TESTS FOR BIOCOMPATIBILITY OF

DENTAL MATERIALS

PRESENTED BY:
DR. DEVANSHI SHARMA
MDS II YEAR
Biocompatibility

• It is the ability of a material to elicit an appropriate

biological response in a given application in the body.
(Craig)

• Biocompatibility is not a property of just a material,

but rather a property of how a material reacts with
its environment. (Phillip’s)
• The ability of a biomaterial to perform its desired
function with respect to a medical (or dental)
therapy, without eliciting any undesirable local or
systemic effects in the recipient or beneficiary of that
therapy, but generating the most appropriate
beneficial cellular or tissue response in that specific
situation, and optimizing the clinically relevant
performance of that therapy.
(Williams 2008)
 DEFINING THE USE OF A MATERIAL

• There are several factors that must be considered

when trying to measure the biological response of a
material.
• The most important factors include
 Location of material
 The duration of material in the body
 Stresses placed on material

Phillips’ –science of dental materials 11th ed.

Location of material

• The location of a material is important to its overall

biological response.
• In general materials that communicate through the
epithelium or lie completely beneath it will need
closer scrutiny when assessing the biological
response than materials that do not penetrate the
epithelium. Similarly, materials that penetrate tooth
enamel will need more scrutiny than materials than
do not.

Phillips’ –science of dental materials 11th ed.

The duration of material in the body

• The duration of the material in the body is important to

the biological response.
• The duration of the presence of a material is an important
factor, because many interactive effects between the body
and material take some time to develop.
• In general, the most stringent tests to measure
biocompatibility are required for materials that are
present for the longest times.
• Long durations give sufficient time for the material to
affect the body and for the body to affect the material in
many complex ways.
Phillips’ –science of dental materials 11th ed.
Stresses placed on material

• stresses placed on the material are important to the

biological response, these stresses may be physical,
chemical, or thermal in nature.
• Short-term, long-term, and fatigue stresses all need
to be considered when assessing the effect of stress
on the biological performance of material.

Phillips’ –science of dental materials 11th ed.

 HISTORICAL BACKGROUND

• Although the concept of the ethical treatment of

patients extends back to the time of Hippocrates
(460-377 BC.), the idea that new dental materials
must be tested for safety and efficacy before clinical
use is much more recent.
• As late as the mid 1800s,dentists tried new materials
for the first time by putting them into patients‘
mouths.

Gottfried schmalz, Biocompatibility of

dental materials, 3rd ed,2009
• Many exotic formulations were used. For example,
Fox developed a "fusible metal“ that consisted of
bismuth, lead, and tin, which he melted and poured
into the cavity preparation at a temperature of
approximately 100˚C.
• Even G.V. Black used patients to test many of his new
ideas for restorative materials, such as early
amalgams.

Gottfried schmalz, Biocompatibility of

dental materials, 3rd ed,2009
• The current philosophy about testing the biological
properties of dental materials in a systematic way
evolved in the 1960s as the need to protect patients
became politically acute and as the number of new
materials increase.
• The concept of protecting the patient as a research
subject is only 40 to 50 years old, and many of the
regulations and ethics in this area still being
challenged and defined today.
Gottfried schmalz, Biocompatibility of
dental materials, 3rd ed,2009
• 1963 = ADA formulated a county for the safety
and biocompatibility of dental materials.
• 1972= A document for biocompatibility test
recommended and“ Standard Practices for
Biological Evaluation of Dental Materials” was
published.
• 1979 = Above document was revised and republished
as document no. 41.
• 1984 = A similar document was produced &
published by F.D.I.
Gottfried schmalz, Biocompatibility of
dental materials, 3rd ed,2009
• Current Organization that regulate these
biocompatibility testings are :
 Food and Drug Administration (FDA—
www.fda.gov)
 American National Standards Institute (ANSI—
www.ansi.org),
 American Dental Association (ADA—
www.ada.org)
 International Organization for Standardization
(ISO—www.iso.ch).
Gottfried schmalz, Biocompatibility of
dental materials, 3rd ed,2009
 BIOCOMPATIBILITY DEPENDS UPON:

– Condition of host.
– Properties of material.
– Context in which the material is used

Gottfried schmalz, Biocompatibility of

dental materials, 3rd ed,2009
 ADVERSE EFFECTS FROM DENTAL MATERIAL

1. Toxicity
2. Inflammation
3. Allergy
4. Mutagenicity
5. Immunotoxicity
6. Cytotoxicity

Gottfried schmalz, Biocompatibility of

dental materials, 3rd ed,2009
TOXICITY

• The first screening test.

• Materials may be
capable of releasing
substances into a
patient's body, and
release of certain
substances in
adequate amounts can
cause overt toxicity.
Gottfried schmalz, Biocompatibility of
dental materials, 3rd ed,2009
INFLAMMATION

• 2nd fundamental type of biological response to a

material.
• The inflammatory response is complex as it
involves the activation of host's immune system .
• Precedes toxicity.

