Saliva remineralizing ability –clinical determination of the remineralization rate of the enamel
In accordance with this methodology about remineralizing ability of saliva judge on its ability to "restore" the enamel of tooth, intentionally damaged by acid. Suppose that saliva will be gradual remineralize the damaged area of enamel,
Equipment:
Hydrochloric acid (HCl) 1 mole/l solution (for enamel etching);
methylene-blue 2% solution.
facilities for cleaning of enamel of teeth from dental deposits,
cotton roll for tooth isolation,
stop-watch
Procedure:
Clean, dry up and isolate a central incisor from saliva
Apply a drop of HCl 1-2 mm in diameter for 60 sec. in the middle of the tooth, 2 mm from the cutting edge.
Wash off solution, dry up an enamel, apply dye on a etched area.
After a 1 minute to take off dye by a cotton tampon,
Estimate intensity of colouring of etched area by comparing with a typographic 10-point scale.
Repeat determination of intensity of colouring every day (without etching!), wipe dye by tampon and determine the point of colouring zone.
Define, how many twenty-four hours does saliva need for the complete reconstruction of enamel. Oriented on a day, when the zone of damage was not painted.( at the next using of dye)
Estimate result
consider, when saliva has good mineralise properties, the CDRRE-test is 1-3 twenty-four hours, when saliva has bad mineralise properties, the CDRRE-test is more than 5 twenty-four hours
Determination of viscidity of saliva
is conducted by the viscometer of Oswald. Investigate 20-30 ml of saliva through 2-3 hours after eating. Cariessusceptible persons have more viscid saliva -6,0-9,0 9-10 units and cariesresistance persons have – only 1,0-4,0 4-5 units.
Determination of acidity of mixed saliva. in practice an indicatory method is comfortable: a standard indicatory paper strake is saturated with saliva, after determine its рН (compare the tint of strake to the standard scale of tints ).
The normal concentration of hydrogen ions in the range of рН = 6,5-7,7. children with рН less than, 7,0 are distinguish in the group of risk. . The pH lower than 6,0 is especially dangerous, because here the loss of mineralizing properties is especially intensive.
Sometimes the acidity of mixed salivacan be determined by means of standard рН-meter, for example рН- meter - 430
The role of resistance of hard tissues on a process of caries development.
Resistance of teeth for caries lesion depend on structure and chemical composition of hard tissues – enamel and dentine.
Development of enamel The enamel is formed with the highly-specialized cells- ameloblasts, which differentiate from the internal epithelial cells of the enamel organ.
There are two stages in a process of development enamel.
pre-eruptive
- secretory stage (formation of organic matrix)
- maturation (initial mineralization) stage
post-eruptive maturation (secondary) (additional tooth minerals derived from the saliva).
The initial event of the secretory stage occurs with an odontoblastic deposition of the first few microns of predentin. This is followed by the initiation of the secretory phase of the ameloblast. The first secreted enamel proteins do not accumulate as a layer, but instead, penetrates into the developing predentin and subjacent odontoblasts. The microenvironment of the ameloblast at this time, is mainly one of proteins and water. As the ameloblast retreats towards the future surface of the tooth, it uses these proteins to form an acellular and avascular matrix template upon which the future hydroxyapatite crystals are to be positioned. This requires a very rigid genetic control over the sequence of events that will extend through matrix formation, crystal nucleation, and crystal growth; as well as rod formation (elongation, widening, and maturation). Towards the end of the secretory stage, the matrix is almost completely degraded. initial mineralization stage The process of enamel initial maturation starts with the increase of Ca2+ and PO3+ concentration. The alkaline phosphatase catalyzes this process. It is characterized by formation of precipitate of insoluble calcium phosphate. The alkaline phosphatase is produced by the secretory cell (ameloblast) under the catalytic influence of vitamin D and some traceelements (Mn, Co, Sr). Process starts from the cutting edge of incisors, and cusps of molars and premolars. Fissures are less mineralized.
post-eruptive maturation (secondary) of tooth enamel.
For the first year after eruption into the mouth, the rods undergo a post-eruptive (secondary) maturation.
Under “maturation " understand a process increase of maintenance of calcium, fluorine and phosphorus and other mineral components of structure of enamel. At microscopic research in an immature enamel there are niches, deepenings, micropores; areas of underdense of "packing" of rods and crystals. Interrods spaces are extended, the borders of enamel rods are undistinct. It forms the microporosity of enamel. (the general volume of pores in immature enamel makes to 6% from all area of it surface
The main source of secondary enamel maturation is saliva This temporary hypomineralization of the enamel with its greater porosity, in part explains why newly erupted teeth are more susceptible to caries than teeth that have been present in the mouth for some time. After eruption (especially after 1,5-2 years), concentration of phosphorus and calcium in all anatomical layers of enamel increases, the calcium and phosphorus content in the superficial (OUTER LAYER) enamel layer is higher than in deep layer. As a result appears a high-mineralized , without rods superficial layer of enamel to 3 mkm. It has a high acidoresistance. Apatities of enamel are presented mainly hydroxyapatites, which are less steady to the action of acids of dental plague. During the maturing change the homogenecity of its structure there is smoothing out of relief of surface, perikimats disappear, an areas without rods appear. The volume of microspaces goes down to 0,1-0,2%, the closeness of enamel increases. The amount of water diminishes.
