the evaluation of multi-pesticide screening ... - phenomenex
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The Evaluation of Multi-Pesticide Screening Methods by GC/MS
Sky Countryman, Jim Archer, Kory Kelly, and Doug SilvaPhenomenex, Inc., 411 Madrid Ave., Torrance, CA 90501 USA
The current work demonstrates the use of two new and unique phases, which have been optimized for the analysis of all classes of pesticides. The phase chemistry improves separation and peak shape for the more polar pesticide compounds when compared to standard 5 % phenyl columns. Selectivity data is compared between a 5ms type phase and the two new columns.
M u l t i - p e s t i c i d e r e s i d u e screening has been evaluated for over 250 different pesticides commonly analyzed from fruits and vegetables (not all data presented here). The unique selectivity offered by the two phases improves resolution
for multi-component analytes providing a more unique elution pattern, which can be used to identify closely eluting analytes.
S ince the phases have orthogonal selectivity, they are also a good choice for dual column methods. Some data is presented for EPA specified testing procedures.
Abstract Figure 1: Multi-Pesticide Residue Analysis using Zebron MultiResidue™-1
Figure 7: Dual Column Organochlorine Pesticide Testing Under EPA Method 8081A
Zebron and MultiResidue are trademarks of Phenomenex, Inc.© 2008 Phenomenex, Inc. All rights reserved.
As world trade increases, the potential threat to other countries’ populations due to contaminated food products increases. Recent deaths caused by food exported from countries like China, emphasize the need for comprehensive testing procedures. The Zebron MultiResidue-1 and the MultiResidue-2 present a comprehensive solution for Multi-Pesticide residue testing by GC/MS and other hyphenated techniques. The columns provide additional confirmation, potential separation from matrix interferences, greater resolution of isomer peaks, and improved chromatography of more polar analytes.
Introduction
Pesticides are widely used by farmers to control pests, weeds, and molds that would otherwise decrease crop production. While this has significantly increased worldwide food product ions, these same pesticides pose significant health and environmental risks. The restrictions for specific pesticides dif fer from one country to the next. As world trade increases, the potential threat to other countr ies’ populations increases. For this reason, pesticides are the subjects of increasing regulation.
Since many different types of pesticides can be used on the same food product, Multi-Residue screening approaches are used to look for multiple classes of pesticide compounds at one t ime. Consider ing that there are more than 500 registered pesticides, no single analysis technique is capable of screening for all possible contaminates. However, gas chromatography (GC) is still the most commonly used method for the majority of the pesticide classes. While analyte specific detectors such as ECD or NPD may be used for screening, Mass Spectrometer (MS) detection must be employed to provide positive confirmation. Zebron MultiResidue™ (MR) co lumns we re spec ia l l y des igned fo r pes t ic ides analysis. The columns were developed using two new
stationary phases that are unlike any other commercially available columns. The phases were designed to provide or thogona l se lect iv i t y to provide maximum resolving power in complex samples. Z e b r o n M u l t i R e s i d u e™ columns provide low bleed on ECD and NPD detectors and both columns are MS certified, so they can also be used with GC/MS for multi-residue pesticide methods.
GC/MS screening of multi-residue pesticide standards was evaluated using the new Zebron MultiResidue™ columns and compared with the results obtained using a standard 5ms type column. Dual column approaches were also evaluated using a chlorinated pesticides sample following EPA Method 8081A.
Results & Discussion
Analysis of 114 different pesticides was done using both Zebron MultiResidue-1 and MultiResidue-2 column (Figures 1 & 2). The Zebron MultiResidue-2 columns showed additional peaks for certain pesticides compared to the Zebron MultiResidue-1 column, however both columns provide increased resolution compared to a standard 5ms type phase.