Gottfried schmalz, Biocompatibility of

dental materials, 3rd ed,2009
ALLERGY

• Occurs when the body

specifically recognizes a
material as foreign and reacts
disproportionately to the
amount of material present.
• The reaction typically involves
T and B lymphocytes and
monocytes or macrophages.

Gottfried schmalz, Biocompatibility of

dental materials, 3rd ed,2009
• In 1963, a new classification scheme was designed by
Philip Gell and Robin Coombs that described four
types of hypersensitivity reactions, known as Type I
to Type V hypersensitivity
• Type I – Anaphylactic
• Type II – Cytotoxic
• Type III – Immunocomplex
• Type IV – Delayed hypersensitivity
• Type V – Serum sickness
 ALLERGIC RESPONSE

a substance as
foreign (Host Disproportionate to
specific) Individuals Dose independent. the amount of
immune system offending substance
recognizes
MUTAGENECITY

 When the components of a material alter the base-

pair sequences of the DNA in cells. These alterations
are termed mutations.
 Metal ions from dental materials such as nickel,
copper, beryllium are known MUTAGEN.
 Root canal sealers , Resin-based materials have
also been identified as having some mutagenic
potential.

Gottfried schmalz, Biocompatibility of

dental materials, 3rd ed,2009
IMMUNOTOXICITY
• Immunotoxicity is defined as adverse effects on the
functioning of the immune system that result from
exposure to chemical substances
• Principle: Small alterations in cells of the immune
system by materials can have significant biological
consequences.
• Immunotoxicity may result from a material causing either
an increase or decrease in cellular function.

Gottfried schmalz, Biocompatibility of

dental materials, 3rd ed,2009
CYTOTOXICITY

• Cytotoxicity is the quality of being toxic to cells.

Examples of toxic agents are a chemical substance,
an immune cell or some types of venom (e.g. from
the puff adder or brown recluse spider).

Gottfried schmalz, Biocompatibility of

dental materials, 3rd ed,2009
 Measuring the biocompatibility

• Dental materials may result in damage to various

tissues. Therefore, a great variety of different test
methods are applied to evaluate the risk of such
damage to ensure material compatibility prior to
market launch.
• common test methods are:
 In vitro tests
 Animal tests
 Usage tests
Gottfried schmalz, Biocompatibility of
dental materials, 3rd ed,2009
 ADA Recommendations for biomaterials
Five types of materials for evaluation
• Type I- materials that may contact parts of body
other than oral cavity
• Type II- materials contacting mucous membrane
of oral cavity
• Type III- materials affecting health of pulp or
adjacent tissue
• Type IV- pulp space filling materials
• Type V- materials affecting hard tissues of the
teeth
Gottfried schmalz, Biocompatibility of
dental materials, 3rd ed,2009
 In vitro tests
• Tests are done in test tube, cell culture dish, or
otherwise outside a living organism.

In vitro tests
Indirect
Direct tests
tests
• Direct tests: material contacts the cell system
without barrier.
• Direct tests can be further subdivided into
 Those in which the material is physically present with
the cells
 Extract from the material contact the cell system
• Indirect tests: when there is a barrier of some sort
between the material and the cell system.
 Agar overlay method
 Millipore filter assay
 Dentin barrier tests
• Advantages:
1. Quick to perform
2. Least expensive
3. Can be standardized
4. Large scale screening
5. Good experimental control
6. Excellence for mechanism of interaction
• Disadvantages:
1. Relevance to the final in vivo use is questionable
2. Lack of inflammatory and other tissue protection
mechanisms in the in vitro environment
3. Cannot predict the overall biocompatibility of a
material

Gottfried schmalz, Biocompatibility of

dental materials, 3rd ed,2009
Test for Cytotoxicity or cell growth

Tests for cell metabolism or cell functions

Mutagenesis assays
CYTOTOXICITY TEST

• CELL CULTURE TEST: Cells are

plated in a well of a cell
culture dish where they
attach. The Material is then
placed in the test system. If
the material is not cytotoxic,
the cells will remain attached
to the well and will proliferate
with time. If the material is
cytotoxic, the cells may stop
growing, detach from the
well.
MEMBRANE PERMEABILITY

• It is the ease with which a dye can pass through

a cell membrane. This test is used on the basis
that a loss in membrane permeability is
equivalent to or very nearly equivalent to cell
death.
• Two types of dyes are used:

vital (eg . Neutral red,Na2 51CrO4)

non vital(eg. Trypan blue, propidium iodide)
TEST FOR CELL METABOLISM OR CELL FUNCTION

