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Flow Injection Mass Spectrometry for High-Throughput Pesticide Residues Measurements Sergio C. Nanita, Anne M. Pentz, James J. Stry, Joseph P. McClory DuPont Crop Protection Stine-Haskell Research Center Newark, Delaware, U.S.A. Lisa Swaim, Bob Plastridge ABC Laboratories Columbia, Missouri, U.S.A. 1. Proof-of-Concept 2. Residue Analysis in/on Fabrics 3. FED-FI-MS Multiresidue Method FPRW, July 21, 2010
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MS analysis without chromatography • Improvements in MS allow direct analysis without HPLC:
DESI Purdue University (R.G. Cooks)
DART
ASAP
JEOL (R. Cody)
Univ. of the Sciences (C.N. McEwen)
• Suitable for analysis of surfaces or solids • Quantitation has been demonstrated • Sample position, angle and surface topography can affect instrument response
Analytical quality
HPLC/MS Flow Injection /MS
In-situ analysis possible! Ambient Ionization/MS
Throughput Potential FPRW, July 21, 2010
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Why change the design of analytical methods? Increased analysis speed and reduction in analytical expenses can be obtained by improving key steps in residue methods Evolutionary
Revolutionary
Extraction Cleanup Instrumentation Improvements
Changes
Increased sample throughput!
QuEChERS
Solvent A
Solvent B
Flow injection mass spectrometry HPLC
Pumps & Mixer
Autosampler & Injector
X
Column
MS
Flow Injection Data Acquisition
System Flush
Autosampler Action
Next Injection
15 s 10 s 25 s
FPRW, July 21, 2010
Total run time ~ 50 seconds
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I. Proof-of-Concept Study
FPRW, July 21, 2010
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Flow Injection Mass Spectrometry Setup Autosampler
Mass Spectrometer
• Bypass the column compartment * • Use 1- meter long red PEEK capillary * 1/16 in. O.D. and 0.13 mm I.D. (part no. 0890-1915, Agilent Technologies) • 1- uL injection * * Nanita, Pentz, Bramble. Anal. Chem. 2009, 81, 3134
FPRW, July 21, 2010
Proof-of-concept study • Typical residue method • SPE Cleanup • Flow Injection used instead of HPLC • Matrix-matched standards • External calibration Corn samples C o n tr o l
m/z 493 m/z 264
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Six compounds tested • Crops Avg Recovery 99–113 • Water Avg Recovery 92–109 • Linear corr. (R) > 0.998 • RSD < 19% • 15-second acquisition time (b )
(a )
0 .0 1 m g /k g (L O Q )
(c ) 0 .1 0 (10
m g /k g x L O Q )
m/z 432 m/z 182 m/z 466 m/z 182 m/z 365 m/z 150 m/z 236 m/z 71 m/z 163 m/z 88
15 seconds FPRW, July 21, 2010
Nanita, Pentz, Bramble. Anal. Chem. 2009, 81, 3134
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II. Application of Fl/MS/MS for Residue Analysis in/on Fabrics
FPRW, July 21, 2010
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Flow Injection Mass Spectrometry Method for Turf Transferable Residue Study - Experimental Fungicide 1. Cotton cloth extraction with methanol (shaking) 2. Direct FI/MS/MS analysis of extract with matrix-matched external standards
