Government of Ontario

CFS - TIS - Toxicology Information Sheet

Centre of Forensic Sciences

Technical Information Sheets

Toxicology


Toxicology Information – PDF, 112 kb


Introduction

The Toxicology Section performs analyses on biological samples (e.g., blood, urine, liver) to determine the absence/presence/concentration(s) of drugs, including alcohol and poisons.

This document is intended as a convenient investigative reference but should not be relied upon as being definitive or exhaustive. Please contact the Centre of Forensic Sciences (CFS) Toxicology Section for assistance with questions of an analytical or toxicological nature by e-mail or telephone 647 329-1400 or 647 329-1430. When calling please ask for the appropriate coordinator:

Coroner’s Coordinator:
CFSToxicologyCoronerCoordinator@ontario.ca

Criminal Coordinator:
toxcrim@ontario.ca

Analytical Decision-making and Capability

The screening methods employed in the Toxicology Section are:

  1. gas chromatography (GC) and gas chromatography/mass spectrometry (GC/MS) screen
  2. immunoassay (IA)
  3. head-space GC analysis for volatiles
  4. Quadrupole Time-of-Flight MS (QTOF)

The targeted/quantitation methods employed in the Toxicology Section are:

  1. GC, GC/MS
  2. liquid chromatography (LC), LC/MS
  3. head-space GC analysis for volatiles

The capability of the screening methods is presented in Appendix 1. While these screening methods have wide-ranging capabilities not all drugs may be reliably detected. Appendix 2 contains a list of compounds that may not be identified by the screening methods but may be detected/quantitated by targeted methods. Many of the compounds contained in this list will not be tested for unless specifically requested. If use of a specific drug is known or suspected it should be noted in the case synopsis.

Decisions regarding which tests to be used in a case are informed by a variety of sources including case type, case history, nature of submitted samples, analytical protocols and capabilities, and discussions with clients. The initial toxicological analyses conducted for a variety of case types are presented in Appendix 3.

Urgent Cases

Requests for expedited analyses must meet specific criteria before being accepted as an urgent case. This process requires authorization by Toxicology Section management.

Examination

All items are visually examined to check the seal numbers (if present), the contents, and the integrity of the packaging.

Instrumentation

Chromatography: Gas Chromatography (GC); Liquid Chromatography (LC)
Chromatography is an analytical technique used to separate compounds based on their chemical and structural properties. GC uses a pressurized gas, while LC uses a pressurized liquid, in the separation of compounds.

Immunoassay (IA)
IA detects compounds in biological fluids using a reaction of an antibody or antibodies to its antigen (i.e., the drug). This technique is primarily a screening technique; however, some IA methods are semi-quantitative, e.g., acetaminophen.

Inductively Coupled Plasma (ICP)
ICP, when coupled with mass spectrometry, is capable of detecting metals and several non-metals at very low concentrations. ICP ionizes the sample and then an MS separates and quantitates the ions.

Mass Spectrometry (MS)
MS detects, identifies, and quantitates compounds. An MS can be coupled with a GC or an LC.

Quadrupole Time-of-Flight-MS (QTOF)
QTOF detects and identifies compounds. A QTOF is coupled with an LC.

Tandem MS (MS/MS)
MS/MS detects, identifies, and quantitates compounds and is commonly coupled to a gas or liquid chromatograph.

Ultraviolet and Visible (UV/VIS) Spectrophotometry
UV/VIS spectrophotometry identifies and/or quantitates a drug based on its UV and/or visible light-absorbing properties.

Colour Tests
Colour tests tentatively identify the presence of drugs in a variety of samples. Chemicals added to the sample will produce an expected colour if the drug is present.

Carbon Monoxide
Carbon monoxide is analyzed by visible spectrophotometry. Results are expressed as % carboxyhemoglobin saturation.

General Toxicology Screen
These analyses, which may include GC, GC/MS, LC, UV spectrophotometry, and colour tests are employed when other analytical methods are not suitable. Samples may include stomach contents, body tissues, and urine. While general toxicology screen methods have a wide-range of capability their sensitivity is low and the methods are generally qualitative.

