**4. Aim and requirements for sealing materials**

### **The aim of sealant application is:**


#### **Requirements for sealing materials:**


250 Contemporary Approach to Dental Caries

ability to release fluorine ions. Therefore the choice of GIC as fissure sealant must result from other reasons than good retention or cariostatic property. Certainly it can be an alternative e.g. in the absence of proper humidity control, with partial tooth eruption or with mentally or physically handicapped patients. Apart from that, glass-ionomer cements have another advantage of simple application technique. Increased efficacy of sealing can be obtained by reapplication of sealant during routine visits, however this will increase cost of treatment. That is why only certain clinical situations can constitute an indication for such

It was observed that resin-modified cements demonstrate much stronger bonding forces with dentine (almost threefold) compared to conventional glass-ionomer cements, but considerably weaker than sealants based on bis-GMA resins. Retention of glass-ionomer cements after 6 months equaled 93%, after 24 months from 82.5% to 86%, and after four-year observation only 35% (50,51,86). Baseggio *et al.* assessed the efficacy of Vitremer modified glass-ionomer cement compared to Fluoishield conventional sealant based on bis-GMA resins containing fluorine. During three-year studies on sealants in second molars in 320 patients aged 2-16 years, retention of sealant modified with glass-ionomer cement was 5.10%, whereas retention of sealant based on bis-GMA resin equaled 91.08%, and total loss of retention was observed in 6.37% and 7.65% of sealed teeth respectively. Caries on masticatory surfaces was observed in 20.06% of teeth sealed with materials modified with glass-ionomer cement and in 8.91% of teeth sealed with materials based on bis-GMA resin

Compomers, also called composite materials modified with polyfunctional acids, contain bis-GMA monomers, deionized glass which constitutes mechanical filler with 42-67% of volume and particle size 0.7-5μm, and monomers containing COOH acidic functional groups. Thanks to glass content they are capable of releasing fluorine contained in glass, but in a much lesser quantity than GICs or resin-modified GICs (63). A two-year assessment of sealants based on bis-GMA resins (Fissurit F, Fissurit FX, compomer Dyract Seal and ormocer Admira Seal) demonstrated that retention of the compomer (Dyract Seal) was much lower compared to the other sealants. However, no significant differences were observed in marginal tightness of assessed sealants and in caries presence on surfaces of sealed teeth (88). In research comparing two materials - one compomer and one based on bis-GMA resin - special emphasis was placed on advantages of compomers, which are easy to use, chemically and physically durable, and they release fluorine. No difference was observed between applied sealant and its retention and between the degree of caries reduction and material retention (quot. after 24). In another study in which GIC-based material with zinc and fluorine was used (Jonosit Seal DMG), after six years of observation in 77.5% of cases retention on occlusal surfaces of teeth was confirmed. Presence of caries was closely related to retention of material. In case of total retention of sealant, caries reduction was 99.6%, whereas with total loss of material after five years caries reduction was 69% (34). Other authors, such as Aranda *et al.* and deLuca-Froga *et al.*, who assessed activity and preventive performance of resin-modified glass-ionomer cements (RMGICs) and composite materials modified with polyfunctional acids, observed that retention index changed after one-year observation, rising from 20% to 95.9% (4,18). An important factor which must be considered in case of glass-ionomer cements used as sealants is the fact that even after clinical loss of material, a small amount of sealant remains at the fissure's bottom with fluorine still being released, thus protecting masticatory surface. Higher grow of caries was observed in the

treatment, for example difficult children or those with a high caries risk (23).

(7).


