Evaluation of interferences between matrix-analyte for the correct identification of the pesticides by GC-QqQ-MS/MS and LC-QqQ-MS/MS

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1 Evaluation of interferences between matrix-analyte for the correct identification of the pesticides by GC-QqQ-MS/MS and LC-QqQ-MS/MS

2 CONTENTS 1. Aim and scope.... Short description Apparatus and consumables.... Chemicals Procedure Sample preparation Extraction methods NL method Ethyl acetate method Citrate QuEChERS Citrate QuEChERS without clean-up Measurement Instrumentation and analytical conditions for the LC- MS/MS system UPLC (Agilent Technologies) triple quadrupole system (Agilent Technologies) Instrumentation and analytical conditions for the GC- MS/MS system A GC system (Agilent Technologies) triple quadrupole system (Agilent Technologies) Evaluation of the commodities Potential false negatives. References... 7 APPENDIX I: MASS TRANSITIONS... APPENDIX II: RESULTS... 1 Page 1 of 1

3 1. Aim and scope This report study the influence of the interferences coming from the matrices of different commodities by using the most applied multiresidue extraction methods in the EU laboratories by LC and GC-MS/MS.. Short description Twenty five homogenous blank samples (apple, aubergine, avocado, banana, broccoli, carrot, cauliflower, celery, cherry, cucumber, green beans, green tea, leek, lemon, lettuce, melon, onion, orange, parsley, pear, pepper, potato, pumpkin, strawberry and tomato) were extracted with acetone, ethyl acetate and acetonitrile and the obtained extracts were analysed by GC-MS/MS and LC-MS/MS. The interferences taken into account were those whose relative abundance were higher than 5% respect to the standard in tomato at.1 mg/kg and their retention time were within ±. min according to the analyte in the standard mix. 3. Apparatus and consumables Automatic pipettes, suitable for handling volumes of 1 µl to 5 µl and 1 ml to 3 ml 5 ml PTFE centrifuge tubes 5 ml PTFE centrifuge tubes 15 ml PTFE centrifuge tubes Vortex Automatical shaker (Agitax, Cirta Lab) Turrax homogeniser Centrifuge, suitable for the centrifuge tubes employed in the procedure and capable of achieving at least 33 rpm Injection vials, ml, suitable for LC and GC auto-sampler. Chemicals Acetone p.a. Petroleum ether Dichloromethane Acetonitrile HPLC grade Ethyl acetate HPLC grade Anhydrous sodium sulphate Anhydrous magnesium sulphate Sodium hidrogenocitrate sesquihydrate Tri-sodium citrate dihydrate Page of 1

4 Sodium chloride Primary Secondary Amine (PSA) Formic acid Ultra-pure water Pesticides standards 5. Procedure 5.1. Sample preparation Following Document No. SANCO/1571/13, the sample was perfectly homogenised by grinding finely at its arrival to the laboratory. 5.. Extraction methods Twenty five blank samples were extracted with four different methods described below to compare the interferences matrix-analyte found NL-method 1. Weigh 15 g ±.1 g of sample in 5 ml PTFE centrifuge tube.. Add ml of acetone and 15 g NaSO. 3. Blend the sample using a Turrax homogeniser for 3 s at 15 rpm (extraction step).. Add ml of petroleum ether and 1 ml of dichloromethane. 5. Blend it again by Turrax for 3 s at 15 rpm (partitioning step).. Centrifuge for 5 min at 37 rpm. 7. Evaporate an aliquot of the extract with a nitrogen stream until dryness. a. for LC analysis evaporate 17 µl extract and reconstitute with 5 µl acetonitrile : water (:, v/v). b. for GC analysis evaporate 17 µl extract and reconstitute with 5 µl of ethyl acetate. With this treatment, 1 ml of sample extract represents. g of sample in LC and in GC the final matrix concentration is 1 g/ml Ethyl acetate method 1. Weigh 1 g ±.1 g of sample in 5 ml PTFE centrifuge tube.. Add 1 ml of ethyl acetate, g MgSO and 1.5 g NaCl. 3. Shake automatically for 15 min.. Centrifuge for 5 min at 37 rpm. 5. For LC, evaporate 5 µl of the extract with a nitrogen stream and reconstitute with 5 µl of acetonitrile : water (:, v/v). For GC analysis, an aliquot of the extract is injected directly. Page 3 of 1