Interpretation

Quantitative results may be expressed as 1) a concentration or 2) as < or > a concentration as appropriate, e.g., when sufficient for interpretation. Blood ethanol interpretations provided in reports are generally limited to cases in which the detected concentration may be associated with fatalities, may be influenced by post-mortem artefacts, may have toxic interactions with other drugs, or in the case of fatal motor vehicle collision, associated with impairment.

Measurement Uncertainty

Measurements made with all scientific instruments are associated with variability. No measurement is exact, but is an estimate of the true value. Calculation of measurement uncertainty (MU) employs statistical methods to determine the range of values within which the quantitative result is likely to reside. The MU provides a reasonable estimate of the variability associated with the analytical method and is based on the analysis of matrix-matched quality control samples. A minimum of 10 such analyses are used. The MU is calculated with a confidence of 95.45 per cent using a k-factor based on the degrees of freedom as determined by the Student’s t-test and the standard deviation of the associated quality control data. The MU is expressed in the same units in which the quantitative result is reported, e.g., ng/mL, mg/L and is reported as: quantitative result ± MU.

Limitations

The focus of this laboratory is drug toxicity. Clinical blood/urine chemistry analysis, e.g., electrolytes, cell counts, gas saturation, creatinine, is not performed in this laboratory. Some specialized screening procedures are sample-specific, e.g., stomach contents. This laboratory does not have validated methods to analyze some sample types, e.g., oral fluid, hair, bile, muscle, brain tissue. There are a variety of analytical issues that may prevent the detection of some of the drugs that this laboratory is commonly capable of detecting, which include:

  • matrix effects
  • degree of putrefaction
  • type of sample (e.g., splenic blood)
  • post-mortem interval
  • storage conditions
  • volume of sample submitted
  • low concentration of the drug/sensitivity of the method

Conversely, some novel, or rarely encountered, drugs not listed in Appendix 1 may be identified by the GC and GC/MS or QTOF screens. In this case, analytical reference material would be acquired (if available) then analysed to confirm identity. Additionally, there are drugs/compounds for which the CFS Toxicology Section does not have a method, examples of which are provided in Appendix 4.


Appendix 1

Drugs that can be reliably detected by screening methods

GC and GC/MS Screen

A
alpha-pyrrolidinovalerophenone (α-PVP)
acetyl fentanyl2
amantadine1
amitriptyline2
amlodipine2
amoxapine2
amphetamine2
amphetamine (4-fluoro)
amphetamine (dimethyl)
anabasine
anileridine1
atomoxetine
atropine/hyoscyamine

B
benzocaine
benzofuran (6-(2-aminopropyl, 6-APB)
benztropine1
benzylpiperazine (BZP)
bromo-dragonfly
brompheniramine2
bupivacaine1
bupropion2
butylone
butyryl fentanyl

C
caffeine1
carbamazepine1
cathinone (cath)
n-ethyl-cath
4-flurometh-cath
3-methoxymeth-cath
4-methyleth-cath
meth-cath
chlorcyclizine
chlordiazepoxide2
chloroquine
chlorpheniramine2
chlorpromazine1
cimetidine
cisapride4
citalopram*2
clomipramine2
clonidine1
clozapine2
cocaethylene
cocaine2
codeine2
cotinine
cyclobenzaprine2
cyproheptadine1

D
desipramine2
dextromethorphan2
dextrorphan*
diazepam2
diazepam (nor)2
dibucaine4
dihydrocodeine
diltiazem2
diltiazem (desacetyl)2
dimethyltryptamine
diphenhydramine2
diphenoxylate1
doxepin2
doxylamine2

E
ephedrine*
ethylone

F
fluoxetine2
fluoxetine (nor)2
flurazepam2
flurazepam (n-desalkyl)2
fluvoxamine2

