Effect of saliva and blood contamination on the shear bond strength of fifth-, seventh-, and eighth-generation bonding agents: An in vitro study.

AIM: To evaluate the bond strength of different adhesives after contamination with saliva and blood at various steps of application.
MATERIALS AND METHODS: Buccal surfaces of 180 human premolars were sliced to expose dentin. The specimens were randomly divided into six groups (n = 30), in which saliva and blood were used as contaminants in three groups each. The groups were further divided according to the generation (fifth - OptiBond Solo Plus Kerr, USA; seventh - OptiBond All-In-One Kerr, USA; eighth - Futurabond DC, Voco, Germany) of the adhesive used. Effect of contaminant application both before and after polymerization of the adhesive was evaluated. Composite cylinders were fabricated on exposed dentinal surfaces and were subjected to shear bond strength test. The results were subjected to one-way analysis of variance and t-test.
RESULTS: The eighth-generation adhesive showed the highest mean shear bond strength, followed by fifth- and seventh-generation adhesive. Salivary contamination resulted in greater decrease of bond strength when contaminated before polymerization. However, the results were vice versa when blood contamination was done, except in the case of fifth-generation adhesive. Blood contamination produced the lowest shear bond strength in all conditions.
CONCLUSION: The eighth-generation adhesives showed the highest and blood contamination the lowest bond strength in all conditions.

INTRODUCTION

The increasing demands for esthetic restorations have led to evolution of modern dental adhesives that have moved from multistep bonding process to simplification, i.e., self-etch and single bottle system. Blending of nanotechnology to adhesive dentistry leads to development of eighth-generation bonding agents (Futurabond DC, Voco, Germany). These are self-etch adhesives with two components that are mixed before application and are applied as single step. These are claimed to have better enamel and dentin bond strength, stress absorption, and longer shelf life.[1]

However, the fact remains that in order to have successful adhesion between resin and the tooth substrate, it is necessary that the adhesive substrate should not be contaminated. Contaminants such as saliva, blood, hemostatic agents, gingival fluid, and handpiece oil can adversely affect the longevity of the restoration owing to microleakage, which in turn leads to sensitivity, tooth discoloration, secondary caries, and eventual loss of the restoration. Moreover, proper isolation remains challenging in cases where cavity margins extend below the gingival tissues.

To the best of our knowledge, no study is available until now evaluating the effect of contaminants such as saliva and blood on the shear bond strength of eighth-generation bonding agents.

Thus, the aim of this study was to compare the influence of salivary and blood contamination at different steps of application on the dentin bond strength of fifth-, seventh- and eighth-generation adhesives. The null hypothesis of this study was that saliva and blood contamination would not affect the dentin bond strength of total- and self-etch adhesives.

MATERIALS AND METHODS

One hundred and eighty extracted human intact maxillary or mandibular premolars were taken. The teeth were cleaned of any residual tissue tag and stored in normal saline. The teeth were embedded in aluminum molds with self-cure acrylic resin till the cementoenamel junction. The buccal surfaces of the teeth were ground on wet 240-grit silicon carbide disks to remove enamel and expose the dentin. Before bonding, fresh whole saliva and blood were collected from a single donor in a sterile beaker and used immediately.

All the teeth were randomly divided into six groups of thirty teeth each:

Groups 1–3: Tooth surfaces contaminated with saliva

Groups 4–6: Tooth surfaces contaminated with blood

Groups 1 and 4: OptiBond Solo Plus, Kerr, USA (fifth generation)

Groups 2 and 5: OptiBond All-In-One, Kerr, USA (seventh generation)

Groups 3 and 6: Futurabond DC, Voco, Germany (eighth generation).

Depending on the stage of contamination, each group was further subdivided into three subgroups (A, B, and C) of 10 teeth each:

Subgroup A (control, n = 10): Respective adhesive application according to the manufacturer's instructions

Subgroup B (n = 10): Contamination with saliva/blood after application of bonding agent and prior to curing. The contaminant was pooled over the area with a microbrush, left for 10 s, and blot dried

Subgroup C (n = 10): Contamination with saliva/blood after curing of bonding agent. The contaminant was pooled over the area with a microbrush, left for 10 s, and blot dried.

