Title
Author(s)
Citation
Issued Date
URL
Rights
Effects of powered toothbrush cleaning on acrylic resin dental
prostheses
Tan, Chow Ming.; 陳昭明.
Tan, C. M. [陳昭明]. (2012). Effects of powered toothbrush
cleaning on acrylic resin dental prostheses. (Thesis). University
of Hong Kong, Pokfulam, Hong Kong SAR. Retrieved from
http://dx.doi.org/10.5353/th_b4833471.
2012
http://hdl.handle.net/10722/174262
The author retains all proprietary rights, (such as patent rights)
and the right to use in future works.
Effects of Powered Toothbrush Cleaning on Acrylic
Resin Dental Prostheses
by
Tan Chow Ming
Dental Materials Science
Faculty of Dentistry
The University of Hong Kong
A thesis submitted to The University of Hong Kong for the
completion of the Master of Science in Dental Materials Science
degree
1st September 2012
Declaration
I declare that, with the exception of the assistance acknowledged, all the work described
in this report is my own original work and that no part of it has been included in any
report, thesis or dissertation submitted to the University of Hong Kong or any other
institution for degree, diploma or other qualification.
Signed: …………………………………
Tan Chow Ming
Acknowledgements
I am deeply grateful to my supervisors Dr J.P. Matinlinna, Dr James K.H. Tsoi
of Dental Materials Science and Dr C.J. Seneviratne of Oral Biosciences for their
guidance and supervision. I wold like thank Dr Christie Y.K. Lung, Dr Alexander T.H.
Tang and Dr May C.M. Wong for their generosity in sharing their expert knowledge
with me throughout my studies.
I would like to extend many thanks to support staff of Dental Material Science
Department Mr Tony Yuen, Mr Paul Lee and Mr Bentley Yeung and Mr Simon Lee as
well as staff from Oral Biosciences Department for their assistance and help to allow the
smooth running of the experiments. My appreciation to Miss. Sarah Wong for patiently
guided me through the procedure in microbiology experiment and Ms Regina Chan for
her ever ready excellent secretarial assistant offered.
I would like to express my hearth felt appreciation to Professor Christopher
Boey C.M. from Department of Paediatrics, Faculty of Medicine, University of Malaya
for his constant encouragement.
Lastly, I thank my colleagues for the many informal chats and suggestions
offered and my families who put up with me being at my computers and laboratory late
in the evenings and on many weekends.
Contents
Declaration……………………………………………………………………………..…………i
Acknowledgements………………………………………………………………………………ii
Table of Contents……………………………………………………………………………….iii
Abbreviations……………………………………………………………………………………iv
Part I: Literature review
1. Poly(methyl methacrylate) PMMA
1.1 Introduction………………………………………………………………....1
1.2 Chemistry of poly(methyl methacrylate)…………………………………...2
1.3 Properties of poly(methyl methacrylate)
1.3.1 Physical properties
1.3.1.1 Surface of PMMA……………………………………..11
1.3.1.2 Thermal properties…………………..………………...15
1.3.2 Mechanical Properties…………………………………………...17
1.3.2.1 Flexural strength……………………………………….18
1.3.2.2 Fracture toughness……………………………………..22
1.3.2.3 Hardness……………………………………………….23
1.3.3. Biological Properties…………………………………………....24
1.4 Processing of poly(methyl methacrylate)
1.4.1 Heat polymerization……………………………………………..25
1.4.2 Chemical polymerization………………………………………..26
1.4.3 Microwave polymerization……………………………………...28
1.4.4 Light activated polymerization………………………………….30
1.5 Mode of polymerization process and its effects on PMMA
1.5.1 Porosity in PMMA……………………………………………....31
1.5.2 Residual methyl methacrylate (MMA)………………………….33
1.6 Water storage and poly(methyl methacrylate)
1.6.1 Effect of water storage on dimensional stability………………..36
1.6.2 Effect of water storage on surface of PMMA…………………...39
2. Hygiene aspects of prostheses made of poly(methyl methacrylate)
2.1 Biofilm and plaque on dental prostheses………………………….41
2.1.1 Candida albicans biofilm………………………………..42
2.2 Denture hygiene…………………………………………………...45
2.3 Mechanical cleaning method for dental prostheses……………….48
2.3.1 Powered denture brush…………………………………..50
2.3.2 Powered toothbrush……………………………………...51
2.3.3Manual denture brushes and manual toothbrushes………55
2.3.4 Sonic and ultrasonic cleaners............................................56
2.4 Chemical cleaning………..………………………………………..58
2.5 Microwave disinfection…………………………………………....63
3. Conclusion…………………………………………………………………………..66
References.…………………………………………………………………………….67
Part II: Laboratory Report
Abstract………………………………………………………………………………..84
1. Introduction………………………………………………………………………....86
2. Materials and methods
2.1 Specimen grouping………………………………………………………...93
2.2 Materials…………………………………………………………………...94
2.3 Specimens fabrication……………………………………………………..95
2.4 Polymerization and post processing……………………………………….99
2.5 Storage condition........................................................................................100
2.6 Powered toothbrush brushing protocol of specimens………………….....101
2.7 Surface roughness test................................................................................105
2.8 Flexural strength test..................................................................................106
2.9 Statistical analysis......................................................................................108
2.10 Microbiology
2.10.1 Materials and methods..............................................................109
2.10.2 Specimen grouping...................................................................111
2.10.3 Candida albicans cultured biofilm growth on specimens........112
2.10.4 Brushing protocol for Candida albicans coated specimens.....114
2.10.5 SEM of Candida albicans coated specimens...........................117
3. Results
3.1 Surface roughness......................................................................................118
3.2 Flexural strength........................................................................................123
3.3 SEM...........................................................................................................128
4. Discussion
4.1 Surface roughness......................................................................................135
4.2 Flexural strength........................................................................................137
4.3 SEM...........................................................................................................139
5. Conclusion...............................................................................................................142
Appendix.....................................................................................................................143
References...................................................................................................................161
Abbreviations
PMMA: poly(methyl methacrylate)
MMA: methyl methacrylate
AFM: Atomic force microscopy
SEM: Scanning electron microscope.
EPS: Extracellular polymetric substance
SFE: Surface free energy
Ra: arithmetic average
Tg: Glass transition temperature
vol: volume
ml: millilitres
wt: weight
kg: kilogram
g: grams
µg: microgram
mm: millimetre
μm: micrometer
h: hour
min: minutes
s: second
M: Moles
PBS: Phosphate buffered saline
YNB: yeast nitrogen base
SAB: Sabouraud's dextrose
psi: pounds per square inch
MPa: megapascal
rpm: revolutions per minute
CAD/CAM: Computer Aided Design/Computer Aided Manufacture
°C: degree Celsius
W: watts
Part I: Literature Review
1. Poly(methyl methacrylate)
1.1 Introduction
Poly(methyl methacrylate) (PMMA) was developed 75 years ago in 1937,
and is still the major material for fabrication of denture bases due to its aesthetic
characteristics, high processing and polishing abilities, relining and rebasing
possibility and low cost [1]. The main disadvantages of PMMA are its
dimensional changes during polymerization, porosity and allergic or cytotoxic
effects from leached and degraded monomers [2].
Prior to the introduction of PMMA as denture base material, vulcanite
(vulcanized rubber) was material of choice in 1855 [3]. Due to its poor aesthetic
qualities and taste it was replaced. Harold M. Vernon and Lester B. Vernon
collaborated with the research and development team of Rohm and Haas
Company in the USA and eventually formulated PMMA which is suitable for the
use in dentistry and replaced vulcanite [1, 3]. PMMA popularity in use as a
material of choice for the construction of dental prostheses continues even until
today. The application of PMMA in the fields of medical and dental science has
been expanding ever since its inception. For example, in ophthalmology, patients
suffer from cataract would need the intraocular lens replacement with an artificial
lens that is usually made from PMMA [4]; in orthopaedic, it is used as bone
cement to anchor cemented arthroplasties to the contiguous bone [5].
1
1.2 Chemistry of poly(methyl methacrylate)
PMMA polymers are derivatives of ethylene containing a vinyl (─C═C─)
group in their structural formula H₂C═CHR,
where R is an organic functional group. Methacrylic acid, the precursor of
methacrylate with the formula of:
CH3
H2C
O
O
OH
Methacrylic acid
is a polar molecule due to the present of carboxyl group. This polarity absorbs
water molecules, which will tend to separate the polymer chains and cause
softening and loss of strength of the polymer. The monomer of PMMA, methyl
methacrylate (MMA), is the ester of methacrylic acid:
CH3
H2C
O
O
Methyl methacrylate
CH3
PMMA is formed through a free radical addition polymerization process
of MMA monomer. The monomers are added to the living end (with radical) of a
growing chain. No change is made to the composition during the addition
polymerization reaction; the monomer and polymer have the same empirical
formulas. The polymerization of monomeric methyl methacrylate to form polymer
of PMMA involves an exothermic reaction.An amount of heat, equivalent to 80
kJ/Mol, is released in reducing the C═C double bond to C─ C single bond.
2
The sequence of conversion from monomer to polymer involves activation,
initiation, propagation, chain transfer and termination. The following is a
description of the process.
The polymerization of polymer PMMA is an addition polymerization
process that requires a source of free radicals and the presence of an unsaturated
group i.e. a double bond Free radicals are generated through activation of radicalproducing molecules by using another chemical, heat, visible light, ultraviolet
light or energy transfer from another compound that acts as a free radical ( R• ). A
free radical is an atom or group of atoms possessing an unpaired electron (symbol
•). The process of producing free radical is called activation, for example, the
decomposition of benzoyl peroxide (Fig. 1.1). The free radical has the ability to
withdraw electron from high electron density double bond in monomer due to the
present of unpaired electron. When the free radical approaches a double bond, an
electron is extracted, and it pairs with the unpaired electron in R• to form a bond
between the radical and the monomer molecule, leaving the other electron of the
double bond or central carbon unpaired. The original free radical bonds to one end
of the monomer molecule and a new free radical site forms at the other end.
O
O
O
O
2
O
O
benzoyl peroxide
free radical
Fig. 1.1 Activation of benzoyl peroxide to generate free radicals ( • )
3
Benzoyl peroxide is the most often used initiator which is activated at
temperature range of 50 ° C to 100 ° C to form two free radicals. These free
radicals react with MMA molecule to initiate polymer chain growth (Fig. 1.2):
O
CH3
O
+
Initiation
H2C
H
O
C
O
O
CH3
C
H
O
O
O
CH3
CH3
Initiation of methyl methacrylate
O
H
O
H
CH3
O
C
CH3
C
O
O
H
C
O
H
O
CH3
O
CH3
Fig. 1.2 Initiation of methyl methacrylate molecules. An unpaired electron of a
free radical attracted to one of the electron in the double bond to form an electron
pair and a covalent bond between the free radical and the monomer molecule.
The remaining unpaired electron makes the new molecule a free radical.
In the propagation stage of the polymerization reaction, the resulting free
radical-monomer complex acts as a new free radical centre when it approaches
4
another monomer to become a free radical. Polymerization continues through the
propagation of the reactive centre, successively adding reactive species to a large
number of methacrylate molecules (Fig. 1.3). In theory, the chain reactions are
supposed to continue until all the monomers have been converted to a polymer.
However, in reality, the polymerization reaction is never totally completed due to
the reaction has passed exotherm and the reaction of polymerization reaches an
equilibrium state [6]. The polymer chain stops to grow any longer when the
reactive centre is destroyed by the termination reactions.
O
CH3
CH3
C
H2C
+
O
O
O
O
O
CH3
CH3
Propagation
CH3
H2C
O
CH3
CH3
O
O
C
O
O
O
O
O
CH3
+
CH3
Chain growth
Repeating
process.
CH3
Fig. 1.3 Propagation and chain growth. When the molecule with a free radical
approaches other methyl methacrylate molecules, the free electron interacts with
the double bond of the methyl methacrylate molecule forming a longer free
radical.
5
During the stage of chain transfer, the active free radical is transferred to a
monomer or inactivated polymer chain and a new free radical for growth is
created (Fig. 1.4).
CH3
CH3
O
CH3
C
O
H2C
+
O
O
O
O
O
CH3
CH3
CH3
O
chain transfer
CH3
O
CH3
CH3
O
O
O
CH3
H3C
C
O
O
CH3
O
+
O
CH3
Fig. 1.4 Chain transfer occurs when a free radical donates a hydrogen atom to
the methyl methacrylate molecule. Resulted in the free radical rearrange to form
a double bond and turns nonreactive, and the MMA monomer becomes a free
radical.
6
The final stage of polymerization reaction is the termination which can
result from chain transfer. Addition polymerization reactions are often terminated
by direct coupling of two free radical chain ends or by the exchange of a hydrogen
atom from one growing chain to another ( i.e. hydrogen abstraction). In direct
coupling, both molecules combine to become deactivated by the formation of a
covalent bond (Fig. 1.5). The exchange of a hydrogen atom from one growing
chain to another growing chain produces a double bond (Fig. 1.6).
O
H
O
CH3
H
H
O
CH3
H H3C
H3C
C
H
C
+
O
O
O
O
CH3
CH3
H
O
O
O
O
H
O
H
H3C
H3C
Termination
O
CH3
H3C
CH3
O
CH3
O
O
O
O
O
O
CH3
CH3
O
O
O
O
CH3
CH3
Fig 1.5 Termination of polymerization occurs when free radicals react to form
stable molecule as well as a transfer process.
7
O
CH3
H
O H
H
CH3
C
H
O
O
O
O
CH3
CH3
H
O
O
O
O
H
C
+
O
O
H
H H3C
H3C
H3C
H3C
Termination after chain transfer
H3C
O
CH3
CH3
O
O
O
CH3
O
O
O
O
O
CH3
CH3
+
O
H
O
O
CH3
CH3
Fig 1.6 The transfer of a hydrogen atom from a growing chain to another chain
forming a double bond.
Addition polymerization reactions are unlikely to completely use up all
monomers; in addition, polymers of high molecular weight are not always formed.
The reason for this is that the presence of inhibitors in the form of impurities
retard the polymerization reaction or react with activated initiator. Inhibitors affect
the storage stability and the working time of the acrylic resin. Commercial dental
acrylic resins usually contain a trace amount of inhibitor such as the methyl ether
of hydroquinone to prevent spontaneous polymerization during storage and the
control of working time in the case of a cold-cured acrylic resin.
In the three dimensional molecular structures of polymers, conformation
relates to the internal movement of molecules and configuration relates to the way
in which a chain is built at any asymmetric carbon. Tacticity refers to the ordering
8
of the configuration sequence of a polymer [6]. MMA monomer polymerized in
system without complex steric constraints produces atactic polymer. This means
the substituents in the polymer chain are randomly arranged along the chain
without any regularity. In a polymer where the relative positions of the substituent
are always the same along the length of the chain, the polymer is said to be
isotactic. In the situation where the substituents are in an alternating configuration
it is called syndiotactic. Atactic polymers cannot crystallize; they are amorphous
and less brittle than crystallize materials.
The constituent of the liquid consists of methacrylate based monomers,
such as methyl methacrylate (MMA), hexamethylene glycol dimethacrylate
(HDMA), hydroxyl ethyl methacrylate (HEMA), or n-butyl methacrylate, along
with
a
cross-linking
agent
(e.g.
ethylene
glycol
dimethacrylate,
trimethylolpropane trimethacrylate, or 1,6-hexanediol dimethacrylate) [7].
However, the liquid monomer of chemically polymerized PMMA has an
additional component of chemically activated accelerator, viz. N, N-dimethyl ptoluidine, which is a tertiary amine to activate the radical formation. Differences
in the composition and structure of the component may influence the physical and
mechanical properties of a PMMA polymer, such as tensile strength, transverse
bend strength, water sorption and solubility.
The MMA has a boiling point of 100.8 °C at normal atmospheric pressure
and its melting point is -48 °C. This means it is highly volatile and also highly
flammable. Extra precautionary measures are necessary when handling a liquid of
such nature. The liquid is kept in a dark glass bottle to extend the shelf life of the
monomer by preventing spontaneous polymerization from the action of light. The
liquid monomer has a density of 0.945 g/mL at 20 °C temperature and the heat of
9
polymerization is 12.9 kcal/mol [8]. Traces of inhibitors such as hydroquinone are
added to extend the shelf life of monomer during storage. The use of MMA as an
organic solvent is excellent, however, it has proven to be highly toxic [9].
PMMA is a clear and transparent resinous thermoplastic. Thermoplastic
resin refer to PMMA can be soften and shaped by heating it above its glass
transition temperature (Tg). The Tg of PMMA is about 72 °C [6]. It is stable to
heat with a softening temperature at 125 °C. Heat decomposition of PMMA can
start to occur at temperature range of 125 ° C and 200 ˚ C, and about 90 % of
polymer decompose
. PMMA has a
Brinell hardness number of 25 to 28 and it may be considered relatively hard.
PMMA is extremely stable. It does not discolour in ultraviolet light and it has
remarkable aging properties. Its tensile strength is approximately 59 MPa and its
specific gravity is 1.19 g/cm³ [8].
10
1.3 Properties of poly(methyl methacrylate)
1.3.1 Physical properties
1.3.1.1 Surface of PMMA
The term surface commonly refers to a boundary define by the outer
atomic layer that separate a bulk solid from an adjacent phase. However, another
approach is to consider the surface as a region of variable depth and having a
degree of flexibility that depends on the nature of the material [10, 11]. Material
surfaces are a special state of matter with unique chemistry, organization,
dynamics and electrical properties. The surface molecules have the direct access
for reaction with adjacent phases and the tendency for surface energy
minimization to reach equilibrium with environment. All processes for breakdown
of materials are initiated at the surfaces, provided bonds within the bulk material
are not disrupted.
The polar or non-polar nature, the hydrogen-bonding capacity, and the
electron donor or acceptor potential seem to have a controlling influence on the
hydrophilic or hydrophobic character and energetic state of the surfaces of
materials. The surfaces of PMMA are hydrophilic and exhibit a tendency to
absorb water by a process of imbibition sorption that is due to the polar nature of
the resin molecules.
A positive correlation exists between the recolonization rate of bacteria
and the substratum surface free energy (SFE). On surfaces with a low surface free
energy the rate of bacterial recolonization is slower and the strength of adhesion
on these surfaces is weaker.
