GC/MS/MS Analysis of Thiabendazole in Grapefruit Extracts Varian

GC/MS/MS Analysis of
Thiabendazole in
Grapefruit Extracts
Varian Application Note
Number 30
Steve Schachterle, Varian Chromatography Systems
Uri Golner and Irina Tannenzapf, Ministry of Agriculture, Israel
Key Words: Saturn 4D, MS/MS, Pesticide, Wave~Board
Introduction
The analysis of grapefruit for thiabendazole (TBZ) is an
economically important analysis. The fungicide is applied
as a postharvest treatment in the citrus packing houses.
Generally the maximum regulatory levels are high (up to
10 ppm), but certain legislations consider TBZ a food
additive when it comes to processed fruit (e.g. juices) and
as such the residue level must not exceed 10 ppb. The
analysis necessary to assure this is indeed a challenging
one due to the complexity of the grapefruit background
matrix.
Matrix
TBZ
Discussion
Conventional GC/MS does not always provide
unambiguous results because the mass spectrum of a
coeluting matrix component may have ions which
coincide with those from TBZ at a concentration far above
the desired detection limit for TBZ. Figure 1 shows the
GC/MS chromatogram for TBZ in a grapefruit extract at
the 500 ppb level. The inset spectra are for the matrix
component and for TBZ. Note that the matrix component
has ions at m/z 201 and 174, the two main ions for TBZ.
Due to the partial chromatographic separation, it is not
possible to measure very low levels of TBZ in the
presence of this interfering component. Figure 2 shows
the GC/MS/MS chromatogram for TBZ in the grapefruit
extract at the 10 ppb level. The background from the
matrix peaks does not appear in the mass chromatogram
for m/z 174. Figure 3 compares the GC/MS total ion
chromatogram with the GC/MS/MS total ion
chromatogram. Note that the GC/MS/MS total ion
chromatogram still contains several peaks; although it
should be noted the level is 20 times lower than the
GC/MS total ion chromatogram. Any ion at m/z 201 that
dissociates under the excitation conditions will produce
ions in the MS/MS chromatogram.
Figure 1: Mass Chromatogram showing 500 ppb of TBZ in
Grapefruit by GC/MS.
Figure 2: 10 ppb of TBZ in Grapefruit by GC/MS/MS
NOTICE: Varian, Inc. was acquired by Agilent
Technologies in May 2010. This document is provided
as a courtesy but is no longer kept current and thus
will contain historical references to Varian. For more
information, go to www.agilent.com/chem.
This includes both molecular ions and fragment ions
produced in the EI ionization process. The real advantage
of MS/MS is that now the m/z 174 mass chromatogram
shows only those m/z 174 ions which were formed by CID
from m/z 201 and not those formed by electron ionization
as seen in the GC/MS chromatogram. Figure 4 shows an
external standard calibration curve prepared by spiking
the grapefruit extract. The linearity is excellent with a wide
dynamic range, even in the presence of the complex
matrix.
Instrument Conditions
Gas Chromatograph
Column:
DB5-ms, 30m x 0.25 mm ID x 1.0 µm film
thickness
Flow Rate:
0.8 ml/min measured at 200°C
Oven Program:
50°C for 1 minute. Program at 10°C/min to
275°C. Hold 13.5 minutes.
Injector:
SPI with high performance insert. 25°C no
hold. Program at 200°C/min to 230°C. Hold
until the end of the run.
Injection Volume:
1 µl
Injection Speed:
1µl/sec
Mass Spectrometer
Experimental
The grapefruit concentrate was prepared for analysis
through a modified Luke procedure (FDA-PAM 232.4) by
a robotic sample preparation system. The extract
contained 1 gram of soluble grapefruit in a final volume of
1 ml. After filtration, the acetone extract was evaporated
with helium and reconstituted with methylene chloride.
Mass Range
Scan/sec
Mult. delay
Target
Threshold
Filament
RF level
Ion Trap Temperature
Excitation voltage
Excitation time
MS
50-210
1
180 sec
25,000
1
20 µA
300 dacs
270°C
-
MS/MS
50-210
1
180 sec
5,000
1
80 µA
450 dacs
270°C
61 volts
40 ms
Conclusion
GC coupled with tandem mass spectrometry (MS/MS)
using an ion trap can be used to identify and quantitate
TBZ at low ppb levels, even in the presence of interfering
components. Compared with standard GC/MS, the
selectivity provided by the additional MS step effectively
eliminates the response from the interfering component
and lowers the detection limit dramatically.
Figure 3: Total ion chromatograms of 500 ppb spike by
GC/MS (top) and GC/MS/MS (bottom).
Figure 4: GC/MS/MS External Standard Calibration Curve
Prepared by Spiking the Grapefruit Extract.
These data represent typical results.
For further information, contact your local Varian Sales Office.
GC/MS30:0694