Introduction to Toxicology

‘there are no safe chemicals, only safe ways

of using them.’
Reference
• The root word “Toxic” is taken from the
Latin toxicus (meaning poisonous)

• Thus literal meaning of the term

“Toxicology” is “the study of poisons”
- the basic science of poisons (old definition)

Poison: Any substance that causes a harmful effect

when administered to a living organism.

Three classes of toxic agents:

1. chemical (cyanide)
2. physical (radiation)
3. biological (snake venom)

"the study of the adverse effects of chemicals or physical

agents on living organisms.
The basic science of poisons that studies toxic substance
with respect to their:
 Source
 Properties
 Mechanism of toxicity
 Toxic effect
 Detection
 Clinical manifestation
 Management
Any substance can be toxic if introduced in a dose capable
Of disturbing the normal physiological homeostasis of the
exposed body.
History Swiss physician Paracelsus (1493-1541)
credited with being -

“one of the founder of modern toxicology”

Paracelsus determined that specific

chemicals were actually responsible for
the toxicity of a plant or animal poison.
He also documented that the body's
response to those chemicals depended
on the dose received.

“All substances are poisons: there

is none which is not a poison.
The right dose differentiates a
poison from a remedy”
Orfila, a Spanish physician, is often
referred to as the founder of
toxicology. It was Orfila who first
prepared a systematic correlation
between the chemical and biological
properties of poisons of the time. He
demonstrated effects of poisons on
specific organs by analyzing autopsy
materials for poisons and their
associated tissue damage. Mathieu Orfila
 The Dose Makes the Poison

An apparently nontoxic chemical

can be toxic at high doses. (Too
much of a good thing can be
bad).

Highly toxic chemicals can be life

saving when given in appropriate
doses. (Poisons are not harmful
at a sufficiently low dose).
 Toxicology Terms
Toxicity - The word “toxicity” describes
the degree to which a substance is
poisonous or can cause injury. The
toxicity depends on a variety of factors:
dose, duration and route of exposure,
shape and structure of the chemical
itself, and individual human factors.

Toxic - This term relates to poisonous or

deadly effects on the body by
inhalation (breathing), ingestion
(eating), or absorption, or by direct
contact with a chemical.
Toxin – The term “toxin” usually is used when talking
about toxic substances produced naturally. A toxin is any
poisonous substance of microbial (bacteria or other tiny
plants or animals), vegetable, or synthetic chemical origin
that reacts with specific cellular components to kill cells,
alter growth or development, or kill the organism.

Toxicant - The term “toxicant” is used when talking about

toxic substances that are produced by or are a by-product of
human-made activities. For example, dioxin (2,3-7,8-
tetrachlorodibenzop-dioxin {TCDD}), produced as a by-
product of certain chlorinated chemicals, is a toxicant. On
the other hand, arsenic, a toxic metal.
 Threshold Dose
• The point at which toxicity first appears

Stochastic Effects
• Effects that occur by chance and which may
occur without a threshold level of dose, whose
probability is proportional to the dose and
whose severity is independent of the dose.
• e.g. carcinogens
 Toxicology Terms
Exposure – Contact providing
opportunity of obtaining a
poisonous dose.

Hazard – The likelihood that the

toxicity will be expressed.
Toxicology is the study of the adverse effects of chemical or
physical agents on living organisms.

Toxicological research examines the cellular, biochemical,

and molecular mechanisms of action as well as functional
effects such as neurobehavioral and immunological, and
assesses the probability of their occurrence.
 Thalidomide
 Introduced in 1956 as
sedative (sleeping pill) and to
reduce nausea and vomiting
during pregnancy
 Withdrawn in 1961

 Discovered to be a human teratogen causing

absence of limbs or limb malformations in
newborns
 5000 to 7000 infants effected
 Resulted in new drug testing rules
1 to 3 infants per 1,000 world
wide??
 WHAT TOXICOLOGISTS DO

- involved in the recognition, identification, and

quantitation of hazard.

- develops standards and regulations to protect

health and the environment.

- involved in safety assessment and use of data as

basis for regulatory control of hazards.

- determines risk associated with use of chemicals.

