Shinkai et al.

Asian Pac J Dent 2014; 14: 1-6

Effects of CO2 laser and fluorides application on root
demineralization
Koichi Shinkai, DDS, PhDa and Shiro Suzuki, DDS, PhDba

Department of Operative Dentistry, The Nippon Dental University School of Life Dentistry at Niigata,b
Niigata, Japan, and Department of Prosthodontics, School of Dentistry, University of Alabama at
Birmingham, AL, USA
Purpose: The effects of CO2 laser irradiation and fluoride application on the acid resistance of tooth root
surfaces were evaluated.
Materials and Methods: A total of 35 extracted human premolar roots were completely coated with
acid-resistant varnish except for a window (2x3 mm) on the proximal surface. The roots were assigned to oneof seven treatment groups as follows: CO2 laser irradiation at 17 J/cm2 (Group 1); irradiation at 25 J/cm2 (Group2); irradiation at 41 J/cm2 (Group 3); 200-minute soak in 0.05% NaF solution (Group 4); 50-minute soak in 0.2%
NaF solution (Group 5); 5-minute application of 2.0% NaF solution (Group 6); and no treatment (control). The
specimens were subjected to a 2-day pH-controlled cyclic demineralization/remineralization process. The
lesion depth on each sample was measured using polarized-light microscopy.
Results: The lesions in groups 3, 4, 5, and 6 were significantly shallower than the lesions in the control
specimens (p<0.01), but there were no significant differences between groups 1 and 2 and the control, or
between groups 1, 2, 3, and 4 (p>0.05). The lesions in groups 5 and 6 were significantly shallower. Group 3
exhibited significantly reduced lesion depth compared to the control.
Conclusion: Fluoride application significantly improved the acid resistance of the root surface. CO2 laser
irradiation at an energy density of 41 J/cm2 improved the acid resistance of the root surface, while CO2 laser
irradiation at energy densities of 15 and 27 J/cm2 had no beneficial effect.
(Asian Pac J Dent 2014; 14: 1-6.)
Key Words: fluoride, laser, tooth demineralization/remineralization, tooth root

Introduction
The elderly population has been steadily increasing, and development of periodontal treatments and dental
home-care techniques has contributed to ensure that individuals retain their natural teeth as they age.1

Duringaging, the root surfaces of teeth gradually become exposed to the oral environment due to gingival recession andconsequently become susceptible to dental caries.2,3

The critical pH at which decalcification occurs on root

surface is the same as for deciduous teeth, and tooth decay in this area is a common problem in older patients.
The use of CO2 laser irradiation to prevent enamel or dentin caries has been well described,4-13 and argon
lasers have also been reported to be effective in enhancing the demineralization resistance of root surfaces.14,15
However, few studies have demonstrated the effectiveness of CO2 laser irradiation in preventing root surface
demineralization.16

Furthermore, the effect of energy density in CO2 laser irradiation on root demineralization

has not been confirmed in previous studies.
It is well known that the application of fluoride to enamel surfaces increases resistance todemineralization,17-23 and several studies have described the effect of fluoride application on root surface
demineralization.24-29

One in vitro study examined the demineralization of human molar roots following

exposure to pH 5.5 buffered solutions containing various concentrations of fluoride, calcium, and phosphate.
The rate of demineralization decreased with the increase of fluoride concentration.23

Acidulated phosphatefluoride (APF) treatment significantly enhanced the resistance to demineralization of root surfaces during invitro acidogenic exposure.24
root surface treatments.

The fluoride concentration and application period may influence the efficacy of

Although these aspects are important, they have not been investigated in previous

studies.

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Therefore, the present study examined the effects of energy density of CO2 laser and fluoride concentrationon the depth of root surface lesions occurring after pH cycling.

The null hypothesis was that the CO2 laser

energy density and the fluoride concentration would not affect the demineralization resistance on root surface.

Materials and Methods
This study was approved by the Ethics Committee of The Nippon Dental University School of Life
Dentistry at Niigata (#ECNG-H-40, 2010).

The study was undertaken using 35 extracted human premolars

with previously unexposed, caries-free root surfaces.