A Teflon tube of 3 mm × 5 mm was applied at the cervical region of all specimens to simulate oral environment and filled with a composite resin (Herculite Precis, Kerr, USA, Shade A2) in two increments. Each increment was compressed firmly and light cured for 40 s using a quartz tungsten halogen light. The Teflon tube was removed and additional curing of resin cylinder was done. To simulate the oral conditions, these specimens were stored in distilled water at 37°C for 24 h. The shear bond strength testing was done in an Instron Universal Testing Machine at the Centre for Advanced Research, Ghaziabad, at a cross-head speed of 0.5 mm/min. A knife edge chisel-shaped shearing rod was used for debonding of resin cylinders. The data obtained were subjected to analysis by one-way analysis of variance, while comparison of means between the groups was done using t-test. Statistical Package for Social Sciences software, 16.0 Version (SPSS, Chicago, illinois, U.S.A) was used for statistical analysis.

RESULTS

The results of the study [Tables 1 and 2] revealed that the eighth-generation adhesives showed the highest mean shear bond strength, followed by the fifth- and the seventh-generation adhesive showed the least. Significant bond strength reduction was seen when samples were contaminated with saliva or blood. All the three adhesives in groups contaminated with saliva resulted in greater decrease of bond strength when contaminated before polymerization as compared to when contaminated after polymerization. The difference between the groups was statistically significant, except in the case of total-etch adhesives. When the three adhesives were contaminated with blood, contamination after polymerization resulted in greater decrease of bond strength than before polymerization in self-etch adhesives (seventh and eighth generation), whereas in the total-etch adhesives, the decrease was more when contaminated before polymerization. The groups showed statistically significant differences. Overall, when saliva and blood were compared, blood contamination produced the lowest shear bond strength in all conditions.

Table 1

Shear bond strengths for tested groups (mean±standard deviation, MPa) after salivary contamination

Table 2

Shear bond strengths for tested groups (mean±standard deviation, MPa) after blood contamination

DISCUSSION

In the present study, the null hypothesis was rejected as saliva and blood were found to lower the dentin bond strength of self- and total-etch adhesives.

When no contamination occurred, the eighth-generation adhesives exhibited the greatest bond strength, followed by the fifth-generation adhesives, but the difference between the two was not significant. The seventh-generation self-etch adhesives exhibited the least dentin bond strength.

The eighth-generation adhesives have highly functionalized SiO2 nanoparticles (0.20 nm) that may have contributed to its greatest bond strength. These nanoparticles facilitate cross-linking of the resin components and intensify its film building properties. According to the manufacturers, the adhesive has superior wetting properties and their nanoparticles could strengthen the hybrid layer for greater bond strength. They also fortify a “stick immediately” effect which affirms that the bond will not be displaced from the cavity while air drying.

Ethanol in OptiBond Solo Plus aids in resin infiltration through the nanospaces of exposed collagen fibers by replacing and competing with moisture. The hybrid layer formed results in stronger micromechanical interlocking between the dentin and the superficially demineralized dentin.[2] Hence, fifth-generation adhesives were found to have better bond strength than the seventh-generation adhesives.

On the other hand, the seventh-generation adhesives form an insubstantial hybrid layer and shorter resin tags (2 μm) because of the low pH of methacrylate monomers when analogized with 37% phosphoric acid.[2] Water as solvent in self-etching systems promotes ionization of the acidic monomers, rendering these adhesives permeable films that are highly vulnerable to the degrading effects of water. After the solvent evaporates, the adhesive membrane left can be very thin, and its mechanical properties may be compromised.[3]

Self-etching adhesives depend on acidic monomers to concomitantly demineralize and infiltrate dentin. The mineral content of the tooth structure must neutralize this acidity to allow complete polymerization of the adhesive layer. With total-etch adhesives, the application of the etchant removes the smear layer. As there is remaining acidity and incapacity to remove the smear layer, the self-etch adhesives have lower bond strength.[2]

Saliva constitutes of more than 99% water and causes an excess of moisture which eventually lowers the bond strength of dentin adhesives.[4]

