Effect of Ethylene Diamine Tetra Acetic Acid and RcPrep to Microstrain of Human Root Dentin

Abstract

Background: Information about microstrain changes in human root dentine after application of chelating agent still limited. This result needed to determine the factors of microstrain which caused by non‑instrumentation of human root dentine formation. Materials and Methods: This experimental conducted in Department of Physic ‑ Faculty of Nature and Mathematics Science, Syiah Kuala University. All 45 specimens randomly divided base on experiment solutions. All specimens were
immersed for 15 min than powdered for X‑ray diffraction (XRD) analysis. Grain size calculated using Scherer equation and microstrain using Hall‑Plot. Result: XRD patterns typically show apatitic and amorf crystallized. A crystal phase of ethylene diamine tetra acetic acid (EDTA) group known as CaOH, RcPrep group as fluorapatite, and Control as Ca‑Cl, XRD pattern of EDTA confirmed as larger crystal than both
control and RcPrep group. XRD pattern of RcPrep group confirmed as larger crystal than the control group. The average value of grain size indicates all treatment groups have larger grain size than control. Microstrain of the control group showed as a lower value than all treatment groups. Microstrain of EDTA group showed as the biggest value than RcPrep and control. Microstrain of EDTA and RcPrep indicate larger than control. Regression analysis of between control and both treatment groups were significant (P < 0.05).
Microstrain of EDTA significantly greater than RcPrep. The relationships between EDTA to Control is weak (R2 = 0.37) same as relationship of control and RcPrep (R2 = 0.37). Conclusion: Chelating agent application into root canal is either change structure or improving microstrain of apatite crystal. Concentration and exposure time of chelating agent may lead as the main factor increasing fracture risk of human root dentine. Application of RcPrep into human root dentine is better than EDTA in relation to decreasing of dentin fracture. EDTA has more capability to increase of microstrain of apatite crystal than RcPrep.

