Journal of
Dental Research
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Changes in Plaque pH in vitro by Sweeteners
B.G. Bibby and J. Fu
J DENT RES 1985 64: 1130
DOI: 10.1177/00220345850640090601
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Changes in Plaque pH in vitro by Sweeteners
B.G. BIBBY and J. FU
Eastman Dental Center, 625 Elmwood Avenue, Rochester, New York 14620
A new in vitro pH method was used to compare the ability of natural
dental plaque to convert nine sweeteners to acid. Tests made in 0.1,
1.0, and 10.0% concentration, or sucrose sweetener equivalents, showed
that isomaltulose gave the lowest pH, followed in order by Lycasin,
mannitol, palatinit, sorbitol, sorbose, and then, with negligible acid
production, aspartame, saccharine, and xylitol. Unlike the other
sweeteners, xylitol and saccharine gave smaller pH depressions in
higher concentrations of test solutions. At the concentrations tested,
none of the sweeteners interfered with acid production from sucrose.
did not interfere with sucrose fermentation. Jensen (1984) tested
several starch hydrolysates and found that they gave depressions of plaque pH comparable to those given by sorbitol.
The foregoing summary of tests relating to acid production
from sweeteners shows that, except for what has been reported
on the sugar alcohols, there is little solid information on the
fermentation of sweeteners in the mouth. For that reason, we
have used a new plaque pH method (Bibby and Krobicka,
1984) to run tests on acid formation from a series of sweeteners.
J Dent Res 64(9):1130-1133, September, 1985
Introduction.
The possibility of combatting caries by using sweetening agents
other than sucrose has prompted comparisons of their abilities
to form acid in the mouth. For this purpose, tests in cultures
of oral bacteria, in saliva, plaque suspensions in fluids, or in
vivo dental plaque have been used. The first of these procedures showed that the majority of the strains of streptococci,
lactobacilli, and actinomyces tested fermented sorbitol but not
xylitol (Edwardsson et al., 1977; Miihlemann et al., 1977;
Drucker and Verran, 1980), but that xylitol-fermenting strains
of streptococci, lactobacilli, or propionibacteria were found in
the mouth (Edwardsson et al., 1977; Gallagher and Fussell,
1979). While Muhlemann et al. (1977) and Drucker and Verran (1980) found that xylitol did not interfere with acid production from other sugars, Vadeboncoeur et al. (1983) obtained
contrary findings. Two strains of S. mutans did not ferment
isomaltulose (Takazoe et al., 1981).
Using saliva, Miihlemann and Schneider (1975) reported no
acid formation from xylitol; neither did it inhibit fermentation
of sucrose. When aspartame was added to incubating glucose
and saliva, there was a slight increase in lactic acid production,
along with a smaller depression of the pH (Mishiro and Ka-
neko, 1977).
Tests made in aliquots of dilute suspensions of plaque generally confirmed the results obtained with bacterial cultures
and saliva. They showed that little, if any, acid was formed
with xylitol. Of the other sugar alcohols tested, mannitol gave
least acid, followed by sorbitol, maltitol, and the starch hydrolysate Lycasin (Hayes and Roberts, 1978; Birkhed, 1978).
In anaerobic tests, isomaltulose was a poor source of acid but
gave more acid than did sorbitol (Maki et al., 1983).
Measurements of pH changes occurring in vivo in plaque on
the teeth have been made either by removing plaque samples
from the mouth for immediate in vitro pH measurement, or by
use of in-dwelling pH electrodes on which plaque has been
allowed to develop. Using the former procedure, Frostell (1973)
found that mannitol and sorbitol gave only small depressions
of the pH and that more acid was produced from Lycasin.
Using in-dwelling pH electrodes, Miihlemann and Boever (1970)
and Miihlemann et al. (1977) reported limited acid production
from sorbitol and sorbose but none from xylitol, which also
Received for publication November 29, 1984
Accepted for publication May 31, 1985
1130
Materials and methods.
pH test.-The procedure used for measuring pH changes in
plaque has been fully described elsewhere (Bibby and Krobicka, 1984). Plaque freshly collected from the teeth was spun
to the bottom of a wire gauze basket into which a combination
microelectrode (Microelectrodes, Inc., MI710), connected to
a pH meter and strip chart recorder, was inserted. Tests were
run in an incubator at a temperature of 37.5°C. The plaque
assembly was first incubated in 1.5 ml of water for 20 min,
then washed with a stream of water from a wash bottle, neu-
tralized in 1.5 ml of stirred saliva, and washed again.
