_______________________________________________________________________________________________________________________________________________________________

Special Issue Article

_______________________________________________________________________________________________________________________________________________________________
Anticaries potential of a stabilized stannous-containing sodium fluoride
dentifrice

ROBERT V. FALLER, BS, SANDRA L. EVERSOLE, AAS & JANET YAN, MS

ABSTRACT: Purpose: To evaluate the anticaries potential of a stabilized stannous-containing sodium fluoride dentifrice
relative to appropriate control products. Methods: A series of in vitro studies was conducted using the following
standard anticaries efficacy measures: (1) fluoride uptake; (2) pH cycling remineralization/inhibition of
demineralization; and (3) surface microhardness. In each study, the stannous-containing sodium fluoride test dentifrice
(1450 ppm F) was compared to a negative control dentifrice (0 ppm F) and a positive control fluoride dentifrice (either
1100 ppm F or 1450 ppm F). Results: Fluoride uptake: The mean fluoride uptake from both the test dentifrice and the
positive control dentifrice was significantly greater than the negative control. There was no statistically significant
difference between the two fluoride dentifrices, although the test dentifrice was directionally higher. pH cycling: The
remineralization measured with the test dentifrice was directionally higher though not significantly different from the
positive control dentifrice. Remineralization by both fluoride-containing dentifrices was significantly greater versus the
negative control. Surface microhardness: The percent increase in surface microhardness measured on enamel surfaces
after treatments with the test dentifrice was found to be significantly higher than that measured for the positive control
and the negative control. (Am J Dent 2010;23 Sp Is B:32B-38B).

CLINICAL SIGNIFICANCE: These data confirm the ability of a stabilized stannous-containing sodium fluoride dentifrice
to fluoridate, strengthen and enhance the remineralization process in subsurface enamel lesions. In addition, these data
strongly suggest the potential for this product to deliver superior enamel care protection against external acid attack.


: Robert V. Faller, Principal Scientist, The Procter and Gamble Company, Advanced Technology and Innovation
Department, Enamel Care Research Group, 8700 Mason-Montgomery Road, Mason, OH 45040, USA. E-:
faller.r@pg.com

Introduction

Fluoride is well known and accepted for its ability to aid
in the reduction and prevention of dental caries.
1-4
Fluorides
ability to incorporate into the crystal lattice of demineralized
enamel results in the formation of more acid resistant forms of
apatite compared to that found in non-fluoridated tooth
mineral.
5,6
One of the most widely practiced and effective
ways to deliver fluoride is via dentifrice.
7

In addition to fluoride, many new dentifrice formulations
now contain additional ingredients, with the intention of
delivering additional consumer benefits. These added ingredi-
ents are usually designed to reduce calculus formation, control
plaque and gingivitis, prevent or remove stains, or alleviate
dentin hypersensitivity.
8
Inclusion of new agents is always
carried out with some degree of caution to ensure that new
ingredients do not adversely influence the anticaries activity
of the fluoride agent.
9,10
Several in vitro anticaries perform-
ance models are widely used and well accepted for the assess-
ment of the anticaries potential of new dentifrice formulas.
Fluoride uptake is one common laboratory assessment used
to measure the level of fluoride incorporated into early carious
enamel samples after a single treatment with the dentifrice
products. Fluoride uptake studies are an important research tool
for assessing the anticaries activity of new formulations and are
used to fulfill regulatory requirements,
11
as well as profes-
sional acceptance guidelines.
12
Increased fluoride incorporation
has been associated with reductions in dental caries in long-
term clinical studies,
11
increased remineralization
13-19
and
increased resistance to acid challenge.
20-24