Full text

32
Journal of International Oral Health 2016; 8(1):32-35
Microstrain of human root dentine … Nasution AI et al
Eect of Ethylene Diamine Tetra Acetic Acid and RcPrep to Microstrain of Human Root
Dentin
Abdillah Imron Nasution1, Cut Soraya2, Sun Nati3, Zulfan M Alibasyah4
Contributors:
1Department of Oral Biology, Faculty of Dentistry, Syiah Kuala,
Indonesia; 2Department of Endodontic, Faculty of Dentistry,
Syiah Kuala University, Aceh, Indonesia; 3Department of
Periodontology, Faculty of Dentistry, Syiah Kuala University,
Aceh, Indonesia; 4Department of Periodontology, Faculty of
Dentistry, Syiah Kuala University.
Correspondence:
Nasution AI. Department of Oral Biology, Faculty of Dentistry,
Syiah Kuala University, Aceh, Indonesia. Kopelma Darussalam,
Kota Banda Aceh, 23211. Tel.: 0651‑7551843/0651‑7551843,
Fax: 0651‑7551843/0651‑7551843. Phone: +618126988519.
Email: nasution@unsyiah.ac.id
How to cite the article:
Nasution AI, Soraya C, Sunnati, Alibasyah ZM. Eect of ethylene
diamine tetra acetic acid and RcPrep to microstrain of human root
dentin. J Int Oral Health 2016;8(1):32‑35.
Abstract:
Background: Information about microstrain changes in human
root dentine after application of chelating agent still limited. This
result needed to determine the factors of microstrain which caused
by non‑instrumentation of human root dentine formation.
Materials and Methods: This experimental conducted in
Department of Physic ‑ Faculty of Nature and Mathematics
Science, Syiah Kuala University. All 45 specimens randomly
divided base on experiment solutions. All specimens were
immersed for 15 min than powdered for X‑ray diraction (XRD)
analysis. Grain size calculated using Scherer equation and
microstrain using Hall‑Plot.
Result: XRD patterns typically show apatitic and amorf crystallized.
A crystal phase of ethylene diamine tetra acetic acid (EDTA) group
known as CaOH, RcPrep group as uorapatite, and Control as
Ca‑Cl, XRD pattern of EDTA conrmed as larger crystal than both
control and RcPrep group. XRD pattern of RcPrep group conrmed
as larger crystal than the control group. The average value of grain
size indicates all treatment groups have larger grain size than control.
Microstrain of the control group showed as a lower value than
all treatment groups. Microstrain of EDTA group showed as the
biggest value than RcPrep and control. Microstrain of EDTA and
RcPrep indicate larger than control. Regression analysis of between
control and both treatment groups were signicant (P < 0.05).
Microstrain of EDTA signicantly greater than RcPrep. The
relationships between EDTA to Control is weak (R2 = 0.37) same
as relationship of control and RcPrep (R2 = 0.37).
Conclusion: Chelating agent application into root canal is either
change structure or improving microstrain of apatite crystal.
Concentration and exposure time of chelating agent may lead as
the main factor increasing fracture risk of human root dentine.
Application of RcPrep into human root dentine is better than
EDTA in relation to decreasing of dentin fracture. EDTA has more
capability to increase of microstrain of apatite crystal than RcPrep.
Key Words: Dentine, ethylene diamine tetra acetic acid, microstrain,
RcPrep, X‑ray diraction
Introduction
Chelating agent is solutions consisting of a synthetic amino
acid which assist widening root canal properly because it has
the ability to softening hard tissue with low negative eect on
the periapical tissues. The most common chelating agents
used include ethylene diamine tetraacetic acid (EDTA) and
EDTA + urea peroxide and Carbowax (RcPrep).1
The application of chelating agent into root canal is known
to lead changes to human root dentine canal dimensions and
capable to decompose body wall of human root dentine canal.
The previous study stated chelating agents change human
root dentine hardness along with the length of application of
the chelating agent.2,3 These events indicate the occurrence
of increasing of microstrain in human root dentine. The
increasing of microstrain identified capable to increasing
fracture risk on human root dentine to lead improving
hypersensitivity during mastication and in worst condition
might tooth detached. Most serious cases of fracture should be
treating by endodontic surgery. One of most famous treatment
by endodontic approach is root resection.4
Information about microstrain changes in human root dentine
after application of chelating agent still limited. Meanwhile,
this initial information also needed to determine the factors
of microstrain which caused by non‑instrumentation of
human root dentine formation.5,6 Therefore, it is necessary to
investigate the dierences microstrain of human root dentine
after chelating agents application.
Materials and Methods
This laboratory experimental conducted in Department of
Physic‑Faculty of Nature and Mathematics Science, Syiah
Kuala University in January 2013. 15 extracted human
premolars were sectioned at the cementoenamel junction
using a diamond bur disc. Every group was then sectioned
horizontally in ⅓ cervical, ⅓ middle, and ⅓ apical to have a total
of 45 specimens. Based on the test solutions used, specimens
were divided randomly into three groups: (1) The EDTA
group: 0.05 mL of 17% EDTA for 15 min (n = 15), (2) the
RcPrep group, 0.05 mL of 15% EDTA‑urea peroxide‑Carbowax
Original Research
Received: 12th August 2015 Accepted: 19thNovember2015 ConictsofInterest:None
Source of Support: Nil

