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Table of Contents   
ORIGINAL RESEARCH  
Year : 2019  |  Volume : 30  |  Issue : 5  |  Page : 777-782
Nanoneedle-like zinc oxide as a filler particle for an experimental adhesive resin


1 Department of Conservative Dentistry, Dental Materials Laboratory - LAMAD, School of Dentistry, Federal University of Rio Grande do Sul, RS, Brazil
2 Department of Genetics, Laboratory of Stem Cells and Tissue Engineering, Brazilian Lutheran University, RS, Brazil
3 Department of Materials, Laboratory of Ceramic Materials, Federal University of Rio Grande do Sul, RS, Brazil

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Date of Submission06-Dec-2016
Date of Decision30-Jan-2018
Date of Acceptance06-Dec-2018
Date of Web Publication18-Dec-2019
 

   Abstract 


Aim: The aim of this study was to develop an experimental adhesive resin with nanoneedle-like zinc oxide (N-ZnO), an inorganic filler, that could avoid particle agglomeration and lead to a homogeneous stress distribution within the material and characterize it. Materials and Methods: N-ZnO particles obtained by a thermal evaporation technique were characterized regarding size and surface area and added at 0 (control), 1, 2, 5, and 10 wt%, to an experimental adhesive resin. The following experimental adhesive resins' properties were assessed: radiopacity, contact angle to conditioned enamel and dentin, color, degree of conversion, flexural strength, resistance to degradation, and cytotoxicity. Statistical analysis was performed using one-way ANOVA and Tukey's post hoc test and paired Student's t-test. Results: Particles presented a mean particle size of 40 nm and a specific surface area of 16 m2/g. N-ZnO10%showed an increased radiopacity when compared to N-ZnO0%. Contact angles were significantly higher for N-ZnO10%at enamel and N-ZnO2%, N-ZnO5%, and N-ZnO10%at dentin. All groups showed color change when compared to N-ZnO0%. Higher the N-ZnO concentration, lower the degree of conversion. There were no significant differences between the groups for flexural strength and resistance to degradation. The addition of N-ZnO showed no difference in cytotoxicity when compared to positive control, N-ZnO0%, and all groups showed higher values than negative control. Conclusions: N-ZnO possibly exceeded potential limitations due to particles' agglomeration and improved the transference and distribution of stress within the material. It could be effectively used as a filler for adhesive resins.

Keywords: Dentin-bonding agents, nanoparticles, zinc oxide

How to cite this article:
Leitune VC, Schiroky PR, Genari B, Camassola M, FelipeA, Samuel SM, Collares FM. Nanoneedle-like zinc oxide as a filler particle for an experimental adhesive resin. Indian J Dent Res 2019;30:777-82

How to cite this URL:
Leitune VC, Schiroky PR, Genari B, Camassola M, FelipeA, Samuel SM, Collares FM. Nanoneedle-like zinc oxide as a filler particle for an experimental adhesive resin. Indian J Dent Res [serial online] 2019 [cited 2020 Aug 14];30:777-82. Available from: http://www.ijdr.in/text.asp?2019/30/5/777/273438



   Introduction Top


The longevity of dental restorations is related to the quality of the hybrid layer formed.[1] Improvements in bond strength and mechanical properties of adhesive systems can be achieved by incorporating inorganic fillers to the adhesive resins.[2],[3] The consequences of it extend beyond mechanical effects and may further alter aspects such as curing properties and radiopacity and endow the material with antibacterial or remineralizing potential.[2],[4] The developed dental adhesives containing zinc oxide (ZnO) may increase the stability of the resin–dentine interface.[5] They have been shown to act as an matrix metalloproteinase inhibitor, preserving the remineralizing capability,[5] to inhibit dentine demineralization[6] and induce phosphate and calcium ion deposition, facilitating dentin remineralization.[7] However, spherical- and fiber-shaped fillers could present a tendency to form agglomerations instead of a homogeneous distribution.[8] These agglomerates can concentrate stress, inducing microcracks and weakening the material.[2]

Nanoneedle-like ZnO (N-ZnO) is an inorganic filler that exhibits nanoneedle-like arms extending from the same center in distinct directions within three-dimensional space.[8] This structure results in a homogeneous stress distribution into the material, which is its main reinforcement mechanism.[9] Furthermore, the needles' arms pointing to different directions induce an easier dispersion in the polymer matrix, avoiding agglomeration.[8] ZnO may also extend the bonding durability due to its antibacterial activity.[10] The aims of this study were to develop an experimental adhesive resin with N-ZnO and to investigate the influence on physico-mechanical properties.


