Abstract
Objective
Recent studies developed antibacterial bonding agents and composites containing a quaternary ammonium dimethacrylate (QADM) and nanoparticles of silver (NAg). The objectives of this study were to investigate: (1) the effect of antibacterial primers containing QADM and NAg on the inhibition of Streptococcus mutans impregnated into dentin blocks for the first time, and (2) the effect of QADM or NAg alone or in combination, and the effect of NAg mass fraction, on S. mutans viability in dentin.
Methods
Scotchbond Multi-Purpose (SBMP) bonding agent was used. QADM and NAg were incorporated into SBMP primer. Six primers were tested: SBMP primer control, control + 10% QADM (mass%), control + 0.05% NAg, control + 10% QADM + 0.05% NAg, control + 0.1% NAg, and control + 10% QADM + 0.1% NAg. S. mutans were impregnated into dentin blocks, then a primer was applied. The viable colony-forming units (CFU) were then measured by harvesting the bacteria in dentin using a sonication method.
Results
Control + 10% QADM + 0.1% NAg had bacteria inhibition zone 8-fold that of control ( p < 0.05). The sonication method successfully harvested bacteria from dentin blocks. Control + 10% QADM + 0.1% NAg inhibited S. mutans in dentin blocks, reducing the viable CFU in dentin by three orders of magnitude, compared to control dentin without primer. Using QADM + NAg was more effective than QADM alone. Higher NAg content increased the potency. Dentin shear bond strength was similar for all groups ( p > 0.1).
Significance
Antibacterial primer with QADM and NAg were shown to inhibit the S. mutans impregnated into dentin blocks for the first time. Bonding agent containing QADM and NAg is promising to eradicate bacteria in tooth cavity and inhibit caries. The QADM and NAg may have applicability to other adhesives, cements, sealants and composites.
1
Introduction
Dental caries is a multifactorial disease in which susceptible dental hard tissues are destructed by acidic by-products from bacterial fermentation of dietary carbohydrates . Though the disease process and the potential to prevent and control the disease are now much better understood, caries is still prevalent and a heavy financial burden around the world . Resin composites are popular filling materials . The chemical and physical properties, polymerization characteristics, and placement techniques of composites have been significantly improved . Nearly 200 million dental restorations are placed annually in the U.S. . However, half of all restorations fail within 10 years, with recurrent (secondary) caries as the main reason for failure . As a result, 50–70% of all restorations were replacements of failed restorations . Replacement dentistry is a major economic burden, considering that the annual cost for tooth cavity restorations in the U.S. was approximately $46 billion in 2005 alone .
With the development of new composites and bonding agents, the concept of complete removal of carious dentin during cavity preparation is no longer supported . This is partly because that the complete caries removal approach could not really yield caries-free cavities, with often residual infection and bacteria remaining in the cavities . On the other hand, minimally invasive techniques are recommended for the treatment of deep caries lesions in order to protect pulp vitality and preserve more tooth tissues . Modern minimally invasive techniques require removing the infected-dentin to achieve its objective of least-invasive surgical approach, leaving behind the affected-dentin in the cavity. Various methods were developed to help remove caries-infected tissue as selectively as possible, while being minimally invasive through maximum preservation of caries-affected tissue . With the preservation of more affected tissue, the cavity likely will contain more residual bacteria . Therefore, it would be highly beneficial to develop antibacterial adhesives to eradicate or greatly reduce the residual bacteria in the affected-dentin and eliminate the harmful effects caused by bacteria.
Adhesives bond composite restorations to tooth structures, and extensive studies have been performed to enhance the adhesive compositions, bond strength, and durability . Antibacterial adhesives containing 12-methacryloyloxydodecylpyridinium bromide (MDPB) were developed, which effectively reduced the growth of oral bacteria such as Streptococcus mutans . Other studies developed several different types of novel antibacterial dental materials . A literature search revealed only a few reports on the development of antibacterial primers: a primer containing MDPB , a primer containing chlorhexidine , and a primer containing quaternary ammonium dimethacrylate (QADM) and nanoparticles of silver (NAg) . The primer containing QADM and NAg showed potent antibacterial effects against dental biofilms with bacteria inoculated on the surface of the cured primer and adhesive resins . However, there has been no report on the effect of the QADM–NAg primer against bacteria impregnated into dentin blocks.
Therefore, the objectives of this study were to investigate the antibacterial effects of primers containing QADM and NAg against S. mutans impregnated into human dentin blocks for the first time, and to determine the effects of different primer compositions. It was hypothesized that: (1) QADM- and NAg-containing primers would be able to inhibit S. mutans inside dentin blocks; (2) combining two antibacterial agents QADM and NAg in primer would be more effective than QADM alone, and increasing the NAg mass fraction would increase the antibacterial potency against S. mutans inside dentin; and (3) incorporating QADM and NAg into primer would not compromise the dentin shear bond strength.
2
Materials and methods
2.1
QADM and NAg incorporation into primer
Scotchbond Multi-Purpose (3M, St. Paul, MN) was used as the parent bonding system and referred as “SBMP”. The SBMP etchant contained 37% phosphoric acid. The SBMP primer contained 35–45% of 2-hydroxyethylmethacrylate (HEMA), 10–20% of a copolymer of acrylic and itaconic acids, and 40–50% of water, according to the manufacturer. The SBMP adhesive contained 60–70% of bisphenol A diglycidyl methacrylate (BisGMA), 30–40% of HEMA, tertiary amines and photo-initiator. In the present study, the primer was modified with antibacterial agents, and the adhesive was used without modification.
Bis(2-methacryloyloxyethyl) dimethylammonium bromide, a quaternary ammonium dimethacrylate (QADM), was synthesized following previous studies . The synthesis employed a modified Menschutkin reaction, in which a tertiary amine group was reacted with an organo-halide. A benefit of this reaction is that the reaction products are generated at virtually quantitative amounts and require minimal purification . Briefly, 10 mmol of 2-( N , N -dimethylamino)ethyl methacrylate (DMAEMA, Sigma, St. Louis, MO) and 10 mmol of 2-bromoethyl methacrylate (BEMA, Monomer–Polymer and Dajec Labs, Trevose, PA) were combined with 3 g of ethanol in a 20 mL scintillation vial. A magnetic stir bar was added, and the vial was stirred at 60 °C for 24 h. The solvent was then removed, yielding QADM as a clear, colorless, and viscous liquid. The QADM was mixed with the SBMP primer at a QADM mass fraction of 10%, following a previous study .
To develop primers containing NAg, silver 2-ethylhexanoate powder (Strem, New Buryport, MA) was dissolved in 2-(tert-butylamino)ethyl methacrylate (TBAEMA, Sigma) at 0.1 g of silver salt per 0.9 g of TBAEMA, following previous studies . TBAEMA was used because it improves the solubility by forming Ag–N coordination bonds with Ag ions, thereby facilitating the Ag salt to dissolve in the resin solution. TBAEMA contains reactive methacrylate groups and therefore can be chemically incorporated into a dental resin . When the Ag solution was mixed with a resin, upon photopolymerization which was accompanied with a simultaneous reduction of the silver salt, Nag were formed in situ in the resin. Previous studies showed well-dispersed NAg with a mean particle size of 2.7 nm in the cured resin . In this study, the Ag solution was mixed with SBMP primer at two different mass fractions of silver 2-ethylhexanoate of 0.05% and 0.1%, respectively, following previous studies . Six primers were tested as listed in Table 1 .