33
Journal of International Oral Health 2016; 8(1):32-35
Microstrain of human root dentine … Nasution AI et al
for 15 min (n = 15), and (3) the control group, 1 mL of 0.9%
saline for 15 min (n = 15).
All specimens were prepared for the determination of
microstrain of root dentin using X‑ray diffraction (XRD)
analysis. Grain size calculated using equation:
D =
KRad
rcos
λ
βθ
Where, D = Grain Size, K = Shape factor constant (0.9),
λ = Wavelength of the X‑ray (1.5405600), cos θ = cosine of half
the 2θ angle, and βr = Broadening of diraction line measured
at half its maximum intensity (radians). The microstrain of root
dentin calculated using equation:
K=
Bcos
sin
r
θ
θ
Wher w, ŋ = Microstrain; Br = Full width at half maximum
in radian; θ = Diraction angle. Furthermore, Microstrain
value statically analyzed using regression test with each
variabel (EDTA and RcPrep).
Results
All patterns were typically apatitic and amorf crystallized,
although some peaks of homogenous apatites were irregular
curves. In general, all specimens showed similar patterns
with the main character looks at the highest intensity in
2θ = 31°‑32°. Crystal phase of control showed as Ca‑Cl,
EDTA group showed as CaOH, and RcPrep group showed
as fluorapatite. General similar pattern of each group as
demonstrate in Figure 1.
The XRD pattern of EDTA conrmed as larger crystal than
both control and RcPrep group. XRD pattern of RcPrep
group conrmed as larger crystal than Control group. Data
normality test using Shapiro‑Wilk using Microsoft Excel
Analyze‑it version 3.90.5 revealed data is normal (P > 0.05).
The average value of grain size calculated using the Scherrer
equation indicates all treatment groups have larger grain
size than control. The largest of grain size crystals founded
in EDTA group. Grain size of each group as demonstrate in
Table 1.
The Microstrain of the control group showed as a lower value
than all treatment groups, while, microstrain value of EDTA
group showed as the biggest value than control and RcPrep.
The average value of microstrain indicates all treatment groups
larger than control. The microstrain values of each group as
demonstrate in Table 2.
Regression analysis of between control and treatment groups
were signicant (P < 0.05). The microstrain was signicantly
greater in 17% EDTA when compared to 15% RcPrep. The
relationships between EDTA application eect to Control in
the category of low and positive relation (R2 = 0.37). Regression
analysis between microstrain of control and RcPrep also
conrmed low and positive relation (R2 = 0.37). Regression
of each microstrain demonstrate in Table 3.
Discussion
The apatite group is a group of similar isomorphous hexagonal
phosphate minerals members were traditionally known as
fluorapatite, chlorapatite, and hydroxylapatite.7,8 It is well
established by XRD analysis that the mineral constituent
of human root dentine is essentially chlorapatite, and that
chlorapatite has a structure differing only in small details
from that of the well‑crystallized mineral hydroxylapatite
and uor‑apatite.9 The same pattern as displayed in 2θ with
high intensity which indicate all specimens is same groups in
apatite crystal. Each HA crystal is arranged by cell unit in lattice
arrangement of Ca and P, and also the lattice arrangement
of O and H.7 Moreover, we suggest chlorapatite is the main
structure of human root dentine. The main dierence, and in
fact, the only fully established one, between the two structures,
is that the hydroxyapatite has a slightly larger unit cell than
uor‑apatite or chlorapatite. Similar with this result, Elliot was
stated that apatites have the general formula, Ca10 (PO4) 6X2
where X is typically F (uorapatite, FAp), OH (hydroxyapatite,
OHAp), or Cl (chlorapatite, ClAp).7
As a displayed on result, the pattern of all specimen displayed
irregular curve and amorphous phases. Dentine is one of the
hard tissues in human body. This hard tissue formed by apatite
crystal. It diers with enamel which has more percentage of
inorganic. Dentin arranged by composition between inorganic
and organic is 70%, organic 20%, and water 10%, whereas
enamel is the hardest tissue in the human body and consists of
approximately 96% inorganic minerals, 1% organic materials,
Table 1: Grain size of each specimen.
Grain size
Control (nm) EDTA (nm) RcPrep (nm)
8.2 8.9 8.8
EDTA: Ethylene diamine tetra acetic acid
Table 2: Microstrain of each specimen.
Microstrain
Control EDTA RcPREP
0.895698047 0.979657791 0.902449062
EDTA: Ethylene diamine tetra acetic acid
Table 3: Regression of microstrain.
Description Regression
statistics EDTA
versus control
Regression statistics
RcPREP versus
control
R20.37 0.38
Coecient value (Fhit) 10.12 10.32
Relationship (Thit) 3.18 3.21
Relationship is signicant if thit>ttable and relationship is signicant if Fhit>Ttable. EDTA: Ethylene
diamine tetra acetic acid