Resolution of specific compounds was compared with the Zebron ZB-5ms phase to demonstrate the increased separation power offered by these new columns. In Figure 3, the elution order for atrazine and propazine is reversed on both the MultiResidue-1 and the MultiResidue-2 columns compared to the ZB-5ms. The MultiResidue-2 demonstrates much longer relative retention for these two compounds and is able to provide baseline resolution. In Figure 4, the MultiResidue-1 provides increased resolution of trifluralin and benefin as well as several of the later eluting compounds. The polarity of the MultiResidue-2 caused the trifluralin to partially co-elute with propachlor, however resolution is substantially increased for the last four compounds.
Asymmetrical peak shape such as fronting is commonly observed for polar pesticides on non-polar phases because the pesticides are not soluble in the phase. This can lead to poor sensitivity and/or co-elutions with other closely eluting pesticides. Figure 5 shows overloading of terbacil and bromacil on the ZB-5ms phase due to the non-polar character of the phase. The phase chemistry of the MultiResidue-1 and MultiResidue-2 was specially designed to match the polarity of pesticide compounds and provide better overall peak shape.
Many pesticides contain a mixture of isomers. Resolving the isomers can provide additional confirmation for samples with interferences in the
chromatographic region of the target analyte. In Figure 6, phosphamidon shows an isomer eluting just after terbufos on the ZB-5ms. If high levels of terbufos were present, it is unlikely resolution between the two compounds would be maintained. The MultiResidue-1 and the MultiResidue-2 columns significantly increase resolution of the phosphamidon isomer ensuring accurate identification.
The US EPA regulates the testing of 20 specific chlorinated pesticides under the official Method 8081A. The method specifies an Electron Capture Detector (ECD), which is extremely sensitive for chlorinated compounds, however, it does not provide any confirmatory information about the peak. To reduce the occurrence of misidentifications, the method requires the use of two GC columns of dissimilar selectivity in a parallel configuration. The EPA considers an analyte’s presence confirmed if it has a peak at the pre-determined retention time on both columns.
The orthogonal selectivity provided by the Zebron MultiResidue-1 and the MultiResidue-2 allows for baseline resolution of all 20 chlorinated pesticides, surrogates, and internal standard in 10 minutes (Figure 7). Two elution order changes are observed between the phases, demonstrating that the phases are different enough to provide accurate confirmation.
Conclusion
Figure 2: Multi-Pesticide Residue Analysis using Zebron MultiResidue™-2
Figure 3: Nitrogen & Phosphorous Pesticides (NPM-102)
Figure 4: Nitrogen & Phosphorous Pesticides (NPM-106)
Figure 5: Nitrogen Pesticides (NPM-633)