Sealing of Fissures on Masticatory Surfaces of Teeth as a Method for Caries Prophylaxis 253

barrier for dye penetration. The barrier resulting from material's hydrophobic properties is

ASPA and Poly F glass-ionomer cements contain polyacrylic matrix, which is hydrophilic thanks to carboxyl groups. These materials are permeable to water and dye solutions. 24 hours after their application it was observed that aqueous dye solutions penetrated through the sealant-enamel interface and through the material. Another study on Concise, Delton, Kerr, Nuva-Seal sealants with the use of aqueous dye solutions with Ca45 and S35 radioactive isotopes did not show dye penetration when enamel was etched and sealant permanently and tightly adhered to enamel. Also extension of enamel curing time up to 30 seconds resulted in 32% decrease of microleakage occurrence compared to teeth etched for a

Studies determining microhardness of sealants demonstrated varied degree of this parameter from initial 7.51±0.62 for Concise EB and 4.78±0.52 for ASPA cement, up to values increasing in time after three months: 16.49±0.29 for Concise EB and 55.28±1.31 for ASPA cement. Compared to sealants based on bis-GMA formula, ASPA cements after three months demonstrated higher stability and a higher flexibility factor. With increased amount of sealant, rigidity of material also increases along with susceptibility to deformation, that is

Solubility and disintegration of bis-GMA type sealing materials in oral environment is low and has no clinical significance. Compared to bis-GMA resins, glass-ionomer cements undergo dissolution and disintegration to a much higher degree. Water absorption of sealing materials is low, which is beneficial as it contributes to closure of marginal fissure

Sealants' resistance to abrasion depends on type of material, content, size and arrangement of filler's particles, as well as on anatomical conditions of sealed teeth. It is assumed that abrasion of sealants based on bis-GMA formula occurs due to impairment of silane bonding agent. Abrasion is a two-phase process: the first phase is characterized by abrasion of polymer and exposure of filler particles, whereas in the second phase they are torn out of the material. Abrasion of sealant on masticatory surface begins on fissure periphery in sites with the thinnest layer of material. Defects on fissure periphery are retention sites, where dental plaque accumulates. The rate of material abrasion also depends on the method of bonding basic ingredients and on the way of material application. Air bubbles trapped in material are sites with decreased resistance to abrasion. The degree of abrasion also depends on activity of separate tooth groups during mastication and on anatomy of tooth and its position in the arch. Sealant abrasion is observed in distal part of superior molars' masticatory surface twice as frequently as in medial parts of those teeth. Moreover, it has been proved that the highest abrasion occurs soon after sealant placement and it decreases with time. The lowest sealant loss was observed in inferior first premolars compared to the other premolar teeth. After 30 months mean value of material volume loss for premolars was 0.43±0.24mm2, whereas mean depth of material loss was 221.8±115.1µm. In *in vitro* studies, abrasion, penetration and deformation four most commonly used bis-GMA resins sealants was assessed. It was proved, that materials were abraded to a different degree. The observed level of material wear increased as follows: Delton-21.5; Kerr-22.3; Nuva Seal-23.9; Carbimet -3.,0 (\*10-4 mm2/mm). The assessed sealants differed considerably in degree of penetration and formability, where the lowest values were observed for Kerr, and the

Young's modulus. The best materials are those with Young's modulus below 10 GPa.

not stable and with time it deteriorates, which results in microleakage.

shorter time.

created by polymerization contraction.


Appropriate period of activity, fast and easy application of sealants, fast bonding and curing should be within time limits accepted by the patient. Long curing may cause anxiety in a child, leading to moisturizing of treatment area, to outflow of material beyond the sealed site and thus to disturbance of proper polymerization (10). It occurs especially with little children, where sealed teeth are not yet fully erupted and covered with a fold of mucous membrane. Therefore chemically cured resins, which are cured from 45 to 90 seconds after initial mixing of material until completion of curing are not recommended. Materials polymerized with halogen light should be used, as they have a more favorable, unlimited activity time and are characterized by fast curing up to 20-40 seconds, depending on intensity of lamp-emitted light.

