journal of dentistry 35 (2007) 773777

available at www.sciencedirect.com

journal homepage: www.intl.elsevierhealth.com/journals/jden

The potential of deciduous and permanent bovine

enamel as substitute for deciduous and permanent
human enamel: Erosionabrasion experiments
Thomas Attin a,b,*, Florian Wegehaupt a, David Gries b, Annette Wiegand a
a
b

Clinic for Preventive Dentistry, Periodontology and Cariology, University Zurich, Zurich, Switzerland
Department of Operative Dentistry, Preventive Dentistry and Periodontology, University of Gottingen, Gottingen, Germany

article info

abstract

Article history:

Aim of the present study was to compare toothbrushing abrasion of eroded human and

Received 17 April 2007

bovine enamel utilizing a toothpaste slurry.

Received in revised form

The surfaces of each 36 teeth from cattle and calves and from each 36 human wisdom

3 July 2007

teeth and deciduous teeth were polished. Each 12 specimens from the respective tooth type

Accepted 4 July 2007

were used for assessing toothbrushing abrasion only (A), erosion only (E) and the combination of erosion and toothbrushing abrasion (EA). The EA samples were subjected to 20 cycles
comprising a demineralization/remineralization procedure directly followed by tooth-

Keywords:

brushing abrasion (100 strokes, 300 g load, toothpaste slurry: 3 ml artificial saliva mixed

Erosion

with 1 g dentifrice). Demineralization in form of erosion was performed with 1% citric acid

Toothbrushing

(1 min), remineralization with artificial saliva (15 min). Between the cycles, the samples

Abrasion

were stored in artificial saliva. Wear of the treated surfaces with reference to untreated areas

Bovine

was determined profilometrically.

The samples subjected to abrasion only (A) did not show a significantly different wear

Human

between the different kinds of teeth. The comparisons of substance loss between teeth of
different species revealed that hard tissue loss of the human deciduous teeth was significantly lower as compared to calves teeth after both erosion and erosionabrasion. Also,
both erosion only and erosionabrasion caused higher enamel loss in cattles teeth than in
human wisdom teeth.
It is concluded that human eroded enamel offers better resistance against brushing than
bovine enamel.
# 2007 Elsevier Ltd. All rights reserved.

1.

Introduction

Numerous studies have been carried out using bovine dental

enamel as substitute for human enamel when demineralization
and remineralization protocols were investigated and also for
evaluating the bond strength of adhesive systems to dental hard
tissues.18 Human enamel has a slightly lower density, and a
lower Vickers hardness, but shows a slightly higher content of

calcium and phosphorus as compared to bovine enamel.9,10

Although some minor quantitative differences exist with
respect to calcium content, an analogous behaviour of calcium
distribution from the outer to inner levels of human and bovine
enamel was observed.11 Bovine enamel reveals a higher
porosity with bigger crystals than human enamel.12,13 This
might be the reason for the observation, that demineralization
in bovine enamel progresses three times faster than in human

* Corresponding author at: University Dental School, Plattenstr. 11, CH-8032 Zurich, Switzerland.
E-mail address: thomas.attin@zzmk.unizh.ch (T. Attin).
0300-5712/$ see front matter # 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.jdent.2007.07.007