Another study evaluates the ability of dentifrice products to
remineralize or reverse early carious lesions using an in vitro
pH cycling protocol designed to simulate the dynamic condi-
tions and natural challenges that occur in the oral cavity over an
extended period of time.
25-28
Decalcified, surface controlled
enamel samples are subjected to saliva soaks, product
treatments, and demineralization treatments to simulate the
effect of product usage over an extended time period. Cross-
sectional analyses of thin sections removed from each sample
post treatment allow analysis of lesion depth and mineralization
measurement on both control (pre-treated) and treated samples.
This approach provides the ability to not only measure lesion
reversal, but also assess the relative ability of each test product
to inhibit lesion progression.
A third well accepted study evaluates changes in surface
microhardness of enamel samples after a 7-day pH cycling
study. Surface microhardness is often used as a means to
measure the ability of a product to harden or remineralize the
outer 15-20 m of enamel surface,
29
or to protect enamel
against erosive tooth surface loss due to dietary acid chal-
lenges.
30
The key limitation of surface microhardness is the
inability of such measurements to provide a clear picture of
what may be happening deep within a lesion. Thus, the com-
bination of cross-sectional x-ray analysis and surface micro-
hardness can provide a unique perspective on the way in which
a specific product works, enabling an assessment of topical
(surface effects) as well as subsurface lesion repair.
In recent years, these methods have been used to evaluate
novel ingredients providing multiple benefits. For example,
White
31
reported on a novel ingredient, sodium hexameta-
phosphate, which simultaneously provides anti-calculus and
tooth whitening benefits. Importantly, the anticaries efficacy
American Journal of Dentistry, Vol. 23, Sp Is B, September, 2010

Table 1. Dentifrice products evaluated in each study.
________________________________________________________________________________________________________
Dentifrice products Major ingredients
________________________________________________________________________________________________________
Fluoride uptake study
Test product
a
1450 ppm F as NaF, SnCl
2
, silica abrasive
Positive control - USP Reference
Standard 1100 ppm F as NaF, silica abrasive
Negative control
b
0 ppm F, silica abrasive
Remineralization/inhibition of demineralization study
Test product
a
1450 ppm F as NaF, SnCl
2
, silica abrasive
Positive control - Crest Cavity
Prevention Dentifrice
(European formula)
a
1450 ppm F as NaF, silica abrasive
Negative control
b
0 ppm F, silica abrasive
Surface microhardness study
Test product
a
1450 ppm F as NaF, SnCl
2
, silica abrasive
Positive control - Crest Pro-Expert
Dentifrice (European formula)
a
1450 ppm F as SnF
2
, silica abrasive
Negative control
b
0 ppm F, silica abrasive
________________________________________________________________________________________________________
a
Procter & Gamble UK, Surrey, UK.
b
Procter & Gamble Company, Mason, OH, USA.

of this multi-benefit toothpaste was not compromised.
32,33

More recently, Crest Pro-Health toothpaste,
a
formulated with
a stabilized stannous fluoride and a silica abrasive system, has
been developed to deliver a wide range of user benefits.
Fortunately, this product does not cause tooth staining com-
monly associated with effective antimicrobial therapies.
34-38

Again, the anticaries efficacy of this unique stannous fluoride,
multi-benefit dentifrice was confirmed via in vitro evalua-
tions.
39
One interesting new aspect of this stabilized stannous
fluoride formulation was its demonstrated ability to protect
tooth surface enamel against dietary, erosive acid attack in a
human clinical study.
40

The aim of the current research was to evaluate the anti-
caries potential of a new sodium fluoride dentifrice with
stannous chloride compared to clinically proven dentifrice
formulations by conducting a series of in vitro studies using
the three standard test methods described above: (1) fluoride
uptake; (2) pH cycling (remineralization/inhibition of demin-
eralization after multiple product treatments); and (3) surface
microhardness after multiple treatments.
11,27,28,41

Materials and Methods
Products evaluated - Table 1 details the products and controls
included in these studies and the major ingredients in each
formula. The positive control in each study varied, depending
upon the objective of the study. The objective of the fluoride
uptake study was to determine whether the test product
provides at least as much fluoride to the enamel as a clinically
proven reference standard. As required by the U.S. FDA,
11
the
positive control for this study was selected to be the sodium
fluoride USP Reference Standard dentifrice.
b
The objective of
the pH cycling remineralization/inhibition of demineralization
study was to determine the impact of stannous chloride on the
mineralization properties of the formula. Here the positive
control was selected to be the European equivalent of the US
reference standard formula (i.e. 1450 ppm F version of the
formulation), as it allowed a direct comparison of these
parameters using formulations with matched levels of total
fluoride (1450 ppm F). The objective of the surface microhard-
ness study was to evaluate the impact of the test product rela-
Anticaries potential of dentifrice 33B



