Constitution of materials for sealing fissures is generally similar to constitution of composite materials used to fill cavities in hard tissues of teeth. The difference is that fissure sealants have more liquid consistence than fillers, which allows penetration into etched fissures. Sealants based on bis-GMA formula contain amine accelerators and an initiator - benzoyl peroxide. Light-cured sealing materials require a lamp emitting wavelengths of 340-400 nanometers to initiate polymerization. Contemporary sealants are made of one ingredient which does not need mixing. It consists of three parts of viscous bis-GMA monomer, which is diluted with one part of MMA monomer (methyl methacrylate) in order to obtain material of relatively low viscosity. The activator is 2% methylbenzyl ether, which in the presence of 1-2% benzoyl peroxide and light initiates polymerization. Research on appropriate selection of monomer and diluent is very important and provides information about obtained physical properties of sealants. Replacement of methacrylate monomer with other monomers leads to changes of viscosity, impacts extension or shortening of curing time, and alters its hydrophilic character. Efficacy of sealing depends on resin's ability to penetrate fissures before its curing, i.e. on the ability to create a mechanical barrier for carious process. In order to produce an adequately strong bonding and secure proper retention, sealant has to flow over the surface of etched enamel and penetrate microfissures on etched surface. However, it is believed that resin's penetrative ability depends on etching pattern and enamel's moisture, surface tension of material, its viscosity and penetration index. Modifications in application of various monomers resulted in change of cuing time.

Studies on sealants' tightness confirmed the occurrence of microleakages for all assessed sealing materials (e.g. Epoxylite 9075, Nuva Seal, ESPE 717, Concise Enamel Bond, Kerr Fissure, Sealant Delton). Microleakage was observed on the sealant-enamel interface, and in case of glass-ionomer cements, dye penetrated through the whole surface of material. Nuva-Seal seemed the most resistant to dye penetration, though after two weeks some number of cases where dye penetration on the sealant-enamel interface occurred could be observed. In most studied cases ESPE 717 and Epoxylite 9075 materials demonstrated microleakage on the sealant-enamel interface and through the material. The occurrence of microleakage was related to hydrophobic structure of materials and their porosity, which constitutes a better

Appropriate period of activity, fast and easy application of sealants, fast bonding and curing should be within time limits accepted by the patient. Long curing may cause anxiety in a child, leading to moisturizing of treatment area, to outflow of material beyond the sealed site and thus to disturbance of proper polymerization (10). It occurs especially with little children, where sealed teeth are not yet fully erupted and covered with a fold of mucous membrane. Therefore chemically cured resins, which are cured from 45 to 90 seconds after initial mixing of material until completion of curing are not recommended. Materials polymerized with halogen light should be used, as they have a more favorable, unlimited activity time and are characterized by fast curing up to 20-40 seconds, depending on

Constitution of materials for sealing fissures is generally similar to constitution of composite materials used to fill cavities in hard tissues of teeth. The difference is that fissure sealants have more liquid consistence than fillers, which allows penetration into etched fissures. Sealants based on bis-GMA formula contain amine accelerators and an initiator - benzoyl peroxide. Light-cured sealing materials require a lamp emitting wavelengths of 340-400 nanometers to initiate polymerization. Contemporary sealants are made of one ingredient which does not need mixing. It consists of three parts of viscous bis-GMA monomer, which is diluted with one part of MMA monomer (methyl methacrylate) in order to obtain material of relatively low viscosity. The activator is 2% methylbenzyl ether, which in the presence of 1-2% benzoyl peroxide and light initiates polymerization. Research on appropriate selection of monomer and diluent is very important and provides information about obtained physical properties of sealants. Replacement of methacrylate monomer with other monomers leads to changes of viscosity, impacts extension or shortening of curing time, and alters its hydrophilic character. Efficacy of sealing depends on resin's ability to penetrate fissures before its curing, i.e. on the ability to create a mechanical barrier for carious process. In order to produce an adequately strong bonding and secure proper retention, sealant has to flow over the surface of etched enamel and penetrate microfissures on etched surface. However, it is believed that resin's penetrative ability depends on etching pattern and enamel's moisture, surface tension of material, its viscosity and penetration index.

Modifications in application of various monomers resulted in change of cuing time.

