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Asian Pac J Dent 2018; 18: 7-14

Effect of nonthermal atmospheric discharge on tooth bleaching
Ayaka Kusanagi, DDS, Masayuki Otsuki, DDS, PhD, and Junji Tagami, DDS, PhD
Cariology and Operative Dentistry, Graduate School of Medical and Dental Sciences, Tokyo Medical
and Dental University, Tokyo, Japan
Purpose: The purpose of this study was to evaluate effect of nonthermal atmospheric discharge on tooth
bleaching in vitro by color measurement using a dental colorimeter and an industrial colorimeter.
Materials and Methods: The air stream of nonthermal atmospheric discharge was exposed on thehematoporphyrin stained paper for 5, 10, 20, 30, and 60 minutes respectively. The air stream without
atmospheric discharge (negative control) and a commercially available tooth bleaching material (positive
control) were also prepared. The L*a*b* values on the treated surface at each step was measured by a dental
colorimeter and an industrial colorimeter. Color difference was calculated from those values. The artificial
discolored bovine teeth samples were prepared and exposed by the air stream of atmospheric discharge for 5, 10,
20, 30, and 60 minutes, and the color change was evaluated.
Results: The nonthermal atmospheric discharge showed bleaching effect for both hematoporphyrin stained
paper and artificial discolored bovine teeth. Although the measured color values of both colorimeters were notconsistent statistically, they showed high correlation.
Conclusion: It was concluded that nonthermal atmospheric discharge showed the bleaching effect and twocolorimeters were useful for measuring color of hematoporphyrin stained paper and artificial discolored bovine
teeth.
(Asian Pac J Dent 2018; 18: 7-14.)
Key Words: atmospheric discharge, color meter, plasma, tooth bleaching

Introduction
Tooth bleaching is one of the most conservative and cost-effective dental treatments to improve or enhance a
person’s smile and it has become one of the most popular esthetic dental treatments. There are two kinds of vital
tooth bleaching treatments. One is dentist-supervised nightguard bleaching (home bleaching) and another isin-office bleaching (office bleaching). An active ingredient of tooth bleaching material is peroxides; mostlyhydrogen peroxide for the office bleaching and carbamide peroxide for the home belching. To accelerate theeffect of office bleaching, high concentration of hydrogen peroxide has been generally contained in thebleaching materials. Photo-activation is also employed to enhance the effect using quartz tungsten halogen lump,
light emitted diode or various lasers as the light source [1]. However, the light exposure for office bleachingmight not improve the bleaching effect and it increased the risk of tooth sensitivity [1].
Recently, the nonthermal atmospheric pressure plasma (NAPP) was proposed for the tooth bleaching toincrease the reaction of hydrogen peroxide in the office bleaching material [2-4]. Those NAPPs were reactedwith not only hydrogen peroxide but also with carbamide peroxide [5,6] and deionized water [7,8] on the tooth
surface and those reactions showed tooth bleaching effect. The NAPP is generated by the atmospheric discharge.
However, there has been no study on the tooth bleaching effect of the NAPP by the atmospheric discharge
without peroxides or water.
The precise color measurement of the tooth is important for the evaluation of the tooth bleaching effect. Twokinds of methods were used for tooth color measurement. One is visual comparison between the target natural
tooth and the tooth shade guides and another is the measurement using a color measuring devise, such as acolorimeter or a spectrophotometer. Although the conventional method using shade guides is useful and widelyused, it is too subjective. Tooth color measurement using a dental colorimeter is able to obtain the objective
results. For dental research concerning tooth color including tooth bleaching, the dental colorimeter and theindustrial colorimeter were often used. Sometimes, it is necessary to compare the color values measured by7

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different colorimeters. However, there are few information about the compatibility of measured values [9].
The purpose of this study was to evaluate the effect of nonthermal atmospheric discharge (NADC) on tooth
bleaching in vitro by color measurement using a dental colorimeter and an industrial colorimeter.