Fig. 1. Enamel cores removed from extracted, human teeth.

tive to a formulation that has been shown in a human clinical
study to be highly protective of the enamel against dietary acid
attack.
40
The negative control was the same silica based placebo
dentifrice containing 0 ppm fluoride for all three studies.
Collection of human saliva - Ten healthy volunteers were
recruited to provide human saliva for these studies. Saliva
samples were collected from the volunteers each day of the
study, pooled and stored under refrigeration until use. All
required precautions were in place to ensure proper handling
of saliva from the point of collection to the ultimate use in the
laboratory studies. Volunteers chewed paraffin wax, and
expectorated any stimulated saliva generated into a plastic
collection vessel over a period that averaged 30-40 minutes
per volunteer per collection period. Saliva was collected early
in the morning from each volunteer on each day of the study
in order to maintain a relatively constant pool of saliva for use
in the study. Once completed, collection vessels were pooled
together, mixed and stored under refrigeration at approxi-
mately 5C until use.
Specimen collection and preparation - Enamel samples were
prepared from human teeth for all studies. Human upper
incisors were obtained from local oral surgeons who collected
the teeth after removing them, typically for orthodontic
reasons. All required precautions were in place to ensure
proper handling of tooth samples from the point of collection
to the ultimate use in these laboratory studies. Available teeth
were individually cleaned and checked for any visible surface
cracks or other imperfections. Those with any visible imper-
fections were discarded. Teeth were stored prior to use under
refrigeration (approximately 5C) in a 1% thymol solution.
Enamel specimens for all studies were prepared by cutting
3 mm-diameter cores from the collected teeth using a
diamond core drill (Fig. 1). Enamel cores were mounted on
plastic rods using dental acrylic (Durabase
c
), covering all
sides except the natural facial surface. Coarse polishing with
600-grit silicon carbide/water slurry was used to remove
approximately 50 m of the outer enamel. After coarse
polishing, the surface of each specimen was polished again
for approximately 1 hour to a mirror finish with Micropolish
No. 3B gamma alumina.
d
Enamel cores found to have surface
imperfections were rejected. Internal studies have shown this
procedure results in the presentation of a renewed enamel sur-
34B Faller et al



face that is essentially free of background fluoride.
Decalcification treatment - Specimens used in these studies
were subjected to a decalcification pre-treatment designed to
produce early caries lesions in the enamel samples. Investi-
gations using decalcified enamel specimens are thought to be
more predictive of potential anticaries efficacy. This is because
fluoride is known to not only prevent demineralization of
healthy enamel, but also promote remineralization of caries
lesions.
14,22,42-44
The remineralization component of anticaries
activity would not be measurable without the initial decalcifi-
cation step that creates spaces in or on the crystal for fluoride to
incorporate or remineralize. As a result, decalcified specimens
are typically used in fluoride uptake studies and have also been
used in in vitro cycling model studies.
17,18,26-28
The importance
of fluoride uptake in demineralized enamel was demonstrated
by Sakkab et al,
41
who found a significant association between
higher levels of fluoride incorporated into demineralized
enamel along with lower caries rates of those treatment groups
with the highest levels of incorporated fluoride.
Enamel specimens were decalcified using a weak acid
containing solution (pH= 5.0) of 0.1 M lactic acid and 0.2%
polyacrylic acid (Carbopol C907
e
) 50% saturated with
hydroxyapatite, and prepared according to the method of
White.
18
This method is known to produce early caries lesions
with depths between 40 and 80 m depending upon the
treatment time.
Fluoride uptake study - The fluoride measurement approach
used here is based on the US Food and Drug Administration
Test Method 40.
11
The overall approach has been shown by
Pfarrer et al
39
to have the sensitivity to measure differences
between clinically effective and clinically ineffective denti-
frice formulations at the 1,100 ppm fluoride level using only
four specimens per treatment group.
Specimen decalcification - Decalcification of the enamel was
carried out for 72 hours at 37C using the approach described
above. Treatment for this period of time produces early caries
lesions with an initial depth in the range of 40 to 50 m.
Determination of indigenous fluoride - After decalcification,
the microdrill biopsy technique described by Sakkab et al,
41

