Untargeted Screening on the X500R QTOF

Transcription

1 Untargeted Screening on the X500R QTOF R. E. Haufler, Paul Winkler, Andrei Schreiber, SCIEX, Concord, ON Canada NEMC August 2016 RUO-MKT

2 Outline X500R technology and performance X500R QTOF Scan modes Application Examples AB Sciex

3 X500R Technology and Performance

4 Super simplified TOF At t=0, all ions are accelerated to the detector Acceleration is constant energy, all ions achieve the same K.E. Since KE=0.5*mass*velocity 2, velocity is mass dependent, therefore the time to impinge on the detector is mass dependent Ions enter the accelerator high m/z low m/z E E=0 Accelerate Detect AB Sciex

5 Resolution is a key requirement Necessary for mass accuracy, but does not guarantee mass accuracy For differentiation of nearby peaks Helps with sensitivity in TOF intensity ±1ppm K K Shift from centroid (ppm) Causes of resolution loss Jitter timing and voltage Path-length variation detector, mechanical variation Trajectory variation ion position, ion velocity variation AB Sciex

6 Mass Resolution The time-of-flight equation Geometry mirror ground NTOF detector TOF y y 0 2 TOF 2 y y 0 d 2 f ( d1, d3, d6, d8, V1, V 3, V 5) Conditions for 2 nd order focusing dff f ( d1, d3, d6, d8, V1, V 3, V 5) liner ground pulser Y Z 3 high voltages ground Shift from mean TOF (ns) grid 6 Pusher plate 4 v y location in accelerator (mm) X AB Sciex

7 ESI QTOF Geometry Parameter Liner length Mirror Length Mirror Stage 1 length Mirror Stage 2 length Accelerator AC section length Accelerator DC section length Liner potential Mirror Plate potential Value 495mm 102mm 36mm 66mm 12mm 60mm +/-6kV -/+2.1kV Mirror Grid Potential 0 Pusher plate potential Beam energy -/+1.6kV 22eV Effective flight path length 2.35m Actual path length 1.95m AB Sciex

8 Space velocity correlation focusing Correlated position and velocity eliminates turnaround time problem Direction of acceleration Resolution and spectral line shapes in the reflecting time-of-flight mass spectrometer with orthogonally injected ions, V. V. Laiko and A. F. Dodonov, RCMS (1994). Improved resolution and substantially higher sensitivity on a QTOF mass spectrometer, A. Loboda, I. V. Chernushevich, and Nic Bloomfield, 57 th ASMS conference AB Sciex Z=distance from collision cell aperture to point of acceleration into TOF analyzer

9 NTOF Analyzer Field-free zone 2 grids This field is always on Double grid prevents field penetration OFF state This field switches on and off ON state Single pulse only, pusher only, no puller AB Sciex

10 Mass Resolution Detector tilter 32,400 31,600 30,800 30,000 29,200 28,400 27,600 32,400 31,600 30,800 30,000 29,200 28,400 27,600 Resolutio AB Sciex

11 Resolution Performance, MS mode detector m/δm(fwhm) m/z(amu) AB Sciex m/z z Δm(amu) Δt(ps) m/δm

12 Resolution Stability 45,000 40,000 35,000 m/δm(fwhm) 30,000 25,000 20,000 15,000 10,000 5, AB Sciex Hours

13 Mass Accuracy Resolution is nothing without mass accuracy 1 Mass accuracy means confidence in the indicated peak position is accurate and stable. A narrow high resolution peak can be an unreliable indicator of peak position. 1 Bob Haufler, NEMC AB Sciex

14 Mass Accuracy: Thermal effects 5 C change at t=0 The high voltage power supplies are independently thermally stabilized AB Sciex

15 Mass Accuracy - Statistical Each sample was acquired every 20 minutes AB Sciex

16 Mass Accuracy - Systematic Linear calibration equation, 2 coefficients: and 0 mass a t 2 t mass error (ppm) m/z (amu) GluFib MSMS ALILTLVS MSMS TOFCAL MS GluFib=[Glu1]-fibrinopeptide B AB Sciex

17 Sensitivity no beam aperturing AB Sciex

18 X500R Scan Modes and Data Acquisition

19 QTOF scan modes Mode Q1 Collision cell TOF Workflow TOFMS Open, all ions pass Off All ions detected (precursors) Targeted, suspect and unknown screening TOFMSMS Set to target precursor On All fragments detected Targeted, suspect and unknown screening SWATH Scanning 25 amu window On All fragments detected Suspect and unknown screening MRM HR Set to precursor On All fragments detected Targeted quantitation and confirmation IDA Scan mode Time Step 1 TOFMS Survey Scan-Obtain precursor ion list Step 2 TOFMSMS Obtain fragment MS of 1 st precursor Step 3 TOFMSMS Obtain fragment MS of 2 nd precursor Step 4 TOFMSMS Obtain fragment MS of 3 rd precursor sec sec sec sec Total ~1sec Cycles repeatedly during chromatography AB Sciex

