Keywords
resin infiltrant - fluoride varnish - microhardness - white spot lesions - minimally
invasive dentistry
Introduction
Dental caries is one of the most common chronic diseases globally, affecting individuals
of all ages and constituting a significant public health issue due to its high rates
of occurrence and its profound impacts on individuals and society.[1]
[2] White spot lesions represent the initial clinical manifestation of dental caries,
characterized by the initial demineralization of enamel in both the surface and subsurface
layers. This condition arises from plaque accumulation in areas prone to stagnation,
particularly in individuals with inadequate oral hygiene. If demineralization activity
is not stopped, it can progress to cavitation, which, if left untreated, may result
in premature loss of primary teeth, which can compromise arch integrity by causing
space loss for the eruption of cuspids and bicuspids and, ultimately, the onset of
malocclusion. In permanent teeth, the caries can advance to the pulp, causing pain
and infection. In advanced stages, dental caries may require more invasive procedures
such as root canal treatment or extraction.[3]
[4]
[5] White spot lesions, also known as initial caries, early enamel caries, or smooth
surface caries, exhibit an opaque white appearance due to the mineral's loss in the
subsurface enamel and altered light reflection as compared with intact enamel.[6] According to Puleio et al,[7] white spot lesions occur in approximately 46% of individuals with fixed orthodontic
devices, primarily resulting from plaque retention around the orthodontic brackets
and bands.
Recently, noninvasive remineralization strategies have been employed to halt the progression
of caries and render the lesion inactive. For decades, fluoride has served as a fundamental
element in the enamel remineralization process. It is widely recognized for its caries-preventive
properties, as it halts demineralization at the enamel surface and facilitates the
formation of fluorapatite. Fluorapatite exhibits lower solubility than hydroxyapatite,
enhancing the enamel's resistance to acid assault.[8] Furthermore, fluoride acts as a mineral reservoir, providing buffering action and
serving as a protective barrier that limits acid contact with the underlying enamel.[9] Topical fluoride treatment, in different formulations, is considered an effective
strategy for halting the progression of carious lesions, supporting its role in preventive
dentistry.[10]
Resin infiltration is an innovative material in minimally invasive dentistry that
effectively bridges the gap between preventive and restorative treatments for carious
lesions that reach the first third of dentin. It is commonly used to treat discolored
enamel resulting from demineralization or developmental defects in enamel. It serves
as a therapeutic approach for incipient carious lesions on both the proximal and smooth
surfaces of primary and permanent teeth. The main features of resin infiltration include
its noninvasive nature, preservation of tooth structure, and the ability to be completed
in a single visit. It halts the advancement of demineralized lesions by sealing the
pores and masking the lesions, limiting future bacterial penetration.[11] In relation to the impact of resin infiltration and fluoride varnish on the microhardness
of white spot lesions across different storage time intervals, no Iraqi study had
been identified. Therefore, the objective of the current study is to assess the effect
of resin infiltrant on the microhardness of white spot lesions across different storage
time intervals in artificial saliva in comparison to the fluoride varnish.
Materials and Methods
Sample Size Determination
Sample size calculation was performed using G*Power version 3.0.10, as described by
Faul et al.[12] The parameters included a statistical power of 85%, a significance level of 0.05,
and an effect size (F) of 0.25. According to Cohen's conventional classification, this effect size corresponds
to a medium effect.[13] Based on these inputs, the required total sample size was calculated to be 36 teeth
(6 teeth per group per time interval).
