Almería 23 rd -25 th October th Joint Workshop of the European Union Reference Laboratories for Residues of Pesticides

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2 Co-extracted matrix components of tomato. LC-TOF-MS Mass 1. Molecular weight of compounds 2. Number interfering compounds 3. Distribution of compounds

3 Co-extracted matrix components LC-TOF-MS Mass Mass 5870 Matrix compounds 8017 Matrix compounds Mass Mass 3419 Matrix compounds 2408 Matrix compounds Compounds with absolute height counts

4 LC-QqQ-MS/MS Skimer After 30 tea injections GC-Q-MS Liner after 40 tea injections

5 Co-extracted matrix components of tomato. LC-TOF-MS Mass m/z: Rt: 4-7 min 2408 Matrix compounds

6 Data base components- tomato matrix compounds Mass m/z: Rt: 4-7 min m/z: Rt: 7-13 min 2408 Matrix compounds DB: 750 components Tomato: 656 matrix compounds

7 Co-extracted matrix components of leek. LC-TOF-MS Mass m/z: Rt: 4-12 min 5870 Matrix compounds

8 Data base components- leek matrix compounds Mass m/z: Rt: 4-12 min m/z: Rt: 7-13 min 5870 Matrix compounds DB: 750 components Leek: 3032 matrix compounds

9 Matrix Nº of co-extracted compounds Rt: 0-17 min Rt: 7-13 min Green tea Orange Chamomille Leek Pepper Tomato

10 Green tea matrix compounds Dil 1/5. LC-TOF-MS Absolute height counts 4873 compounds 1970 compounds 2903 compounds

11 Orange matrix compounds Dil 1/5. LC-TOF-MS Absolute height counts 5394 compounds 2324 compounds 3070 compounds

12 TIC LC-TOF-MS. Orange and Green tea. Dil 1/5

13 Green tea Orange matrix 4873 compounds 5394 compounds Absolute height counts Absolute height counts 1732 compounds 1214 compounds Absolute height counts Absolute height counts

14 LC-TOF-MS XIC ± 20 ppm Fenuron at 100 µg/l Tomato Orange Number of co-extracted matrix compounds Rt: min Tomato 25 compounds Orange 101 compounds Mass Profiler Professional Agilent Tech.

15 LC-TOF-MS XIC of tomato matrix compounds and Fenuron Fenuron (100 ppb) LC-TOF-MS XIC of orange matrix compounds and Fenuron Fenuron (100 ppb)

16 Number of matrix coeluting compounds vs matrix suppression Orange matrix Dilution 1/30 Nº of coleuting Pesticide Rt (min) Suppression compounds (± 0.05 min) Ʃ Compounds height (counts) CINOSULFURON E+06 FENPROPIDIN E+07 PROPOXUR E+06 CARBOFURAN E+06 FORCHLORFENURON E+05 FLUOMETURON E+06 ISOPROTURON E+07 METALAXYL E+07 OFURACE E+06 HEPTENOPHOS E+06 SPINOSYN A E+07 METAZACHLOR E+07 BUPIRIMATE E+07 CYPRODINIL E+07 TRIADIMENOL E+06 FLAZASULFURON E+06 PROMECARB E+06 AZOXYSTROBIN E+06

17 Data base components- orange matrix compounds Mass m/z: Rt: 4-8 min m/z: Rt: 7-13 min 8017 Matrix compounds DB: 750 components Orange: 2743 matrix compounds

18 Data base components- orange matrix compounds. Dilution 1:10 Mass 5539 Matrix compounds DB: 750 components Orange: 1765 matrix compounds

19 Data base components- orange matrix compounds. Dilution 1:50 Mass Matrix Nº of compounds Rt: 0-17 min Nº of compounds Rt: 7-13 min TIC Area Rt: 7-13 min Orange E+09 Orange 1: E+09 Orange 1: E+09 Tomato E+09 Peeper E Matrix compounds DB: 750 components Orange: 1082 matrix compounds