 Areas of Toxicological Study Descriptive Toxicology
 Toxicity testing for safety and
regulatory needs
Descriptive  Appropriate tests based on chemical
type, use and anticipated exposure
 Observational data generate
Mechanistic
hypotheses for mechanistic studies

Regulatory Regulatory Toxicology

 Decides if chemical poses
sufficiently low risk to allow
Mechanistic Toxicology marketing.
 Study of physiological, biochemical and  Uses data from descriptive and
molecular mechanisms by which toxic mechanistic studies
effects occur  Specific enforcement
 Mechanistic data are useful in the
responsibilities
design and production of safer
- Food and Drug Administration
Alternative chemicals for drugs and
- Environmental Protection Agency
therapeutics
-Consumer Product Safety Commission
 Differential toxicity mechanisms for age,
- Department of Transportation
sex, genotype and species variability
 Other Areas of Toxicology
Forensic toxicology is a hybrid of analytic chemistry and
fundamental toxicological principles. It is concerned primarily
with the medicolegal aspects of the harmful effects of chemicals
on humans and animals.

Clinical toxicology designates an area of professional emphasis in

the realm of medical science that is concerned with disease
caused by or uniquely associated with toxic substances.

Environmental toxicology focuses on the impacts of chemical

pollutants in the environment on biological organisms.

Ecotoxicology is a specialized area within environmental

toxicology that focuses more specifically on the impacts of toxic
substances on population dynamics in an ecosystem.
Toxicokinetics describes the changes in the concentrations of a
toxicant over time due to the uptake, biotransformation,
distribution and elimination of toxicants
STORAGE

ABSORPTION ELIMINATION
lungs liver
skin kidney
ingestion G I tract
lungs

BIOTRANSFORMATION
Toxicodynamics
describes the dynamic interactions of a toxicant with a
biological target and its biological effects. A biological
target, also known as the site of action, can be binding
proteins, ion channels, DNA, or a variety of other
receptors.
CLASSIFICATION OF TOXIC AGENTS

The term toxin generally refers to toxic substances that are

produced by biological systems such as plants, animals, fungi,

or bacteria.

The term toxicant is used in speaking of toxic substances that

are produced by or are a by-product of anthropogenic

(human-made) activities.
 CLASSIFICATION OF TOXIC AGENTS

• Toxic agents are classified in a variety of ways,

depending on the interests and needs of the
classifier.
• For example:
Toxic agents can be discussed in terms of their target
organs (liver, kidney, hematopoietic system, etc.)
Use (pesticide, solvent, food additive, etc.)
Source (animal and plant toxins)
effects (cancer, mutation, liver injury, etc.)
 CLASSIFICATION OF TOXIC AGENTS

• Toxic agents may also be classified in terms of:

 Their physical state (gas, dust, liquid)
 Their chemical stability or reactivity (explosive,
flammable, oxidizer).
 General chemical structure (aromatic amine,
halogenated hydrocarbon, etc.)
 Poisoning potential (extremely toxic, very toxic,
slightly toxic, etc.).
Exposure
For toxic effects to occur, a chemical must reach a particular target
site at a sufficient concentration for a sufficient duration.
Toxicologists usually divide the exposure of experimental animals to
chemicals into four categories:

Acute – Less than 24 hours – generally a single dose Oral

intubation/gavage, dermal, IP, SC, IV Inhalation usually 4 hr
Subacute – Repeated exposure for 1 month or less
Subchronic – Repeated exposure for 1 to 3 months
Chronic – Repeated exposure for greater than 3 mos
 Acute and Chronic Exposures
Can Lead to Different Outcomes

Benzene
Acute exposure – CNS narcosis
Chronic exposure – bone marrow damage and leukemia

Anticoagulant pesticides (1st generation)

Acute exposure – no effect
Chronic exposure – loss of blood clotting ability

Cigarette Smoke
Acute exposure – Nervous system stimulation (nicotine)
Chronic exposure – Cancer of mouth, pharynx, larynx, lung,
esophagus, pancreas and bladder; emphysema
 Exposure: Pathways
• The major routes (pathways) by which toxic
agents gain access to the body are
– Ingestion (Gastrointestinal Tract)
– Inhalation (Lungs)
– Dermal/Topical (Skin)
– Injection
• intravenous, intramuscular, intraperitoneal

• Typical Effectiveness of Route of Exposure

iv > inhale > ip > im > ingest > topical
 Effects of Chemical Exposure

A broad range of effects can be found with many drugs and

chemicals with sufficient exposure.

Allergic Reactions occur in hypersensitive individuals or after

sensitization in allergic or sensitized persons.
Often requires binding of chemical (hapten) to endogenous protein
in order to be recognized by the immune system.
Reaction ranges from skin irritation to anaphylactic shock

Idiosyncratic Reactions occur in individuals who have genetic

polymorphisms that lead to structural changes in biomolecules,
making them very sensitive or insensitive to a chemical.
 Effects of Chemical Exposure

Immediate vs Delayed Toxicity –

Immediate toxic effects can be defined as those that occur or

develop rapidly after a single administration of a substance,
whereas delayed toxic effects are those that occur after the lapse
of some time. Carcinogenic effects of chemicals usually have a
long latency period, often 20 to 30 years after the initial
exposure, before tumors are observed in humans. For example,
diethylstilbestrol (DES).

