Tahar AbdAlaziz Suliman MD, PhD

Clinical Toxicology & Neurology (Pediatric)

 Toxicology
• The study of poisons
• Poisons are chemical/physical
agents that produce adverse
responses in biological organisms
Any substance can be toxic if
introduced in a dose capable of
disturbing the normal physiological
homeostasis of the exposed body.
OTHER TERMS
Toxicants: are toxic substances
from chemicals

• Toxins: are poisonous substance

produced within living cells or
organisms
– Venom
Toxicology
The science of poisons that studies toxic
substances with respect to their:
• Sources
• Properties
• Mechanism of toxicity
• Toxic effects
• Detection
• Clinical manifestations
• Management
Sources
• Chemical Source: the commonest
source e.g. drugs, corrosives
• Plant Source: e.g. hashish,
cocaine
• Animal Source: the least but most
serious source. Venomous animals
such as scorpions, spiders,
snakes, wasps.
 Venomous & Poisonous Animals
Venomous animals deliver or inject venom
into other organisms, using a specialized
apparatus of some kind (usually fangs or a
stinger). The venom is produced in a gland
attached to this apparatus.
Poisonous animals do not deliver their
toxins directly. The entire body, or large
parts of it, may contain the poisonous
substance. These organisms may be
harmful when eaten or touched.
Wasp sting (VENOM)
Androctonus bicolor
Cerastes vipera
Tetradotoxin in Puffer Fish
 Sites of Toxic Actions
• Local (non-specific):
Wherever the poison contacts the
biological system it starts its harmful
effects. It does not require specific site
or receptor to elicit its effects such as
toxicity by acids or alkalis.
• Remote (systemic):
The poison affects a system far from its
portal of entry.
• Local & Remote:
The poison has the capacity of acting
locally and systematically.
Oxalic acid is an example of these poisons.
 Duration and frequency of exposure
• Acute: application of a single or short-
term (less than a day) dosing of a
substance to cause toxicity
• Sub-acute: toxicity is expressed after
repeated applications for a duration
less than half-life expectancy of the .
• Chronic: Expression of toxic symptoms
only after repeated exposure to a
chemical in doses regularly applied to
the organism for a time greater than
half of its life-expectancy
Chronicity
Index
•The ratio of the acute to
chronic LD50 dosage

• Compounds with strong cumulative

properties have larger chronicity
index.
 Types of Toxic
Mechanisms
• Direct: the poison itself can cause toxic
effects as in corrosives.
• Indirect: toxicity results from the
interaction of the poison with the biological
activity within biological system.
– Binding to cell membrane to change in their function or
structure thus affecting their normality.
– Interference with enzymatic actions.
– Formation of metabolites which are more toxic than the
parent poison.
– Effects on DNA
 Classification of toxic
agents
1) According to the Target organ they are
acting on it (hepatotoxic, Nephrotoxic)
2) According to their Use (food additive,
drug, pesticide)
3) According to their Source (animal or
plant)
4) According to their Effects (carcinogen,
mutagen)
5) According to their Physical state
(gas, liquid)
6) According to their Chemistry
(Amine, hydrocarbon)
7) According to their Poisoning
potentiality (extremely toxic, slight
toxic, etc)
8) According to their Biochemical
mechanism of action (alkylating
agent, AchE inhibitor)
 Factors Affecting Action of Poison
A) Factors related to the poison:
• Dose: a basic principle in toxicology.
– Dose is the amount of chemical that comes into
contact with the body or gets inside the body.
– The increase of dose will increase the severity
of toxicity.
• Physical status: gaseous state is more toxic
than liquid state than the solid state.
• Purity: this depends on the impurity of the
poison; if the impurities are more toxic
than the poison, the toxicity will be more
and vice versa.
B) Factors related to the
individual:
– Age
– Health
– Sensitivity
– Sex
C) Factors related to mode of
exposure:
– Inhalation > IM > ingestion > Skin contact
D) Factors related to
 Forensic
• Forensic Toxicology
toxicologist is concerned with
the detection and estimation of poisons
for legal purposes:

– Tissues and body fluids obtained at

autopsy
– Blood, urine, or gastric material
obtained from a living person
•Poisoning as a cause of death can be proven only with
toxicologic analyses that demonstrate the presence of the
poison in the tissues or body fluids of the deceased.