774

journal of dentistry 35 (2007) 773777

enamel.14,15 Moreover, it was shown that surface-abraded,

irradiated bovine enamel has a reduced solubility as compared
to non-abraded, irradiated bovine enamel.16 This feature was
not observed, when testing the influence of surface-abrasion on
demineralization of irradiated human enamel. Nevertheless, it
was proved that human and bovine enamel behaves similarly to
acidic challenges and remineralizing conditions.17 Thus, Mellberg18 estimated bovine enamel as a suitable substitute for
evaluation demineralization of enamel. Due the similar
characteristics mentioned above, bovine enamel is also
frequently used for investigating erosions or erosio-abrasive
effects, which occurs when an acidic erosive challenge of about
pH 23 is followed by a mechanical impact applied on the tooth
surface.4,5,19 A typical erosio-abrasive challenge is represented
by brushing of a tooth surface previously been subjected to an
acidic substrate. The acidic attack leads to a loss of dental hard
tissue leaving back a demineralized, softened surface, which is
highly susceptible to additional tissue loss in case of abrasion.20,21 Additionally it was shown that also the surfaces of
artificial initial caries lesions, created with solutions with pH 5,
show a higher susceptibility than sound enamel.22
It has been suggested that caries progression is faster in
deciduous than in permanent teeth.23,24 One reason for this
observation might be that deciduous enamel has a higher
content of carbonate (type A and B), which might be one
factor contributing to the faster caries progression in
deciduous teeth.25 However, there is conflict in the literature
about the question, whether deciduous enamel behaves
different to an erosive attack as compared to permanent
enamel.2630 In these studies, the erosive demineralization of
these two tooth substrates was compared. However, no
information is given regarding the susceptibility to erosion
abrasion of deciduous enamel as compared to enamel
prepared from permanent teeth. Moreover, it might be
reasonable to perform erosion or erosionabrasion tests in
deciduous teeth of calves as a substitute for deciduous
human teeth.
As known from dental restorative materials, the susceptibility of different materials to abrasion is often not wellcorrelated to the chemical composition or physical properties
of the materials, such as hardness.3133 For example, substance loss of different materials of same hardness due to
abrasion is increased when structural patterns of one material
allow for removal of bigger particles or crystals than another
material.32 In this sense, it is also conceivable that an erosive
attack alters some structural patterns of the dental hard
tissue, such as crystal density or size, leading to a more or less
higher susceptibility of the surface to a subsequent abrasive
challenge. These alterations might be different for deciduous
and permanent enamel as well as for human and bovine
enamel.
As yet, no systematic study has been conducted to evaluate
the suitability of bovine teeth from calves or cattle to serve as a
substitute for deciduous teeth and permanent human teeth
under erosionabrasive influences.
Thus, aim of the present study was to compare the enamel
loss of human permanent and deciduous teeth with the
enamel tissue loss of cattles and calves enamel as induced by
toothbrush abrasion with a toothpaste slurry, erosion and the
combination of erosion and abrasion.

The working hypotheses for the present study were that

bovine enamel shows no difference as compared to human
enamel in enamel loss due to:
(1) erosion,
(2) toothbrush abrasion of a previously eroded surface.

2.

Material and methods

In the study each 36 freshly extracted incisors from both cattle

(23 years of age) and calves (less than 12 month of age) as well
as 36 wisdom teeth and 36 deciduous teeth (incisors and
molars) were used. After extraction, the teeth were stored in
0.5% thymol solution until required. A single cylinder (4 mm in
diameter) was prepared from the central labial aspect of each
tooth by means of a trephine mill (Komet, Lemgo, Germany).
The specimens were embedded in acrylic resin (Palavit G,
Kulzer Wehrheim, Germany) using a steel mould of 25 mm
diameter. The enamel surfaces of the specimens were ground
flat and polished with water-cooled carborundum discs (800,
1000 and 1200, 2400 and 4000 grit; Water Proof Silicon Carbide
Paper, Struers, Erkrath, Germany). Grinding and polishing was
performed with a digitally controlled automatic grinding
device ensuring complete flatness of the prepared surfaces
(Exact Mikroschleifsystem Mikro 40, Exakt Apparatebau,
Norderstedt, Germany). The preparation steps resulted in
removal of around 300 mm of the outermost enamel layer. The
thickness of enamel removed was controlled with a micrometer (Mitutoyo, Tokyo, Japan). Finally, acrylic resin was
applied onto the bottom of the embedded specimens in order
to align the polished surface parallel to the bottom. Before
inclusion in the experiment, the polished surfaces were
checked for exact flatness and deficiencies with a profilometer
(Perthometer Concept, Mahr, Gottingen, Germany). Specimens
with deficiencies, such as grooves or cracks, or with an
unsufficient flatness (more than 0.1 mm difference in height of
later reference surface to center of polished area) were
discarded.
Then, a part of the enamel surfaces was covered with tape
(Tesa, Beiersdorf, Hamburg, Germany) exposing an area of
1.4 mm  3.0 mm in the centre of the enamel specimens. The
covered areas stood as reference surfaces, which were later
not subjected to the de- and remineralizing solutions and
allowed for measuring the substance loss.
Each 12 specimens from the respective tooth type were
later used for assessing abrasion only (A), erosion only (E) and
the combination of erosion and abrasion (EA).
The specimens subjected to erosionabrasion (EA) were
submitted to a cycling treatment comprising de- and remineralization steps as well as toothbrushing. The complete cycle
was run every day during a 20-day period and included the
following steps:
1. Demineralization for 1 min (storage in 1% citric acid with
pH 2.3, 20 ml for each specimen).
2. Rinsing with distilled water
3. Remineralization for 15 min (storage in artificial saliva,
20 ml for each specimen).
4. Toothbrushing abrasion.

