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Forensic
Applications
Barbiturates in Urine for GC or GC/MS Confirmations
Using 200mg 10mL HyperSep Verify-CX Extraction Column
(Part Number: 60108-742)
Prepare Sample
To 2mL of urine add internal standard(s)* and 1mL of
100mM phosphate buffer (pH 5.0)
Mix/vortex
Sample pH should be 5.0±0.5
Adjust sample pH accordingly with 100mM monobasic
or dibasic sodium phosphate
Condition Verify-CX Extraction Column
3mL of CH
3
OH then aspirate
3mL of DI H
2
O then aspirate
1mL of 100mM phosphate buffer (pH 5.0) then
aspirate
NOTE:
Aspirate at <3"Hg to prevent sorbent drying
Apply Sample
Load sample at 1 to 2mL/minute
Wash Column
3mL of DI H
2
O
1mL of 100mM acetic acid
Dry column (5 minutes at >10"Hg)
2mL of hexane
Elute Barbiturates
3mL of hexane/ethyl acetate (50:50); Collect eluate at
1 to 2mL/minute
Dry Eluate
Evaporate to dryness at <40°C
Reconstitute with 100µL ethyl acetate
Optional Derivatization
Add 25 to 50µL of 0.2M TMPAH***
Reaction occurs in the injection port
Quantitate
Inject 1 to 2µL onto gas chromatograph
For mass spectrometry, monitor the following ions (GC/MS):
Underivatized
Drug
Primary Ion** Secondary Tertiary
Amobarbital
156
141
157
Butalbital
156
141
157
Butalbital
168
167
181
Hexobarbital*
221
157
236
Pentobarbital
156
141
197
Phenobarbital
204
232
117
Secobarbital
168
167
195
Thiopental
172
157
173
Derivatized
Drug
Primary Ion** Secondary Tertiary
D
5
-butalbital
201
214
Butalbital
196
195
209
Amobarbital
169
184
185
Pentobarbital
169
184
112
13
C
4
-secobarbital
200
185
Secobarbital
196
195
181
D
5
-phenobarbital
237
151
Phenobarbital
232
146
175
* Suggested internal standard
** Quantitation ions (target ions in bold)
*** Part number TS-49301 (MethElut reagent 12x1mL vials)
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