Studies on sealants' tightness confirmed the occurrence of microleakages for all assessed sealing materials (e.g. Epoxylite 9075, Nuva Seal, ESPE 717, Concise Enamel Bond, Kerr Fissure, Sealant Delton). Microleakage was observed on the sealant-enamel interface, and in case of glass-ionomer cements, dye penetrated through the whole surface of material. Nuva-Seal seemed the most resistant to dye penetration, though after two weeks some number of cases where dye penetration on the sealant-enamel interface occurred could be observed. In most studied cases ESPE 717 and Epoxylite 9075 materials demonstrated microleakage on the sealant-enamel interface and through the material. The occurrence of microleakage was related to hydrophobic structure of materials and their porosity, which constitutes a better






intensity of lamp-emitted light.

barrier for dye penetration. The barrier resulting from material's hydrophobic properties is not stable and with time it deteriorates, which results in microleakage.

ASPA and Poly F glass-ionomer cements contain polyacrylic matrix, which is hydrophilic thanks to carboxyl groups. These materials are permeable to water and dye solutions. 24 hours after their application it was observed that aqueous dye solutions penetrated through the sealant-enamel interface and through the material. Another study on Concise, Delton, Kerr, Nuva-Seal sealants with the use of aqueous dye solutions with Ca45 and S35 radioactive isotopes did not show dye penetration when enamel was etched and sealant permanently and tightly adhered to enamel. Also extension of enamel curing time up to 30 seconds resulted in 32% decrease of microleakage occurrence compared to teeth etched for a shorter time.

Studies determining microhardness of sealants demonstrated varied degree of this parameter from initial 7.51±0.62 for Concise EB and 4.78±0.52 for ASPA cement, up to values increasing in time after three months: 16.49±0.29 for Concise EB and 55.28±1.31 for ASPA cement. Compared to sealants based on bis-GMA formula, ASPA cements after three months demonstrated higher stability and a higher flexibility factor. With increased amount of sealant, rigidity of material also increases along with susceptibility to deformation, that is Young's modulus. The best materials are those with Young's modulus below 10 GPa.

Solubility and disintegration of bis-GMA type sealing materials in oral environment is low and has no clinical significance. Compared to bis-GMA resins, glass-ionomer cements undergo dissolution and disintegration to a much higher degree. Water absorption of sealing materials is low, which is beneficial as it contributes to closure of marginal fissure created by polymerization contraction.

Sealants' resistance to abrasion depends on type of material, content, size and arrangement of filler's particles, as well as on anatomical conditions of sealed teeth. It is assumed that abrasion of sealants based on bis-GMA formula occurs due to impairment of silane bonding agent. Abrasion is a two-phase process: the first phase is characterized by abrasion of polymer and exposure of filler particles, whereas in the second phase they are torn out of the material. Abrasion of sealant on masticatory surface begins on fissure periphery in sites with the thinnest layer of material. Defects on fissure periphery are retention sites, where dental plaque accumulates. The rate of material abrasion also depends on the method of bonding basic ingredients and on the way of material application. Air bubbles trapped in material are sites with decreased resistance to abrasion. The degree of abrasion also depends on activity of separate tooth groups during mastication and on anatomy of tooth and its position in the arch. Sealant abrasion is observed in distal part of superior molars' masticatory surface twice as frequently as in medial parts of those teeth. Moreover, it has been proved that the highest abrasion occurs soon after sealant placement and it decreases with time. The lowest sealant loss was observed in inferior first premolars compared to the other premolar teeth. After 30 months mean value of material volume loss for premolars was 0.43±0.24mm2, whereas mean depth of material loss was 221.8±115.1µm. In *in vitro* studies, abrasion, penetration and deformation four most commonly used bis-GMA resins sealants was assessed. It was proved, that materials were abraded to a different degree. The observed level of material wear increased as follows: Delton-21.5; Kerr-22.3; Nuva Seal-23.9; Carbimet -3.,0 (\*10-4 mm2/mm). The assessed sealants differed considerably in degree of penetration and formability, where the lowest values were observed for Kerr, and the

