Contents

Introduction
Relevance to dentists
Tests for evaluation of biocompatibility
Craigs classification of tests
Advantages disadvantages of tests
Standards that regulate the measurement
biocompatibility

of

Pulpal responses to specific agents and techniques.

Reaction of oral soft tissues to restorative materials
Reaction of bone and soft tissues to implant material
Biocompatibility of metals

Biocompatibility is defined as the ability of a

material to elicit an appropriate biological response in a
given application in the body.

The term Biocompatible is defined in Dorlands

Medical Dictionary as being harmonious with life and
not having toxic or injurious effects on biologic function.

In general, biocompatibility is measured on the basis of

Toxicity [such as pulp and mucosal response],

Inflammation,
Allergic responses and
Mutagenic reactions.

Requirements of a biocompatible dental

material
It should
not be harmful to the pulp and soft tissues.
not contain toxic diffusible substances that can be released and
absorbed into the circulatory system to cause a systemic toxic
response.
be free of potentially sensitizing agents that are likely to cause an
allergic response.
have no carcinogenic potential.
Whether a material is biocompatible or not is dependent on what
physical function.

RELEVANCE TO DENTISTS

Dentists potential concerns about biocompatibility can be

organized in to 4 areas:
1) Safety of the patient
Ex: Alloys, resins and cements.
2) Safety of Dental staff
Ex: Amalgam Mercury vapour.
Chronic exposure to latex and resin based materials.

3) Regulatory compliance issues

Biocompatibility issues are closely linked to regulations
that affect dental practice.
Ex: Dental amalgam.
Because of the biologic concerns about mercury,
regulators have considered monitoring and restricting
amount of mercury in waste water for dental practice.

4) Legal Liability
Biocompatibility issues also influence liability issues that
affect dental practitioners.
Because dental materials can affect the well-being of
patients and dental auxillaries, practitioners assume a
legal risk when using these materials.

ELEMENTS INVOLVED AND INTERFACES

Tests for evaluation of Biocompatibility

The purpose of biocompatibility tests is to eliminate any

potential product or component of a product that can
cause harm or damage to oral and maxillofacial tissues.
Biocompatibility tests are classified on three levels, with
the most rapid and economical occurring at primary level.
Group I : PRIMARY TESTS
Group II: SECONDARY TESTS
Group III: PRE-CLINICAL USAGE TESTS

Tests for evaluation of Biocompatibility:

Craig classification
Biocompatibility Tests
In vitro tests

Animal tests

Usage tests

Skin sensitization tests

Direct

Indirect

Mucous membrane irritation tests

Cytotoxicity tests
Tests for cell metabolism or cell function
Tests that measure immune function
Muagenesis assays

Group I : PRIMARY TESTS

The primary tests consist of cytotoxic evaluations either in
direct or indirect methods.
Genotoxicity Test
Mammalian or non mammalian cells, bacteria, yeasts or
fungi are used to determine whether gene mutations,
changes in chromosomal structures or other deoxyribo
nucleic acid or genetic changes are caused by the test
materials, devices and extracts from materials.

Group II: SECONDARY TESTS

At this level the product is evaluated for its potential to

create systemic toxicity, inhalation toxicity, skin irritation
and sensitization and implantation responses.
In systemic toxicity tests such as oral median lethal dose
test, test sample is administered daily to rats for 14 days
either by oral gavage or by dietary inclusion. If 50% of
animals survive, the product has passed the test.

The inhalation toxicity tests are performed in an exposure

chamber with aerosol preparations by releasing the spray
material around head and upper trunk of animals.

The animals are subjected to 30 seconds of continuous

spray released at 30 minute interval. After 10 consecutive
exposures, the animals are observed over a 14 days
period. If any animal dies within 2-3 minutes, the agent is
considered very toxic.

Implantation tests

The animal species is selected according to the size of the

implant test specimen and the intended duration of the test
in relation to the life span of animal.
Short term tests ( 12 weeks) Mice, rats, guinea pigs
Long term tests ( Weeks) dogs, rabbits, sheep, goats.

Group III : PRE CLINICAL USAGE TESTS

Pulp and Dentin Usage test

This test is designed to assess the biocompatibility of dental
materials placed in dentin adjacent to the dental pulp. Class V
cavity preparations are done to leave 1mm or less of tubular
dentin between the floor of cavity preparation and pulp.
The appropriate number of cavities are restored, and some are
retained for control specimens.

