Forensic medicine-

Toxicological Analysis
Dr. Muhammad Naeim, PhD
 Learning outcome

Students must be able to:

• Explain types of toxicological
investigations
• Explain different areas in the analysis of
poisons
 Types of toxicological investigations
Different types of proposes for toxicological analysis:
• Poison analysis in sudden and unexpected death (postmorterm
toxicology)
• Suspected poisoning
• Bio analysis involving living persons for driving under influence of
drug, use of drugs in sport, suspected drug assisted assault
• Employment, sport screening
• Testing in prison for drugs/poison
• Death of fire
 HOMEWORK
• Define forensic toxicology.
 Areas in the analysis of poisons

Determines the absence or presence of drugs and their

metabolites, chemicals such as ethanol and other
volatile substances, carbon monoxide and other gases,
metals, and other toxic chemicals in human fluids and
tissues, and evaluates their role as a determinant or
contributory factor in the cause and manner of death/
poisoning.
 Areas in the analysis of poisons
• Postmortem drug testing – this is used in death
investigations to establish whether or not drugs were the
cause or contributing factor in death.
• Workplace drug testing – this consist of evaluation of
biofluids, primarily urine and blood from employees or
job applicants for drug content.
• Evaluation of Contraband – this is applied when
evidence seized is being linked to the illicit drug trade.
Police agencies needed labs that could link materials
to illegal drug trafficking and manufacture.
• The substance may also have been diluted by
its dispersal through the body----sample under
investigation may only contain micrograms or
nanograms of active constituent.

• During the post mortem examination, the

forensic toxicologist will assist the forensic
pathologist in collection and documentation
of any evidence of forensic value.
• A forensic toxicologist is normally presented with
preserved samples of body fluids, stomach
contents, and organ parts.
• They will have access to the pathologist's
report which should contain information on
various signs and symptoms as well as other
postmortem data.
• The toxicologist needs a through knowledge of
how the body alters or metabolizes drugs
because few substances leave the body in
the same state as they entered.
 Collection of Samples
• Care must be taken.
• The shorter the delay between death and collection of
samples the better.
• Many toxins will be broken down naturally by decay and
cellular autolysis.
• Large quantities of specimen is needed--- extraction
may be ineffective, thus re-extract
• Specimens of many different organ are required since
drugs and poison displays varying affinities for body
tissues.
FORENSIC PATHOLOGIST ROLES
 Specimen Collection
Blood

– the most satisfactory method for obtaining samples is

from venous puncture of the femoral vein.
– An alternate site to collect venous blood is to incise
the jugular.
– this is typically done after the neck has been
exposed under the sub-clavicular incision.
– The correlation between blood concentration and
effects of the drug is very high in blood testing.
 Specimen Collection
Urine –
• in pre-employment screening urine is most often used to
test for presence of drugs.
– This is typically because of the types of test that can
be run and the cost effectiveness of the testing.
– One draw back to urine is the correlation between
drug concentration in urine and drug effects is usually
poor (why? Bioactive concentration vs amount of
elimination)
– Urine should also be collected in post mortem
investigations since some toxins show in higher levels
in urine.
– At autopsy, a catheter or suprapubic puncture will
suffice.
 Specimen Collection
Bile –
• this can be useful in cases where morphine and
chlorpromazine are suspected toxins.
• These toxins are concentrated by the liver and
excreted into the gall bladder.
• Direct collection of bile into a bottle is advised because
bile is too viscous to be sucked through a needle.

Liver –
• the liver filters the blood and is most heavily involved in
drug metabolism. (hepatotoxicity)
 Specimen Collection
Cerebrospinal Fluid –

• not often collected for toxins, but may be needed for

microbiological analysis.
• If needed it should be collected by lumbar or cisternal
puncture.
– Cisternal puncture = Passage of a hollow needle
through the posterior anto-occipital membrane into
the cerebellomedullary cistern
 Specimen Collection
Vitreous Humor –

• sometimes useful in bodies with appreciable post

mortem decay (decomposed).
• The fluid in the eye resist putrefaction longer than other
body fluids.
• Vitreous can also be used at estimating time of death.
• Puncture should be made through the sclera with a 5
Gauge needle.
 Specimen Collection
• Gastric Contents –

• This is typically done in a sudden death in which the

decedent has large quantities of a lethal agent in his
stomach.
• undigested pills or liquids that were ingested just
before the death
• Contents should be emptied into a wide mouth jar.
– In the case of suicide, large amounts of toxins in the
gastric tract may make this point apparent.
 Specimen Collection
Hair –
• Not common in forensics for a number of reasons.
– Drugs only present in extremely low concentrations.
– The benefit of hair is that its is a good timeline for
drug usage since it is only eliminated when hair is cut.
• Hair is also useful for other toxicant or exposure
determinations.
• In cases of suspected chronic poisoning that results in
death.