5 With this treatment, 1 ml of sample extract represents. g of sample in LC and in GC the final matrix concentration is 1 g/ml Citrate QuEChERS 1. Weigh 1 g ±.1 g of sample in 5 ml PTFE centrifuge tube.. Add 1 ml of acetonitrile. 3. Shake automatically for min.. Add g of MgSO, 1 g NaCL, 1 g trisodium citrate dehydrate and.5 g sodium hydrogenocitrate sesquihydrate. 5. Shake automatically for min.. Centrifuge for 5 min at 37 rpm. 7. Transfer 5 ml of the supernatant into 15 ml PTFE centrifugue tube containing 75 mg MgSO and 15 mg PSA.. Vortex for 3 seconds. 9. Centrifuge for 5 min at 37 rpm. 1. For GC, evaporate 5 µl of the extract and reconstitute with 5 µl of ethyl acetate. For LC analysis, 5 µl of the extract were diluted with µl of water. With this treatment, 1 ml of sample extract represents. g of sample in LC and in GC the final matrix concentration is 1 g/ml Citrate QuEChERS without clean-up This procedure was the sample than the previous one excluding the steps number 7 and Measurement Both LC and GC systems were operated in multiple reaction monitoring mode (MRM). Selected reaction monitoring (SRM) experiments with two SRM transitions were used. The mass transitions used are presented in Appendix I. 5.. Instrumentation and analytical conditions for the LC- MS/MS system Liquid chromatograph (19 UPLC, Agilent Technologies) Column: Zorbax Eclipse Plus C.1 mm x 1 mm and 1. µm particle size (Agilent) Mobile phase A: Water (.1% formic acid) Mobile phase B: Acetonitrile (.1% formic acid and 5% HO) Column temperature: 35ºC Flow rate:.3 ml/min Injection volume: 5 µl Page of 1

6 Mobile phase gradient for pesticides analyse Time [min] Mobile phase A Mobile phase B % % % % 15 % 1% 17 % 1% Re-equilibration with initial mobile phase:.5 minutes Triple quadrupole mass spectrometer (9, Agilent Technologies) Ionisation mode: Positive and negative mode Drying gas temperature: 1 C Drying gas flow: 13L/min Nebuliser pressure: 5 psi Sheath gas temperature: 375ºC Sheath gas flow: 1 L/min Fragmentor: 3 V Capillary voltage: 3 V (in positive and negative mode) Nebuliser and collision gas: nitrogen 5.5. Instrumentation and analytical conditions for the GC- MS/MS system Gas chromatograph (79 A GC system, Agilent Technologies) Column: HP 5MS UI 15 m.5 mm ID and.5 µm Injection mode: Splitless Ultra-Inert liner with a glass wool frit Injection volume: µl Injector temperature: held at C (.1 min) and then ramped up to 3 C at C/min. Carrier gas: helium at constant pressure (1.1 psi) Carrier gas purity: % Oven temperature: 7 C for 1 min, programmed to 15 C at 5 C/min, then to C at C/min and finally to C at 1 C/min and kept at ºC for.7 min Triple quadrupole mass spectrometer (7, Agilent Technologies) Ionisation mode: electron impact ionisation Transfer line temperature: C Ion source temperature: C Quadrupoles temperature: 15ºC Page 5 of 1

7 Quenching gas: helium Quenching gas flow:.5 ml/min Collision gas: nitrogen Collision gas flow: 1.5 ml/min Collision gas purity: % Solvent delay: minutes. Evaluation of the commodities. The 5 different commodities were extracted with the four extraction methods and their extracts were injected both in LC and GC systems. Each of the matrices was evaluated taking into account the presence of a signal in the ±. min retention time of the compound and with intensity at least 5 % of the standard signal at.1 mg/kg. To develop this study in LC, 1 pesticides were monitored. In Appendix II, Figure 1 is represented the percentage of compounds with some interference, either in one transition (blue) or in both transitions (green). As can be seen, the highest percentage did not reach % in any case. In general, not big differences could be observed. The extracts were also injected in GC and their data compiled in Figure, taking into account the pesticides. In this case, the presence of interferences that could affect to identification/quantification is significantly bigger than in LC. In general, the highest percentage of interferences was found when NL-method was applied, going up to 1% for the case of avocado matrix. In Figure 3 all the results obtained per extraction method, the 3 compounds (LC and GC), two transitions per each and the 5 matrices are plotted considering ±. min (blue) and ±.1 min (red) as retention time window. In the view of the results, the number of interferences was very low (close to 1 % in LC and % in GC as maximum, both for NL-method). Reducing retention time window to ±.1 min, the probability to find a peak of the matrix interfering with the pesticide was also decreased, but not drastically. 7. Potential false negatives. It was considered a potential false negative a not resolved interference observed in the blank from the pesticide peak which affected to the ion ratio making the difference higher than 3 % from expected one, as it is established at the AQC procedures. These results have been represented in Figure. In LC the worst result was obtained for avocado with NL-method, and it was around % the number of potential false negatives. For the rest, they were below or close to %, except for green beans with citrate QuEChERS and leek with citrate QuEChERS without clean-up. The compound showing more potential false negatives is fenthion-sulfoxide (interfering in 7 out of 1 Page of 1