20 Dilution effect on matrix suppression LC-TOF-MS 100 pesticides Orange matrix Spiked level 100 µg/l Serial dilution

21 Dilution effect on suppression decrease LC-TOF-MS XIC ± 20 ppm Diethofencarb at 100 µg/l Solvent Orange matrix No Dilution Dilution: 1/5 Dilution: 1/10 Dilution: 1/15

22 Orange matrix Dil 1/10. QuEChERS vs MiniLuke. LC-TOF-MS LC-TOF-MS TIC of orange matrix extract QuEChERS method MiniLuke method Number of co-extracted matrix compounds QuEChERS 4495 compounds MiniLuke 4117 compounds 4 th Joint Workshop of the European Mass Union Profiler Reference Professional Laboratories Agilent Tech. for Residues of Pesticides

23 Orange matrix Dil 1/10. QuEChERS vs MiniLuke. LC-TOF-MS Number of co-extracted matrix compounds Rt 0-17 min QuEChERS 4495 compounds MiniLuke 4117 compounds Mass Profiler Professional Agilent Tech. QuEChERS 1635 compounds Rt < 6 min Rt 6-12 min Rt > 12 min MiniLuke 1565 compounds QuEChERS 2514 compounds MiniLuke 2211 compounds QuEChERS 346 compounds MiniLuke 341compounds

24 Citrate QuEChERS 2 g sample Add 10 ml acetonitrile + TPP Shake Add 4 g MgSO g NaCl + 1 g NaAc 2 H 2 O g NaAc 3/2 H 2 O Shake automatically Centrifugue 5 min r.p.m. Take 3 ml aliquot Add 750 mg MgSO mg PSA mg C18 Shake Centrifugue Acidify Modified QuEChERS method 2 g sample + 4 ml H 2 O Shake 30 sec, wait 30 min Add 10 ml acetonitrile + TPP Shake Add 4 g MgSO g NaCl + 1 g NaAc 2 H 2 O g NaAc 3/2 H 2 O Shake automatically Centrifugue 5 min r.p.m. Take 3 ml aliquot Add 150 mg CaCl 2 anh mg PSA Shake Centrifugue Acidify AOAC method 5 g sample Add 15 ml acetonitrile Shake Homogenize with Polytron Centrifuge and take the supernatant Re-extractact and combine extracts Concentrated to about 1 ml SPE with TPT Cartridge Evaporate to about 0.5 ml Add IS Evaporate to dryness Make up to 1.5 ml with adecuate solvent Analysis 0.2 g sample/ml Analysis 0.2 g sample/ml Analysis 3.33 g sample/ml

25 LC-TOF-MS TIC of Green tea extracts QuEChERS method Modified QuEChERS with CaCl 2 Almería 23 rd -25 th October 2013 Green tea extracts. CaCl2 vs QuEChERS method. LC-TOF-MS Number of co-extracted matrix compounds CaCl compounds QuEChERS 6008 compounds Mass Profiler Professional 12 1 Agilent Tech

26 Green tea extracts. CaCl2 vs QuEChERS method. LC-TOF-MS Number of co-extracted matrix compounds CaCl compounds QuEChERS 6008 compounds Mass Profiler Professional Agilent Tech. Rt < 6 min Rt 6-12 min Rt > 12 min CaCl compounds QuEChERS 1926 compounds CaCl compounds QuEChERS 3085 compounds CaCl2 889 compounds QuEChERS 997 compounds

27 Green tea extracts. LC-TOF-MS LC-TOF-MS TIC of Green tea extracts QuEChERS method Modified QuEChERS with CaCl 2 AOAC method Methods CaCl 2 AOAC QuEChERs Ʃ Area 8.34E E E+09 Number of co-extracted matrix compounds CaCl compounds 2003 QuEChERS 6008 compounds 363 AOAC 2641 compounds