• Other specimens : nail, saliva, sweat, bone marrow,

maggots, lung, brain, kidney, bone
 Toxicological Investigation
• Case examples: poisoning death,
suspected poisoning, illegal drug
abuse
• 3 steps
– ④ Obtaining case history and suitable
specimens (pathologist)
– ② Toxicological analyses
– Interpretation of Analytical findings
③
① Case History and Specimens
In case of poison death (postmorterm Toxicology)
• Collect facts/history
• Gross autopsy findings
• Drugs available to the decedent
• Interval between the onset of symptoms and
death
• Analyses performed
• Analyzed Specimens
 :
Toxicological Analyses
• Factors to consider
– Amount of specimen available
– Nature of the poison sought
– Possible biotransformation of the poison

• GC/MC or HPLC is most widely used for most

drug identification
③ Interpreting Findings
• The goal of a forensic toxicologist is to confer with
the pathologist in determining the cause and
manner of death/ suspected poisoning case.
• In simple terms we infer that the death/poisoning
is due to a specific toxic substance when
appropriate quantities of that toxicant are found.
• When other findings conclude cause of
death/poisoning we may specify the toxic
substance as a contributing factor to
death/poisoning.
 The Screening Step

• A screening test is normally employed to provide the

analyst with quick insight into the likelihood that a
specimen contains a drug substance.
• Quick test to narrow down possibilities
• Positive results arising from a screening test are
considered to be tentative at best and must be verified
with a confirmation test.
• The most widely used screening tests are immunoassay
(colour test; spot test), thin layer chromatography, gas
chromatography, UV spectrophotometry, Breathalyzer
etc..
 Screening Tests
• Physical tests -- boiling point, melting point, density, and

• :
refractive index
Crystal tests -- treatment with a chemical reagent to
produce crystals
• Chemical spot tests -- treatment with a chemical

• :
reagent to produce color changes
Chromatography (thin-layer or gas) -- used to separate
components of a mixture
 The Confirmation Step
• For a substance to be reported as positive, at least
two different analytical techniques must be used.
• Confirmation step is fundamental principle in forensic
toxicology
• Determines exact identity
• Gas chromatography/mass spectrometry is generally
accepted as the confirmation test of choice.
• The GC separates the sample into its components, while
the MS represents a unique “fingerprint” pattern that
can be used for identification.
• Once the drug/toxic substance is extracted and
identified, the toxicologist may be required to provide
an opinion on the drug’s effect on an individual’s
natural performance or physical state.
• Quantification of the substances required---prepared
standard of known concentration
 Spot test

Urine Testing: The Most Commonly Used Drug

Test
• The forensic toxicologist must devise an
analytical scheme (standard operating
procedures) that will successfully detect, isolate,
and specifically identify toxic drug substances.
• Once the drug has been extracted from
appropriate biological fluids, tissues, and organs,
forensic toxicologist can proceed to identify the
drug/toxic substance present.
• Drug extraction is generally based on a large
number of drugs being either acidic or basic.
 Present screening of drugs
• Forensic Toxicologists may face an general unknown----thus, mass
screening is carried out first, if positive the work towards individual
targets (eg. Drugs), confirm and quantify
• Immunoassay techniques often form the first part of toxicological
analysis for screening.
examples: RIA, ELISA, EMIT, CEDIA(cloned enzyme donor
immunoassay, FPIA(fluorescence polarization immunoassay).
• Extraction techniques: Older methods of extraction of drugs from
biological matter is called liquid/liquid extractions, are still common
and this involves extractions done under controlled pH for a group of
drugs or for one particular drug. Extractions are also done for
specific drug or group of drugs indicated by the results from
immunoassay screening method.
• Other extraction; SPE, SPME, SFE
• The extracts are then subjected to various chromatography
procedure such as GC,HPLC and more recently CZE and MS for
screening, quantitative determinations as well as for confirmation
THANK YOU