acid is formed or (in lower quantities) other acids – propionic, acetic or butyric.
Exposing the hydroxyapatite structure to acids leads to an increase in the concentration of calcium, magnesium, phosphate, hydrogen phosphate, carbonate and bicarbonate in the microscopic tear of the beginning tooth decay. In the presence of increased amount of acids in the oral cavity (pH 5.5), dicalcium phosphate dihydrate and octacalcium phosphate form directly form the ions released during the dissolution of hydroxyapatite. When the effect of acids diminishes, neutral pH is gradually reached. Dicalcium phosphate dihydrate hydrolyzes to octacalcium phosphate in the presence of calcium, bicarbonate and carbonate ions, all coming from the dissolved hydroxyapatite, dental plaque or saliva. Both apatite forms (dicalcium phosphate dihydrate and octacalcium phosphate) can form apatite or carbonate hydroxyapatite under such conditions. In the presence of magnesium ion (coming from dissolved hydroxyapatite, dental plaque, saliva, food or drinks), the magnesium immediately after the dissolution of hydroxyapatite or dicalcium phosphate dihydrate substitutes the calcium ion in the tricalcium phosphate thus inhibiting the formation of octacalcium phosphate. The result is the inability of dicalcium phosphate dihydrate and octacalcium phosphate to form hydroxyapatite. Low concentration of fluoride ions (normally acquired during the hygiene of the oral cavity form toothpaste or mouthwash) reacts with the dissolution products of hydroxyapatite and forms fluoroapatite or fuorohydroxyapatite. These fluoridated structures of apatite inhibit the dicalcium phosphate hydrate and octacalcium phosphate structures.
High concentration of fluoride ions found in many fluoride gels (containing calcium fluoride) leads to a recovery of hydroxyapatite or fluorhydroxyapatite. Calcium fluoride can act as a reservoir of calcium ions, supplying the hydroxyapatite in the enamel during the attacks of acids. With the help of two fluoride ions it stops further dissolution of hydroxyapatite structure – fluorhydroxyapatite forms. Calcium fluoride can hydrolyzes to fluorhydroxyapatite, but only in the presence of hydrogenphosphate or phosphate ions.
In general, the presence of fluoride ions suppresses the acidic environment, makes calcium phosphate in dicalcium phosphate dihydrate and octacalcium phosphate less soluble and facilitates the formation of fluorhydroxyapatite.
The process of demineralization of crystalline structure of hydroxyapatite or other apatite structures begins with dissolution of a central core and proceeds towards the periphery. The center (so-called central core of the crystal) is affected first, due to a higher concentration of carbonates and imperfections in the symmetry of the crystal lattice. The origin of structure disruption can be noticed by a fine cut found on the terminal end of the hydroxyapatite crystal. Dissolution continues in the direction from the central core, causing a deepening of the tear in the crystal. If the pathological process is not soon stopped it causes total destruction of hard teeth tissues. Under the laboratory conditions, the core can be dissolved by lactic acid in a few minutes. Peripheral shell is able to withstand a complete dissolution in order of hours.
Subchapter Author: Václav Pavlíček