• Some in vitro tests for biocompatibility use the

biosynthetic or enzymatic activity of cells to assess
cytotoxic response.
• Tests that measure deoxyribonucleic acid (DNA)
synthesis or protein synthesis are common examples
of this type of test.
• These test are not routinely use to assess
biocompatibility of materials.
MTT test:[3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium
Bromide]

• Measures the activity of cellular dehydrogenase

• MTT is a yellow, soluble molecule
• Chemical MTT is used to know the enzymatic action
of cell.
• If the cell is able to reduce the MTT the resulting
Formazen formed is proportional to the enzymatic
activity.
• Formazen quantified by dissolving it and measuring
the optical density of the resulting solution,
alternatively the formazen can be localized around
the test sample by light or electron microscopy
• Other formazen generating chemicals NBT, DXTT,
WST, Alamar Blue.
• Alamar blue tests quantitatively measure cell
proliferation using a fluorescent indicator that allows
the continuous monitoring of cells over time.
MUTAGENESIS ASSAY

• Mutagenesis assays assess the effect of

materials on a cell’s genetic material.
• Genotoxic mutagens directly alter the DNA of the
cell through various types of mutations.
AMES TEST

• The test uses several strains of the bacterium

Salmonella typhimurium that carry mutations in
genes involved in histidine synthesis
• It is a auxotrophic mutant, so that they require
histidine for growth.
• The variable being tested is the mutagen's ability to
cause a reversion to growth on a histidine-free
medium
• The plate is incubated for 48 hours
• Eg ; amalgam & drugs
STYLE’S CELL TRANSFORMATION TEST

• Non- mammalian test

• This assay quantifies the ability of potential
carcinogens to transform standardized cell lines so
they will grow in soft agar
• Untransformed fibroblasts normally will not grow
within an agar gel, whereas genetically transformed
cells will grow below the gel surface
 Indirect tests

• Tests that use barriers [Indirect tests]

• Cytotoxicity tests measure the toxicity when material
is indirect contact with the cell culture
• Agar overlay method
• Millipore filter assay
• Dentin barrier tests
Agar over lay method
• Agar forms a barrier between the cells and the
material which is placed in the top of the agar
• Nutrients, gas and soluble toxic substances can
diffuse through the agar
• Solid test samples or liquid samples absorbed on to
filter paper can be tested with this assay for up to
24hrs
Millipore filter assay

• Establish monolayer of cells on the filter, Culture

medium is placed and this mixture is allowed to gel
over the cells
• Filter- monolayer gel is detached and turned over so
that the filter is on top for placement of solid or
soluble test samples for 2 or more hours
• After exposure the toxicity in the Millipore filter test
is assessed by the width of the cytotoxic zone around
each test sample
Dentin barrier tests

• It shows the improved correlation with the

cytotoxicity of dental materials in usage tests in teeth
• Incorporation of dentin disks between the test
samples and the cell assay system
• Then the cytotoxicity is measured and the testing
material placed on one side of the dentin disk in the
devised used to hold the dentin disk
• Cells can also be grown in the collection side of the
disks. Collection fluid is also in the other side.
Immortalized pulpal fibroblats can be used as target
cells.
• Components of the material may diffuse through the
dentin and effect of medium on cell metabolism can
be measured
• To assess the rate of diffusion the collection fluid can
be circulated into and out of the collection
ANIMAL TESTS
ANIMAL TESTS
• Animal tests is done by placing a material into an
intact organism of same type.
• Common animals used are mice, rats, hamsters,
or guinea pigs, although many types of animals
have been used including sheep, monkeys, pigs,
cats and dogs.
• Animal tests are distinct from usage tests
(which are also often done in animals) in that the
material is not placed in the animal with regard to
its final use.
 ANIMAL TEST

SYSTEMIC TOXICITY TEST (SHORT/LONG TERM)

MUCOUS MEMBRANE IRRITATION TEST

IMMUNE SENSITIZATION TEST

MUTAGENECITY/ CARCINGENICITY TEST

IMPLANTATION TEST
Mucous Membrane Irritation test

• Mucous Membrane Irritation test determines if

a material causes inflammation to mucous
membranes or abraded skin.
• Done with hamster cheek-pouch tissue or rabbit
oral tissue
Skin Sensitization Test
• Skin Sensitization Test in guinea pigs (guinea
pig maximization test), the materials are injected
intradermally to test for development of skin
hypersensitivity reactions
Implantation Tests

• Implantation Tests used to evaluate materials

that will contact subcutaneous tissue or bone..
• Used for testing implants & endodontic materials.
• Material placed subcutaneously, intramuscularly,
or as a bone implant at lateral cortex of femur or
tibia or both.
• Histopathological examination has to be done
• Observation period may be upto 1 year.
Systemic Toxicity Test