   Materials and Methods Top


Experimental adhesive resins

Adhesive resins were obtained by mixing 50 wt% bisphenol A glycol dimethacrylate, 25 wt% triethylene glycol dimethacrylate, 25 wt% 2-hydroxyethyl methacrylate, camphorquinone, and ethyl 4-dimethylaminobenzoate (Sigma Aldrich, St Louis, MO, USA) at 1 mol%. The N-ZnO particles were obtained by a thermal evaporation technique as described previously[11] and added at different concentrations: 1, 2, 5, and 10 wt%. An adhesive resin without filler was the control. To improve the adhesion interface between filler particles and the matrix, they were submitted to the salinization process with 5% silane (Ɣ-methacryloxypropyltrimethoxysilane, Sigma Aldrich, Milwaukee, WI, USA) and 95% acetone by weight. Particles were stored for 24 h at 37°C for complete solvent evaporation. All components were weighed using an analytical balance (AUW220D, Shimadzu, Kyoto, Japan), mixed, and ultrasonicated. To perform photoactivation, the light-emitting diode unit Radii Cal (1200 mW/cm2, SDI, Australia) was used.

Particle size and surface area

A scanning electronic microscopy (TM3000 model, Hitachi High-Technologies Co., Japan) was used to evaluate the N-ZnO size and morphology. Specific surface area was determined by the Brunauer–Emmett–Teller method, using an automated gas absorption system (Quantachrome NOVA1000 Autosorb, Boynton Beach, FL, USA).

Radiopacity

Radiopacity was measured according to ISO 4049[12] standard, except for the specimens' dimensions (6.0 mm × 1.0 mm). Radiographic images were obtained using a digital system with phosphorous plates (VistaScan, Dürr Dental GmbH and Co., KG, Bietigheim-Bissingen, Germany) at 70 kV and 8 mA, with 0.4 s of exposure time and a focus-film distance of 400 mm. In each film was positioned one specimen from each group (n = 5), and an aluminum step-wedge was simultaneously exposed. Images were saved in the TIFF format and processed using Photoshop software (Adobe Systems Incorporated, CA, USA). The pixel density of the aluminum step-wedge and the specimens were obtained in a standardized area of 2 mm2.

Contact angle

Contact angles were measured by the sessile drop method using an optical tensiometer (Attension Theta; Biolin Scientific, Finland). For this, a drop of 2 μL of each experimental adhesive resin was dispensed on acid-etched and rinsed dentin (n = 5) and enamel (n = 3) slices. The drop image was captured 10 s after the deposition with a micro-video system. The contact angles were calculated using Photoshop software (Adobe Systems Incorporated, San Jose, USA).

Color measurements

A spectrophotometer (CM-2500d, Konica-Minolta, Osaka, Japan), with 10° sphere illumination geometry and specular component excluded, was used to measure the reflectance spectra from 400 to 700 nm at a wavelength interval of 10 nm. The xenon flash lamp D65 was used as light source. Discs of experimental resin (n = 3) were produced (10.0 mm × 1.0 mm). Corrected reflectivity spectra were calculated by the Kubelka–Munk reflectance model.[13] The corrected reflectivity spectra at all visible wavelengths were used to calculate CIE tristimulus (X, Y, and Z) values. CIE tri stimulus values were converted at infinite thickness to color parameters (L*, a*, and b*). The calculation of color difference between the experimental adhesive resins was carried out by equation 1 as follows:



Where L* refers to lightness, an achromatic coordinate, ranging from black (0) to white (100); a* refers to a chromatic coordinate representing the green-red axis; and b* refers to a chromatic coordinate representing the blue-yellow axis. ΔE represents the corresponding color change.