8 matrices), followed by chlorpyrifos methyl, fenhexamid, dichlorvos and phosmet from the list of compounds evaluated. In GC the worst result was obtained for avocado with NL-method, with around % of compounds evaluated showing potential false negatives. In general, NL-method was the extraction method providing higher percentage of compounds with potential false negatives. In the case of leek and banana the method with higher number of potential false negative was citrate QuEChERS without clean-up and in orange was ethyl acetate. Phenothrin was the compound generating potential false negatives in almost all matrices tested, followed by molinate, fenpropidin, tolylfluanid, chlorbromuron and isoprothiolane.. References Ana Lozano, Barbara Kiedrowska, Jos Scholten, Marijke de Kroon, André de Kok, Amadeo R. Fernández-Alba. Miniaturisation and optimisation of the Dutch mini-luke extraction method for implementation in the routine multi-residue analysis of pesticides in fruits and vegetables. Food Chemistry 19 (1) 1 Samanta Uclés, Noelia Belmonte, Milagros Mezcua, Ana B. Martínez, M. Jesus Martinez-Bueno, Miguel Gamón & Amadeo R. Fernández-Alba. Validation of a multiclass multiresidue method and monitoring results for 1 pesticides in fruits and vegetables by gas chromatography-triple quadrupole mass spectrometry. Journal of Environmental Science and Health, Part B (1) 9, Ana Lozano, Łukasz Rajskia, Noelia Belmonte-Valles, Ana Uclés, Samanta Uclés, Milagros Mezcua, Amadeo R. Fernández-Alba. Pesticide analysis in teas and chamomile by liquid chromatography and gas chromatography tandem mass spectrometry using a modified QuEChERS method: Validation and pilot survey in real samples. Journal of Chromatography A, 1 (1) Analytical quality control and method validation procedures for pesticide residues analysis in food and feed. Document Nº SANCO/1571/13. Page 7 of 1

9 APPENDIX I: MASS TRANSITIONS Table 1. Detection and chromatographic parameters for the selected compounds analysed by LC-MS/MS. # Name Rt (min) SRM1 CE1 (ev) SRM CE (ev) 1,-D 7. 19> >13 15 Acephate >13 5 1> Acetamiprid.3 3>1 3>5 15 Aldicarb > > Aldicarb-Sulfone 1.7 3>1 5 3> 1 Aldicarb-Sulfoxide 1.7 7>13 5 7>9 1 7 Azinphos-Methyl >1 31>13 Azoxystrobin 9.3 >37 1 >3 9 Benfuracarb >5 1 11>195 1 Bifenazate > >17 11 Bitertanol 1. 33>9 5 33> Boscalid > > Bromuconazole >159 37>7 1 Bupirimate.9 317>7 317>1 15 Buprofezin 1.7 3>1 1 3> Carbaryl 7.9 >15 1 >17 17 Carbendazim 1. 19> >13 1 Chlorantraniliprol.3 >53 1 > 19 Chlorfenvinphos > >99 Chlorpyrifos-Methyl >9 1 3> Clofentezin > >1 Clomazone.59 >1 1 >15 3 Cyazofamid >1 1 35>1 15 Cymoxanil >1 199> Cyproconazole >15 3 9>7 1 Cyprodinil 7.9 >93 >77 7 Cyromazine. 17> > Demeton-S- Methylsulfoxide 1. 7>19 7>19 9 Diazinon > >153 3 Dichlorvos.53 1> > Dicrotophos. 3>11 3>7 3 Diethofencarb 9.1 > 5 > Difenoconazole 1. > >51 3 Diflubenzuron >15 311> Dimethoate.31 3> > Dimethomorph.7 3>31 3>15 37 Diniconazole >159 3>7 3 Dithianon 13. 9> 3 9> Dodine.5 > >57 Page of 1