• Material is administered to test animals eg: rat

(orally or i.v.). If >50% animals survive –material
is safe.
 USAGE TEST
• Usage tests are performed in animals or humans.
• A usage test requires that the material be placed in an
environment clinically relevant to the use of the material in clinical
practice. If the test is performed in humans, it is called a clinical
trial.
• These tests are usually performed on larger animals with anatomy
that more closely resembles that of humans.
• The human clinical trial is therefore the “Gold standard” of usage
tests.
 Dental Pulp Irritation Tests
• Pulp compatibility of a material is investigated on
teeth of experimental animals or on human teeth
that have to be extracted for orthodontic reasons.
• In both cases, class V cavities are prepared as
atraumatically as possible and are then filled with the
test material. This approach is equivalent to the
future mode of application on patients.
• After a period of days to several months, the teeth
are removed and histologically prepared, and the
pulps are microscopically evaluated for signs of acute
or chronic inflammation and odontoblast reaction
(including dentin neogenesis)
• In addition, the space between test material and the cavity
wall is investigated for bacterial penetration.
• These methods can be modified in such a way that the pulp
is exposed or part of the pulp is removed before the
material is applied.
• Assessment: The most important causes of pulp damage
resulting from a restorative procedure (in addition to cavity
preparation) are the following:
 Toxic substances released from the material
 Bacteria and their toxins between the material and the
cavity
• The pulp can react to these irritations in the following ways:
1. Inflammation
2. Tertiary dentin formation
• Limitations:
• Dentin sclerosis reduces the dentin’s permeability,
but low-molecular substances may diffuse even
through sclerotic dentin.
• However, sound teeth, mostly without obliterated
dentin, are used in pulp/dentin tests.
• Thus, there may be a discrepancy between the
clinical situation (below a carious lesion with dentinal
sclerosis) and the usage test with filling materials.
• The diffusion of potentially damaging substances
through sclerotic dentin toward the pulp may be
reduced.
• In addition, the target organ of the pulp/dentin test is
the pulp of sound (test) teeth and not the
“predamaged” pulp, as is frequently the case in
patients.
• A chronic inflammation in the patient’s tooth pulp
may impair the defensive capacity of the pulp,
rendering it more susceptible to toxic material
components.
Periapical Tissue Damage and Endodontic Usage Test

• The literature includes descriptions of animal models

(e.g., primates, dogs) that allow the application of a
given material into the root canal according to
endodontic techniques after a usual root canal
preparation.
• Compatibility is assessed by histologic evaluation of
the periapical tissues.
• Assessment: The classic endodontic usage test is very
elaborate and includes the same technical and ethical
problems as the pulp/dentin test using large
experimental animals.
• Relatively few studies using this test method are
available in the literature.
• The presented findings, however, document a good
correlation with clinical observations.
• In particular, stimulating effects on special cells can
be determined, such as the influence of calcium
hydroxide compounds on periapical cementoblasts.
• Otherwise, implantation tests, in which Teflon tubes
are filled with the experimental material and
subsequently implanted, may be used as alternatives
• Such tests are especially useful when assessing the
claimed bioactive effects of test materials.
 Dental Implants in Bone

• Materials used for dental implants are inserted into

the jaw of test animals (intraosseous implants).
• For this, penetration of the epithelial barrier,
equivalent to the treatment of patients, is simulated
on experimental animals.
• Appropriate animals are, among others, primates,
dogs, miniature pigs, guinea pigs, and rats.
• Tissue reaction is assessed histologically, with the
tissue in contact with the implant being particular
interest
• A good correlation of these findings with patients’
situations can be expected.
 Mucosa and Gingival Usage Tests

