Postmortem

Forensic
Toxicology

Teri Martin
teri.martin@jus.gov.on.ca
September 23, 2003
 Outline

• Definitions and purpose of postmortem tox

• Samples of forensic interest
• Handling and storage of samples
• Pitfalls in postmortem toxicology
• Interpretation of results
 Postmortem Forensic Toxicology

• Qualitative and quantitative analysis of drugs

or poisons in biological specimens collected at
autopsy

• Interpretation of findings in terms of:

• Physiological effect at time of death
• Behavioural effect at time of death
 Quantitative vs. Qualitative

• Qualitative analysis – determines the presence

or absence of a drug or poison in a submitted
sample

• Quantitative analysis – determines the amount

of drug or poison that is present in the
submitted sample
 Postmortem Forensic Toxicology

Types of cases:
• Suspected drug intoxication cases
• Fire deaths
• Homicides
• Driver and pilot fatalities
• Therapeutic drug monitoring
• Sudden infant death (SIDS)
Samples of Forensic Interest
 Issues in Specimen Collection

• Selection
• Multiple, varied sites of collection
• Collection
• Appropriate method of collection
• Adequate volumes for analysis
• Storage and handling

Important to ensure analytical results are

accurate and interpretations are sound
 Typical autopsy specimens

• Blood
• Urine
• Stomach contents
• Bile
• Liver
• Hair
• Vitreous humor
 Blood

• Antemortem  ideal blood sample

• Postmortem blood is not truly “blood”

• Anatomical site of collection at autopsy

should be noted
 • Central sites
Subclavian
• Heart

Heart
• Peripheral sites
• Femoral
Iliac
• Iliac
• Subclavian
Femoral

• Other sites
• Head blood
• Hematoma blood
 Hematoma

• Extravascular blood clot

• Protected from metabolism
• Analysis will indicate what drugs were
present in the blood at the time of formation
 Hematoma case example

• A 26 year old man was found dead at the bottom of

a staircase. Death was due to physical injuries.
• Question as to alcohol use prior to fall down stairs
• No urine available at autopsy
• Alcohol not detected in femoral blood
• Alcohol in hematoma blood  150 mg/100 mL
• The deceased had been drinking prior to receiving
the head trauma.
• The deceased had survived for several hours after
the injury.
 Hematoma

• Caution: There may be a delay between the

incident which resulted in hematoma and the
actual formation of the hematoma
• Therefore, this alcohol concentration does not
necessarily indicate the BAC at the time of
the fall down the stairs.
 Urine

• Produced by the kidneys

• Blood filtered by the kidneys
• Stored in the bladder until voided

• Qualitative - the presence of a drug in the

urine of an individual indicates that some
time prior to death the drug or poison was
present in the blood of the individual
 Stomach contents

• Visual examination may reveal tablets

• Drugs that have been orally ingested may be detected in
stomach contents
• Caution: drugs administered by other routes may also
diffuse into stomach contents from the blood
• Generally qualitative:
• Stomach contents are not homogeneous
• Only a portion of stomach contents collected (unmixed?)
• Useful for directing further analysis
 Case Example

• A 26 year old woman is found dead in bed

• Numerous medications in her home:
• Amitriptyline, Oxycodone, Morphine, Paroxetine,
Diphenhydramine, Pseudoephedrine, Phenobarbital,
Codeine, Temazepam, Diazepam
• Only 3 mL of blood collected at autopsy
• Qualitative analysis of stomach contents:
• Amitriptyline: detected
• Nortriptyline: detected
• Quantitation can now be performed in blood
 Liver

• Drug metabolism occurs in the liver

• Both parent compounds and metabolites may
be present in higher concentrations in the
liver than in the blood  ease of detection
• Limitation is that drugs are not uniformly
distributed throughout the liver  confounds
interpretation
 Bile

• Digestive secretion
• Continuously produced by the liver
• Stored in the gallbladder

• Qualitative - the presence of a drug in the

bile of an individual indicates that sometime
prior to death, the individual was exposed
to the drug
 Vitreous humor
• Fluid that occupies the space between the
lens and the retina of the eye.
• Sequestered from putrefaction, charring and
trauma, microorganisms.
• Useful in cases where decomposition is
advanced, body is exhumed or in fire deaths
• Limitation is blood:vitreous ratio may not
be known
 Hair

• Recent specimen of interest

• Metabolism does not occur in hair
• Can provide a historical record of drug or
poison exposure
• Pros and cons of hair analysis still being
uncovered  racial variability?
 Case Example

Poklis, A. 2002. Abstract SOFT, Dearborn, Michigan.