Thereafter, the electrode assembly was immersed in 1 ml of
a 0.1, 1.0, or 10% solution of a sweetener and the pH changes
recorded for 20 min, at which time the pH had stabilized at
its lowest point. After washing in a stream of water and neutralization in saliva, the test was repeated in the next higher
concentration of sweetener. After three to six sweetener tests,
a final run was made in 10% sucrose to determine whether the
plaque sample had retained its full activity throughout the series of tests. If the pH was not depressed to 4.5 or below, the
preceding tests were regarded as unsatisfactory.
Plaque.-Plaque samples were collected with dental instru-
ments from all accessible tooth surfaces of adult volunteers
who had suspended oral hygiene for 24 to 48 hr and who had
not eaten for two hr prior to plaque removal.
Test materials.-The sweeteners used and their sweetness
relative to sucrose (s= 1) were: Lycasin, mainly sorbitol
(s=0.5); sorbitol, C6HI406(s= 0.5); mannitol, C6H1406 (s=
0.5); isomaltulose, C12H22O11 (s = 0.4); palatinit, C12H22011
(s=0.85); sorbose, C6HI206 (s=0.9); xylitol, C5H1205
(s = 1.0); saccharine, C7H5NO3S (s = 550); and aspartame,
C14H18N20S (s= 172). These were obtained from chemical
supply houses, except Lycasin, isomaltulose, and palatinit, which
were donated by the Lifesavers Company. All except the saccharine and aspartame were prepared as 0.1, 1.0, and 10%
aqueous solutions. Based on the known sweetness of the saccharine and the aspartame, these were diluted in de-ionized
water to give solutions with the sweetness equivalents of 0.1,
1.0, and 10% sucrose. The test solutions were kept refrigerated
and were warmed before use.
Two series of experiments were run. In the first, the plaque
responses to sorbitol, mannitol, or xylitol were compared with
those given by sucrose, fructose, maltose, raffinose, and corn
starch. In the second, a comparison was made with the six
other available sweetening agents.
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Vol. 64 No. 9
SWEETENERS AND
i
m! Li_
PLAQUE pH IN VITRO
1131
Plaque pH in Sweeteners
7.0
6.0
a
0
-5
0.
5.0
4.0
sorbitol mannitol
saccharin aspartame
xylitol
lycasin
AGENT
SWEETENING
Fig. 1 Mean pHs, with standard deviations, of plaque after incubation for 20 min in 0.1, 1.0, or 10% solutions of various sweeteners or, for saccharine
and aspartame, sweetness equivalents of sucrose solutions.
isomalt.
sucrose
palatinit
sorbose
Sucrose & Xylitol Fermentation in Same Plaque
8.0
mmmmm.
" p
1
10% xylitol
)
O
7.01
5% xylitol &
10% sucrose
0
I
Q.
w
aU
<
6.0
-J
a.
5.0
4.0
I r
m
0
r
Ia
I
r
.
on
AI
.·.·.·.1
10% sucrose
20% xylitol &
10% sucrose
°\
L
I
i~
5 10 15 20 0 5 10 15 20 0
TIME,
Fig. 2-Effect
BR
I~~~~
l
a
a
m
minutes
xylitol to sucrose solutions.
plaque pH
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of adding
I
a
a
5 10 15 20 0 5 10 15 20
J Dent Res September 1985
BIBBY & FU
1132
pH in Same Plaque in Saccharine or Sucrose
8.0
I
I
-.~g:9'
1.0%
EQ
0.1%
W
::
10%
EQ
EQ
1.0%
EQ &
1.0% Sucrose
: .:
............
|~~~~~>
0~~~~~~~
0
I
7.O0h
0.
a
13
6.01-
5.01-
4.0
I
0
a
a
a
I
I
i
I
I
'
II
I-
I
I·
I.....
I
I· I1
I
I
-
5 10 15 20 0
TIME,
Fig. 3-Effect on plaque pH of adding saccharine (given as
Results.
The minimum pHs developed in plaque after 20 min of
incubation in 0.1, 1.0, and 10% solutions of different sugars
and sugar alcohols are shown in Table 1. It can be seen that
the pHs given by the 10% solutions of sugars and corn starch
were all less than 4.5, whereas those of the sugar alcohols
were consistently higher. With the exception of xylitol and
saccharine, for which the results were opposite, the lowest pHs
were given by the 10% solutions.