was used to measure the indigenous fluoride level in each
enamel core and to stratify the samples into treatment groups.
Specimens were mounted on the microdrill stage and sampled
using a miniature carbide end-milling tool. The biopsy tech-
nique removed a small portion of the core, leaving behind a
cylinder with a diameter of approximately 450 m 10 m
and a height of approximately 75 m, ensuring sampling of the
enamel through the depth of the demineralized lesion area and
into the underlying sound enamel. The depth of each sampling
was verified using the Leitz Axioskop microscope
f
fitted with a
digital Microcode II.
g
Two drillings were removed from each
specimen. The powder created during drilling was collected,
combined, and dissolved in 150 L of 0.5 M perchloric acid.
A solution of the dissolved powder for fluoride ion selec-
tive electrode analysis was prepared by adding 150 L of total
ionic strength adjustment buffer
h
and the pH was adjusted to
approximately 5.4 by the addition of 100 L of 1N sodium
hydroxide. Sample solutions were analyzed by reading the
millivolt potential with a calibrated fluoride ion-specific
electrode (Model 96-09-00
i
). Fluoride concentration was de-
American Journal of Dentistry, Vol. 23, Sp Is B, September, 2010
























Fig. 2. Treatment cycle used in pH cycling study: remineralization/ inhibition
of demineralization study.

termined from a calibration curve obtained on the same day as
the analysis. Specimens were rank-ordered by indigenous
fluoride level and stratified into treatment groups. No statis-
tically significant differences between groups were observed
for indigenous fluoride level (P< 0.05) by the least significant
difference (LSD) method.
Specimen treatment - Treatment slurries were made by
thoroughly mixing 10g of dentifrice and 30g of pooled, whole
human saliva using a non-aerating mixing technique. The
resultant slurry was centrifuged for 15 minutes at 10,000 rpm
to produce a clear supernatant free of suspended solids.
Groups of specimens (n= 4) were suspended in supernatants
for a period of 30 minutes with mixing.
Analysis of post-treatment fluoride levels - After treatment,
the samples were thoroughly rinsed and again subjected to
microdrill biopsy to determine post-treatment fluoride levels.
Indigenous fluoride levels were subtracted from post-
treatment fluoride levels to determine fluoride uptake. Data
collected were reported as micrograms of fluoride per unit
area sampled (g/cm
2
). The mean fluoride uptake for each
treatment was calculated and then compared using the LSD
test at a 5% significance level ( = 0.05).
pH cycling study: Remineralization/inhibition of deminerali-
zation - The in vitro pH cycling model used in this study (Fig.
2) is designed to simulate the natural challenges that occur in
the oral cavity.
13,18,25
Demineralized enamel specimens were
exposed to pooled, human saliva to initiate the formation of a
pellicle coating on the enamel surfaces. Specimens were treated
four times a day with a prepared slurry of the dentifrice (one
part test product: three parts pooled saliva) to simulate dilution
of product that occurs during actual product use. All specimens
were challenged each day for a 3-hour period with a demineral-
ization solution to simulate the drop in pH that occurs after
ingestion of fermentable carbohydrates
45
and which is capable
of initiating and stimulating the overall caries process. This
*indicates saliva soak was replenished with fresh saliva
American Journal of Dentistry, Vol. 23, Sp Is B, September, 2010




















Fig. 3. Preparation of controlled enamel specimen for treatment and
sectioning for x-ray analysis. Cores of enamel were removed from extracted,
human teeth.