Sealing of Fissures on Masticatory Surfaces of Teeth as a Method for Caries Prophylaxis 255

more, the amount of sealant's dose used in the study did not impact BPA concentration in blood serum. Further research is necessary to establish the impact of BPA from sealants on

Chemical compounds contained in sealing materials may have a destructive influence on genetic material in DNA and RNA; it was proved that as many as 14 components of composite materials indicate genotoxicity. On the other hand it was not demonstrated if materials based on bis-GMA and UDMA (urethane dimethacrylate) indicate mutagenicity. It is known that TEGDMA (triethylene glycol dimethacrylate) used as componomer in nontoxic concentrations has mutagenic properties, it impairs proper cell structure, e.g. through interactivity with cell membrane, and its functionality, e.g. decreasing cell's glutathione level, which is responsible for cell structure protection and detoxication. Apart from that it changes expression level of many genes which are important for proliferation processes, control of cell cycle and cell death as well as for DNA replication and repair.

Dentine bonding systems, used to increase adhesion of sealant to hard tissues of teeth, including glutaraldehyde, proved mutagenic and genotoxic in some *in vivo* studies (89).

Data from available studies indicate that concentrations necessary to evoke mutagenic reaction in *in vivo* conditions are much higher than those which evoke a reaction in a patient. Data on mutagenic impact of materials show that their components act mainly in *in vitro* conditions. Such situation occurs especially in case of dentine bonding systems and lightactivated glass-ionomer cements. It was proved that resin-modified glass-ionomer cements (Vitrebond) evoked genotypic reactions. The cause was 2-phenylindole chloride which had

Therefore all materials introduced into oral cavity in certain conditions may harmfully

Sealing procedure should be performed not later than four months after eruption of first permanent molars, at the age of 5-6 years, but practically even after 10 years of age premolars at the age of 8-12 years, especially if caries is observed in deciduous teeth and first permanent molars, second permanent molars at the age of 11-13 years, and the procedure can be performed up to the 15th year of age. In a population with a high caries

Sealant should be applied in teeth with narrow and deep fissures and in teeth with developmental abnormalities on masticatory surfaces. Sealing depends mainly on meticulousness during clinical procedure, i.e. isolation of teeth from moist oral environment, thorough cleaning of occlusal surfaces, etching, rinsing, drying and

This is a very important stage of the procedure. Maintaining the area dry is critical for successful sealing procedure. Usage of dental dam, saliva ejector and alternatively lignin

impact the organism through releasing specific doses of chemical compounds.

risk, also deciduous molars and permanent premolars should be sealed.

estrogen balance.

cytotoxic effect on cell cultures.

**4.1.1 Tooth selection** 

application of sealant.

rolls is recommended.

**4.1 Technique of sealing procedure** 

**4.1.2 Isolation of teeth to be sealed** 

highest values were observed for Nuva Seal, which was conditioned by material composition. For example, Kerr had 40% of quartz filler particles by volume, which increased its resistance to abrasion and decreased the risk of damaging the surface layer of resin susceptible to abrasion. Due to the necessity of avoiding mastication impairment, sealing material should be present only in anatomical fissures and crevices. Therefore its excess should be removed from occlusal surface.

