Effects of Acid Etching on Dentin Surface: SEM Morphological Study

Adriana Bona MATOS
Regina Guenka PALMA
Cintia Helena Coury SARACENI
Departamento de Dentística, Faculdade de Odontologia, Universidade de São Paulo, São Paulo, SP, Brasil

Braz Dent J (1997) 8(1): 35-41 ISSN 0103-6440

| Introduction | Material and Methods | Results | Discussion | Conclusions | Acknowledgments | References |

The aim of this in vitro study was to evaluate, using scanning electron microscopy (SEM), the effects of acid solutions on dentin surface, and to analyze the depth of demineralization that the acid solutions cause on dentin, using different acids. Fifteen 3-mm thick dentin discs were prepared from the middle third of human molars. Standard smear layer was prepared on the dentin surface using 600 grit sandpaper, for 1 minute. The acids used were 10%, 35% and 37.5% phosphoric acid and 10% maleic acid, for 15 seconds, washed and dried. The control group received no treatment. Dentin discs were fractured, observed on the horizontal surface and also on the fractured surface to evaluate the depth of demineralization. Specimens were immersed in 4% glutaraldehyde in phosphonate buffer, and prepared for SEM examination at 2000X and 4000X magnification. Acid etching of dentin, regardless of the concentration of phosphoric acid, caused removal of the smear layer, exposing the apertures of dentinal tubules. This was not observed when 10% maleic acid was used. At the fractured surface, one could observe an increase in demineralization of the width of dentinal tubules, to a specific depth of about 8.19 to 11 mm, except for 10% maleic acid.

Key Words: dentin etching, acid solutions, SEM.


The idea of using phosphoric acid on dental surfaces was first introduced by Buonocore (1955), who observed that adhesion to metal surfaces by paints improved when acids were used. Various studies (Brännstrom and Johnson, 1974; Buonocore, 1956; Pashley et al., 1992, 1993; Gwinnett, 1994; Swift et al., 1995) about the use of demineralizing agents on dentin surfaces in order to improve bond strength of esthetic restorative materials have been developed.

Currently, despite great advances in restorative dentistry concerning adhesive materials, bonding between restoration and dental structures has not been totally understood or achieved. One of the main problems with dentin bonding is the presence of microleakage at the restoration/tooth interface.

Researchers found that marginal leakage still occurs when a perfect seal of the demineralized surface is not attained. Thus, a weak zone of demineralized dentin susceptible to failure is created (Sano et al., 1994).

The great challenge when using adhesive systems that indicate total etching is to obtain a dentin demineralization depth, promoted by acids, that is compatible with the depth of diffusion of hydrophilic primers. If the primers diffuse into all demineralized dentin, the bonding agent will be able to impregnate the total depth of the dentin, creating an effective hybrid layer (Nakabayashi, 1992). In the beginning, it was thought that the thicker the hybrid layer and the longer the resin tags, the better the achieved adhesion. However, it is now believed that adhesion is not attributed to long resin tags, but to a continuous hybrid layer. It is still controversial if adhesive systems penetrate the total demineralization depth of dentin (Sano et al., 1994, 1995).

It was established that acid etching agents remove smear layer to different degrees, increase the width of dentinal tubule apertures (Saraceni et al., 1994) and demineralize dentin in depth.

The purpose of this in vitro study was to verify the action of acid solutions on dentin surfaces, as well as to evaluate and measure the extent of demineralization depth, using scanning electron microscopy (SEM).

Material and Methods

Three-millimeter thick dentin discs were obtained from the middle third of extracted human molars that were immersed in saline solution. Dentin discs were prepared with diamond points at high speed with water cooling to remove coronal enamel and the roots were sectioned with Carborundum discs. Standard smear layer was obtained with 400 grit sandpaper abrasive discs, followed by 600 grit discs, for 1 minute each (Pashley et al., 1993), and maintained immersed in saline solution.

The following acids were applied to the dentin surface: 10% phosphoric acid (Aelitebond), 35% phosphoric acid (Scotchbond Multi-Purpose Plus), 37.5% phosphoric acid (Optibond), and 10% maleic acid (Scotchbond Multi-Purpose). The control group received no acid treatment. The conditioning time for all test groups was 15 seconds.

Samples were then immersed in 4% glutaraldehyde in phosphonate buffer (0.075 M, pH 7.3) for 12 hours, under refrigeration to perform collagen fixation. Samples were fractured, submitted to dehydration procedures using different ethanol graduations until 100%. The critical point was obtained and samples were gold sputtered.

Prepared dentin discs were observed by scanning electron microscopy (SEM) at 2000X and 4000X magnification and then photographed.

Photomicrographs were observed from the horizontal view to analyze the smear layer removal and from the cross-section view (fractured surface) to measure depth of demineralization.


The control group was untreated (Figure 1). Photomicrographs showed removal of the smear layer and demineralization of dentinal tubule apertures with all three concentrations of phosphoric acid (Figures 2,3,4), but 10% maleic acid (Figure 5) only removed the superficial smear layer (Table 1).

Figure 1 - Control group (2000X)

Figure 2 - Dentin surface after etching with 10% phosphoric acid (top, 2000X; bottom, 4000X).

Figure 3 - Dentin surface after etching with 35% phosphoric acid (top, 2000X; bottom, 4000X).

Figure 4 - Dentin surface after etching with 37.5% phosphoric acid (top, 2000X; bottom, 4000X).

Figure 5 - Dentin surface after etching with 10% maleic acid (top, 2000X; bottom, 4000X).