Figure 6: Organophosphorous Pesticides (SPM-844)
Peak No. I Sample Analyte 1 Dichlorvos 2 EPTC 3 Butylate 4 3,5-Dichlorobenzoicacid(methylester) 5 Vernolate 6 Pebulate 7 Mevinphos 8 4-Nitrophenol(methylester) 9 MevinphosIsomer 10 Trichlorfon 11 Dicamba(methylester) 12 MCPP(methylester) 13 Molinate 14 Tebuthiuron 15 MCPA(methylester) 16 DEET 17 Tetraethylpyrophosphate(methylester)
Peak No. I Sample Analyte 18 Demeton 19 Dichloroprop(methylester) 20 Trifluralin 21 Thionazin 22 Cycloate 23 Benefin 24 Propachlor 25 Ethoprop 26 Chlorpropham 27 2,4-D(methylester) 28 Sulfotep 29 Naled 30 Phorate 31 Dicrotophos 32 Pentachlorophenol(methylester) 33 Profluralin 34 DemetonIsomer
Peak No. I Sample Analyte 35 Prometon 36 Atraton 37 Monocrotophos 38 AtratonIsomer 39 Silvex(methylester) 40 Terbufos 41 Propazine 42 Diazinon 43 Pronamide 44 Atrazine 45 Simazine 46 Terbuthylazine 47 Dioxathion 48 Fonofos 49 Dimethoate 50 2,4,5-TMethylester 51 Disulfoton
Peak No. I Sample Analyte 52 Chloramben(methylester) 53 PhosphamidonIsomer 54 Secbumeton 55 Dichlofenthion 56 2,4-DB(methylester) 57 Terbacil 58 Dinoseb(methylester) 59 Alachlor 60 Chlorpyrifosmethyl 61 Phosphamidon 62 Ronnel 63 Prometryn 64 Ametryn 65 Bentazon(methylester) 66 Aspon 67 Simetryn 68 Metribuzin
Peak No. I Sample Analyte 69 Methylparathion 70 Terbutryn 71 Metolachlor 72 Malathion 73 DCPA 74 Fenitrothion 75 Chlorpyrifos 76 Trichloronate 77 Triadimeton 78 Pichloram(methylester) 79 Isopropalin 80 Fenthion 81 MGK-264Isomer 82 Parathion 83 Merphos 84 Bromacil 85 ClofenvinfosIsomer
Peak No. I Sample Analyte 86 MGK-624 87 Pendimethalin 88 Diphenamid 89 Clofenvinfos 90 Butachlor 91 Crotoxyphos 92 Stirofos 93 Tokuthion 94 Oxadiazon 95 MerphosOxide 96 Napropamide 97 Fenamiphos 98 Oxyflurofen 99 Acifluorfen 100 Carboxin
Peak No. I Sample Analyte 101 Ethion 102 Tricyclazole 103 Fensulfothion 104 Carbofenotion 105 Famfur 106 Norflurazon 107 Hexazinone 108 EPN 109 Phosmet 110 Leptophos 111 Azinphos-Methyl 112 Fenarimol 113 Azinphos-ethyl 114 Coumaphos
Peaks for Figure 1 and Figure 2:
ConditionsColumn: ZebronMultiResidue-2Dimensions: 30meterx0.25mmx0.20µmPart Number: 7HG-G017-10Injection: Splitlessat260°C,1µLCarrier Gas: [email protected]/min (ConstantFlow)Oven Program: 80°Cfor0.5minto150°C@ 10°C/minto240°C@4°C/min to320°C@15°C/minfor3minDetection: MSD@320°C;45-400amu
6 8
1
23
4
5
10 12 14 16 18 20 22 24 26 28 30 32 34 min
16183
10 12 14 16 18
69
8
10
11 12
13
14
15
16
17
18
19
2023
24
25
26
27
28
29
30
31
32
33
34
35-37
40
45
46
47-49
54
53
5051,52
38-39
41
42-44
21,22
7
26 28 30 32
92
94
95
96,97
98
99
111,112
114
113
100
101
102
103104 105
106
107
108
109
110
93
20
55
22 24
56,57
5962
58
60,61
63
64
65
66,67
70
68,69
71 72
73
75
77
7674
78
79-81 87
82
85
83,84
86
88
89
90
91
min min
min
ConditionsColumn: ZebronMultiResidue-1Dimensions: 30meterx0.25mmx0.25µmPart Number: 7HG-G016-11Injection: Splitlessat260°C,1µLCarrier Gas: [email protected]/min (ConstantFlow)Oven Program: 80°Cfor0.5minto150°C@ 10°C/minto240°C@4°C/min to320°C@15°C/minfor3minDetection: MSD@320°C;45-400amu