In contrast, most substances produce immediate toxic effects but

do not produce delayed effects.
 Effects of Chemical Exposure
Reversible vs Irreversible Effects
Some toxic effects of chemicals are reversible, and others are
irreversible. If a chemical produces pathological injury to a tissue,
the ability of that tissue to regenerate largely determines
whether the effect is reversible or irreversible. Thus, for a tissue
such as liver, which has a high ability to regenerate, most injuries
are reversible, whereas injury to the CNS is largely irreversible
because differentiated cells of the CNS cannot divide and be
replaced. Carcinogenic and teratogenic effects of chemicals, once
they occur, are usually considered irreversible toxic effects.
 Effects of Chemical Exposure
Local vs Systemic Toxicity –
Another distinction between types of effects is made on the basis of the
general site of action. Local effects are those that occur at the site of first
contact between the biological system and the toxicant. Such effects are
produced by the ingestion of caustic substances or the inhalation of
irritant materials.
Systemic effects require absorption and distribution of a toxicant from its
entry point to a distant site, at which deleterious effects are produced.
Most substances except highly reactive materials produce systemic
effects.
Corrosives and irritants act locally, little goes systemic. Acids (GI) Cl2
(lung). Systemic effects in sensitive (not always highest concentration)
tissues. CNS, circulatory and blood, visceral organs. Pb, DDT

For some materials, both effects can be demonstrated. For example,

tetraethyl lead
 Effects of Chemical Exposure
The toxic effects observed following single or repeated exposure to a chemical
are often quite different...

1. Acute Effects: rapidly developing, reaching a maximum with severe

symptoms.
(exposure to high doses of CN- will kill within a few minutes..... )

2. Subacute Effects: symptoms generally not as severe, but toxic effects often
same as acute....

3. Chronic Effects: progresses at a slow and varying rate; may be mistaken

for other diseases. Often difficult to determine cause-and-effect unless in
laboratory....
 asbestos-caused cancer may be delayed 20-30 years....
 acute exposure may result in either acute or chronic effects....
(acute exposure to asbestos may lead to cancer....)
 chronic exposure may result in either chronic or acute effects...
(chronic lead exposure may lead to subacute or acute symptoms....)
4. Accumulative Effects: occurs two ways...
 accumulation of toxin: exposure to heavy metals (lead,
mercury) that have long half-lives result in disease due to
metal accumulation....
 accumulation of effect: low level exposure to
organophosphate pesticides depresses acetylcholine
esterases to a point where symptoms occur....
5. Delayed Effects: effect may occur only after long
exposure; agent cannot be found in blood or tissues.
Damage to system already done....
 radiation sickness
 Chemical Interactions
Because of the large number of different chemicals an individual
may come in contact with at any given time (workplace, drugs,
diet, hobbies, etc.), it is necessary, in assessing the spectrum of
responses, to consider how different chemicals may interact with
each other. Interactions can occur in a variety of ways.
Additive – combined effect is the same as the sum of effects when
given alone. Ex. OPs
Synergistic - combined effects are much greater than the sum of
effects when given alone Ex. CCl4 and EtOH
Potentiation – exposure to a chemical with no toxicity increases
the toxicity of another compound Ex. CCl4 and isopropanol
Antagonism – co-administration of two chemicals interferes with
the toxicity of both or one of them Ex. Antidotal therapies
 Antagonism
Functional – Chemicals counterbalance each other by exerting
opposite effects on a physiological function. Ex. Convulsions
treated with benzodiazepines

Chemical (or inactivation) – Chemical reaction between two

compounds leads to less of the toxic compound. Ex. Chelators
and metals Ex. Antivenins

Dispositional – Disposition of toxic chemical is changed so that

concentration and/or duration is diminished. Ex. Ipecac, charcoal,
pH alteration, metabolism induction or inhibition

Receptor – Chemicals compete for the same receptor, decreasing

effective binding of toxic compound. Ex: Naloxone and morphine
Ex: Tamoxifen and estradiol Ex: OPs Atropine and AChR
 Tolerance

A state of decreased responsiveness due to a prior exposure to

the same or a structurally similar chemical in an individual

Dispositional Tolerance – A decreased amount of chemical

reaches the site where the effect is produced Ex. CCl4 –
metabolism inhibition Ex. Cd and metallothionein

Receptor Tolerance – Same amount of chemical reaches the site,

but target receptor response decreased Ex. OPs and muscarinic
AChR
 Resistance

A change in the susceptibility to a chemical at the population

level.