Informed consent was obtained from the patients under aprotocol approved by the institutional review board of The Nippon Dental University School of Life Dentistry at
Niigata.

The teeth were cleaned and stored in 0.01% thymol solution at 4˚C until use.

The crown wasremoved approximately 2 mm below the cemento-enamel junction, and the root was longitudinally sectioned at
the center of the buccal surface.

The root surface was lightly planed immediately before carrying out theexperiment.
The roots were assigned to one of seven treatment groups as follows: CO2 laser irradiation at 17 J/cm2(Group 1); irradiation at 25 J/cm2 (Group 2); irradiation at 41 J/cm2 (Group 3); 200-minute soak in 0.05% NaF
solution (Group 4); 50-minute soak in 0.2% NaF solution (Group 5); 5-minute application of 2.0% NaF solution(Group 6); and no treatment (control).

Exposure was limited to a rectangular area (2x3 mm) on the root surface

created by coating the remaining surfaces with acid-resistant varnish (Protect varnish, Kuraray Noritake Dental,
Tokyo, Japan).

The exposed area was irradiated using a CO2 laser (Opelaser 03SIISP, Yoshida, Tokyo, Japan)

whose output was in the form of a continuous series of pulses.
defocused to a 1 mm spot size.

The 10.6 µm-wavelength, 0.5 W beam was

The irradiation cycle consisted of a pause time of 10 ms and an irradiation timeof 5 ms (Group 1), 10 ms (Group 2), or 50 ms (Group 3).

A computer-controlled X-Y stage (Shot-602,

Sigma-koki, Tokyo, Japan) was used to ensure uniform exposure.
traversed at 1 mm/s through the laser beam.

The root was fixed to the stage, which wasGroups 5 and 6 were treated with NaF in a room temperature(23˚C) bath stirred at 120 rpm.
The specimens were subjected to a 2-day pH-cycling process in which five roots from each group were
immersed daily in a demineralizing solution (pH 4.7, containing 0.05 M acetic acid, 2.2 mM calcium, and 2.2
mM phosphate ions) for 18 hours, and then in a remineralizing solution (pH 7.0, containing 0.15 M potassium
chloride, 1.5 mM calcium, and 0.9 mM phosphate ions) for 6 hours.
and stirred at 120 rpm.

The solutions were maintained at 37˚C

The roots were irrigated with deionized water for 5 minutes during transfer between

solutions and at the conclusion of the cycling process.
The samples were sectioned perpendicular to the root surfaces and through the center of each window usinga hard-tissue microtome (Isomet, Buehler, Lake Bluff, IL, USA).
approximately 200 µm were obtained from each window.

Three sections with a thickness ofEach section was ground to a thickness ofapproximately 100 µm using a whetstone (#2,000).
The sections were examined at 200x magnification using a polarized light microscope (Eclipse LV100POL,
Nikon, Tokyo, Japan).
Japan).

Digital photomicrographs were obtained using a CCD camera (DS-L2, Nikon, Tokyo,

The lesion depth in each section was determined by measuring the width between the original root

surface and the deepest position of the lesion using the camera control software.
The experimental results were analyzed using ANOVA and Tukey’s multiple comparison tests for post hoc

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Asian Pac J Dent 2014; 14: 1-6

analysis (p<0.05).

All analyses were carried out using the SPSS 12.0 software package for Windows (SPSS,

Chicago, IL, USA)

Results
The mean and standard deviation of the lesion depths for each experimental group are presented in Table 1.
ANOVA testing indicated significant differences between the mean lesion depths in the experimental groups andthe control.

The lesions in groups 3, 4, 5, and 6 were significantly shallower than the lesions in the control

specimens (p<0.002).
and 3 (p<0.05).

The lesions in groups 5 and 6 were significantly shallower than the lesions in groups 1, 2,Group 3 exhibited significantly reduced lesion depth compared to the control, but slightcarbonization was observed on the irradiated root surface.

However, there were no significant differencesamong groups 1, 2 and the control, or among groups 1, 2, 3, and 4 (p>0.05).
Table 1.