DuPont
ABC Laboratories Acq time = 15 s Cloth: 69cm x 99cm LOQ 1.0 ug/cloth 0.15 ng/cm2
R = 0.9992 RSD = 8%
Acq time = 60 s Cloth size: 69cm x 99cm LOQ 1.0 ug/cloth 0.15 ng/cm2
R = 0.9997 RSD = 5%
FPRW, July 21, 2010
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III. Fast Extraction & Dilution Flow Injection Mass Spectrometry
FED-FI-MS
FPRW, July 21, 2010
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• Can fast sample prep methods be used with flow injection MS? - Yes, with required modifications
• Can a multiresidue method with high-throughput sample preparation and analysis be engineered with flow injection MS? - Yes, with simpler procedures
QuEChERS • Simplest sample
J.P. McClory’s Method (DuPont) • Fast extraction, X 2
prep method • Dehydration & partition w/salts
• Final volume measured
• Dispersive SPE
• HPLC/MS/MS
High-Throughput Flow Injection MS Method (DuPont)
• Not compatible with FI-MS • Increased MS sensitivity allows significant extract dilution, direct flow injection, and less sample preparation FPRW, July 21, 2010
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QuEChERS • Simplest sample
J.P. McClory’s Method (DuPont) • Fast extraction, X 2
prep method • Dehydration & partition w/salts
• Final volume measured
• Dispersive SPE
• HPLC/MS/MS
High-Throughput Flow Injection MS Method (DuPont)
Fast Extraction & Dilution Flow Injection Mass Spectrometry
FED-FI-MS FPRW, July 21, 2010
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Fast Extraction & Dilution Flow Injection Mass Spectrometry
FED-FI-MS Method for samples with LOW water content
Method for samples with HIGH water content
10-g pecan sample extracted with 10 mL of organic solvent, 2 min in GenoGrinder with 3 metal beads • Fat freeze-out (1h) • Centrifuge 10 min • Dilute 1/50 • Flow injection MS LOQ = 0.05 mg/kg LOD ~ 0.02 mg/kg
Diluted 1/50
5-g lemon sample extracted with 25 mL of organic solvent, 2 min in GenoGrinder with 3 metal beads • Centrifuge 10 min • Dilute 1/10 • Flow injection MS
Diluted 1/10
LOQ = 0.05 mg/kg LOD ~ 0.02 mg/kg
Extraction solvent = Methanol Dilution solvent = 98.5% Methanol, 1.5% conc. ammonia FPRW, July 21, 2010
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Fast Extraction & Dilution Flow Injection Mass Spectrometry
FED-FI-MS Prototype Multiresidue Method 12 compounds, 4 pesticide classes
Classic Sulfonylurea Herbicides
Triflusulfuron methyl, m/z 493 264 & 238 Azimsulfuron, m/z 425 182 & 156 Chlorimuron ethyl, m/z 415 186 & 121 Sulfometuron methyl, m/z 365 150 & 199 Chlorsulfuron, m/z 358 141 & 167 Flupyrsulfuron methyl, m/z 466 182 & 156 Carbamate Insecticides Methomyl, m/z 163 88 & 106 Oxamyl, m/z 237 72 & 90
Novel
Pyrimidine Carboxylic Acid Herbicides Aminocyclopyrachlor, m/z 214 68 & 101 Aminocyclopyrachlor, m/z 228 68 & 41 methyl Anthranilic Diamide Insecticides Cyantraniliprole, m/z 475 286 & 444 Chlorantraniliprole, m/z 365 285 & 453
FPRW, July 21, 2010
Validation Raw Data Example, Pecan, Azimsulfuron • Quantitative ion transition shown: m/z 425 m/z 182 • Confirmatory ion transition monitored: m/z 425 m/z 156