Sample
The present study utilized 36 intact, caries-free premolar teeth that were extracted
for orthodontic purposes, excluding those with cracks, restorations, or any other
defects. The extractions were performed with care to minimize trauma, and the teeth
were subsequently rinsed with deionized water and wiped with acetone to remove any
debris. After that, the teeth were polished with rubber cups using nonfluoridated
pumice and a conventional low-speed handpiece. Each tooth was then inspected under
the magnifying lens to ensure there were no fractures or cracks. The samples were
kept in 20 mL of deionized water containing 0.1% thymol at room temperature until
use. This solution helped prevent bacterial and fungal growth while minimizing enamel
brittleness.[14]
[15]
Sample Preparation
A 6-mm diameter circular window was standardized on the buccal side of the samples
using an orthodontic ruler. To identify the middle area of the buccal surface, two
imaginary lines were drawn: the first line extending from the tip of the buccal cusp
to the cervical line and the other line connecting the mesial to the distal surface
of the tooth at the most pronounced curvature of these teeth. Following that, a circular
adhesive tape measuring 6 mm was cut and affixed to the sample buccal surface. The
samples were then coated with an acid-resistant nail polish. Afterward, the adhesive
tape was removed, creating a circular window on the buccal surface of each sample.
Grit paper (400) was positioned within a specialized manual device, and the exposed
area of each sample was ground and polished in a single direction 10 times. This method
facilitated the attainment of a flat surface on each sample suitable for microhardness
measurement.[16]
White Spot Lesion Formation
The pH-cycling procedure was employed to induce white spot lesion on the enamel of
each sample. The demineralizing solution was prepared by combining 0.075 M/L acetic
acid, 1.0 M/L calcium chloride, and 2.0 mM/L phosphate chloride, with the pH adjusted
to 4.3 at 37°C. The remineralizing solution was prepared by mixing 0.9 mM/L potassium
phosphate, 1.5 mM/L of calcium nitrate, and 150 mM/L potassium chloride, with a pH
of 7 at 37°C. The samples were submerged in 20 mL of the demineralization solution
for 6 hours at 37°C in an incubator. After this period, the samples were washed with
de-ionized water for 1 minute. The samples were then submerged in a remineralizing
solution for 17 hours at 37°C and stored in an incubator. This entire procedure was
replicated on a daily basis for a period of 10 days. The samples were then thoroughly
rinsed with de-ionized water, after which the sample was examined using a stereomicroscope
to detect any microscopic alteration related to caries development, which may be visually
detected as white spots upon drying.[17]
Treatment Groups after White Spot Lesion Formation
In this study, 36 tooth samples were utilized and assigned into two groups (n = 18): one group treated with resin infiltrant and the other with fluoride varnish.
Each group was then subdivided into three subgroups (n = 6) based on storage time in artificial saliva (1-week, 2-week, and 4-week). The
resin infiltrant was applied following the manufacturer's instructions as follows:
Icon-etch (15% HCL gel) was applied first for 2 minutes and then air-dried for 30 seconds.
The second step involved the application of Icon-dry (99% ethanol) for 30 seconds,
followed by air-drying. The final step involved the application of Icon-infiltrant
for 3 minutes, which was then light-cured for 40 seconds using a Woodpecker LED.Q
curing unit (Guilin Woodpecker Medical Instrument Co., Ltd., Guilin, China) with a
light intensity of 1,000 to 1,200 mW/cm2, before the treated samples were stored in artificial saliva ([Fig. 1]). In the second group, fluoride varnish was applied as a thin layer over the circular
window on the buccal surface of each tooth sample ([Fig. 2]). The detailed composition and manufacturer information of the materials used in
this study are presented in [Table 1].
Fig. 1 Application process of the resin infiltrant following the manufacturer's instructions.
Fig. 2 Application of fluoride varnish on the circular window of the buccal surface.
Table 1
Composition and manufacturer information of the materials used in the study
|
Material
|
Manufacturer
|
Composition
|
|
Resin infiltrant
|
DMG, Hamburg
(Germany)
|
Icon-etch: 15% hydrochloric acid.
Icon-dry: 99% ethanol (alcohol).
Icon-infiltrant: triethylene glycol dimethacrylate.