A selective process (evolution) by which sensitive individuals

do not survive and only those with agenetic trait that
accommodates the chemical survive.

Subsequent selection through numerous generations fixes the

trait, stabilizing the resistant population.
 Dose-Response Relationships

„Dose: The amount of agent actually deposited within the body.

Response: The biological response to an agent.
Since the basic question in toxicology is how dose is related to
toxicity, most toxicology studies are designed to investigate, how
living creatures react as doses vary from high to low levels.
Then at various intervals, the animals are examined for the presence
or absence of effects.
These effects may be behavioral changes, alterations in the
compositions of body tissues or fluids (example blood, serum, urine,
etc), or structural changes in parts of the organism.
Toxicity is a relative phenomenon and that it depends not only on
the toxic properties but on the dose of the compound
administered. This relationship between the dose of a compound
and the response it elicits is a fundamental concept in toxicology.
The interpretation of the dose response relationship is based on
certain assumptions:
1. the response is proportional to the concentration at the target
site;
2. the concentration at the target site is related to the dose;
3. the response is causally related to the compound administered.
Dose-Response Relationships
Fundamental relationship that brings together the dose of a
chemical and the resulting effect.

There are two types of dose–response relationships:

 the individual dose–response relationship, which describes the

response of an individual organism to varying doses of a chemical,
often referred to as a “graded” response because the measured
effect is continuous over a range of doses.
 a quantal dose–response relationship, which characterizes the
distribution of individual responses to different doses in a
population of individual organisms.
Graded Dose-Response – the Individual Level
• Individual dose–response
relationships are characterized by a
dose related increase in the severity
of the response.
• The dose relatedness of the response
often results from an alteration of a
specific biochemical process.
• For example, Fig in this slide shows
the dose–response relationship
between different dietary doses of the
organophosphate insecticide
chlorpyrifos and the extent of
inhibition of two different enzymes in
the brain and liver:
acetylcholinesterase and
carboxylesterase.
 Exposure to ethanol
Graded responses between no effect and death
Information obtained from graded dose
response Curve (DRC):
 The position of graded log dose response curve (DRC)
as the index of the drug/agent potency.

 The height of DRC curves serves as an index of drug

efficacy. The more the height of DRC, the more the
efficacy of the drug/agent.

 The gap between the therapeutic effect of DRC and the

adverse effect DRC indicates the safety margin of the
therapeutic range.
At the Population Level – The Quantal Dose-Response

• In contrast to the “graded” dose–response relationship

that occurs in individuals, the dose–response relationships
in a population are by definition quantal—or “all or
none”—in nature.
• That is, at any given dose, an individual in the population is
classified as either a “responder” or a “nonresponder.”
• Although these distinctions of “quantal population” and
“graded individual” dose–response relationships are useful,
the two types of responses are conceptually identical.
• The ordinate in both cases is simply labeled the response,
which may be the degree of response in an individual or
system or the fraction of a population responding, and the
abscissa is the range in administered doses.
 The Normal Distribution

•The bars represent the percentage of animals that responded at each dose.
•One can clearly see that only a few animals responded to the lowest dose and the
highest dose.
•Larger numbers of animals responded to doses intermediate between these two
extremes, and the maximum frequency of response occurred in the middle portion
of the dose range.
•Thus, we have a bell-shaped curve known as a normal frequency distribution.
•The reason for this normal distribution is that there are differences in
susceptibility to chemicals among individuals; this is known as biological
variation.
 Cumulative Frequency Distribution

•If the numbers of individuals responding at each consecutive

dose are added together, a cumulative, quantal dose– response
relationship is obtained.

•Frequency converted to cumulative gives sigmoid curve

Information obtained from quantal dose-
response curve:
 The ED50 for a specialized quantal effect can serve
as an index of drug potency.

 Slope of the curve is an index of the potential

variability among the responding individuals.