The animals are sacrificed after 7 days, 28 3 days and

70 5 days.
After routine histopathologic processing, specimens are
graded for degree of inflammatory response to prevalence
of reparative dentin formation in pulp and the number of
microorganisms (microleakage) entrapped in the
surrounding cavity walls and cut dentinal tubules.

Pulp Capping and Pulpotomy Usage test

In this test procedure pulp is merely exposed for the pulp

capping evaluation and is partially removed for
pulpotomy assessment.
A Ca(OH)2 product is used as a negative control. The
animals are sacrificed after 7 2 days 70 5 days.
Observations are made of dentinal bridge formation
adjacent to or sub adjacent to the applied material.

Pulp capping

Pulpotomy

Endodontic usage test

Root canals are replaced by obturating test material and control

material. ZOE alone or ZOE combined with a sealer is used as
control material.
The animals are sacrificed after 28 3 days and 13 Weeks.
For a biocompatible material, one should observe minimal or
no response.

Standards that regulate the measurement

of Biocompatibility

ADA established guidelines 1926.

In 1972 the council on Dental materials, instruments, and
equipment of ANSI/ADA approved Document No. 41 for
recommended standard practices for biological evaluation
of Dental materials.
Three categories of tests are described in the 1982 ANSI /
ADA document: initial, secondary and usage tests.

Initial tests include invitro assays for cytotoxicity, red

blood cell membrane lysis (hemolysis) mutagenesis and
carcinogenesis at the cellular level.
Secondary tests (in-vivo) for inflammatory or
immunogenic potential.
Finally, materials that pass secondary tests and still hold
potential are subjected to one or more in vivo usage tests.

ISO 1993

It contains 12 parts, each dealing with a different aspect of

biological testing.
This standard divides tests into initial and
supplementary tests to assess the biological reaction to
materials.
-Initial tests for cytotoxicity, sensitization, systemic
toxicity.
-Supplementary tests are tests such as chronic toxicity,
carcinogenicity and biodegradation.

Pulpal Responses to specific Agents and

Techniques

I. AMALGAM

While placing a conventional amalgam restoration,

pressure of condensation will intensify the initial
minimal inflammatory response and it will subsequently
increase the formation of reparative dentin.

II Chemically Cured Resin Composites

The addition of mineral fillers to the direct filling chemically cured resin
composites did not reduce their potential for creating severe pulp responses.
If not lined properly, they cause chronic pulpitis that persists for an
indefinite time.

III Visible light Cured Resin composites

Level of pulp response to resin composite restorations is especially intensified

in deep cavities when an incomplete curing of resin permits a higher
concentration of residual unpolymerized monomer to reach the pulp.
Precautions:
1) Use twice the recommended time exposure
2) Cure in increments.

IV Zinc phosphate cement

As a base more toxic than as a luting agent.

A young tooth with wide open dentinal tubules is more
susceptible.
Coat the dentin with two coats of an appropriate varnish, dentin
bonding agent, liner, or a thin wash of calcium hydroxide.

V. Zinc oxide eugenol cement

In vitro, eugenol from ZOE, depresses cell respiration and reduces

nerve transmission with direct contact.
The concentration of eugenol in cavity preparation just below ZOE
is bactericidal, the concentration on the pulpal side of dentin may
be less.
This lower concentration reportedly suppresses nerve transmission
and inhibits synthesis of prosta glandins and leukotrienes. [anti
inflammatory]

VI. Glass lonomer cement

Polyacrylic acid and related poly acid are much weaker

than phosphoric acid, as polymers, they possess higher
molecular weights that may limit their diffusion through
the dentinal tubules to the pulp.

VII. Conditioning (etching) Agents

Conditioning procedures are used with both resin composite systems and
GICs.
Conditioning techniques that are associated with weaker acids, shorter
periods of application, and elimination of subbing and scrubbing procedures
produce a minimal pulp response.

VIII. Bonding Agents

Many of these reagents are cytotoxic to cells invitro if
tested alone.
When placed on dentin and rinsed with tap water between
applications of subsequent reagents, cytotoxicity is
decreased.
Long term in vitro studies suggest that components of
many bonding agents permeate up to 0.5 mm of dentin to
cause significant suppression of cellular metabolism for up
to 4 weeks after their application.