Every substance of foreign origin which is introduced into oral environment may have an adverse impact and induce certain biological or health effects. Almost all organic ingredients can be washed out from polymerized sealing material by organic solvents, e.g. methanol, ethanol or water. Released formaldehyde and metacrylic acid are particularly dangerous, the latter creating an inhibitory oxygen layer in the surface layer of sealant after polymerization and it can release into oral environment for a long time (up to 115 days after polymerization). Admittedly it does not cause toxic effect, but it may contribute to a local allergic reaction. During monomer particle disintegration, alcohol may be formed (e.g. from bis-GMA resin - bivalent alcohol). Further metabolism of alcohol occurs in digestive tract, leads to release of bisphenol A (BPA), which is a constituent of many compounds, such as bis-GMA, bis-EMA (bisphenol ethoxylate dimethacrylate), TEGDMA (triethylene glycol dimethacrylate), applied in sealing materials. Bisphenol A was detected in saliva of patients subjected to prophylactic fissure sealing. The substance belongs to the group of chemical compounds called xenoestrogens, which - joining with estrogen receptors - imitate the activity of natural hormones, thus they may impact human health. In a study by Olea *et al.* (56) on the presence of bisphenol A, it was detected at the level of 90-931µg/ml in saliva samples collected from patients one hour after sealing. The study also demonstrated that sealing material exposed to 100oC temperature for 30 minutes in buffers with pH 1 and pH 13, released bisphenol A in concentrations causing increased proliferation of MCF-7 cells up to one hour after application, which indicates induction of para estrogenic activity. In other studies authors assessing seven different sealants *in vitro* did not confirm the presence of bisphenol A in any of the assessed materials. However, determinable amounts of TEGDMA were detected in all studied sealants. It can also be supposed that with proper proceeding, bisphenol A will not be released if substances reacting with each other have a sufficient degree of purity. Only in products containing bis-GMA, small amounts of bisphenol A are released, and it occurs only directly after material curing. In most studies bisphenol A was detected in saliva only directly after application of sealant, and this was the case especially with "older" generation of Delton sealant, which still contained bis-DMA (bisphenol A dimethacrylate) in concentrations not affecting the organism. Approximate calculations of bisphenol A released from sealants showed that its level is lower than 1.5%, and such concentrations are too small to cause carcinogenic effect. A study assessing the presence and concentration of BPA (bisphenol A) in blood serum and saliva in 30 individuals aged 18-40 years before sealing and one, three and twenty-four hours after sealing (Delton Pit&Fssure Sealant Dentsplay) indicated the presence of bisphenol A in all examined patients before sealing at the level from 0.07 to 6.00 ng/ml. The level of bisphenol A three hours after sealing had the highest value and after twenty-for hours it returned to the same value as before sealing. The highest recorded concentrations of bisphenol A after a single application and after four subsequent applications were 3.98mg/ml and 9.08mg/ml respectively. Bisphenol A was not observed in blood serum at any stage of the study (89). Exposures to BPA from other sources than resins contributed to a change of BPA level in saliva. What is more, the amount of sealant's dose used in the study did not impact BPA concentration in blood serum. Further research is necessary to establish the impact of BPA from sealants on estrogen balance.

Chemical compounds contained in sealing materials may have a destructive influence on genetic material in DNA and RNA; it was proved that as many as 14 components of composite materials indicate genotoxicity. On the other hand it was not demonstrated if materials based on bis-GMA and UDMA (urethane dimethacrylate) indicate mutagenicity. It is known that TEGDMA (triethylene glycol dimethacrylate) used as componomer in nontoxic concentrations has mutagenic properties, it impairs proper cell structure, e.g. through interactivity with cell membrane, and its functionality, e.g. decreasing cell's glutathione level, which is responsible for cell structure protection and detoxication. Apart from that it changes expression level of many genes which are important for proliferation processes, control of cell cycle and cell death as well as for DNA replication and repair.

Dentine bonding systems, used to increase adhesion of sealant to hard tissues of teeth, including glutaraldehyde, proved mutagenic and genotoxic in some *in vivo* studies (89).

Data from available studies indicate that concentrations necessary to evoke mutagenic reaction in *in vivo* conditions are much higher than those which evoke a reaction in a patient. Data on mutagenic impact of materials show that their components act mainly in *in vitro* conditions. Such situation occurs especially in case of dentine bonding systems and lightactivated glass-ionomer cements. It was proved that resin-modified glass-ionomer cements (Vitrebond) evoked genotypic reactions. The cause was 2-phenylindole chloride which had cytotoxic effect on cell cultures.

Therefore all materials introduced into oral cavity in certain conditions may harmfully impact the organism through releasing specific doses of chemical compounds.

## **4.1 Technique of sealing procedure**
