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Clinical/Forensics
Fast Screening and Confirmation for Gamma-Hydroxybutyrate (GHB)
(con tinued from page 3)
the ir euphori c and seda tive properties. 1,4
butanediol and GEL are quickly metabolized to
GHE after ingestion and are analyzed as such.
Because GHE is endogenous in humans, and has a
half-life of one hour or less after injection, it is very
important to collect biofluids (typically blood or
urine) quickly for toxicological investigation.
Analytical methods for GHB usually emplo y gas
chromatography and mass spectrometry for quan
tification and confirmation . The methodology
described here establishes a headspace GC-FID
screening procedure followed by confirmation and
quantification by headspace
GC/MS,
and was
developed by the FBI Chemistry Unit.' We have
adapt ed Rtx®-BACI and Rtx®-BAC2 columns
with court-tested and pro ven performance in
blood alcoho l analyses- and new, highly inert
Rxi™-SMS columns to the methods.
A typical headspace GC-FID blood alcohol system,
using an Rtx®-BACI column or an Rtx®-EAC2
column, can be adapted for GHB screening. For
the analysis, GHB is converted to gamma-butyro
lacton e (GBL) to impro ve chromatography, and
alpha -methylene-gamma-butyrolactone (AMGB)
is used as the internal standard. Figure 1 illustrates
the conversion reaction of GHE to GBL. Figure 2
shows that either Restek column is suitabl e for
GHB screening , providing Gaussian peak shapes,
baseline resolut ion , and an analysis time of less
than 5 minutes.
A sample yieldin g positive screening results
requi res confirmation and qu ant ification by
GC/MS. The confirmation and quantification
analysis incorp orat es the same headspace and GC
conditions, including conversion of GHB to GEL,
but GBL-d6 is the required internal standard. To
illustrate GBL and GEL-d6 separation and peak
shape on an RxjTM-5ms column we analyzed Iul, of
a standa rd, using GC/MS. (Figure 3). Thi s typical
liquid injection shows the two compounds are par
tially resolved on the Rxi™-Sms column, and pos
itively ident ified using full scan. Then, extracted
ion data (EI) were obtained(Figure 4). After posi
tive identification, GHB is quantified by compa r
ing the areas of the deuterated and und euterated
GEL extracted ions.
Figure 1 Conversion of GHB to GBL.
- - - - .HO
~
~OH
gamma-hydroxybutyrate
gamma-butyrolactone
(GHB)
(GBL)
2006.03
Figure 2 Symmetric peak for GHB, and baseline resolut ion from an
inte rnal standard in less tha n 5 minutes, using an Rtx
llD
-BACl or
Rtx
llD
-BACl Column .
Rtx
llD-BACl
column
Sharp peaks and baseline
resolution in less than 5 minutes
12 samples per hour!
1. gamma-hydroxybutyrate (GHB) / gamma-butyrolactone (GBl)
2. alpha-methylene-gamma-butyrolactone (AMGBl)
i
I
I
I
I
I
I
4
6
8
10
Time(min)
GC_PIlOO870
2
Rtx
llD
-BAC2 column
1
I
0
I
I
2
I
i
4
1
Time(min)
I
6
1
I
8
i
I
10
GC..
PHOO871
Column:
Rtx'-BACl 30m, 0.32mm !D,
1.8IJm
(cat.# 18003) and
Rtx' -BAC2 30m, 0.32mm!D, 1.2pm(cat.# 18002),
connectedvia universal"Y" Press-Tight' connector (cat.# 20405)
Sample:
GHB, GBL,cx-methylene--y-butyrolactone(AMGBl), lO/Jg/ ml each in water
Inj.:
1.0mlheadspace, split (splitratio 1:10), l rnrnsplit inlet liner (cat.# 20972)
Inj. temp.:
200' C
Carrier gas:
helium, constant pressure
linear velocity:
44cm/sec.
@
50' C
Oven temp.:
50' C(3 min.) to 150'C
@
20' C/min. (hold7 min.)
Det:
F!D
@
240'C
Headspace autosampler: Teledyne/Tekmar HT3
Sample/ platen temp.:
100' C
Sample equilibration:
15min.
Mixing time:
5 min.
Vial pressure:
10psig
Vial pressurization
time:
2min.
l oopfill time:
2min.
Transfer line temp.:
120'C
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Website :
www.chromtech.net.auE-mail :
info@chromatech.net.auTelNo : 03 9762 2034 . . . in AUSTRALIA