H
haloperidol1
hydrocodone2
hydroxychloroquine
hydroxyzine1

I
ibogaine
imipramine2


K
ketamine2

L
lamotrigine2
laudanosine
levamisole
lidocaine
loratadine
loxapine2

M
maprotiline1
meclizine1
mefloquine1
meperidine2
meperidine (nor)2
mephedrone2
mepivacaine1
methadone2
methamphetamine2
methamphetamine (4-fluoro)
methedrone
methotrimeprazine2
methylenedioxyamphetamine (MDA)2
methylenedioxyethylamphetamine (MDEA)2
methylenedioxymethamphetamine (MDMA)2
3,4-methylenedioxypyrovalerone (MDPV)
methylone
methylphenidate2
metoclopramide1
metoprolol2
midazolam2
mirtazapine2
moclobemide1

N
nicotine1
nortriptyline2

O
olanzapine2
orphenadrine2
oxybutynin1
oxycodone2

P
paroxetine2
pentadrone
pentazocine2
pentoxyphylline
pentylone
phenacetin
phencyclidine (PCP)2
phenethylamines (2C-B, 2C-B-Fly, 2C-T-7, PEA)
pheniramine2
phenmetrazine
phentermine1
piperazine, 1-3 chlorophenyl (mCPP)
piperazine, trifluoromethylphenyl (TFMPP)
p-methoxyamphetamine (PMA)2
p-methoxymeth-amphetamine (PMMA)
procainamide1
procaine1
prochlorperazine4
procyclidine1
propoxur1
propoxyphene2
propranolol2
protriptyline2
pseudoephedrine2
pyrilamine (mepyramine)1

Q
quetiapine2
quinidine1

R
ropinirole
ropivacaine

S
scopolamine (hyoscine)1
selegiline
sertraline2
strychnine1

T
tapentadol
terbinafine
thioridazine1
ticlopidine
tramadol2
tranylcypromine1
trazodone2
trifluoperazine1
trihexphenidyl2
trimethoprim4
trimebutine
trimipramine2
triprolidine2

V
varenicline
venlafaxine2
venlafaxine (O-desmethyl)
verapamil2

X
xylometazoline

Z
zolpidem2
zopiclone breakdown product

*The GC and GC/MS screen is not capable of distinguishing racemates, therefore compounds such as dextrorphan/levorphanol, citalopram/escitalopram, and ephedrine/pseudoephedrine cannot be separated.

QTOF Screen

The QTOF screen is a powerful and sensitive method that can reliably detect the drugs included in the following methods (details are listed in Appendices 5 and 6):

  • LC-MS/MS Mix 1
  • LC-MS/MS Mix 2
  • LC-MS/MS Mix 3

In addition, the QTOF screen can identify U-47700 and psilocin. The list of drugs potentially identifiable by QTOF is too extensive to list within this document. For questions about a specific drug not listed, please contact the appropriate case coordinator.

Immunoassay Tests (known cross-reactivity)

Barbiturates:

amobarbital2
butalbital2
pentobarbital2
phenobarbital2
secobarbital2

Head-space GC-FID analysis for volatiles (screen and quantitation)

acetone
ethanol
isopropanol
methanol
n-propanol (not quantitated)


Appendix 2

Compounds that may not be identified by screening methods, but might be detected and/or quantitated by targeted methods.

A
acebutolol4
acepromazine4
amiloride4
antipyrine (phenazone)1
atenolol4
atracurium4
azacyclonol4

B
bromocriptine4

C

carbaryl1
carbon monoxide6
chlorpropamide4
chlorzoxazone4
cyanide2

D
dantrolene4
diclofenac4
dipyridamole4

E
ethopropazine4
ethylene glycol8

F
fenfluramine1
fenodipine4
fenoprofen4
formic acid5

G
guaifenesin4

I
indomethacin4

K
ketoconazole4

L
labetolol4
lithium7
loperamide4

M
mefenamic acid4
methaqualone1
methocarbamol4
metronidazole4
mexiletine1

N
nabumetone4
nadolol4

O
oxprenolol4

P
papaverine4
pericyazine4
phenylbutazone4
phenyltoloxamine1
physostigmine1
pimozide4
pindolol4
pipotiazine4
piroxicam4
prazosin4
propafenone4