Low surface free energy bacteria adhere
preferentially to low surface free energy surfaces (e.g. poly(tetrafluoroethylene) or
11
Teflon), and bacteria with high surface free energy prefer high surface free energy
substrata (e.g. enamel). The rate of bacterial colonization of intraoral hard
surfaces is positively correlated with surface roughness (SR). Surface roughness
exhibits more influence on plaque accumulation and plaque composition compare
to surface free energy [12]. The surface free energy of heat polymerized PMMA is
36.5 ergs/cm2, compared to Teflon which is 24.0 ergs/cm2.
Surface roughness is a 2-dimensional parameter of a material surface
measured by roughness measuring system such as the so-called Stylus system and
is described as arithmetic average roughness (Ra). It is the arithmetic mean of the
departures of profile from the mean line. It is a distance measurement between the
peak and valley part of a material surface and does not represent the
morphological configuration of the surface. The morphology of surface describe
the pattern of a material surface such as a porous surface, grid-like surface and
texture surface, it is a 3-dimensional parameter. The evaluation of surface
morphology is usually carried out under scanning electron microscope (SEM) and
atomic force microscopy (AFM) examination [13].
Quantitative study of a surface roughness can be analysed with a surface
roughness profilometer with a diamond stylus. The profilometer will give a
reading of Ra which is the arithmetic average of the absolute values of the
measure profile height of surface irregularities, measured from a mean line within
a preset length of specimen. The output measurement in unit µm of can be
obtained when the stylus needle passing across a length of 0.8 mm at 1 mm/s.
Another output reading from profilometer is root-mean-square roughness (Rrms).
It has been known that bacteria react to surface topographical features
which are larger than the bacterial cells. Bacteria have shown preferential
12
adherence to the bottom of the crevices rather than the top [14]. A general
perception regards that rough surfaces colonize more rapidly than smooth surfaces
due to more surface area available for attachment and that the greatest initial
accumulation is in the bottoms of roughness elements because of protection from
shear. However, it is still uncertain whether this preferential adherence at bottom
of pits, in microscopic and nanoscopic scale, is a function of preferential
attachment of microorganism or the end result of ineffective cleaning [15].
There are controversies as to the bacterial response to micro-scale surface
features. Some studies showed that the lowest Ra value resulted in the most
hygienic surfaces [16]. One study [17] recommended searching for optimal
surface smoothness for all intraoral hard surfaces for the reduction of bacteria
colonization and plaque formation. This was based on the investigation on
standard implant abutment and surface roughened titanium implant abutment in
patient subjects, which demonstrated that rough surfaces harboured 25 time more
bacteria accumulation compared to smooth surface [17]. However other
investigators showed that surface roughness had no effect on the number of
bacteria adhered [18].
Rough surfaces can be formed through the original surfaces imprint of
mould used or created through frictional abrasion using some abrasive. In the
latter case, frictional work is defined as the loss of energy that leads to
deformation and dissipation within the contacting element and under some
circumstances mechanical or chemical damage of the interface zone. This has
resulted in significant chemical, morphological, mechanical and topographical
modification of the interfacial zone. Heat and debris are always produced in this
interaction at the interfacial zone.
13
In the case of brushing a PMMA based dental prosthesis using a powered
toothbrush, the spinning of brush head carrying the bristles can cause kinetic
energy transformation where the polymer molecules on interface zone can
experience bond stretching from heat energy dissipated and may result in plastic
strain. Once sufficient amount of plastic strain has accumulated, this will result in
the appearance of surface craze and ultimately the cumulative effect of
catastrophic fracture of the bulk material under service conditions.
The studies of materials revealed that the rougher the surface, the higher
the surface free energy count. The more difficult it will be for the denture wearers
to thoroughly clean and maintain their prostheses devoid of biofilm and plaque on
the denture. The rough surface finish on the fitting surface of denture promotes
the retention of Candida albicans [19, 20].
Surface roughness influences the adhesion of micro-organisms to enamel
surfaces [21] and tooth surfaces [22], probably because of the greater surface
area provided and the provision of protected sites for colonization. In addition,
rough surface may aid mechanical retention of microorganism during the early
plaque formation in the depth of surface irregularities found on enamel [23]. The
valleys on the rough surface provided microorganism shield from shearing force
therefore promoting the growth and survival of microorganism in plaque and
biofilm on the dental prostheses.
When Candida albicans was incubated on the surfaces of acrylic resin
specimens, results showed more microorganisms on the rougher surfaces than on
the smoother surfaces. Pre-coating denture base materials with saliva reduced
Candida albicans adhesion on PMMA specimens [24].
14
1.3.1.2 Thermal properties.
Glass transition temperature of PMMA exhibits a variation of
which was observed. A characteristic for all polymeric materials is the glass
transition temperature (Tg), which is the temperature at which a transition to a
softer and more flexible material occurs. The glass transition temperature is
related to the degree of polymerization [25]. The interatomic bonds that hold the
different polymer chains together contain valence electrons that continuously
move back and forth. These movements of electrons cause a varying electron
density that exists along the chain at different times and location. A situation of
disequilibrium is created and the adjacent chains adapt their electron densities
along the chain to balance these unbalanced charge densities. These interactions
cause the development of interatomic induction force which is also known as van
der Waals (or London forces) among the chains. These forces together with
hydrogen bonding can form polar bonds between the polymer chains. These polar
bonds are weaker than the primary bond along the polymer chains. The weaker
polar bonds are broken when the polymer is heated to its Tg or higher temperature
and this will allow the polymer molecular chains to move more freely relative to
each other. The increased mobility can impact on the physical properties which
include strength, modulus of elasticity and thermal expansion. At or near the Tg
temperature the strength and elastic modulus of PMMA decrease, but the thermal
expansion increases.
Thermal conductivity of PMMA is around 6 X 10-4
•g-1•cm-2 which is
considered low. The significance of this is that when heat is generated during
polymerization; it can be difficult for this heat to dissipate away. As a result,
temperature within the bulk material can reach above the boiling point and the
15
quality of final denture could be compromised. In the mouth, the insulating effect
can lead to under stimulation of the mucosa cover by the denture plate possibly
promoting the occurrence of denture stomatitis.
Coefficient of thermal expansion for PMMA is high at approximately 80
ppm/°C. The linear coefficient of thermal expansion for PMMA is 76 x 10-6 /
g
-
. Thermal diffusivity of PMMA is 0.124 mm2/sec.
Colour and appearance are of considerable importance to dental prostheses
end user, especially at the delivery of final finished prostheses. Every patient
wants to look as naturally and aesthetically pleasing as possible with their dental
prostheses in use. The ability to manipulate colour of PMMA polymeric material
is a significant quality well suited for such aesthetic demand in dentistry.
Colouring pigments and veins can be incorporated in the powder of PMMA, in
order to make the finished product well accepted by clinician and end user. On the
other hand, it may be made clear and exceptionally transparent for ophthalmology
purpose.
16
1.3.2 Mechanical properties.
“Mechanical properties” is a description expressing the response of a
material to externally applied force in a scale-independent manner [6]. Strength of
a material or a design of dental prosthesis generally refers to the mechanical
property that ensures the prosthesis serves its intended function effectively, safely
and for a reasonable time period. Effectively speaking, the strength is the ability
of the prosthesis to resist induced stress without fracture or permanent
deformation. Plastic deformation occurs when the elastic stress limit within the
material is exceeded. Deformation may be observed as elastic or reversible when
the force applied is within range of proportional limit of a material.
Stress and strain are often used as the units of expression about the
mechanical properties of a material. Stress is defined as force per unit area acting
on a given plane of material. The types of stress in a functional setting for
prosthesis include tensile stress, shear stress and compressive stress [8]. The
average level of stress at which a material exhibits initial plastic deformation or at
which fracture occurs in test specimen of the same shape and size can be defined
as the strength of a material [8]. Factors that influence strength include strain rate,
the shape of the test specimen, the surface finish which denotes surface flaws and
the testing environment. Strain is defined as the change in length per unit original
length [6]. It is the relative deformation of an object subjected to a stress. In
particular, the tensile strength of PMMA is typically 50 MPa or less. The elastic
modulus is low; the flexural modulus is within the range of 2200-2500 MPa [26].
Dental PMMA resins are visco-elastic materials because these materials
act as rubbery solids that recover elastic deformation once the stresses induced are
17
removed. When PMMA resins are subjected to a sustained load, the materials
exhibit an initial deformation, and additional plastic deformation may occur over
time under continuous load. The additional deformation is called creep.
1.3.2.1 Flexural strength.
The transverse strength or also known as flexural strength of a material, is
a measure of stiffness and resistance to fracture.
Flexural strength provides an indication of the material performance under
static loading. This property would only be directly correlated for PMMA in
which the elastic range extended to the point of fracture and the viscoelastic
properties were not strain rate dependant. Flexural strength can be measured and
calculated through performing a three-point bending test (Fig. 1.7).
Fig.1.7 Schematic of a 3-point bending test. The three points are the two supports
at the bottom and the central loading point on the top.
18
Impact strength is another property include in the evaluation of material.
Robinson J.G. et al. [27] conducted an investigation into PMMA on these two
properties by creating surface defects on specimens. They suggested that the use
of small surface defects and high test rate which was 100mm min -¹ crosshead
speed, would give useful information on the behaviour of denture base materials
under stress. Nevertheless, the outcome of this investigation appeared to suggest
that a flexural test alone was not able to fully characterize the mechanical
properties of PMMA. This is comprehensible because PMMA is a type of
biomaterial that is unique in many of its characteristics. For example, recently it is
used in CAD-CAM milling to produce longer term intermediate implant
supported prostheses. Thus, there are still potential characteristics to be
discovered or to be expanded in its use. Currently, the International Standard
Organization (ISO 1567) (1988) and the British Standard specification 1989 (BS
2487) for denture base resins still use the three-point bending test and have
specified transverse deformation limits which are from 1 to 2.5 mm for a force of
15–35 N and 2–5 mm for a force of 15–50 N. The minimum average breaking
force of acrylic resin is 55 N.
PMMA has relatively weak flexural strength, with a typical value of
flexural modulus of 2200 – 2500 MPa [26]. Attempts were made to strengthen it
by reinforcing PMMA with incorporation of fibres, with some degree of success.
The reinforcement resulted in a tougher material with high impact strength and
improved fracture resistance. The incorporation of various types of fibres or beads,
such as carbon [28, 29], polyethylene [30, 31] , glass [32, 33] , E-glass [34],
aramid [35, 36] and poly(methyl methacrylate) [37, 38] into acrylic resin has
been an attempt to improve its mechanical properties. These fibres are available in
19
various blends, e.g. continuous, woven and chopped. Metal inserts in the form of
wires, meshes and plates have been incorporated into dentures in an attempt to
reinforce areas potentially vulnerable to fracture [39, 40] .
Flexural strength tests can reflect the loading characteristics of a denture
base in a clinical situation [41]. Studies by Regli et al. and Swoope et al. [42, 43]
showed that dentures in service undergo only small deformations and Ladizesky
et al. [44] reported that flexural modulus should be measured at similar small
deformations. The calculated flexural modulus with small deflection may be
regarded as Young's (elastic) modulus of the material.
Deflection of specimen during the flexural test is dependent on the
thickness and type of materials [45, 46]. When comparing the actual flexure
strength of various materials these factors need to be taken into account.
Three-point bending test can be performed with a universal testing
machine to measure the maximum load at fracture of a test material, expressed in
the unit MPa. The device consists of a loading wedge and a pair of adjustable
supporting wedges placed apart from each other at specified distance. The
specimens may be cantered on the device where the loading wedge is set to travel
at a crosshead speed of 1 mm/min or at a pre-determined rate, engaged the centre
of the upper surface of specimens. Specimens are loaded until fracture occurred.
20
Flexural strength in MPa is calculated using the following formula:
where 𝑭 is the maximum load (N) exerted on the specimen, 𝓵 is the distance (mm)
between the supports, 𝒃 is the width (mm) of the specimen measured, and 𝒉 is the
height (mm) of the specimen measured.
Flexural tests can also be conducted through a four-point bending test
using the same type of specimens: two loading noses are used at equal spacing
from their adjacent support points, with a distance between the load points of onethird or one half of the support span [47]. The major difference between fourpoint and three-point bending test is the location of the maximum bending
moment and the maximum axial fiber stress. The maximum axial fiber stress in
four-point bending test is uniformly distributed between the loading noses,
whereas, the maximum axial fiber stress in three-point bending is located
immediately under the loading nose [48].
Comparing the three-point and four-point bending tests in measuring the
flexural strength and modulus of denture base polymers, Chitchumnong et al.
showed that the three-point test always gave higher values [45]. This suggested
that the test method had an influence on the parameter flexural strength. In the
four-point bending test, the two loading elements apply a more uniform load to
the beam that prevents V-shaped buckling of the beam, and avoids stress
concentrations in the midline when a single loading element is used.
21
1.3.2.2 Fracture toughness.
The fracture toughness of a material is a measure of the ability of the
material to resist propagation of a preformed crack. A fracture on materials always
occurs by initiation and propagation of a crack. The inherent resistance against
cracks propagation can be measured by introducing a crack of predetermined size
and shape notch and then measuring the stress required for this crack to grow. A
calculation made on a parameter known as K1c of the test polymeric material.
Although determination of fracture toughness is sensitive to the specimen
geometry and loading conditions, it provides two mechanical parameters namely
“fracture toughness” which relates to the sensitivity of crack initiation, and “work
of fracture” which relates to the resistance to crack propagation [12].
The Izod impact test is a popular method to evaluate the fracture resistance
of denture base. Results of impact tests have often been used to compare flexural
properties with fracture toughness. However, it was found that there are poor
correlations between the impact test and flexural strength or flexural modulus
results [41, 46]. Zappini et al. demonstrated that results of impact tests showed
only moderate correlation with fracture toughness [12]. They suggested that
fracture toughness test would be more reliable than impact tests to determine
mechanical properties of acrylic denture resins.
22
1.3.2.3 Hardness
The hardness test has been considered as a simple and useful method to
assess mechanical properties of polymeric materials because it is very sensitive to
monomer content of dental polymers [49, 50]. Hardness has been known to
correlate with mechanical properties such as tensile strength for many metals [51].
For PMMA base acrylics, Vickers hardness measurement has been used to
investigate changes in the surface physical properties after long-term immersion
in water or in a disinfection solution [52-54]. The surface hardness was positively
correlating with the flexural parameters and negatively correlating with fracture
toughness [51].
23
1.3.3. Biological properties
Biocompatibility is defined as the ability of a material to perform with an
appropriate host response in a specific application [55]. PMMA based resins are
widely used resins in dentistry, especially in fabrication of dentures and
orthodontic appliances. Other uses include bone cements, acrylic glass, and
artificial fingernails and nail varnish. Literature regarding the biocompatibility of
PMMA resins mainly consists of in vitro studies. Various factors in the mouth
such as saliva characteristics, chewing, thermal, and chemical dietary changes
may influence the biological behaviour of these materials.
PMMA is considered to be biocompatible material, and it has been
rigorously trialled and tested in the field of biomaterial. However, allergy may
occur due to the leachable components mainly attributed to the monomers and
benzoic acid [56]. Most allergic reactions are associated with cold-cured acrylic
resin due to the presence of high residual monomer or benzoic acid [57].
Investigation and experiments were carried out to minimize the present of residual
methyl methacrylate by subjecting the dental prosthesis to additional curing cycles
but this can cause dimensional distortion as the internal processing stress are
released. Lung et al. [58] investigated the curing conditions and the concentration
of residual monomer, and found that the extended time at high temperature
employed for polymerization would decrease the residual monomer concentration.
This extended time of cure can compensate for lower processing temperature than
100 °C which is recommended by manufacturers [59, 60] .
24
1.4 Processing of poly(methyl methacrylate).
1.4.1 Heat polymerization
Heat-polymerized PMMA for dental use is supplied in the form of powder
polymer and liquid monomer components. The advantages of powder-liquid
formulations are the ability to process by the dough technique and the
polymerization shrinkage can also be minimized. The beads or granules in powder
are already polymerized. Another noteworthy point for powder-liquid formulation
is that the heat from polymerization reaction is reduced because a large proportion
of mixture is in the polymeric form, thus preventing the potential overheating.
The constituents of the power-liquid formulation are depicted in the Table 1.
Table 1 Constituent of heat polymerized PMMA. Adapted from [26].
Power
Liquid
1. Beads or granules of PMMA.
1. Methyl methacrylate monomer.
2. Benzoyl peroxide as initiator.
2. Hydroquinone as inhibitor.
3. Pigment and dye for colouring.
3. Ethylene glycol dimethacrylate as
4. Titanium/zinc oxides as opacifiers.
cross-linking agent.
5. Dibutyl phthalate as plasticiser.
6. Nylon or acrylic, or other synthetic
fibers.
The heating process used to control polymerization is called the
polymerization cycle or curing cycle. A well-controlled polymerization cycle can
prevent the effect of uncontrolled temperature rise, such as boiling of monomers
25
or the material porosity. One technique involves processing the polymer resin at a
constant temperature in a bath at 74 ° C for 8 h without terminal boiling. A second
technique is processing at 74 ° C in a water bath for 8 h and then increasing the
temperature up to 100° C for 1 hour. A third technique involves processing the
resin at 74 ° C for approximately 2 h and then increasing the temperature of the
water bath to 100 ° C and processing for an additional 1 hour. In literature there
were many combinations of processing cycle studies and proposals. The purpose
of such extensive investigations was aimed at minimising the presence of residual
methyl methacrylate monomer [58].
The denture flask should be cooled slowly to room temperature after the
completion of the polymerization cycle. Rapid cooling may result in warping due
to the excessive internal stress created because of difference in thermal
contraction of resin and investment medium.
1.4.2 Chemical polymerization
Many terms are used to assign the same process, these include chemicalcured, cold-cured, self-cured and auto-polymerized. The chemical reaction of the
chemical cure is the same as heat polymerization except that the polymerization is
initiated by a tertiary amine instead of using heat.
Chemically polymerized PMMA materials involve a chemical activator
such as N,N-dimethyl-p-toluidine and N,N-dihydroxyethyl-p-toluidine [61].