cycling protocol has been used previously to demonstrate
significant differences in marketed product performance.
26,28
It
exhibits excellent dose response sensitivity, and provides a
useful tool for predicting potential clinical effectiveness of
fluoride-containing formulations.
26-28

Specimen pretreatment - Prior to use, enamel samples were
stored in a 1% thymol solution under refrigeration (approxi-
mately 5C). The samples were decalcified according to the
method of White
18
for 96 hours producing early caries lesions
with a depth of approximately 50-80 m. Specimens were then
thoroughly rinsed with deionized, distilled water. Approxi-
mately 1/3 of the surface of each specimen was covered with an
acid resistant nail polish (Fig. 3) to serve as a reference area
during the subsequent analysis. Specimens were individually
numbered and placed in specially designed holders (four
specimens per group) which allowed each group to be treated
simultaneously.
Specimen treatment - Enamel samples were stored for 1 hour in
vessels containing pooled human saliva to initiate the formation
of a pellicle layer on the surface of the samples. Specimens were
then subjected to four treatment sequences per day for a total of 5
treatment days (Fig. 2). The treatment sequences consisted of a
dentifrice slurry treatment [one part dentifrice: three parts fresh
pooled, human saliva (w:w)] followed by saliva remineralization.
A demineralization period occurred between the second and third
treatments each day. The demineralization solution was the same
as that used to form the initial caries lesions.
Saliva baths were refreshed three times each day as follows:
(1) in the morning after the first treatment; (2) after the daily
period of demineralization; and (3) at the end of each work day.
Saliva baths were continuously stirred with a mechanical
magnetic system. Each group of specimens was removed four
times a day and treated with a slurry consisting of three parts
(15g) of fresh pooled human saliva to one part (5g) dentifrice.
Slurries were prepared fresh for each treatment and were mixed
with a mechanical magnetic stirring system to make slurries
homogenous for a period of approximately 4 minutes prior to
the actual treatment of demineralized samples. Each treatment
lasted 1 minute. Between the second and third treatments each
Anticaries potential of dentifrice 35B














Fig. 4. Results of transverse microradiographic analysis are used to determine
Z by comparing (a) the lesion control area to (b) the treated (remineralized)
area (x200 magnification).

day, each group of five samples was stored for 3 hours in a
fresh volume (~18 ml) of the demineralization solution (Fig. 2).
Specimen analyses - After 5 days of treatment, specimens were
rinsed well in distilled deionized water and stored refrigerated
in a humid environment. Specimens were cut plano-parallel
using a hard tissue sectioning saw (Silverstone-Taylor hard
tissue microtome
j
). Each section was cut to allow the control
and treated portion to be represented for analysis (Fig. 3). A
thin section (~100 m) was removed from each specimen and
placed flat on a specially designed holder that fits adjacent to an
aluminum stepwedge (steps ranged from approximately 25-200
m in thickness). These sections were then exposed to CuK
radiation, and microradiographs were taken using Kodak
SO253 Holographic film.
k
Standard black and white film
developing methods were used. Radiographs were analyzed
using Transverse Microradiography,
l
a computer-based image
analysis system using the aluminum stepwedge for calibration.
Sound enamel beneath each lesion was normalized to a value of
87 volume% mineral. Mineral profile scans were made from
the areas of interest (control and treated). Analysis of the lesion
body, known as the Delta Z (Z) value for each area, was
calculated as the difference between sound enamel and the
lesion. The change in mineral from the initial lesion to the
lesion exposed to the pH cycling treatments was recorded as the
remineralization value (Z). Images representing controlled
and treated specimens are presented in Fig. 4.
Surface microhardness study - This study was a 7-day cycling
study incorporating four treatments per day to simulate 2 weeks
of product use, two brushings per day.
Specimen preparation and treatment - Specimen preparation
for the surface microhardness study was the same as that used
for the pH cycling: Remineralization/inhibition of demineral-
ization study. The only differences were the following: (1) the
inclusion of surface microhardness measurements before and
after the study; and (2) the extension of the study to 7 treatment
days (14 treatments) to simulate 2 weeks of product use.
Determination of surface microhardness on human enamel pre-
pH cycling - In our laboratory we used Vickers hardness as a
preferred method for assessing surface microhardness, whereas
Knoop hardness is routinely employed for cross-sectional
microhardness assessment. The Vickers diamond
d
provides the
ability to measure the impact of the diamond in two directions,
which in our technical judgment provides a more useful
measure on enamel surfaces than the single length measurement
36B Faller et al


















Fig. 5. Vickers Hardness Indentations. Three measurements taken at three
distinct points on tooth enamel surface, then averaged.