S

sotalol4
sufentanil1

T
terazosin4
terfenadine4

tiaprofenate4
timolol4
tolbutamide4
toluene5
triamterene4

V
valproic acid5

Y
yohimbine4

Z
zuclopenthixol4

Methods used for the quantitation of compounds identified in the preceding appendices are denoted as follows: 
1 GC-NPD
2 LC-MS/MS
3 GC-MS
4 LC-DAD
5 GC-FID
6 Visible spectrophotometry
7 ICP-MS
8 Qualitative


Appendix 3

Initial analyses presented by case type

Homicide: Ethanol, Screen, LC-MS/MS Mix 3, IA cannabinoids
Attempted murdera: dependent upon case history
Sexual assaulta: dependent upon case history
Alcohol-impaired driving: Ethanol
Drug-impaired driving: Screen, IA cannabinoids, UDM, GHBa
Possible drug-related death: Ethanol, Screen, LC-MS/MS Mix 3
Death of child < 5 years of age: Ethanol, Screen, LC-MS/MS Mix 3, IA cannabinoids, IA acetaminophen and salicylate
Mandatory inquest: Ethanol, Screen, LC-MS/MS Mix 3, IA cannabinoids
SIU death investigation: Ethanol, Screen, LC-MS/MS Mix 3, IA cannabinoids
Fire-related deathb: CO (whole blood required)
Confirmation of ketoacidosis: Ethanol (includes acetone), BHB
Fatal motor vehicle collision (driver) and aviation death: Ethanol, Screen, LC-MS/MS Mix 3, IA cannabinoids, COc

a dependent upon case history
b other analyses may be performed dependent upon evidence/suspicion of intoxication
c if fire is involved


Appendix 4

Examples of drugs/compounds for which this laboratory does not have a method

Animal toxins

a-bungarotoxin
connotoxin
maurotoxin
tetrodotoxin

Anesthetic gases
chloroform
diethyl ether
halothane
isoflurane
nitrous oxide

Curare-related toxins
alloferine
toxiferine
tubocurarine

Other
insulin
lead, mercury
polychlorinated biphenyls (PCB)
succinylcholine
thallium
xylazine


Appendix 5

Capability of quantitative methods

Barbiturate method (LC-MS/MS)
amobarbital
butalbital
pentobarbital
phenobarbital
phenytoin
primidone
secobarbital

Acid drug method (UPLC-DAD)
furosemide
ketorolac
naproxen
salicylic acid

LC-MS/MS Mix 1
amlodipine
diltiazem
diltiazem (desacetyl)
flurazepam       
lamotrigine
metoprolol
propranolol
zaleplon
ziprasidone
zolpidem
zopiclone

LC-MS/MS Mix 2
brompheniramine
chlorpheniramine
diphenhydramine
doxylamine
ephedrine
orphenadrine
pheniramine
phenylephrine
promethazine
pseudoephedrine
trimeprazine
triprolidine

LC-MS/MS Mix 3
6-monoacetylmorphine (6-MAM; qualitative)
alprazolam
amitriptyline
amphetamine
benzoylecgonine
bupropion
chlorpheniramine
citalopram
clonazepam
clonazepam (7-amino)
cocaine
codeine
cyclobenzaprine
dextromethorphan
diazepam
diazepam (nor)
diphenhydramine
fentanyl
flunitrazepam (7-amino)
fluoxetine
fluoxetine (nor)
flurazepam (n-desalkyl)
hydrocodone
hydromorphone
ketamine
ketamine (nor)
lorazepam
meperidine
meperidine (nor)
mephedrone
methadone
methamphetamine
methylenedioxyamphetamine
methylenedioxyethylamphetamine
methylenedioxymethamphetamine
midazolam
mirtazapine
morphine
nortriptyline
olanzapine
oxazepam
oxycodone
oxymorphone
paroxetine
pseudoephedrine
quetiapine
risperidone
sertraline
temazepam
tramadol (cis)
trazodone
venlafaxine
zopiclone