Benzoyl peroxide is the initiator that generates free radicals. An accelerator, such
as a tertiary amine, sulfinic acid, or substituted barbituric acid is required in
chemically polymerized process. The most important combination is an amine–
26
peroxide redox system. The amine forms a complex with benzoyl peroxide to
reduce the thermal energy required to break the initiator into free radical at room
temperature. Polymerization process continues the same as heat-cured PMMA
involving four stages: activation, initiation, propagation and termination.
The chemical polymerization method is less efficient than the heat
polymerization. The amount of uncured residual MMA in post chemical-cured
PMMA is high compared to other methods of polymerization [62]. As a result,
PMMA produced through chemical polymerization has a lower molecular weight
and lower strength properties as well as low colour stability when compared to
heat polymerized PMMA. The lower molecular weight might relate to the
lowering of glass transition temperature (Tg) 70-80 °C. However, this fact remains
debatable [63]. The implication for this is a material with a lower glass transition
temperature which could be particularly prone to distortion when exposed to high
temperatures, which may happen when hot water is used for cleaning or soaking
dental prostheses. Polishing denture creates frictional heat on the surfaces of
PMMA made prostheses. Such concentrated heat on the surface can adversely
affect the fit of denture as a result of warpage. Dental prostheses made with
chemically polymerized resins have poorer mechanical properties, including lower
transverse bending strength, impact strength, fatigue resistance and abrasion
resistance, than those made with heat polymerized acrylic resins.
27
1.4.3 Microwave polymerization
The first report of the use of microwave energy to polymerize denture base
materials was by Nishi in 1968 [64]. Kimura et al. [65] reported that it was
possible to cure acrylic resin in a very short time using this technique. In some
other studies [65, 66], the light fibre-reinforced plastic flask was substituted for
the heavy brass flask and compress, and the water-bath curing tank was replaced
with a microwave oven. The supply of microwave energy to a liquid monomer,
which is polar in its nature, would cause the MMA to be oriented in an
electromagnetic field. The rapid flipping of such polarized molecules is supposed
to generate heat resulting from molecular friction. Free radicals are initiated and
polymerization reactions with monomers may continue. The microwave heating
might evenly distribute heat energy within the entire thickness of the bulk material
of PMMA. There will be no concerns in regard to the excessive rise of
temperature to cause boiling of monomers leading to porosity. Thus, rapid heating
cycles can be used to polymerize PMMA in a shorter time compared to water-bath
heating.
The major advantages of the microwave heating over the water-bath
heating are equal heating and rapid temperature rise throughout the bulk of
PMMA material. Peyton [67] reviewed the polymerization methods and a careful
control of the temperature during processing was the only important factor when
all factors were considered for its advantageous use. In terms of the flexural
strength, the microwave polymerized, injection-moulded, polyurethane-based
polymer offered no advantage over the existing heat polymerized PMMA denture
base [12].
28
Polymerization method determines the mechanical properties of the
denture base PMMA. So et al. showed that microwave heat treatment of coldcured acrylic resins with glass fibre reinforcement increased the mechanical
property [34]. However, Schlosberg et al. [68] found no statistical difference
between the heated water bath polymerized and microwave polymerized PMMA
denture base material in term of transverse strength, Knoop hardness, density and
residual monomer in test specimens. Thus, the observed beneficial improvement
was with post polymerization treatment with microwave of self-curing PMMA
resin. Flexural strength of selected chemically polymerized PMMA was shown to
be optimised by post-polymerization microwave irradiation [69].
Microwave generated heat polymerization may also affected by, e.g. the
volume of the investment gypsum, amount of water contained in the gypsum,
powder/liquid ratio of the resin, thermal conductivity of the flask, and microwave
translucency of the flask material.
A shorter polymerization time, and less
residual monomer are considered as two of the advantages of microwave
polymerization [49].
29
1.3.4 Light activated polymerization.
Light activated polymerizing PMMA materials are derived partly from
PMMA and partly from urethane dimethacrylates (UDMA) [70].
Photons from a light source activate the initiator to generate free radicals
which may initiate the polymerization process. A visible light source is the
preferred choice nowadays because of the safety concerns compared to ultraviolet
light. When visible light of the blue-to-violet region is irradiated towards these
materials, camphorquinone and an organic amine will generate free radicals. The
supplies of free radicals are limited by such factors as light intensities, angle of
illumination and distance of resin from the light source. The monomer-polymer
conversion is dependent on the duration of the light irradiation [71].
Machado et al. [72] observed that heat polymerized PMMA showed less
porosities in processed specimens compared to light activated polymerized
denture base polymer viz. Triad VLC (Dentsply Trubyte, York PA). It has been
proposed that internal porosities could concentrate stresses in the matrix and
contribute to the formation of microcracks under loading. These air-filled voids
presented an air-inhibited layer of incompletely polymerized matrix at the void
surface [73].
30
1.5 Mode of polymerization and its effects on PMMA
1.5.1 Porosity in PMMA.
Porosity in the final processed material has a great impact on the
properties of dental prostheses constructed. This can made the denture prone to
fracture, easy staining with dietary colouring, promote denture plaque
accumulation and finally denture cannot be fitted well. It has been proposed that
internal porosities my concentrate stresses in the matrix and contribute to the
formation of microcracks under loading [74].
Causes of porosity in PMMA base denture base material are attributed to
factors such as polymerization shrinkage, volatilization of monomer, incorrect
liquid-powder ratio and pressure application during processing, as well as thermal
contraction [8].
Polymerization shrinkages or contraction porosity may happen because the
density of polymer and monomer differs, and in addition to that, the
intermolecular bonds change from van der Waals to covalent after the
polymerization reaction. The stronger covalent bond entails a shorter
intermolecular distance, hence the closer packing of polymer molecules. The
calculated percentage shrinkage was found to be 21 vol.% in one study [75].
Volatilization of monomers also known as gaseous porosity is due to the
evaporation of the monomer, which can happen when the temperature of heated
water bath reaches temperature above 100 C. The evaporation temperature
.
C. Within the thicker area of dental prostheses, the added heat
release from the resultant exothermic polymerization reaction will cause the
temperature rising quickly above the boiling point of the monomer [8].
31
Granular porosity occurs mainly due to the loss of monomer during
mixing of the powder and the liquid monomer. It is advisable to keep the mixture
of polymer-monomer dough in a tight-lid cover container before packing it into a
mould which might minimise granular porosity in polymerized PMMA [76].
Thermal contraction occurs within the whole bulk PMMA material when
the curing temperature changes from 100 °C to post-polymerization bench cooling
temperature of
C. Voids or air bubbles inclusion during mixing and packing of
dough into mold can contribute to porosities in PMMA. When pressure is
insufficiently applied to the moulding flask during processing, porosity may occur
within cured denture. It was suggested that the screw knob on the press-flask
which hold the curing flasks, needs to be tighten to 37 N m throughout the
polymerization process to ensure that sufficient pressure be applied to maintain
the dimensional stability and prevent porosities within the bulk material of
PMMA [77]. The principle is correct; however, the 37 N m is arbitrary and may
not be applicable and suitable for all situations.
Porosities found on dental prostheses have many negative effects to the
denture wearer such as staining, harbouring the growth of microorganisms [78]
and compromising the strength of PMMA [26, 79] . A report in literature [80]
suggested that porosities on dental prosthesis had favour the retention and
subsequent progressive released of allergen persulfate, which is also a chemical
usually present in denture cleanser to cause allergic contact chelitis.
32
1.5.2 Residual methyl methacrylate (MMA)
The conversion of monomers to a polymer is never complete and some
unreacted monomer call residual monomers are left in the PMMA polymer [81].
Residual monomer refers to those substances (monomers, additives, and reaction
products) that are not firmly incorporated (whether reacted or unreacted) in the
polymer network from which they may leach out.
Monomer–polymer conversion and the residual monomer content are two
essentially important aspects of leachable substances from processed PMMA. The
rate of monomer–polymer conversion indicates the number of unsaturated C=C
double bonds converted to saturated single C―C bonds during polymerization.
The concentration of residual monomers and elutable additives are dependent on
several parameters and interrelated factors such as polymerization method,
polymerization cycle and post polymerization treatment [82].
It has been known that leaching of residual methyl methacrylate (MMA)
can cause allergic reactions in sensitized subject. On the other hand, residual
MMA can have a
plasticizing effect on the acrylic resin leading to altered
mechanical properties [58]. Residual MMA was found to leach out from bulk
material and the amount leached was proportional to the amount of residual MMA
present. Cold-cured PMMA resin leached more monomer compare to heat-cured
PMMA. The major loss of residual MMA was during the the first few days of
immersion but became almost constant after 2 weeks. This diffusive loss of
residual MMA was also found to increase with the immersion temperature [58,
83].
33
The amount of residual monomers in the polymerized resin was dependent
on the type of the polymerization process and the amount of the monomer in the
mixture ratio and the processing method [70], and especially dependent on the
duration of curing [84].
The amount of residual monomers in cold-cured resin is about 10 times
that of a heat-cured denture base resin, and the polymer in a cold cured-resin also
has a low molecular weight [8]. Baker et al. demonstrated that detectable MMA
l
lv
lv w
8 μg/ L
x
μg/ L
lv y l
g
,
the total
MMA
l b
detected for up to one week after insertion of a cold-cured appliance with the
l w
l
ly
b
g μg/ L [85].
PMMA base acrylic dentures may release MMA over long periods of time,
but the level of leachable substances is difficult to detect. Studies have shown that
MMA was not the only substance that was released from acrylic denture bases.
Other substances that could be detected included methacrylic acid, benzoic acid,
phthalates, dibutylphthalate, dicyclohexyl phthalate, phenyl benzoate, and phenyl
salicylate [86], as well as formaldehyde [56]. Even though the amount released
for formaldehyde was lower than MMA, its toxicity was higher [87, 88].
These reported leaching of residual MMA have shown mild to strong
allergic reactions in skin-patch tests. Besides the allergenic behaviour of MMA, it
was also found to be an irritant to skin and oral mucosa [89-91]. T
“b
g
” syndrome is a symptom related to such irritant and allergic reaction of
mucosa in the oral cavity is associated to the eluded residual MMA from dental
prostheses made of PMMA [92].
34
The presence of residual monomer is inevitable. An extensive research and
investigation by Lung et al. pointed out that residual monomers are an inevitable
consequence of the equilibrium of a chemistry of system. Moreover, they
suggested to take an advantage of the relative rates of depolymerisation and
diffusive loss of monomers to attain lower values of residual MMA with a
suitable choice of the processing conditions, namely time and temperature. The
suggestion of a processing temperature of 95 °C and overnight processing was
considered appropriate in laboratory conditions. Another suggestion from the
same literature review was a processing temperature at 70
C for 7 h with
terminal boiling was suggested to give the lowest value of residual monomers [58].
Residual monomers of chemical polymerizing resin decreased when post
polymerization treatment with microwave irradiation was conducted [69, 93].
35
1.6 Water storage and poly(methyl methacrylate)
1.6.1 Effects of water storage on dimension stability
The American Dental Association (ADA) Specification No. 12 for denture
base polymer recommends that the increase in mass of the polymer should not be
more than 0.8 mg/cm2 of surface after immersion in water for seven days at
37±1°C [94]. The order of water sorption is within the range of 1.0-2.0 wt.%. The
noncrystalline structure of PMMA possesses a high internal energy. This allows
the molecular diffusion to occur in the resin because less activation energy is
required when compared to a crystalline structure of a polymer. Polymers with
crystalline structure possess a highly ordered short range structure and very strong
intermolecular covalent bonds. This makes penetration of water molecules
through breaking the strong intermolecular covalent bond in crystalline structure
of the polymer quite impossible.
Although the exact mechanism through which a PMMA polymer reaches
water saturation is unknown, a 2-phase process was proposed base on the results
of a study by Takahashi et al. [95]. When a denture polymer is immersed in water,
soluble constituents, such as unreacted monomers, plasticizers, and initiators leach
out. The resultant microvoids formed are filled with water molecules by inward
diffusion. Both the outward leakage of the soluble constituents and the inward
diffusion of water are time-dependent processes, and finally may reached an
equilibrium state [95].
According to work the done by Braden [96], the kinetics of water sorption
for PMMA follows the mathematical law of diffusion and could be described by
two physical parameters i.e. the equilibrium water absorption and the diffusion
36
coefficient. The diffusion coefficient of PMMA will determine the time required
for saturation in water immersion or drying out in air. Braden also found that the
equilibrium value of water sorption for PMMA was independent of temperature,
but the temperature has an influence on the diffusion coefficient by an increment
of two times factor at the temperature range 22.5 °C to 37 °C.
Water molecules may act as plasticizers in a PMMA polymer when
subjected to water storage. The water molecules can penetrate into the spaces
between the polymer chains and push them farther apart. Thus, the intermolecular
distances of polymers increase and the intermolecular bonds, such as van der
Waals forces, are weakened. As a result, the weight and volume increase to cause
an expansion. The greater absorption of water by the PMMA material, the greater
will be the associated dimensional change [97] and decrease in flexural strength
[53]. Because the expansion effectively increases the free volume and the chain
segments polymer can move about relatively easily followed by the decrease of
the elastic modulus of the material. Degradation occurs in a PMMA denture base
material under water storage which may be due to leaching of monomer [98].
Factors that may influence water sorption induced dimensional changes
include the type of resin, thickness of material and the amount of crosslinking
polymer species. For example, a heat polymerized PMMA polymer takes a longer
time than a chemically polymerized polymer for water sorption to reach saturation
because of the lower diffusion coefficient of water [95]. However, Arima et al.
[99] found no significant differences between highly cross-linked chemical
polymerizing relining resins and heat polymerizing denture base resins [50].
There seems to be little agreement as to how long PMMA polymers should
be immersed prior to their mechanical testing. Although the international standard
37
guidelines [100] points to 50 hours of water immersion, Takahashi et al. [101]
suggested that the equilibrium strength of some denture polymers may well
exceed 30 days and they recommended a 4-month water immersion protocol
whereas, the water sorption mainly occurs during the initial 14 days [95]. In the
study by Chow et al. wa
w
w
b
y
the linear expansion
was 0.45 % [102].
38
1.6.2 Effects of water storage on surface of PMMA
Von Fraunhofer et al. [101] demonstrated that at room temperature water
could absorb into acrylic denture base material and cause changes in
microhardness of the surface. Absorption of water into PMMA resins is facilitated
by the polarity of poly(methyl methacrylate) molecules; water molecules penetrate
the mass and occupy position between the polymer chains.
A craze is a localised region of plastic deformation of polymer which may
be filled with small voids. As the voids in the crazed region grow these tiny voids
become separated by thin fibrils of polymer, ultimately the fibrils fail and a crack
is formed. In fact, the mechanism of crazing found on these surfaces is related to
solvent attack and water sorption. PMMA is saturated with water through the
process of water sorption, once it is exposed to dry environment, drying will take
place progressively from the surface extending into the depth of the full thickness
of bulk material. This can result in a water concentration gradient within the bulk
material of PMMA. The concurrent effect of drying is the shrinkage of material
on the air contacting surfaces. The outward surfaces are dry while the internal
section is wet, which fact can lead to build up of tensile stresses on the surface
that, on the other hand, can cause surface crazing through action of solvents such
as MMA. PMMA processed dental prostheses which are totally well soaked in
water or thoroughly dry, will be free from the surface crazing effect.
Crazing generally begin at the surface of PMMA resin and is oriented at
right angles to tensile forces. Micro cracks formed in this manner will progress
internally from the surface into bulk material. Heat generated during polishing of
39
dental prosthesis, different contraction around metals component or attack by
solvents such as alcohol may lead to crazing.
Zappini et al. [12] reported that the fracture pattern of Lucitone® 199, a
denture base through heat polymerized PMMA, exhibited large amounts of
permanent deformation and crazing before fracturing. Crazing on surface of
PMMA material would promote an ultimate material fracture. Precaution taken to
stop crazing can extend the service life of a PMMA made denture.
40
2. Hygiene aspects of prostheses made of poly(methyl methacrylate).
2.1 Biofilm and plaque on dental prostheses
In the oral cavity, an acquired pellicle forms rapidly on non-shedding
surfaces. This acquired pellicle is composed of primarily host derived salivary
substances which include high molecular weight mucins. This is to be followed by
a microorganism adhering to a component of the acquired pellicle and thus
denture plaque accumulates through continuous adhesion, aggregation, and
growth of attached microorganism on the denture surfaces [103].
Biofilms are defined as microbial communities encased in a matrix of
extracellular polymeric substance (EPS) and, displaying phenotypic features that
differ from their planktonic or free-floating counterparts [104]. The plaque and
biofilms on a denture or dental prostheses are composed of a complex mixture of
fungi, bacteria and desquamated epithelial cells from the host. This biofilm acts as
a protective reservoir for oral microorganisms. Studies have demonstrated that
different species of oral and non-oral pathogens are associated with denture
plaque. These include Candida spp., Staphylococcus spp., Streptococcus spp.,
Lactobacillus spp., Pseudomonas spp., Enterobacter spp. and Actinomyces
spp.[105, 106]. Mature biofilm on natural and denture teeth may have similar
total numbers of bacteria but different species proportions [107].
The continuous swallowing or aspiration of micro-organisms from denture
plaque may pose a risk of upper respiratory tract infection in vulnerable group
such as immune compromised patients or debilitated elderly people [108].
Nikawa et al. (1998) proposed the term ‘ l q
’ which should
be used because the microbiota and its pathogenicity of denture plaque resemble
41
those of dental plaque formed on the tooth surface. Moreover, they suggested that
‘
l
’ w
l
b
red
‘
’, because the inflammation of (palatal) mucosa is not induced by the
denture, but by wearing the denture or by plaque accumulation on the denture
[109].
In view of these facts presented, the efficient mechanical plaque removal
is one of the important oral health measures to prevent the onset and progression
of denture related diseases for individuals.
2.1.1 Candida albicans biofilm
The development of a Candida biofilm consists of four steps. The first
step is the adhesion of a microorganism onto a surface [110]. The second involves
discrete colony formation, and organization of cells. The third step is the secretion
of extracellular polymeric substances (EPS) and maturation into a threedimensional structure. The final step is the dissemination of progeny biofilm cells.