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10 uL solvent wash injection
1 uL solvent blank
10 uL solvent wash injection
10 uL solvent wash injection
Pecan control A
Pecan control B
3x ≈ 1800
3x ≈ 1800
Lowest STD 0.7 ng/mL
LOQ-level STD 1.0 ng/mL
0.05 mg/kg fortification B
0.05 mg/kg fortification C
Detection threshold must be defined for each analyte
0.05 mg/kg fortification A
0.05 mg/kg fortification D
FPRW, July 21, 2010
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Validation Raw Data Example, Pecan, Azimsulfuron
STD 5.0 ng/mL
STD 10 ng/mL
0.50 mg/kg fortification A
0.50 mg/kg fortification B
0.50 mg/kg fortification C
0.50 mg/kg fortification D
STD 50 ng/mL
STD 100 ng/mL
10 uL solvent wash injection
10 uL solvent wash injection
• Quantitative ion transition shown: m/z 425 m/z 182 • Confirmatory ion transition monitored: m/z 425 m/z 156
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10 uL solvent wash injection
1 uL solvent blank
FPRW, July 21, 2010
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FED-FI-MS Matrix-Matched Standard Calibration LOW Water Content (Pecan)
Representative calibration curve, azimsulfuron R = 0.9993 y = 1.17x104 + 798
Analyte Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Sulfometuron methyl Chlorsulfuron Flupyrsulfuron methyl
Correlation, R 0.9989 0.9993 0.9994 0.9996 0.9990 0.9988
y y y y y y
Linear Equation 4 = 3.62x10 (x) + 376 = 1.17x104 (x) + 798 4 = 1.08x10 (x) - 1130 = 2.47x104 (x) - 961 3 = 3.84x10 (x) + 447 = 3.37x104 (x) + 761
Analyte Methomyl Oxamyl Aminocyclopyrachlor Aminocyclopyrachlor methyl Cyantraniliprole Chlorantraniliprole
Correlation, R 0.9998 0.9991 0.9996 0.9997 0.9992 0.9997
y y y y y y
Linear Equation 3 = 6.43x10 (x) - 529 = 3.75x103 (x) -179 3 = 9.71x10 (x) + 793 = 3.38x104 (x) + 1440 3 = 3.11x10 (x) - 165 = 6.76x103 (x) - 230
FPRW, July 21, 2010
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FED-FI-MS Matrix-Matched Standard Calibration HIGH Water Content (Lemon)
Representative calibration curve, azimsulfuron R = 0.9999 y = 1.32x104 + 2240
Analyte Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Sulfometuron methyl Chlorsulfuron Flupyrsulfuron methyl
Correlation, R 0.9997 0.9999 1.0000 0.9999 0.9999 1.0000
y y y y y y
Linear Equation 4 = 2.93x10 (x) + 2470 = 1.32x104 (x) + 2240 4 = 1.07x10 (x) + 321 = 2.07x104 (x) + 1510 3 = 3.45x10 (x) + 1940 = 2.46x104 (x) + 3260
Analyte Methomyl Oxamyl Aminocyclopyrachlor Aminocyclopyrachlor methyl Cyantraniliprole Chlorantraniliprole
Correlation, R 0.9993 0.9997 0.9999 0.9999 0.9998 0.9999
y y y y y y
Linear Equation 3 = 2.86x10 (x) + 1240 = 1.61x103 (x) + 1830 3 = 7.91x10 (x) + 2680 = 2.04x104 (x) + 5070 3 = 3.32x10 (x) + 557 = 6.20x103 (x) + 488
FPRW, July 21, 2010
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FED-FI-MS Method Validation for LOW Water Content (Pecan) Sulfonylureas Errorbars bars==11 in ineach eachdirection direction Error N = 8 fortified samples
120
Pyrimidine carboxylic Carbamates acids
Anthranilic diamides
Recovery (%)
100
80 70
60
40
20
0
FPRW, July 21, 2010