10 # Name Rt (min) SRM1 CE1 (ev) SRM CE (ev) Epoxiconazole > > Ethion > >171 1 Ethirimol.53 1>1 1>3 5 3 Ethoprophos 9. 3> >97 3 Fenamidone >9 31>5 5 5 Fenamiphos 9.5 3>3 1 3>17 Fenamiphos-Sulfone.95 33> 1 33>1 7 Fenamiphos-Sulfoxide 5. 3>9 3>1 Fenarimol > 331>59 9 Fenazaquin > > Fenbuconazole >15 337> Fenhexamid >97 5 3> Fenoxycarb 1.3 3>11 5 3> 53 Fenpropimorph 7.7 3>17 3 3> Fenpyrazamide >7 1 33>3 55 Fenpyroximate >3 1 >17 5 Fenthion >7 79> Fenthion-Sulfone > > Fenthion-Sulfoxide > 1 95> Fipronil > >5 Flonicamid.31 3>199 3> Fluazifop 9.7 3> 15 3>5 Flubendiamine >7 15 1>5 3 Fludioxonil 9.3 7>19 3 7>15 3 Flufenacet 1. 3> > Flufenoxuron >15 9>11 5 Fluopyram > 397>173 7 Fluquinconazole >37 37>1 5 Flusilazol 1. 31>7 1 31>15 9 Flutriafol 7.5 3>95 5 3>7 1 7 Formetanate 1. >15 > Fosthiazate 7.59 > 1 >1 7 Haloxyfop 1.3 3>31 1 3> 73 Hexaconazole > >7 7 Hexythiazox > 1 353> Imazalil.5 97> > Imidacloprid 3. 5>9 15 5> Indoxacarb >1 5>3 5 7 Ioxynil.1 37> > Iprovalicarb >3 31>119 1 Isofenfos methyl > > Isoprocarb.19 19> >95 15 Isoxaflutole > > 5 3 Kresoxim-Methyl >7 31> 1 Page 9 of 1

11 # Name Rt (min) SRM1 CE1 (ev) SRM CE (ev) Linuron 9.1 9>1 9> Lufenuron 1. 59> >3 1 Mandipropamid 9. 1>35 1>3 7 Mepanypirim 9.9 >7 1 >191 1 Meptyldinocap >193 95> Metalaxyl 7. > 5 > Metconazole 1. 3>15 3>7 91 Methamidophos 1.1 1>15 1 1>9 1 9 Methidathion >15 33> Methiocarb 9.1 >11 1 > Methiocarb-Sulfoxide. >15 1 >17 95 Methomyl.9 13>1 13> 9 Methoxyfenozide > > Metobromuron.11 59> >1 1 9 Monocrotophos 1. >193 5 > Myclobutanil 9. 9>15 9> Nitempyram >5 1 71> Omethoate 1.1 1>13 5 1>15 1 Oxadixyl.1 79> > Oxamyl >9 5 37>9 1 1 Oxyfluorfen 1. 3>5 5 3> Paclobutrazol.9 9>15 3 9>7 1 1 Penconazole 1. >159 > Pencycuron >15 39>9 1 Pendimethalin 13.7 >1 > Phenthoate >7 31>79 11 Phosalone >1 3> Phosmet 9. 31>1 31> Phoxim >19 99> Pirimicarb.7 39> >7 11 Pirimiphos-Methyl >1 3>1 115 Prochloraz.75 37>3 1 37> Profenofos 1 375> > Propamocarb >1 1 19> Propaquizafop 1.17 > >1 119 Propargite >31 3>57 1 Propiconazole 1.5 3> > Propoxur 7. 1>1 5 1> Propyzamide 9.1 5>19 1 5> Proquinazid > >9 1 Prothioconazole >3 15 3>1 15 Prothiofos 1. 35>1 35>11 1 Pymetrozine.9 1>15 1>51 17 Pyraclostrobin >19 3>13 Page 1 of 1