Some studies have reported that Candida biofilm formation on PMMA
occurs in three phases namely an early phase, intermediate phase, and the
maturation phase [111]. The early phase was characterized by the presence of
adhering and developing blastospores into distinct microcolonies. Intermediate
phase can be seen between 18 to 24 h and the Candida biofilm appears as a
mixture of yeasts, germ tubes, young hyphae and an extracellular polymeric
42
substance (EPS). The mature Candida biofilm consists of dense network of yeasts,
pseudohyphae, and hyphae embedded in a thick EPS layer.
Fig. 2.1 Scanning electron micrograph of different morphology of Candida
albicans on surface of PMMA specimen.
Denture wearers harbour the growth of Candida albicans when the
hygiene practices and maintenance standard are poor. The prevalence of Candida
albicans present on denture was reported by Meurman et al. at 78.4% of the
population [112].
Denture plaque containing Candida may cause denture-induced stomatitis,
oral candidiasis, as well as Candida-associated caries, and periodontitis of
43
abutment teeth. However, only limited evidence is available on the cariogenic
potential of Candida [113].
The presence or absence of dental prosthesis determines the prevalence
and concentration of Candida in different sites of the oral cavity. In healthy
denture-wearing carriers Candida is found distributed on the dorsum surface of
tongue, fitting surface of the upper denture and in some cases the palate [114].
Evidence from previous studies showed that presence of dental prostheses [78] or
removable orthodontic appliances [115] increases the density of Candida
colonization in all sites in the oral cavity.
A major clinical implication of Candida biofilm is its higher resistance to
antifungal agents [116-118].
44
2.2 Denture hygiene
The purpose of denture hygiene practise is to minimize or totally eliminate
the fungal and bacterial attachment and activity on dentures, in addition to
reducing food-related stains on the denture surfaces. This would prevent the
occurrence of denture stomatitis on the oral mucosa covered by the denture [119].
Denture cleaning may be divided into mechanical cleaning method and
chemical cleaning methods [120, 121]. A mechanical cleaning method such as
brushing or the use of ultrasonic water bath are the most effective means in
removing denture-plaque and biofilms from the surfaces of dental prostheses.
Qualities of a good denture cleaning agent are: ease of use, effective removal of
deposit on surfaces of denture without damage to the denture, bactericidal and
fungicidal, non-toxic, non-irritant, chemically defined and stable, regulatoryapproved on a global basis and cost-effective [122].
Chemical cleaning methods using denture cleansers are popular among the
denture-wearers for the prevention of colonization by Candida albicans and
related Candida species, as well as to impede denture plaque formation. Effective
plaque removal from soiled dental prostheses from a period of in-service use
requires a degree of manual dexterity that is often lacking especially among
elderly patients and those individual who are ill and frail [123]. The value of good
dental and denture hygiene, as shown in a systematic review by van der MaarelWierink et al. [124] in the frail older people, could reduce aspiration pneumonia.
Another systematic review on the subject of cleaning denture for adults by
Souza Raphael et al. pointed out that no evidence conclusively states that any
denture cleaning method is more beneficial for the health of denture bearing areas
45
’
and preference when compared with each other [124,
125]. There is a lack of evidence on this subject of effective denture cleaning
method.
Professional denture hygiene instruction goes hand in hand with the
advanced designs of devices in achieving an optimal effect on denture hygiene
practice. The same has been shown in the study on maintaining dental hygiene.
When professional instruction was not given, the potential of the electric powered
toothbrush was underutilized, compare to the manual toothbrushes [126].
Currently, a plethora of information from various sources contributes to
the controversy of the easiest or most effective cleaning method to maintain
denture hygiene and much of it can be controversial. There is a need in this field
of studies to clear this confusion, and this current investigation at hand may help
to advance the understanding on the subject of mechanical cleaning methods one
step further.
A study conducted by Sumi et al. pointed out that elderly people may have
difficulty in mechanically removing dental plaque due to diminished manual
dexterity, impaired vision, or ill
.T
l
’
bl y
perform self-
care gradually decreases with advancing age, and the role of the caregiver in daily
oral care becomes increasingly a necessity. However, optimal oral care by
caregivers is not always possible because of the time constraints, difficulties
involved in brushing other individuals’
,l k
, and the lack of
perceived need. The recognition of this issue faced by the elders and care givers
have prompted them to make a device through modification of a powered
toothbrush. The results from their investigation were positive and promising,
46
especially in helping those concerned subjects and care givers to achieve good
dental plaque control [127]. Briefly, the use of a powered toothbrush is not just
an aid to the elderly, it can help the care givers substantially in delivering better
services as well. Denture hygiene practise may further improve from the use of
powered devices such as powered toothbrushes. No quantitative and qualitative
investigation on the positive contribution from such usage is yet reported.
47
2.3 Mechanical cleaning method for dental prostheses
Brushes of various designs and made are use in the mechanical cleaning
method (Fig. 2.1). Broadly, they can be classified into manual and powered
brushes plus the sonic cleaners. The mechanical movement of bristles
perpendicular to the surface of dental prostheses creates a scarping motion to get
rid of the layer of an attached biofilm.
The efficacy of brushing with regard to plaque removal is dictated by three
main factors namely the design of the brush, the skill of the individual using the
brush and the frequency and duration of use [128].
48
Varieties in
Bristles stiffness, form and arrangement.
Size and shape of head &
handle design.
Manual
denture
brush
Manual
toothbrush
Brushes for cleaning dental prostheses
Powered
denture
brush
Powered
toothbrush
Direct
Replaceable batteries
Rechargeable batteries
Power
source
Motion of head
Reciprocating
Dual motion
Arcuate
Circular
Vibratory
Sonic
Ultrasonic
Fig. 2.1 Diagram showing types of brushes for cleaning dental prostheses.
49
2.3.1 Powered denture brush
The powered denture brush is a purpose-designed and manufactured
device, commercially marketed in the United States. The designer Jim Harrison
found that denture wearers often lacked the ability, knowledge or dexterity to
maintain the hygiene of their prostheses. He conceived of the idea for a purposedesigned powered denture brush from a manual denture brush. He and his team
members designed and developed the Power Dent Pulse™, the brand name of the
powered denture brush. It looked like a manual denture brush except with a large
handle to hold batteries. To date, no research has been conducted to investigate
efficacy on denture plaque removal and the influences that may impact on the
properties of acrylic resin of its usage. This investigation initiated intends to
explore the gap in this area of research by means of using the battery powered
toothbrush which has a simple rotational movement bristles head.
50
2.3.2 Powered Toothbrush
The origin of powered toothbrushes date back to the 18th century. They
were originally designed as a mechanical toothbrush [129] by a Swedish
l k
k
D S
’
l
b
[130], but it was not until the 1960s
that electrically powered toothbrushes were introduced [131] and became
established as a preferred alternative to manual toothbrushes. These designs
‘b k
’
l
l
b
.
Powered toothbrushes have been indicated for young children,
handicapped or patients with special needs who lack the dexterity to brush
manually. Patients wearing fixed orthodontic appliances and hospitalized or
institutionalized patients who need a care worker or nurse to carry out oral
hygiene, would benefit from using powered toothbrushes. A powered toothbrush
may also be considered for any individual who wish to improve the tooth brushing
technique and enhance interest in the oral hygiene practice. It may increase
motivation and compliance with oral and denture-hygiene measures. It is desirable
to extend these benefits to individuals wearing dental prostheses through the use
of powered toothbrush as a mean of mechanical cleaning to remove accumulated
plaque and biofilm. Mechanisation of tools, in any form, is certain to produce
some form of efficiency in the process application. There is currently a favourable
trend in the continuing advancement and improvement of devices of personal
hygiene in general, and oral or denture hygiene.
The parts of powered toothbrush consist of a head, bristles and a handle
which house the batteries. The head which is connected to the shank is detachable
from the handle and also replaceable. In general, power toothbrush heads are
51
smaller than manual toothbrush heads. They are available in a full size or a
compact size head. The shape of a replaceable head may be circular, rectangular
or diamond-like.
The bundles of bristles are arranged either in rows (as for a conventional
toothbrushes) or in a circular pattern mounted in a rounded head. Bristles are also
arranged as more compact single tufts which facilitate interproximal cleaning and
brushing in less accessible areas of the mouth.
Bristles used in the manufacturing of brushes for personal hygiene
maintenance are commonly made of nylon e.g. Tynex® by DuPont. The hardness
or stiffness, end-rounding, shape and type of bristles have influences on the
efficacy of plaque removal from surfaces. The factors such as the material, length,
thickness, compactness and tip geometry of bristles, might affect abrasion by
tooth brushing. The same brushing abrasion effect is accurately correlated in tests
carried out on PMMA
’ surfaces as well.
The motion of the brush head includes a multitude of different movements
e.g. reciprocating, circular and sonic vibratory. Some advance models (e.g
Vitality™ Dual Clean by Oral-B, Proctor & Gamble) may be a combination of
two types of motion. Range of sonic vibration is 18,000 to 32,000 brush strokes
per minute.
The speeds at which the head or the bristles move differ according to the
designs of powered toothbrushes. The number of strokes per minute varies from,
for example, as low as 1,000 cycles per minute for a replaceable battery type, to
about 3,600 oscillations per minute for an arcuate model. The ultrasonic bristles
vibrate at 31,000 brush strokes per minute.
52
The power source that supply direct current to the powered tooth brushes
is commonly replaceable batteries or rechargeable batteries.
The force applied during the use of power toothbrush for brushing teeth in
the mouth is less compare to manual toothbrush [132]. Findings of van der
Weijden [132] showed that the force used with an electric powered toothbrush
was significantly less than with a manual brush when performing tooth-brushing.
They also recorded the forces loads for powered toothbrushes which were in the
range of 80-90 g and in the 250 g when manual toothbrushes were used. A
contemporary powered toothbrush may feature a pressure sensor to avoid gingival
trauma from overzealous brushing. Pressure sensor incorporated into a powered
toothbrush enables the user to self-adjust the applied force-pressure when
brushing force is over the pre-set limit. The brush head flexes back with an
audible warning buzz and light when a predetermined brushing force threshold is
breached.
A timer is incorporated in some powered brushes to allow the users
beware of the time recommended for tooth brushing and not exceed the duration.
Many studies [59, 133-135] attributed the harmful and undesirable adverse effects
of tooth brushing to prolonging as well as to a longer accumulated total time of
brushing. These features can help to minimise the damaging effects of surface
abrasion and surface wear of substrate caused by brushing.
Tooth brushing abrasion is influence by variables such as the frequency of
brushing, the force applied during brushing, the characteristics of bristle, the
bristle stiffness and the end-rounding of bristle ends [18].
53
The subject of abrasion to dental hard tissue and gingiva is an inquiry that
challenges many researchers to find a reliable conclusion. The same is true for the
effects of denture hygiene practice impact on the surface quality of dental
prostheses. There is still some debate about the conclusions made. However,
unequivocal consensuses among the investigators have concluded that the
brushing of dental prostheses using brush with toothpaste can abrade the surface
of acrylic resins [57, 136]. Factors that were attributed to abrasion include
frequency of brushing, applied brushing forces, bristles characteristics such as
stiffness and end-rounding, and composition of dentifrice used [18]. Brushing
PMMA specimens using a powered toothbrush with water alone did not cause
increase in surface roughness [136]. The linear motion of bristles caused more
wear and abrasion on PMMA specimens compare to rotary motion of bristles
[137].
According to a previous study, the occurrence of a polishing effect on
resins may be possible when the denture was brushed using a toothbrush with low
abrasion toothpaste [138].
Cleaning and brushing teeth using powered toothbrushes have been proven
to reduce dental plaque and gingival bleeding score in clinical trials. A powered
toothbrush is effective and efficient compared to the use of manual toothbrushes
in reducing dental plaque score [139]. However, there were inconclusive findings
from other investigators stating that there was no different in efficacy between
powered toothbrushes and manual toothbrushes [140]. This may be due to
differences in experimental designs and conditions [18].
54
2.3.3 Manual denture brushes and manual toothbrushes
These mainly consist of purpose-designed denture brushes with a large and
a small bristles area opposing each other back to back. The longer, circularly
arranged tufts of bristle are used to clean the tissue surface or small hard-to-reach
surfaces of the denture. The flat rectangular portion is used for cleaning the
polished and the occlusal surfaces. A regular toothbrush with regular stiffness of
bristles can be used for cleaning dentures as long as its design permits access to
all the surfaces of dentures. These toothbrushes are easily and readily available
alternative means to clean dental prostheses, as well as conveniently within reach
and comparatively cheap for the public. The difficulty to adapt a regular size
toothbrush into the recesses and curves of the fitting surface may sometimes result
in an inadequately clean denture.
The choice of bristles on any toothbrush used for dental prostheses
cleaning should neither be too soft nor too hard in stiffness. The use of a
toothbrush with hard bristles and excessive pressure during overzealous brushing
can result in damaged surfaces of PMMA denture base material to the extent that
the fit and aesthetic of the prosthesis may be compromised. The physical and
mechanical properties can be adversely affected as a consequence, which may
lead to a catastrophic fracture which may happen below a normal load of function.
55
2.3.4 Sonic and ultrasonic cleaners
Sonic and ultrasonic devices are mechanical aids generally used by the
dental professionals. A chemical solution may be used together with an immersion
medium to increase the efficacy. Sonic cleaners operate through the generation of
audible sonic energy wave; ultrasonic cleaners operate by means of highfrequency sound waves. The processes are generally known as sonication where
particles are dislodged from adhering to surfaces. The sonic type of cleaners are
more common among the general population and they produce less mechanical
agitation of solution than do the ultrasonic found in most dental operators. It has
been reported that ultrasonic sonic cleaning done in the dentist’ office was more
efficient in plaque removal than either a commercial chemical immersion cleanser
or a sonic cleaner at home [141].
There are two mechanisms of action proposed for ultrasound generated
cleaning, the first being the movement of liquid resulting from sound waves
transferred to the liquid (vibration), and the second, the cavitation and collapse of
bubbles formed by vibration of the unit [142].
Budtz-Jorgensen et al. reported that although ultrasonic treatment by itself
did not reduce the number of microorganisms that could be cultured from soiled
dentures, it did enhance the effectiveness of disinfecting solutions in which
dentures were immersed during the ultrasonic treatment [143].
There is disagreement as to the effectiveness of these mechanical cleaners
in reducing plaque on denture. The effectiveness of ultrasound generated cleaning
is contradictory; it may be attributed to the mechanical action of the device [144]
or to the chemical solutions used [141]. The combination of an ultrasonic cleaning
56
method with brushing or with a chemical immersion method has been suggested
as an effective alternative for cleaning complete dentures; however, this
effectiveness has not been clinically tested [145].
57
2.4 Chemical cleaning
The use of chemical solutions for soaking dental prostheses is a popular
alternative to mechanical cleaning. It requires less effort and compliance from
individuals and the solution may reach surfaces on the prostheses that may be
inaccessible to denture brush. Commercial denture cleansers are classified into the
following groups: neutral peroxides with enzymes, enzymes, acids, hypochlorites,
peroxides, crude drugs, and mouth rinses for dentures [121, 146] . Denture
cleansers are widely used to prevent colonization by Candida albicans and related
Candida species, and to prevent denture plaque formation in addition to the
primary function of plaque removal from dentures [147].
Daily use of denture cleansers as proposed by Palenik et al.[141], if
inappropriately done by ill-informed individual, can affect the physical and
mechanical properties of the PMMA denture base materials. For some individuals
or in circumstances that could raise the concern of safety issues, it may be best to
avoid any use of such chemicals or microwave irradiation disinfection. A case in
point may be the incapacitated seniors with concomitant dementia and frail
condition. Simply brushing a dental prosthesis with a suitable powered toothbrush
and water could possibly be a more viable and safer choice in such a situation.
Sodium hypochlorite is effective as a denture cleanser or disinfection
agent because of its ability to dissolve organic material which is made up of
accumulated biofilm on the dental prostheses. It is bactericidal and fungicidal
which can fulfil the objective of disinfection of dental prostheses. However, it is
not suitable for dental prostheses with metal components due to corrosion of these
metal parts. Dental prostheses should be soaked in a hypochlorite solution for
58
only 10 to 15 min and not over night because hypochlorite has an unpleasant taste
and odour. Bleaching of an acrylic resin denture by sodium hypochlorite has been
reported by McNeme et al. [148] and Moore et al [149]. No matter how brief the
contact time between the metal components, much as the low concentration of
hypochlorite used, the cumulative effect of regular long term use cannot be
underestimated.
Glutaraldehyde disinfection of acrylic resin denture was investigated by
Polyzios et al. [150]. They used a microwave energy polymerized PMMA
specimens immersed in a solution of 2% alkaline glutaraldehyde (Cidex-7,
Johnson & Johnson. East Windsor, NJ) for 1 h or 12 h. They had compared the
chemical method vs. using microwave irradiation disinfection with respect to the
dimensional stability, flexural properties and hardness of acrylic resin. Their
results showed that flexural properties were not affected by these two methods of
disinfection. However, contacting glutaraldehyde on eyes or skin can cause
irritation (American Dental Association, 1985). There is also possibility of some
residual solution in the acrylic resin because of the porosities found on the
surfaces of prostheses. It is important that the selection of the disinfection
solutions should be based on its disinfection effectiveness and compatibility with
the oral tissues. Shen et al. studied the effect of two glutaraldehyde base
disinfectants, namely alkaline and phenol buffered, on flexural strength and
rigidity of denture base resins and reported that the flexural strength was not
affected by immersion time or the type of disinfectant used [151].
The alkaline and neutral peroxides act through an oxygen-releasing
mechanism, which loosens debris and removes light extrinsic stain on surfaces.
The effervescent tablets sold over-the-counter belong to this group. When these
59
agents are added to water they produced an alkaline solution of hydrogen peroxide
[152]. This peroxide solution subsequently releases oxygen and the
oxygen
bubbling effect would enable minor mechanical cleaning [143]. The oxidising
agents incorporated can remove extrinsic stain and serve as antibacterial agents. A
study by Nikawa et al. revealed the ability of the peroxide cleansers to decrease
Candida albicans biofilm activity [146].