Results of lesion depths under various conditions of laser irradiation and fluoride application

SpecimengroupsConditions of laser irradiation

Fluoride
application

Lesion depth (µm)(Mean ± S.D.)*Power

DurationInterval

Energy

1

0.5 W

5 ms

10 ms

17 J/cm2

None

41.0 ± 3.4 a,b

2

0.5 W

10 ms

10 ms

25 J/cm2

None

42.3 ± 2.9 a,b

3

0.5 W

50 ms

10 ms

41 J/cm2

None

38.4 ± 10.8 b

4

None

0.05% NaF

37.9 ± 4.2 b,c

5

None

0.2% NaF

32.6 ± 3.4 c,d

6

None

2.0% NaF

28.3 ± 3.2 dNone

45.9 ± 2.7 aControl
None* The same superscript letters indicate no significant difference (p>0.05).

Fig. 1 Representative polarized light microphotographs of experimental groups(a) Group 1, (b) Group 2, (c) Group 3, (d) Group 4, (e) Group 5, (f) Group 6, and (g) Control

Representative PLM images of demineralized lesions are provided in Fig. 1.
the root surfaces coated with acid-resistant varnish.

The cementum was intact on

In all groups, stripes parallel to the bottom of the lesion

were observed over most of the lesion, indicating remineralization during the pH cycling procedure.

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widths of lesions in groups 1, 2, 3, and 4 were approximately equal to the mean lesion width of the control
samples, while lesions in groups 5 and 6 were smaller.

Discussion
In previous studies, tooth surface acid resistance has generally been evaluated using transverse
microradiography (TMR).11,27,28

We assessed the acid resistance of root surfaces in terms of the lesion depth

measured using digital polarized light microphotography (PLM).
incapable of measuring mineral loss.
root surface after pH cycling test.
study.

16

The PLM method is easier than TMR but isHowever, the PLM method is sufficient to evaluate the acid resistance ofThe design of the pH cycling test was based on tests carried out in a previous

A two-day cycle was conveniently brief, and may be useful to detect significant differences among theexperimental groups.

In this study, the laser output was in the form of a continuous series of pulses, and 0.5 W

laser beam was defocused to a 1 mm spot size.

The irradiation cycle consisted of an interval time of 10 ms, anda duration time of Group 1, 2 and 3 were 5, 10, and 50 ms, respectively.

A computer-controlled X-Y stage wasused to ensure uniform exposure, and the specimen fixed to the stage was moved at 1 mm/s during the laser
irradiation.

Therefore, the laser energy density of Group 1, 2, and 3 were 17 J/cm2 (0.25 J/cm2 per pulse), 25

J/cm2 (0.5 J/cm2 per pulse) and 41 J/cm2 (2.5 J/cm2 per pulse), respectively.
There are many articles described about effects of fluoride application and laser irradiation on theprevention of tooth root-surface caries.14-16,24-29

Although these previous reports suggested that fluoride

application and laser irradiation were effective in prevention of root caries, their results did not clearly show theeffects of energy density of CO2 laser and fluoride concentration on the root-surface demineralization.
Therefore, this study focused to examine how energy density of CO2 laser and fluoride concentration influencedemineralization of root-surface after pH cycling.

This study simultaneously examined the effects of a fluoride

application of various concentrations and a laser irradiation of various energy densities on tooth root
demineralization.

From the results of this study, the lesion in the group applied 2.0% NaF (group 6) was

significantly shallower than the lesions in the group applied 0.05% NaF (group 4), but there were no significant
differences between the lesions of groups irradiated various energy densities of laser (groups 1, 2, and 3).

Thus,

the high concentration fluoride application demonstrated greater demineralization resistance of root-surface
compared to the laser irradiation.

The null hypothesis that the fluoride concentration would not affect the root

surface demineralization resistance was rejected; however, the null hypothesis about CO2 laser energy density
was accepted.
The results concerning laser irradiation showed that the lesion depth in group 3 which used CO2 laser
irradiation at an energy density of 41 J/cm2 was significantly lower than that in the control.

