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©2014 Waters Corporation
INTRODUCTION
Pesticides are widely used in the production of fruit and
vegetables across the globe. Governments, food
producers and food retailers have a duty to ensure they
are not present in final products for consumption. Most
countries have regulations governing pesticide residues in
food. For pesticides in food products, legislation imposes
Maximum Residue Limits (MRLs) which lead to the
requirement for analytical techniques that are sensitive,
selective and reproducible. Multi-residue pesticide analysis
is challenging due to the low limits of detection required
in a diverse range of food commodities. As there are
currently in excess of 1000 pesticides in use, laboratories
are under increasing pressure to broaden the range of
pesticides determined in ever shorter turnaround times.
Therefore, the analytical methods they employ need to
use efficient but low selectivity sample preparation
methods combined with high selectivity and sensitivity
MS/MS methods. Typically, this analysis is carried out
using a dedicated GC-MS/MS system with an EI source.
As shown by Portoles et al1, EI causes extensive
fragmentation of some pesticides leading to poor
sensitivity and selectivity. APGC is a soft ionization
technique which generates high relative and absolute
abundance molecular ions resulting in highly sensitive and
selective MRM transitions. Furthermore, the APGC source
is interchangeable with the LC electrospray source
enabling a single MS instrument to be used for the
analysis of both LC and GC amenable pesticides.
In this study, we demonstrate sensitive, accurate and
repeatable results for the analysis of pesticides in
QuEChERS extracts of strawberry, pear and spinach below
the regulatory limits.
Quantitative Analysis of Pesticides in QuEChERs Extracts Using APGC/MS/MS Douglas Stevens1, Dominic Roberts2 , Ramesh Rao2 1Waters Corporation, 34 Maple Street, Milford, MA 01757, USA. 2Waters Corporation, Altrincham Road, Wilmslow, UK.
METHODS
Strawberry, pear and spinach samples were
homogenized using a domestic food blender. The
samples were then extracted using the QuEChERS
(CEN method 15662 DisQuE #186004831) protocol to
generate blank matrix extract in acetonitrile. A nine
point calibration range from 0 to 50 ng/mL
(equivalent to µg/kg) was prepared by addition of a
mixed pesticide standard in acetonitrile to each
matrix. To test the repeatability at low concentration,
each matrix was fortified with the pesticide mix at 1
µg/kg (1ppb final concentration in aliquot). A
deuterated internal standard, chrysene-d12, was
added to give a fixed concentration of 2 ng/mL to
each vial prior to analysis and was used as an
injection standard to correct for injection volume
variation. All standards were analyzed in triplicate and
the low level spike in each matrix was analyzed ten
times using the Waters® Xevo TQ-S with the APGC
source using the conditions described below.
GC Conditions
MS Conditions
References
1. Portoles, Tania, Laura Cherta, Joaquim Beltran, and Felix Hernandez. "Improved gas chromatography–tandem mass spectrometry determination of pesticide residues making use of
atmospheric pressure chemical ionization." Journal of Chromatography A 1260 (2012): 183-192
2. Young, Michael, Tran, Kim Van, Shia, Jeremy C. “Multi-Residue
Pesticide Analysis in Ginseng Powder”. Waters application note
#720005006EN (2014)
3. Giroud, Barbara, Antoine Vauchez, Emmanuelle Vulliet, Laure Wiest, and Audrey Bulete. "Trace level determination of pyrethroid and neonicotinoid insecticides in beebread using
acetonitrile-based extraction followed by analysis with ultra-high-performance liquid chromatography–tandem mass spectrometry."
Journal of Chromatography A 1316 (2013): 53-61
RESULTS & DISCUSSION
Analysis of 20 GC amenable pesticides, difficult to analyze
in EI due to excessive fragmentation, was performed
using positive ion MRM mode. By varying source
conditions either charge exchange or protonation can be
selected for an APGC analysis. For the analysis of
pesticides, protonation provides more efficient ionization
than charge exchange. Therefore, a vial of water was
added to the source to promote protonation. The MRM
transitions with optimized cone voltages and collision
energies are shown in Table 1. Two transitions were
monitored for each pesticide to increase method
specificity. The high intensity of the precursor/molecular
ion generated by APGC makes it possible to use specific
and sensitive MRM transitions. In contrast, many pesticide
MRM transitions used with EI MS/MS use lower m/z, less
specific fragment ion as the precursor. The inherent
specificity provided by use of the molecular ion as the
precursor in an MRM transition over the use of a fragment
ion results in more confident detection of lower levels of
analytes even in these complex matrices prepared with a
simplified, generic sample preparation technique.
Table 1. Summary of the 20 pesticides analyzed, MRM
conditions and method performance results
Since the sensitivity of this system is well beyond
regulatory requirements, a practical application of this
performance is to dilute samples, thereby, further
reducing matrix effects on chromatography and
minimizing the amount of material injected on
column. This in turn reduces the frequency of column
trimming, extends the useful life of the column and
increases the interval between source cleanings which
is already measured in months. The net effect of all of
these factors is increased up-time and utilization for
the system.
CONCLUSION APGC on Xevo TQ-S is sensitive, accurate and
reproducible for pesticides that are difficult to analyze using conventional EI GC/MS/MS
Soft ionization provided by this technique produces
abundant molecular ions for selective and sensitive MRM transitions
Routine and sensitive multi-residue pesticide
analysis of QuEChERS extracts from fruit and vegetables, using the same workflow used for LC/MS/MS analysis of pesticides, is possible with this system
System can covert between GC and LC operation in
minutes allowing comprehensive analysis of both GC and LC amenable pesticides on a single instrument
Figure 2. Typical matrix matched calibration curve for
endosulfan sulphate in strawberry matrix
Figure 3. Mean calculated concentration of pesticides spiked
at 1 μg/kg in 3 different food matrices (n=10)
Table 2. Mean concentration of each pesticide (n=10) in the
three sample matrices
Each sample type, including matrix matched stan-
dards and replicates, was analyzed on three different
days. Figure 2 shows a typical calibration curve and
residuals plot for endosulfan sulphate generated from
the triplicate injection of the matrix matched calibra-
tion standard in strawberry extract. The response is
linear from 0.05 to 50 ng/mL with a correlation coeffi-
cient R2 of 0.994. All of the residuals are less than
15% demonstrating excellent linearity and repeatabil-
ity. The limits of detection and linearity achieved for
all 20 pesticides are summarized in Table 1. The limits
of detection ranged from 0.01 to 0.5 ng/mL with ex-
cellent linearity (R2 >0.99) for all. This demonstrates
that the method can easily achieve the regulatory
limits and is applicable to routine quantitative analy-
sis.
To assess the accuracy and precision of the method
each sample matrix was spiked at 1 µg/kg (10 times
below the blanket MRL of 10 µg/kg) and ten replicate
injections made. The concentration of each pesticide
was calculated using matrix matched calibration
curves. Table 2 shows the mean calculated concentra-
tions for each pesticide in all three samples matrices.
The accuracy of the method is excellent with all
measured concentrations within 5% of the true con-
centration.
The %RSD for all pesticides is also shown to be very
good at < 5%. This demonstrates that the method is
precise, accurate and reproducible across different
sample matrices analyzed on different days. The sen-
sitivity and overall performance characteristics of
APGC on Xevo TQ-S currently exceeds existing regu-
lations related to pesticide residue analysis.
Figure 1. Photo of UPLC and APGC on Xevo TQ-S
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