Toxicology milestones
The Thermo legacy
1939: DDT Excellent insecticide= excellent lifesaver
1956: Minimata Mercury poisoning
1961: society of toxicology founded
1970:EPA founded and ban of cocaine in the US
1972: DDT is banned 1937: ban on marihuana
1928: ban on doping
1984: Bhopal 1995: Sarin attack in Tokyo
1999 Dioxine crisis in Belgium
1948: Foundation ATLAS MAT
1958: start of Carlo Erba SID and MAT CH4 first commercial succesful GCMS
1969: first splitless injection by Grob on Carlo Erba GC
1969: Finnigan Model3000GCMS 1975: first capillary GC by Carlo Erba
1987: MAT90 first PC controlled GCMS
1990s: MD800 and GC8000
Joined company forces
1920 1950 1968 1980 1990
YOUR NEEDS TODAY ARE STILL ADDRESSED Sample prep: as less as possible; as quick as possible
Software: Complete with data validation
Instrumentation: Rugged and robust; fast and reliable
Total workflow: Optimized production with high quality results
•Sample preparation reagents and solutions for various matrices
•Blood, Urine, Oral Fluid hair and other bodily fluids
•Accurate separation of components analogues
•Avoid misinterpretation of target compounds
•Avoid metabolic product exclusion or interference
•Opioids - Keto-moeity derivitization
•Removal of noise associated with sample matrix components
•Higher recovery, Better reproducibility
•Less noise = Better chromatography
•Hyper Sep Verify - CX
•Enhance target compound stability
•Better detection
SAMPLE PREP: INJECTING BLOOD, SWEAT AND TEARS ON A
GC Sample Matrix
Derivatization
Solid Phase Extraction
Multiple Derivitization
KEYPOINTS AND SOLUTIONS FOR THE PRODUCTIVE LAB a large number of samples in the trays
Up to 648 2-mL vials and multiple 100 mL solvent bottles stations
Unattended, injection modes changes in a sequence: liquid – liquid Large Volume - HS - SPME
ATC (Automatic Tool change) Station for automatic recognition and syringe exchanges during
operation
Reliable, reproducible injections with microvolume samples using the bottom sensing sensor
<2%RSD and up to 3 injections possible with 5 ml sample volume in vial.
Advanced sample handling and sample preparation
Dilution, derivatization, standard addition, internal standard additions, etc.
Barcode reading for avoiding writing mistakes
Set up possible for two GC’s
Completely adaptable to your wishes:
cooled/heated trays;
larger washing stations;
HS ovens;
vortex for mixing;
and more……….
just ask us for implementation of your idea!
KEYPOINTS FOR THE LAB: Modular approach:
change injectors and detectors in 3 minutes only
disassemble and sonicate the injector parts
Injection modes include:
SSL
SSL with backflush
PTV
PTV with backflush
all are compatible with Large Volume without any hardware change
Fast cooling of the oven and PTV for fast run to run times;
from 350C to 40C in approximately 4 minutes
extra fast cooling with cryo option for oven and injector
Methodology is completely compatible with third party vendors
little RT difference
same injection parameters
Quick start up; Fast methodology; Easy maintenance
Generate results shortly after installation;
Ensure production over a long period of time
THE ISQ: SINGLE QUAD MS
Heated source: ion volume, repellor, lenses, RF prefilter
Solid, highly inert material ensures reliable performance in all
ionization modes, including EI and CI
Dual filament cartridge
Two filaments in same magnetic orientation
