Transcript Lecture 2 ppt - Professor Monzir Abdel

Poisons and Drugs
Prof. Monzir S. Abdel-Latif
Chemistry Department
Islamic University of Gaza
http://www.monzir-pal.net
Areas of Toxicology
Mechanistic Toxicology

A mechanistic toxicologist is concerned with
identifying and understanding the mechanisms by
which chemicals exert toxic effects on living
organisms. For example, the relative toxic potential
of organophosphate insecticides in humans,
rodents, and insects can be accurately predicted on
the basis of an understanding of common
mechanisms (inhibition of acetylcholinesterase).
Descriptive Toxicology

A descriptive toxicologist is concerned directly with
toxicity testing, which provides information for safety
evaluation and regulatory requirements. The appropriate
toxicity tests in cell culture systems or experimental
animals are designed to yield information to evaluate risks
posed to humans and the environment from exposure to
specific chemicals. Toxicologists in the chemical industry,
however, must be concerned not only with the risk posed
by a company’s chemicals (insecticides, herbicides,
solvents, etc.) to humans but also with potential effects on
fish, birds, and plants, as well as other factors that might
disturb the balance of the ecosystem.
Regulatory Toxicology

A regulatory toxicologist has the responsibility for
deciding, on the basis of data provided by
descriptive and mechanistic toxicologists, whether
a drug or other chemical poses a sufficiently low
risk to be marketed for a stated purpose and the
subsequent human or environmental impacts
resulting from its use.
Some Regulatory Bodies in USA

In the USA, the Food and Drug Administration (FDA) is responsible
for allowing drugs, cosmetics, and food additives to be sold in the
market according to the Federal Food, Drug and Cosmetic Act
(FFDCA). The U.S. Environmental Protection Agency (EPA) is
responsible for regulating most other chemicals according to the
Federal Insecticide, Fungicide and Rodenticide Act (FIFRA), the
Toxic Substances Control Act (TSCA), the Resource Conservation
and Recovery Act (RCRA), the Safe Drinking Water Act, and the
Clean Air Act. In 1996, the U.S. Congress passed the Food Quality
Protection Act (FQPA) which fundamentally changed the pesticide
and food safety laws under FIFRA and FFDCA requiring stricter
safety standards particularly for infants and children, who were
recognized as more susceptible to health effects of pesticides.
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. The expertise
of forensic toxicologists is invoked primarily to aid
in establishing the cause of death and determining
its circumstances in a postmortem investigation.

Clinical toxicology designates an area of
professional emphasis in the field 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. Although toxicologists
concerned with the effects of environmental
pollutants on human health fit into this definition, it
is most commonly associated with studies on the
impacts of chemicals on nonhuman organisms such
as fish, birds, animals, and plants.
Local versus systemic effects
Adverse effects may be local or systemic. Local
effects occur at the site of exposure of the
organism to the potentially toxic substance.
Corrosives always act locally. Irritants frequently
act locally. Most substances that are not highly
reactive are absorbed and distributed around the
affected organism causing general injury at a target
organ or tissue distinct from the absorption site.
Accumulation Site and Affected Organ
The target organ is not necessarily the organ of
greatest accumulation. For example, adipose (fatty)
tissue accumulates organochlorine pesticides to
very high levels but does not appear to be affected
by them. Some substances produce both local and
systemic effects. For example, tetraethyl lead
damages the skin on contact, and is then absorbed
and transported to the central nervous system
where it causes further damage.
Effects of a chemical can accumulate even if the
chemical itself does not. There is some evidence
that this is true of the effects of organophosphate
pesticides and other neurotoxins on the nervous
system. This may lead to poor functioning of the
nervous system in humans in old age. Because of
the time difference between exposure and effect,
establishing the relationship between such delayed
effects and the toxin is often difficult.
Allergy
Adverse effects related to allergies appear to be increasing.
Allergy (hypersensitivity) is the name given to disease
symptoms following exposure to a previously encountered
substance (allergen) that would otherwise be classified as
harmless. Essentially, an allergy is an adverse reaction of
the altered immune system. The process, which leads to
the disease response on subsequent exposure to the
allergen, is called sensitization.
Regenerative Versus Non-regenerative
Tissue
Another important aspect of adverse effects to be considered
is whether they are reversible or irreversible. For the liver,
which has a great capacity for regeneration, many adverse
effects are reversible, and complete recovery can occur.
For the central nervous system, in which regeneration of
tissue is severely limited, most adverse effects leading to
morphological changes are irreversible and recovery is, at
best, limited. Carcinogenic effects are also irreversible, but
suitable treatment may reduce the severity of such effects
CHARACTERISTICS OF
EXPOSURE