Degree of conversion

The degree of conversion was evaluated using Fourier transform infrared spectroscopy with a Vertex 70 (Bruker Optics, Ettlingen, Germany) spectrometer equipped with an attenuated total reflectance device consisting of a diamond crystal of 2-mm diameter (Platinum ATR-QL, Bruker Optics, Ettlingen, Germany). A support was coupled to fix the light-curing unit and standardize the distance between the fiber tip and sample at 5 mm. The sample (3 μL) was directly dispensed onto the diamond crystal and light activated for 20 s (n = 3). Absorbance spectra were obtained before and immediately after light polymerization using Opus 6.5 software (Bruker Optics, Ettlingen, Germany). The degree of conversion was calculated as described previously.[14]

Flexural strength

The specimens (n = 8) were fabricated in customized stainless steel molds according to ISO 4049[12] standard, except for dimensions (12 mm × 2 mm × 2 mm). The experimental adhesive resins were placed into the mold positioned on the top of an acetate strip. Each side of the specimens was then light polymerized for 20 s. Afterward, specimens were stored in distilled water at 37°C for 24 h. Flexural strength test was performed with a universal testing machine (EMIC, São José dos Pinhais, PR, Brazil) at a crosshead speed of 1.0 mm/min, and the values were calculated as described previously.[14]

Softening in ethanol

To determine the degradation in solvent, specimens (n = 5) were embedded in an acrylic resin and polished in a polisher (Model 3v, Arotec, Cotia, SP, Brazil) with a felt disc saturated with alumina suspension (Alumina, 1.0 μm, Arotec, Cotia, SP, Brazil), after the specimens were stored and dried at 37°C for 24 h. Specimens were subjected to a microhardness test in which three indentations (10 g/10 s), 100 μm apart from each other, were assessed using a microhardness tester (HMV 2, Shimadzu, Tokyo, Japan). The initial Knoop hardness number (KHN1) was registered, specimens were subjected to softening in absolute ethanol for 2 h at 37°C, then the hardness test was repeated, and the final hardness number was measured (KHN2). The percentage difference of KHN1 and KHN2 was calculated (ΔKHN%).

Cytotoxicity

Cytotoxicity was determined by cell viability (3- [4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide] [MTT] test) according to ISO 10993-5[15] standard, using mononuclear cells obtained from peripheral blood. For each experimental adhesive resin, 5 × 105 cells in 150 μl of Dulbecco's Modified Eagle's Medium (DMEM) in (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) (HEPES) (Sigma Aldrich, St Louis, MO, USA) (free acid, 2.5–3.7 g/l) – Ham's nutrient mixture F-12 and Dulbecco's modified Eagle's essential medium (HDMEM), supplemented with 10% fetal bovine serum (Cultilab, São Paulo, SP, Brazil) were used. The cells were maintained in contact with the specimens (n = 3) for 24 h at 37°C and atmospheric humidity of 5% CO2. The HDMEM with the same cell amount was used as negative control. The rate of viable cells was quantified using the MTT colorimetric assay after an incubation period of 24 h, and it was measured at 540 nm absorbance using the Multiskan EX Microplate Reader (Labsystems, USA).

Statistical analysis

Data normality was evaluated using the Kolmogorov–Smirnov test. Statistical analysis was performed using one-way ANOVA and Tukey's post hoc test at the 0.05 level of significance for all tests. For softening in ethanol, a paired Student's t-test was used too.