IX. Liners, Varnishes and Non-resin Cements

Calcium hydroxide cavity liners come in many forms,

ranging from saline suspensions with a very alkaline pH
(above 12) to modified forms containing zinc oxide,
titanium dioxide and resins.

The initial response of exposed pulpal tissue to highly alkaline

aqueous pulp capping agents.
Necrosis to a depth of 1mm

Coagulation of hemorrhagic exudates of superficial pulp.

Neutrophil infiltration.

after 5-8 weeks only slight inflammation remains.

Necrotic zone

Dystrophic calcification
Stimulus for dentin bridge
formation

X Bleaching Agents

These agents usually contain some form of peroxide (generally

carbamide peroxide).
Invitro studies have shown that peroxides can rapidly traverse
the dentin in sufficient concentrations to be cytotoxic.
Studies showed that peroxides rapidly even penetrate intact
enamel and reach the pulp in a few minutes and occurrence of
tooth sensitivity is very common.

Reaction of other Oral soft tissues to restorative

materials

Released products of restorative materials also contribute

either directly or indirectly to this inflammation,
particularly in areas where the washing effects of saliva
are less such as inter proximal areas, in deep gingival
pockets or under removable appliances.

Cements cytotoxicity - decreases substantially with

time. The buffering and protein binding effects of saliva.
Composites are initially very cytotoxic in invitro tests of
direct contact with fibroblasts. This is primarily from
unpolymerized components in the air inhibited layer that
leach out from the materials.
Amalgam restorations carried in to gingival crevice may
cause inflammation of gingiva because of products of
corrosion or bacterial plaque.

Denture base material

Denture base materials, especially methacrylates immune hypersensitivity reactions of gingiva and mucosa.
The greatest potential for hypersensitization is for dental
and laboratory personnel who are exposed repeatedly to a
variety of unreacted components.

Soft denture liners

Plasticizers which are incorporated into some materials to

make them soft and flexible, are released in vivo and in
vitro.
In animal tests, several of these materials have caused
significant epithelial changes, presumably from the
released plasticizers.

Reaction of Bone and soft tissues to implant

materials

There are four basic materials used in implant

fabrication : ceramics, carbon, metals and polymers

Reactions to Ceramic Implant Materials

Most ceramic implant materials have very low toxic

effects on tissues, either because they are in an oxidized
state or are corrosion resistant.

These are toxic and are non imunogenic and non

carcinogenic.

Reactions to pure Metals and Alloys

A variety of implant materials has been used, including
stainless steel, chromium cobalt molybdenum and titanium and
its alloys.
Titanium is a pure metal which forms a thin film of
various titanium oxides, which is corrosion resistant and allows
bone to osseo-intregrate in the soft tissue.
Peri-implantitis is now a documented disease around
implants and involves many of same bacteria as periodontitis.

BIOCOMPATABILITY OF METALS
BERYLLIUM

Although the beryllium concentration in dental alloys

rarely exceeds 2 wt % the amount of beryllium vapor
released in to the breathing space during melting of Ni-CrBe alloys may be significant over an extended period.

The risk of beryllium vapor exposure is greatest for dental

technicians during alloy melting especially in the absence
of an adequate exhaust and filtration system.

High levels of beryllium have been measured during

finishing and polishing when a local exhaust system was
not used. They were reduced to levels considered safe
when exhaust fan was used.
Exposure of beryllium may result in acute and chronic
forms of beryllium disease BERYLLIOSIS

Clinical features

Symptoms range from coughing, chest pain and general

weakness to pulmonary dysfunction.

Contact dermatitis
Chemical pneumonitis

NICKEL

It is a great concern to dental patients with a known allergy to

this element.
Dermatitis resulting from contact with nickel solutions was
described as early as 1989.
Inhalation, ingestion and dermal contact of nickel or nickel
containing alloys are common because nickel is found in
environmental sources such as air, soil and food as well as in
synthetic objects such as coins, kitchen utensils, and jewellery.

Nickel allergy was determined by a standard patch test consisting of 5%

Nickel sulfate solution or 5% Nickel sulfate solution on a petrolatum base, in
centre portion of a square band-aid.
This is applied on medial aspect of upper arm, which was cleaned with a
alcohol swab. This is left in place for 48hr undisturbed. A plain band-aid acts
a control.
It is read after 20 min.
0 no reaction.
+ erythema is seen.
++ erythema, papules are seen.
+++ erythema, papules, vesicles are seen.
++++ edema with vesicles is seen.