Trif
lus ul
Ox Me Am Flu Az i Ch Su Ch Am am lorim lfom lors t ho p yr ms i no ino yl my s ulf ulfu c yc c yc ulfu e tu u ro furo l lop lo p uro ro n ro n r on ne nm y y n r ra th m ac h me eth chl eth yl th lo yl o yl yl rm r eth yl
Cy
an t ra
Ch lor a n tra nil ip ro le ro le
nil ip
Overall Recovery = 101%
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FED-FI-MS Method Validation for HIGH Water Content (Lemon) Sulfonylureas Errorbars bars == 11 in in each each direction direction Error N = 8 fortified samples
120
Pyrimidine carboxylic Carbamates acids
Anthranilic diamides
Recovery (%)
100
80 70
60
40
20
0
T rif l us
Flu Ch Su Ch Az Am Am Me Ox im s lfom lori lor s tho am py rs in o ino mu u lfu yl ulf u my c yc cy c ulf u e tu ul fu ro n l ro n ro n lop lop r on r on r on eth yr a yr a me m m c c e y eth thy l hl o hl o thy l yl r rm l eth yl
FPRW, July 21, 2010
FED-FI-MS Proficiency/ Blind Test for LOW Water Content (Pecan) Chemist #1 (auditor) Fortified samples Chemist #2 (analyst) Extracted samples & reported results
n.a. = Analyte not added LOQ = 0.05 mg/kg LOD ~ 0.02 mg/kg FPRW, July 21, 2010
Cy
ant ra
Ch lora n tr a nil ro le ip ro le
nil ip
Overall Recovery = 91%
Sample Name Pecan #1 Pecan #1 Pecan #1 Pecan #1 Pecan #1 Pecan #1 Pecan #1 Pecan #1 Pecan #1 Pecan #1 Pecan #1 Pecan #1 Pecan #2 Pecan #2 Pecan #2 Pecan #2 Pecan #2 Pecan #2 Pecan #2 Pecan #2 Pecan #2 Pecan #2 Pecan #2 Pecan #2 Pecan #3 Pecan #3 Pecan #3 Pecan #3 Pecan #3 Pecan #3 Pecan #3 Pecan #3 Pecan #3 Pecan #3 Pecan #3 Pecan #3
Analyte Peak N ame Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole
Calculated Analyte Analyte Concentration Added Reported (ng/mL) (mg/kg) (mg/kg) 17.7 N o Peak N o Peak 9.43 N o Peak 1.15 N o Peak N o Peak N o Peak N o Peak 21.8 1.06 N o Peak N o Peak 21.6 N o Peak 1.2 N o Peak N o Peak N o Peak 12.1 12.2 N o Peak N o Peak 18.3 8.89 11.7 N o Peak N o Peak 11.9 N o Peak N o Peak N o Peak 1.51 1.71 2.08
0.900 n.a n.a 0.500 n.a 0.075 n.a n.a n.a n.a 1.100 0.050 n.a n.a 1.300 n.a 0.075 n.a n.a n.a 0.600 0.600 n.a n.a 0.900 0.600 0.600 n.a n.a 0.700 n.a n.a n.a 0.070 0.070 0.095
0.89 < LOD < LOD 0.47 < LOD 0.058 < LOD < LOD < LOD < LOD 1.09 0.053 < LOD < LOD 1.08 < LOD 0.060 < LOD < LOD < LOD 0.61 0.61 < LOD < LOD 0.92 0.45 0.59 < LOD < LOD 0.60 < LOD < LOD < LOD 0.076 0.086 0.10
Accuracy: Passed, Failed, 22 or Percent % 99 Passed Passed 94 Passed 77 Passed Passed Passed Passed 99 106 Passed Passed 83 Passed 80 Passed Passed Passed 102 102 Passed Passed 102 75 98 Passed Passed 86 Passed Passed Passed 109 123 105
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FED-FI-MS Proficiency/ Blind Test for LOW Water Content (Pecan) Chemist #1 (auditor) Fortified samples Chemist #2 (analyst) Extracted samples & reported results
n.a. = Analyte not added LOQ = 0.05 mg/kg LOD ~ 0.02 mg/kg FPRW, July 21, 2010
FED-FI-MS Proficiency/ Blind Test for LOW Water Content (Pecan) Chemist #1 (auditor) Fortified samples Chemist #2 (analyst) Extracted samples & reported results
n.a. = Analyte not added LOQ = 0.05 mg/kg LOD ~ 0.02 mg/kg FPRW, July 21, 2010