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Journal of International Oral Health 2016; 8(1):32-35
Microstrain of human root dentine … Nasution AI et al
and 3% water.10,11 Although both tissues are comprised of a
calcium apatite mineral phase and a protein component, they
dier to overall structure include crystal size and shape, the
nature of the proteins present, and the relative proportions
of mineral and protein components. Dierences in structural
organization and composition give rise to mineralized tissues
with dierent properties that are well‑suited for their intended
biological purpose.12,13 Similar with this statement, our result
has found the grain size of these hexagonal‑member apatites
with the formula Ca5(PO4)3 X (X: Cl, OH, F) as found in
this experiment: D (nm)=8.2; 8.9; 8.8 for chlorapatite;
hydroxylapatite, and fluorapatite. The result has suggest,
chlorapatite is main structure of human root dentine,
application of both chelating agents EDTA and RcPrep into
root canal will change size, structure and microstrain of crystal
human root dentine.
Apatite crystal is Ca5 (PO4)3(X) which X is able to substitute
by other element. This substitution changes of lattice
arrangement of cell unit which improving grain size of apatite.
The apatite lattice is very tolerant of substitutions, vacancies,
and solid solutions, for example, X can be replaced by other
elements14 Thus, every cell units of apatite suggested ion Ca
position on the hexagonal corner to formed calcium column.
Ca position is perpendicular to C‑axis. Ca position is also on
cell central canal which formed Ca triangles. Spaces between
Ca columns were placed by two ions PO4 on the hexagonal
side. The F ion position is higher than OH position on C‑axis.13
The XRD data of human root dentin which applied to RcPrep
have slimmer peak than other group. This found suggest
human root dentin which applied to RcPrep have better
crystallinity. Calderyn,12 was reported that OH position on
C‑axis between ¼‑⅓, while ion F position is ¼ above the Ca
triangle. Application of RcPrep may cause a remineralization
in dentine crystals by F ion and aect crystal more strength
than EDTA application.
The main dierence, and in fact, the only fully established
one, between the two structures is that the hydroxyapatite
has a slightly larger unit cell than that of the uorapatite.
These conrm as human root dentin which applied to RcPrep
achieved dimensions close to enamel whereas has better
crystallinity than dentine. The previous study also stated 20%
of 85% organic of dentine is arranged by collagen.14,15 This
organic as conrm in XRD diractogram displayed as graph
or curve pattern is more amorf than enamel. This fact also
conrms every mineral have specic diractogram. Otherwise,
EDTA known has the function to clean the root canal as well as
bleaching agent which commonly used in dentistry, but EDTA
proved reducing mineral content of human root dentine.
Application of EDTA may cause a reduction in the calcium
crystals and increasing percentage of microstrain. These results
are consistent with Calt and Serper which states, chelating
agents have ability to demineralized human hard tissue.16
The increasing of microstrain which imply by EDTA which
properly known has capability to dissolving calcium which
is found in the building blocks of dentin. Calcium pulled
Figure 1: All specimens showed similar patterns with the main character looks at the highest intensity (↓) in 2θ = 31°‑32°.
X‑ray diraction (XRD) pattern of RcPrep group conrmed as better crystal than both control and ethylene diamine tetra acetic
acid (EDTA) group. XRD pattern of EDTA group conrmed as better crystal than control group.

35
Journal of International Oral Health 2016; 8(1):32-35
Microstrain of human root dentine … Nasution AI et al
out from crystal arrangement consequently affects crystal
structure. Microstain that occurs in dentin due to changes in
the structure or position of atoms will inuencing root dentin
resistance to accepting mechanical forces.17 A large grain
size of crystal in dissolving dentine and microstrain value,
automatically increasing of susceptibility of dentine to fracture.
Conversely, both grain size and microstrain of uorapatite as
an element which resistance to acid will constant to withstand
the mechanical forces.
The results of this study indicate that the application of
EDTA is increasing of susceptibility of human root dentine
to fracture. Similar with this statement, analyzed data
between microstrain controls when compare to EDTA
and RcPrep, statistically showed the signicant eect and
positive relationships. In other word, the increasing in
microstrain is aected by both chelating agents application.
In line with the results, analysis by regression conrms the
relationship of chelating agent to improving fracture‑risk is
positive and weak. It indicates chelating agent application
both EDTA and RcPrep predicted have 37‑38% factor which
able to increasing of fracture‑risk of human root dentine.
Concentration and exposure time of chelating agent may
lead to the main factor to increasing fracture risk of human
root dentine.
In addition, chelating agent application into root canal
will change the structure and improving of microstrain.
Compilation of microstrain between treatment groups to
chlorapatite as control showed application EDTA increasing
microstrain on crystal up to 9% while application of RcPrep
only 1%. Tsukamoto et al. stated that the crystallinity is strongly
inuenced by the crystal size and microstrain.18
Microstrain obtained from the small size of microstructures on
a uorapatite root dentine was lower up to 1% while has larger
grain size compared to normal [Table 2]. Based on this result,
application of RcPrep into human root dentine is better than
EDTA in relation to decreasing of dentin fracture.
Conclusion
Chelating agent application into root canal is either change
structure or improving microstrain of apatite crystal.
Concentration and exposure time of chelating agent may
lead as the main factor increasing fracture risk of human root
dentine. Application of RcPrep into human root dentine is
better than EDTA in relation to decreasing of dentin fracture.
EDTA has more capability to increase of microstrain of apatite
crystal than RcPrep.
Acknowledgments
The author would like to thank all the undergraduate students
of Faculty of Dentistry, Syiah Kuala University, Aceh, Indonesia
for their participation in this study.
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