The use of the peroxide preparation on a regular basis may help to prevent
the formation of stain and calculus if accompanied by brushing and rinsing. In
some commercial formulations enzymes may be added to ‘enhance’ their
antibacterial action. Enzymes are effective in destroying the plaque matrix and
allowing the accumulated plaque be easier brushed or rinsed off [153]. This type
of chemical cleaning is generally considered as safe and effective for cleaning all
type of dental prostheses including those with metal components. However,
precaution needs to be taken seriously, not to ingest the tablets by accident. The
elderly or visually impaired may mistake the tablets as antacid tablets and small
children may be vulnerable as well.
The proper care of denture-soaking container is critical in a situation
where the critically ill individual is with suppressed immunity. The chemical
soaking solutions can serve as growth media for pathogenic microorganisms that
are formed on dental prostheses. Well cleaned dental prostheses may be recontaminated when placed in the holding containers with such solutions. The recontaminated dental prostheses may become a source of infection for such
patients. The hassle of cleaning the holding container and discarding the soiled
solution is a pertinent issue well worth the effort to be carried out conscientiously.
60
Diluted acids are used as aids to clean dental prostheses. They include 3%
to 5 % hydrochloric acid with or without phosphoric acid, and clear coloured
household vinegar. An acid solution dissolves the inorganic material which
accumulated on the dental prostheses and is good for the removal of persistent
stains not removed by regular cleaning methods. These acids should be handled
with extra care. The ability of acids to corrode metals is a disadvantage that limits
their use to prostheses without any metal parts.
Chlorhexidine is commonly prepared as the digluconate or acetate salt in
suitable concerntrations in aqueous solutions. Chlorhexidine destroys bacteria by
breaking their membranes and inducing cytoplasmic precipitation [154]. It is a
cationic molecule capable of interacting with inorganic human dentine particles
and it also bonds to negatively charged surfaces, such as the bacterial cells wall
[155, 156]. Chlorhexidine gluconate is an antiseptic agent with a broad spectrum
of antimicrobial activity including Candida albicans and other common nonalbican Candida species [157, 158].
Chlorhexidine gluconate at concentration 0.12% with immersion duration
of 15 min and 2.0% with an immersion duration of 5 min have been shown to be
effective for denture cleansing in clinical trials, compared to using water as the
control [159]. A major drawback with chlorhexidine is extrinsic staining on dental
prostheses after a repeated use, making it unsuitable for a daily routine use.
As the facts presented demonstrate that chemical method still fall short of
ideal cleaning and the inherent shortcomings of chemical contain within these
cleansers, therefore, chemical denture cleansers should be used as an adjunct with
61
other methods of cleaning. In vivo studies are lacking the information whether the
daily use of a cleanser may cause mucosal irritation and allergy [80].
Comparison on efficacy of the various denture-cleaning products is
difficult due to the lack of research studies comparing a variety of products used
on dentures in vivo. Available research results are difficult to compare because of
the wide variations in materials and methods as well as in the quantification of
results. Nikawa et al.[121] had attempted discussion on this subject and suggested
a recommendation addressing this issue on comparing efficacy of the denturecleansers.
62
2.5 Microwave disinfection.
The use of a standard microwave oven as an efficient method for cleaning
and disinfecting denture was first suggested by Rohrer and Bulard [160]. The
proposed mechanism for the action of disinfection was the thermal effect and the
non-thermal effect. The thermal effect on Candida albicans was studied by
Campanha [87] and it led to a suggestion that the direct effect of heat could alter
cell structures, modify cell membrane permeability and cause cell death. The
effects are the direct causes of heat generated by microwave radiation upon the
organic
matter.
The
non-thermal
effect
involved
the
destruction
of
microorganisms at a temperature lower than the thermal destruction point [88].
Microwave irradiation is selectively absorbed by certain biochemical molecules
such as proteins, nucleic acids and protein-lipopolysaccharide compounds of cell
membranes which can lead to cell destruction [161].
It seems that the microwave disinfection is simple, easy to use, effective,
quick and a cheap method for the denture disinfection [162, 163]. The incidence
of drug induced resistance for fungi and other microorganism can be eliminated.
However, it cannot be used if the dental prostheses contain any metal components.
Water and other solution have been used as media during microwave disinfection
of prostheses to prevent detrimental dimensional and structural changes [160]. It
was suggested that placing the dental prosthesis in a cup of water to obtain an
even distribution of heat was more effective for disinfection [164]. Furthermore,
the microwave energy might be able to further cure the denture and alter the
mechanical and physical properties, such as hardness, flexural strength and colour
of the denture [34]. Therefore, many other relevant points are to be considered
before embarking on this method of cleaning dentures.
63
Studies suggested that the exposure time required for microwave
disinfection ranges from 1 min to 20 min and the power setting varies from the
low of 350 W to the high of 800 W [60] and the immersion media used during
microwave irradiation disinfection may include water [164] or sodium
hypochlorite. A point of concern is that such heated liquid could possibly pose a
danger of scalding to an individual especially aged and frail. Controversy and
mixed result still appear in reported experiments when dentures have been
colonized by microorganism in laboratory compare to when it is taken directly
from the patient’s mouth [165, 166].
Both short term and long term effects of microwave use on denture
materials are inconsistent and no consensus has been agreed on an acceptable
standardised protocol for the microwave oven therapy. There still exist potential
risk for causing structural damages to PMMA denture base material on its
application [60]. The adverse effects of microwave irradiation on the mechanical
properties of PMMA depend on the duration of exposure, the power setting of
microwave oven, immersion solution, frequency of use and the polymerization
process whether auto or microwave.
The efficacy of microwave disinfection of the dental prostheses seems to
be temporary, re-infection and re-growth of plaque biofilm can readily occur as
the mouth remains contaminated. It seems unreasonable to microwave a dental
prosthesis daily or a few times a week as a replacement for regular brushing.
When regular cleaning is performed prior to microwave disinfection, the results
tend to favour better disinfection and denture related stomatitis control [60].
64
Residual biofilms remaining on oral mucosa could lead to fungal regrowth and re-colonization, regardless of how thoroughly cleaned the prosthesis is
when putting it back in the mouth. The oral mucosa needs to be swabbed clean as
well which implies the prosthesis is off the mouth and cleaning with liquid soap
and a powered toothbrush is a logical action step to achieve a better denture
plaque removal.
The present philosophy of making a killing of microorganism requires a
balanced thinking of sustaining a reasonable length of material life span through
guarded material deterioration.
There is an increasing acceptance of the
hypothesis that the microflora should not be eliminated but should instead be
prevented from shifting from a favourable ecology to an ecology favouring oral
disease [167].
65
3. Conclusion
The use of battery powered toothbrushes as a mean of mechanical cleaning
method of PMMA prostheses has a good potential to be promoted as a good and
safe device, especially for the elderly patient and their care-takers. The use of such
devices requires judicious care and consideration from the dental health care
providers and the end users. Both professionals and patients are definitely in need
of quality information to assist to make a well informed choice from the great
varieties of options on brushing devices available on the market.
66
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82
Part II: Laboratory Report
This part has been submitted as the manuscript with title “Effect of powered
toothbrush cleaning on acrylic resin dental prostheses” to the journal,
Gerodontology.
Effects of Powered Toothbrush Cleaning on Acrylic Resin Dental Prostheses
Abstract
Introduction
Optimal denture hygiene maintenance requires brushing and other supplementary
methods to effectively remove plaque and biofilm build-up on surfaces of dental
prostheses. Brushing denture using manual brushes is the most common practise
among denture wearers. Evaluation on the effects of mechanical cleaning using a
powered toothbrush on poly(methyl methacrylate) (PMMA) denture base material
was performed.
Materials and methods
Heat polymerized PMMA specimen beams with the dimension of 45.0 x
6.5 x 4.5 mm were fabricated in stainless steel moulds. PMMA of the brand
Trevalon C (Dentsply Ltd. De Tray Division Weybridge, UK) was mixed at the
powder to liquid ratio of 2.4 : 1 and processed according to manufacturer’s
instruction. Specimens were kept in water storage for 0, 1, 7, 15, 30 and 60 days
was assigned to control and then tested at random. Test specimens underwent a
focal area of brushing action using battery powered toothbrush at 2 N applied
force for 22 min with water as the medium. Observations were made on the
surface roughness measurement (Ra) and flexural strength computed through
performing 3-point bending test. Specimens had Candida albicans biofilm coated
and efficacy of powered toothbrush removing the biofilm was investigated. The
84
results of mean surface roughness value and flexural strength were analysed by
using 2-way ANOVA and Tukey post hoc test at 5% significance level.
Results
The mean ± standard deviation of the Ra value of surfaces of PMMA specimens
before and after brushed for 22 min with water were 0.29 ± 0.06 μm and 0.27 ±
0.06 μm respectively. The lowest mean flexural strength of 87.37 ± 9.23 MPa was
recorded for specimen group after brushing plus water storage for 7 days. The
highest mean flexural strength was 103.72 ± 8.52 MPa for specimens without
water storage after brushed. The experiment showed that PMMA specimens
recorded decrease surface roughness value and no observable significant changes
on flexural strength after brushing with power toothbrush. However, flexural
strength and surface roughness value were significantly lower in specimens group
after 7 days in water storage compared to control with no storage. Mean flexural
strength for 15, 30 and 60 days storage reached a plateau thereafter. SEM
micrographs of post-brushed specimens revealed satisfactory removal of Candida
albicans biofilm.
Conclusion:
Power toothbrush seemed a suitable device for cleaning dental prostheses made of
poly(methyl methacrylate) when brushing was performed with liquid medium
added.
85
1. Introduction
Poly(methyl methacrylate) (PMMA) base dental prostheses deteriorate
over the useful time span of dental prosthesis. The deterioration is related to its
inherent properties as a polymer, the physical wear and tear in the oral
environment during function as well as methods of denture hygiene practised.
When the dentures are inappropriately cleaned, which lead to surface and bulk
material flaw, this can contribute to the material deterioration within PMMA and
give way to ultimate failure. It is of interest to investigate the influence and the
effect of using powered toothbrush denture cleaning protocol on PMMA denture
base material.
The oral cavity is a unique environment that promotes formation,
accumulation and deposition of biofilm on the surfaces of natural tissue and
artificial prostheses. Microbial biofilms form on the teeth is known as dental
plaque whereas the biofilm on dentures are termed denture plaque [1]. Matured
biofilm on natural and denture teeth may have similar total numbers of bacteria
but proportion of sepsis may differ [2].
Persistent long term accumulation of denture plaque can cause denture
related stomatitis [3] in denture wearer and aspiration pneumonia in frail elderly
through continuous swallowing or aspiration of microorganism present on denture
[4]. In particular, the old and frail elderly people have unique requirement in their
need to maintain denture hygiene. Their frail conditions have made the task of
personal hygiene and dental hygiene very challenging for themselves and for their
caregivers [5]. They require some form of efficiency enhancement design
incorporated in denture brushes so as to help them cope with brushing denture.
86
The simple, compact designed, highly versatile and affordable generic brand
battery powered toothbrush suggested here, may be suitable for the old frail as
well as their caregivers to help with better daily oral and denture hygiene
maintenance.
A study conducted by Sumi et al. pointed out that elderly people may
have difficulty in mechanically removing dental plaque due to diminished manual
dexterity, impaired vision, or chronic illness [5, 6]. The older person’s ability to
perform self-care gradually decrease with advancing age, and the role of the
caregiver in daily oral care becomes increasingly a necessity. However, optimal
oral care by caregivers is not always possible because of time constraints,
difficulty involved in brushing other individuals’ teeth, lack of cooperation, and
the lack of perceived need. The recognition of this issue faced by the elders and
care givers prompted them to make a device through modification of a powered
toothbrush. The results from their investigation were positive and promising,
especially in helping those concerned subjects and care givers to achieve good
dental plaque control [6].
The presence of distressing oral malodour is of great concerned to denture
wearer which is the direct consequence of inefficient cleaning of denture [7].
There is a need to consistently perform good standard of denture hygiene and oral
hygiene practice in order to prevent the development of pathology affecting
general health and oral health [1, 8]. An early study described the methods of
cleaning dental prosthesis as two broad category of mechanical and chemical
means [7]. The addition to these groups which can be considered new are
ultrasonic solution or water-bath cleaning and microwave generated denture
sterilization in microwave oven [9].
87
A mechanical cleaning method such as brushing or use of ultrasonic water
bath are the most effective in removing denture-plaque and biofilm from surfaces
of dental prostheses according to previous studies [10, 11]. Qualities of a good
denture cleaning agent are, but not limited to: the ease of use, effective removal of
deposit on surfaces of denture without damage to the denture, bactericidal and
fungicidal, non-toxic, non-irritant, chemically defined and stable, regulatoryapproved on a global basis and cost-effective [12]. Battery operated powered
toothbrush can satisfy most of the required qualities, except bactericidal and
fungicidal.
The mechanical cleaning method with brushes involves mechanically
moving bristles across the surfaces of denture. Toothpaste or powder cleanser can
be added together during brushing, in order to increase cleaning efficiency [13].
The use of toothpaste carries within it the risk of abrasion created on denture
surfaces, due mainly to its abrasive nature of the filler particles. Tap water is used
as rinsing medium during and after cleaning with brushes. The cleaned dental
prostheses will either be worn in the mouth or stored in a container filled with an
immersion medium. The types of immersion media use may contain any
combination or single constituent of antiseptic chemical, or antifungal and
antimicrobial substances plus chemical solvent for example alcohol. From the
denture wearer’s perspective and the manufacturer’s interest, colouring dye and
flavouring chemical are a must-have formulated addition into these over-thecounter personal hygiene consumer products sold in a great number of
conveniently located retailing business outlets. The efficacy of using such solution
to clean and to “kill” bacteria in denture plaque still remains an elusive pursuit to
most denture wearers and an inconclusive confusion to the dental profession.
88
During the physiologic function of the denture in the mouth and
throughout the useful life span of the dental prosthesis, it is constantly subjected
to physical load as well as chemical interaction with the immersion media. When
the denture is left outside the mouth and kept in an immersion solution, surfaces
of denture may be influenced by the immersion medium. The dentist often
advices the denture wearer to meticulously clean the denture daily. However,
when such measures are being over zealously under taken by the patient, the
longevity of the prosthesis would be compromised [13, 14]. The brushes, cleaning
pastes, denture-soaks and other liquid chemicals that can have long duration
contact with the surfaces of the PMMA denture base material will certainly speed
up the process of deterioration of PMMA made dental prostheses. Adding to this
list are the chemical substances that denture wearer might consume in the diet, the
PMMA is constantly subject to chemical challenges at the molecular level. The
qualities of the physical and surface properties of PMMA denture base material
are certainly affected. Cracks, craze lines and voids or surface irregularities buildup on the surfaces of denture will cause stress fatigue during normal functional
load and lead to may result in the eventual denture breakage [14].
Factors that could affect the surface properties and the mechanical
properties of PMMA include the magnitude of force applied during brushing, the
stiffness of the bristles, the type of brushing media used concurrently, the total
duration of brushing, the type of cure of PMMA used and the temperature setting
of the brushing performed whether under cold or hot water condition. The
resultant observable increase in surface roughness was reported when brushing
was performed using a type of toothpaste [15].
89
Previous researches emphasized cleaning the denture to remove denture
plaque biofilm and the efficiency and efficacy were the main primary target
outcomes [15, 16]. There is a lack of information and research carried out on the
subject of parallel damage affecting the surfaces of dental prosthesis made of
PMMA that contribute to material substance deterioration which cause the
ultimate failure of prosthesis. The research paper by Harrison et al. [15] was
referred to for the experimental protocol and the design of current study. The
current study follows the same applied for brushing. Some variations were made
to suit the current laboratory condition. The force applied on the brush head was
set at 204 g equivalent to 2 N of the normal brushing force.
Denture wearers have very limited choice of suitable brushes to choose
from the available consumer market, especially in this region. The brushes are
either with brush head too large in size or too soft in bristles which are unsuitable
for brushing and cleaning dentures. Dentures soaking effervescent tablets seem
the only choice left and using these alone has been proven to be inconclusive in its
efficiency to totally remove denture plaque and Candida biofilm [16-18]. These
tablets are meant for adjunctive use with mechanical brushing [10, 19-21].
The purpose of this experiment at hand is intended to fill the current gap in
researches on mechanical cleaning method of dental prostheses. The null
hypothesis for this study was set that the use of powered toothbrush to clean
dental prosthesis has no impact on the mean flexural strength and mean surface
roughness value of poly(methyl methacrylate) denture base specimens. These two
parameters of investigation formed the first and second part of a three-part
research experiment, microbiology constituted the third part.
90
The aim of the microbiological experiment was to qualitatively evaluate
the efficacy of battery powered toothbrush for the in vitro removal of Candida
albicans biofilm formed on denture acrylic surfaces.
In principle, Candida albicans can adhere and colonise PMMA surfaces to
form biofilms [22, 23]. Mature biofilms are recognized as matrix-enclosed
microbial population adhering to each other and they characteristically display a
phenotype that is different from planktonic cells, such as increase resistance to
antimicrobial agents and protection from host defences [24].
The purpose of denture hygiene practice is to reduce and to eliminate
microbial biofilm formation. Mechanical cleaning and chemical disinfection have
been recommended for effective removal of denture plaque and Candida biofilms
[19, 25].
The structure of Candida albicans and other yeasts as observed by
scanning electron microscopy (SEM) has been described earlier by Barnes et al.
[26]. SEM micrographs were used to evaluate the effectiveness of denture
cleaning using ultrasonic device compared to effervescent tablets immersion
method [27]. It has been established that it is feasible to use SEM in determining
the morphologic type and distribution of surface coating and microorganisms on
the surface of dentures [28].
SEM images a sample by scanning it with a high-energy beam of electrons,
and it is capable of imaging at a significantly higher resolution than light
microscopes. SEM allows the evaluation of detailed surface topography and
morphology of Candida albicans biofilm at high magnification but involves
91
degradation of the hydrated structural features (e.g. matrix) due to the fixation and
dehydration steps performed during sample preparation [29].
The advantages of SEM examination include a deeper depth of field,
which allows more of a specimen to be in focus at one time compare to light
microscope that only manage relative shallow depth of field. Besides, the SEM
has much higher resolution and higher level of magnification level at thousands of
times. The images produce are strikingly clear which makes SEM most useful for
the examination of Candida biofilm architecture and surface topography in greater
detail. In the current experiment, the absent of Candida cells and biofilm in SEM
micrographs would
confirm their thorough and complete removal effected
through the experimental brushing protocol [27].