The content of calcium and phosphate in a saliva influences on mineralization and remineralization intensity of enamel and individual resistance to caries development.
The average calcium content in saliva is 0.04-0.08g/l, its content may vary from 0.006 to 0,123g/l. In general the saliva contains half the calcium content of the blood serum. Salivary calcium is in both ionized and bound state.
Saliary inorganic phosphate content is 0.06-0.65g/l. In average, salivary iorganic phosphate content is 2-4 times higher than that of the blood serum.
The major mechanism of homeostatic maintenance of mineral exchange in the oral cavity is the condition of salivary oversaturation with the hydroxyapatite Ca2+ and HPO4 2-. Salivary oversaturation with calcium and phosphate salts interferes enamel dissolution, as the saliva is already oversaturated with enamel constituents. The parotid saliva unlike other secretions is often oversaturated with it, with this more intensive maxillary caries affection is connected. The oversaturation of saliva with calcium and phosphate salts promotes diffusion of these ions to enamel, because their active concentration in saliva greatly exceeds such one in enamel, and the state of oversaturation promotes their absorption on the dental surface, consequently the speed of the first ion exchange phase in hydroxyapatite increases.
The state of salivary oversaturation with hydrozyapatite is closely connected with the salivary pH. With pH reduction the degree of oversaturation abruptly decreases. Salivary oversaturation is maintained to 6,0…..6,2, at the further acidulation it quickly loses nonsaturation with hydroxyapatite, it becomes able to dissolve it quickly and loses its mineralizing properties; so pH 6,0….6,2 is a critical one, when the saliva from the oversaturation state changes into the nonsaturated state, the liquid changes from the mineralizing into the demineralising one. The pH lower than 6,0 is especially dangerous, because here the loss of mineralizing properties is especially intensive.
The enamel is made up of billions of crystals that in turn make up millions of individual rods. Enamel rods can be correctly called enamel prisms. The peripheral layer of the prism is very resistant to acids.
Each rod that extends from the dentoenamel junction (DEJ) to the tooth surface is completed start-to-finish by one ameloblast. Around each rod there is an enveloping protein matrix. These enveloping protein wraps of both the enamel rods and crystals are the main channels for diffusion for demineralizing acids and remineralizing electrolytes. The inorganic phase of enamel is based on the mineral, hydroxyapatite (HAP) Са10(РО4)6(ОН)2, made up mainly of calcium (Ca), phosphate (PO4) and hydroxyl (OH) ions. It also contains trace amounts of other elements that happen to be in the bloodstream during enamel formation, in fact more than 40 elements have been identified in analysis of enamel. There are other types of apatite crystals: carbonateapatite, and fluorideapatite. Hydroxyapatite and carbonateapatite are less stable than fluorideapatite.
The final enamel is approximately 95% inorganic, and 5% organic material and water. 96% of inorganic substances and 1-2% of organic The early matrix has 20% of protein, and it is reduced to 0,3% simultaneously with enamel maturation. This 5% porosity forms (organic material 1-2% and water up to 4 %) a network of channels for fluid diffusion of ions and small molecules that are dispersed throughout the entire enamel cap. The space available for this diffusion is found between the rods and even between the crystals. To further extend this intra enamel network throughout the enamel, there are morphologic structures in the enamel with a high protein content, such as the striae of Retzius, enamel lamellae, enamel tufts, pores, and enamel spindles. These several diffusion channels probably serve two very important purposes in preserving the teeth: (1) to permit physiological remineralization throughout life, and (2) the voids and protein content in the enamel help to prevent fractures under biting pressures. Unfortunately, these same channels of diffusion also serve another purpose, the conducting of plaque acids into the enamel interior to cause demineralization.
High enamel hardness, its low solubility, determining its resistance to caries, is provided only under the conditions of maintenance the connection of dental enamel with pulp through its influence on the liquor flow speed. The removal of tooth pulp causes decrease of microhardness and acid-resistance of the tissue. The calcium and phosphorus content in dental tissues decreases.
The change of ion concentration on the enamel surface may because of potential changes stimulate the pulp. Here the odontoblasts may change liquor concentration at the enamel-dentine junction, they normalize the rest potential changing the liquor flow. that is why the influence of the pulp on preserving enamel properties can be described as liquor flow change as a reaction to osmotically-active substances, pH changes and other factors.
The major physiological enamel properties are resistance, solubility and permeability.