• Tissue response to materials with direct contact of

gingival and mucosal tissues is assessed by
placement in cavity preparations with subgingival
extensions.
• The material’s effect on gingival tissues are observed
and responses are categorized as slight, moderate, or
severe, depending on the number of mononuclear
inflammatory cells
• A difficulty with this type of study is the frequent
presence of some degree of pre existing
inflammation in gingival tissue due to the presence of
bacterial plaque, surface roughness of the restorative
material, open or overhanging margins, and over- or
undercontouring of the restoration.
 Diagnostic Tests on Patients
• Contrary to the test methods that are used to
characterize a (new) material and which have been
described so far, diagnostic tests on patients are used to
more deeply analyze claimed or real unwanted side
effects in individual subjects (individual compatibility).
• This branch of biocompatibility studies has become very
important during recent years, since many materials do
not cause clinically manifest reactions in the vast
majority of the population but may generate claimed or
real disease symptoms linked to materials in single
cases. The assumption of an individual compatibility
for dental materials is based on these observations.
Allergy Tests
• The patch test, originally developed and described by
Jadassohn , is the most important allergy test
regarding dental materials.
• This test can be applied to identify delayed type
hypersensitivity (type IV reactions) as the cause for
an allergic contact dermatitis.
• Immediate reactions (type I reaction, such as asthma)
can be diagnosed by the prick test.
Patch Test
• Adhesive tapes containing the potential allergens at
concentrations that are just high enough to trigger
the allergic reaction (but which are nonirritating) are
adhered to the clinically sound skin of the patient’s
back.
• The most important allergens are combined in so-
called standard series at ready-made concentrations
and are commercially available. Special series include
dental materials.
• The patient should avoid excessive sweating or
exposure to sun as well as scratching of the back, and
should not have a shower or bath.
• During the following days, after the tape has been
removed, the skin is evaluated for test reactions:
redness, itching, blisters, etc.
• Skin reactions are assessed after 2 and 3 days but
later checks (after 5 and 7 days) are also necessary to
detect late reactions, since immunocompetent T
lymphocytes occasionally require several days before
they cause a visible allergic reaction.
• The patch test is the primary method for the
detection of a delayed-type hypersensitivity (type IV
reaction) allergy to dental materials.
Prick Test
• This test is used to detect “immediate-type” allergies (type
I reactions).
• The allergen is applied as a drop to the skin, and then the
skin is “pricked” through the drop.
• Test substances/extracts (e.g., containing natural latex) are
offered by various manufacturers. After 5–30 min, the skin
reaction is assessed (redness, formation of weals).
• Although the risk of provoking an immediate allergic
reaction by the test itself is very minor, it cannot be
completely excluded.
• Therefore, this test should be executed only by qualified
personnel.
Radioallergosorbent Test (RAST)
• The RAST belongs to the group of in vitro tests for
diagnosing an allergy.
• It is used to diagnose immediate- type allergies (IgE
mediated) by identifying an allergen-specific IgE in the
patient’s blood.
• Because the RAST is an in vitro test, the patient will not
exposed to the risk of sensitization by the test itself.
• However, with atopic patients or through other circulating
antibodies, this test may render results that are
inconsistent with clinical findings.
• This test can be used for diagnosing suspected allergies to
medication or latex, potentially in combination with the
prick test.
valuation of Pulp Sensitivity
• The sensibility test of the pulp may demonstrate
functional nerval structures.
• This method is used for pulp diagnosis and is mainly
based on the application of cold and of electric
current.
• The threshold of pulp nerves regarding electric current
varies between 20 and 100 μA, whereas this value for
periodontal structures ranges between 176 and 250
μA.
• Thus, it is possible to differentiate between an
irritation of nerves in the pulp and in the
periodontium.
• Thermal examination is performed with sticks of ice,
CO2-snow (– 78.5 oC), or cold sprays, which, for
instance, contain propane, butane or similar
substances (– 22 to – 50 oC). Dichlorine–difluorine–
methane, which has been used previously, has been
discontinued for environmental reasons.
• These tests are frequently applied in biocompatibility
tests to determine material-associated pulp damages
in clinical studies.
• However, a decisive limitation of sensibility tests is that
they only indicate the presence of functioning nerval
structures. But these tests cannot be used to prove
vitality or specific inflammatory reactions of the pulp.
 CORRELATION AMONG IN VITRO, ANIMAL, AND USAGE TESTS

• No single test is used to evaluate the biocompatibility of a

new material.
• Three phases are generally recognized in the testing of a
new biomaterial
Primary test (In vitro)
(e.g the first test conducted to evaluate a new casting alloy
are in vitro cytotoxicity and mutagenicity test.)
↓
Secondary test (In vivo)
↓
Usage test
• Question about the accuracy of primary and secondary
test have challenged the wisdom of linear paradigm
• In 1977 Mjore et.al. Challenged the linear testing
paradigm and showed that the in vitro and animal test
did not necessarily predict the results of usage tests
• Therefore most researchers have adopted an alternative
paradigm .The basic linear paradigm is present but the
need to consider non liner thinking is also infused.
 Standards
• 1972 – Council on Dental Materials, Instrument
and equipments (later called Council on
Scientific Affairs) of ANSI/ADA approved
document No. 41 for recommended standard
practices for biological evaluation of dental
materials.
• 1982 – ANSI/ADA document No.41 was updated
to include tests for mutagenicity.
• 1992 – ISO 10993 was first proposed but
modified versions are updated periodically.
• 2002 - ISO 10993 consisted of 16 parts, each
addressing a different area of biological testing.
Allergic Responses to dental materials:
Allergic contact dermatitis or stomatitis
• This is most common adverse reaction to dental
materials
• The interval between exposure to the causative agent
and the occurrence of clinical feature varies between 12-
48 hrs.
• It usually occurs where body surface makes direct
contact with the allergens.
• E.g.: - monomers of bonding agent, acrylic components
of dental cements.
• Industry workers who handle these materials are also
affected.
Allergy to latex Products: -
• In 1991, FDA issued a bulletin in respond to the
increasing number of latex-related allergic reactions.
• Malten & associates (1976) reported increasing
incidence of hypersensitivity and suggested that the
polyether in latex rubber gloves was causative agent
• Dermatitis of the hands (eczema) in the most
common adverse reaction.
• Reactions vary from localized rashes and swelling to
wheezing and anaphylaxis.
• To avoid these reactions to latex products, vinyl
gloves may be used.
• Blink horn and Leggate (1984) and Axelsson et al
(1987) reported adverse reaction to rubber dam
involving respiratory distress, edema, and chest
pains.
• The definitive diagnostic test for these is patch test.
• The suspected allergen is applied to skin with intent
to produce reaction in around 48-96 hrs.
Mercury Controversy: -

