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Forensic
Applications
Gamma-Hydroxybutyrate (GHB) in Urine without Conversion to
Gamma-Butrylactone (GBL)
Using 200mg 6mL HyperSep Retain-AX Extraction Column
(Part Number: 60107-412)
Sample Preparation
200µL of urine
Add internal standard* and 100µL of 100mM phosphate
buffer (pH 6.0)
Mix/vortex
Condition HyperSep Retain AX Extraction Column
1 x 3mL CH
3
OH
1 x 3mL DI H
2
O
1 x 1mL 100mM phosphate buffer (pH= 6.0)
Apply Sample
Place test tubes into vacuum manifold for collection
The sample loading and wash are both collected
Decant sample onto column. Aspirate at ~1"Hg
Wash Column
Add 1mL of CH
3
OH/NH
4
OH (99:1) to original sample
test tube
Mix/vortex
Decant wash onto column
Evaporate to dryness at 60°C using a stream of air or N
2
Sample Clean-Up
Add 200µL of dimethylformamide
Add 1mL of hexane saturated with dimethylformamide
Mix by inversion for 5 minutes
Centrifuge at 3,000rpm for 5 minutes
Transfer lower dimethylformamide layer to a clean test tube
Evaporate to dryness at <50°C using a stream of air or N
2
Derivatize
Add 100µL ethyl acetate and 100µL BSTFA
(with 1% TMCS)**
Mix/vortex
Analysis
Inject 1 to 2µL onto GC/MS
For mass spectrometry monitor the following ions:
Compound
Primary Ion *
Secondary
Tertiary
GHB-D6-di-TMS*
239
240
241
GHB-di-TMS
233
234
235
* Suggested internal standard for GC/MS: GHB-D6
** Part number TS-38831
Recommended GC Column
Part Number
TraceGOLD TG-35MS 30m x 0.25m x 0.25µm
26094-1420