12 # Name Rt (min) SRM1 CE1 (ev) SRM CE (ev) 1 Pyrethrin > >13 19 Pyridaben > >17 13 Pyridate > > Pyrimethanil. >13 >17 13 Pyriproxyfen 1.3 3>15 3> Quinoclamine.13 >15 5 >77 13 Quinoxyfen 11. 3>7 5 3> Quizalofop-ethyl >71 373> Rotenone >13 395> Spinosyn A. 733>1 733>9 13 Spinosyn D >1 77>9 139 Spirodiclofen 1. 11> > Spiromesifen > >55 11 Spirotetramat > >7 1 Spiroxamine >1 9>1 13 Tebuconazole 9.9 3>15 3>7 1 Tebufenozide > > Tebufenpyrad >15 33> Teflubenzuron > > Terbuthylazine 9.5 3> >1 1 Tetraconazole > >7 19 Thiabendazol 1.9 >175 3 > Thiacloprid >1 1 53>1 151 Thiamethoxam.5 9>11 1 9>11 15 Thiodicarb.7 355>1 355> 153 Tolclofos-Methyl >9 1 31> Triadimenol 9 9>7 5 9> Triazophos > 1 31>1 15 Trichlorfon.97 57>1 57> Trifloxystrobin 1.9 9> 1 9>1 15 Triflumuron >15 359> Triticonazole > > Zoxamide > >159 Page 11 of 1

13 Table. Acquisition and chromatographic parameters for the selected compounds analysed by GC-MS/MS. # Name Rt (min) SRM1 CE1 (ev) SRM CE (ev) 1,'-DDE 11.5 >17 3 >11,'-DDT +,'-DDD >15 35> ,5-Dichloroaniline > >9 3-Chloroaniline.7 17> >1 1 5,'-DDE 1. >17 3 >11,'-DDT >15 35>199 7 Acrinathrin >11 5 9>11 Alachlor.5 1>1 1 1>13 9 Aldrin 9. 93>1 93>57 1 Ametryn.3 7>15 5 7>1 11 Anthraquinone 9.1 >1 1 >15 1 Atrazine.59 15>5 1 15> Azoxystrobin 1.3 3>39 1 3>15 1 Benalaxyl >15 >17 15 Bifenox > >1 5 1 Bifenthrin 1. 11>1 1 11> Biphenyl >1 15>1 1 Bixafen > > Boscalid >11 1 1>7 5 Bromopropilate >15 31>155 1 Bupirimate 1. 73> >1 15 Buprofezin > >1 1 3 Butralin 1.1 >17 >19 1 Butylate > >1 3 5 Cadusafos > >73 1 Captan 1. 19> > Carbofuran.5 1>19 1 1>1 1 Carbophenothion >13 1 3> Carbosulfan 1.7 1>19 1 1>1 1 3 Chinomethionat > 1 > Chlorbromuron 3. 33>1 5 33>5 1 3 Chlordane > 373> Chlorfenapyr 1.5 7>7 15 7> 5 3 Chlorfenvinphos 1.3 7>159 7>1 35 Chlorobenzilate > > Chlorothalonil 7.39 >31 > Chlorpropham > > Chlorpyrifos > > 5 39 Chlorpyrifos-Methyl.3 >93 >71 1 Chlorthal-Dimethyl > >1 35 Page 1 of 1

14 # Name Rt (min) SRM1 CE1 (ev) SRM CE (ev) 1 Chlozolinate > >1 5 Coumaphos 15. 1>1 1 3> Cyfluthrin >17 5 > 1 Cypermethrin >17 5 9>11 5 Cyproconazole >111 1 >15 1 Cyprodinil 1.3 > > Deltamethrin >93 53>17 5 Desmethyl-Pirimicarb.1 15>9 1 >15 9 Diazinon 7. 3> > Dichlofluanid 9.1 >13 17> Dichlorvos.93 15> > Diclobutrazole 1.5 7> >1 53 Dicloran. >17 5 >1 5 Dicofol > > Dieldrin >3 35>3 5 Diethofencarb 9.1 7> > Dimethenamid.11 3> > Dimethipin.55 11>5 1 11> Diphenylamine > >1 Disulfoton 7.7 1>19 5 1>1 1 1 Disulfoton-Sulfoxide 3. 15>97 3 1> DMST.7 1>79 1> Dodemorph > 15>97 1 Endosulphan, alpha > 15 1> 5 5 Endosulphan, beta > 1 39> 15 Endosulphansulphate >9 5 37> 7 Endrin > > 5 EPN > > Epoxiconazole 1. 19> > Ethion > > Ethofumesate >11 5 7> Ethoprophos > > Ethoxyquin. >17 15 > Etofenprox > > Etrimfos 7.5 9>11 5 9>153 7 Fenamidone 1.5 >1 3>13 77 Fenarimol > > Fenazaquin >15 5 1> Fenbuconazole > >1 15 Fenchlorphos.7 5>7 1 5> 3 1 Fenhexamid >7 177>113 1 Fenitrothion 9. 77> 5 77>19 Page 13 of 1