In this study, saliva contamination in the total-etch adhesives both before and after polymerization caused a great depletion of dentin bond strength, but the difference between the two steps of contamination was statistically insignificant. Pashley et al.[5] and Fritz et al.[6] attributed the depletion of adhesion before curing to the occlusion of open dentinal tubules by salivary proteins.[456] Moreover, increase in the contact angle could further decrease the bond strength.[6] On the contrary, according to some studies, the humidity of the saliva prevents the collapse of the collagen network by providing a wet dentin surface and eliminates influence of saliva contamination on the bond strength of this adhesive to dentin.[789101112]

Salivary contamination following adhesive polymerization also leads to reduced bond strength owing to the deposition of salivary glycoprotein over the superficial deficiently polymerized adhesive layer. This may have created a physical barrier lessening the effective copolymerization.

Salivary contamination of the self-etch adhesives (seventh, eighth generation) showed statistically significant differences between the control and the contamination before polymerization. Their bond strength reduction could be attributed not only to the macromolecules occluding the dentinal tubules but also to the mixture of water with the adhesive.[13]

In the present study, the hydrophilic nature of resins, glycerol phosphate dimethacrylate (GPDM) in the seventh-generation adhesive and hydroxyethyl methacrylate (HEMA) molecules in the eighth-generation adhesives, causes them to retain water within the adhesive film and becomes more dispersed in water. Thus, they are incompetent to participate in chain growth during polymerization.[14] In addition, the use of an air stream might not remove water from GPDM/HEMA mixtures adequately.[1516] Moreover, salivary proteins might also interfere with infiltration of hydrophilic monomers during the hybridization process,[17] thus lessening the dentin bond strength. However, in contrast to our study, some authors reported that hydrophilic nature of the recent adhesives and application of their multiple coats may allow them to exert their function to some degree in the presence of saliva by displacing or propagating through it and forming the hybrid layer.[18]

Statistically significant decline in bond strength was also seen when salivary contamination occurred after curing of the adhesive. This is in agreement with Fritz et al.,[6] who attributed the findings to adsorption of salivary glycoproteins to the poorly polymerized adhesive surface, thus intercepting adequate copolymerization. On the contrary, Hitmi et al. stated that there is no salivary diffusion after the curing of adhesive.[17]

Our study showed that self-etch adhesives may be less sensitive to salivary contamination compared to fifth-generation bonding agents. Similar results have been reported by Sheikh et al., who attributed the findings to the hydrophilic nature of self-etch adhesives and their inherent acidity which helps them displace through the salivary macromolecules and preserve the bond strength.[19]

Similar bond strength reduction was observed when using blood as the contaminant in all the tested bonding agents at all steps of contamination both before and after polymerization in all generations of adhesives. Moreover, on comparing saliva with blood, blood contamination produced the lowest shear bond strength in all conditions. It could be because blood creates a greater mechanical barrier than saliva owing to the difference in the type and amount of inorganic and organic elements in the blood.[20]

It was also observed that blood contamination after polymerization caused greater reduction in bond strength when compared to contamination before polymerization. These findings are in accordance with those of Kaneshima et al., who stated that blot drying of blood contaminants after adhesive curing could interfere with copolymerization by forming a barrier between the adhesive and the restorative material. They also stated that decontamination procedures such as blot drying before adhesive curing are able separate blood from adhesion surface. Moreover, the acidic primer in self-etch adhesives could clean blood and retain the bond strength more as compared to contamination after curing.[21]

However, this was not the case in total-etch adhesives. Blood constitutes of heavier macromolecules such as platelets and fibrinogen which occluded the dentinal tubules and lead to shallower resin tag formation. This contamination in turn causes great reduction in bond strength before adhesive curing. Furthermore, some reaction between blood constituents and exposed collagen was reported to degrade primer infiltration.[20]

Owing to the inherent limitations of an in vitro study, the effect of contamination on the bonding ability of these adhesive systems to dentin warrants further investigation.

CONCLUSION

Within the limitations of this study, the following conclusions can be drawn:

Blood and saliva contamination had a negative impact on the shear bond strength to the dentin

Blood contamination affected the bond strength more than the saliva contamination

The bond strength was affected more if contamination occurred after adhesive application.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.