• Controversy has raged over biocompatibility of

amalgam restorations because of the presence of
elemental mercury.
• Recognized symptoms of chronic mercury poisoning
are weakness, fatigue, anorexia, weight loss,
insomnia, etc.
• The lowest level of total blood mercury at which
nonspecific symptom occur is 35 µg/mL.
Minimizing Dental Iatrogenesis:

• Iatrogenesis is defined as the creation of side effect,

problems, or complications resulting from treatment
by a physician or dentist.
Cavity preparation:
• Stanley HR (1994) reported that low hand piece
speed (6000-20,000 rpm) with air water spray, a
cavity preparation 2 mm from the pulp, elicits
minimal pulp lesion.
• If preparation is less than 1 mm of the pulp, intensity
of response increases.
• Histopathologically, thermal insult results in loss of
cytoplasmic continuity of odontoblasts and
displacement of odontoblast nuclei into the dentinal
tubules due to dehydration.
• The generation of heat within the pulp is the most
severe trauma that restorative procedures impart on
the pulp.
• If the insult is extensive and cell rich zone of pulp is
damaged, reparative dentin formation may be
impaired.
• The pulp is a tissue of low compliance according to
Goodies et al (1989) because it is encased in hard
dentinal walls, it consists of a large amount of
connective tissue with a small blood supply and has
no possibility of developing a collateral circulation.
For these reasons, the pulp is vulnerable to thermal
damage during and after extensive restorative
procedures.
• In a cavity preparation with a diamond bur, the entire
surface of the bur is in contact with the tooth surface
thus generating frictional heat but in the case of TC
bur the flutes themselves may allow a slight cooling
action with a greater cutting efficiency.
• According to Cohen, ‘Blushing’ of teeth during or
after cavity or crown preparation is attributed to
frictional heat. Coronal dentin develops a pinkish hue
very soon after dentin is cut. This represents vascular
stasis and is reversible
Pulpal reaction to restorative materials
Pulp responses to specific agents

Bleaching agents
• These are used in non-vital and vital teeth.
• These agents contain peroxides
• These agents may be in contact with teeth for several
minutes to severe hours.
• Peroxides can penetrate the intact enamel and reach the
pulp.
• Occurrence of tooth sensitivity is very common with the
use of these agents.
• Bleaching agents will also damage the gingiva, if not
isolated properly.
Amalgam
• Swerdlow and Stanley (1962) reported that the pulp
response to amalgam placement is due to
condensation pressure.
• Little pulpal response is elicited when cavity is
prepared with high-speed air-water spray technique
• However, when cavity is restored with amalgam the
pressures of condensation will intensify the response
• Boremark and associates (1968) showed that
radioactive mercury reached the pulp in humans
after 6 days if no cavity liner was used.
• Implantations tests show that low copper amalgams
are well tolerated, but high copper amalgam cause
severe reaction.
• Liners are suggested to avoid pulpal reaction.
• Amalgam based on gallium rather than mercury have
been developed that are free of mercury.
• The following pulp reactions may occur immediately
after application/condensation of amalgam in deep
cavities with a remaining dentin thickness (RDT) of
less than 0.5 mm
 Reduced number of odontoblasts
 Odontoblast nuclei in dentin tubules
 Dilated capillaries
 Slight to severe inflammatory cell infiltration in the
odontoblast layer
Visible light-cure Resin composites
• The level of the pulp response to resin composite
restorations is especially intensified in deep cavity
preparations when an incomplete curing of resin
permits a higher concentration of residual
unpolymerized monomer to reach the pulp.
• Visible light-cured systems were developed to
provide greater depth of cure, shorter curing time,
less porosity and more wear resistant composite
restoration.
• A more conservative cavity preparation with
incremental placement of the resin composite is
highly recommended to minimize the pulp response.
• No pulp damage is to be expected if resin-based
composites or adhesives are applied in shallow or
medium cavities, even after prior acid-etching of the
dentin (total etch/total bonding technique).
• In these situations, adhesives may serve as sealants
and thus as protection against potentially penetrating
bacteria
• In deep cavities, however, especially if microexposure
of the pulp cannot be excluded, the use of a calcium
hydroxide preparation applied on the deepest part of
the cavity is still recommended.
• If a calcium hydroxide suspension is used for this
purpose, then it should be covered by suitable glass
ionomer cement.
Zinc Phosphate Cement
• If zinc phosphate is used instead of ZOE to cement a
crown or inlay, the phosphate cement is forced into
the dentinal tubules
• After 3-4 days, it creates a wide spread three
dimensional inflammatory lesion involving all the
coronal pulp tissue.
• A young tooth with wide and open dentinal tubules is
more susceptible to intense response than an older
tooth, which has produced sclerotic and reparative
dentin that blocks the tubules.
• Zinc phosphate cements elicits strong to moderate
cytotoxic reactions that decrease with time after
setting. Leaching of zinc ions and a low pH is cause of
these effects.
• Initial pH on setting is 4.2 at 3 minutes
• The best protection against phosphoric acid
penetration is provided by coating the dentin with
two coats of an appropriate varnish, a dentin-
bonding agent, or a thin wash of calcium hydroxide.
• Calcium hydroxide plugs the dentin tubules and
neutralizes acids; hydrophilic resin primers infiltrate
the collagen mesh produced by acid-etching of the
dentin and seal the patent dentin tubules.
• These procedures eliminate 90% of the severity of
the adverse pulp responses, making them similar to
those of polycarboxylate cement.
Zinc Oxide Eugenol cements