15 # Name Rt (min) SRM1 CE1 (ev) SRM CE (ev) 3 Fenpropathrin >15 5>1 1 Fenpropidin.93 9> >9 3 5 Fenpropimorph 9.7 1>7 1 1>11 1 Fenthion 9.5 7>19 7>19 7 Fenvalerate/Esfenvalerate RR/SS > >9 Fenvalerate/Esfenvalerate RS/SR > >9 9 Fipronil > >13 9 Fipronil-Desulfinil.71 3> > Fipronil-Sulfone >55 33> Flamprop-Isopropyl >15 5 3> Flamprop-Methyl 1.9 7>15 3> Flonicamid 5. 17> > Fluacrypirim >1 3 15> Fluazifop-p-Butyl 1. >91 15 >3 97 Flucythrinate I > > Flucythrinate II > > Fludioxonil 1. >17 3 > Fluopicolide >1 173> Fluopyram > > Fluquinconazole 15. 3>9 3> 3 13 Flusilazole >15 33>15 1 Flutolanil > >1 15 Flutriafol > >95 1 Fluvalinate-tau >55 1 5> 17 Folpet 1. 17> >7 5 1 Fonofos.9 137>19 5 > Formothion >93 5 >15 11 Fosthiazate > > HCB.19 >9 5 >1 11 HCH, alpha-.3 19> > HCH, beta-. 19> > Heptachlor.3 7>37 1 7> Heptachlorepoxid, cis >1 13> Heptachlorepoxid, trans > > Heptenophos.9 1> > 1 11 Hexaconazole 11. 1>159 1> Indoxacarb >13 1 >1 1 Iprodione >5 1 31>5 11 Iprovalicarb I > >9 1 1 Iprovalicarb II > > Isazofos > >1 5 1 Isocarbophos >1 1 3>1 Page 1 of 1

16 # Name Rt (min) SRM1 CE1 (ev) SRM CE (ev) 15 Isofenphos-Ethyl 1. 13> > Isofenphos-Methyl > > Isoprothiolane >5 15 1> Isopyrazam > >33 19 Kresoxim-Methyl 1. >11 5 > Lambda-Cyhalotrin > > Lindane.7 19> > Malaoxon. 17> > Malathion > >15 13 Mecarbam > > Mepanypirim >7 3 > Merphos >57 19> Metalaxyl.7 >13 >1 13 Metazachlor 1. 9> > Metconazole >9 15>99 1 Methidathion >5 5 15> Methiocarb 9. 1> > Methiocarb-Sulfone.75 11> > Methoxychlor 1.5 7>19 5 7>115 1 Metolachlor 9. 3>1 1> Mevinphos > > Molinate.5 1>55 1 1> Myclobutanil > > Napropamide >7 3 71> Nuarimol > > Ofurace >15 3> Orthophenylphenol.39 17> > Oxadixyl > > Paclobutrazol >15 1 3>17 15 Paraoxon-Methyl >79 5 3>1 155 Parathion-Ethyl > > Parathion-Methyl.3 3> > Pebulate > > Penconazole 1.5 >157 5 > Pendimethalin >1 1 5> Pentachloroaniline >19 5 3> Permethrin I > > Permethrin II > > Phenothrin I >1 13> Phenothrin II 1. 13>1 13> Phenthoate 1. 7>11 1 7> 5 1 Phorate >19 31> Phorate-Sulfone > >13 1 Phosmet 1.7 1>77 3 1> Page 15 of 1