The enamel caries-resistance is ability to resist cariogenic factors. It is stipulated for optimum content of mineral components, phosphorus and calcium in enamel structure. The formation of resistance of enamel to caries is connected with its constant changes happening in the period of primary mineralization (before the tooth eruption) and during enamel maturation (after the tooth eruption).
The main index of enamel resistance is also Ca/P correlation. Healthy enamel of young persons has lower index Ca/P than the enamel of elderly. Usually this correlation is 1,67. The index Ca/P decreases at first signs of enamel demineralization.
Under the physiological conditions two processes simultaneously happen in the enamel- decalcination and mineralization. The criterion of this process proceeding into the pathological process is calcium reserve exhaustion and decrease of Ca/P correlation lower than 1,33, what is proved by the enamel inability to resist decalcination. At this phase ( protein matrix loss) remineralization is impossible, otherwise matrix restoration is necessary. That is why protein matrix maintenance and the maintenance of optimum correlation Ca/P must be one of the aims of prevention and therapy of different caries stages.
The existing data indicate that acid solubility (carious decalcination) of enamel is a complex chemical process accompanied by the changes in shape, size and orientation of apatite crystals with the previous decline of calcium content in the carious decalcination areas. Within reasonable limits of enamel dissolution the phosphorus loss also is happening. The caries-resistant tooth areas (tubercles, cusps) are highly-mineralized, they contain more calcium, while the areas affected by caries (fissures, precervical area) are hypomineralized and contain less calcium. The superficial enamel layer is the least soluble. In its deeper layers its solubility fluctuates, but it is higher than in the superficial layer.
According to the hydroxyapatite composition, (Са10(РО4)6(ОН)2) the molar correlation of the major ingredients (Ca and P) may fluctuate within the borders of 1,3-2,0 at the maintenance of the major mineral properties. This apatite property allows preserving integrity of the crystal structure at Ca ion loss (within some limits). At this some free areas or isomorphic Ca2+ substitution appear in the apatite crystalline grid.
Under the action of acids on hydroxyapatite, H+-ions substitute extra Ca2+-ions from the apatite crystalline grid.
Ca10(PO4)6 (OH)2 + 2H+
+
2HOH→Ca9
(H3
+O)2
(PO4)6(OH)2+Ca2+Ca10(PO4)6 (OH)2 + 2H+ + 2HOH→Ca9 (H PO4)2 (PO4)4(OH)2+Ca2+
In the first reaction Ca2+ is substituted by 2 ions of hydroxonium H3 +O, in the second Ca2+ is oxidized and displaced by the H+-ions with partial formation of monosubstituted P. In all cases the Ca/P index decreases to 1,30, this is considered as mineralization degree decrease, i.e. demineralization start. The oxyapatite structure is maintained, though its ability to resist the acid action decreases because of Ca2+-ions content.
This way, the ability to resist the action of acid depends on the index Ca/P excess over the minimum (1,30). Hydroxyapatite with the Ca/P index equal to 1,67 , is able to resist acid influence before substitution of two Ca2+ ions for the H+-ions in it. Oxyapatite with the Ca/P index equal to 1,30, can’t resist acid action, its structure is preserved.
Ca8 (H3 +O)4 (PO4)6(OH)2 + 4H+→2Ca2+ + 6CaHPO4 +6 H2O
To explain the established fact of selective decalcination of intact enamel in the process of acid dissolution we may guess that on the early stages of this process the two reactions take place:
1) the gradual and stoichiometric destruction of crystalline grid
Ca10 (PO4)6(OH)2 + 8H+→10Ca2+ + 6HPO4 2- + 2H2O
cation exchange on the enamel surface
Ca10 (PO4)6(OH)2 + 2H3O+←!!!!! Ca9 (H 3O)2(PO4)6 (OH)2 + Ca2+
The cation exchange (Ca2+ for H3 O+) from the demineralising solution at the second reaction type (in contrast to the first type reaction) is a reversible process and it doesn’t cause crystalline grid destruction of apatite.
The given scheme of two reactions of the mechanism of acid enamel solution explains reaction of enamel under conditions of prolonged calcium deficiency (1) and under the short-lasting action of H+-ions. If Ca2+ ions which constitute the enamel , didn't possess the capability for exchange, every action of acid agent would cause the immediate and irreversible enamel destruction.
Because of the ion-exchanging process H+-ions may be absorbed by the enamel without its structure destruction. At this Ca/P index decreases because of Ca2+ ions leaving the crystalline grid. So, enamel acts as a specific buffer system in relation to the acids forming in the oral cavity. As ion exchange is a reversible process, calcium-deficiency enamel apparatuses may remineralise; at this crystalline apatite grid is build at the expense of salivary Ca2+ ions, and absorbed hydrogen ions continuously leave enamel, in this way, Ca/P index becomes normal.