• ZOE is recommended as a nontoxic reference substance

in respective to other materials.
• Cox CF et al 1987 stated that eugenol from ZOE fixes
cells, depresses cell respiration and reduces nerve
transmission with direct contact.
• ZOE may form a temporary seal against bacterial
invasion
• It inhibits the synthesis of prostaglandin and leukotriens
(anti-inflammatory)
• Interaction of eugenol with vallinoid receptors on nerve
cells playing an important role in nociception.
Glass Ionomer Cement
• When GIC first introduced, the pulpal response were
classified as bland, moderate, less irritating than silicate
cement and zinc phosphate cement.
• The blandness of GIC is attributed to absence of strong
acids of toxic monomers.
• Polyacrylic acid and polyacids are much weaker than
phosphoric acid and possess higher molecular weight that
limit their diffusion through dentinal tubules to the pulp.
• Tobias et al. (1978), found that glass ionomer cements
were less irritating than zinc phosphate cement,
equivalent in irritancy to polycarboxylate cement and
more irritating than zinc oxide cement.
• Smith and Rusa (1986) compared the initial activity of
GIC with zinc polycarboxylate and zinc phosphate
cements and found a general rise in pH for all
cements during first 15 minutes. However, the initial
reactions of GIC’s were slower.
• It is recommended that if there is less than 0.5-mm
residual dentin or a pulp exposure, an appropriate
lining of calcium hydroxide should be placed prior to
the placement of a glass ionomer
Conditioning (etching) agents:

• Before a resin composite or a GIC restorative material

is placed, surface contaminants must be removed to
permit the micro mechanical attachment or the ionic
exchange of the dental material with the tooth
structure.
• Brannstrom and Nordenvall (1977) noted no
significant difference between dentinal surface
conditioned for 15 seconds and those conditioned for
2 seconds and thus recommended shorter
conditioning times.
• Brannstrom (1981), showed that conditioning of
dentin and removal of smear layer allows the ingress
of bacteria and the outward flow of dentinal fluid
within the tooth – material interfacial region
resulting in biofilm formation that interfaces with
adhesion.
• Some scientists recommend that smear layer should
remain but in modified form, where as some other
propose that the smear layer be completely
removed.
• Bowen and colleagues (1982) introduced mordanting
solution (acidified ferric oxalate), that appeared to
dissolve the original smear layer and replace it with a
more uniform ‘artificial’ (altered) smear layer.
• With the use of less concentrated acids with higher
molecular weights and shorter time intervals for
conditioning, pulp response is minimized.
Bonding Agents

• Bonding agents do not appear to be toxic

• To enhance bonding to composite, a fast setting
visible light cured, low viscosity (unfilled) resin primer
is applied that infiltrates the demineralized dentin
surface and the exposed collagen mesh to form
hybrid layer.
• The plugging of the dentinal tubules prevents the
penetration of toxic components to the pulp from
composite restorations.
Influence of Patient Age on Pulp response

• As permanent teeth endure the effects of abrasion,

caries and restorative procedure, the pulp becomes
reduced in size because of deposition of secondary
dentin, pulp stones and clarifications.
• At age 55 years, the volume of pulp tissue is one fifth
that at age 25 years and contains only one fifth of its
former blood supply (Stanley, 1990)
• If an inflammation develops in pulp of an aging
patient, that pulp has less defense in resolving a
lesion and resisting infection.
ENDODONTIC MATERIALS

GUTTA PERCHA
• Only highly purified gutta-percha should be used in
patients with a latex allergy. If necessary, synthetic
gutta-percha points can be applied (e.g.,
Synthapoints).
• It may be concluded from data that
thermomechanical compaction (condensation),
specifically at a higher rotational speed
(>10,000/min), may damage the periodontal tissues.
 Histopathologic evaluation of subcutaneous tissue
response to three endodontic sealers in rats Ali R. Farhad et
al Journal of Oral Science, Vol. 53, No. 1, 15-21, 2011
evaluate the subcutaneous biocompatibility of three root canal
sealers (AH Plus,epiphany,grossman sealer).
RESULT:Grossman endodontic sealer had the most severe
inflammatory response followed by the AH Plus, Epiphany.