Materials and Methods
System of NADC
The experimental system of NADC used in this study was shown in Fig 1A. It was consisted from amultifunction synthesizer (Wave Factory WF1943 1CH, NF, Kawasaki, Japan), a high speed bipolar amplifier(HSA4051, NF), a digital phosphor oscilloscope (TDS 3014C, Tektronix, Tokyo, Japan), an air pump
(SPP-15GA, Techno Takatsuki, Takatsuki, Japan) and a discharge tube (experimental).

Fig. 1 Experimental apparatus
A. Overall image: a, multifunction synthesizer; b, high speed bipolar amplifier; c, digital phosphor oscilloscope,d, air pump; e, air flow meter; f, discharge tube
B. Discharge: Air flow through the discharge exposed the experimental surface

Preparation of hematoporphyrin (HP) stained paper
The hematoporphyrin (HP) stained paper was prepared by following procedure according to a previous study[10]. The 0.24 g of hematoporphyrin (Wako, Osaka, Japan) was dissolved in 300 mL of ethanol for preparing 0.1
wt% hematoporphyrin ethanol solution. The glossy white photo paper (GL-101A4100, Cannon, Tokyo, Japan)
was immersed in the solution for 5 minutes and naturally dried in a darkroom. The color of the stained paper wasmeasured by an industrial colorimeter (NR-11, NR, Nippon Denshoku, Tokyo, Japan) and the portion of L*
value between 48 and 53 was chosen and employed for this study. The HP stained paper was trimmed with asuitable size with approximately 2 × 2 cm and a vinyl tape with a hole of 5 mm in diameter was put for
determining the experimental area. The L*a*b* values of each HP paper sample were measured using an
industrial colorimeter (NR) and a dental colorimeter (Shade eye NCC, SE, Shofu, Kyoyo, Japan).
Bleaching procedure
DC(+) group
The HP paper was placed on a flat table keeping a distance of 5 mm from the tip of a cylinder of NADC inwhich metal electrodes were set (Fig. 1B). Plasma was generated by discharge with a peak voltage of −5 V, apeak current value of −10 A and a frequency of 1.3 kHz. The 5 L/min of air flow through metal electrodes was
exposed to surface of sample from the tip of tube (Fig. 1B). After exposure for 5, 10, 20 30, and 60 minutes,
L*a*b* values of the exposed area of HP stained paper were measured using two colorimeters.

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DC(−) group (negative control group)
HP paper was places on the table as well as DC(+) group. The discharge was turned off and the air stream was
exposed for 5, 10, 20 30, and 60 minutes followed by color measuring using two colorimeters.
BL group (positive control group)
A commercially available office bleaching product (Pyrenees, Mitsubishi Gas Chemical, Tokyo, Japan) wasused as a positive control. It contained low concentration hydrogen peroxide (approximately 3.5%) and visible
light activating titanium dioxide photo catalyst. The contents of two capsules of Pyrenees were mixed well. The
mixed liquid was applied on the HP stained paper and light was irradiated by a light unit (Cosmo Blue, GC,
Tokyo, Japan). The light source of the light unit was violet light emitted diode (LED) with 405 nm of the peakwavelength, and the light intensity was 55 mW/cm2. The bleaching liquid was changed at 5 minutes and every
10 minutes, and color was measured at 5, 10, 20 30, and 60 minutes.
Number of the specimen in each group was 10 (n = 10). The experimental surface of each step was also
recorded by a digital camera. The color difference (ΔE) between the baseline (0 minute) and the eachexperimental period was calculated, according to the following equation;
ΔE = [(ΔL*)2 + (Δa*)2 + (Δb*)2]1/2
where ΔL, Δa, and Δb are difference values of L*, a*, and b* between the baseline and each experimental periodof the bleaching respectively.
Artificial discolored bovine teeth
Extracted bovine lower incisors were thawed in the running tap water. After removing soft tissues by a scalpel,
the labial surface was ground by a wet silicon carbide (SiC) papers #400-1,000 to obtain flat enamel surface.
Then, 5 × 5 mm specimen was prepared by a diamond cutting saw with copious water. The specimen wasembedded in a dental acrylic resin (Unifast, GC) and polished by #1,200 SiC paper. The black tea was extracted
from two teabags (Lipton Yellow Label, Unilever Japan, Tokyo, Japan) in 100 mL of boiled water. The samples
were immersed in the tea extract for 7 days at 37˚C kept in an incubator. The solution was changed at 4th day.
The color of the stained enamel surface was measured by a colorimeter (NR) and the samples of L* value
between 43 and 58 were selected for this study.
The stained enamel surfaces were exposed as well as DC(+) group of the experiment using HP stained paper.
The color was measured using two colorimeters at 5, 10, 20, 30, and 60 minutes. Ten specimens were preparedand evaluated (n = 10).
Statistical analysis
The L*, a*, b* and ΔE values of each group in HP stained paper experiments were statistically analyzed by
two-way and one-way analysis of variance (ANOVA), then Tukey’s HSD at confidential level of 0.05% (p =0.05). The L*, a*, b* and ΔE values in bovine teeth experiments were analyzed by one-way ANOVA andTukey’s HSD. Obtained data by the dental colorimeter (NR) and the industrial colorimeter (SE) were comparedby Kendall's coefficient of concordance (Kendall's W) for assessing agreement among raters and Spearman's
rank correlation coefficients (Spearman's rho) for measuring rank correlation. Commercially available software(SPSS Statistics ver.21.0, IBM, Armonk, NY, USA) was used for these analyses.