Table 2. Baseline hardness values (all specimens).
________________________________________________________________________________________________________
Specimen HV1 HV2 HV3 Mean HV
________________________________________________________________________________________________________
1 42.5 31.3 39 37.60
2 45.1 35.7 40.7 40.50
3 38.8 33.4 27.5 33.23
4 31.2 48.1 32.7 37.33
5 63.8 65 69 65.93
6 53.9 51.2 30.2 45.10
7 64.3 71.4 72.2 69.30
8 26.7 28 28.6 27.77
9 48.6 41.7 41.6 43.97
10 52.1 50.7 42.7 48.50
11 28.5 18.4 22.7 23.20
12 27.1 26.8 33.6 29.17
13 30.2 51.1 45.6 42.30
14 40.8 37.4 34.3 37.50
15 43.2 48.8 53 48.33
________________________________________________________________________________________________________

Table 3. Baseline surface microhardness measurements (by group).
________________________________________________________________________________________________________
Group Specimen Initial HV Mean HV/Group
________________________________________________________________________________________________________
1 10 48.50
2 40.50
1 37.60
8 27.77 38.59
2 15 48.33
13 42.30
14 37.50
12 29.17 39.33
3 6 45.10
9 43.97
4 37.33
3 33.23 39.91
________________________________________________________________________________________________________

of the Knoop diamond.
After the treatment phase, each specimen was analyzed for
surface microhardness with a Buehler Micromet microhardness
tester
m
using a Vickers diamond at a weight of 200g and dwell-
time of 15 seconds. Hardness indentations were made three
times on each surface by visually dividing the specimen into
three equal, pie shaped parts and taking an indentation within
each piece of the pie (Fig. 5). Hardness numbers were recorded
for each of these three indentations, and the number averaged
for each specimen (Table 2). Average values were used to place
specimens in groups such that the initial surface hardness of
each group was not significantly different (Table 3). Average
hardness values in the range of VHN = 25-50 were targeted, as
American Journal of Dentistry, Vol. 23, Sp Is B, September, 2010


Table 4. Comparison of fluoride uptake tesults.
________________________________________________________________________________________________________
Statistical
Dentifrice product Mean fluoride uptake (S.D.) ranking*
________________________________________________________________________________________________________
Test product
(1450 ppm F as NaF + SnCl
2
) 14.18 (3.68) a
Positive control - USP reference
standard (1100 ppm F as NaF) 12.21 (0.64) a
Negative control placebo -0.72 (1.14) b
________________________________________________________________________________________________________
*Means with the same letter are not significantly different (= 0.05, N=4) by
the S-N-K test.

Table 5. Comparison of remineralization of caries lesions.
________________________________________________________________________________________________________
Z* (Volume % mineral Statistical
Dentifrice product x m) (SEM) ranking**
________________________________________________________________________________________________________
Test product (1450 ppm F as NaF + SnCl
2
) -424.87(225.70) a
Positive control (1450 ppm F as NaF) -410.67 (94.01) a
Negative control placebo 187.67 (91.98) b
________________________________________________________________________________________________________
*Z represents the change in mineral level. A more negative value for Z
indicates that more mineral was incorporated into the initial caries lesion.
** Different letters indicate results are significantly different (P< 0.05, N= 4) by
the Fisher LSD test.

