Evaluation of In-Office Bleaching on Enamel and Dentine: An
FTIR Study
AYCA DOGAN1, KURTULUS GOKDUMAN2, SUKRAN BOLAY3, FERIDE
SEVERCAN4
1 Department of Cosmetic Technology, Kocaeli University, Izmit, TURKEY
2 Department of Biotechnology , Middle East Technical University, Ankara, TURKEY
3 Department of Restorative Dentistry, Hacettepe University, Ankara, TURKEY
4 Department of Biology , Middle East Technical University, Ankara, TURKEY
Abstract: In recent years bleaching of vital teeth has become popular among both dentist and
patients. Different bleaching agents were used for this purposes. They are either applied professionally
at high dose (office bleaching) or by patient at lower dose (home bleaching). However the effects of
bleaching on teeth is still unclear and controversial. In the present work we studied the effect of a high
concentration bleaching agent (35% hydrogen peroxide) which is called as office bleaching, on human
enamel and dentine using Fourier Transform Infrared (FTIR) Spectroscopic Technique. The results
revealed differences in the signal intensity/area values and peak positions of some bands in between
the treated and control tissues. In addition, the OH stretching band of hydroxyapatite at 3566 cm-1
appeared in the spectra of enamel tissue which was absent in dentin. The relative amount of
carbonate and phosphate changed for the treated groups. In conclusion office bleaching caused some
alterations in the structure and concentrations of enamel. However, these changes was not significant
in dentin tissue.
Key-Words: Office bleaching; Tooth whitening; Hydrogen peroxide; Enamel; Dentine;
FTIR
Spectroscopy
1 Introduction
The technique of bleaching or whitening teeth
was first described in 1877[1]. Since then
bleaching of teeth has been in use, with little
changes in science or technique during that
time[2]. For example in 1937, Ames described
a technique for treating mottled enamel by
using a combination of hydrogen peroxide,
ether, and heat [3]. However current in-office
bleaching technique is basically the same as
the technique developed between 1880-1916,
which uses 35% hydrogen peroxide with
rubber dam isolation[2].
The history of “modern day” tooth
bleaching, however, began in 1989. In 1989
Haywood and Heyman introduced the
nightguard vital bleaching method[4]. When
home ‘nightguard’ bleaching using carbamide
peroxide was introduced in 1989, it appeared
that the in-office approach would become less
popular. However, at the present time there has
been a recent resurgence in-office bleaching,
primarily due to aggressive marketing of
various ‘high tech’ light sources such as lasers
and plasma arc lights, coupled with claims of
reducing bleaching time, even to a single
office visit.
In-office bleaching product contains 3035% hydrogen peroxide[2]. The mechanism of
the action of bleaching agents is thought to be
due to the ability of hydrogen peroxide to form
oxygen free radicals that interact with
adsorbed colored organic molecules and
oxidize these macromolecules and pigment
stains, producing dental discoloration into
smaller and lighter molecules.[5]
In the present work we aimed to study the
effect of a high concentration bleaching agent
(35% hydrogen peroxide) which is called as
office bleaching, on human enamel and dentine
composition using Fourier Transform Infrared
(FTIR) Spectroscopic Technique. We used
FTIR spectroscopy because with this technique
biological systems can be observed
atmolecular level without any damage in
structural components[8-11].
450 cm-¹ region. As seen from the figures
dramatic changesModeare
observed for enamel.20.14.1061 15:59
File # 1 : TESTMI~1
=
Fourteen human premolars newly extracted for
periodontal reasons were used. They were
randomly divided into control and treatment
groups of seven specimens. The specimens of
experimental groups were exposed to 35%
hydrogen peroxide. The control and
experimental groups were stored in distilled
water for 7 days. The roots of each tooth were
sealed with nail varnish to prevent the
penetration of bleaching agent. The roots were
sectioned to obtain flat buccal and lingual
enamel surfaces from half of the crown. The
specimens were sectioned with a high-speed
diamond rotary instrument using water and air
spray. Enamel and dentine specimens were
ground in liquid nitrogine and then, were
investigated in the form of KBr pellets by
using FTIR spectroscopic technique. A PerkinElmer spectrometer was used with 4 cm-1
resolution for this purpose.