journal of dentistry 35 (2007) 773777

775

5. Rinsing with distilled water.

6. Remineralization for the rest of the day (storage in artificial
saliva).
The artificial saliva was mixed according to the formulation
given by Attin et al.34. Toothbrushing was performed in an
automated brushing machine described in detail previously35
applying 100 brushing strokes (100 strokes per minute) with a
load of 300 g. During the 20-day experimental period a total of
2000 brushings strokes were applied an each specimen of
group EA and A. A toothbrush with medium bristle stiffness
(elmex 39, Gaba, Munchenstein, Switzerland) was used for
brushing. The toothbrush has a multi-tufted, flat design with a
total of 1248 filaments. Each filament is 0.22 mm in diameter
with a length of 88 mm. The brushing machine allowed a
linear motion of the toothbrushes on the enamel specimens.
Additionally, the plate, on which the specimens were fixed,
made slow movements (20 per minute) of 2 mm in a direction
perpendicularly to the motion of the toothbrushes. This
feature avoided that the toothbrush run in the identical track
on the enamel samples during the brushing procedures. The
toothpaste slurry used consisted of 3 ml artificial saliva mixed
with 1 g dentifrice (elmex, Gaba) with RDA: 38  9 (according to
Imfeld et al.36).
The samples subjected to erosion only (E) were brushed,
and the samples submitted to abrasion only (A) were not
demineralized. The remaining steps of the cycle were run by
these samples in the same way as described for group EA.
After the total of 20 cycles, the adhesive tape was removed
and the subsatnce loss was registered with a profilometer
(Perthometer concept, Mahr, Gottingen, Germany). The areas
protected with the tape during cycling were used as reference
areas allowing for measuring the depth of the groove resulting
from the chemical and/or mechanical impact. Three traces on
each specimen were recorded and averaged.
The data were proved for normal distribution (KolmogoroffSmirnoff test) and comparisons between the different
groups were conducted with two-sided t-tests. The level of
significance was set at P  0.05 with adjustment of calculated
P-values according to Bonferroni-Holm.

3.

Results

P-values given in the results section refer to the P-values

adjusted according to Bonferroni-Holm. In Fig. 1, hard tissue
loss due to abrasion, erosion and erosionabrasion in
deciduous teeth, calves teeth, wisdom teeth and cattles
teeth is given.
The statistical analysis for the impact of the different
treatments within the same kind of teeth showed that hard
tissue loss induced by abrasion only (A) was significantly
lowest as compared to both the erosion (E) and erosion
abrasion (EA) treatment. The combination of erosion and
abrasion (EA) caused the significantly highest loss of enamel.
The comparisons between the different kinds of teeth
showed that the treatment abrasion only resulted in a nonsignificantly different enamel loss. Also, the comparisons
within the same species showed no significant differences
between calves teeth and cattles teeth for erosion only

Fig. 1 Mean hard tissue loss (mm) and standard deviation

due to abrasion, erosion and erosio-abrasion in deciduous
teeth, calves teeth, wisdom teeth and cattles teeth.
Columns of the same treatment group indicated with the
identical letter were not statistically significantly different.
Each column represents the mean value of n = 12
specimens.

(P = 0.1800), and between deciduous teeth and wisdom teeth

for erosion only (P = 0.4013) and erosionabrasion (P = 0.7706).
However, calves enamel yielded significantly higher abrasion
for erosionabrasion than cattles enamel (P = 0.0126).
The comparisons of substance loss between teeth of
different species revealed that hard tissue loss of the human
deciduous teeth was significantly lower as compared to calves
teeth after both erosion (P = 0.0000) and erosionabrasion
(P = 0.0000). Also, both erosion only (P = 0.0000) and erosion
abrasion (P = 0.0348) caused higher enamel loss in cattles
teeth than in human wisdom teeth.