Filament lens assures that both filaments will remain protected, for
longer life even with redundancy
The S-Shaped Ion Guide
Reduces neutral noise created by excited helium neutrals striking
the detector, resulting in lower detection limits and better selectivity
Autotune:
Complete and standardized autotune algorithm for excellent day to
day variability
THE ISQ: TRUE FAST SCANNING d:\training\fs2802 28/02/2010 11:33:35
new 1pgOFN, at scan rate 2500 amu/s, factory spec
RT: 3.502 - 3.753 SM: 5B
3.55 3.60 3.65 3.70 3.75
Time (min)
0
10000
20000
0
10000
20000
0
5000
10000
15000
20000
Re
lative
Ab
un
da
nce 0
5000
10000
15000
624 625623
621 627
620 628619
1201 12031200
12061198
12091196
1212
2262 226322602267
22572270225522732251
45314530 4533
454145224546451645544512
NL:1.79E4
m/z= 271.500-272.500 MS fs2802
NL:2.04E4
m/z= 271.500-272.500 MS fs2803
NL:2.28E4
m/z= 271.500-272.500 MS fs2806a
NL:2.40E4
m/z= 271.500-272.500 MS fsre2801
Theoretical
(scan/sec)
Actual
(scans/sec)
9.62 9.67
18.52 18.58
35.00 35.00
70.13 70.00
THE WIRELESS SOURCE: EASY MAINTENANCE
Step 1. Insert removal tool
Step 2. Remove source
Step 3. Hot source is held in tool Step 4. Push source out of tool
COCAINE AND MAJOR METABOLITES IN URINE Total ion chromatogram of cocaine and it metabolites from an extracted urine sample at 150 ng/mL
Confirmation
analysis
THC IN URINE
Validated based on guidelines published by EWDTS
- Linearity, Carryover, Inter/Intraday Precision and Robustness
Confirmation
analysis
THC in urine
Precision
- Intra-and inter-batch precision of less
than 4% CV at 6 and 18.75ng/mL
Robustness
-100% of all QC samples at passed
across a study spanning 7 weeks in over
100 batches.
Minimal Instrument Maintenance
-Daily septa replacement
-Daily tune
-liner replacement - as needed
-1 Ion source for the duration of the
experiment
Confirmation
analysis
TRIPLE QUADRUPOLE MS – PRINCIPLE OF
OPERATION Quantitation of target compounds in matrix samples
Q1 selects the precursor ion TCDD ion: m/z 319.90
Q3 selects the product ion fragments to m/z 256.90
Argon
Collision Gas
Select Fragment Detect
Ion S
ourc
e
Dete
cto
r
SRM: selected
reaction monitoring
RT: 25.70 - 26.46 SM: 5G
25.7 25.8 25.9 26.0 26.1 26.2 26.3 26.4
Time (min)
0
20
40
60
80
100
0
20
40
60
80
100
Re
lative
Ab
un
da
nce
0
20
40
60
80
10026.36
26.3226.18 26.4025.83 25.89
25.9825.72 25.79 26.0926.04 26.25
RT: 26.03SN: 18RMS
RT: 26.11SN: 9
26.35
26.17 26.4525.79 26.0625.70 25.97 26.3026.2025.83 25.92
RT: 26.02SN: 136880RMS
RT: 26.11SN: 108979RMS
NL: 6.62E8
TIC F: MS 2407-14
NL: 2.54E6
m/z= 291.5-292.5 F: MS 2407-14
NL: 1.37E5
m/z= 256.5-257.5 F: + c SRM ms2 [email protected] [ 190.00-300.00] MS ICIS 2407-06
MS/MS vs SIM for detecting pcb’s in soil samples.
RT: 19.80 - 43.03 SM: 5G
20 22 24 26 28 30 32 34 36 38 40 42
Time (min)
0
5
10
15
20
25
30
35
40
45
50
55
60
65
70
75
80
85
90
95
100
Re
lative
Ab
un
da
nce
26.94
26.36
32.12
27.75 31.8633.24
30.6633.94
34.31
28.07
28.6325.6334.63 36.1330.0925.57
37.1624.3740.42
38.4823.54
23.30
40.9221.45
41.47
21.39
NL:7.79E8
TIC F: MS 2407-14
PCB’s in MS/MS
s/n 14000
Complete FS
chromatogram
Selective ion mode
s/n 18
PCB’s in FS
Increased S/N:
-Lower detection limits
- automatic integration by
software much more repeatable
and reliable
SHORT OVERVIEW
Dual Filaments
for Prolonged
Uptime
Repellor for
longer source
uptime
Heated ion guide
protects main
quadrupole sets so
they never have to be
cleaned or replaced.