In recent years, there has been an increase in the preference of atomic
force microscopy (AFM) in biomaterial imaging due to its advantages over the
other types of spectroscopy such as fluorescence microscopy and confocal
scanning laser microscopy (CSLM). AFM provides not only a three-dimensional
image of a sample but also a three-dimensional surface profile. In the AFM
imaging, the samples do not require any special treatment such as coatings with
metal or carbon, and no drying is required. Additionally, the images can be
obtained in the ambient air or even in a liquid environment [30].
92
2. Materials and methods
2.1 Specimen groups
A total of 244 pieces of PMMA specimens beam were required for this
experiment. Specimens were kept in water storage for 0, 1, 7, 15, 30 and 60 days.
12 PMMA specimens from each test group were subjected to brushing protocol
and 12 specimens from control group were not.
Surface roughness value (Ra) of 12 specimens for all groups was analysed before
brushed and after brushed. Specimens for this test were from the test group of
flexural strength test prior to loading into universal testing machine. Table 1
showed the specimen allocation for this study.
Table 1 Specimen allocation
Day(s) in
Water
Storage
Testing
Parameter
0, 1, 7, 15, 30, 60
Surface Roughness value
Ra (μm)
Flexural strength
(MPa)
Before brushing
S
p
e
c
i
m
e
n
Test
n=12
After brushing
Control
n=12
After brushing
No brushing
93
2.2 Materials
Poly(methyl methacrylate) of the brand Trevalon C (DENTSPLY Ltd. De
Tray Division Weybridge, UK) in liquid monomer and powder polymer forms and
of the heat polymerized type was used for fabrication of PMMA specimens beam.
Stainless steel moulds which can be assemble and disassemble with 6
compartments for specimens were used. (Fig. 2.1 (a) & (b))
(a)
(b)
Fig.2. 1(a) and (b) Stainless steel mould for specimens fabrication.
94
2.3 Specimens fabrication
Heat cured PMMA specimens beam with the dimensions of 45.0 x 6.5 x
4.5 mm were fabricated in a stainless steel mould. The PMMA was mixed at the
powder to liquid ratio of 2.4 : 1.0 and process according to manufacturer’s
instruction and Prince Philip Dental Hospital, Dental Technology Laboratory
heat-cured poly(methyl methacrylate) processing protocol.
Firstly, the stainless steel (SS) moulds were pre-coated with a thin layer of
petroleum jelly (Vaseline®, USA) for easy detachment of the cured specimens. A
small portion of dough was placed on the SS mould and firmly pressed with
fingers at the beginning which showed uneven distribution of pressure and dough
material into mould spaces (Fig. 2.2). Then, incremental packing pressure was
applied to stainless steel moulds range from 1500-4500 psi (105-316 kg·cm-2).
The fastening screws of moulds were progressively tightened at successive trial
packing while the flash of excess dough was squeezed out (Fig. 2.3). This
progressive screw tightening step was to facilitate the pressure release and prevent
a back pressure build-up which could cause porosity due to extreme air-traps
while packing the dough into SS mould. The excess PMMA dough that flowed
out of the mould as flash during pressing was folded back overlay the dough in
the mould after the hydraulic press was released and the cover plate of the mould
uncovered (Fig. 2.4). This step was repeated twice and the fastening screws were
tightened in each successive step. The layers of overlay became thinner with each
successive pressing and compacting which facilitated fabricating more compact
dough specimens (Fig. 2.5).
95
Fig. 2.2 A lumpy dough placed on mould caused uneven pressure distribution.
Fig.2.3 Pressed and tightened fastening screw through sequential steps.
Fig.2.4 Flash folded back overlay mould followed by pressing.
Fig.2.5 Further compaction of thinner fold-over flash and dough.
96
At final pressing, the pressure was maintained for 10 minutes. Each
folding back and overlay of mould with flash of PMMA dough followed by
pressure application would make the dough stage PMMA specimens more
compact where air bubbles included during mixing and packing could be
compressed and expelled. An example of insufficiently compacted dough in
specimens is showed with large quantity of visible air bubbles (Fig. 2.6).
Fig. 2.6 Close up view of specimens removed from mould at 1st press-packing to
show air bubbles at base which appeared as white spots in specimens at dough
stage packing.
The SS moulds were transferred to a spring flask clamp (Ash™, England).
The tightening screw on the clamp was tightened to a torque of 36 N m [31], this
is to ensure sufficient pressure applied to minimized porosities in PMMA
specimens.
97
Fabricated specimens were not subject to surface finish polishing protocol.
Specimens were group assigned to control and test at random selection.
Specimens with excessive porosity throughout or polymerization
shrinkage concentrated on one surface were excluded (Fig. 2.7). The inclusion of
these porous specimens could result in extreme data to be generated from
experiment which would make subsequent erroneous interpretation inevitable.
Specimens that were included in experiment had minimal porosity (Fig. 2.8).
Fig. 2.7 Specimen excluded due to excessive porosities.
Fig. 2.8 An example of specimens without porosity included in experiment.
98
2.4 Polymerization and post processing
Heat curing of PMMA was carried out in a polymerization hot-water bath
(Model 1491, Loborwelt Leleux Gmbh, Germany). The stainless steel moulds
with packed PMMA dough were secured in a spring clamp and placed into the
water bath at room temperature thereafter heated to 100 °C and maintained at this
temperature for 40 minutes. At the end of polymerization, the stainless steel
moulds in clamp were removed from hot-water bath and left on bench to cool
overnight at room temperature.
The moulds were removed from the clamp the following day and
disassembled to retrieve the polymerized specimen beams. Flash and fins of
excess PMMA on specimens were trimmed and removed using sharp blades and
fine silicon carbide paper No. 500 (Struers, Denmark) to render the edges of
specimens even and smooth. This was to ensure a more uniform and
homogeneous measurement of all dimensions.
Each specimen was labelled
accordingly to indicate the date of fabrication, the specimen group and the
specimen number.
99
2.5 Storage condition
Specimens were totally immersed in a plastic container filled with
deionized water covered with plastic wrap (Parafilm®), and stored in a waterjacket incubator (Model 3157, Forma Scientific, Ohio, USA) at a constant
temperature of 37 ⁰C with a 99 % relative humidity. The duration of storage in
deionized water (Milli-Q® RG Ultra-pure water system, Millipore, Massachusetts,
USA) in an incubator before other testing was 1 day, 7 days, 15 days, 30 days and
60 days. Specimens with no water storage conducted were assigned as the control
group for comparison to test groups and it was kept in water storage. Specimens
were tested dry at room temperature after immediate removal from container filled
with deionized water.
100
2.6 Powered toothbrush brushing protocol of specimens
The powered toothbrush used was a generic brand battery powered with
rotating brush head obtained off-the-shelf from a local supermarket, at the
purchased price of USD 5.00. The rotating head measured 13.5 mm in diameter,
with a total of 28 turfs of Tynex®(DuPont, USA) nylon bristles, arranged in
circular pattern of 2 distinct height where the groups centrally placed is slightly
shorter than the peripheral flank. The peripheral group was longer and only lined
in single circular row. Each bristle was approximately 150 µm in diameter and
end-rounded (Fig. 2.9). The peripheral turfs of bristles were trimmed down to
make all bristles same level (Fig. 2.9). This was done carefully using a pair of
sharp
scissor
under
a
microscope
magnification
(20x)
(Fig.
2.9).
Fig. 2.9 Arrow points to periphery bristles trimmed even to central tufts (20x).
This was to ensure more even distribution of applied on surface of
specimen during brushing. The head of the powered brush rotated at a rotational
101
speed of 2540 (±5) rpm measured with a stroboscope (Gr 1538-A, Strobotac,
General Radio, MA., USA).
Test specimens underwent a single focal area of brushing action using
battery powered toothbrush with a rotating brush head at a vertically applied force
of 2 N (equivalent to 204 g) [15], measured at the bristles-specimen interface.
The applied force was in the form of weight placed over the handle of powered
toothbrush, perpendicularly to the surface of the specimen beam to be brushed.
The duration of the brushing was 22 min continuous with 5 ml of deionized water
as the medium.
Brushing station which housed the experimental powered toothbrush and
force gauge was originally conceived, designed and constructed in laboratory (Fig.
2.10). The toothbrush was fixed in the holder of the brushing station, allowing an
alignment of the refillable brush head parallel to the surface of the samples.
Weight placed over the handle of powered toothbrush would register a reading of
applied brushing force of 2 N in force gauge (Model NK-10, AIGU® GTGY
Group Co. Ltd., Hong Kong). The applied force on PMMA specimens was a
constant at 2 N throughout, a screw at the bottom of toothbrush support frame
allowed such a fine control (Fig.2.10).
The specimen holder was designed to hold two specimens at one time and
the brush head was centred on the midpoint of two specimens in line with the
push lever of the force gauge. Thus, the central portion of specimens were
brushed leaving the two tail ends unbrushed (Fig. 2.11). The central area of a
specimen was the point of application of the crosshead of universal testing
machine and the stylus of profilometer used.
102
Fig.2.10 The brushing station, a PMMA beam specimen holder and a force gauge
with weight distributed to maintain stability of the station.
103
Fig. 2.11 Two pieces of specimens fitted into holder with centring marks clearly
mark out for brushing target. Deionized water (5 ml for each 22 min session) was
contained within the specimens’ holder stage.
Each brush head was used for a cumulative time of 66 min before
replacing with another. The purpose was to reduce excessive bristles wear of any
one particular brush head refill. New brush heads refill unpacked from packaging
were subjected to pre-conditioning which involved a single session of 22 min
brushing with water as the medium. This was to age the brand new nylon bristles,
making them less stiff compared to those used brush head refills.
104
2.7 Surface roughness test
The profilometer consists of a stylus that traverses across the surface, and
an amplified trace of the profile is provided in surface roughness value Ra. The Ra
value is the arithmetical average of all deflection of the profile through the mean
sample length, measured in micrometres (µm).
A set of mould was randomly picked up for an analysis of surface
roughness on the surfaces of stainless steel component plates with profilometer
(Surtronic3+, Taylor-Hudson Limited, Leicester, England). This served as
reference to the Ra surface roughness of the specimens. The surface analysis was
set with a cut-off length (or sampling length) of 0.8 mm, and evaluation length of
4.0 mm. Evaluation length is the length over which the values of surface analysis
parameters are assessed. Cut-off length is the length of the reference line used for
identifying the irregularities characterizing the surface.
Measurements were taken for specimens before and after brushing. Three
measurement readings from 3 parallel locations, 1 mm apart were taken on target
spot. The quadrant at 9-12 o’clock position within the semi-circular brushed area
was
earmarked
for
surface
roughness
evaluation
(see:
Fig.
2.12).
Fig. 2.12 Schematic illustration of surface roughness value Ra measurement.
105
2.8 Flexural strength test
The dimensions of specimens were measured using a scientific calliper (±
0.02 mm) with analogue output (Model No. 1143M, Moore & Wright, Sheffield,
England). Three measurement readings were taken for each dimension of length,
width and height prior to the start of 3-point bending test. An average
measurement calculated to be applied for flexural strength calculation.
A universal testing machine (ElectroPuls™ E3000, Instron Industrial
Products, Grove City, PA, USA) was use to performed the 3-point bending test on
the specimens (Fig. 2.13). The span length between 2 supports was 20.0 mm and
the crosshead speed was set at 1.0 mm/min during loading. The load was applied
to fracture specimens into two. Load and deflection were recorded with Console
software (Instron Industrial Products) and the load and deflection curves were
plotted. The maximum load to fracture was applied to formula (1) to calculate the
flexural strength of these PMMA specimens beam:
(1)
Where
σ: Flexural strength in 3-point bending test
𝑭: Maximum load on the load-deflection curve (N)
𝓵: Span length between the supports (mm)
𝒃: Width of the specimen measured (mm)
𝒉: Height of the specimen measured (mm)
106
Fig. 2.13 A PMMA specimen for 3-point bending test set up with a loading wedge
above and two support wedges below. The loading wedge crosshead speed was
set at 1 mm/min and span length between supports was 20.0 mm.
107
2.9 Statistical analysis
Data collected on the mean value and the standard deviation of the flexural
strength and surface roughness value (Ra) were calculated and statistically
analysed using software Predictive Analytics Software (PASW) Statistic 18.0
(Statistical Package for Statistical Science Inc., Chicago, IL, USA). The level of
significant α was set at 0.05. Analysis of data was perform with 2 –way ANOVA
and the Tukey multiple comparison post hoc analysis were perform (p<0.05) to
compare the testing in different water storage days.
108
2.10 Microbiology
2.10.1 Materials and methods
The work scheme of the microbiology experiment was represented in an
illustration (Fig. 2.14). PMMA specimens of the control group for the flexural
strength test which were tested and fractured into two parts were used for this part
of the investigation. The specimens with the dimension 22.0 x 6.5 x 4.5 mm were
trimmed, fitted and arranged into a 6-well polystyrene tissue culture plate
(Iwaki™, Tokyo, Japan) (Fig. 2.15 ). Each well could fit 4 specimen beams, and
they required immobilization to prevent specimens from floating in culture
medium.
Fig. 2.14 Schematic illustration of PMMA specimens cultured with Candida
albicans biofilm experiment.
109
Fig. 2.15 PMMA specimens in 6-well tissue culture plate.
110
2.10.2 Specimen grouping
Specimens for the microbiology experiment were grouped as shown in Table 2.
Table 2 Specimens allocation for microbiology experiment.
Brushing performed with moistened bristles
S
p
e
c
i
m
e
n
Brushed
Duration
(second)
10
20
30
30 with 0.5 g toothpaste
Test
n=2
2
2
2
2
Control
n=2
No brushing performed
Specimens were kept in water-jacket incubator with total immersion in deionized
water at constant temperature of 37 ºC for 30 days before brushing test conducted.
This step was to simulate PMMA in oral environment under constant contact with
saliva in the mouth.
111
2.10.3 Candida albicans cultured biofilm growth on specimens
Sterilization of PMMA specimens was necessary to ensure the
uninterrupted growth of Candida albicans biofilm. For this purpose specimens
were immersed in 0.05 % sodium hypochlorite for three minutes with ultrasonic
agitation. This was followed by washing and rinsing of the specimens with sterile
distilled water for 10 min in ultrasonic water bath. This step was repeated three
times to ensure the total removal of residual sodium hypochlorite on the surfaces
of PMMA specimens.
Standard inoculum for Candida biofilm formation was prepared as
previously described [32]. Subculture of Candida albicans was incubated on
Sabouraud dextrose agar (SDA) (Gibco, Paisley,UK) at 37 ºC overnight. The
Candida albicans subculture was used for the subsequent broth culture. The broth
culture was prepared by inoculate a large loopful (3-5 colonies) of Candida strain
using a sterile wire loop and dissolved in 20 ml liquid medium of yeast nitrogen
base (YNB) ( Difco Laboratory Inc., USA) with 50mM glucose. The broth culture
was kept in a temperature control shaker (Stuart SI500, Bibly Scientific Limited,
Staffordshire, UK) set at 80 rpm and incubated at 37 ˚C for 24 hour. At the end of
24h, the broth culture was then centrifuged at 3500 rpm for 10 min at 37 ºC and
resultant cells pellets were washed twice with phosphate-buffer saline (PBS)
solution. The cells pellets were resuspended in 15 ml of a medium containing
YNB with 100mM glucose. A final suspension of approximately 107 cells/ml of
Candida albicans was prepared through dilution.
The PMMA specimens in the tissue culture plate were immersed in this
final suspension. The plate was placed in a temperature control shaker rotating at
112
80 rpm and incubated for 90 min at 37 ºC, to allow Candida albicans to adhere
onto the PMMA specimens.
Subsequently, the specimens were washed with PBS twice to remove nonadherent cells. Culture medium containing YNB with 100 mM glucose was added
to cover specimens in plate and continued biofilm development in a temperature
control shaker at 80 rpm with incubation temperature at 37 ºC for 48 hours.
At the completion of biofilm development after 48 hours, the culture
medium was poured away and the specimens were washed lightly with PBS
solution twice to remove non-adherent cells.
113
2.10.4 Brushing protocol for Candida albicans coated specimens
The specimens were carefully removed from the culture plate and secured
in the holder of the brushing station, prepared for the brushing test.
A clean new brush head was used for each brushing test. It was necessary
to disinfect the bristles before the test to ensure that the brushed surfaces on
specimens were free from any contamination with microorganisms. The procedure
to disinfect the bristles of brush heads involved immersion into a sodium
hypochlorite solution 6.25 % (Clorox®) and placement in a ultrasonic bath
(BioSonic UC100, Whaledent, West Sussex, UK) for 5 minutes (Fig. 2.16) This
was followed by rinsing the brush heads in sterile distilled water for 15 minutes in
a ultrasonic bath, and this step was repeated three times to ensure a complete rinse
off residual sodium hypochlorite from within bristles in the brush heads.
Fig. 2.16 Disinfection of brush heads and bristles of powered toothbrush.
114
Specimens from the test group were subjected to brushing whereas the
control group was spared intact. Brushing force applied on test specimens was set
at 2 N but the length of time varied. Two pieces of securely positioned specimens
were subjected to brushing at one time (Fig. 2.17).
Fig. 2.17 A close-up view of specimens in a holder with a brush head aligned to
the top surface of two pieces of specimens. The same holder in previous
experiment was modified to accommodate shorter specimens. The green material
is silicone impression material (Genie™, Sultan Healthcare Inc. USA) to hold
specimens in place, the rest is cold-cured acrylic resin. Tightened metal screw
fastened the specimens to provide additional stability.
115
The brushing protocol proposed was set for 10 s, 20 s, 30 s and 30 s with
toothpaste as depicted in Table 3. The control group was not brushed, thus the
biofilm was intact.
Table 3 Brushing protocol
Test Group.
Brushing with moistened bristles
brushes.
10 s
Control group.
20 s
No brushing
30 s
30 s with 0.5g toothpaste
During the brushing procedure, the PMMA specimens with Candida
biofilm and the heads of the powered toothbrush were enclosed within a layer of
cling-wrap to prevent the sputter of microorganism, which could lead to
contamination of surrounding environment. The specimens and the wrap were
later discarded as bio-hazard waste. The used brush heads were then sterilised in
sodium hypochlorite at a concentration of 6.25 % with ultrasonic agitation.