•Presence of poisons can be demonstrated

only by chemical methods of isolation and
identification.

•If toxicological analyses are avoided, death

may be ascribed to poisoning without definite
proof.
Analytical
toxicology
deals with the detection,
identification, and
quantification of poisons.
 SAMPLES REQUIRED FOR TOXICOLOGICAL
ANALYSIS FROM AUTOPSIES
•Blood
– The best place at autopsy is from femoral & iliac veins,
then axillary veins in consequence
• NO SAMPLES FROM:
1. Jugular veins: may be contaminated by reflux
from upper thorax
2. General body cavity: highly contaminated by
intestinal contents
3. Heart or great vessels in chest: postmortem
diffusion of drugs & alcohol from the stomach
or aspirated vomit contaminate these sites
•Urine
20-30 ml urine in sterile
container without preservatives
•Faeces
Used in heavy metals as arsenic,
lead, mercury
• Vomit and Stomach contents
• Organs
– The most common organ saved for
analysis is liver
– Bile can be helpful in morphine and
chlorpromazine
– Lungs in some cases of solvents
•Hair & Nails
– Heavy metal poisonings
– Recently, prolonged use of opiates
 METHODS OF ANALYSIS
• QUALITATIVE METHODS

A) COLOR TEST
This is a rapid, easily performed,
qualitative, screening test, but not
specific method
• Can be used as bed side rapid test
• Examples:
1- Ferric chloride test for
salicylates (pink-purple)
2- Zwikker test for barbiturates
(purple color)
3- Formaldehyde-Sulfuric acid test
for BZD (orange)
4- Mandalin Test for opioid (brown
color)
B) CHEMICAL TEST
• Reinsch Test is an initial indicator
to detect the presence of one or
more of the following Heavy Metals
in a biological sample
– Antimony
– Arsenic
– Bismuth
– Selenium
– Thallium
– Mercury
• QUANTITATIVE METHODS
A) CHROMATOGRAPHY
1- Thin-Layer Chromatography (TLC):
Mobile phase (a mixture of organic solvents such
as chloroform, and methanol) is run across a
Stationary phase (silica gel spread on a glass plate).
2- Gas Chromatography- Mass spectrometry:
Stationary phase is a liquid and the mobile phase (a
carrier gas) is an inert gas such as helium or
nitrogen.
Spotting Running
3- High-Performance Liquid Chromatography (HPLC):
In HPLC the stationary phase is a column packed with
solid particles and the mobile phase is a liquid solvent.

