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Toxicology in alternative matrices

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Calvin Chong

on 16 December 2013

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Transcript of Toxicology in alternative matrices

Toxicology:
traditional method applied in alternative matrices
vs
alternative methods applied in traditional matrices

A summary of presentations
in the TIAFT 2013 Scientific meeting
Calvin Chong
"Traditional methods"
HPLC-DAD
GC-MS
LC-MS/MS

Thin-layer chromatography
Colourimetric tests
Immunoassays

"Alternative methods"
MALDI-MS(/MS)
GC-MS/MS

"Traditional matrices"
Serum
Urine
Vitrous humour
Hair

"Alternative matrices"
Hair
Dried blood spot
Nails
Breath

Hair

Used since 1979 for forensic case, clinical case, workplace drug testing, abstinence and compliance, particularly in pain therapy.
The traditional aspect

Enzyme-linked immunoassays, Capillary electrophoresis, GC-MS, LC-MS/MS
The traditional methods

MALDI-Imaging MS (since ca. 2011)
The not so traditional methods discussed
MALDI-MS
Sample
Preparation
Wash with 0.5% SDS, then sonicated with distilled water and methanol alternatively for three times
Affixed with conductive adhesive tape to an indium tin oxide-coated glass slide
Manual razor-cutting of hair
Visible wave length acquisition on a flatbed scanner prior to matrix deposition
7 mg/mL alpha-cyano-4-hydroxycinnamic acid in ACN & 0.2% trifluoroacetic acid
MALDI
TOF-IMS
Matrix-assisted laser-desorption ionization Time-of-flight imaging-MS
Scanning each "Pixel" through with a TOF scan, with SRM configured with parent mass and fragment mass (e.g. 150/119 for methamphetamine)
Pros and cons of MALDI-IMS
Pros
Allows more precise definition of sequence of drug use in hair
Even a single dose of zolpidem was detectable (on Bruker FTICR-MS)
Essentially non-destructive
Cons
Very low through-put (requires manual razor-cutting as laser cutting and freeze fracturing may cause loss of analyte such as methamphetamines)
Expensive equipments (and not already available in TRL)
Does not differentiate well in ring-substituted isomers (require chromatographic-MS confirmation)
Dried
Blood
Spots

Most commonly used in newborn screening. Also utilized in measurement of hormones, vitamin D, haemoglobinpathies, and even HIV infections
The traditional aspect

It was discussed in two different aspects:
As a means of sample preparation (Ambach et al, Rossi et al, Cuypers et al)
As a sample type (Wood et al)
Discussion points
Ambach L et al. (Univ of Bern)
Rapid and simple LC-MS/MS screening of 64 novel psychoactive substances using
dried blood spots

Rossi S et al. (Univ of Sacred Heart, Rome)
Sample Prep: Preliminary study on the application of different techniques to fresh and cadaveric blood for the determination of drugs of abuse and benzodiazepines

Cuypers E et al. (Catholic University Leuven)
Are dried blood spots usable in postmortem forensic toxicology?

Wood M, Dargan P et al.
(Waters corp, Guys & St Thomas)
Broad toxicological screening results from suspected GHB/GBL overdose cases
.
Dried blood spots as a sample preparation step
Pipetting 10 uL of whole blood into dried blood spot cards and left to dry
Extracted with MeOH, followed by blow-dry and reconstitution in mobile phase
Followed by LC-MS/MS
LOD:
1.0-10 ng/mL with cathinones, piperazines, tryptamines, etc on an ABI 5500 QTrap (Ambach et al)
5 ng/mL for majority of benzodiazepines (Rossi et al)
Dried blood spot as a specimen type
Utilizes MS(E) technique from Waters corp's UPLC-TOF/MS to do general unknown screening, for GHB/GBL as well as DOA such as mephedrone, methamphetamine, and ketamine
Probably useful esp for cases with low sample volume, and where whole blood is available
Poetzsch S et al. (Univ of Zurich)
Systematic Investigations of MALDI-mass spectrometry imaging for the use in single hair analysis.

Kamata T et al. (Osaka Police)
Single-hair analyses of hypnotics on the supposition of a single administration in drug-facilitated crimes.

Miki A et al. (Osaka Police)
Hair analysis for drugs by mass spectrometry imaging: a new perspective to investigate drug use history.


Shen M et al. (Ministry of Justice, PRC)
Mass imaging of ketamine in single scalp hair by MALDI-CASI-FTMS

Duvivier W et al. (Wageningen, Netherlands)
A new approach in hair forensics
Nails
Drugs are incorporated into nails in both ways: germinal matrix (accounts for 80%) and nail bed (accounts for 20%)
Nails are slow growing
1cm/3 months for fingers; 1cm/6 months for toenails

5 - 10 mg of nail
extraction by mechanical pulverization followed by sonication and extraction with methanol
AB Sciex QTrap 5500 / Phenomenex C18 Column
Analytical method

Madry M et al (Univ of Zurich)
Incorporation mechanisms of zolpidem in fingernails - a systematic investigation
after a single dose
Performance
Characteristics
Detection Window: Approximately 3-4 months
Three 'peaks' corresponding to
Sweat deposition
Nail bed deposition
Germinal matrix deposition
Confirmation requires repeated sampling
Probably more useful for monitoring than for diagnosis, and most likely less useful than hair analysis
And compared with hair...
Less specimen available
Essentially, 20 potential sample site per person
Less hydrophobic matrix
Probably poorer retention of drugs
More easily washed out
More easily contaminated by sweat
May be worthwhile as a bridge between body fluid analysis and hair analysis
Cost-effectiveness issues
Exhaled
Breath

Exhaled breath contains (1) water, (2) water soluble volatile compounds, and (3) aerosol particles, which contain surfactants (carriers of non-polar compounds) among other ingredients.
The science
Discussion points
Beck et al. (Karolinska Institutet)
Detection of drugs of abuse in exhaled breath using a device for rapid collection - comparison with plasma, urine, and self-report in 47 intoxicated drug users.
Extraction from commercial collecting device
Commercial specimen collecting device already available
Essentially, a device that allows air to be blown in through a piece of filter paper
Extraction by ACN followed by SPE at basic pH (SPEC DAS, pH 10)
Sensitivity of Assay
The good: able to detect methadone in all specimens collected from a group of patients given methadone
The ugly: the detection window using an LC-MS/MS platform for THC is only about 2 hours following inhalation
Remains an investigatory matrix
Exhaled breath has been explored as a potential matrix since ca. 2010. initially with the subject blowing directly into a SPE...
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