3 Results and Discussion
The tooth mainly composed of enamel,
dentine, dentine-enamel junction and pulp.
Enamel is the hardest tissue found in the
human body. Mature enamel is highly
mineralized [12]. It contains 96% inorganic
material, 1% organic material and 3% water by
weight. The inorganic component is mainly
calcium phosphate in the form of
hydroxyapatite crystals. Other elements
present are small amounts of carbonate,
magnesium, potassium, sodium and fluoride.
Dentine is softer than enamel. The composition
of dentine is approximately 70% inorganic
material, 20% organic material and 10% water
by weight. The main inorganic component is
hydroxyapatite, and
the main organic
component is Type I collagen[13].
Figure 1 shows the representative
FTIR spectra for control groups in the 40001000 cm-¹ region for enamelThe bands are
labelled on the figure and their band
assignments are given in Table I.
Figure 2 shows normalized A) treated
and untreated enamel and B) treated and
untreated dentin IR spectra in 3900-2000 cm-¹
region. Figure 3 demonstrates normalized A)
treated and untreated enamel and B) treated
and untreated dentin IR spectra in the 1900-
Sample Description: 18.09.2004 1/100kbr+testmine2 1.cekim
Scans =
Res = 4,000000
Apod =
6
2
1.5
ABSORBANCE
2 Materials and Methods
5
2
1
10
92
2
.5
7
2
2
4000
Absorbance / Wavenumber (cm-1)
8
2
34
2
1
3000
3000
2000
2000
1000
1000
WAVENUMBER (cm-1)
Figure1. The representative FTIR spectra of
A) enamel and B) dentin of control group in
4000-1000 cm-¹ region
Table 1. Major absorptions in IR spectra of
control enamel.
Band Enamel Dentine
#
Frequency Frequency Definitions of the
spectral assignment
(cm-1)
(cm-1)
1 3567
2 3369
_
3363
OH stretching
O-H and N-H group
stretching vibration:
polysaccharides,
protein
3 1637
1652
H2O and organic
material-enamelAmide I (protein C=O
stretch)-dentin4 1544
1547
Amide II (protein NH bend, C-N stretch)
5-7 1200-900 1200-900 ۷1۷3 PO4 stretching
(mineral)
8 890-850 890-850 ν2 CO3-2 (mineral)
type B
9-10 700-450 700-450 ۷4 PO4 bending
(mineral)
450
File # 2 : CONTRO~1
Mode =
20.14.1061 17:35
Sample Descrip tio n: 13.10.2004 1/100KBr+controlmine 3 3.cekim
Scans =
Res = 4,000000
Apod =
25,5
Control Enamel
Treated Enamel
A
1
24
A
22
ABSORBANCE (a.u.)
ABSORBANCE (a.u.)
20
18
16
.5
Control Enamel
Treated Enamel
14
A 12
10
0
8
6
4
2
-.5
File # 2 : CD1
Mode =
3500
3500
Sample Descrip tio n: 13.10.2004 1/100KBr+controldentin 3 1.cekim
20.14.1061 22:13
3000
3000
2500
2500
WAVENUMBER (cm-1)
Absorbance
Scans = / Wavenumber (cm-1)
Res = 4,000000
-0,1
2000
2000
1900,0
1800
1900
1700
1600
1500
1400
1500
1300
1200
cm-1
1100
1000
1100
900
800
WAVENUMBER (cm-1)
Apod =
700
700
600
500 450,0
5,10
1
Control Dentine
Treated Dentine
B
Control Dentine
Treated Dentine
4,5
B
ABSORBANCE (a.u.)
ABSORBANCE (a.u.)
4,0
3,5
3,0
.5
A
2,5
2,0
1,5
1,0
0,5
0
-0,03
3500
3500
Absorbance / Wavenumber (cm-1)
3000
3000
2500
2500
2000
2000
WAVENUMBER (cm-1)
Figure 2. The average normalized A)
treated and untreated enamel and B)
treated and untreated dentine IR spectra in
the 3900-2000 cm-¹
Mineral matrixes of the enamel and
tisues are composed in its majority of crystals
of carbonated hydroxyapatite and the absorbed
components in the infrared region are the
hydroxyl (OH-), carbonate (CO3-2) and
phosphate radical (PO4-3). [14]. In the spectra
of enamel samples, the hydroxyl stretching
vibration is seen as a shoulder at 3566 cm-¹
which has a high degree of crystallinity,
because this vibration assigned to the OH
groups which located within the crystalline
channels of calcium hydroxyapatite. [15]. The
broad band at 3400 cm-¹ corresponds to NH
stretching vibrations of Amide A and
intermolecular OH bonding as seen from Fig 2.