Sample Name Pecan #4 Pecan #4 Pecan #4 Pecan #4 Pecan #4 Pecan #4 Pecan #4 Pecan #4 Pecan #4 Pecan #4 Pecan #4 Pecan #4 Pecan #5 Pecan #5 Pecan #5 Pecan #5 Pecan #5 Pecan #5 Pecan #5 Pecan #5 Pecan #5 Pecan #5 Pecan #5 Pecan #5 Pecan #6 Pecan #6 Pecan #6 Pecan #6 Pecan #6 Pecan #6 Pecan #6 Pecan #6 Pecan #6 Pecan #6 Pecan #6 Pecan #6
Sample Name Pecan #7 Pecan #7 Pecan #7 Pecan #7 Pecan #7 Pecan #7 Pecan #7 Pecan #7 Pecan #7 Pecan #7 Pecan #7 Pecan #7 Pecan #8 Pecan #8 Pecan #8 Pecan #8 Pecan #8 Pecan #8 Pecan #8 Pecan #8 Pecan #8 Pecan #8 Pecan #8 Pecan #8 Pecan #9 Pecan #9 Pecan #9 Pecan #9 Pecan #9 Pecan #9 Pecan #9 Pecan #9 Pecan #9 Pecan #9 Pecan #9 Pecan #9
Analyte Peak Name Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole
Analyte Peak Name Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole
Calculated Analyte Analyte Concentration Added Reported (ng/mL) (mg/kg) (mg/kg) No Peak 9.64 No Peak No Peak No Peak 15.4 No Peak No Peak No Peak No Peak No Peak 38.9 No Peak 1.1 No Peak No Peak 1.41 19.9 No Peak No Peak No Peak No Peak 7.54 No Peak No Peak No Peak No Peak 0.871 26.5 No Peak 0.925 No Peak 11.5 No Peak No Peak 22.5
n.a 0.600 n.a n.a n.a 0.875 n.a n.a n.a n.a n.a 1.900 n.a 0.080 n.a n.a 0.075 1.200 n.a n.a n.a n.a 0.300 n.a n.a n.a n.a 0.065 1.500 n.a 0.060 n.a 0.600 n.a n.a 1.100
< LOD 0.48 < LOD < LOD < LOD 0.77 < LOD < LOD < LOD < LOD < LOD 1.95 < LOD 0.055 < LOD < LOD 0.071 1.00 < LOD < LOD < LOD < LOD 0.38 < LOD < LOD < LOD < LOD 0.044 1.33 < LOD 0.046 < LOD 0.58 < LOD < LOD 1.13
Calculated Analyte Analyte Concentration Added Reported (ng/mL) (mg/kg) (mg/kg) No Peak 1.19 No Peak No Peak No Peak No Peak 0.903 39.5 No Peak No Peak 1.32 No Peak 1.48 No Peak No Peak 1.05 No Peak No Peak No Peak 30.6 1.56 1.19 No Peak No Peak 1.24 No Peak 0.987 No Peak 27.4 No Peak 9.43 1.68 No Peak 25.5 No Peak No Peak
n.a 0.080 n.a n.a n.a n.a 0.060 2.000 n.a n.a 0.070 n.a 0.080 n.a n.a 0.065 n.a n.a n.a 1.600 0.080 0.070 n.a n.a 0.080 n.a 0.065 n.a 1.500 n.a 0.600 0.080 n.a 1.400 n.a n.a
< LOD 0.060 < LOD < LOD < LOD < LOD 0.045 1.98 < LOD < LOD 0.066 < LOD 0.074 < LOD < LOD 0.053 < LOD < LOD < LOD 1.53 0.078 0.060 < LOD < LOD 0.062 < LOD 0.049 < LOD 1.37 < LOD 0.47 0.084 < LOD 1.28 < LOD < LOD
Accuracy: Passed, Failed, 23 or Percent % Passed 80 Passed Passed Passed 88 Passed Passed Passed Passed Passed 103 Passed 69 Passed Passed 95 83 Passed Passed Passed Passed 127 Passed Passed Passed Passed 68 89 Passed 77 Passed 97 Passed Passed 103
Accuracy: Passed, Failed, or 24 Percent % Passed 75 Passed Passed Passed Passed 75 99 Passed Passed 94 Passed 93 Passed Passed 82 Passed Passed Passed 96 98 86 Passed Passed 78 Passed 75 Passed 91 Passed 78 105 Passed 91 Passed Passed
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FED-FI-MS Proficiency/ Blind Test for LOW Water Content (Pecan) Chemist #1 (auditor) Fortified samples Chemist #2 (analyst) Extracted samples & reported results
n.a. = Analyte not added LOQ = 0.05 mg/kg LOD ~ 0.02 mg/kg
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Sample Name Pecan Pecan Pecan Pecan Pecan Pecan Pecan Pecan Pecan Pecan Pecan Pecan