Initiators and stabilizers
|
|
Fluoride varnish (FluoroDose)
|
Centrix Inc. (United States)
|
5% Sodium fluoride (NaF) or 22,600 ppm of fluoride, Beeswax, coloring agent.
|
|
Artificial saliva
|
Prepared in laboratory
|
0.030 g glucose, 0.580 g NaCl, 0.002 g ascorbic acid, 0.170 g CaCl2, 0.330 g KH2PO4, 0.340 g Na2HPO4, 0.160 g NaSCN, 0.160 g NH4Cl, 1.270 g KCl, 0.200 g urea, and 2.700 g mucin in 1,000 mL distilled water.
|
Abbreviations: CaCl2, calcium chloride; KCl, potassium chloride; KH2PO4, potassium dihydrogen phosphate; Na2HPO4, sodium hydrogen phosphate; NaCl, sodium chloride; NaSCN, sodium thiocyanate; NH4Cl, ammonium chloride; ppm, parts per million.
Microhardness Testing
Microhardness measurements were conducted utilizing a Vickers microhardness testing
device (Model: TH715, SN: 0003, Beijing Time High Technology Ltd, China) in the Department
of Material Engineering at Technology University ([Fig. 3]). A load of 500 g for 30 seconds was used.[18] The enamel microhardness was initially measured at the baseline, followed by assessment
after induction of carious lesions using the pH cycling method. Finally, measurements
were taken after the samples were treated with study materials at various storage
times (1-week, 2-week, and 4-week). Three Vickers indentations were performed on each
sample, and the hardness value was calculated by averaging three readings.[19]
Fig. 3 Vickers microhardness testing device.
Statistical Analysis
Statistical analysis was performed using Statistical Package for the Social Sciences
(SPSS) version 28, with the significance level set at p ≤0.05. Data are shown as means ± standard deviations. A paired t-test was employed to compare the mean enamel microhardness at the baseline, after
demineralization, and after treatment for each group at each time interval. An independent
sample t-test was used to compare the effect of the materials after the treatment.
Results
The mean microhardness values of intact enamel, subsequent to demineralization through
the pH cycling procedure and following the application of resin infiltrant, are represented
in [Table 2]. A statistically significant reduction in the enamel microhardness was observed
following the demineralization process for all storage time intervals. Subsequently,
an elevation in the mean microhardness values was recorded following the treatment
with resin infiltrant at various storage times, which was statistically significant
([Fig. 4]). The values of the paired t-test are seen in [Table 3].
Table 2
Microhardness mean values and standard deviation of enamel surface treated with resin
infiltrant at different storage times
|
Phases
|
1-week
mean ± SD
|
2-week
mean ± SD
|
4-week
mean ± SD
|
|
Sound enamel
|
303.6 ± 26.7
|
301.1 ± 11.7
|
295.3 ± 27.3
|
|
Demineralization
|
94.4 ± 72.9
|
87.7 ± 42.7
|
132.9 ± 54.6
|
|
Posttreatment
|
154.1 ± 83.6
|
185.1 ± 65.7
|
243.4 ± 71.05
|
Abbreviation: SD, standard deviation.
Fig. 4 Microhardness of the resin infiltrant at different storage times for the baseline,
pH cycling, and after treatment.
Table 3
Paired t-test among phases of microhardness for resin infiltrant group
|
Phases
|
1-week
|
2-week
|
4-week
|
|
MD
|
p-Value
|
MD
|
p-Value
|
MD
|
p-Value
|
|
Sound × demineralization
|
209.13
|
0.001
|
213.4
|
0.001
|
162.4
|
0.001
|
|
Sound × treatment
|
149.5
|
0.001
|
116
|
0.001
|
51.9
|
0.001
|
|
Demineralization × treatment
|
−59.7
|
0.001
|
−97.3
|
0.001
|
−110.5
|
0.001
|
Abbreviation: MD, mean difference.
Note: Paired t-test. p-Value <0.05 is considered significant.
The microhardness means values for sound enamel, subsequent to the pH cycling procedure
and after treatment with the fluoride varnish, are presented in [Table 4]. A statistically significant reduction in the microhardness was observed following
the pH cycling procedure. Conversely, a statistically significant increase in microhardness
was noted after the treatment for all storage time intervals ([Fig. 5]). The values of the paired t-test are shown in [Table 5].