   Results Top


The morphology of the N-ZnO obtained is shown in the scanning electron microscopy micrograph as depicted in [Figure 1]. The mean particle size was 40 nm with 16 m2/g of specific surface area. It is possible to observe the needle shape of the nanoparticles.
Figure 1: Scanning electron microscopy of nanoneedle-like zinc oxide particles showing the nanoneedle-like structure (×40,000)

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The radiopacity values are presented in [Table 1]. The addition of 10% N-ZnO significantly increased the radiopacity of the experimental adhesive resin in comparison to N-ZnO0%(P < 0.05).
Table 1: Mean and standard deviation values of radiopacity (mmAl) and contact angles (°) at dentin and enamel for experimental adhesive resins

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The contact angles within the dentin were significantly higher with the addition of N-ZnO at 2% or more. In the enamel, the values were significantly higher for the N-ZnO10% group [Table 1].

All groups showed a color change (ΔE) when compared to N-ZnO0%, and the higher the N-ZnO concentration, the higher the ΔE (P < 0.05). Changes in the parameters L*, a*, and b* resulted in an increase in lightness and green, as well as a decrease in yellow [Table 2].
Table 2: Mean and standard deviation values of L*, a*, and b* color parameters and color differences (Δ) between the experimental adhesive resins and the control group

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The degree of conversion decreased as the concentration of N-ZnO increased (P < 0.05), ranging from 65% (N-ZnO0%) to 33% (N-ZnO10%). For flexural strength, there were no significant differences between the experimental groups and the control group (P > 0.05). The degree of conversion and flexural strength values are shown in [Table 3].
Table 3: Mean and standard deviation values of degree of conversion (%) and flexural strength (MPa) for experimental adhesive resins

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All groups showed a reduction on microhardness after the immersion in ethanol, but no difference were observed between groups for KHN1 and the percentage of reduction (P > 0.05). The softening in ethanol results are shown in [Table 4].
Table 4: Microhardness values before (Knoop hardness number) and after immersion in solvent (KHN2) and percentage of microhardness variation (ΔKHN%)

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The addition of N-ZnO showed no difference in cytotoxicity when compared to the positive control, N-ZnO0%(P > 0.05). All groups showed higher values than the negative control (P < 0.05) [Figure 2].
Figure 2: Cytotoxicity of experimental adhesive resins in absorbance at 570 nm. Different capital letters indicate significant differences (P < 0.05)

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   Discussion Top


Fillers are incorporated into adhesive resins to increase bond strength and decrease degradation over time, as well as to improve or functionalize material properties.[2],[3],[4],[16] In this study, an experimental adhesive resin with N-ZnO particles was developed and its physico-mechanical properties were evaluated. The N-ZnO structure consists of needle-like arms extending in different directions[8] with a mean particle size of 40 nm and a specific surface area of 16 m2/g. Hence, N-ZnO is supposed to enhance the interaction with resin matrix and change the stress distribution as the concentrated stress at the tip of one particle needle is transferred to the other needles.[9],[17] Radiopacity was increased by adding N-ZnO, while flexural strength, resistance to degradation, and cytotoxicity were not altered. The degree of conversion decreased, the contact angle increased, and the color changed.

The addition of inorganic particles could provide radiopacity to the adhesive resin.[4],[14] Although the experimental adhesives did not meet the ISO requirements (1 mmAl),[12] the N-ZnO10% group showed higher radiopacity than the control group. An increased radiopacity could improve the differential diagnosis with secondary caries and demineralized dentin that remains radiographically detectable under restorations after selective caries removal.[18] Radiopacity is particularly important for nanofilled adhesives that are applied in thicker layers due to their higher viscosity.[2] Higher viscosity may also decrease the wetting of dentin and enamel.[19] The addition of 10% of N-ZnO increased the contact angle on the acid-etched enamel, while for conditioned dentin, it significantly increased by adding 2% of N-ZnO. Higher amounts of filler may increase the adhesive resin's viscosity to a point that infiltration into conditioned substrate could be compromised, reducing the bond strength.[20] However, a study that evaluated a commercial adhesive with similar composition showed comparable results, with contact angles on superficial and deep-conditioned dentin of 40 and 42, respectively.[21]

