Triumphs and Challenges of High-Resolution Mass Spectrometry in Comprehensive Residue Screens

Transcription

1 Triumphs and Challenges of High-Resolution Mass Spectrometry in Comprehensive Residue Screens Florida Department of Agriculture Mark Crosswhite, Ph.D.

2 outline ur Goal identify and quantify quickly and reliably UHPLC v HPLC easy and cost effective improvement ur Two Platforms rbitrap and Triple Quad advantages of high resolution MS on the Exactive high resolution and high mass accuracy allows for post acquisition data mining limitations of high resolution MS on the Exactive cannot separate compounds with same exact masses no mass filtering fragments can be produced from any ions in the cell Resolution elucidate fragment formulas and structures use trusted triple quad fragments even though their identities are often unknown Exactive Plus vs. ABSciex real samples detection quantitation false positive realized Current and Future Strategies

3 outline ur Goal identify and quantify quickly and reliably UHPLC v HPLC easy and cost effective improvement ur Two Platforms rbitrap and Triple Quad advantages of high resolution MS on the Exactive high resolution and high mass accuracy allows for post acquisition data mining limitations of high resolution MS on the Exactive cannot separate compounds with same exact masses no mass filtering fragments can be produced from any ions in the cell Resolution elucidate fragment formulas and structures use trusted triple quad fragments even though their identities are often unknown Exactive Plus vs. ABSciex real samples detection quantitation false positive realized Current and Future Strategies

4 UHPLC Capacities of orthogonal analyses are multiplicative, not additive. Especially helpful with duty cycle issues in MRM MS (triple quad) and high resolution MS where there may not be MS/MS capabilities. propazine vs sebuthylazine m = Dimethomorph (E and Z) Acquity UPLC BEH C18 Cl Cl 1.7 µm 2.1 x 150 mm Flow rate.200 ml/min UHPLC HPLC UHPLC Time (min) Time (min) Time (min)

5 outline ur Goal identify and quantify quickly and reliably UHPLC v HPLC easy and cost effective improvement ur Two Platforms rbitrap and Triple Quad advantages of high resolution MS on the Exactive high resolution and high mass accuracy allows for post acquisition data mining limitations of high resolution MS on the Exactive cannot separate compounds with same exact masses no mass filtering fragments can be produced from any ions in the cell Resolution elucidate fragment formulas and structures use trusted triple quad fragments even though their identities are often unknown Exactive Plus vs. ABSciex real samples detection quantitation false positive realized Current and Future Strategies

6 representation of quadrupole in ms/ms mode Q1 Q2* Q3 mass filters fragments mass measures ion optics representation of rbitrap C-trap HCD cell* Sees what it looks for (MRM) ω=(qk/m) (1/2) orbitrap rbitrap sees everything all the time (full scan) Allows postacquisition data mining * fragments generated by colliding analyte with neutral gas

7 outline ur Goal identify and quantify quickly and reliably UHPLC v HPLC easy and cost effective improvement ur Two Platforms rbitrap and Triple Quad Advantages of High Resolution MS on the Exactive high resolution and high mass accuracy allows for post acquisition data mining limitations of high resolution MS on the Exactive cannot separate compounds with same exact masses no mass filtering fragments can be produced from any ions in the cell Resolution elucidate fragment formulas and structures use trusted triple quad fragments even though their identities are often unknown Exactive Plus vs. ABSciex real samples detection quantitation false positive realized Current and Future Strategies

8 what high resolution MS will do Phosmet [M + H + ] + = RT = min required resolution = m/(δm) Azinphos methyl [M + H + ] + = RT = min S S P S S P Triazophos [M + H + ] + = RT = min required resolution = Hexaconazole [M + H + ] + = RT = min Cl S P H Cl

9 resolving power simazine ( ) carbaryl ( ) RP required = thiabendazole ( ) resolvable with new rbitrap RP required = 4698 carbaryl ( ) m/z m/z

10 m/z m/z simulation v observed of a 1:10 mixture triazophos [M + H + ] + = hexaconazole [M + H + ] + = simulation resolution observed , , ,

11 outline ur Goal identify and quantify quickly and reliably UHPLC v HPLC easy and cost effective improvement ur Two Platforms rbitrap and Triple Quad advantages of high resolution MS on the Exactive rbitrap high resolution and high mass accuracy allows for post acquisition data mining limitations of high resolution MS on the Exactive cannot separate compounds with same exact masses no mass filtering fragments can be produced from any ions in the cell Resolution elucidate fragment formulas and structures use trusted triple quad fragments even though their identities are often unknown Exactive Plus vs. ABSciex real samples detection quantitation false positive realized Current and Future Strategies

12 what high resolution MS alone will not do Secbumeton [M + H + ] + = RT 8.23 min Prometon [M + H + ] + = RT 8.21 min Terbumeton [M + H + ] + = RT 8.32 min Prometryn [M + H + ] + = RT min S Terbutryn [M + H + ] + = RT min S

13 outline ur Goal identify and quantify quickly and reliably UHPLC v HPLC easy and cost effective improvement ur Two Platforms rbitrap and Triple Quad advantages of high resolution MS on the Exactive rbitrap high resolution and high mass accuracy allows for post acquisition data mining limitations of high resolution MS on the Exactive cannot separate compounds with same exact masses no mass filtering fragments can be produced from any ions in the cell Resolution elucidate fragment formulas and structures use trusted triple quad fragments even though their identities are often unknown Exactive Plus vs. ABSciex real samples detection quantitation false positive realized Current and Future Strategies

14 Extracted Ion Chromatogram (upper) Mass Spectra (lower) 8.21 PRMET STADARD Θ * Time (min) V Collision Energy 15 V Collision Energy Θ V Collision Energy * m/z