28 LC-TOF-MS TIC of Green tea extracts Modified QuEChERS with CaCl 2 AOAC method Almería 23 rd -25 th October 2013 Green tea extracts. CaCl2 vs AOAC method. LC-TOF-MS Number of co-extracted matrix compounds CaCl compounds AOAC 2641 compounds 4 th Mass Profiler Professional Agilent Tech. Joint Workshop of the European Union Reference Laboratories for Residues of Pesticides

29 Green tea extracts. CaCl2 vs AOAC method. LC-TOF-MS Number of co-extracted matrix compounds Rt 0-17 min CaCl compounds AOAC 2641 compounds CaCl compounds Mass Profiler Professional Agilent Tech. Rt < 6 min Rt 6-12 min Rt > 12 min AOAC 589 compounds CaCl compounds AOAC 1365 compounds CaCl2 889 compounds AOAC 687 compounds

30 Tomato matrix compounds. LC-TOF-MS Absolute height counts 3304 compounds Dimethoate D6 Rt: 6.38 ± 0.5 min 37 interfering compounds Malathion D10 Rt: 10.5 ± 0.5 min 13 interfering compounds TPP Rt: ± 0.5 min 15 interfering compounds

31 Orange matrix compounds. LC-TOF-MS Absolute height counts 5616 compounds Dimethoate D6 Rt: 6.38 ± 0.5 min 37 interfering compounds Malathion D10 Rt: 10.5 ± 0.5 min 13 interfering compounds TPP Rt: ± 0.5 min 15 interfering compounds

32 Green tea Dil 1/5 matrix compounds. LC-TOF-MS Absolute height counts 4155 compounds Dimethoate D6 Rt: 6.38 ± 0.5 min 37 interfering compounds Malathion D10 Rt: 10.5 ± 0.5 min 13 interfering compounds TPP Rt: ± 0.5 min 15 interfering compounds

33 Data base components- orange matrix compounds. Dilution 1:50 Mass Matrix Nº of compounds Rt: 0-17 min Nº of compounds Rt: 7-13 min TIC Area Rt: 7-13 min Orange E+09 Orange 1: E+09 Orange 1: E+09 Tomato E+09 Peeper E Matrix compounds DB: 750 components Orange: 1082 matrix compounds

34 Sampling efficiency μlc-ms/ms

35 High temperature: 500ºC Nitrogen flow: 40 L/min at 8 kg pressure Gas velocity: m/sec (1000 km/h, almost Match1)

36 μlc-ms/ms Time Ion J. Chem. Phys (1979) Field induced ion evaporation from liquid surfaces at atmospheric pressure

37 μlc-ms/ms Observing Thermal Focusing 2,5 cm Temperature = 550 ºC

38 J. Abian et al. J. Mass Spectrom. 34, (1999)

39 μlc-ms/ms Sampling efficiency

40 μlc-ms/ms Ion

41 μlc-ms/ms Observing Thermal Focusing 2-4 mm Temperature = 550 ºC

42 μlc-ms/ms Sampling efficiency 35 μl/min Atmospheric pressure ion sources. Thomas R. Covey et al. Mass spectrometry Reviews, 2009, 28,

43 μlc-ms/ms Ion Ion Ion Ion Ion

44 Most used I.D. Column (mm) Flow (µl/min) Nebulizer I.D. column (mm) Flow (µl/min) Sensitivity Theoretically Gain Standard LC Standard micro LC Micro

45 Thanks for your Attention

46 Data base components orange matrix compounds. Dilution 1:50 LC-MS/MS Mass Matrix Nº of compounds Rt: 0-17 min Nº of compounds Rt: 7-13 min TIC Area Rt: 7-13 min Orange E+09 Orange 1: E Matrix compounds Orange 1: E+09 Tomato E+09 Peeper E+09 Miner 3D Enterprise DB: 227 components Orange: 1082 matrix compounds

Validation Data for the Analysis of 1-Napthylacetic acid and 1-Naphtylacetamide in tomato and zucchini Using Mini-Luke, Ethyl Acetate and QuEChERS methods Followed by LC-QqQ-MS/MS UNIVERSITY OF ALMERIA

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