LC-MS/MS Mix 4
acetyl fentanyl
alprazolam (hydroxyl)
amoxapine
bromazepam
buprenorphine
butyryl fentanyl (qualitative)
chlordiazepoxide
chlorpromazine
clobazam
clomipramine
clozapine
demoxepam
desipramine
desomorphine
diltiazem
diltiazem (desacetyl)
doxepin
doxylamine
duloxetine
etizolam
flunitrazepam
flunitrazepam (N-desmethyl)
flurazepam
fluvoxamine
imipramine
levorphanol
loxapine
MDPV (qualitative)
methotrimeprazine
methylone
methylphenidate
naloxone
naltrexone
nitrazepam
nitrazepam (7-amino)
O-desmethylvenlafaxine
olanzapine
orphenadrine (qualitative)
PCP
pentazocine
pheniramine
promethazine
propoxyphene
triazolam
triazolam (hydroxy)
trimipramine
ziprasidone
zolpidem

Antiepileptic method (LC-MS/MS)
baclofen
gabapentin
topiramate
vigabatrin

LC-MS/MS Mix 1
amlodipine
diltiazem
diltiazem (desacetyl)
flurazepam       
lamotrigine
metoprolol
propranolol
zaleplon
ziprasidone
zolpidem
zopiclone

LC-MS/MS Mix 2
brompheniramine
chlorpheniramine
diphenhydramine
doxylamine
ephedrine
orphenadrine
pheniramine
phenylephrine
promethazine
pseudoephedrine
trimeprazine
triprolidine

LC-MS/MS Mix 3
6-monoacetylmorphine (6-MAM)
alprazolam
amitriptyline
amphetamine
benzoylecgonine
bupropion
chlorpheniramine
citalopram
clonazepam
clonazepam (7-amino)
cocaine
codeine
cyclobenzaprine
dextromethorphan
diazepam
diazepam (nor)
diphenhydramine
duloxetine (qualitative)
fentanyl
flunitrazepam (7-amino)
fluoxetine
fluoxetine (nor)
flurazepam (n-desalkyl)
hydrocodone
hydromorphone
ketamine
ketamine (nor)
lorazepam
meperidine
meperidine (nor)
mephedrone
methadone
methamphetamine
methylenedioxyamphetamine
methylenedioxyethylamphetamine
methylenedioxymethamphetamine
midazolam
mirtazapine
morphine
nortriptyline
olanzapine
oxazepam
oxycodone
oxymorphone
paroxetine
pheniramine
pseudoephedrine
quetiapine
risperidone
sertraline
temazepam
tramadol (cis)
trazodone
venlafaxine
zopiclone

Antiepileptic method (LC-MS/MS)
baclofen
gabapentin
topiramate
vigabatrin

Cannabinoid method (LC-MS/MS)
D-9-tetrahydrocannabinol (THC)
11-nor-carboxy-D-9- tetrahydrocannabinol (Carboxy-THC)

Digoxin method (LC-MS/MS)
digoxin
digitoxin (qualitative)


Appendix 6

Capability of targeted qualitative methods

Urine Drug Mix (UDM; LC-MS/MS)
6-monoacetylmorphine (6-MAM)
acetylfentanyl
acetylnorfentanyl
alprazolam
amitriptyline
amlodipine
amoxapine
amphetamine
baclofen
benzoylecgonine
bromazepam
brompheniramine
buprenorphine
buprenorphine glucuronide
bupropion
chlordiazepoxide
chlorpheniramine
citalopram
clobazam
clomipramine
clonazepam
clonazepam (7-amino)
clozapine
cocaethylene
cocaine
codeine
codeine-6-glucuronide
cyclobenzaprine
demoxepam
desipramine
desomorphine
dextromethorphan
diazepam
diazepam (nor)
diltiazem
diltiazem (desacetyl)
diphenhydramine
doxepin
doxylamine
duloxetine
ephedrine
fentanyl
flunitrazepam
flunitrazepam (7-amino)
flunitrazepam (N-desmethyl)
fluoxetine
fluoxetine (nor)
flurazepam
flurazepam (n-desalkyl)
fluvoxamine
gabapentin
heroin
hydrocodone
hydromorphone
hydromorphone-3-glucuronide
hydroxyalprazolam
hydroxytriazolam