17 # Name Rt (min) SRM1 CE1 (ev) SRM CE (ev) 19 Phosmet-Oxon >77 3 1> Phthalimide. 17> > Picolinafen 1.1 3> > Picoxystrobin > > Pirimicarb 7. 3>1 1 1>9 17 Pirimiphos-Methyl > > Procymidone 11 3>9 3>55 17 Profenofos > > Prometon.5 5>13 3 5> Prometryn.7 1>1 1 1> 179 Propaphos >1 1 > Propargite > > Propazine.7 1>17 9> Propiconazole I > > Propiconazole II > >191 1 Propyzamide > > Prosulfocarb.77 1> 3 51>1 5 1 Prothiofos > > Pyraclostrobin >77 13>1 1 1 Pyrazofos > > Pyridaben >117 17> Pyrifenox >7 1 > 191 Pyrimethanil > > Pyriproxyfen >7 1 13>9 193 Quinalphos 1.5 1> > Quinoxyfen >7 5 37> Quintozene.3 95> > Secbumeton 7.5 5>19 5 5> Spirodiclofen > >19 19 Spiromesifen >5 3 7> Sulfotep 5.95 >1 1 3>1 1 Sulprofos > > Tebuconazole >15 5>153 1 Tebufenpyrad > >7 5 3 Tecnazene > >13 Tefluthrin > > Terbufos. 31> >175 1 Terbumeton.79 19>15 5 5> Terbutryn 9. 1>15 3 1>17 1 Tetrachlorvinphos > > Tetraconazole > 3 33>1 3 1 Tetradifon > > Tetrahydrophthalimide > 5 151>1 1 Tetramethrin I 1.3 1>77 3 1>17 15 Page 1 of 1

18 # Name Rt (min) SRM1 CE1 (ev) SRM CE (ev) 13 Tetramethrin II 1.3 1>77 3 1> Tolclofos-Methyl. 5>5 15 5> 5 15 Tolylfluanid >91 3> Triadimefon 9.7 >11 5 > Triazophos > >1 1 1 Trifloxystrobin 13. >19 3 > Trifluralin 5. 3> 1 >1 15 Vinclozolin.33 1> >19 Page 17 of 1

19 APPENDIX II: RESULTS Citrate QuEChERS without Clean-up Citrate QuEChERS without Clean-up transitions 1 transition transitions 1 transition Citrate QuEChERS Citrate QuEChERS Ethyl Acetate Ethyl Acetate transitions 1 transition transitions 1 transition NL-Method NL-Method Figure 1. Percentage of possible interferences analyzed by LC for each of the four methods in the ±. min retention time range. In blue, percentage of compounds presenting the interference in only one transition, and in green, percentage of compounds with interferences in both transitions. Page 1 of 1

20 Citrate QuEChERS Citrate QuEChERS without Clean-up without Clean-up transitions 1 transition transitions 1 transition Citrate QuEChERS Citrate QuEChERS Ethyl Acetate Ethyl Acetate transitions 1 transition transitions 1 transition NL-Method NL-Method Table. Percentage of possible interferences analyzed by GC for each of the four methods in the ±. min retention time range. Page 19 of 1

21 % of Interferences % of Interferences % of Interferences,5 LC-QqQ-MS/MS,5,, 1,5 1,5 1,,5. min.1 min 1,,5, 1 SRM SRM 1 SRM SRM 1 SRM SRM 1 SRM SRM, 1 SRM SRM 1 CQ* CQ Ethyl Acetate NL-method CQ* Q-MS/MS,5 GC-QqQ-MS/MS, 1,5. min.1 min 1,,5 M 1 SRM SRM 1 SRM SRM Ethyl Acetate NL-method, 1 SRM SRM 1 SRM SRM 1 SRM SRM 1 SRM SRM CQ* CQ Ethyl Acetate NL-method Figure 3. Total percentage of interferences per extraction method (citrate QuEChERS without clean-up, CQ*, citrate QuEChERS, CQ, ethyl acetate and NLmethod), considering ±. min (blue) and ±.1 min (red) for retention time, and showing results found for one transition (1 SRM) or both transitions ( SRM). Page of 1

22 Strawberry LC-QqQ-MS/MS QC CQ wtih without no Clean-up Citrate QuEChERS Ethyl Acetate NL-method Dutch mini-luke GC-QqQ-MS/MS C wtih without no Clean-up trate QuEChERS hyl Acetate method tch mini-luke Figure 5. Potential false negatives detected in LC and GC for the selected matrices and with the four extraction methods: citrate QuEChERS without cleanup (blue), citrate QuEChERS (green), ethyl acetate (yellow) and NL-method (pink). Page 1 of 1

COMMISSION REGULATION (EU)

L 269/8 Official Journal of the European Union 13.10.2010 COMMISSION REGULATION (EU) No 915/2010 of 12 October 2010 concerning a coordinated multiannual control programme of the Union for 2011, 2012 and