Objectives . To evaluate the effect of four collagen cross-linking agents, Glutaraldehyde (GD), Grape Seed Extract (GSE), Genipin (GE) and Tannic Acid (TA) on the Reduced Modulus of Elasticity (Er) and Nanohardness (H) of demineralized dentin.

Materials and methods . The occlusal surfaces of two human molars were ground flat and finely polished to expose dentin. Each tooth was then sectioned into 4 equal segments and these segments were etched with 32% Phosphoric Acid Etchant for 20 s to superficially demineralize dentin. Initial measurements were performed before treatment to establish baseline values. The segments were then treated with the cross-linking agents for 1 min, 10 min and 60 min of cumulative exposure time. Nanomechanical properties were assessed at each time point. The cross-linkers used were: 5% Glutaraldehyde (GD), 6.5% Grape Seed Extract (GSE), 0.5% Genipin (GE) and 20% Tannic Acid (TA); with the pH of the agents maintained between 7.2 and 7.4. The samples were kept hydrated in Hank’s Solution at all times, including during testing. The reduced modulus Er (GPa) and nanohardness H (GPa) of the demineralized dentin were evaluated using a Triboindenter (Hysitron) by making a total of 12–15 indentations at three seperate sites. The data were statistically analyzed using two-way ANOVA and Tukey tests ( p < 0.05).

Results . For Er, a statistically significant interaction was observed between treatment agent and times ( p < 0.0001). There was a statistical difference between all the cross-linking agents ( p < 0.0001) as well as the treatment times ( p < 0.0001). GSE and TA showed the maximum increase in Er values, ranging from 0.198 GPa (±0.072) to 0.690 GPa (±0.203) for GSE and 0.232 GPa (±0.078) to 0.843 GPa (±0.208) for TA for post etch testing to 60 min of treatment respectively. A statistically significant interaction was also observed for hardness values between the treatment agents and treatment times ( p < 0.0001). There was no statistical difference in H values between GD and GE ( p = 0.54) or TA and GSE ( p = 1.000), while GSE and TA presented statistically higher values ( p < 0.05). Overall, hardness increased as exposure time increased, ranging from a lowest baseline value of 0.008 GPa (±0.003) to 0.029 GPa (±0.012) after treatment for 60 min with GSE.

Conclusions . The use of collagen cross-linkers increased the nanomechanical properties of demineralized dentin. Different hardness (H) and reduced modulus (Er) values may indicate distinct interaction mechanisms between agents and the dentin matrix. Increase in exposure time resulted in gradual increase in the mechanical properties. GSE and TA showed the greatest increase in H values while TA presented the most remarkable changes in Er values.

Research supported by NIH-NIDCR (DE017740).