To characterize fully the potential hazard of a
specific chemical agent, we need to know not only
what type of effect it produces and the dose
required to produce that effect but also information
about the agent, the exposure, and its disposition in
the subject. Two major factors that influence
toxicity as it relates to the exposure situation for a
specific chemical are the route of exposure, the
duration, and frequency of exposure.
Route and Site of Exposure

The major routes (pathways) by which toxic agents gain
access to the body are the gastrointestinal tract (ingestion),
lungs (inhalation), skin, and other parenteral (other than
intestinal canal) routes. Toxic agents generally produce the
greatest effect and the most rapid response when given
directly into the bloodstream (the intravenous route). An
approximate descending order of effectiveness for the
other routes would be inhalation, intraperitoneal,
subcutaneous, intramuscular, intradermal, oral, and
dermal.
Duration and Frequency of Exposure


Toxicologists usually divide the exposure of experimental
animals to chemicals into four categories: acute, subacute,
subchronic, and chronic.
Acute exposure is defined as exposure to a chemical for
less than 24 hours. Whereas acute exposure usually refers
to a single administration, repeated exposures may be
given within a 24-hours period for some slightly toxic or
practically nontoxic chemicals. Acute exposure by
inhalation refers to continuous exposure for less than 24
hours, most frequently for 4 hours.
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Repeated exposure is divided into three categories:
subacute, subchronic, and chronic.
Subacute exposure refers to repeated exposure to a
chemical for 1 month or less.
Subchronic exposure refers to repeated exposure to a
chemical for 1 to 3 months, and
Chronic exposure refers to repeated exposure to a
chemical for more than 3 months, although usually this
refers to studies with at least 1 year of repeated dosing.
Toxicity testing
Dose response and Concentration response
A dose–response (concentration–response)
relationship is defined as the association between
dose (concentration) and the incidence of a defined
biological effect in an exposed population, usually
expressed as percentage. Historically the defined
effect was death.
The classic dose–response or concentration–response
relationship is shown in the figure below. This is a
theoretical curve and in practice such a curve is
rarely found. Curves of this kind form the basis for
the determination of the LD50 or the LC50 (the
median lethal concentration). The LD50 and LC50
are specific cases of the generalized values LDn
and LCn. The LDn is the dose of a toxicant lethal to
n% of a test population. The LCn is the exposure
concentration of a toxicant lethal to n% of a test
population.
the LD50 is the statistically derived single dose of a
chemical that can be expected to cause death in
50% of a given population of organisms under a
defined set of experimental conditions. Similarly,
the LC50 is the statistically derived exposure
concentration of a chemical that can be expected to
cause death in 50% of a given population of
organisms under a defined set of experimental
conditions.
Approximate Acute LD50s of Some Representative Chemical Agents
agent
LD50, mg/kg∗
Ethyl alcohol
10000
Sodium chloride
4000
Ferrous sulfate
1500
Morphine sulfate
900
Phenobarbital sodium
150
Picrotoxin
5
Strychnine sulfate
2
Nicotine
1
Tetrodotoxin
0.10
Dioxin (TCDD)
0.001
Botulinum toxin
0.00001
Workplace Hazardous Materials
Information System (WHMIS)
D1A- Poisonous product that causes serious and
immediate effects.
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Oral LD50 < 50 mg/kg
Dermal LD50 < 200 mg/kg
Inhalation LC50 < 2500 ppm 4 hours (gas)
Inhalation LC50 < 1500 ppm 4 hours (vapor)
and a saturated vapor concentration > 2 times the LC50
value.
Inhalation LC50 < 0.5 mg/l or < 500 mg/m3 4 hours (dust,
mist, fume)
D1B- Toxic material that causes serious and immediate
effects .