On the other hand,the other experimental groups which used the laser irradiation at an energy density of lower than 27 J/cm2
showed lower lesion depth compared to the control, but there were no significant differences among them.
From these results, it was speculated that CO2 laser irradiation at high energy density might be effective inincreasing resistance for demineralization of root-surface, but CO2 laser irradiation at low energy density might
produce no beneficial effect on root-surface.
However, since a carbonization may occur on the irradiated root-surface at high energy level, appropriate
energy density of CO2 laser should be establish when using the laser for prevention of tooth root caries.

A

series of previous studies demonstrated reduced enamel and dentin demineralization following laser treatment in

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a caries model.4-13

In the case of enamel, a decrease in carbonate and phosphate after laser application appears

to be the cause of the reduced demineralization.

On the other hand, the mechanism of demineralization

inhibition in cementum and dentin of tooth root following laser irradiation is yet clarified.

Nelson et al.4

reported that fusion and melting occurred on the root dentin surface after CO2 laser irradiation at a power densityof 50 J/cm2.

Additionally, an increase in inorganic content occurring on the laser-irradiated dentin surface wasrelated to increased dentin resistance to demineralization.

However, chemical changes in the dentin substrate

resulting from elevated temperatures during laser irradiation are responsible for the decrease in carbonate content
and protein deposition.

Therefore, CO2 laser irradiation levels that are sufficiently high to cause carbonizationin dentin would not be suitable for application to root surfaces.
Previous studies have reported that fluoride exerts a beneficial effect on demineralization/remineralizationof the root-surface even at low concentrations.25,27,28

Similar results were obtained in the present study.

It iswell-known that the mechanisms of protection against tooth root caries depend on the deposition of fluoride on
the root surfaces, and the deposition of fluoride enhance tooth root demineralization resistance by creating afairly amount of fluoridated hydroxyapatite.25

Petersson et al.28 examined remineralization of root caries

lesions after in vitro treatment with various concentration fluorides, and concluded that daily application of adentifrice slurry containing 1,400 ppm fluoride combined with twice daily rinsing with a 250 ppm fluoride
solution significantly remineralize tooth root caries lesions in vitro.

Hong et al. reported that the effect of afluoride varnish on root surface demineralization/remineralization was dependent upon the exposure period andindependent of the concentration.29

In this study, three fluoride application procedures were decided frompoints of view as follows: 5-minute application of 2.0% NaF solution (9,000 ppm) was assumed as anapplication of fluoride varnish in clinic (group 6), 50-minute soak in 0.2% NaF solution (900 ppm) was assumed
as a tooth brushing with fluoride paste (group 2), and 200-minute soak in 0.05% NaF solution (225 ppm) was
assumed as a mouth washing using fluoride rinse (group 3).

The results of this study showed that each meanlesion depth in groups 4, 5, and 6 was significantly shallower than that in the control group.

Furthermore, themean lesion depth tended to decrease with increasing fluoride concentration regardless of application period.
Accordingly, these results suggested that the fluoride application was effective in increasing demineralization
resistance of root-surface, and the efficacy was dependent upon the concentration rather than the application
period.

Therefore, it is speculated that an application of fluoride varnish such as 2.0% NaF solution in clinic

may be more effective in preventing root caries than tooth brushing and mouth washing with fluoride at home.
A periodic application of a fluoride varnish in clinic is recommended for preventing root caries.
Effectiveness of the combination of fluoride and laser treatment has been reported for the inhibition of root
caries.16

However, the combination details such as factors of fluoride concentration and laser energy density

have not investigated in the previous study.

Further researches evaluating effects of combined various

concentration fluoride and various energy density CO2 laser on demineralization resistance of root surface arepromising.
Acknowledgment

This research was supported in part by a Grant-in-aid for Scientific Research (A) from the Japan Society for the
Promotion of Science (No. 21249091).

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Correspondence to:Dr. Koichi Shinkai
Department of Operative Dentistry, The Nippon Dental University School of Dentistry at Niigata
1-8 Hamaura-cho, Chuo-ku, Niigata 951-8580, Japan
Fax: +81-25-265-7259 E-mail: shinkaik@ngt.ndu.ac.jp
Accepted January 30, 2014.
Copyright ©2014 by the Asian Pacific Journal of Dentistry.
Online ISSN 2185-3487, Print ISSN 2185-3479

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