B) IMMUNOASSAYS
• Enzyme-multiplied immunoassay technique (EMIT).
frequently used to detect the presence of certain drugs
in urine.
• Polarization immunoassay (FPIA)
• Radioimmunoassay (RIA)
 HPLC
System
Column
Thin-Layer Chromatography (TLC)
This is a separation technique .
It was described in details by Ergon Stahl
(1969).
In TLC a mobile phase (a mixture of
organic solvents such as chloroform,
and methanol) is run across a
stationary phase (silica gel spread on a
glass plate).
 TLC
The samples to be analyzed are spotted
near the bottom portion of the plate
and allowed to dry. Then the plate is
placed upright into a chamber, with the
bottom of the plate (where the sample
has been spotted) in contact with the
mobile phase. The mobile phase will
then draw up across the plate by
capillary action.
As the solvent moves past the samples, the
components of the samples will migrate,
with the speed of migration dependent
upon the relative affinity of the
components for the mobile phase
compared to the stationary phase.
When the leading edge of the solvent
reaches the top of the plate, it is removed
from the solvent and allowed to dry. The
location of the sample components can
then be visualized.
Stahl provided methods for 264 stains
or dyes that can be applied for the
required component such as
ninhydrin will react with
amphetamine to give pink color.
Alternatively, a fluorescent dye can be
incorporated in the solid phase, so
that ultraviolet light can reveal the
sample components as dark spots
against the a bright background.
The results of TLC can be quantified by
using the retention factor (Rf)
which is the ratio of the distance that
a sample component moves to the
distance that the leading edge of the
solvent moves.
Rf = Sample distance movement
Solvent distance movement
Sample components can be identified
by comparing their Rf to the Rf of
known substances or by using a table
of Rfs given according to the mobile
and stationary phases used.
Gas Chromatography- Mass
spectrometry
The stationary phase is a liquid
The mobile phase (a carrier gas) is an inert gas
such as helium or nitrogen.
There are 2 types of columns used in GC:
• Packed column: the liquid is coated onto
particles packed into a stainless steel or
glass column.
• Capillary column: the liquid is coated onto
the walls of the column itself , which is
narrow and made of glass.
• Samples are injected into a heated port,
where they are vaporized and carried
into the column along with the carrier
gas. A detector then produces a signal as
sample components exit the column.
• When the detector is hooked up to a
recorder, a gas chromatogram
(recording) can be produced. This is a
plot of electronic signal versus time,
which shows series of peaks that
corresponds to the components of the
sample.
• The time it takes for a substance to pass
thorough a column (retention time Rt) can be
compared to standards to identify that
substance. Also, since the area under each
peak is proportional to the concentration of
that substance, comparison with a standard of
known concentration allows estimation of the
concentrations.
• Flame photometric detectors increase the
sensitivity over flame ionization.
• Electron capture detector uses radioactive
source and it can detect picograms of DDT due
to presence of 5 chlorine atoms in the
molecule.
Mass Spectrometry (MS)
Although chromatography allows
identification of substances based on Rf
and Rt with standards, definitive
identification requires additional analysis.
Mass spectrometry is one of these
methods; it is used in combination with
gas chromatography. As the sample
components exit the GC column, they are
routed into a vacuum chamber in the
mass spectrometer, where they are hit
with a beam of electrons.
This knocks electrons off the sample
molecules, creating positive electrons
and breaking them into fragments.
These fragments are then passed
through an electromagnetic field,
which separates them by their
mass/charge ratio. The resulting
spectrum plotting the abundance and
mass/ charge ration of each fragment
is specific for a given substance.
High-Performance Liquid Chromatography (HPLC)

In HPLC the stationary phase is a column packed

with solid particles and the mobile phase is a
liquid solvent.
As the mobile phase is pumped through the
column, the sample is injected. A detector then
identifies the components as they exit the
column. Components are identified by their Rt,
the length of time they take to pass through the
column. And the results are compared with
standards.
 Mechanisms of Toxicity
Step1: Delivery from the site of exposure to
the target

Step2: Reaction of the ultimate toxicant

with the target molecule

Step3: Cellular dysfunction and resultant

Toxicities

Step 4: Repair or Dysrepair

Step1- Delivery from the site of exposure to the target

• Delivery is the movement of the toxicant from the site of

exposure to the site of its action
• Intensity of a toxic effect depends on the concentration &
persistence of the ultimate toxicant at its site of action.

• Ultimate toxicant is the chemical that reacts with the

endogenous target molecule (e.g. receptor, enzyme,
DNA, protein, lipid) or critically alters the biological
environment, initiating structural and/or functional
alterations that result is toxicity.
 Factors affecting Delivery

• Absorption • Presystemic
elimination

• Distribution • Distribution
toward the away from the
target target

• Reabsorption • Excretion

• Detoxication
• Toxication
 EXPOSURE SITE
Skin, GIT, Inhalation,
Injection
TOXICANT
D
E
L
I Presystemic elimination
Absorption
V Distribution away from
Distribution toward the
target
E the target
Reabsorption R Excretion
Toxication Y Detoxication

ULTIMATE TOXICANT
TARGET MOLECULE
Protein, Lipid, Microfilament,
DNA, Receptor
 Absorption versus Presystemic Elimination

Absorption is the transfer of a chemical from the site of

exposure into the systemic circulation.

The rate of absorption is related to:

1) The concentration of the chemical at the absorbing surface
2) The area of the exposed site
3) The characteristics of the epithelial layer through which
absorption takes place (e.g., the thickness of the stratum
corneum in the skin)
4) The intensity of the subepithelial microcirculation
5) The physicochemical properties of the toxicant: In
general, lipid-soluble chemicals are absorbed more readily
than are water-soluble substances.
 Presystemic Elimination means removal of the
toxicant during its transfer from the site of
exposure to the systemic circulation.