Table 6. Comparison of surface microhardness.
________________________________________________________________________________________________________
Hardness value (HV) % Statistical
Dentifrice product Initial Final HV Increase ranking*
________________________________________________________________________________________________________
Test product (1450 ppm
F as NaF + SnCl
2
) 39.33 128.89 89.56 227.7 a
Positive control
(1450 ppm F as SnF
2
) 38.55** 70.50 31.94 82.8 b
Negative control placebo 38.59 45.31 6.72 17.4 c
________________________________________________________________________________________________________
* Different letters indicate results are significantly different (P< 0.05, N=4), by
the Fisher LSD test.
** n= 3, due to loss of one specimen during treatment phase of study.

this range of values has generally produced specimens with
excellent reliability and performance in this model system (data
on file, P&G). The remaining specimens with microhardness
values outside this preferred range (specimens 5, 7 and 11)
were discarded. This procedure ensured the baseline level of
demineralization was consistent across all groups at the start of
the study.
Determination of surface microhardness on human enamel
post-pH cycling - Each specimen was re-analyzed for surface
microhardness. Hardness numbers were again recorded three
times on each specimen in an area adjacent to each of the
original indentations using a Vickers diamond at a weight of
200g and dwell time of 15 seconds. The average hardness
numbers were calculated for each specimen.
Analysis - Difference in hardnesss values (post - pre) was calcu-
lated and reported. The percent change in surface micro-
hardness was calculated.
Results

Results in Table 4 show that mean fluoride uptake from
enamel samples treated with the test product containing
stannous chloride was found to be directionally higher, though
not significantly different, from that measured from samples
treated with the positive control, the USP Reference standard.
As expected, fluoride uptake from the negative control
containing no fluoride was found to be significantly lower than
Vickers Hardness
Indentations
American Journal of Dentistry, Vol. 23, Sp Is B, September, 2010



either of the fluoride-containing formulas.
Results in Table 5 show the remineralization measured from
enamel samples treated with the test product containing
stannous chloride was found to be directionally higher, but not
significantly different, from the positive control. As expected,
remineralization from the negative control with no fluoride
agent was found to be significantly lower than the other pro-
ducts. In fact, the positive Z value measured for the placebo
dentifrice is an indication of further demineralization resulting
over the course of the study resulting from the use of the
negative control dentifrice.
Results in Table 6 show a significant increase in surface
microhardness measured on enamel surfaces treated with the
test dentifrice compared to those treated with the positive
control. As expected, the findings from surfaces treated with
the negative control were found to be significantly lower than
surfaces treated with the other products tested.

Discussion

Results show that this combination of in vitro studies is a
useful way to evaluate the anticaries potential of new dentifrice
formulations. Fluoride uptake studies provide an indication of
the bioavailability of fluoride in the diluted formula and its
ability to incorporate into early caries lesions relative to
clinically proven formulations. The in vitro pH cycling protocol
used in these studies is designed to simulate the dynamic
conditions and natural challenges that occur in the oral cavity
over an extended period of time.
13,18,25
This allows for an
assessment of changes in the enamel properties over time, after
a series of saliva soaks, product treatments, and acid treatments.
X-ray analysis of untreated and treated lesion areas provides an
assessment of the ability of each product to progress or reverse
the size and intensity of the early caries lesions produced by the
decalcification pre-treatment. Surface microhardness assess-
ments before and after cycling provide an assessment of the
extent of protection of the enamel surface after continued
exposure to each of the test products. This combination
approach provides a more holistic technical assessment of the
anticaries potential of a new dentifrice formulation, compared
to any one of the measures taken individually.
The mean fluoride uptake values measured in this study
after treatment of the demineralized enamel samples were
found to be between -0.72 and 14.18 g F/cm
2
, depending on
the dentifrice. The negative control did not contain any
fluoride, and enamel samples treated with this paste showed a
net loss in average fluoride content, compared with the baseline
values. In this study, the highest fluoride uptake was achieved
after use of the dentifrice containing a combination of sodium
fluoride and stannous chloride. Although this product did
contain slightly higher levels of total fluoride compared to the
positive control (1450 ppm F vs. 1100 ppm F, respectively), the
purpose of this test was to determine if the new product would
deliver fluoride to demineralized enamel at a level that was at
least equal to a clinically proven fluoride control dentifrice. The
1100 ppm F (NaF) dentifrice included in this study was a
clinically proven formulation that serves as a gold standard in
anticaries effectiveness. Delivery of fluoride at a level at least
equivalent to that formulation, as the new product does, helps
ensure effectiveness of the new formulation. It is clear from the
Anticaries potential of dentifrice 37B




