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Forensic
Applications
Antidepressant/Painkillers in Blood and Urine
Using 60mg 3mL HyperSep Retain-CX Extraction Column
(Part Number: 60107-303)
Sample Preparation
To 1mL of 100mM phosphate buffer (pH= 6) add
internal standard*
Add 1mL of blood or urine
Add 2mL of 100 phosphate buffer (pH= 6)
Mix/vortex
Sample pH should be 6.0±0.5
Adjust pH accordingly with 100mM monobasic or
dibasic sodium phosphate
Mix/vortex
Centrifuge as appropriate
Condition HyperSep Retain-CX Extraction Column
1 x 3mL CH
3
OH
1 x 3mL H
2
O
1 x 1mL 100mM phosphate buffer (pH= 6)
NOTE:
Aspirate at <3"Hg to prevent sorbent drying out
Apply Sample
Load sample at 1 to 2mL/minute
Wash Column
1 x 3mL DI H
2
O
1 x 3mL 1% acetic acid
1 x 3mL methanol
Dry column (5 minutes at >10"Hg)
Elute
1 x 3mL ethyl acetate;
ACN:ammonia(78:20:2 v/v)
Collect eluate at 1 to 2mL/minute
Evaporate eluate under a gentle stream of nitrogen <40ºC
Analysis
Reconstitute sample in 100µL of methanol
Inject 5µL onto GC/MS
Compound
MRM Transition
Amitriptyline
278.8/91.1
Amitriptyline-D3*
281.2/91.2
Diphenhydramine
256.2/167.1
Diphenhydramine-D3*
259.2/167.1
Doxepin
280.2/107.1
EDDP
278.2/234.2
EDDP-D3*
281.4/234.3
Methadone
310.2/105.1
Methadone-D9*
319.2/268.3
Nortriptyline
264.2/91.1
Norpropoxyphene
326.2/44.1
Norpropoxyphene-D5*
331.1/267.1
Propoxyphene
340.2/58.1
Propoxyphene-D11*
351.3/64.0
Sertraline
308.1/161.0
Tramadol
264.2/58.1
Tramadol-D3*
268.2/58.0
Venlafaxine
278.2/58.2
Zolpidem
308.2/235
* Suggested internal standards
Recommended HPLC Column
Part Number
Hypersil GOLD 3µm, 150 x 2.1mm
25003-152130