.
116
2.10.5 SEM of Candida albicans coated specimens
Specimens with the attached biofilm from brushed and non-brushed
groups were prepared for scanning electron microscopy (SEM) by fixing them for
2 hours in a 2.5% glutaldehyde (v/v) (BDH Lab. Supplies, UK) and then
dehydrated for 15 minutes at each concentration in a graded ethanol series (70, 85,
95 and absolute alcohol). The dry specimens were mounted on aluminium stubs
and sputter coated with gold in an ion sputter coater (JFC-1100 JEOL, Tokyo,
Japan) and each sample was examined with a Hitachi S-3400N (VP-SEM)
scanning electron microscope at Electron Microscope Unit, Faculty of Medicine,
The University of Hong Kong. Observation was made in high-vacuum mode at 15
kV.
Images of high magnification were taken on the specimens from control
(non-brushed) group and tested (brushed) groups. For the brushed specimens, the
zooming target under the scanning electron microscope was the brushed areas that
need to be scanned and evaluated.
117
3. Results
3.1 Surface roughness
The mean ± standard deviation of the Ra value of surfaces of PMMA
specimens before brushing was 0.29 ± 0.06 μm. The mean of the Ra values after
brushing for 22 min with water was 0.27 ± 0.06 μm (Fig. 3.1), which were
significantly smaller than that of before brushing (p = 0.022). Among the groups
of specimen, the number of days in water storage affect the Ra before and after
brushing differently (p = 0.006) with lowest Ra recorded from specimen groups
after 7 days in water storage compared to no storage. A 2-way ANOVA (Table 4)
was conducted to examine the effect of before-after brushed factor and the
duration of water storage factor on the surface roughness value mean. There did
not appear to be any interaction between the factors of duration in water storage
and before-after brushing (p = 0.826). Tukey analysis revealed that the surface
roughness value of 7 days water storage was significantly different from 0, 15, 30
and 60 days water storage (Table 5, Fig. 3.2).
118
Fig.3.1 Boxplot of mean surface roughness value Ra (µm) for specimen before and
after brushed (p<0.05).
119
Table 4 Result of 2-way ANOVA of surface roughness value of before and after
brushed specimens in different days of water storage.
Dependent Variable: Surface roughness
Type III Sum of
Source
Squares
df
Mean Square
F
Sig.
a
11
.008
2.219
.014
20.230
1
20.230
5.812E3
.000
Storage
.059
5
.012
3.386
.006
Brushed
.019
1
.019
5.319
.022
Storage * Brushing
.008
5
.002
.432
.826
Error
.835
240
.003
Total
21.150
252
.920
251
Corrected Model
Intercept
Corrected Total
.085
a. R Squared = .092 (Adjusted R Squared = .051)
120
Table 5 Result of Tukey post hoc multiple comparison of surface roughness value
Ra with various duration in water storage
121
Mean surface roughness Value Rₐ (µm)
0.4
0.35
Mean Rₐ (µm)
0.3
0.25
0.2
Before Brushing
After Brushing
0.15
0.1
0.05
0
0 day
1 day
7 days
15 days
30days
60 days
No. of days in water storage at 37 ° C
Fig. 3.2 Bar chart of mean surface roughness value Ra (µm) in various duration
of water storage with error bars denoting standard deviation.
122
3.2 Flexural strength.
Table 6 and Fig. 3.3 showed the mean flexural strength of specimen with varying
the water storage condition. The mean from the group of brushed specimen and
underwent 7 day in water storage recorded the lowest value of 87.37 ± 9.23 MPa.
The highest recorded value of 103.72 ± 8.52 MPa was from the brushed specimen
group without water storage. The mean value of specimens from non-brushed
specimen in 15 days water storage group was relatively higher than other storage
day groups.
Table 6 Mean flexural strength results (Test: brushed; Control: non-brushed)
Day(s) in Water
Storage
Specimen
1
2
3
4
5
6
7
8
9
10
11
12
0 day
1 day
7 days
15 days
Control
112.82
128.56
87.53
120.92
120.86
84.66
96.04
86.52
115.20
99.85
103.22
71.71
30 days
60 days
Test
114.68
97.75
91.59
96.77
102.39
97.41
108.08
117.8
97.68
106.1
115.02
99.36
Control
82.78
98.59
110.65
108.62
108.15
99.52
96.18
88.16
81.67
100.8
103.6
94.2
Test
103.54
111.52
88.99
88.18
81.98
104.85
102.87
112.73
119.32
101.55
89.32
93.03
Control
100.29
111.26
105.41
96.79
91.88
68.31
81.33
97.73
80.60
87.07
104.44
83.69
Test
111.03
92.62
88.11
86.48
81.52
86.64
94.10
87.55
74.24
79.33
84.47
82.39
Control
84.38
86.94
80.45
106.48
107.82
95.12
86.61
80.18
97.44
88.45
96.72
74.78
Test
84.78
101.46
83.93
101.33
85.66
87.60
64.36
88.76
84.32
86.91
95.06
91.35
Test
81.22
89.77
105.74
89.69
105.05
104.68
84.93
99.40
109.54
78.50
99.45
103.97
Control
86.90
70.30
80.72
83.81
98.22
92.52
93.67
88.65
88.07
87.68
95.87
74.46
Test
95.72
100.88
89.52
96.44
92.10
97.52
85.48
100.41
98.13
85.92
89.68
92.97
Control
75.80
107.43
87.68
73.51
82.10
82.57
102.23
111.41
89.87
112.04
83.03
80.12
Mean
103.72
Standard deviation 8.52
97.74
9.68
99.82
11.53
92.40
12.25
87.37
9.23
90.44
10.39
87.96 102.32 95.99
9.66 17.62 10.62
86.74
8.36
93.73
5.28
90.65
13.93
123
Mean flexural strength of PMMA specimens
140
*
**
Flexural Strength (MPa)
120
100
80
Test
60
Control
40
20
0
0 day
1 day
7 days
15 days
30 days
No. of days in water storage at 37 °C
60 days
Fig. 3.3 Bar chart of mean flexural strength with error bars represents standard
deviation. Horizontal lines denoting significant difference in means from Tukey
post hoc multiple comparison of days in water storage (p<0.05) (Test=brushed,
Control=non- brushed)
A 2-way ANOVA (Table 7) was conducted to examine the effect of brushing and
water storage on the flexural strength. There was a significant interaction between
the effect of brushing and the factor on number of days in water storage (p =
0.003). There was no significant difference in flexural strength between before
and after brushing the specimens (p = 0.452). There was a significant difference in
the flexural strength among groups of specimens in different days in water storage
(p = 0.006). Therefore, Tukey HSD’s test was performed to compare all
conditions including duration of days in water storage. The flexural strengths of
various days in water storage conditions are summarized in Table 8 and Fig.3.4.
124
Table 7 Result of 2-way ANOVA of flexural strength between before-after
brushing and duration in water storage.
Type III Sum of
Source
Squares
df
Mean Square
F
Sig.
a
11
409.819
3.369
.000
1274482.945
1
1274482.945
1.048E4
.000
Storage
2095.628
5
419.126
3.446
.006
Brushed
69.278
1
69.278
.570
.452
2343.099
5
468.620
3.853
.003
Error
16055.440
132
121.632
Total
1295046.390
144
20563.445
143
Corrected Model
Intercept
Storage * Brushed
Corrected Total
4508.005
a. R Squared = .219 (Adjusted R Squared = .154)
125
Table 8 Result of Tukey post hoc multiple comparison for flexural strength under the
factor duration of water storage
126
Fig. 3.4 Estimated marginal mean of flexural strength.
127
3.3 SEM
Observations under SEM revealed presence of Candida albicans biofilms
on the surfaces of specimens. Specimen without brushing showed presence of
thick biofilm layer consisted of dense network of yeasts, pseudohyphae and
hyphae embedded in a thick EPS layer (Fig. 3.5).
Fig. 3.5 SEM micrograph on a specimen with no brushing, an intact biofilm
showed intricate network of yeast cells and hyphae (x3700).
128
Specimens with surface irregularities and porosities demonstrated the ingrowth of Candida albicans into micro crevices (Fig. 3.6 & Fig. 3.7). These
Candida albicans cells and hyphae were inaccessible to the rotating brush bristles.
Fig. 3.6 Porosities (red arrows) on a PMMA specimen (x500).
Fig. 3.7 Candida albicans cells grew into porosities on a PMMA specimen
(x1000).
129
A distinct brushed area on specimens was visible (Fig. 3.7) compared to
non-brushed specimens (Fig. 3.8) after fixation and dehydration. SEM micrograph
at lower magnification showed the same distinct brushed area (Fig.3.9).
Fig. 3.7 Mark out dash line area brushed for 20 s.
Fig. 3.8 Control group non-brushed with intact biofilm.
Fig. 3.9 The semi-circle dash line area was brushed for 20 s (x32).
130
Candida albicans cells were evident on surface of PMMA specimens
after brushing with powered toothbrush for 10 s under the SEM evaluation
(Fig.3.9).
Fig. 3.9 A specimen from test group after 10 s of brushing performed, cells and
hyphae were found (x500).
The micrographs of specimens after 20 s and 30 s brushing showed present
of scanty number of Candida albicans cells and some crushed cell debris (Fig.
3.11 and Fig. 3.12) compared to 10 s brushing. Less cells debris was observed on
specimen surface after 30 s brushing protocol compare to 20 s brushing (Fig 3.12).
131
Fig. 3.11 A specimen after 20 s of brushing showed the presence of Candida cells
and some crushed cell debris (x1000).
Fig.3.12 A test specimen after 30 s of brushing performed (x500).
132
Fig. 3.11 illustrated the SEM micrograph taken of specimen after 30 s
brushing with toothpaste. It revealed the total absent of yeast cell and present of
filler particles from toothpaste which were evenly distributed. The present of filler
particles were due to no rinsing procedure in the brushing protocol.
Fig.3.13 A specimen was brushed for 30 s with toothpaste showed presence of
well scattered filler particles (red arrows) and absence of Candida cell (x2000).
Evaluation of the clean PMMA surface of specimen without Candida
albicans growth revealed the topography of PMMA surfaces were even and
uniform with linear layers stacking pattern. Some layers appeared more extruded
(Fig.3.13 and Fig. 3.15). This was a contrast to the polished surface of PMMA
with fine silicon carbide paper No. 500 (Struers, Denmark) as shown in Fig.3.14
which appeared relatively rough surface topography with innumerable overt
scored lines.
133
Fig. 3.14 Contrast between polished and unpolished surface. Area below red
dotted line was polished with No. 500 silicon carbide paper, area immediately
above red dotted line was manifestation of surface shrinkage of PMMA which was
indented and preclude polishing.
Fig. 3.15 Clean surface of PMMA specimen without Candida albicans biofilm.
134
4. Discussion
4.1 Surface roughness
Surface roughness is associated with the surface finishing of a dental
prosthesis. The highly polished surface of PMMA dental prosthesis has a low
surface roughness value Ra compared to a coarse finish surface. In this experiment
the specimens of PMMA were not subjected to post fabrication finishing and
polishing. Thus, there was an observation of surfaces of PMMA specimens after
brushing protocol that showed a decrease in the surface roughness value Ra. This
might have been due to the polishing phenomena of the nylon bristles on the
surfaces of polymer. The same observation was reported by Harrison et al.[15].
This would suggest that the rotating motion of nylon bristles of the experimental
powered toothbrush on the surface of PMMA, did not pose an increased risk of
abrading the surface during brushing action, when water was used as medium.
However, the favourable effect may be lost in a situation where surface of PMMA
deteriorates in service condition and abrasion effect would be probable. When
toothpaste was used during brushing, the surface roughness of PMMA acrylic
resin increased due to the abrasive action of filler particles in a toothpaste [33].
The experiment with Candida albican showed that the removal of
Candida biofilm was more thorough when some toothpaste was added during
brushing. It is noteworthy that toothpastes may not be suitable in daily denture
cleaning routine with a powered toothbrush. Brushing with a non-abrasive liquid
medium such as soap and mouth rinses would avoid the risk of abrasion, but the
Candida biofilm would not be thoroughly removed. The supplementary chemical
cleaning method such as hypochlorite and chlorhexidine immersion might be
necessary in order to totally eliminate Candida biofilms [16].
135
The force of 2 N equivalents to a load of 204 g applied on the brushed
area throughout the experiment appeared sufficient to remove biofilm and did not
cause visible damage to the PMMA surfaces. Previous research has demonstrated
that the force applied with powered toothbrush in dental hygiene practise was
lower compared to using manual tooth brushing [34]. The forces recorded for
powered toothbrushes were in the range of 80-190 g (as load) and in the 250 g
range when manual toothbrushes were used [34]. The actual force applied in
individual varies greatly during the cleaning process, correlation between cleaning
forces applied intraorally and extraorally may be an overinterpretation. However,
it was reasonable not to go far beyond these ranges of force indicated in previous
researches [35-37]. It would be of interest to research into the forces applied by
individual when performing denture brushing with a powered toothbrush.
The specimens were not subjected to polish finishing, this might have
contributed to a higher surface roughness value before brushing. However, a
report stated that surface roughness was influenced to the greatest extent by
finishing and polishing procedure and to a lesser extent by acrylic resin material
itself [38]. The SEM evaluation of a PMMA specimen in present study confirmed
this, as shown in Fig. 3.14 and Fig. 3.15. Therefore, it was reasonable to avoid
adding this variable into the current experiment by not polishing finished PMMA
specimen beams. It was interesting to note that current study with unpolished
finish specimens showed a reduction in surface roughness value after brushing
whereas others studies reported no change [15, 39, 40] with polished finish
PMMA specimens, when brushing was carried out with water only. This
phenomena appeared to be worthy of further examination.
136
4.2 Flexural strength
The data from this experiment showed that the PMMA beams after 7 days
in water storage would experience a decline in flexural strength compared to no
water storage. Specimens in 15, 30 and 60 days of water storage produced
relatively constant mean flexural strength. This may be attributed to the plasticiser
effect of water molecules acting on PMMA polymer molecules creating a change
in strength of the bulk material. This plasticiser influence started from the
beginning until it reached a plateau at about the day 15 in water storage.
There seemed to be little agreement as to how long PMMA polymers
should be immersed prior to their mechanical testing. Despite the international
standard guidelines points to 50 hours of water immersion [41], Takahashi et al.
[42] suggested that the equilibrium strength of some denture polymers may well
exceed 30 days. Although they recommend a 4-month water immersion protocol,
the water sorption mainly occurs during the initial 14 days [43]. In the study by
Chow et al. water sorption saturation was attained about 30 days after immersion
in water at a constant temperature of 37 °C and the linear expansion was 0.45 %
[44].
The factor on effect of temperature between control group and test group
was not convincingly accounted for in the present experiment. The specimens
from control group which had not undergone storage in water were kept exposed
to room temperature whereas the test groups were kept at 37 °C. An assumption
was made that the influence of the difference would be negligible; on the other
hand, this significance might have been overlooked.
137
The effect and influence of brushing using a powered toothbrush on
PMMA specimens beam seemed inconclusive from the findings gathered in the
current experiment. There was uncertainty appeared during the data analysis (Fig.
3.4) where the lines plot exhibited confusing pattern of intersection, making
meaningful interpretation of interaction among factors impossible. The hypothesis
accounted for this uncertainty might be due to other confounding factors which
failed to be recognized. Another possibility might be the experimental set-up was
not sensitive enough to detect minor differences in mean flexural strength. The
parameter of surface microhardness might seem to be a viable alternative to be
investigated [45] after having subjected the polymer surface to powered
toothbrush brushing.
Although the same material was used, separate batches of specimens were
fabricated. This might account for some differences between these sample groups
during testing for flexural strength as well as surface roughness value.
138
4.3 SEM
SEM examination of specimens revealed the presence of thick layers of
matured Candida albicans biofilm on surfaces of PMMA for non-brushed
specimens. The biofilm consisted of intricate networks of yeast cells and hyphae
deeply embedded into porosities, cracks, and crevices of the PMMA specimens.
Brushing with a powered toothbrush for 10 s, 20 s, 30 s and 30 s plus
using some toothpaste could not remove the Candida from the porosities.
However, on more even surfaces where irregularities were absent, all protocols of
brushing showed the thick layer of Candida albicans biofilm was cleanly brushed
away. The result of the present study was consistent with Harrison et al. [15].
They demonstrated that powered toothbrush cleaning could remove Candida
biofilm when observed under SEM evaluation.
SEM micrographs showed that Candida albicans biofilm was removed
from PMMA specimen surfaces with different brushing duration experimented.
More efficient removal was seen with 30 s compared to 10 s brushing duration.
The removal was not totally complete for brushing time especially in 10 s and 20
s, since some cells and cell fragments remained attached on the
surface
irregularities of specimens. The reason for this might be that the Candida albicans
cells were shielded away from the movement of bristles during brushing due to
the uneven surface topography in microscopic scale. Another reason postulated
could be the bristles maintained no contact with the surface of PMMA specimens
during brushing. This condition of non-contact exited because assumption was
made that when the 2 N forces was applied to the powered toothbrush, the
pressure was equally loaded and evenly distributed to all bristles in the brush head
onto the target brushed area. This was certainly not the case in such an experiment
139
set-up. Human error might have contributed to this, where the seating of
specimens into the holder and lowering the brush head was performed without any
aid of precision control and monitoring, it was entirely carried out under human
visual judgement that was prone to error [46, 47].
There was uncertainty concerned the re-deposition of Candida albicans
cells and cell debris back to surface irregularities during brushing after their
removal. This could contribute to erroneous interpretation of result in current
study. A constant supply of running stream of water to rinse off dislodged cells
and debris might have allayed this concern.
Brushing with some added toothpaste seemed to produce a more
favourable result as shown in the scanning electron micrographs (Fig. 3.13) when
compared to brushing alone (Fig.3.12). No remains of Candida cell were found
during SEM observation base on this brushing protocol. However, previous
studies have confirmed that the surfaces of PMMA specimens were abraded
leading to increased surface roughness especially when toothpaste was added and
the brushing time was extended [33, 36, 48]. Thus, increased surface roughness
would lead to increased denture plaque retention and microorganism
accumulation on the denture surfaces [49], as well as to promote the initial
adhesion of microorganism [50].
