Temporal relationship between sucrose –associated changes in

1. Original paper by: G.C. Vale; C.P.M.Tabchoury ; R.A.Arthur ; A.A.Del Bel Cury ;
A.F.Paes Leme ;J.A.Cury
Source:Journal of the European Organization for Caries
Research(ORCA),MARCH2007;41:406-412
PRESENTED BY :ROSHNI MAURYA

2.  Sucrose –unique cariogenic carbohydrate
 fermented to acids ; metabolized into EPS by
microorganisms of dental biofilm.

3.  Lower average pH induced by sucrose fermentation
triggers a shift in balance of resident plaque microflora
to a more cariogenic one (MARSH,2003)
 Low pH is responsible for enamel –dentin
demineralization, whose detection is time dependent
 Insoluble EPS change biofilm matrix, its porosity.
[Dibdin & Shellis, 1988]
 EPS may enhance bacterial adherence to the tooth
surface, plaque formation & accumulation.
 Relationship b/w EPS & En. Demin. Has been well
supported by in situ studies[Cury et al.,2000]

4.  EPS are considered critical virulence factors in
dental biofilm formed in presence of
sucrose[BOWEN, 2002]
 Simultaneously, dental biofilm exposed to
sucrose presents reduced conc. Of Ca,Pi & F.[Cury
et al.,1997],responsible for solubility equi.of
mineral tissue, their conc. in plaque fluid
determine pH below which enamel/dentin
demineralize.
 Although, a strong relationship b/w these
changes has been found[Cury et al.,1997,2000;
Aries et al.,2006], temporal relationship b/w these
changes in biofilm & en. Demin. Has not been
established, since in previous work they have
both been measured at end of experiment.

5.  The study aimed to evaluate the timing of
the effects of sucrose on biochemical and
microbiological composition of dental
biofilm and its relationship with enamel
demineralisation.

6.  Experiment design : study approved by Research & Ethics
Committee of FOP- UNICAMP (Protocol no.053/2004), crossover
blind design ,conducted in 3 phases.
 During these phases,12 volunteers wore acrylic intraoral palatal
appliances, each containing 6 enamel slabs (3 on each side).
 Volunteers were randomly assigned into 3 groups.
 In these groups, the slabs were treated e/o 8 times/day with
 Distilled & deionized water (-ve control)
 Mix. Of 10% glucose+ 10% fructose solution (active control)
 20% sucrose solution
 After 3,7,14 days of each phase, the biofilms formed were
collected from 2 enamel slabs at random for microbiological &
biochemical analyses ,for evaluation of mineral change in enamel.

7.  After a wash a washout period of least 7 days, new enamel
slabs were placed in the appliance for the next treatment.
 After 3 phases, all volunteers did all the treatment assigned.

9.  216 human enamel slabs (4x4x2mm) were obtained from middle
third of buccal & lingual surfaces of sound human impacted third
molars.( Cury et al.,2000)
 Surface of enamel was cleaned using pumice & polished to remove a
layer of 50micromt(Cury et al.1997).
 Slabs were randomly divided into 3 grps, acc. To treatments.
 An acrylic resin intraoral palatal appliance ,cont. 6 new slab placed
separately , 3 of them located on rt.& left post. Sides of appliance,
made for each volunteer for each phase of study.
 Plastic meshes were fixed to acrylic surface of appliances over
cavities to protect enamel slab surfaces from mechanical attrition,
leaving 1mm space for dental biofilm accumulation.

10.  12 volunteers (17-27 yrs old) fulfilled :
 Inclusion criteria : mean stimulated saliva flow rate>=0.7ml/min;
ability to comply with exp. Protocol
 Exclusion criteria: antibiotic use of last 2 mons before starting
the study,
 Use of any form of medication modifying salivary secretion,
 Use of fixed /removable orthodontic appliances,
 Periodontal disease
 General/ systemic illness
 Volunteers were informed about the procedures, written consent
was obtained prior to comm. Of study.

