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Supporting Information
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Uptake and translocation of organophosphates and other emerging contaminants in
crops used for forage and food.
Eggen, T*., Heimstad, E., Stuanes, A.O., Norli, H.R.
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*Corresponding author, E-mail: Trine.Eggen@bioforsk.no
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Experimental Chemical analyses
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Tables and figures
Table S1.Compound retention time, time segments, quantitation and qualification ions.
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Table S2. Recoveries, relative standard deviation (RSD) and limit of quantification
(LOQ) for three different plant species, n=5 replicates
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Figure S1 Distribution of neutral and charged species of NBBS and DEET with pH
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Experimental
Chemical analysis
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Chemicals: Acetonitrile, pestiscan, was purchased from Lab-Scan (Poch SA, Gliwice,
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Poland). Sodium chloride (pa) and calcium chloride (pa) was purchased from Merck
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(Darmstadt, Germany). Anhydrous magnesium sulfate was purchased from J.T. Baker
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(Phillipsburg, NJ). Superclean PSA (primary-secondary amine), Superclean ENVI-Carb
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(graphitizised carbon black), and DSC-18 Sorbents, as well as sodium sulfate p.a., were
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purchased from Sigma-Aldrich (St. Louis, USA). Internal standard 2-brombiphenyl (purity
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98.5) was purchased from (Dr. Ehrenstorfer, Augsburg, Germany).
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Extraction of plant materials: The sample preparation followed the QuEChERS method [1,
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2]. Subsamples of 1-5 g dried plant materials was added 10 mL bi-distilled water, 10 mL
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acetonitrile and 1.0 µg of 2-brom-biphenyl as internal standard into 50 mL polypropylene
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centrifuge tubes. The samples were homogenized on Polytron (Kinematica AG) for 1 min.)
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Samples were added 4 g anhydrous MgSO4 and 1 g NaCl and the tubes were shaken 1 min
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and centrifuged for 5 min at 3000 rpm. After phase separation, all extracts were subjected to
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dispersive extraction technique by use of 900 mg anhydrous MgSO4 and 150 mg PSA. For
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clean up of seeds, 50 mg DSC-18 sorbent was added and for carrot, meadow and barley, 15
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mg of Envi-Carb was added. For seeds an extra cleanup step to remove high molecular
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compounds like fatty acids, their esters and to some extent plant sterols was performed by
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freezing the samples at -20oC for 2 hours before carefully decantation of the none frozen
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acetonitrile phase into a tube containing 1 g of CaCl2. The tubes were shaken 1 min and
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centrifuged at 3000 rpm in 5 min.
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Extraction of soil: 2.0-5.0 g subsample of dry soil and 1.0 g anhydrous MgSO4 was added
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10.0 mL of acetonitile and 1 µg of internal standard into 50 mL poly-propylene tubes and
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extracted in an “end over end “device for 30 min. The sample was centrifuged at 3000 rpm for
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5 min and analyzed without any further clean up.
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Instrumentation: The extracts were analyzed using an Agilent 6890 N gas chromatograph
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connected to an Agilent 5973 mass spectrometer with an inert ion source operated in selected
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ion monitoring mode. The GC was equipped with a Gerstel Programmable Temperature
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Vaporising Injector (Mühlheim Ruhr, Germany) with a sintered liner, injection volume 5
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µ
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mL/min . At 1.89 min the split valve was closed and the injector temperature was raised by
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270oC/min to 280 oC and held for 1.2 min. For information about retention time,
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quantification ions and recovery, please see Table S1 and S2.
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o
C in 1.8 min with a vent flow at 60.0
Separation was performed using a fused silica J&W Scientific HP-5MSI (0.25 mm i.d. x
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30 m) with 0.25 µm film thickness. The column was connected to a 2.5 m methyl de activated
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pre-column of same internal diameter as the analytical column (Varian inc. Lake Forest CA,
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USA). The temperature program was 90oC, held for 1 min, 20oC/min to 160oC, held for 0
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min, then 5oC/ min to 280oC, held for 5 min. A recovery test (with five replicates of barley
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leaf, carrot root and fescue leaf spiked with 0.01 µg/g were performed and a recovery in the
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ranged of 60-392 % was obtained. For DEET in fescue leaf and TBP in carrot- and fescue leaf
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recoveries higher that 120 % were found. High recoveries are related to interferences from
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matrix which co elute with the compounds and give an overestimation of the quantitative
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result. However, in the real samples the concentrations was higher than 0.01 µg/g and
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interferences therefore became insignificant. The calibration curves covered the range from
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0.001 to 1.0 µg/ml with regression coefficients from 0.996 to 0.999. Reagent blanks and
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positive control samples spiked at 0.2 µg/g of each test compound were included in each run.
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The recoveries show acceptable results between 70-120 % [2]. Except for DEET in fescue
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leaf and TBP in carrot- and fescue leaf, the limit of quantification could be set to 0.01µg/g
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with a signal to noise higher than 10. In fescue leaf the LOQ had to be increased to 0.05 µg/g.
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No LOQ for DEET in fescue leaf and TBP in carrot root was established due to interferences.
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Tables and figures
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Table S1. Summary of compounds retention time, time segments, quantitation and
qualification ions.
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Compound
ion)b
Rt (min)
Window/start time (min)
Quantitation (qualifying
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2-Brombiphenyl (IS)
10.9
I/10
232 (234)
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DEET
10.8
I
162 (176)
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TBP
11.7
I
211 (155,167,183)
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TCEP
13.6
II/13.1
249 (251,223)
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NBBS
13.9
II
213 (158,184)
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TCPP
14.1a
II
277(279,291) ____
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a
Retention time for the first eluting isomer
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b
A compound is regarded as identified when the area of the qualifying ion is within ± 30 %
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tolerance limit of the quantitation ion [2]
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Table S2. Recoveries, relative standard deviataion (RSD) and limit of quantification (LOQ) for three
different plant species, n=5 replicates.
Barley leaf
Recovery
(%)
DEET
118
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74
a
b
Carrot root
RSD
(%)
4
LOQa
Fescue leaf
RSD
(%)
17
LOQa
0.01
Recovery
(%)
98
RSD
(%)
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LOQb
0.01
Recovery
(%)
392
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TBP
104
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0.01
136
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_
126
7.1
0.05
TCEP
96
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0.01
96
12
0.01
60
11.8
0.05
NBBS
96
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0.01
88
25
0.01
82
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0.05
TCPP
115
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0.01
113
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0.01
110
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0.05
S/N>10 estimated from the spiked samples at 0.01 µg/g
S/N>10 estimated from the real samples
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Figure S1.
pH.
Distribution of neutral and charged species of DEET (A) and NBBS (B) with
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B
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Reference:
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1.
Quechers http://www.quechers.com.
2.
Pihlström, T. Method Validation and Quality Control Procedures for Pesticide Residues
Analysis in Food and Feed; 2007.
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