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Similar criteria is used as for the D1A. The criteria is as
follows :
Oral LD50 > 50 mg/kg < 500 mg/kg
Dermal LD50 > 200 mg/kg < 1000 mg/kg
Inhalation LC50 > 1500 ppm < 2500 ppm 4 hours (vapor)
Inhalation LC50 > 0.5 mg/l < 2.5 mg/l 4 hours
or > 500 mg/m3 < 2500 mg/m3 4 hours(Dust, Fumes or
Mist(
Another important value that may be derived from
the relationship shown is the threshold dose or
concentration, the minimum dose or concentration
required to produce a detectable response in the
test population. The threshold value can never be
derived with absolute certainty and therefore the
lowest observed effect level (LOEL) or the NOEL
have normally been used instead of the threshold
value in deriving regulatory standards.
The use of the LD50 in the classification of
potentially toxic chemicals has been described; it
must be emphasized that such a classification is
only a very rough guide to relative toxicity. The
LD50 tells us nothing about sub lethal toxicity. Any
classification based on the LD50 is strictly valid
only for the test population and conditions on
which it is based and on the related route of
exposure.
LD50 as a Measure of Toxicity
The LD50 tells us nothing about the shape of the
dose–response curve on which it is based. Thus,
two chemicals may appear to be equally toxic since
they have the same LD50, but one may have a
much lower lethal threshold and kills members of
an exposed population at concentrations where the
other has no effect
Fate of the LD50
At their November meeting in 2001, the OECD Joint
Meeting of the Chemicals Committee and Working
Party on Chemicals, Pesticides and Biotechnolgy,
agreed that the LD50 for acute oral toxicity, known
as OECD Test Guideline 401 and heavily criticised
by the animal welfare protectorate shall be
abolished and deleted from the OECD manual of
internationally accepted Test Guidelines.
The determination and use of the LD50 are likely to
decline in future as fixed dose testing (The Fixed
Dose Procedure (FDP), proposed in 1984)
becomes more widely used. In fixed dose testing,
the test substance may be administered to rats or
other test species at no more than four dose levels:
the possible dose levels are preset legally to equate
with a regulatory classification or ranking system.
Dosing is followed by an observation period of 14
days. The dose at which toxic signs are detected is
used to rank or classify the test materials.
FDM versus LD50
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1.
2.
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The new OECD FDM (Test Guideline 420) departs
from the conventional method (Test Guideline 401) in
two principal ways:
FDM considers non-lethal endpoints; and
it does not develop a quantitative relationship between
dose and lethality .
The method relies upon clear signs of toxicity
("evident toxicity"), a non-lethal endpoint as well as
lethality, as the basis for acute oral toxicity
assessment.
Acute Oral Toxicity - Fixed Dose
Procedure

This Guideline is intended primarily for use with rat. Groups of
animals of a single sex (normally females) are dosed in a stepwise
procedure using the fixed doses of 5, 50, 300 and 2000 mg/kg
(exceptionally 5000 mg/kg). The initial dose level is selected on the
basis of a sighting study as the dose expected to produce some signs
of toxicity without causing severe toxic effects or mortality. Further
groups of animals may be dosed at higher or lower fixed doses,
depending on the presence or absence of signs of toxicity or
mortality. This procedure continues until the dose causing evident
toxicity or death is identified, or when no effects are seen at the
highest dose or when deaths occur at the lowest dose. The test
substance is administered in a single dose by gavage using a
stomach tube or a suitable intubation canula. Animals should be
fasted prior to dosing. A total of five animals of one sex will
normally be used for each dose level investigated.
Fixed Dose testing

In this procedure the test substance is given at one of the
four fixed-dose levels (5, 50, 300, and 2000 mg/kg) to five
male and five female rats. The objective is to identify a
dose that produces clear signs of toxicity but no mortality.
Depending on the results of the first test, either no further
testing is needed or a higher or lower dose is tested: If
mortality occurs, retesting at a lower dose level is
necessary (except if the original dose chosen is 5 mg/kg).
If no signs of toxicity occur at the initial dose, it is
necessary to retest at a higher dose level. The results are
thus interpreted in relation to animal survival and evident
toxicity and it becomes possible to assign the chemical to
one of the OECD classification categories.
Fixed dose testing (no mortality)
The initial test dose level is selected with a view to
identifying toxicity without mortality occurring.
Thus, if a group of five male and five female rats is
tested with an oral dose of 300 mg/ kg body weight
and no clear signs of toxicity appear, the substance
should not be classified in any of the defined
categories of toxicity. If toxicity is seen but no
mortality, the substance can be classed as
‘harmful’.
If mortality occurs, retesting with a dose of 50 mg/
kg body weight is required. If no mortality occurs
at the lower dose but signs of toxicity are detected,
the substance would be classified as ‘toxic’. If
mortality occurs at the lower dose, retesting at 5
mg/ kg body weight would be carried out and if
signs of toxicity were detected and mortality
occurred, the substance would be classified as
‘very toxic’.
Again: Fixed Dose Versus LD50

In an international study involving 33 laboratories in 11 countries,
the acute oral toxicity to the rat of 20 substances and preparations
was evaluated using a fixed-dose procedure and the results
compared with those obtained for the test materials using the
classical LD50 test. The study has shown that the fixed-dose
approach to acute oral toxicity testing: (1) produces consistent
results that are not substantially affected by inter-laboratory
variations; (2) provides adequate information for risk assessment
purposes on signs of toxicity, including their nature, time to onset,
duration and outcome; (3) uses fewer animals than the current
internationally agreed OECD procedure; (4) subjects animals to less
pain and distress than the classical LD50 test and causes less
compound-related mortality.
Workplace Hazardous Materials
Information System (WHMIS)