8 10 12 14 16 18 20 22 24 26 28 30 32 34
1
23
28
29
26 3130
37
18
36
49
32
43
53
52
50
51
39
40
61
60
59
65
62
63
64,67,69
66
68
70
72
73
84
80,76
77
27
16
2
4
3
8,5
5457
58
56
55
9
6
11
12-14
15
1016 18 22 2491
94,95
98
100102
99101
103
104105
106
107
108
109
110
111
112,113
11496,97
92
93
90
33,35 41,45,44
42,47
46,48
71,74,75,78
79,82
7
17min
min
Conditions (are the same for both columns)
Injection: Splitlessat250°C,1µLCarrier Gas: [email protected]/min(ConstantFlow)Oven Program: 100°Cfor0.5minto220°C@35°C/minto 340°C@20°C/minfor2minDetection: ElectronCapture(ECD)@350°CSamples: 1.Tetrachloro- 12. EndosulfanI m-xylene(TCMX)(surr) 13. 4,4’-DDE 2.1-Bromo-2- 14. Dieldrin nitrobenzene(IS) 15. Endrin 3.a-BHC 16. 4,4’-DDD 4.g-BHC(Lindane) 17. EndosulfanII 5.b-BHC 18. Endrinaldehyde 6.d-BHC 19. 4,4’-DDT 7.Heptachlor 20. Endosulfansulfate 8.Aldrin 21. Methoxychlor 9.Heptachlorepoxide 22. Endrinketone 10.g-Chlordane(trans) 23. Decachlorobiphenyl 11.a-Chlordane(cis) (DCB)(surr)
4
1
23
4
5
6 7
8
9 10111213
14
151617
18
19 20
21
2223
16153
6 8 10 min
ZebronMultiResidue-130meterx0.32mmx0.50µm
3
16154
4
1
2
3
4
5
6
7
8 9
1011
1213
14
151617
18
1920
21
2223
5 6 7 8 9 min
ZebronMultiResidue-230meterx0.32mmx0.25µm
11 12 13 14 15 16 17 18 19 20 min
1
2
3
5
6
4
7
8
9
ZB-MR-130meterx0.25mmx0.25µm
11 12 13 14 15 16 17 18 19 min
1
2
3
4
6
5
7
8
9
ZB-5ms30meterx0.25mmx0.25µm
15 20 25 30 35 40 min
12
3
56
4
7
8
9
5 4
ZB-MR-230meterx0.25mmx0.20µm
Samples: 1. EPTC 2. Mevinphos 3. Ethoprop 4. Atrazine 5. Propazine 6. Prometryn 7. Terbutryn 8. Triadimefon 9. Diphenamid
14 16 18 20 22 24 26 28 30 32 min
12
3
4
5
6
7
8
ZB-MR-130meterx0.25mmx0.25µm
14 15 16 17 18 19 20 21 min
1
2
3
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5
6
78
ZB-5ms30meterx0.25mmx0.25µm
2422 26 28 30 32 34 36 38 min
12
3
4
5
6
7
8
ZB-MR-230meterx0.25mmx0.20µm
Samples: 1. Propachlor 2. Trifluralin 3. Benefin(Benfluralin) 4. Profluralin 5. Isopropalin 6. Pendimethalin 7. Oxadiazon 8. Oxyfluorfen
1
2
34
5
6
7
12 14 16 18 20 22 24 26 28 30 32 min
ZB-MR-130meterx0.25mmx0.25µm
14 15 16 17 18 19 20 21 22 min
1
2
3
4
5
6
7Overloading due toPolarity Mismatch
ZB-5ms30meterx0.25mmx0.25µm
22 24 26 28 30 32 34 36 38 40 42 44 min
1
2
3
5
6
4
7
ZB-MR-230meterx0.25mmx0.20µm
Samples: 1. DEET 2. Terbacil 3. Metribuzin 4. Bromacil 5. Triadimefon(Amiral) 6. Tricyclazole 7. Hexazinone
3
2
5
47
6 8
9
10
17 18 19 20 21 22 23 24 25 26 min
1
ZB-MR-130meterx0.25mmx0.25µm
16 17 18 19 20 21 22 23 24 25 min
1
3
2
45
7
6
8
9
10
ZB-5ms30meterx0.25mmx0.25µm
18 20 22 24 26 28 30 32 34 min
1
2
3
45
8
76
9
10
ZB-MR-230meterx0.25mmx0.20µm
Samples: 1. Terbufos(Terbuphos) 6.Famophos(Famphur) 2. Phosphamidon(isomer) 7.Carbophenothion 3. Phosphamidon 8.Phosmet 4. Chlorfenvinphos 9.Leptophos 5. Ethion 10.Azinphos-ethyl
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