In view of the facts presented, the mechanical cleaning method using
powered toothbrush requires supplementary chemical or other cleaning methods
in order to achieve optimal level of denture hygiene [10, 20].
Powered toothbrush offers the advantages of an increased ease and
efficiency, especially for the frail elderly who may have lost some degree of
manual dexterity [5, 6, 51]. A powered toothbrush is intended as one device for
140
two purposes; cleaning dentures extraorally and brushing the teeth intraorally.
Other methods of cleaning dental prostheses, such as chemical soaking, could not
measure up to powered toothbrush in this unique characteristic of both intra- and
extraoral applications.
However, extrapolating findings from current laboratory investigations to
clinical outcome is always difficult and must be exercised with caution.
Evaluation on biofilm removal may be expected to show differences in results
from clinical use.
141
5. Conclusion
Within the limit of these experiments the following conclusion may be made:
1. A battery powered toothbrush with a simple mono direction rotational
movement could be a suitable device for hygiene maintenance of acrylic resin
dental prostheses.
2. The results suggested that surfaces of PMMA specimens after brushing with a
powered toothbrush demonstrated no adverse effect in flexural strength and no
increase in surface roughness value (Ra) when water was used as medium.
3. In vitro Candida albicans biofilm cultured on surfaces of PMMA specimens
can be removed by brushing with powered toothbrush.
4. Further research is required to expand and redefine the experiments.
142
Appendix
1. Surface Roughness Test
Table 9 Raw data of surface roughness value Ra for PMMA specimens
Water storage
Brushed
0 day
1 day
7 days
15 days
30 days
60 days
No.
Before
After
Before
After
Before
After
Before
After
Before
After
Before
After
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
0.22
0.26
0.28
0.38
0.26
0.22
0.26
0.30
0.30
0.32
0.34
0.38
0.32
0.38
0.26
0.26
0.24
0.20
0.36
0.36
0.24
0.26
0.24
0.32
0.28
0.34
0.32
0.24
0.34
0.30
0.28
0.30
0.28
0.38
0.32
0.36
0.30
0.20
0.22
0.28
0.26
0.34
0.22
0.30
0.34
0.32
0.32
0.18
0.36
0.28
0.20
0.40
0.38
0.26
0.26
0.24
0.36
0.20
0.32
0.24
0.36
0.26
0.18
0.40
0.28
0.16
0.32
0.32
0.24
0.30
0.32
0.16
0.26
0.26
0.20
0.26
0.24
0.42
0.38
0.32
0.30
0.26
0.24
0.20
0.26
0.24
0.26
0.28
0.20
0.20
0.28
0.22
0.28
0.34
0.30
0.18
0.28
0.26
0.20
0.30
0.34
0.32
0.28
0.18
0.34
0.22
0.24
0.30
0.18
0.22
0.22
0.24
0.24
0.24
0.22
0.20
0.18
0.30
0.26
0.20
0.22
0.30
0.24
0.24
0.20
0.32
0.32
0.28
0.24
0.30
0.24
0.34
0.32
0.30
0.30
0.22
0.28
0.32
0.22
0.38
0.32
0.36
0.26
0.30
0.36
0.24
0.34
0.22
0.34
0.24
0.26
0.26
0.28
0.34
0.26
0.26
0.22
0.32
0.36
0.22
0.24
0.24
0.34
0.26
0.32
0.28
0.30
0.32
0.26
0.30
0.34
0.22
0.34
0.20
0.32
0.32
0.30
0.38
0.28
0.34
0.24
0.32
0.30
0.32
0.36
0.34
0.32
0.32
0.32
0.24
0.22
0.28
0.20
0.32
0.26
0.22
0.24
0.28
0.24
0.36
0.28
0.26
0.38
0.36
0.30
0.32
0.34
0.36
0.26
0.20
0.30
0.26
0.42
0.38
0.42
0.24
0.14
0.20
0.16
0.32
0.36
0.32
0.26
0.32
0.22
0.24
0.40
0.38
0.42
0.38
0.30
0.16
0.22
0.24
0.36
0.30
0.16
0.14
0.32
0.24
0.30
0.38
0.26
0.24
0.30
0.26
0.28
0.42
0.30
0.40
0.32
0.24
Mean
0.29
0.29
0.28
0.28
0.26
0.24
0.30
0.28
0.31
0.28
0.31
0.28
Standard Deviation
0.06
0.05
0.07
0.07
0.05
0.04
0.05
0.04
0.04
0.06
0.09
0.08
R
a
(
µ
m
)
1.1 Surface roughness statistical output
Between-Subjects Factors
Days in Water Storage
Before and After Brushed
Value Label
N
0
0 day
42
1
1 day
42
2
7 days
42
3
15 days
42
4
30 days
42
5
60 days
42
0
Before Brushed
126
1
After Brushed
126
143
Before and After Brushed
Surface roughness
Before Brushed
Statistic
Mean
.2919
95% Confidence Interval for Lower Bound
.2811
Mean
Upper Bound
.2923
Median
.3000
.004
Std. Deviation
After Brushed
.00548
.3027
5% Trimmed Mean
Variance
Std. Error
.06146
Minimum
.14
Maximum
.42
Range
.28
Interquartile Range
.10
Skewness
-.110
.216
Kurtosis
-.585
.428
Mean
.2748
.00522
95% Confidence Interval for Lower Bound
.2644
Mean
Upper Bound
.2851
5% Trimmed Mean
.2742
Median
.2600
Variance
Std. Deviation
.003
.05863
Minimum
.14
Maximum
.42
Range
.28
Interquartile Range
.08
Skewness
Kurtosis
.224
.216
-.262
.428
144
Descriptive Statistics
Dependent Variable:Surface roughness
Days in
Water
Before and After
Storage
Brushed
Mean
Std. Deviation
N
0 day
Before Brushed
.2924
.05744
21
After Brushed
.2933
.04662
21
Total
.2929
.05167
42
Before Brushed
.2848
.06809
21
After Brushed
.2781
.07040
21
Total
.2814
.06849
42
Before Brushed
.2638
.05162
21
After Brushed
.2371
.03964
21
Total
.2505
.04742
42
Before Brushed
.2971
.04703
21
After Brushed
.2800
.04472
21
Total
.2886
.04615
42
Before Brushed
.3067
.04487
21
After Brushed
.2819
.05582
21
Total
.2943
.05157
42
Before Brushed
.3067
.08610
21
After Brushed
.2781
.07561
21
Total
.2924
.08132
42
Before Brushed
.2919
.06146
126
After Brushed
.2748
.05863
126
Total
.2833
.06056
252
1 day
7 days
15 days
30 days
60 days
Total
Levene's Test of Equality of Error Variances
a
Dependent Variable:Surface roughness
F
df1
3.016
df2
11
Sig.
240
.001
Tests the null hypothesis that the error variance of
the dependent variable is equal across groups.
a. Design: Intercept + Storage + Brushed +
Storage * Brushed
145
Estimated Marginal Means
1. Days in Water Storage
Dependent Variable:Surface roughness
Days in
95% Confidence Interval
Water
Storage
Mean
Std. Error
Lower Bound
Upper Bound
0 day
.293
.009
.275
.311
1 day
.281
.009
.263
.299
7 days
.250
.009
.233
.268
15 days
.289
.009
.271
.307
30 days
.294
.009
.276
.312
60 days
.292
.009
.274
.310
2. Before and After Brushed
Dependent Variable:Surface roughness
95% Confidence Interval
Before and After
Brushed
Mean
Std. Error
Lower Bound
Upper Bound
Before Brushed
.292
.005
.282
.302
After Brushed
.275
.005
.264
.285
146
3. Days in Water Storage * Before and After Brushed
Dependent Variable:Surface roughness
Days in
95% Confidence Interval
Water
Before and After
Storage
Brushed
0 day
Before Brushed
.292
.013
.267
.318
After Brushed
.293
.013
.268
.319
Before Brushed
.285
.013
.259
.310
After Brushed
.278
.013
.253
.303
Before Brushed
.264
.013
.238
.289
After Brushed
.237
.013
.212
.263
Before Brushed
.297
.013
.272
.323
After Brushed
.280
.013
.255
.305
Before Brushed
.307
.013
.281
.332
After Brushed
.282
.013
.257
.307
Before Brushed
.307
.013
.281
.332
After Brushed
.278
.013
.253
.303
1 day
7 days
15 days
30 days
60 days
Mean
Std. Error
Lower Bound
Upper Bound
Tests of Normality
a
Before and
After Brushed
Surface
Before
roughness
Brushed
After Brushed
Kolmogorov-Smirnov
Statistic
df
Sig.
Shapiro-Wilk
Statistic
df
Sig.
.105
126
.002
.981
126
.071
.115
126
.000
.980
126
.064
a. Lilliefors Significance
Correction
147
Homogeneous Subsets
Surface roughness
Tukey HSD
Days in
Subset
Water
Storage
N
1
2
7 days
42
.2505
1 day
42
.2814
15 days
42
.2886
60 days
42
.2924
0 day
42
.2929
30 days
42
.2943
Sig.
.159
.2814
.918
Means for groups in homogeneous subsets are
displayed.
Based on observed means.
The error term is Mean Square(Error) = .003.
Profile Plots
Fig. 6.1 Line graph of estimated marginal mean in various days of water storage
148
2. Flexural Strength Test statistical output
Between-Subjects Factors
Days in water storage
Before and After Brushed
Value Label
N
0
0 day
24
1
1 day
24
2
7 days
24
3
15 days
24
4
30 days
24
5
60 days
24
0
Before brush
72
1
After Brushed
72
Tests of Normality
a
Days in water
storage
Flexural
strength
Kolmogorov-Smirnov
Statistic
df
Shapiro-Wilk
Sig. Statistic
df
Sig.
0 day
.106
24 .200
*
.972
24
.726
1 day
.091
24 .200
*
.980
24
.903
7 days
.186
24
.032
.933
24
.112
15 days
.158
24
.127
.943
24
.190
30 days
.102
24 .200
*
.976
24
.821
60 days
.088
24 .200
*
.979
24
.876
a. Lilliefors Significance
Correction
*. This is a lower bound of the true significance.
149
Test of Homogeneity of Variance
Levene
Statistic
Flexural strength
df1
df2
Sig.
Based on Mean
2.004
5
138
.082
Based on Median
1.604
5
138
.163
1.604
5
104.878
.166
2.005
5
138
.082
Based on Median and with
adjusted df
Based on trimmed mean
150
Descriptive Statistics
Dependent Variable:Flexural strength
Days in
water
Before and After
storage
Brushed
Mean
0 day
Before brush
97.7433
9.67561
12
After Brushed
1.0374E2
8.55863
12
Total
1.0074E2
9.44458
24
Before brush
92.4000
12.52819
12
After Brushed
99.8233
11.53115
12
Total
96.1117
12.37070
24
Before brush
90.4475
10.39052
12
After Brushed
87.3733
9.23938
12
Total
88.9104
9.74305
24
1.0232E2
17.62734
12
After Brushed
87.9600
9.66076
12
Total
95.1421
15.71840
24
Before brush
86.7392
8.36010
12
After Brushed
95.9950
10.65278
12
Total
91.3671
10.49042
24
Before brush
90.6492
13.93243
12
After Brushed
93.7308
5.28456
12
Total
92.1900
10.42450
24
Before brush
93.3839
13.10185
72
After Brushed
94.7711
10.81627
72
Total
94.0775
11.99168
144
1 day
7 days
15 days
30 days
60 days
Total
Before brush
Std. Deviation
N
151
Levene's Test of Equality of Error
Variances
a
Dependent Variable:Flexural strength
F
df1
2.729
df2
11
Sig.
132
.003
Tests the null hypothesis that the error
variance of the dependent variable is equal
across groups.
a. Design: Intercept + Storage + Brushed +
Storage * Brushed
Tests of Between-Subjects Effects
Dependent Variable:Flexural strength
Type III Sum of
Source
Squares
df
Mean Square
F
Sig.
a
11
409.819
3.369
.000
1274482.945
1
1274482.945
1.048E4
.000
Storage
2095.628
5
419.126
3.446
.006
Brushed
69.278
1
69.278
.570
.452
2343.099
5
468.620
3.853
.003
Error
16055.440
132
121.632
Total
1295046.390
144
20563.445
143
Corrected Model
Intercept
Storage * Brushed
Corrected Total
4508.005
a. R Squared = .219 (Adjusted R Squared = .154)
152
Estimated Marginal Means
1. Days in water storage
Dependent Variable:Flexural strength
Days in
95% Confidence Interval
water
storage
Mean
Std. Error
Lower Bound
Upper Bound
0 day
100.744
2.251
96.291
105.197
1 day
96.112
2.251
91.659
100.565
7 days
88.910
2.251
84.457
93.364
15 days
95.142
2.251
90.689
99.595
30 days
91.367
2.251
86.914
95.820
60 days
92.190
2.251
87.737
96.643
2. Before and After Brushed
Dependent Variable:Flexural strength
95% Confidence Interval
Before and After
Brushed
Mean
Std. Error
Lower Bound
Upper Bound
Before brush
93.384
1.300
90.813
95.955
After Brushed
94.771
1.300
92.200
97.342
3. Days in water storage * Before and After Brushed
Dependent Variable:Flexural strength
Days in
95% Confidence Interval
water
Before and After
storage
Brushed
0 day
Before brush
97.743
3.184
91.446
104.041
After Brushed
103.744
3.184
97.446
110.042
Before brush
92.400
3.184
86.102
98.698
After Brushed
99.823
3.184
93.526
106.121
Before brush
90.447
3.184
84.150
96.745
After Brushed
87.373
3.184
81.076
93.671
Before brush
102.324
3.184
96.026
108.622
After Brushed
87.960
3.184
81.662
94.258
Before brush
86.739
3.184
80.441
93.037
After Brushed
95.995
3.184
89.697
102.293
1 day
7 days
15 days
30 days
Mean
Std. Error
Lower Bound
Upper Bound
153
60 days
Before brush
90.649
3.184
84.351
96.947
After Brushed
93.731
3.184
87.433
100.029
Multiple Comparisons
Flexural strength Tukey HSD
95% Confidence
Interval
(I) Days in water
(J) Days in water
storage
storage
0 day
1 day
7 days
15 days
30 days
Std.
(I-J)
Error
Sig.
Upper
Bound
Bound
13.8395
3.18371 .004
2.6260
21.0407
15 days
5.6017 3.18371 .495
-3.6057
14.8090
30 days
9.3767
3.18371 .043
.1693
18.5840
60 days
8.5538 3.18371 .085
-.6536
17.7611
0 day
-4.6321 3.18371 .693
-13.8395
4.5753
7 days
7.2012 3.18371 .217
-2.0061
16.4086
15 days
.9696 3.18371 1.000
-8.2378
10.1770
30 days
4.7446 3.18371 .671
-4.4628
13.9520
60 days
3.9217 3.18371 .821
-5.2857
13.1290
3.18371 .004
-21.0407
-2.6260
1 day
-7.2012 3.18371 .217
-16.4086
2.0061
15 days
-6.2317 3.18371 .372
-15.4390
2.9757
30 days
-2.4567 3.18371 .972
-11.6640
6.7507
60 days
-3.2796 3.18371 .907
-12.4870
5.9278
0 day
-5.6017 3.18371 .495
-14.8090
3.6057
1 day
-.9696 3.18371 1.000
-10.1770
8.2378
7 days
6.2317 3.18371 .372
-2.9757
15.4390
30 days
3.7750 3.18371 .843
-5.4324
12.9824
60 days
2.9521 3.18371 .939
-6.2553
12.1595
0 day
4.6321 3.18371 .693
Lower
-4.5753
7 days
1 day
Mean Difference
11.8333
-11.8333
*
*
*
*
0 day
-9.3767
3.18371 .043
-18.5840
-.1693
1 day
-4.7446 3.18371 .671
-13.9520
4.4628
7 days
2.4567 3.18371 .972
-6.7507
11.6640
15 days
-3.7750 3.18371 .843
-12.9824
5.4324
60 days
-.8229 3.18371 1.000
-10.0303
8.3845
154
60 days
0 day
-8.5538 3.18371 .085
-17.7611
.6536
1 day
-3.9217 3.18371 .821
-13.1290
5.2857
7 days
3.2796 3.18371 .907
-5.9278
12.4870
15 days
-2.9521 3.18371 .939
-12.1595
6.2553
30 days
.8229 3.18371 1.000
-8.3845
10.0303
Based on observed means.
The error term is Mean Square(Error) = 121.632.
*. The mean difference is significant at the .05 level.
Homogeneous Subsets
Flexural strength
Tukey HSD
Subset
Days in water storage
N
7 days
24
88.9104
30 days
24
91.3671
60 days
24
92.1900
92.1900
15 days
24
95.1421
95.1421
1 day
24
96.1117
96.1117
0 day
24
Sig.
1
2
1.0074E2
.217
.085
Means for groups in homogeneous subsets are displayed.
Based on observed means.
The error term is Mean Square(Error) = 121.632.
155
Fig 6.2 Box plot of flexural strength in various days in water storage
156
Table 10 Surface roughness value of SS mould-plates
Mean Ra (µm) on surfaces of SS mould
Plate label
Side A
Side B
B3
0.30
0.40
B5
0.29
0.30
B7
0.26
0.35
B9
0.24
0.23
B11
0.18
0.28
3. Images from experiment
Fig. 6.3 Disassembled stainless steel mould with fasten screw shown
157
Fig. 6.4 Fracture surfaces of specimen, solid black arrows denote tension areas,
red arrows point to compression area (x10)
Fig. 6.5 Black arrow points to area brushed for 22 min with brand new brush
head showing noticeable scratch marks, blue arrow points to target area (dash
line) brushed for 22 min with aged bristles, no scratches observed.
158
Fig. 6.6 Processed specimen beams after removal from mould shows surface
porosities on top side (vertically displayed) and the underside of specimens had
no porosity (horizontally arranged)
Fig. 6.7 Hydraulic press and spring clamp with screw knob
159
Fig. 6.8 Generic brand battery powered toothbrush used in current experiment
Fig. 6.9 Candida albicans of matured biofilm (x7000)
160
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