According to the CIELAB system, N-ZnO induced a color change in the experimental adhesive resins. The color was mostly influenced by changes in parameter L*. These color differences can be explained by the materials' translucency. Translucency depends on light absorption and scattering that are, in turn, influenced by resin matrix composition and filler loading.[13],[22] Scattering is mainly due to the refractive index mismatch between the organic matrix and the filler particles. When the refractive indexes of both are closer, a translucent material is obtained. When they are not closer, translucency depends on the filler concentration.[22] The refractive index of the polymeric matrix used was reported to range from 1.47 to 1.59 when uncured and from 1.50 to 1.62 after polymerization,[4] which are lower than the ZnO refractive index of 2.2.[23] In addition, the higher the filler content, the higher the light dispersion and the lower the light transmittance.[24] Indeed, the color change was directly proportional to the filler loading. The enhanced scattering can also affect the curing process due to reduced light transmittance.[24] In this study, the degree of conversion decreased by increasing the N-ZnO content. When compared to commercial adhesive systems, the addition of up to 5% did not negatively alter this property.[25] A low degree of conversion may reduce the mechanical properties.[26] Uncured monomers can be released,[27] and polymer permeability and water sorption could increase,[26] affecting its resistance to degradation.[28] However, there was no difference among the groups for flexural strength and resistance to degradation.

The maintenance of flexural strength, despite the reduced degree of conversion, can be explained by the filler particles' reinforcement capability.[2] Fillers could serve as obstacles during crack extension,[10] and nanoparticles improve the mechanical properties as a consequence of stress transfer through the interfaces.[29] The N-ZnO morphology ensures a surface area that is even higher than that of spherical nanoparticles with a smaller size.[11] Moreover, an enhanced distribution of tension through particles' needles may have contributed.[9],[17] This finding also suggests that N-ZnO was homogeneously dispersed within resin matrix because agglomerations could give rise to stress concentration and local defects, compromising the flexural strength.[10],[30] With an increased filler amount, interparticle interactions become more likely, forming agglomerates with larger diameters.[2],[10],[30] Spherical nanoparticles are likely to form agglomerates, as well as fiber fillers that are usually easily entangled since their distribution is not randomly oriented.[8] The N-ZnO geometry enables random distribution because, besides its high surface area, the arms point to different directions.[8],[10] Thus, studies have demonstrated that polymers with nanoneedle particles present improved mechanical properties[8],[10],[17] and better results than that with particles presenting other geometries.[17]

It has been reported that the higher the degree of conversion, the lower the amount of softening in ethanol.[28] The N-ZnO did not influence this property. It suggests that the increased amount of unreacted monomers did not negatively compromise the resistance to degradation. This finding could be explained by the fillers' low tendency to degradation when compared to resin matrix,[16] which could have overcome the lower degree of conversion due to an increased filler content.

It is equally important for adhesive resins to be noncytotoxic, considering that they can be in close proximity to the pulp tissue, and that resin-based materials are not inert in the oral environment.[27] The presence of N-ZnO did not compromise the cytotoxicity. These findings corroborate the outcomes of other studies that tested the ZnO biocompatibility.[31]

The incorporation of N-ZnO in the adhesive resin possibly exceeded potential limitations due to the particles' agglomeration on the incorporation of spherical or fiber fillers and improved the transference and distribution of stress within the material.


   Conclusions Top


It could be concluded that, especially considering the N-ZnO particular morphology and nanosize, it could be effectively used as filler for adhesive resins. Incorporation of N-ZnO has the potential to promote the strengthening of the adhesive resin, which could improve clinical performance and restoration longevity.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
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Sanchez FA, Takimi AS, Rodembusch FS, Bergmann CP. Photocatalytic activity of nanoneedles, nanospheres, and polyhedral shaped ZnO powders in organic dye degradation processes. J Alloys Compounds 2013;572:68-73.  Back to cited text no. 11
    
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Correspondence Address:
Dr. Fabrício Mezzomo Collares
Department of Conservative Dentistry, Dental Materials Laboratory, School of Dentistry, Universidade Federal do Rio Grande do Sul, Rua Ramiro Barcelos, 2492 - Rio Branco, 90035-003 - Porto Alegre, RS
Brazil
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/ijdr.IJDR_779_16

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