 Preliminary study of the inflammatory response to

subcutaneous implantation of three root canal sealers D.
Silva-Herzog et al International Endodontic Journal 2011
evaluate RoekoSeal was the most biocompatible sealer,
producing the least amount of inflammatory exudate, when
compared to sealapex and AH Plus.
ZOE SEALERS

• Patients with an allergy to eugenol (or to fragrances)

should not be treated with materials containing
eugenol, isoeugenol, or Peru balm.
• Data show that ZOE sealers are characterized by a
moderate local toxicity, which is significantly
increased if paraformaldehyde is added.
• A number of case reports document that
paraformaldehyde-containing ZOE sealers may cause
an aspergillosis of the maxillary sinus when the root
canals of upper posterior teeth are overfilled and the
sealers are pressed into the maxillary sinus
Epoxy-based sealers

• Epoxy-based sealers are initially toxic, but toxicity

considerably declines when the materials are set, and
then no tissue reactions, or only slight ones, are
observed.
• Epoxy-based sealers are initially toxic, but toxicity
considerably declines when the materials are set, and
then no tissue reactions, or only slight ones, are
observed.
Calcium-Hydroxide-Based Sealers

• CH sealers are characterized by a low toxicity, which

occurs only in the initial period after application.
• There is clear indication that these materials may
stimulate the formation of hard tissue.
• However, an inferior marginal adaptation together
with microleakage due to increased solubility is a
potential risk to be considered for this group of
materials.
Endodontotic procedures

• As a consequence of pathologic changes in the dental

pulp, the root canal system can harbour numerous
irritants.
• As irritants are released from the root canal system
into the periradicular tissue, granulation tissue
proliferates and replaces normal periradicular tissues.
• Removal of irritants from the root canal system and
its total obturation results in repair of the
periradicular tissue to its normal architecture.
• Grossman indicated that ideal root canal filling
material should meet the following requirements-
 It should seal the canal laterally as well as apically
 It should not shrink after being inserted
 It should be impervious to moisture
 It should be bacteriostatic or at least not encourage
growth
 It should not irritate periradicular tissue.
 It should be neither mutagenic or carcinogenic
• The sealers form a fluid – tight seal at the apex by
filling
 The minor interstices between solid material and the
canal wall
 The patent accessory canals.
• A minimal reaction was found when the canal was
not overfilled with sealant.
• When the teeth were overfilled with sealant, there
was persistent chronic inflammatory response.
• Gulati et al 1990 stated that cytotoxicity of zinc oxide
eugenol might be attributed to the fact that
eugenolate formed hydroxide in contact with tissue
fluids and released free eugenol which was
responsible for toxicity
• Mittal et al 1999 attributed the toxicity of Sealapex to
polymeric resin and for Endoflas FS to presence of
eugenol and para monochlorophenol.
• AH26 resin sealer cause moderate to severe toxicity.
This toxicity is due to release of formaldehyde during
the initial setting reaction between bisphenol A
resinand hexamethylene tetramine. (Span Burg et al
1993).
 Sealer efficacy
• Although all root canal sealers leak to some extent, there
is a critical level of leakage that is unacceptable for healing
and may result in endodontic failure.
• Leakage may occur at the interface of dentin and sealer, at
interface of solid core and sealer, through the sealer itself
etc.
• If the apical surface can be blocked principally by a solid
core material success is improved over time.
• In most studies when obturation was done without sealers
the leakage results were enormously greater.
• Without question, all the materials used to seal root canal
irritate peri-radicular tissue if allowed to escape from the
canal.
• M. Mittal, Satish Chandra and Shallen Chandra
Comparative tissue toxicity evaluation of four
endodontic sealers. Journal of Endodontics.Vol.21,
No.12, 1995, 622-62. They evaluated the response of
the tissue histologically to four root canal sealers Zinc
Eugenol, Tubliseal, Sealapex and Endoflas. By
injecting them into the subcutaneous connective
tissue of dorsal surface of rats.
• This study concluded all the sealers caused some
inflammation that decreased with time. Overall,
Sealapex showed least inflammatory reaction
compared with other sealers.
 REFERENCES

• Gottfried schmalz, Biocompatibility of dental

materials, 3rd ed,2009.
• Phillips’ –science of dental materials 11th ed.
• Basic dental materials John J Manappallil- 4th ed.