Results
The typical color change of the HP stained papers in DC(+) and BL groups were shown in Fig. 2. The bleaching

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effect was found in both groups. The HP stained papers in DC(−) showed no change visually. The change of
L*a*b* values of each group was shown in Fig. 3.

Fig. 2 Typical image of the change of HP paper in DC(+) and BL groups

Fig. 3 Change of average L*a*b* values in each group
A, DC(+) group measured by NR;
B, DC(+) group measured by SE;
C, DC(−) group measured by NR;
D, DC(−) group measured by SE;
E, BL group measured by NR;
F, BL group measured by SE

In DC(+) and BL groups, L* values were gradually increased and a* and b* values were decreased. For L*,
a*, and b* values before exposure (time 0), there were no statistical differences among all experimental groups
measured by both color meters respectively (p > 0.05). For L*, a*, and b* values among three groups, there were
statistically differences in factor of experimental group and experimental time measured using both NR and SE

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by two-way ANOVA (p < 0.05). The L* values in DC(+) and BL groups were much increased until 30 minutes.
The L* values of DC(+) and BL groups showed statistically higher than those of DC(−) group. The a* values in
DC(+) until 5 minutes and those of BL groups until 20 minutes were much decreased. The b* values in DC(+)
and BL groups were much decreased until 30 minutes. The a* and b* values of BL groups showed statisticallylowest followed by those of DC(+) group, then DC(−) group. For ΔE values as shown in Fig. 4, there were
statistically differences in factor of experimental groups and experimental time measured using both NR and SE
by two-way ANOVA. The ΔE values of BL groups showed statistically highest and followed by those of DC(+)
group, then DC(−) group.
The Kendall's W of L*, a*, and b* values measured by NR and SE were 0.000, 0.886, and 0.506 respectively.
Spearman's rhos of them are significant (p < 0.05) and were 0.980, 0.968, and 0.967 respectively.

Fig. 4 The ΔE values in each experimental time

A, Measured by NR;

B. Measured by SE

The typical image of the change of bovine tooth by NADC was shown Fig. 5. The stain was slightly bleachedby exposed time. Change of L*a*b* values of bovine tooth by DC was shown in Fig. 6. The Kendall's W of L*,
a*, and b* values measured by NR and SE were 0.735, 0.308, and 0.934 respectively. Spearman's rhos of them
are significant (p < 0.05) and were 0.949, 0.790, and 0.821 respectively. The change of ΔE values in eachexperimental time was shown in Fig. 7. The ΔE value was increased accompanied by increasing L* value.