#10 #10 #10 #10 #10 #10 #10 #10 #10 #10 #10 #10
Analyte Peak Name
Analyte Analyte Calculated Concentration Added Reported (mg/kg) (mg/kg) (ng/mL)
Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole
24.3 No Peak 0.947 26.9 No Peak No Peak 8.76 1.48 1.74 No Peak No Peak No Peak
1.200 n.a 0.065 1.500 n.a n.a 0.600 0.080 0.080 n.a n.a n.a
1.22 < LOD 0.047 1.35 < LOD < LOD 0.44 0.074 0.087 < LOD < LOD < LOD
Accuracy: Passed, Failed, or Percent % 102 Passed 72 90 Passed Passed 73 93 109 Passed Passed Passed
Summary Results for 10 Pecan Samples • FED-FI-MS passed for all 12 analytes • Zero false positive • Zero false negative • 120 accurate results (10 samples x 12 analytes) • Average accuracy for all compounds detected = 91%
FPRW, July 21, 2010
FED-FI-MS Proficiency/ Blind Test for HIGH Water Content (Lemon) Chemist #1 (auditor) Fortified samples Chemist #2 (analyst) Extracted samples & reported results
n.a. = Analyte not added LOQ = 0.05 mg/kg LOD ~ 0.02 mg/kg FPRW, July 21, 2010
Sample Name Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon
#1 #1 #1 #1 #1 #1 #1 #1 #1 #1 #1 #1 #2 #2 #2 #2 #2 #2 #2 #2 #2 #2 #2 #2 #3 #3 #3 #3 #3 #3 #3 #3 #3 #3 #3 #3
Analyte Peak Name Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole
Analyte Analyte Calculated Concentration Added Reported (mg/kg) (mg/kg) (ng/mL) No Peak 9.44 0.543 42.9 No Peak 0.421 No Peak 1.34 No Peak No Peak No Peak 26.4 50.8 0.681 No Peak No Peak No Peak 17.4 0.86 No Peak 16.2 No Peak No Peak No Peak No Peak 1.25 No Peak 14.9 No Peak 28.6 No Peak No Peak No Peak 12.5 2.04 29.5
n.a. 0.600 0.040 2.600 n.a. 0.030 n.a. 0.080 n.a. n.a. n.a. 1.600 3.000 0.040 n.a. n.a. n.a. 1.000 0.050 n.a. 1.000 n.a. n.a. n.a. n.a. 0.070 n.a. 0.800 n.a. 1.600 n.a. n.a. n.a. 0.800 0.110 1.600
< LOD 0.47 0.027 2.15 < LOD 0.021 < LOD 0.067 < LOD < LOD < LOD 1.32 2.54 0.034 < LOD < LOD < LOD 0.87 0.043 < LOD 0.81 < LOD < LOD < LOD < LOD 0.063 < LOD 0.75 < LOD 1.43 < LOD < LOD < LOD 0.63 0.10 1.48
Accuracy: Passed, Failed,26 or Percent % Passed 78 68 83 Passed 70 Passed 84 Passed Passed Passed 83 85 85 Passed Passed Passed 87 86 Passed 81 Passed Passed Passed Passed 90 Passed 94 Passed 89 Passed Passed Passed 79 91 93
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FED-FI-MS Proficiency/ Blind Test for HIGH Water Content (Lemon) Chemist #1 (auditor) Fortified samples Chemist #2 (analyst) Extracted samples & reported results
n.a. = Analyte not added LOQ = 0.05 mg/kg LOD ~ 0.02 mg/kg FPRW, July 21, 2010
FED-FI-MS Proficiency/ Blind Test for HIGH Water Content (Lemon) Chemist #1 (auditor) Fortified samples Chemist #2 (analyst) Extracted samples & reported results
n.a. = Analyte not added LOQ = 0.05 mg/kg LOD ~ 0.02 mg/kg FPRW, July 21, 2010
Sample Name Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon
#4 #4 #4 #4 #4 #4 #4 #4 #4 #4 #4 #4 #5 #5 #5 #5 #5 #5 #5 #5 #5 #5 #5 #5 #6 #6 #6 #6 #6 #6 #6 #6 #6 #6 #6 #6
Sample Name Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon
#7 #7 #7 #7 #7 #7 #7 #7 #7 #7 #7 #7 #8 #8 #8 #8 #8 #8 #8 #8 #8 #8 #8 #8 #9 #9 #9 #9 #9 #9 #9 #9 #9 #9 #9 #9