The results of the pH tests using Lycasin, sorbose, isomaltulose, palatinit, saccharine, and aspartame, each tested with
four or more plaque samples, are summarized in Fig. 1. This
TABLE 1
MEANS OF 20-MINUTE pH READINGS GIVEN IN THREE TESTS,
EACH WITH A DIFFERENT PLAQUE, IN SOLUTIONS OF
SUGARS AND SUGAR ALCOHOLS
Solution Concentrations (w/v)
Substrate
0.1 %
1.0 %
10 %
Sucrose
5.42
4.82
4.15
Fructose
5.47
4.39
4.14
Maltose
5.24
4.53
4.22
Raffinose
5.75
5.03
4.30
Sorbitol
6.30
6.17
5.82
Mannitol
5.67
5.54
5.22
6.07
6.13
6.33
Xylitol
Corn Starch (Cooked)
5.55
4.85
4.29
5
I
a
III
-
I-
I
I
i
5 10 15 20
10 15 20 0
minutes
sucrose sweetness
it
0~~~
.M.,"'
5 10 15 20 0
:, ,
equivalents) to sucrose.
shows that metabolism of isomaltulose resulted in a low pH
similar to that given by sucrose. The highest pHs were found
with xylitol, saccharine, and aspartame, followed by sorbose
and Lycasin. Contrary to the responses given with the other
sweeteners, xylitol and saccharine gave higher pH readings as
the concentrations of the test solutions increased. An expanded
series of tests in xylitol using one or more plaque samples
from six different subjects showed that this was a consistent
finding (Table 2). These responses to xylitol were not changed
by three prior incubations in water and saliva designed to eliminate any acidogenic carbohydrate or inherent acid from the
plaque. The addition of up to 20% of xylitol to sucrose solutions did not change the pH given by plaque from that found
with sucrose alone. Typical findings in sequential tests in the
same plaque are shown in Fig. 2. Similarly, additions to 10%
sucrose of up to 10% of isomaltulose, sorbose, or palatinit or
10% sucrose sweetness equivalents of saccharine or aspartame
also failed to interfere with acid production of plaque. Typical
TABLE 2
MEAN pHs OF PLAQUE BEFORE AND AFTER TESTING IN
XYLITOL SOLUTIONS OF DIFFERENT CONCENTRATIONS
pH Before Test
pH After Test
pH of Xylitol
Solutions
N
10
10
pH in Xylitol Solutions (means
+
0.1%
7.27 + 0.35
6.32 + 0.37
1.0 %
7.43 + 0.28
6.43 ± 0.39
10 %
7.50 + 0.22
6.75 + 0.22
5.49
5.34
5.06
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SD)
Vol. 64 No. 9
SWEETENERS AND PLAQUE pH IN VITRO
findings with a single plaque tested in saccharine are given in
Fig. 3.
Discussion.
particularly as they concern the sugar
with published reports, but our findalcohols,
ings of mannitol fermentation, of active acid production from
isomaltulose, and lack of xylitol interference with sugar fermentation are contrary to those of Birkhed (1978), Maki et al.
(1983), and Vadeboncoeur et al. (1983), who ran tests in bacterial cultures or dilute solutions of plaque. We believe that
our results obtained with whole plaque samples give a more
meaningful indication of what is likely to happen on tooth
our results,
are in agreement
Most of
surfaces in vivo.
Our finding that xylitol depressed the pH of plaque can
probably be accepted as a true finding, since it was a consistent
result in our tests and also because Muhlemann et al. (1977)
recorded a similar fall of about one pH unit in their tests on
in vivo plaque. Such slight acid formation in plaque could be
accounted for by the presence of some xylitol-fermenting organisms in plaque (Gallagher and Fussell, 1979). It is more
difficult to explain why, in contrast to what was found with
other sweeteners, the dilute (0.1%) solutions of xylitol and
saccharine gave the greatest depressions of plaque pH and smaller
pH falls in the higher concentrations (Table 2, Figs. 1, 2, &
3). The demonstration in our supplementary tests-that the
minimum pH achieved with the different xylitol concentrations
was not related to the pH of the test solution (Table 2), the
order of testing, or residual acids or fermentable substrates in
the plaques-shows that our findings were unlikely to be the
result of an experimental artifact. This conclusion is made
more probable by the fact that Muhlemann et al. (1975) recorded a lower pH with a 10% than with a 20% xylitol.
To us, the most likely explanation for the reversal of pH
effects shown by xylitol, saccharine, and, to some extent, aspartame is that interactions between those substrates and the
plaque gave rise to basic products of some sort. Such an effect
has been suggested by Mishiro and Kaneko (1977), who showed
that the breakdown of the constituent amino acids of aspartame
neutralized the acid formed by glucose fermentation in saliva.