imipramine
ketamine
ketamine (nor)
lamotrigine
levorphanol
lidocaine
lorazepam
lorazepam glucuronide
loxapine
meperidine
meperidine (nor)
mephedrone
methadone
methamphetamine
methylenedioxyamphetamine
methylenedioxyethylamphetamine
methylenedioxymethamphetamine
methylenedioxypyrovalerone
methylone
metoprolol
midazolam
mirtazapine
morphine
morphine-3-glucuronide
morphine-6-glucuronide
naloxone
naltrexone
nitrazepam
nitrazepam (7-amino)
norfentanyl
nortriptyline
O-desmethylvenlafaxine
olanzapine
orphenadrine
oxazepam
oxazepam glucuronide
oxycodone
oxymorphone
paroxetine
pentazocine
phenazepam
phencyclidine
pheniramine
propoxyphene
propranolol
pseudoephedrine
quetiapine
risperidone
sertraline
tapentadol
temazepam
temazepam glucuronide
topiramate
tramadol (cis)
trazodone
triazolam
trimipramine
venlafaxine
zaleplon
ziprasidone
zolpidem
zopiclone

Glossary

Abbreviations
Analytical results are reported in terms of mg/100 mL, mg/L, or ng/mL, as shown below:

g - gram
mg - milligram
µg - microgram
L - litre
mL - millilitre
ng - nanogram

Breakdown Product
A compound produced either inside or outside the body that may or may not be pharmacologically active.

Carboxyhemoglobin saturation

The percentage of hemoglobin bound by carbon monoxide.

Central Nervous System Depression (CNS depression)
A lowering of the functional activity of the brain and/or spinal cord. Depression of the respiratory and the cardio-regulatory centres are most relevant toxicologically.

Coroner's Case Analytical Summary
Contains analytical results with general notes regarding the reported drug concentrations. In absence of a note, the detected drug concentration is considered to be toxicologically insignificant. No comment on therapeutic efficacy is implied by the detection of a drug.

(D) Could Cause Death
The detected drug concentration could cause death.

Detected
The drug has been identified in the sample. Identification is based on criteria specific to the analytical technique.

Fatal Reference
A minimum drug concentration at which death has been reliably reported in the forensic literature.

(I) Interpretive Data Note
Provides additional information concerning the result.

Inconclusive
The presence or absence of a drug could not be determined.

Metabolite
The product of enzymatic conversion of a drug within the body to a different compound that may or may not be pharmacologically active.

No [other] significant findings by a [method name(s)]
This comment is inserted to provide a reference to the methods that were used. Appendix 1 and 5 above can be used to identify compounds not listed and that were either not detected or the results were deemed to not be toxicologically significant, e.g., caffeine or nicotine. This may also apply to endogenous compounds, e.g., acetone < 2 mg/100 mL.

Not Detected
The drug is either not present or is present but at an amount that cannot be discerned from other constituents in the sample.

Post-mortem redistribution
A phenomenon that refers to a change (either an increase or a decrease) in blood drug concentration after death; post-mortem redistribution may occur regardless of sampling site but is commonly observed as increased drug concentrations in heart blood as compared to femoral blood.

Putrefaction
The decomposition of organic material that involves micro-organisms.

Report
Contains a comprehensive summary of analytical results accompanied by interpretative conclusions.

(T) Toxicologically Significant
The detected drug concentration may produce toxicity.

Tentative
A drug, or class of drugs, has been identified using a non-specific screening method or has not been confirmed by additional methods. For positive identification further analysis is required.

Therapeutic
The detected drug concentration is generally considered to not be toxicologically significant. The use of this term does not imply clinical efficacy.

Traces
The drug was detected at a concentration below that which can be reliably quantitated. The use of this term does not imply clinical efficacy.