The frequency of this band shifted to lower
values in treated enamel which indicates that
NH groups were involved in a new set of
hydrogen bonds of weaker strength[16,17].
19001800
1900,0
1700
1600
15001400
1500
1300
1200
cm-1
11001000
1100
900
800
700
700
600
500 450,0
WAVENUMBER (cm-1)
Figure 3. The averaged normalized
A)
treated and untreated enamel and B)
treated and untreated dentine IR spectra in
the 1900-450 cm-¹ region.
Dentine is mainly composed of type I
collagen as the organic material. Type I
collagen constitues around 90% of the dentin
protein fraction.
Therefore the amide I
absorbance of proteins mainly due to collagen
and it is observed at 1655 cm-1 .Carbonate
bands overlaps the amide II bands at around
1545 cm-1 . Therefore, in the present study the
amide II band of the protein matrix were not
taken into consideration[14,15].
The ۷2 carbonate (870-880 cm-¹) bands and
۷1۷3 phosphate (900-1200 cm-1 ), ۷4 phosphate
(520-650 cm-1) in the enamel spectrum and
560-605 cm-¹ in the dentin spectrum)
stretching bands which arise from mineral
components are observed. (14)
Investigations of mineral crystallinity
focused essentially on the ۷1۷3 phosphate band
between 1200-900 cm-1 [16-18]. As seen from
figure 3, there is a decrease in the intensity of
۷1۷3 phosphate band indicates a decrease in
mineral components in the treated enamel and
dentine tissue. However, these changes are
more dramatic for enamel samples. Moreover,
similar decrease is observed in the ۷4
phosphate band for treated enamel and dentine
samples.
After deconvolution, ۷1۷3 PO4 band is
divided into a high and low freguency domain.
The 1020 cm-1 component is characteristic of
poorly crystalline apatites and the 1030 cm-1
component of the more crystalline ones.
Carbonate plays an important role in
affecting crysttallinity and solubility of the
mineral matrix as seen from figure 3 The
intensity of carbonate band at 870-880 cm-¹
was seen to decreased implying that there is a
decrease in carbonate content in treated enamel
samples[16-18]. This decrease in CO3-2
content may disturb or disorganize the apatite
lattice. However, this decrease are not great for
dentine tissues.
The mineral to matrix ratio is
indicative of the relative quantity of mineral
present in calcified tissues[19]. The intensity
of ۷1۷3 phosphate stretching and the amide I
bands were evaluated to determine the relative
ratio of mineral and protein matrix. Intensity of
the mineral to matrix ratio decreased from 56,
218 to 15,455 after the treatment of Hydrogen
Peroxide for enamel tissue. However, this ratio
was not dramatically decreased after the
treatment of Hydrogen Peroxide for dentine
tissue. These changes can be ccclearly seen
seen from Fig3. .
4
CONCLUSION I
In the present study it was determined that
bleaching treatment led to a significant loss of
calcium and phosphate in treated enamel tissue
by using FTIR spectroscopic method. Loss of
mineral components observed from the PO4
bands indicates that demineralization took
place in the enamel samples. This fact led to
decrease in hardness of enamel samples [13].
Furthermore, the decrease in total carbonate
content was associated with lower mineral
crystallinity. Hydrogen peroxide bleaching
treatment gave no statistically significant
changes in dentine tissues.
The adverse effects of hydrogen
peroxide on enamel were evident in specimens
bleached in vitro but presence of saliva could
prevent the demineralizing effect of bleaching
agents in clinical conditions. It might be
proposed that remineralization of bleached
enamel is improved by application of highly
concentrated fluorides. It was found that the
frequent use of fluoride dentifrice resulted in
greater benefit in enamel surface rehardening,
with a similar effect on fluoride uptake, when
compared with its combination with a single
fluoride varnish application. These studies are
under investigation in our laboratory.
This study was supported by METU research
found: BAP-2004 07 02 -00-131.
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