Fig. 5 Typical image of the change of bovine tooth by DC

Fig. 7 The ΔE values in each experimental time

A, Measured by NR

Fig. 6
B. Measured by SE
Change of L*a*b* average values of bovine tooth by DC

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Discussion
In this study, HP stained paper and artificial discolored bovine teeth were used for evaluation of bleaching effect.
This experimental method was already used for research of tooth bleaching [10,11]. The evaluation using HP
stained paper is sensitive and is suitable for the screening test. However, it is difficult to predict the tooth
bleaching effect from only the results of evaluation using HP stained paper. It is necessary to evaluate thebleaching effect using teeth. Since original extracted bovine teeth were very bright and whitish, they were
stained by black tea extract for 7 days before the experiment. Tooth discoloration is classified as extrinsic,
intrinsic, or a combination of both, and tea is one of the typical extrinsic chromogens [12]. In this study, bovine
teeth were stained from both enamel surface and pulp chamber. Evaluation using artificial discolored bovine
teeth can be expected to acquirer the results more clinically than using HP stained paper.
Hydrogen peroxide is widely used as an active ingredient of office bleaching. When office bleaching material
is applied on the tooth surface, hydrogen peroxide in bleaching material is reacted and produces water andoxygen molecules, which reaction is accelerated by heat, catalyst and light irradiation. The concentration of HP,
pH of bleaching material, application times also affect the bleaching effect. During this reaction of HP, free
radicals such as oxygen (O·), hydroxyl radical (OH·), perhydroxyl radical (HO2·) and super oxide anion (O2-·)
are released [13]. Tooth discoloration is caused by extrinsic and/or intrinsic chromogen molecules [12]. Those
free radicals react with the chromogen molecules in the teeth and those molecules were degraded to smaller,transparent and soluble molecules, then chromogens were removed from discolored tooth and the tooth isbleached. However, the exact mechanism of tooth bleaching by peroxides has not been clear [2,4].
In this study, the plasma was generated by NADC and was contributed the bleaching effect. Plasma is thepartially ionized gas [9] and the plasma technology has been widely used for industry [14]. This technology has
been tried to be applied medicine including dentistry [14,15] such as surface treatments of implant [16,17],
ceramics [14] and dentin [18], and polymerization of acrylic resin [19] and composite resin [20], in addition,
tooth bleaching [2-7].
In the previous studies on tooth bleaching using NAPP, tooth surface was directly applied by the plasma
accompanied with hydrogen peroxide, carbamide peroxide or deionized water. The chemical reaction ofhydrogen peroxide and carbamide peroxide in the tooth bleaching material is accelerated by NAPP and plays
bleaching effect as the mechanism above mentioned [2-5]. The bleaching effect was also shown by deionizedwater and NAPP [6,7]. The NAPP with water is able to produce reactive species, such as hydrogen peroxide andozone [21,22], but also free radicals [21-23]. Those products are thought to contribute the bleaching effect.
In this study, the plasma generated by NADC was applied without peroxides or water.

And the plasma by

glow discharge was not directly applied on the experimental surfaces and air flow through the glow discharge
was exposed. Although the mechanism of the bleaching in this study is not clear, it can be speculated thathydrogen peroxide and free radicals would be produced by plasma with oxygen and water in the air and would
be delivered and reach the surface of experimental samples, then the exposed surfaces were bleached. Further
study is necessary for revealing the bleaching mechanism.
The DC(+) group in HP stained paper experiment showed bleaching effect. However, this effect was less thanBL group. The office bleaching material used in this study (Pyrenees) contains low concentration of hydrogen
peroxide and visible light-activating titanium dioxide photo catalyst. This bleaching effect was comparable withthat of another product containing higher concentration of hydrogen peroxide [10,24].