11.  Sugar sol. were prepared every 48 h,Vol. were instructed to remove
appliances from oral cavity & drip one drop of treatment solutions
onto each enamel slab acc.to treatment protocol, 8 times a day , at
predetermined times) (8.00,9.30,11.00,14.00,15.30,17.00,19.00&
21.00h)
 Excess of fluid was removed with gauze, after 5 min appliance was
reinserted in mouth. Vol. were also instructed to wear app. all time,
removing them only during meals
 a washout interval of 1 wk was established b/w exp. Phases[Cury et
al., 1997,2000]
 During a 7 day pre –exp period, & during washout periods & exp.
Phases, Vol. brushed their natural teeth & app. Except for the area
of enamel slabs , with nonfluoridated toothpaste.
 Vol. lived in an optimally fluoridated city ( 0.70mg F/ l, for the
region)

12.  On 3rd,7th,14th days of each exp. Phase,approx.10h after last
exposure to treatments, plastic meshes were removed using a
small scalpel, dental biofilm was collected with a sterile plastic
spatula from 2 enamel slabs at random one from each side of
app.
 On 3rd day, biofilm was collected from ant. Left & cent. Rt. Slabs
 On 7th day, from cent. left & post. Rt. Slabs
 On 14th day, from post. left & ant.rt. Slabs
 Plastic spatula was used to homogenize the sample & collect an
aliquot for micro. analysis, rest of biofilm was used for
biochemical analysis.

14.  At end of each exp. Phase, enamel slabs were removed from
app. Longitudinally sectioned 1mm from one lat. Edge,
embedded in acrylic resin , cut surfaces were exposed &
polished
 Cross-sectional microhardness(MH) values were converted to
mineral content (VOL.%) [Featherstone et al., 1983], &
integrated mineral loss( Z) for each treatment & time of
biofilm formation was cal.( Curey et al. 2000)

15.  Variables were analysed using split –plot
analysis (ANOVA), considering the treatments
as plots & time of biofilm formation as
subplots, with volunteers considered as
statistical blocks.

22.  The findings showed that undisturbed dental biofilm exposed 8
times/ day to sucrose significantly provoked an increase in
enamel Z as a function of time compared with –ve control
treatment group.[ Holeman et al.,1988;Tenuta et al.,2003,
respectively],showing that dental plaque completely undisturbed
for more than 1 wk causes detectable en.demin.
 Data also showed that biofilm treated with sucrose induced higher
enamel loss than exposed to glucose + fructose solution.
 The greater effect of sucrose on caries development in
comparison with its monosacc. Glucose & fructose may be
attributed to diff. in biofilm composition,as these carbohydrates
did not differ in acidogenicity. [Tenuta et al.,2006]
 Sucrose was able to sign. Inc. MS & LB counts in dental biofilm
compared with –ve control grp, but diff. with regard to glu.& fruc.

23. grp was not statistically significant
 Regarding biochemical changes induced by sucrose in
biofilm, the conc. of insoluble EPS was only the diff. from –ve
control and active control treatments.[Cury et al.,2000;
Tenuta et al., 2006],consistent with the role of polysacc. On
caries deve., enhancing bacterial adherence to enamel[
Bowen, 2002], changing the properties of biofilm matrix
[Dibdin & Shellis, 1988]
 Lowe Ca, Pi & F conc. In whole biofilms exposed to sucrose &
glu. + fruc. Compared with –ve control grp & absence of a
significant difference b/w carbohydrate treatment agrees
with previous studies [ Cury et al.,2000; Tenuta et al.,2006a]

24.  In summary ,among variables analysed, insoluble EPS
seems to be only the differential change induced by
sucrose on dental biofilm composition compared
with its monosaccharide components , although sign.
Ena. Demin. Occurs only after 7 days of biofilm accu.,
changes in biofilm composition are observed earlier.

26. THANK U!