Table 4
Microhardness mean values and standard deviation of enamel surface treated with fluoride
varnish at different storage times
|
Phases
|
1-week
mean ± SD
|
2-week
mean ± SD
|
4-week
mean ± SD
|
|
Sound enamel
|
262.8 ± 62.3
|
257.1 ± 79.4
|
252.1 ± 40.1
|
|
Demineralization
|
66.7 ± 28.6
|
141.2 ± 53.9
|
69.6 ± 32.2
|
|
Post treatment
|
140 ± 38.4
|
186.4 ± 46.4
|
129.8 ± 78.8
|
Abbreviation: SD, standard deviation.
Fig. 5 Microhardness of the fluoride varnish at different storage times for the baseline,
pH cycling, and after treatment.
Table 5
Paired t-test among phases of microhardness for fluoride varnish group
|
Phases
|
1-week
|
2-week
|
4-week
|
|
MD
|
p-Value
|
MD
|
p-Value
|
MD
|
p-Value
|
|
Sound × demineralization
|
196.1
|
0.001
|
115.9
|
0.001
|
182.5
|
0.001
|
|
Sound × treatment
|
122.8
|
0.001
|
70.70
|
0.001
|
122.3
|
0.001
|
|
Demineralization × treatment
|
−73.3
|
0.001
|
−45.22
|
0.001
|
−60.18
|
0.001
|
Abbreviation: MD, mean difference.
Note: Paired t-test. p-Value <0.05 is considered significant.
After treatment, an independent t-test revealed that 4-week remineralization period showed statistically significant
differences between study materials (p < 0.001). However, no significant differences were found at 1-week and 2-week remineralization
periods (p = 0.5 and p = 0.9), respectively ([Table 6] and [Fig. 6]).
Table 6
Independent sample t-test between groups after intervention
|
Groups
|
Time intervals
|
MD
|
p-Value
|
|
Resin infiltrant
|
1-week
|
14.1
|
0.5
|
|
Fluoride varnish
|
|
Resin infiltrant
|
2-week
|
−1.3
|
0.9
|
|
Fluoride varnish
|
|
Resin infiltrant
|
4-week
|
113.6
|
<0.001
|
|
Fluoride varnish
|
Abbreviation: MD, mean difference.
Note: Independent sample t-test. p-Value <0.05 is considered significant.
Fig. 6 Microhardness of enamel surface for both groups after treatment.
Discussion
Preventing dental caries is more crucial than its treatment. In children and adolescents,
preventive strategies can effectively halt early caries development and facilitate
the remineralization of damaged dental surfaces. The use of preventive materials to
remineralize the white spot lesions and early enamel decay can slow or even prevent
the development of cavities while maintaining the structural integrity of the tooth.
Microhardness measurement is commonly used as a criterion to evaluate the effectiveness
of various remineralizing agents in halting demineralization and reversing lesions.[20] In the present study, carious lesion initiation was achieved using the pH cycling
method, which has been validated in previous studies[17]
[21] for its ability to induce enamel demineralization. This method is widely recognized
for its simplicity, reliability, and effectiveness in simulating the development of
dental caries. A statistically significant reduction in the enamel microhardness was
noted following the pH cycling in both groups, indicating enamel demineralization
and the initiation of a carious lesion. When the pH of the surrounding environment
drops below the critical threshold of 5.5, it creates an acidic environment that promotes
demineralization by causing the loss of calcium and phosphate ions. This process leaves
micropores in the enamel, resulting in decreased hardness.[22] Following demineralization, studies[23]
[24] have shown that enamel microhardness significantly decreases while lesion depth
increases due to mineral loss. Conversely, after applying remineralizing treatments,
an increase in surface microhardness and a corresponding reduction in lesion depth
have been observed, reflecting the restoration of mineral content and enamel integrity.
Enamel microhardness was evaluated for each material across different time intervals.
Each material was analyzed separately over time, and comparisons were made between
materials at each time point. Therefore, the results reflect main effects only.
