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Forensic
Applications
Carboxy-delta-9-THC (pKa 4.5) in Urine for GC/MS Confirmations
Using 200mg 10mL HyperSep Verify-CX Extraction Column
(Part Number: 60108-742)
Prepare Sample – Base Hydrolysis of Glucuronides
To 2mL of urine add internal standard* and 100µL of 10
M NaOH
Mix/vortex
Hydrolyze for 20 minutes at 60°C
Cool before proceeding
Adjust sample pH to 3.0 with approximately 1.0mL
of glacial acetic acid (check pH to ensure that the pH
value is
≈
3.0)
Condition Verify-CX Extraction Column
3mL of CH
3
OH
3mL of DI H
2
O
1mL of acetate buffer (pH 3.0)
NOTE:
Aspirate at <3"Hg to prevent sorbent drying
Apply Sample
Load sample at 1 to 2mL/minute
Wash Column
2mL of DI H
2
O
2mL of 100mM HCl/acetonitrile (95:5)
Dry column (5 to 10 minutes at >10"Hg/full flow for
positive pressure manifold)
200µL of hexane then aspirate (additional step to remove
any residual moisture)
Elute Carboxy THC
3mL of hexane/ethyl acetate (50:50)
Collect eluate at 1 to 2mL/min
NOTE:
Before proceeding, ensure there are no water droplets at
the bottom of the collection tube (this may increase drying time
and decrease BSTFA derivatizing efficiency)
Dry Eluate
Evaporate to full dryness at <40°C under a stream of
nitrogen
Derivatize**
Add 50µL of ethyl acetate and 50µL of BSTFA
(with 1% TMCS)***
Mix/vortex then react for 20 minutes at 70°C
Remove from heat source to cool
NOTE:
Do not evaporate BSTFA
Quantitate
Inject 1 to 2µL onto gas chromatograph
For GC/MS monitor the following ions:
Compound
Primary Ion** Secondary Tertiary
D
3
-Carboxy-delta
9
-THC
374
476
491
D
9
-Carboxy-delta
9
-THC*
380
479
497
Carboxy-delta
9
-THC
371
473
488
* Suggested internal standard
** Quantitation ion
*** Part number TS-38831 (10 x 1mL ampules)
Recommended GC Columns
Part Number
TRACE TR-DoA5 Column, 15m x 0.25mm x 0.25µm
26AF130P
TRACE TR-DoA35 Column, 15m x 0.20mm x 0.33µm
26AC497P