Realtime monitoring of dentine dissolution by citric acid

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Realtime monitoring of dentine dissolution by
citric acid Siân B Jones1, R Peter Shellis1,
Gareth D Rees2 and Michele E Barbour1 1University
of Bristol, UK, 2GlaxoSmithKline, Weybridge, UK
Objectives The objective of this study was to
investigate dentine dissolution in citric acid in
real time. The method used is well established
for investigating enamel dissolution, but has not
previously been used to investigate dentine.
Dentine is a more complex tissue than enamel, and
its dissolution kinetics are not well
understood. Aim 1 To apply a pH-stat technique
to investigate dentine dissolution in
real-time. Aim 2 To determine dentine
dissolution rate as a function of pH, acid
concentration and added calcium or phosphate. Aim
3 To further develop the model to investigate
inhibitory effect of pre-treatment with fluoride
on the rate of dissolution.
Methods Dentine sections were prepared from
sound third molar roots, polished, rinsed and
ultrasonicated. Dissolution was investigated in
0.3, 1 and 1.5 citric acid solution at pH 3.20
and in 1 citric acid at pH 2.90 3.75 . The
addition of 20 mM calcium or phosphate to 1
citric acid was investigated at pH 3.20 and 3.75.
Some dentine specimens were pre-treated with 1500
ppm fluoride (NaF) for 30 min prior to measuring
dissolution in citric acid solution. The reaction
vessel was well stirred and maintained at 36 C.
The smear layer on the dentine surface was
investigated by SEM. Specimens with no acid
treatment were compared against specimens treated
with 1 citric acid at pH 3.75 for 30 seconds.
The cross-sectional area of each dentine specimen
was measured, and dissolution rates normalised to
area. The data from the pH-stat gave real-time
dissolution rate as the amount of H added to the
solution correlated to dissolution of the dentine
surface. The amount of H added 30 min after
commencement of the reaction, when the reaction
started after 10 nmol of H was added, was
recorded. ANOVA with Tukey as post hoc or
Mann-Whitney U test was carried out at p lt 0.05.
Figure 1. Real-time
read-out showing amount of H required to keep
solution at a constant pH. Value taken after
1800 s.
a
b
Figure 2. a) SEM image of smear layer on dentine
surface and b) after removal of smear layer
following 30 s acid treatment.
Results The dissolution rate of dentine
decreased as a function of exposure time (Figure
1). There are several possible ways to analyse
this data for this investigation the amount of
H added after 1800 s was chosen. The SEM
images (Figure 2) show the smear layer on the
surface as a consequence of specimen preparation
but this is removed following a 30 s acid
treatment exposing the characteristic dentinal
tubules. Figures 3 to 6 show the effect of
various parameters on dentine dissolution. The
concentration of the citric acid had no
significant effect on dentine dissolution.
Dissolution rate was negatively correlated with
pH. The addition of calcium or phosphate had no
significant effect on the rate of dissolution at
either pH value. Figure 7 shows that pre-treating
dentine with 1500 ppm fluoride had an inhibitory
effect on dentine dissolution.

• Conclusions
• pH stat is an effective method to investigate
  dissolution rate of dentine in real time
• Dissolution rate was negatively correlated with
  pH
• Acid concentration had no statistically
  significant effect on dentine dissolution rate
• Addition of 20 mM calcium or phosphate had no
  statistically significant effect on dissolution
  rate in citric acid at pH 3.20 or 3.75
• Pre-treatment with 1500 ppm fluoride for 2 min
  resulted in a 20 reduction in dissolution rate

This research was supported by GlaxoSmithKline