Curved ion
guide enables
off-axis optics
to effectively
eliminates
neutral noise
Choose between
nitrogen or argon for
collision gas.
Full source
removal without
venting, including
ion guide surface.
No wire
connections are
necessary, and
never vent to clean
your instrument.
Super fast analyzers switch
between Full Scan, SIM and
MRM in milliseconds.
KEY POINTS FOR THE PRODUCTIVE LAB
•Repeller
•Allows effective tuning even when parts get contaminated
•Increased ion abundance stability and tune-ability
•Dual filament cartridge
•Two filaments for reducing instrument downtime
•Both filament operate in the same magnetic orientation for an increased life cycle
•Alignment insure to get same ionization performance while switching filaments
•Increased temperature all along the ion path
•Reduces residence of the matrix that form the basis of ion burns
•Dual heated zone for more ruggedness
•Increased diameter prefilter and lenses
•Dilutes effect of contamination
The TSQ 8000 features the ExtractaBrite ion source, the first
ever source on a GC-triple quadrupole in which all parts
that require periodic maintenance can be removed and
inserted while maintaining vacuum, greatly reducing
downtime for source maintenance.
UNSTOPPABLE PRODUCTIVITY – VENT-FREE
MAINTENANCE
DOA IN ORAL FLUID
Results of analysis: DOA in oral fluid
Confirmation
analysis
• Methadone D9
• Methadone
• Cocaine D3
• Cocaine
• BZE D3
• BZE
• Nordiazepam D5
• Nordiazepam
• THC D3
• THC
• Oxazepam D5
• Oxazepam
• Dihydrocodeine
• Codeine D3
• Codiene
• Lorazepam D4
• Lorazepam
• Diazepam
• Morphine D3
• Morphine
• 6am D3
• 6am
• Temazepam D5
• Temazepam
• THC Acid D3
• THC Acid
Run time 9.5 mins
THC IN ORAL FLUID
Agilent application note 2D GCMS: selectivity in chromatography Sample prep: - 400 ul oral fluid - SPE clean up - derivatization with BSTFA and TCMS - evaporate to dryness - add 25 ul acetonitrile - 4ul injection
- injected on column 11ng
Thermo solution GC-HSRM: selectivity in detection Sample prep: - 200 ul oral fluid - SPE clean up - derivatization with BSTFA - evaporate to dryness - add 50ul toluene - 2ul injection
- injected on column 16ng
Results of analysis: THC in oral fluid at 0.2ng/ml
Confirmation
analysis
DETERMINATION THCA IN HAIR •Delta 9 THC (THC) is the most commonly abused illicit drug.
•The metabolite THCA indicates the ingestion of THC as opposed to passive contamination of the
hair through the air.