results that the addition of stannous chloride to this formulation
has not interfered with the uptake of fluoride into the enamel.
Results of the remineralization/inhibition of demineraliza-
tion study clearly demonstrate the ability of the stannous-
containing sodium fluoride test dentifrice to promote the
mineralization of enamel, as evidenced by significant reversal
of the demineralized enamel lesions compared to the negative
control. Both fluoride containing products performed signifi-
cantly better than the placebo (negative) control. Remineraliza-
tion values for the test dentifrice were not significantly different
from the matched control without stannous chloride. Results of
this study confirm the ability of the test dentifrice to reminer-
alize softened enamel, and further confirm that the inclusion of
stabilized stannous does not adversely impact the minerali-
zation process.
Results of the surface microhardness study clearly demon-
strate the ability of the two fluoride containing dentifrices to
significantly harden softened enamel relative to the negative
control. Interestingly, the test product containing both sodium
fluoride and stabilized stannous chloride showed a significantly
higher surface microhardness than the positive control with
sodium fluoride alone. Considering the fact that a formulation
similar to the positive control toothpaste (identical except for a
lower level of total fluoride, i.e. 1100 ppm F rather than 1450
ppm F) has been demonstrated to provide superior enamel
protection against erosive acid attack,
40
these results are
particularly striking. This enhanced level of enamel surface
microhardness suggests that this sodium fluoride dentifrice with
stannous chloride could provide an even higher level of
protection against erosive acid challenge than that measured
previously by Hooper et al.
40
Previous studies have demonstra-
ted the ability of both stannous fluoride and stannous chloride
to deposit a stannous containing phase onto enamel surfaces.
46

Data from this study appears to support the previous obser-
vations. The test product should be evaluated further to determine
its ability to protect enamel against erosive acid attack.
Each of the models included in this series of studies
contains key elements important to the caries process. In addi-
tion, each of the models allows us to understand more fully
how different products are able to interact with enamel to
deliver potential benefits. Taken as a whole, the results strongly
support the anticaries potential of the new, stabilized stannous-
containing sodium fluoride dentifrice, and strongly suggest the
potential for this toothpaste to provide significant benefits with
respect to protection of the enamel against tooth surface loss
due to erosive acid challenges. Further studies to determine
whether the highly significant enhancement in surface micro-
hardness observed with this formulation will provide enhanced
surface protection effects are warranted.
a. Procter & Gamble Company, Cincinnati, OH, USA.
b. US Pharmacopeia, Rockville, MD, USA.
c. Reliance Manufacturing Company, Worth, IL, USA.
d. Buehler Limited, Lake Bluff, IL, USA.
e. B.F. Goodrich Company, Cleveland, OH, USA.
f. Leica Microsystems Inc., Bannockburn, IL, USA.
g. Boeckeler Instruments, Inc., Tucson, AZ, USA.
h. Ricca Chemical Co., Arlington, TX, USA.
i. Orion, Beverly, MA, USA.
j. Scientific Fabrications, Littleton, CO, USA.
k. Eastman Kodak Company, Rochester, NY, USA.
l. Inspektor Research Systems BV, Amsterdam, The Netherlands.
38B Faller et al




Acknowledgement: To Anita Guy for assistance with the manuscript
development.
Disclosure statement: Mr. Faller, Ms. Eversole, and Ms. Yan are full-time
employees of the Procter & Gamble Company.
Mr. Faller is a Principal Scientist and Ms. Eversole is a Principal Researcher,
The Procter & Gamble Company, Health Care Research Center, Mason, OH,
USA. Ms. Yan is a Principal Researcher at the Procter & Gamble Company
(Beijing) Technology Co., Ltd., Beijing, China.

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