• 30 year old woman, previously in good health

• Nausea, vomiting, diarrhea, rash, fever
• Weakness in hands and feet  Guillian Barre?
• Hospitalized with hypotension, seizures
• Misplaced laboratory result  Arsenic!
• Sequential hair analysis for arsenic showed
chronic arsenic poisoning over 8 month period
 Non-biological submissions

• Used to direct analysis of biologicals

• May indicate the nature of substances that
may have been ingested, inhaled or injected
• Examples:

• Containers found at the scene

• Syringes
• Unidentified tablets or liquids
 Autopsy specimens of limited value

• Pleural fluid
• Chest cavity blood
• Gutter blood
• Samples taken after embalming
• Samples taken after transfusion in hospital
• “Spleen squeezings”
• “Esophageal scrapings”
 Chest Cavity Fluid

• Not readily definable

• Most likely to be collected if:
• Traumatic injury to the chest
• Advanced decomposition
• A “contaminated” blood sample, chest
cavity fluid may contain fluids from
stomach, heart, lungs etc.
 Samples taken after embalming

• Methanol is a typical component of

embalming fluid
• Most drugs are soluble in methanol
• Embalming process will essentially “wash”
the vasculature and tissues
• Qualitative analysis can be performed on
body tissues
 Case Example

A 72 year old woman, given meperidine to

control pain following surgery, later died in
hospital. The woman was in poor health and it is
possible that death was due to natural causes.
However, coroner requests toxicology to rule out
inappropriate meperidine levels.
BUT:
• Body had been embalmed
• Liver and spleen submitted
Storage and Handling
 Proper specimen handling

• Identification of samples
• Continuity
• Contents
• Specimens delivered to lab without delay
• Specimens should be analyzed as soon as possible
• Storage areas should be secure
 Storage and Handling

• Not feasible to analyze specimens immediately

• Sample should be in well-sealed container
• Sample containers must be sterile
• Use of preservatives and anti-coagulants
• Refrigeration vs. Freezing
• Both inhibit bacterial action; esp. freezing
• Freezing results in  prep time
• Freeze-thaw cycle may promote breakdown
 Storage of Samples

• Preservative
• Sodium fluoride
• Anti-coagulants
• Sodium citrate
• Potassium oxalate
• EDTA
• Heparin
• Not imperative for postmortem blood samples
 Determining analyses

• Case history
• Medical history
• Autopsy findings
• Symptomatology
• Experience of the toxicologist
• Amount of specimen available
• Nature of specimens available
• Policies of the organization
Pitfalls in Postmortem
Forensic Toxicology
 Decomposition

• Autolysis
• The breakdown of cellular material by enzymes
• Putrefaction
• A septic/infectious process
• The destruction of soft tissues by the action of
bacteria and enzymes
• Traumatic deaths may demonstrate  putrefaction
 Decomposition

• Fewer samples available for collection

• Quality of samples is diminished
• Putrefaction produces alcohols
• Ethanol
• Isopropanol
• Acetaldehyde
• n-propanol
 Postmortem redistribution
• A phenomenon whereby increased
concentrations of some drugs are observed in
postmortem samples and/or site dependent
differences in drug concentrations may be
observed

• Typically central blood samples are more prone

to postmortem changes (will have greater drug
concentrations than peripheral blood samples)
 Possible mechanisms of
postmortem redistribution

• Diffusion from specific tissue sites of higher

concentration (e.g. liver, myocardium, lung) to
central vessels in close proximity
• Diffusion of unabsorbed drug in the stomach to
the heart and inferior vena cava
• Diffusion of drugs from the trachea, associated
with agonal aspiration of vomitus
 Case Example

• 37 year old man found dead in his home

• Cause of death identified at autopsy as asphyxia due
to choking; white pasty material lodged in throat
• Heart blood • Femoral blood
• Morphine: 20 000 ng/mL • Morphine: 442 ng/mL
• Amitriptyline: 0.36 mg/dL • Amitriptyline: 0.01 mg/dL

• Examination of esophageal and tracheal contents

revealed presence of both morphine and
amitriptyline
 Susceptible Drugs

Drugs most commonly associated with postmortem

redistribution:

1. are chemically basic

2. have large volumes of distribution
 Volume of distribution

• Review from last lecture:

• Volume of distribution is the amount of drug in
the whole body (compared to the amount of drug
in the blood)
• If a drug has a large volume of distribution, it is
stored in other fluids and tissues in the body
 Susceptible Drugs

• Tricyclic • Narcotic
antidepressants Analgesics
• Amitriptyline • Codeine
• Nortriptyline • Oxycodone
• Imipramine • Propoxyphene
• Desipramine

• Antihistamines • Doxepin
• Diphenhydramine • Digoxin
 Example: Digoxin

p. 60, Principles of Forensic Toxicology

• A 33 year old white female is admitted to
hospital after taking 60 digoxin tablets
• An antemortem blood sample collected 1 hour
prior to her death indicates a blood digoxin
level of 18 ng/mL
• Heart blood digoxin concentration obtained at
autopsy is 36 ng/mL
 Example: Digoxin