In the group treated with resin infiltrant, the results demonstrated a statistically
significant elevation in the enamel mean microhardness compared with the demineralized
stage for all time intervals. This is due to the inherent capacity of the resin infiltrant
to penetrate the microporosities present in the demineralized enamel, facilitated
by its low viscosity, which creates a diffusion barrier within the enamel lesion.
This process ultimately leads to rehardening of the demineralized tissue, thereby
enhancing its mechanical integrity and preventing cavitation. These outcomes aligned
with the studies conducted by Torres et al and Paris et al.[25]
[26] However, the microhardness did not reach the baseline level, a result that is in
accordance with the finding of Neres et al.[27] This may be due to certain regions of demineralized enamel that remained unfilled
by the resin during the polymerization process, a phenomenon attributed to the material
shrinkage, as discussed by Torres et al.[25] Additionally, the organic matrix of resin infiltrant, predominantly consisting of
triethylene glycol dimethacrylate (TEGDMA), may not always form a consistent polymer
chain from this monomer.[28] Furthermore, the mechanical characteristics of this monomer are comparatively inferior
to those of alternative monomers; a condition resulting from the lack of strong intermolecular
secondary bonds and aromatic structure.[29] The results of the present study demonstrated that the resin infiltrant group showed
the highest microhardness at 4-week followed by 2-week, while it provided lowest microhardness
at 1-week. This can be explained by the ability of the TEGDMA monomer of resin infiltrant
to continue polymerization even after the initial curing, which can increase hardness
over time. This post-curing polymerization effect strengthens the infiltrated enamel
by creating a more cross-linked polymer matrix, peaking at around 4-week.[30] These findings are in contrast with the findings of Pintanon et al,[31] who concluded that resin infiltrant provides an immediate increase in the microhardness
of white spot lesions, which then decreases over time.
When sodium fluoride varnish was applied, a notable increase in mean microhardness
values was observed compared with the demineralized phase. This can be attributed
to the chemical interaction between the fluoride ion and the enamel surface, resulting
in the formation of fluoride-containing compounds, primarily calcium fluoride. These
compounds serve as reservoirs of fluoride ions, subsequently hindering the further
dissolution of enamel during an acidic attack.[32]
[33] The results of the present study aligned with the findings of Kamal et al and Oliveira
et al.[34]
[35]
After treatment, there were no significant differences between the two groups at both
1-week and 2-week intervals. This may be due to the short duration, which is not enough
to show a clear difference between the materials, or it may be due to the physical
or chemical properties of each material, which may lead to similar effects in the
early stages, as both may interact with the surface in the same ways, without deep
penetration or sufficient interaction to show a significant difference. However, at
the 4-week mark, a significant difference with resin infiltrant demonstrating a greater
effect, this finding is aligned with the result of Aziznezhad et al,[36] who found that surface microhardness was significantly higher after treatment with
resin infiltrant compared with fluoride varnish. Furthermore, the study conducted
by Manav Özen et al[37] supports the long-term effectiveness of resin infiltrant in improving enamel surface
hardness compared with fluoride-based treatment. This can be explained by the formation
of homogenous resin–hydroxyapatite complex, exhibiting enhanced surface hardness.
The formation of resin–hydroxyapatite complex occurs through the encapsulation of
hydroxyl-apatite crystals by resin infiltration material.[38] In contrast, the fluoride varnish exhibited a decrease in effectiveness, as its
effect gradually disappeared when the applications stopped, necessitating regular
reapplication to maintain its effects.[39] Although the study by Kashash et al[40] focused on color change, it similarly showed that fluoride varnish had less long-term
effectiveness compared with resin infiltration. In our study, while microhardness
showed no significant difference between materials during the first 2 weeks, a significant
improvement was observed with resin infiltration at 4 weeks, supporting the concept
that resin infiltration provides better long-term outcomes in both structural and
esthetic aspects.
Conclusion
In conclusion, both resin infiltrant and fluoride varnish positively contributed to
the enhancement of the enamel microhardness. However, resin infiltrant demonstrated
a more consistent increase in the microhardness over time, indicating its potential
for effective long-term remineralization.