Analyte Peak Name Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole
Analyte Peak Name Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole
Calculated Analyte Analyte Concentration Added Reported (ng/mL) (mg/kg) (mg/kg) 0.911 No Peak 1.08 No Peak 25.3 No Peak 1.92 37.5 1.39 No Peak No Peak No Peak 24.6 0.539 No Peak No Peak No Peak No Peak 18.5 No Peak 16.5 0.869 No Peak No Peak No Peak No Peak 61 No Peak No Peak No Peak 40.8 No Peak No Peak 1.76 1.98 No Peak
0.050 n.a. 0.060 n.a. 1.400 n.a. 0.100 2.000 0.120 n.a. n.a. n.a. 1.400 0.030 n.a. n.a. n.a. n.a. 1.200 n.a. 1.000 0.070 n.a. n.a. n.a. n.a. 3.600 n.a. n.a. n.a. 2.600 n.a. n.a. 0.110 0.110 n.a.
0.046 < LOD 0.054 < LOD 1.27 < LOD 0.096 1.88 0.070 < LOD < LOD < LOD 1.23 0.027 < LOD < LOD < LOD < LOD 0.93 < LOD 0.83 0.043 < LOD < LOD < LOD < LOD 3.05 < LOD < LOD < LOD 2.04 < LOD < LOD 0.088 0.099 < LOD
Calculated Analyte Analyte C oncentration Added Reported (ng/mL) (mg/kg) (mg/kg) No Peak 41 No Peak No Peak No Peak No Peak 2.11 No Peak 1.8 No Peak 33.6 No Peak No Peak No Peak 1.79 15.6 31.8 No Peak No Peak No Peak No Peak 2.12 No Peak 1.23 32.6 No Peak 37.4 No Peak 0.874 0.368 No Peak 21.8 No Peak No Peak 15.3 No Peak
n.a. 2.400 n.a. n.a. n.a. n.a. 0.130 n.a. 0.120 n.a. 2.000 n.a. n.a. n.a. 0.100 0.800 1.800 n.a. n.a. n.a. n.a. 0.110 n.a. 0.080 1.800 n.a. 2.000 n.a. 0.050 0.030 n.a. 1.200 n.a. n.a. 0.800 n.a.
< LOD 2.05 < LOD < LOD < LOD < LOD 0.11 < LOD 0.090 < LOD 1.68 < LOD < LOD < LOD 0.090 0.78 1.59 < LOD < LOD < LOD < LOD 0.11 < LOD 0.062 1.63 < LOD 1.87 < LOD 0.044 0.018 < LOD 1.09 < LOD < LOD 0.770 < LOD
Accuracy: Passed, Failed,27 or Percent % 92 Passed 90 Passed 91 Passed 96 94 58 Passed Passed Passed 88 90 Passed Passed Passed Passed 78 Passed 83 61 Passed Passed Passed Passed 85 Passed Passed Passed 78 Passed Passed 80 90 Passed
Accuracy: Passed, Failed,28 or Percent % Passed 85 Passed Passed Passed Passed 85 Passed 75 Passed 84 Passed Passed Passed 90 98 88 Passed Passed Passed Passed 100 Passed 78 91 Passed 94 Passed 88 60 Passed 91 Passed Passed 96 Passed
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FED-FI-MS Proficiency/ Blind Test for HIGH Water Content (Lemon) Chemist #1 (auditor) Fortified samples Chemist #2 (analyst) Extracted samples & reported results
n.a. = Analyte not added LOQ = 0.05 mg/kg LOD ~ 0.02 mg/kg
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Sample Name Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon Lemon
#10 #10 #10 #10 #10 #10 #10 #10 #10 #10 #10 #10
Analyte Peak Name Triflusulfuron methyl Azimsulfuron Chlorimuron ethyl Flupyrsulfuron methyl Sulfometuron methyl Chlorsulfuron Aminocyclopyrachlor Aminocyclopyrachlor methyl Methomyl Oxamyl Cyantraniliprole Chlorantraniliprole
Analyte Analyte Calculated Concentration Added Reported (mg/kg) (mg/kg) (ng/mL) 1.67 59.1 No Peak 0.53 No Peak No Peak No Peak 1.29 No Peak 33.6 No Peak 1.4
0.100 3.400 n.a. 0.040 n.a. n.a. n.a. 0.080 n.a. 2.000 n.a. 0.080
0.084 2.96 < LOD 0.027 < LOD < LOD < LOD 0.065 < LOD 1.68 < LOD 0.070