First pass through the GIT mucosal cells and liver:

GIT mucosa and the liver may eliminate a significant

fraction of a toxicant during its passage through these
tissues, decreasing its systemic availability.
For example:
• Ethanol oxidation (Alcohol dehydrogenase) in the gastric
mucosa
• Morphine glucuronation in intestinal mucosa and the liver
 Distribution to and Away from the Target

During the distribution phase toxicants enter

the extracellular space and may penetrate
into cells.
– Lipid-soluble compounds move readily into cells
by diffusion.

– Highly ionized and hydrophilic toxicants (e.g.

aminoglycosides) are largely restricted to the
extracellular space unless specialized membrane
carrier systems are available to transport them.
  Mechanisms Opposing Distribution
to a Target
• Binding to plasma proteins: toxicants such as DDT
cannot leave capillaries by diffusion because they
are bound to plasma lipoproteins.
• Specialized barriers: brain capillaries lack
fenestrae and are joined by extremely tight
junctions. BBB prevents the access of hydrophilic
chemicals to the brain except for those that can be
actively transported.
• Accumulation in storage (as adipose tissue): some
chemicals accumulate in tissues (i.e., storage sites)
where they do not exert significant effects.
 Excretion versus Reabsorption

• Excretion is the removal of toxicants

from the blood and their return to
the external environment.
• Reabsorption is the reuptake of filtrated
toxicants by renal tubules across their tubular
cells into the peritubular capillaries.
• Reabsorption by diffusion is dependent on the
lipid solubility of the chemical.
 Excretion versus Reabsorption
• For organic acids and bases, diffusion is inversely
related to the extent of ionization, because the
nonionized molecule is more lipid-soluble.

• The ionization of weak organic acids such as salicylic

acid and phenobarbital and bases such as amphetamine,
procainamide, and quinidine is strongly pH-dependent
in the physiologic range.

• Reabsorption is affected by the pH of the tubular fluid.

• Acidification favors excretion of weak organic bases

• Alkalinization favors elimination of weak organic acids.

 Toxication versus Detoxication
• Toxication (metabolic activation): biotransformation
to harmful products.

Usually, toxication makes toxicants reactive toward

endogenous molecules with susceptible functional
groups.

Sometimes, toxication may have physicochemical

properties to alter biological processes or structures.
e.g, oxalic acid formed from E. glycol may cause
acidosis.

Occasionally, chemicals acquire structural features and

reactivity by biotransformation that allows for a more
efficient interaction with specific receptors or
enzymes.
 Toxication (cont.)
Increased reactivity may be due to conversion into

• Electrophiles: molecules containing an electron-

deficient atom with a partial or full positive charge
that allows it to react by sharing electron pairs
with electron-rich atoms in nucleophiles.
• Free radicals: molecules that contain one or more
unpaired electrons in its outer orbital.
• Nucleophiles: uncommon mechanism for activating toxicants.
– formation of cyanide from amygdalin.
 Detoxication
• Biotransformations that eliminate the
ultimate toxicant or prevent its formation
 Step2- Reaction of the ultimate
toxicant with the target molecule

• This reaction leads to the injury

to the target molecule itself, cell
organelles, cells, tissues and
organs, and even the whole
organism.
Step3- Cellular dysfunction and resultant toxicities

• The reaction of toxicants with a target molecule may result

in impaired cellular function.

• Normally, each cell has defined programs:

– Programs determine the destiny of cells—that is,

whether they undergo division, differentiation (i.e.,
produce proteins for specialized functions) or apoptosis

– Programs control the activity of differentiated cells,

determining whether they secrete more or less of a
substance, whether they contract or relax, and whether
they transport and metabolize nutrients at higher or
lower rates.
 Step 4- Repair or Dysrepair

• Many toxicants alter macromolecules,

which, if not repaired, cause damage
at higher levels in the organism.

• The organism trials to repair the damaging

effects from toxicants on molecular,
cellular, and tissue levels