In order to apply this protocol to classification under
WHMIS, the following interpretations should be
applied:
1.
Where mortality in the FDM is 50% at one of the fixed
dose levels, it can be assumed that the LD 50 for the
controlled product, when tested in accordance with
OECD 401, would likely fall below that fixed dose level.
Consequently, it follows that:
Where mortality in the FDM is 50%
A.
at a dose of 5 or 50 mg/kg b.w. one would classify the controlled
product in D1A.
B.
at a dose of 500 mg/kg b.w. one would classify the controlled
product in D1B, provided there was no data available to show that
lethality at 50 mg/kg b.w. exceeded 50%.
C.
at a dose of 2000 mg/kg b.w., one would need to review test data
at the next lower dose level (500 mg/kg) to determine if lethality
at that level is 50%.
Where there is 100% survival with or
without evident toxicity
a. at a dose of 5 or 50 mg/kg b.w. one would need to
review test data at the next highest dose level (50 or 500
mg/kg b.w.) to determine if lethality at that level is ³ 50%.
b. at a dose of 500 or 2000 mg/kg b.w. the LD 5o is not
very likely to be 500 mg/kg b.w., therefore the product
would not be classified for acute oral toxicity under
WHMIS Class D1.
3. Where mortality is < 50% but > 0% at one of the
fixed dose levels, it can be assumed that in most
cases, the LD 50 value for the substance, when
tested in accordance with OECD 401, would fall
above that fixed dose level. Consequently, it
follows that:
Where mortality is < 50% but > 0% at
one of the fixed dose levels
a. at a dose of 5 mg/kg b.w. - one would need to
review the lethality data for the next highest fixed
dose (50 mg/kg) to determine the % lethality at that
level. If mortality at the next fixed dose level is
50% the controlled product would be classified
D1A.
Where mortality is < 50% but > 0% at
one of the fixed dose levels
b. at a dose of 50 mg/kg b.w. - one would need to review the
lethality data for the next highest fixed dose (500 mg/kg
b.w.) to determine the % lethality at that level. If mortality
at the next fixed dose level is ³ 50% the controlled product
would be classified in D1B, as described in 1(b), above.
However, if lethality at 50 mg/kg b.w. is 40% (i.e. if the
data are pooled and 4/10 animals died, especially if the
majority of them were of one sex) and no data are
available at 500 mg/kg b.w., the conservative approach
would be to assume that the LD 50 value for the substance
might be below 50 mg/kg b.w., and consequently it would
be prudent to classify in D1A.
Where mortality is < 50% but > 0% at
one of the fixed dose levels
c. at a dose of 500 mg/kg b.w. - if lethality at 500 mg/kg b.w.
is 40% (i.e. 4/10 animals died) and no data are available at
the next highest fixed dose (2000 mg/kg b.w.), the
conservative approach would be to assume that the LD 50
value for the substance might be below 500 mg/kg b.w.,
and consequently it would be prudent to classify in D1B.
If lethality at 500 mg/kg b.w. was lower than 40%, it
would be reasonable to assume that the substance is not
classified under WHMIS class D1.
d. at a dose of 2000 mg/kg b.w. - not classified for acute oral
toxicity under WHMIS Class D1.
Acute Toxic Class Method

1.
2.
3.
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The test consists of a stepwise procedure with the use of three animals of a single
sex (normally females) per step. Absence or presence of compound-related
mortality of the animals dosed at one step will determine the next step
no further testing is needed,
dosing of three additional animals, with the same dose
dosing of three additional animals at the next higher or the next lower dose level.
The starting dose level is selected from one of four fixed levels (5, 50, 300, and
2000) and it should be that which is most likely to produce mortality in some of
the dosed animals. When there is no information on a substance to be tested, it is
recommended to use the starting dose of 300 mg/kg. This procedure is
reproducible, uses very few animals and is able to rank substances in a similar
manner to the other acute toxicity testing methods.
Acute Oral Toxicity: Up-and-Down
Procedure
The test consists of a single ordered dose progression in which animals
of a single sex (normally females) are dosed, one at a time. The first
animal receives a dose step below the level of the best estimate of
the LD50 (when no information is available to make a preliminary
estimate of the LD50, the suggested starting dose is 175 mg/kg). If
the animal survives, the dose for the next animal is increased by a
factor of 3.2 times the original dose; if it dies, the dose for the next
animal is decreased by a similar dose progression. The preferred
species is the female rat and the test substance is administered in a
single dose by gavage (if not possible the dose may be given in
smaller fractions over a period not exceeding 24 hours).