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The bleaching effect in discolored bovine teeth experiment was slight and less than that in HP stained paper.
Office bleaching was effective inside the dentin through the enamel [11]. It seems to be difficult to penetrate air
stream and products by NADC to the dentin through enamel. The bleaching effectiveness of NADC may belimited on the enamel surface.
There are two types of color measuring devices; spectrophotometer and colorimeter. In this study, a dental
colorimeter and an industrial colorimeter were used and obtained data were compared. Colorimeters measuretristimulus values and filter light in red, green and blue areas of the visible spectrum [9]. Although colorimeters
is less accurate than spectrophotometers, they were widely used for both dentistry and general industry, because
they are generally less expensive than spectrophotometers and still useful. The colorimeters can obtainparametric data as well as spectrophotometer, which could be easily applied for statistical analysis. ShadeEye
NCC (SE) used in this study is designed for measuring the shades of natural teeth and ceramic restorations. It
was reported that the shade selecting using SE could make better results than visual method for theuncomplicated cases [25].
For measuring the tooth color, it is sometimes difficult to apply an industrial colorimeter, especially forclinical situation. Although a dental colorimeter is useful for measuring tooth shade, it is not clear that a dental
colorimeter can be measure the color of non-tooth samples. It was necessary to know the compatibility between
a dental colorimeter and an industrial colorimeter. This study evaluated the compatibility of these colorimeters.
Although the Kendall's W of a* in HP stained paper experiment and that of b* in bovine teeth experiment werehigh, those of L* in HP stained paper experiment and a* in bovine teeth experiment were low. Therefore, it isdifficult to compare the L*, a*, and b* values measure by both colorimeters directly. Since Spearman's rho of the
L*, a*, and b* values measure by both colorimeters in both experiments were very high (0.790-0.980) andstatistically significant, both colorimeter can utilize for the experiments using HP stained paper and discoloredbovine teeth and can evaluate the change of color by bleaching.
Tooth bleaching using peroxides is not risk-free. Tooth hypersensitivity is a major adverse effect of the tooth
bleaching [13]. There is no evidence of toxic and carcinogenetic risks of hydrogen peroxide used at exposurelevels of tooth bleaching [13]. However, only limited long-term clinical data are available on the side effects of
tooth bleaching. Many studies reported no deleterious effects on bleached enamel and dentin surfaces concerning
structure and hardness [13]. The change of temperature and morphology after bleaching by NAPP with hydrogen
peroxide [2-4], carbamide peroxide [5,6], and deionized water [8] were reported and there were no deleterious
effects. This study suggested the possibility of tooth bleaching by NADC technology without peroxides or water.
Further studies on the safety and efficacy are necessary to establish the clinical application. It was concluded that
NADC showed the bleaching effect and two colorimeters were useful for measuring color of HP stained paper
and artificial discolored bovine teeth.
Conflicts of interest
None

References
1.
2.
3.

He LB, Shao MY, Tan K, Xu X, Li JY. The effects of light on bleaching and tooth sensitivity during in-office vital bleaching: A
systematic review and meta-analysis. J Dent 2012; 40: 644-53.
Lee HW, Kim GJ, Kim JM, Park JK, Lee JK, Kim GC. Tooth bleaching with nonthermal atmospheric pressure plasma. J
Endod 2009; 35: 587-91.
Sun P, Pan J, Tian Y, Bai N, Wu H, Wang L, et al. Tooth whitening with hydrogen peroxide assisted by a direct-current cold
atmospheric-pressure air plasma microjet. IEEE Trans Plasma Sci 2010; 38: 1892-6.

13

Kusanagi et al.

4.
5.
6.
7.
8.
9.
10.
11.
12.
13.
14.
15.
16.
17.
18.
19.
20.
21.
22.
23.
24.
25.