Acid conditioning of dental structures was introduced by Buonocore (1955), who tested application of phosphoric acid on enamel surface. This procedure provided an increase of microscopic surface for bonding, resulting in micro-mechanical retention of restorations to dental structures, rather than chemical adhesion. This procedure is still a fundamental step during esthetic restorative treatment.

Buonocore et al. (1956) was again the first to use acid on dentin surface trying to improve adhesion of restorative materials, using 7% chlorhydric acid for 1 minute. He did not achieve good results with this technique, but it is very important to observe that, at that time, the esthetic restorative materials available were mostly acrylic resin and silicate cement, completely different from current ones. The unsuccessful pulpal result was attributed to acid etching and dentin conditioning was contraindicated.

Only a few years later, when Bowen (1963) developed Bis-GMA molecule studies about dentinal bonding were started again. Acid conditioning of enamel was recognized and considered an established procedure. These former adhesion principles were based on chemical bonding; however, since the bond strength was weak and easily suffered hydrolysis, the results were not satisfactory.

Studies aimed at the development of an effective dentin bonding agent (DBA), and one could observe the alterations. The most usual classification of DBA divided them into generations. The fourth generation of DBA, which is the current one, indicates acid etching of enamel and dentin at the same time, a technique usually called “total etching” (Kanca, 1992), application of a hydrophilic primer, and, as the last step, application of the bonding resin. It is important to point out that it is not yet understood that the achieved adhesion is promoted by micro-mechanical retention, and not by chemical means.

The dentin bonding system is applied on the dentinal surface and it is expected to form a hybrid layer. This hybrid layer was first described by Nakabayashi and was defined as formed by the definitive interlocking of collagen fibers and the dentin bonding agent. This layer was supposed to be continuous and to prevent microleakage (Nakabayashi, 1992; Tay et al., 1995)

Smear layer is composed by debris of cut dental hard tissues, saliva, bacteria, oil from high- and low-speed handpieces, and is formed every time dentin is cut or abraded. It is related to the use of rotatory and manual instruments during cavity preparation and its thickness varies around 5 mm, depending on the type of instrument used (Eick, 1992; Pashley, 1992). Smear plugs have the same composition as the smear layer and are located inside dentinal tubules. To form a hybrid layer it is necessary to remove superficial smear layer and also smear plugs.

Acid etching was defined as any change promoted on dentin after smear layer formation (Bertolotti, 1992).

After dentin conditioning, this tissue becomes mineral poor and rich in proteins, with exposes collagen and increases fluid flow and permeability (Gwinnet, 1994). Some objectives of acid etching are: to remove the weakness of smear layer, in order to promote bonding to subjacent dentin matrix; to demineralize dentin matrix to provide infiltration of resin; to expose inter- and peritubular dentin; and to clean dentin surface, that becomes moisture free (Pashley et al., 1992, 1993).

Analyzing dentin conditioning agents, acid solutions have been considered chemical agents. It is known that any acid solution shows all ideal characteristics of an etching solution, but it is important to know these characteristics in order to choose a solution which is closest to the ideal one. To consider an etching agent ideal, it is necessary to have several features, such as: be isotonic; have neutral pH; be non-toxic to the pulp and gingival tissues; be chemically compatible to restorative material and be able to chemically improve dentin surface, preparing it for adhesion (Pashley et al., 1992; Bertolotti, 1992).

Another factor that must be considered is the diffusion of acid solution through the dentin surface and, consequently, its etching capacity. This factor depends on the viscosity of the acid solution and the molecular weight of the acid. Related to viscosity, Pashley et al. (1992) state that gel solutions, applied for the same time and at the same concentration as a liquid solution, provides less etching effects. Thus, for a gel solution to produce the same etching pattern as a liquid solution, a longer application time is necessary. The same authors state that solution diffusion is inversely proportional to molecular weight, for example, polyacrylic acids diffuse less than acids with lower molecular weight, such as, phosphoric and nitric (Pashley et al., 1992).

In this study, it was observed that different concentrations of phosphoric acid demineralize dentin surface to different degrees. Greater demineralization was found with concentrations of 10% and 35% and less demineralization with 37.5%, in accordance with the results of Bertolloti (1992) in relation to 10% and 35% concentrations. Although, Gwinnett (1994) observed that 10% phosphoric acid applied for 20 seconds produced tissue modifications of approximately 5 mm, this study pointed to a mean demineralization extent of 10.19 mm, with the acid solution being applied for 15 seconds. Phosphoric acid provides a clean and well defined etching pattern, with increased width of funnel-shaped dentinal tubules (Saraceni et al.,1994; Perdigão and Swift, 1994).

Maleic acid removed the smear layer, but not smear plugs. Our results are in accordance with those of Bertolotti (1992) and Gwinnett (1994). We also found that this acid did not demineralize dentin in its depth. Other studies agree with our results, adding that maleic acid can provide an alteration of the tissue of approximately 1 mm when applied for 15 seconds (Bertolotti, 1992; Gwinnett, 1994).


Observing from the horizontal view, all acid solutions used removed superficial smear layer, increasing the width of dentinal tubule apertures to greater or lesser degrees. When observing the fractured surface, acid solutions provided a mean demineralization in depth of dentinal tubules of 9.83 µm, except for maleic acid.


We acknowledge the discipline of Oral Pathology of the School of Dentistry of the University of São Paulo, and the School of Medicine for the use of the SEM equipment. We thank André Guilherme Jorge and Prof. Dr. Edson Liberti who performed the scanning electron microscopy and photomicrographs of the samples.


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Correspondence: Prof. Dr. Edmir Matson, Departamento de Dentística, Faculdade de Odontologia, Universidade de São Paulo, 05508-900, São Paulo, SP, Brasil.

Accepted January 6, 1997
Electronic publication: September, 1997