The aim of endodontic treatment is to eliminate microorganisms and their byproducts from tooth root canal system and also establish a durable seal all along the canals to prevent recontamination. Gutta-percha is the most commonly used material to fill the canals. It is a solid material and cannot provide a full adaptation to root canal walls. Therefore,  gutta-percha is used as a core and endodontic sealers are used to fill the irregularities and penetrate into dentinal tubules to provide proper fluid-tight (1). Sealer penetration to dentinal tubules has three major benefits: first, it increases the contact surface between filling material and dentin improving sealing ability (2). Second, this interlocking may enhance mechanical retention of the filling material (3-5). And third, it can prevent bacteria from colonizing within the tubules by separating them from nutritional resources (4, 6, 7). Measuring dentinal tubule penetration of endodontic sealers is used to evaluate their performance and sealing ability (4). Endodontic sealers ability to enter tubules is affected by their physiochemical properties (8, 9). There are evidences showing that smear layer plays a negative role in sealer penetration by occlusion of dentinal tubules (3, 10, 11). So, a sealer with good flow rate, film thickness, surface tension, solubility and viscosity may be able to penetrate into dentinal tubules if they are open (12).

Nano-ZnO sealer is a ZnO-based endodontic sealer that has nanoparticles in its composition. Nanoparticles are of interest in endodontics because of their tubular penetration, antimicrobial activity, and reduced leakage plus enhanced physiochemical properties (13). As far as we know, there is no study on penetration depth of this new nanoparticle sealer. Therefore, the purpose of this study was to evaluate the tubular penetrability of Nano-ZnO sealer in comparison with AH26 (a commonly used resin-based sealer) and Pulp Canal Sealer EWT (another ZnO-based sealer without nanoparticles) by means of scanning electron microscopy. The null hypothesis (H₀) was that none of these sealers penetrate into dentinal tubules.

Materials and Methods

Twenty human single-rooted premolars were selected and immerged in NaOCl 5.25% for 1 hour to remove all organic debris from root surface. Mesio-distal radiographs were taken to ensure all teeth have one canal. Teeth were decoronated by a diamond disk (Jota AG-Ruthi Switzerland) to standardize the root length to 12 mm. A #15 k-file (Dentsply/Maillefer, Ballaigues, Switzerland) was inserted through the canal until it was seen at the apical foramen. Working length was determined 1 mm shorter than this length. Root canals were prepared by a crown-down technique using NeoNiTi rotary files (Neolix, châtres-la-Forêt, France) according to the manufacturer’s instructions: the coronal one-third was prepared by a NeoNiTi C1 file (25/.12) and the middle and apical one-thirds were prepared by a NeoNiTi A1 file (25/.08). The electric motor (NSK, Japan) was used at the speed of 350 rpm and torque of 1.5 NCm according to the manufacturer’s instructions. Canals were irrigated between files using 1 ml of NaOCl 5.25%.  In order to remove the smear layer, 1 ml of EDTA 17% was applied for 1 minute followed by 3 ml of NaOCl 5.25% at the end of the instrumentation. 3 ml of normal saline was used as final irrigation and canals were dried using sterile paper points (Meta Biomed, Chungbuk, Korea).

Then, teeth were randomly divided into three groups of 6 teeth in each group as follows:

Group 1. Teeth were obturated with gutta-percha and the resin-based sealer AH26 (Dentsply, DeTrey, Germany).

Group 2. Teeth were obturated with gutta-percha and Pulp Canal Sealer (Sybron Kerr, Romulus, MI, USA).

Group 3. Teeth were obturated with gutta-percha and Nano-ZnO sealer.

Two teeth were sliced longitudinally and examined under the microscope to ensure the removal of the smear layer.

All 3 sealers were prepared according to the manufacturer’s instruction on a glass slab with a metal spatula until a homogenous consistency was obtained. A lateral obturation technique was used in a way that the canal walls were coated with sealer by using a size B spreader (Dentsply/Maillefer, Ballaigues, Switzerland). A #25 master gutta-percha cone with 0.02 taper (Sure Dent, Korea), coated with sealer was inserted into the canal followed by sealer coated accessory cones until the entire canal was filled. The excess gutta-percha was removed by a heat-carrier and then orifices were sealed by 2-3 mm of Cavit (Coltosol, Ariadent, Tehran, Iran). All specimens were incubated for 72 hours at 37^ᵒc and 100% humidity to ensure sealers setting were completed.

A low-speed, water-cooled, automatic cutting machine CNC (Nemo, Mashhad, Iran) was used to section teeth at 2 levels (4 and 8 mm from the apex) horizontally. Specimens were then sonicated in distilled water to remove smear layer. In order to prepare specimens for SEM examination, we used Perdigao et al. (14) method:

1. Specimens were placed in a composition of glutaraldehyde 2.5%, paraformaldehyde 2% and phosphate buffer (pH=7.4) for 12 hours and dried with hexamethyldisilazane.
2. Then specimens were embedded in epoxy resin (AXON, France) with mixing ratio of 100 to 32.
3. After 48 hours, the surfaces of specimens were polished with 180 to 3000 grit sandpapers followed by a polishing wool pad
4. After placement in ethanol 100% for 5 minutes, specimens were placed in HCl 6% for 30 seconds to get demineralized and then placed in NaOCl 1% for 10 minutes to remove all organic debris.
5. Specimens were kept in ultrasonic cleaner to remove surface contaminations.
6. All specimens were mounted on aluminum stubs and coated with gold and palladium by using an Emitech SC7620 Sputter Coater (Quorum Technologies, Laughton, East Sussex, UK) for 3 minutes.

At the end, specimens were evaluated under a scanning electron microscope (LEO1450VP, Carl Zeiss AG, Oberkochen, Germany) at 20 kv and 205 nm resolution.

First, specimens were examined at ×200 magnification to see the whole sample surface. Then the area with the deepest sealer penetration was selected and examined at ×500 and ×5000 magnifications.

Results

The normal distribution of data was assessed using the Shapiro-Wilk tests. The data only in AH26 group in Coronal and Apical had a normal distribution (P>0.05).

Table I shows penetration depth of three sealers.

There was no statistically significant difference between sealers penetration depth in apical section (P-value = 0.302)

Table I. Comparison of penetration depth (µm) of 3 sealers in 2 different zones