4.

Discussion

There are few studies in the literature, which have investigated the differences between bovine and human enamel with
respect to susceptibility to demineralization or erosion.14,37,38
Also, few studies have compared the different behaviour of
permanent and deciduous human enamel towards erosive
attacks.2630 The results of the present study indicate that both
erosion and erosionabrasion was higher in bovine enamel
than in human enamel. However, the differences within the
same species, i.e. deciduous human teeth vs. wisdom teeth
and cattles teeth vs. calves deciduous teeth, were more or
less negligible and only significant for the comparison of
erosionabrasion of cattles teeth vs. calves teeth.
In the present study, enamel samples were polished in
order to allow standardized profilometric measurements with
flat reference surfaces. Additionally, existence of the cementum layer on top of bovine enamel renders it necessary to
remove this layer by polishing. It has also to be realized that
polishing of the human deciduous teeth resulted in removal of
the prismless outermost surface layer. Flattening of the

776

journal of dentistry 35 (2007) 773777

enamel surface has an impact on abrasion resistance, as shown

by Slop et al.39 These authors reported that an initially high
abrasion could be recorded during the first 10,000 strokes, so
that in their study a linear increase of enamel loss with
increasing brushing strokes was seen only above 20,000 strokes.
Therefore, the curve representing the relationship between
enamel removal and applied brushing strokes consisted of two
parts: a non-linear (up to about 20,000 strokes with about 0.5 mm
enamel loss) and a linear part (above 20,000 strokes). It was
assumed that this effect was due to the flattening procedure,
which might have already touched the crystallites of the
outermost enamel layers of about 0.4 mm, rendering these
crystallites more susceptible to abrasion. In the present study, a
total of 2000 brushing strokes were applied to the samples. This
means that in the experimental group with abrasion only (group
A) the brushing procedure was limited to the non-linear part of
the curve as presented by Slop et al.39 In the group reflecting the
combination of erosion and abrasion (EA) the aspect of surface
flattening might have been less important, since the erosive
attacks already led to an enamel loss of about at least 2 mm as
shown by the group erosion only (E). This means that in the
EA-group the brushing procedure took place in the linear part of
the curve as presented by Slop et al.39
Human deciduous enamel contained significantly more
carbonate than permanent enamel.25 The carbonate ion
occupies two different positions in the hydroxyapatite
structure, namely the hydroxide position (A) and the phosphate position (B). In both positions, the carbonate ion causes
distortion in the hydroxyapatite structure. Especially the
amount of carbonate in position A is increased in deciduous
enamel, which is assumed to distort the lattice more than in
the phosphate position and is less tightly bound than in the B
position. This feature might contribute to the increase in the
susceptibility of human deciduous enamel as compared to
permanent enamel.25 Also, human deciduous enamel is more
porous than human permanent enamel.40 Thus, the obtained
finding was unexpected that those two substrates behaved
similar under the chosen conditions of acid treatment
(demineralization) and mechanical impact by toothbrushing.
However, previous studies did also show similar changes of
human permanent and bovine teeth in erosion models.26,27 It
should be noticed that outline and arrangement of enamel
rods in human deciduous teeth closely resemble the appearance in permanent teeth.41 This morphological pattern might
explain, why human deciduous teeth behaved similar and
showed same resistance as permanent enamel under all
treatment conditions in the present study, such erosion,
abrasion and the combination of erosion and abrasion.
When bovine teeth are used as substitutes for human teeth,
permanent incisors of 23-year-old cattle are usually used. As
mentioned in the introduction, structural aspects and composition of cattle teeth has been documented and compared to
human permanent teeth. However, information on structure
and composition of calves teeth are not available in the
literature. The present investigation now showed that first
dentition bovine teeth taken from calves behave similar to a
pure erosive attack as compared to permanent bovine teeth,
but different with respect to erosionabrasion.
In conclusion, both erosion and erosionabrasion in bovine
teeth led to higher enamel loss than in human teeth. Thus, the

working hypotheses of the present study have to be rejected.