• Postmortem increase in blood digoxin

concentrations is suspected to be due to the
release of the drug from the myocardium

• Postmortem levels > Antemortem levels

• Heart blood levels > Femoral blood levels
 Postmortem redistribution

• Coping with the problem of postmortem

redistribution:

• Analysis of both central blood and peripheral blood

in cases where postmortem redistribution may be a
factor

• Compilation of tables to determine average and

range of postmortem redistribution factors for
drugs
 Incomplete Distribution

• Site dependent differences in drug levels due to

differential distribution of drugs at death
• Has been noted in rapid iv drug deaths
• Example:
• Intravenous injection of morphine between the toes
• Fatal amount of drug reaches the brain
• Full distribution of the morphine throughout the
body has not occurred
• Femoral concentration > Heart concentration
 Drug Stability

• Knowledge of a drug’s stability is necessary to

facilitate interpretation of concentrations
• Breakdown of drugs may occur after death and
during storage via non-enzymatic mechanisms

• Cocaine  Benzoylecgonine (Hydrolysis)

• LSD  degradation due to light sensitivity
• Others ?
 Example: Bupropion

• Bupropion, an antidepressant, was identified

and confirmed during a GC drug screen
• Blood analyzed using a quantitative analysis:
• Bupropion  not detected
• Review of the literature:
• Laizure and DeVane, 1985. Ther. Drug. Monit.
• “Bupropion showed a log linear degradation that
was both temperature and pH dependent…”
 Evaporation of volatiles

• Ethanol
• Carbon monoxide
• Cyanide
• Toluene
• Other alcohols
 Example: Carbon Monoxide

Ocak et al. 1985. J. Analytical Toxicology. 9: 202-206

• Effects of storage conditions on stability of CO

• No significant change in % CO saturation in capped

samples stored at room temperature or 4oC
• Significant losses in % CO saturation in uncapped
samples stored at room temperature and at 4oC

• Mechanism for loss  diffusion

Interpretation
 Interpretation

Therapeutic, toxic or fatal? How do you know?

• Compare measured blood concentrations with

concentrations reported in the literature:
• Clinical pharmacology studies
• Incidental drug findings
• Plasma  blood

• Consider case history:

• Symptoms observed by witnesses?
• Tolerance of the individual to the drug
 Blood:plasma ratios

• Knowledge of the blood:plasma ratio can be

very important when applying information from
clinical studies to postmortem forensic tox

• Cocaine, blood:plasma ratio is 1.0

• Phenytoin, blood:plasma ratio is 0.4
• Ketamine, blood:plasma ratio is 1.7
• Hydroxychloroquine, blood:plasma ratio is 7.2
 Example: THC
• Six healthy male volunteers recruited for a study
of the pharmacokinetics of THC in humans
• Smoked a “high-dose” THC cigarette
• 15 minutes after cessation of smoking, plasma THC
concentrations averaged 94.8 ng/mL

• The plasma:blood ratio for THC is 1.8

• Plasma contains 1.8x as much THC as whole blood
• The results of this study correspond to a blood THC
concentration averaging 53 ng/mL
 Importance of History: Tolerance

• Drug concentrations in non-drug related deaths

may overlap with reported drug concentrations
in fatal drug intoxications

• Methadone example:
• Naïve users - deaths due to methadone are
associated with blood levels > 0.02 mg/100 mL
• Patients on methadone maintenance – peak blood
concentrations may range up to 0.09 mg/100 mL
 Interpretation

Acute vs. Chronic Ingestion: Can you tell?

• Parent:metabolite drug concentration ratio may

be of assistance in differentiating between
acute and chronic ingestion of a drug
 Example: Amitriptyline

Case 1 Case 2

Amitriptyline: 0.4 mg% Amitriptyline: 0.04 mg%

Nortriptyline: 0.02 mg% Nortriptyline: 0.08 mg%

Parent >> Metabolite Parent < Metabolite

Suggestive of acute Slow death and/or

overdose and rapid death chronic administration
 Interpretation

Metabolites are produced when drugs are

biotransformed (converted) into other
chemicals, more easily excreted from the body

Metabolite drug concentrations may be the

more useful measure of exposure or toxicity
 Metabolites: Exposure

The parent compound may be a prodrug or may

have a shorter t1/2 than the metabolite:

• Clorazepate  nordiazepam
• Flurazepam  N-desalkylflurazepam
• Heroin  morphine
 Metabolites: Toxicity

The metabolite may have  toxicity over the parent

compound:

• Acetaminophen  N-Acetylbenzoquinoneimine
• Meperidine  normeperidine
• Methanol  formic acid
• Ethylene glycol  oxalic acid  calcium oxalate