20 Untargeted Analysis Known unknowns (suspect screening) spectra exists in library (MSMS and possibly retention time) Library match Formula finder Isotope pattern MSMS pattern Compare retention time if LC method is standard Obtain standard for confirmation and quantitation Unknowns (unknown screening) no spectra exists in library Formula finder Isotope pattern ChemSpider MSMS pattern Obtain standard based on ChemSpider results for confirmation AB Sciex

21 IDA details - Data Acquisition Setup AB Sciex

22 IDA Data AB Sciex

23 IDA Data AB Sciex

24 Sample - Control Comparison in SCIEX OS Software Quick example: unknown screening of food dye compounds Select samples Select control AB Sciex

25 Unknown Screening in SCIEX OS Software Setup: Peak finding Library searching Traffic lights Formula finding Peak detection sensitivity (fast to exhaustive) AB Sciex

26 Results Review and Filtering in SCIEX OS Software Table is Filtered by Formula Finding Results and Sorted by Area Control XIC TOF-MS MS/MS Sample XIC TOF-MS MS/MS AB Sciex Sample was diluted with water/sugar solution (10.000x) to simulate dissolved cake icing

27 Review of Formula Finding Results in SCIEX OS Software Evaluation of TOF-MS and MS/MS Error and ChemSpider Hit Count Formula with 5 Hits Zoom Link to ChemSpider AB Sciex

28 Review of ChemSpider Results in SCIEX OS Software Automatic In-silico Fragmentation of Structure Found in ChemSpider MS/MS AB Sciex Tentative identification of Erythrosine in RED food color

29 Examples

30 Three Isomers Analysis by MRM HR 3 compounds with same formula but separated by LC conditions AB Sciex

31 Sebuthylazine MSMS Spectra AB Sciex

32 Suspect screening: water samples Cocaine human metabolite: Benzoylecgonine Caffeine Sildenafil (viagra) Water samples from lakes, rivers, drinking water and a few other sources around the world AB Sciex

33 Benzoylecgonine in Different Water Samples ? Berlin ng/l Zermatt Canoe Zürich Philadelphia Rio de Janeiro Roma Santa Fe Scarborough Toronto Wien St. Pete Beach Creeks Drinking water Lakes AB Sciex Rivers BE and other drug metabolites are detected in rivers in major cities, concentration can be used to estimate consumption. Sea

34 Caffeine in Different Water Samples ? Philadelphia ng/l Mountain hut in Switzerland Vienna Drinking water Lakes Zürich Berlin Roma Rio de Janeiro Dresden Wien St. Pete Beach Creeks Rivers Sea Caffeine or other compounds consumed in large amounts are markers for human activity, i.e. artificial sweetener AB Sciex

35 Sildenafil in Different Water Samples ng/l Wien Creeks Holy water from a church in Roma Drinking water? Lakes Rio de Janeiro Rivers LOQ Sea Concentration of sildenafil detected were very low (<10 ppt), one exception! AB Sciex

36 Data of Holy Water (Direct Injection 100 µl) Compound µg/l LOQ (µg/l) Acetaminophen Benzoylecgonine (metabolite of cocaine) Caffeine Carbamazepine 0.21 < Codeine Dextromethorphan Diazepam EDDP (metabolite of methadone) Erythromycin Morphine Sildenafil Thiabendazole < AB Sciex

37 Summary The X500R technology and performance Technology to achieve high mass resolution (>30,000 resolution) N-geometry. Detector tilter. Space-velocity correlation focusing. Technology to achieve high mass accuracy(<1ppm accuracy for m/z>50amu) Thermal stabilization. Simplified accelerator with no puller. X500R scan modes IDA for suspect and untargeted screening. SWATH for suspect and untargeted screening. MRM HR for targeted quantification and confirmation. Application examples Unknown food dye compound MRM HR identification of identical m/z isomers Detection of human activity compounds in various water samples AB Sciex

38 Trademarks/Licensing AB Sciex is doing business as SCIEX. For Research Use Only. Not for use in diagnostic procedures AB Sciex. The trademarks mentioned herein are the property of AB Sciex Pte. Ltd. or their respective owners. AB SCIEX is being used under license AB Sciex

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