This fact should be considered when interpreting data from
studies, in which bovine teeth were used as substitutes in
erosion or erosionabrasion models.

references

1. Reis AF, Giannini M, Kavaguchi A, Soares CJ, Line SR.

Comparison of microtensile bond strength to enamel and
dentin of human, bovine, and porcine teeth. Journal of
Adhesive Dentistry 2004;6:11721.
2. Nakamichi I, Iwaku M, Fusayama T. Bovine teeth as possible
substitutes in the adhesion test. Journal of Dental Research
1983;62:107681.
3. Hannig C, Hamkens A, Becker K, Attin R, Attin T. Erosive
effects of different acids on bovine enamel: release of
calcium and phosphate in vitro. Archives of Oral Biology
2005;50:54152.
4. Buzalaf MA, de Moraes Italiani F, Kato MT, Martinhon CC,
Magalhaes AC. Effect of iron on inhibition of acid
demineralisation of bovine dental enamel in vitro. Archives
of Oral Biology 2006;51:8448.
5. Rios D, Honorio HM, Magalhaes AC, Buzalaf MA, Palma-Dibb
RG, Machado MA, da Silva SM. Influence of toothbrushing on
enamel softening and abrasive wear of eroded bovine
enamel: an in situ study. Pesquisa Odontologica Brasileira
2006;20:14854.
6. Attin T, Meyer K, Hellwig E, Buchalla W, Lennon AM. Effect
of mineral supplements to citric acid on enamel erosion.
Archives of Oral Biology 2003;48:7539.
7. Meyer-Lueckel H, Schulte-Monting J, Kielbassa AM. The
effect of commercially available saliva substitutes on
predemineralized bovine dentin in vitro. Oral Diseases
2002;8:1928.
8. Kielbassa AM, Shohadai SP, Schulte-Monting J. Effect of
saliva substitutes on mineral content of demineralized
and sound dental enamel. Supportive Care in Cancer
2001;9:407.
9. Braden M. Heat conduction in normal human teeth. Archives
of Oral Biology 1964;32:47986.
10. Esser M, Tinschert J, Marx R. Materialkennwerte der
Zahnhartsubstanz des Rindes im Vergleich zur humanen
Zahnhartsubstanz. Deutsche Zahnarztliche Zeitschrift
1998;53:7137.
11. Davidson CL, Boom G, Arends J. Calcium distribution in
human and bovine surface enamel. Caries Research
1973;7:34959.
12. Moriwaki Y, Kani T, Kozatani T, Tsutsumi S, Shimode N,
Yamaga R. The crystallinity change of bovine enamel during
maturation. Japanese Journal of Dental Materials 1968;9:7885.
13. Arends J, Jongebloed WL. Crystallites dimensions of enamel.
Journal de Biologie Buccale 1978;6:16171.
14. Featherstone JD, Mellberg JR. Relative rates of progress of
artificial carious lesions in bovine, ovine and human
enamel. Caries Research 1981;15:10914.
15. Flim GJ, Arends J. Diffusion of 45Ca in bovine enamel.
Calcified Tissue Research 1977;24:5964.
16. Kielbassa AM, Hellwig E, Meyer-Lueckel H. Effects of
irradiation on in situ remineralization of human and bovine
enamel demineralized in vitro. Caries Research 2006;40:
1305.
17. Koulourides T, Housch T. Hardness testing and
microradiography of enamel in relation to intraoral de- and
remineralisation. In: Edgar WM, Leach SA, editors.
Demineralisation and remineralisation of the teeth. Oxford: IRL
Press; 1986. p. 25572.