15 Extracted Ion Chromatogram (upper) Mass Spectra (lower) 8.32 TERBUMET STADARD Θ * Time (min) V Collision Energy 15 V Collision Energy Θ V Collision Energy * m/z

16 trusted triple quad fragments real detect.040 ug/g ABSciex 5500 azoxystrobin frag intensity (arb) azoxystrobin frag ug/g 0 V collision energy 15 V collision energy 45 V collision energy Exactive Plus Time extracted ion chromatogram azoxystrobin + H + extracted ion chromatogram mass = extracted ion chromatogram mass =

17 outline ur Goal identify and quantify quickly and reliably UHPLC v HPLC easy and cost effective improvement ur Two Platforms rbitrap and Triple Quad advantages of high resolution MS on the Exactive rbitrap high resolution and high mass accuracy allows for post acquisition data mining limitations of high resolution MS on the Exactive cannot separate compounds with same exact masses no mass filtering fragments can be produced from any ions in the cell Resolution elucidate fragment formulas and structures use trusted triple quad fragments even though their identities are often unknown Exactive Plus vs. ABSciex real samples detection quantitation false positive realized Current and Future Strategies

18 B 3688 compound matrix ABSciex 5500 quantitation (μg/g) Exactive plus quantitation (μg/g) percent difference exact mass (H+ adduct, Da) measured mass (Da) mass error (ppm) azoxystrobin eggplant pyraclostrobin blueberry dimethoate blueberry thiabendazole pear pyrimethanil strawberry flonicamid strawberry fluopicolide onion azoxystrobin onion thiabendazole apple imazalil banana

19 B 3704 compound matrix ABSciex 5500 quantitation (μg/g) Exactive plus quantitation (μg/g) percent difference exact mass (H+ adduct, Da) measured mass (Da) mass error (ppm) methamidophos collards * acephate collards dinotefuran collards thiabendazole papaya chlorantraniliprole collards chlorantraniliprole peas azoxystrobin papaya boscalid blackberries boscalid papaya * possible suppression in source or interference in mass measurement

20 B 3706 compound matrix ABSciex Exactive 5500 plus quantitation quantitation (μg/g) (μg/g) percent difference exact mass (H+ adduct, Da) measured mass (Da) mass error (ppm) methamidophos acephate bell peppers bell peppers * carbendazim okra azoxystrobin bell peppers azoxystrobin bell peppers boscalid tomato methoxyfenozide bell peppers bifenazate bell peppers difenoconazole bell peppers * remember the extracted ion chromatograms using trusted triple quad fragments

21 B 3707 compound matrix ABSciex Exactive 5500 plus quantitation quantitation (μg/g) (μg/g) percent difference exact mass (H+ adduct, Da) measured mass (Da) mass error (ppm) phorate sulfoxide potatoe trifloxystrobin strawberry pyrimethanil strawberry novaluron strawberry propamocarb squash pyraclostrobin bell pepper pyraclostrobin tomato pyraclostrobin strawberry penthiopyrad bean propamocarb cucumber

22 propoxur (internal control) from B 3707 measured mass of propoxur +H + ppm error average error

23 summary information of real samples f ~ 125 samples screened for ~ 250 pesticides per sample on the Exactive Plus we found: no false positives when compared to ms/ms analysis on an ABSciex 5500 (everything confirmed) no false negatives when compared to an ABSciex 5500 (Exactive Plus did not miss anything at the established LDs) the largest percent difference and average percent difference in quantitation between the Exactive Plus and the ABSciex 5500 was 60 and 4 respectively. one false positive on the ABSciex 5500 (not their fault)

24 real sample false positive realized 302.1/97.2 fenhexamid B3716_R1_01_57782 intensity (arb) 302.1/55.0 fenhexamid B3716_R1_01_ E E 6 extracted ion chromatogram of fenhexamid +H + on Exactive Plus B 3716_R1_01_57782 Time

25 outline ur Goal identify and quantify quickly and reliably UHPLC v HPLC easy and cost effective improvement ur Two Platforms rbitrap and Triple Quad advantages of high resolution MS on the Exactive rbitrap high resolution and high mass accuracy allows for post acquisition data mining limitations of high resolution MS on the Exactive cannot separate compounds with same exact masses no mass filtering fragments can be produced from any ions in the cell Resolution elucidate fragment formulas and structures use trusted triple quad fragments even though their identities are often unknown Exactive Plus vs. ABSciex real samples detection quantitation false positive realized Current and Future Strategies

26 our current strategy and flow if no detections screen with Exactive Plus for ~250 compounds if analyte found above LD report report using ABSciex 5500 identification and quantitation using ms/ms, ion ratios and a 6 point calibration curve

27 our next strategy and flow if analyte found at <.5 tolerance screen with Exactive Plus for ~500 compounds if analyte found at >.5 tolerance report report using ABSciex 5500 estimate and characterization- 1 point calibration standard, and characteristic fragments for confirmation identification and quantitation using ms/ms, ion ratios and a 6 point calibration curve

28 our following strategy and flow if analyte found at < tolerance screen with Exactive Plus for ~750 compounds if analyte found at > tolerance Exactive Plus report using ABSciex 5500 quantitation and characterization- 6 point calibration curve, and characteristic fragments for confirmation identification and quantitation using ms/ms, ion ratios and a 6 point calibration curve

29 our ideal strategy and flow routine screen with Exactive Plus for ~1000 compounds if needed/special case Exactive Plus report using ABSciex 5500 quantitation and characterization- 6 point calibration curve, and characteristic fragments for confirmation identification and quantitation using ms/ms, ion ratios and a 6 point calibration curve

30 acknowledgements Walter Hammack Ghislain Gerard Casey Burrows Erin McAllister Jon Pelt