Over all, our findings show that sweeteners have a greater
acid-forming capacity than is indicated in a number of reports.
A probable reason for this is that most of the earlier tests were
made in cultures of bacteria, saliva, or dilute suspensions of
plaque in which, because of the smaller numbers of bacteria
present, the bacterial challenge would be weaker than that offered by the plaque samples used in our tests. Our plaque
contained many times more bacterial cells and probably a wider
variety of bacterial types and a broader and more powerful
enzymatic potential than in all the other types of test except
1133
those made on plaque in vivo. Since our findings with sweeteners and other agents are in almost entire agreement with
those recorded in telemetric pH tests on in vivo plaque (Bibby
and Krobicka, 1984), we feel that our technically simpler pH
method can be relied upon to give useful information on the
pH responses of in vivo plaque.
REFERENCES
BIBBY, B.G. and KROBICKA, A. (1984): An in vitro Method for
Making Repeated pH Measurements on Human Dental Plaque, J
Dent Res 63:906-909.
BIRKHED, D. (1978): Automatic Titration Method for Determination
of Acid Production from Sugars and Sugar Alcohols in Small Samples of Dental Plaque Material, Caries Res 12:128-136.
DRUCKER, D.B. and VERRAN, J. (1980): Comparative Effects of
the Substance-Sweeteners Glucose, Sorbitol, Sucrose, Xylitol and
Trichlorosucrose, on Lowering of pH by Two Oral Streptococcus
mutans Strains in vitro, Arch Oral Biol 24:965-970.
EDWARDSSON, S.; BIRKHED, D.; and MEJARE, B. (1977): Acid
Production from Lycasin, Maltitol, Sorbitol and Xylitol by Oral
Streptococci and Lactobacilli, Acta Odontol Scand 35:257-263.
FROSTELL, G. (1973): Effect of Mouth Rinses with Sucrose, Glucose, Fructose, Lactose, Sorbitol and Lycasin on the pH of Dental
Plaque, Odont Revy 24:217-266.
GALLAGHER, I.H.C. and FUSSELL, S.J. (1979): Acidogenic Fermentation of Pentose Alcohols by Human Dental Plaque Microorganisms, Arch Oral Biol 24:673-679.
HAYES, M.L. and ROBERTS, K.R. (1978): The Breakdown of Glucose, Xylitol and Other Sugar Alcohols by Human Dental Plaque
Bacteria, Arch Oral Biol 23:445-451.
JENSEN, M.E. (1984): Human Plaque pH Responses to Hydrogenated Starch Hydrolysates, IADR Abst 63:No. 1150.
MAKI, Y.; OHATA, I.; TAKAZOE, Y.; MATSUKOBO, Y.; TAKAESU, V.; TOPITSOGHOU, A.; and FROSTELL, G. (1983):
Acid Production from Isomaltulose, Sucrose, Sorbitol, and Xylitol
in Suspensions of Human Dental Plaque, Caries Res 17:335-339.
MISHIRO, Y. and KANEKO, H. (1977): Effect of a Dipeptide, Aspartame, on Lactic Acid Production in Human Whole Saliva, J
Dent Res 56:1427-1429.
MUHLEMANN, H.R.; SCHMID, R.; NOGUCHI, T.; IMFELD, T.;
and HIRSCH, R.S. (1977): Some Dental Effects of Xylitol Under
Laboratory and in vivo Conditions, Caries Res 11:263-276.
MUHLEMANN, H.R. and BOEVER, J. (1970): Radiotelemetry of
the pH of Interdental Areas Exposed to Various Carbohydrates.
In: Dental Plaque, W.D. McHugh, Ed., Edinburgh: Livingstone,
pp. 179-186.
MUHLEMANN, H.R. and SCHNEIDER, P. (1975): The Effect of
Sorbose on pH of Mixed Saliva and Interproximal Plaque, Helv
Odont Acta 19:76-80.
TAKAZOE, I.; KOZEI, OL; JUNICHI, S.; KAZUMASA, S.; TATSUYA, I.; and YOSHIKAZA, N. (1981): Non-cariogenic Sweeteners, UK Patent No. 2,186,203A.
VADEBONCOEUR, C.; TRAHAN, L.; MOUTON, C.; and MAYRAND, D. (1983): Effect of Xylitol on the Growth and Glycolysis
of Acidogenic Oral Bacteria, J Dent Res 62:882-884.
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