journal of dentistry 35 (2007) 773777

18. Mellberg JR. Hard-tissue substrates for evaluation of

cariogenic and anti-cariogenic activity in situ. Journal of
Dental Research 1992;71:9139. (spec no.).
19. Amaechi BT, Higham SM. In vitro remineralisation of
eroded enamel lesions by saliva. Journal of Dentistry
2001;29:3716.
20. Attin T, Koidl U, Buchalla W, Schaller HG, Kielbassa AM,
Hellwig E. Correlation of microhardness and wear in
differently eroded bovine dental enamel. Archives of Oral
Biology 1997;42:24350.
21. Davis WB, Winter PJ. The effect of abrasion on enamel and
dentine and exposure to dietary acid. British Dental Journal
1980;148:2536.
22. Kielbassa AM, Gillmann L, Zantner C, Meyer-Lueckel H,
Hellwig E, Schulte-Monting J. Profilometric and
microradiographic studies on the effects of toothpaste and
acidic gel abrasivity on sound and demineralized bovine
dental enamel. Caries Research 2005;39:3806.
23. Peyron M, Matsson L, Birkhed D. Progression of approximal
caries in primary molars and the effect of Duraphat
treatment. Scandinavian Journal of Dental Research
1992;100:3148.
24. Shwartz M, Grondahl HG, Pliskin JS, Boffa J. A longitudinal
analysis from bite-wing radiographs of the rate of
progression of approximal carious lesions through human
dental enamel. Archives of Oral Biology 1984;29:52936.
25. Sonju Clasen AB, Ruyter IE. Quantitative determination of
type A and type B carbonate in human deciduous and
permanent enamel by means of Fourier transform
infrared spectrometry. Advances in Dental Research
1997;11:5237.
26. Lussi A, Kohler N, Zero D, Schaffner M, Megert B. A
comparison of the erosive potential of different beverages in
primary and permanent teeth using an in vitro model.
European Journal of Oral Sciences 2000;108:1104.
27. Lippert F, Parker DM, Jandt KD. Susceptibility of deciduous
and permanent enamel to dietary acid-induced erosion
studied with atomic force microscopy nanoindentation.
European Journal of Oral Sciences 2004;112:616.
28. Amaechi BT, Higham SM, Edgar WM. Factors influencing the
development of dental erosion in vitro: enamel type,
temperature and exposure time. Journal of Oral Rehabilitation
1999;26:62430.
29. Hunter ML, West NX, Hughes JA, Newcombe RG, Addy M.
Relative susceptibility of deciduous and permanent dental

30.

31.

32.

33.
34.

35.

36.

37.

38.

39.
40.

41.

777

hard tissues to erosion by a low pH fruit drink in vitro.

Journal of Dentistry 2000;28:26570.
Hunter ML, West NX, Hughes JA, Newcombe RG, Addy M.
Erosion of deciduous and permanent dental hard tissue in
the oral environment. Journal of Dentistry 2000;28:25763.
Attin T, Vataschki M, Hellwig E. Properties of resin-modified
glass-ionomer restorative materials and two polyacidmodified resin composite materials. Quintessence
International 1996;27:2039.
Mair LH, Stolarski TA, Vowles RW, Lloyd CH. Wear:
mechanisms, manifestations and measurement. Report of a
workshop. Journal of Dentistry 1996;24:1418.
Wright KHR. The abrasive wear resistance of human dental
tissues. Wear 1969;14:26384.
Attin T, Buchalla W, Gollner M, Hellwig E. Use of variable
remineralization periods to improve the abrasion resistance
of previously eroded enamel. Caries Research 2000;34:4852.
Wiegand A, Wegehaupt F, Werner C, Attin T. Susceptibility
of acid-softened enamel to mechanical wear
ultrasonication versus toothbrushing abrasion. Caries
Research 2007;41:5660.
Imfeld T, Sener B, Lutz F. Mechanical effects on dentin of the
best selling toothpastes of the Swiss market. Evaluation of
the potential cleaning effect, relative dentin abrasion and
surface roughening. Acta Medicinae Dentium Helvetica
1998;3:549. (in German).
Meurman JH, Frank RM. Progression and surface
ultrastructure of in vitro caused erosive lesions in human
and bovine enamel. Caries Research 1991;25:817.
Rios D, Honorio HM, Magalhaes AC, Delbem AC, Machado
MA, Silva SM, Buzalaf MA. Effect of salivary stimulation on
erosion of human and bovine enamel subjected or not to
subsequent abrasion: an in situ/ex vivo study. Caries
Research 2006;40:21823.
Slop D, de Rooij JF, Arends J. Abrasion of enamel. I. An in
vitro investigation. Caries Research 1983;17:2428.
Linden LA, Bjorkman S, Hattab F. The diffusion in vitro of
fluoride and chlorhexidine in the enamel of human
deciduous and permanent teeth. Archives of Oral Biology
1986;31:337.
Radlanski RJ, Renz H, Willersinn U, Cordis CA, Duschner H.
Outline and arrangement of enamel rods in human
deciduous and permanent enamel. 3D-reconstructions
obtained from CLSM and SEM images based on serial ground
sections. European Journal of Oral Sciences 2001;109:40914.