•Hair is a very complex and variable matrix even after sample cleanup
Small Samples
typically 20mg
Low Concentrations
40 fg on column
Low Level Detection
Cutoff of 0.05 pg/mg
Overcome Matrix Interference
Accurate Quantitation
Detection limit of 20% of cutoff
Confirmation
analysis
•Sensitivity of GC Quantum XL to THCA
•On-Column Injection of 8fg
•Use of a 0.05 pg/mg cutoff
•Demonstrates the sensitivity sufficient for published cutoffs
DETERMINATION THCA IN HAIR
MSTFA Derivatized, EI, SRM Transition: 371 > 289
• NCI - Specific transitions can be defined in NCI (Methane used as reagent gas),
enhancing selectivity for both THC-COOH and the corresponding ISTD D3
Confirmation
analysis
•Demonstrates very low noise level relative to the signal
•Zoom in the baseline by 100 times before we start seeing significant noise
•Possible to drop the concentration by a factor of 10 and still achieve the selectivity to accurately quantitate THCA
THCA IN HAIR - NCI TRANSITIONS
PFAA Derivatized, Negative Ion Chemical Ionization - 50 fg THCA and ISTD-D3
Confirmation
analysis
METHODOLOGY: SHORT DESCRIPTION •Hair strand is washed with deionized water and acetone, dried and cut into small pieces;
•20 – 30 mg hair snippets are powdered in a ball mill
•1ng EtG-D5 as internal standard is added
•15 min ultrasonic extraction with 1.5ml deionized water
•SPE clean up
•Derivatization with PFPA
•PTV injection: 250C (5.5min)-14.4C/sec-280C- backflush on 5min
•GC: 80C (3min)-25C/min-270C (4min)
•He: 2ml/min
•Sample volume 1ul
•Transferline: 200C
•Source Temperature: 140C
•CI gas: methane 2ml/min
DETERMINATION OF ETG IN HAIR
Maintenance Classic injection with SSL and
single quad
‘Smart’ injection with PTV backflush and
Quantum
Improvement
Liner Every 20 inj 100 inj 5
Precolumn 20 inj 100 inj 5
Analytical column 400-500 inj 1000-1500 inj 3
Ion volume 400-500 inj 1000-1500 inj 3
Added bonus: • GC Run time: 2 times shorter • LOD: 10 times better
GC/MS
Confirmation and Quantitation of an Expanded Panel of Sympathomimetric Amines in Urine
Using Single Quadrupole GC/MS
High-Throughput Confirmation and Quantitation of a THC Metabolite in Urine Using Single
Quadrupole GC/MS
Confirmation and Quantification of Phenclidine in Urine Using the ISQ Single Quadrupole
GC-MS
Confirmation and Quantification of Benzoylecgonine in Urine Using the ISQ Single
Quadrupole GC-MS
Confirmation and Quantitation of Cocaine and Major Metabolites in Urine Using the ISQ
Single Quadrupole GC-MS
Quantum (TSQ) XLS
Confirmation and Quantitation of THC in Oral Fluid Using MRM Method
Quantitation of EtG in Hair using PTV backflush-GC-NCIMSMS
APPLICATION NOTES
HOT LIST: MEPHEDRONE - NEW DESIGNER
DRUG • Mephedrone is illegal since 2010 across Europe.
• The drug is very popular as a party drug and so far little analytical experiments have been performed.
• Recently published by Westphal and Junge
• Massenspektrometrische, infrarotspectroskopische und NMR spektroskopische Daten von Mephedron, butylon und
Methylon sowie einigen ihrer Derivate
• System: TSQ7000. Predecessor of QuantumXLS
TRACEFINDER: METHOD DEVELOPMENT • Import from Xcalibur processing method, or from
a datasheet; or through automated library search
• Automatic input for quan and qual ions
• Add quality controls for the method:
• Linearity demands
• Internal standard recovery
• Blancs and negatives
• Quality control samples
• Ion ratio demands
• LOD,LOQ and LOR
• Add automated library search for metabolites
Data evaluation
TRACEFINDER: PRODUCTION • Batches and batch templates for quick sample
list set up
• Sample types include specimens, negatives,
calibrators
• Colour coded flags for QC controls as set uo in
method development
• Active link between reporting and data
• Update RT; integration; ion ratio; QC settings in
• Local method: this set of data
• Master method: all following set of data
• Choose between over 30 report types
• Export to LIMS
• All results, data, methodology in one directory
Data evaluation
CONCLUSION: A TOTAL SOLUTION FROM PREP
TO REP
• Fast and easy sample preparation procedures are available
• A robust and sensitive instrumentation is provided
• Coupled with complete method information for fast start up
• Data handling is fast and the software provides fully embedded qualtity controls