Accuracy: Passed, Failed, or Percent % 84 87 Passed 68 Passed Passed Passed 81 Passed 84 Passed 88
Result Summary for 10 Lemon Samples • FED-FI-MS passed for all 12 analytes • Zero false positive • Zero false negative • 120 accurate results (10 samples x 12 analytes) • Average accuracy for all compounds detected = 84%
FPRW, July 21, 2010
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Advantages of Flow Injection Increased throughput
Less solvent
• 15 s data acquisition time. • >1000 injections/day possible.
• ~ 0.4 L/1000 sample methods injections. • Eliminates the need for columns, • No liquid waste generated. and other HPLC components.
Simpler instrumental
Requirements for High-Throughput Quantitation by Flow Injection • Matrix-matched standards • Highly-sensitive MS instrument (e.g. API-5000) with orthogonal spray/inlet ion source preferred
Spray
Inlet
• Number of analytes limited by triple quadrupole MS scan speed, up to 20 analytes (40 ions) tested • Highly selective fragmentation (avoid trivial neutral losses like H2O, NH3, CO2), and carefully rule out interferences by pesticide class • Multiple ion transitions must be monitored • Avoid non-volatile salts like those containing Na+, K+ (volatiles are ok, like NH4Ac) FPRW, July 21, 2010
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FED-FI-MS
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Sample Throughput Potential
Prototype method/ estimated
Potential/ to be proven
Fast Extraction & Dilution (FED), one chemist
up to 72 samples/day
≥ 96 samples/day
Flow Injection Mass up to 1,300 Spectrometry (FI-MS) injections/day
2,800 – 5,700 injections/day
Based on experiments with fortified controls; dry ice homogenization done previously by other personnel. Estimate includes solvent blank injections for rinsing/cleaning. Based on 2 – 4 injections per minute.
Throughput-limiting steps FED Sample weighing FI-MS Autosampler speed FPRW, July 21, 2010
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FED-FI-MS What’s next? Prototype FED-FI-MS Method Applied/Proven Multiresidue FED-FI-MS
Method Performance • Ruggedness testing • Expansion to other analytes • Expansion to other matrices • Inter-laboratory trials • Extraction efficiency studies • Incurred residue sample tests (vs. HPLC)
Quantitative Calibration • Internal standards • Standard addition method Contact information Sergio C. Nanita DuPont Crop Protection Phone: 1-302-451-5806 Email:
[email protected] FPRW, July 21, 2010
Instrumentation
• Accurate Mass Instruments - Time-of-Flight (TOF) - Orbitrap MS • Try faster autosamplers! • Improve LOQ & LOD
Lots of work! Get onboard! Collaborate!
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Acknowledgements Anne Pentz Fred Bramble Joe McClory Jim Stry John May Teri Gray Steven McEuen Lisa Swaim* Roger Winters* Emily Vogl* Bob Plastridge* Del Koch* Clark Chickering*
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Also, helpful discussions with: Steve Cheatham Bob Dietrich Dave Ryan
*ABC Labs
FPRW, July 21, 2010
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