Asian Pac J Dent 2018; 18: 7-14

Claiborne D, Mccombs G, Lemaster M, Akman MA, Laroussi M. Low-temperature atmospheric pressure plasma enhanced
tooth whitening: The next-generation technology. Int J Dent Hyg 2014; 12: 108-14.
Nam SH, Lee HW, Cho SH, Lee JK, Jeon YC, Kim GC. High-efficiency tooth bleaching using non-thermal atmosphericpressure plasma with low concentration of hydrogen peroxide. J Appl Oral Sci 2013; 21: 265-70.
Nam SH, Lee HJ, Hong JW, Kim GC. Efficacy of nonthermal atmospheric pressure plasma for tooth bleaching. Sci World J
2015; 2015: 18-23.
Lee HW, Nam SH, Mohamed AAH, Kim GC, Lee JK. Atmospheric pressure plasma jet composed of three electrodes:
Application to tooth bleaching. Plasma Process Polym 2010; 7: 274-80.
Kim MS, Koo IG, Choi MY, Jung JC, Eldali F, Lee JK, et al. Correlated electrical and optical studies of hybrid argon
gas-water plasmas and their application to tooth whitening. Plasma Process Polym 2012; 9: 339-45.
Chu SJ, Trushkowsky RD, Paravina RD. Dental color matching instruments and systems. Review of clinical and research
aspects. J Dent 2010; 38(Suppl 2): 2-16.
Suemori T, Kato J, Nakazawa T, Akashi G, Igarashi A, Hirai Y, et al. Effects of light irradiation on bleaching by a 3.5%hydrogen peroxide solution containing titanium dioxide. Laser Phys Lett 2008; 5: 379-83.
Kishi A, Otsuki M, Sadr A, Ikeda M, Tagami J. Effect of light units on tooth bleaching with visible-light activating titanium
dioxide photocatalyst. Dent Mater J 2011; 30: 723-9.
Hattab FN, Qudeimat MA, Al-Rimawi HS. Dental discoloration: An overview. J Esthet Dent 1999; 11: 291-310.
Minoux M, Serfaty R. Vital tooth bleaching: biologic adverse effects-a review. Quintessence Int 2008; 39: 645-59.
Arora V. Cold atmospheric plasma (CAP) in dentistry. Dentistry 2014; 4: 189.
Kim JH, Lee MA, Han GJ, Cho BH. Plasma in dentistry: a review of basic concepts and applications in dentistry. Acta
Odontol Scand 2014; 72: 1-12.
Scacchi M, Merz BR, Schär AR. The development of the ITI Dental Implant Sysytem. Part 2: 1998-2000: Steps into the next
millennium. Clin Oral Implants Res 2000; 11(Suppl 1): 22-32.
Duske K, Koban I, Kindel E, Schröder K, Nebe B, Holtfreter B, et al. Atmospheric plasma enhances wettability and cellspreading on dental implant metals. J Clin Periodontol 2012; 39: 400-7.
Ritts AC, Li H, Yu Q, Xu C, Yao X, Hong L, et al. Dentin surface treatment using a non-thermal argon plasma brush forinterfacial bonding improvement in composite restoration. Eur J Oral Sci 2010; 118: 510-6.
Nishigawa G, Maruo Y, Oka M, Oki K, Minagi S, Okamoto M. Plasma treatment increase shear bond strength between heat
cured acrylic resin and self-curing acrylic resin. J Oral Rehabil 2003; 30: 1081-4.
Chen M, Zhang Y, Yao X, Li H, Yu Q, Wang Y. Effect of a non-thermal, atmospheric-pressure, plasma brush on conversionof model self-etch adhesive formulations compared to conventional photo-polymerization. Dent Mater 2012; 28: 1232-9.
Liu J, He B, Chen Q, Li J, Xiong Q, Yue G, et al. Direct synthesis of hydrogen peroxide from plasma-water interactions. Sci
Rep 2016; 6: 38454.
Gorbanev Y, O’Connell D, Chechik V. Non-thermal plasma in contact with water: The origin of species. Chem - A Eur J
2016; 22: 3496-505.
Bruggeman P, Schram DC. On OH production in water containing atmospheric pressure plasmas. Plasma Sources Sci
Technol 2010; 19: 45025.
Suyama Y, Otsuki M, Ogisu S, Kishikawa R, Tagami J, Ikeda M, et al. Effects of light sources and visible light-activatedtitanium dioxide photocatalyst on bleaching. Dent Mater J 2009; 28: 693-9.
Li Q, Wang YN. Comparison of shade matching by visual observation and an intraoral dental colorimeter. J Oral Rehabil
2007; 34: 848-54.

Correspondence to:Dr. Masayuki Otsuki
Cariology and Operative Dentistry, Graduate School for Medical and Dental Sciences,
Tokyo Medical and Dental University,
1-5-45, Yushima, Bunkyo-ku, Tokyo 113-8549, Japan
Fax: +81-3-5803-0195 E-mail: otsuki.ope@tmd.ac.jp
Accepted December 28, 2017
Copyright ©2018 by the Asian Pacific Journal of Dentistry.
Online ISSN 2185-3487, Print ISSN 2185-3479

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