 The ratio of LD50 and ED50 (the therapeutic index) is

an index of the safety margin of the drugs.
 Determination of LD50

• Chemicals is administered to mice or rats (orally/ip) at several doses

(usually 4/5) in the lethal range
• Draw a dose response curve (% of death vs dose in log scale) and mid
point represent LD50
• To linearize data, response (death) is converted to probits (probability
unit)
50% response = 5 probit
16% ,, = 4 ,,
84% ,, = 6 ,,
• A plot of % of population responded in probit units, against log dose
yields a straight line.
• The LD50 is determined by drawing a vertical line from the point of line
where the probit unit = 5 (50% mortality
LD50
 APPROXIMATE ACUTE LD50 s(mg/Kg) of Some
Rep.Chemical Agents
Sodium Chloride 4,000
Ferrous Sulfate 1,500
Morphine Sulfate 900
Phenobarbital Sodium 150
Picrotoxin 5
Strychnine Sulfate 2
Nicotine 1
d-tubocurarine 0.5
Tetrodotoxin 0.10
Digoxin 0.001
Botulinum toxin 0.00001
 Disadvantages of LD50

• A crude index of toxicity not reflecting other

side effects

• A large number of animals required

• A large variation occurs

• Very little information concerning chronic

toxicity
 Evaluating the Dose–Response Relationship

Therapeutic Index (TI)

The therapeutic index (TI) in its broadest sense is defined as the ratio of
the dose required to produce a toxic effect and the dose needed to elicit
the desired therapeutic response.
The therapeutic index of a drug is an approximate statement about the
relative safety of a drug.
Larger the ratio, the greater the relative safety.
LD50
TI = in animal studies, or for humans,
ED50
TD50
TI =
ED50
Significance of TI:

• It provides crude measure of the safety of a drug, more

TI, more safety (eg Antibiotic, TI=4 & anti-cancer, TI=2).
• For evaluation of new drugs TI has greater significance.

Limitations:

• It is based on animal toxicity data, which may not reflect

forms of toxicity that are important clinically.
• It takes no account of idiosyncratic reactions.
 Evaluating the Dose–Response Relationship

Margins of Safety is the ratio of TD1 verses ED99

An index of a drug's effectiveness and safety. It is calculated as the amount
of drug that is toxic/lethal to 1% of animals (TD1/LD1) divided by the
amount of drug that causes a beneficial effect in 99% of the animals (ED99)
.

For many chemicals and effects there will be a dose below which no
effect or response is observed. This is known as the threshold dose.
The chronicity index of a chemical is a unitless value obtained by
dividing its 1-dose TD50 by its 90-dose (90-day) TD50, with both
expressed in milligrams per kilogram per day.

Potency versus Efficacy

To compare the toxic effects of two or more chemicals, the dose

response to the toxic effects of each chemical must be
established. One can then compare the potency and maximal
efficacy of the two chemicals to produce a toxic effect.

• Efficacy: Maximum effect that a drug

can produce regardless of dose. Is the
ability of a drug to produce the desired
therapeutic effect.
• Potency: Amount of a drug that is
needed to produce a given effect.
NOAELs/The No Observed Adverse Effect Level
A dose, below which, the probability of an individual responding is
zero.

Dose - Response Curve

100

90
80

70

60
% deaths

50
40

30

20
LOAEL
10 LD50
0
1 10 100 1000 10000
Dose (mg/kg)

NOAEL
The NOAEL is important for setting exposure limits. For
example, the acceptable daily intake (ADI) is based on the
NOAEL. This is a factor used to determine the safe intake
for food additives and contaminants such as pesticides
and residues of veterinary drugs and, hence, to establish
the safe level in food.
The ADI is determined by the use of a suitable safety
factor which may be any factor up to 1000, but is usually
100:
 Dose-Response Curves for Essential Compounds

Hormesis – is a dose response phenomenon

characterized by a low dose stimulation, high
dose inhibition, resulting in an inverted U-
shaped dose response.

For example, for substances that are required

for normal physiologic function and survival
(e.g., vitamins and essential trace elements
such as chromium, cobalt, and selenium), the
shape of the “graded” dose–response
relationship in an individual over the entire
dose range is actually U-shaped.
 Using Dose-Response Curves

Dose-response curves provide valuable information to the

toxicologist. Examination of these curves and their supporting data
provides a basis for comparing pesticide toxicity threshold levels as
well as medium (LD ) and maximum effective doses.
50

The slope of the dose-response curve is of critical importance. A

steep curve indicates only a slight difference between a nontoxic
dose and a toxic dose; in such a case, even a small increase in dose
produces a significant change of effect. Conversely, a somewhat flat
dose-response curve indicates that a relatively large increase in dose
has little effect.
The assessment of toxic substances,

A toxicity assessment provides an estimate of how much of

a substance causes what kind of harm. Toxicity
assessment is a major component of risk assessment. A
toxicity assessment is a tool to investigate the potential for
a substance to cause harm--and how much causes what
kind of harm.
A human health risk assessment includes four steps, which
begin with planning:
Planning - Planning and Scoping process. ...
Step 1 - Hazard Identification. ...
Step 2 - Dose-Response Assessment. ...
Step 3 - Exposure Assessment. ...
Step 4 - Risk Characterization.