1Fourier Transform Ion Cyclotron Resonance Mass Spectrometry: A PrimerMass Spectrometry Reviews, 1998, 17, 1-35Presented by Peter Abraham and Alex HuffmanSeptember 12, 2002Presentation Outlinen Overview of ICR principlesn FT-ICR MS design and theoryn Capabilities and specificationsn Applicationsn Brief Intermission2Ion Cyclotron Resonance (ICR)n When an ion enters a uniform magnetic field (Bo), it will assume a circular path in the plane perpendicular to Bo.n Angular frequency is the cyclotron frequency (ωc)n ωcindependent of vICRn An ion with m=100 Da will travel ~30 km in a 1 sec. observation period.n Improved signal-to-noise ratios and better resolution3Excitation and Detection of ICR Signaln ICR motion not useful unless we can detect a signaln Phase of ions randomn Radius of of ions too small→ Need spatially coherent ion packetSingle Frequency Excitationn Excitation achieved by applying a uniform electric field oscillating at, or near the ωcof a given ionn ions with same m/z accelerated to same radius4Single Frequency Detectionn “Image” Chargen independent of Bostrengthn increases linearly with rn increases linearly with ion chargeTime Domain Excitation Waveformn Detection signal proportional to ωcn frequency proportional to mass (FT)n average over time to get better signal-to-noise mass resolution5Broadband Excitationn Excitation E creates multiple excitation frequenciesn not good mass resolutionBroadband Excitationn frequency sweepn simple time domainn Stored waveform inverse Fourier transformn simple frequency domain6Broadband DetectionIon-Neutral Collisionsn Collision dampening (F= qE + qv x B –fv)7Axial Confinement: Electrostatic Trapping Potentialn experiments performed in cells, not infinite conductive sheetsn E and B fields hold ions on two axes (x,y)Radial Ion “Magnetron” Rotationn ICR perturbed by of B and r-component of electrostatic trapping potential↓B8Magnetron MotionTrap Configuration9Mass Calibrationn “Internal” calibrationn ions of known m/z inserted with analyte subject to same perturbationMass Resolutionn ωc/dωc= - m/dm n Resolution (resolving power in text):m/∆m50%or ωc/ ωc, 50%n m/∆m50%= -qBo/m∆ωc, 50%n Linearly related to B010Mass Resolution (continued)n At low pressure limit, peak width independent of m/z, but peaks closern ICR frequency varies with (m/z)-1Mass Resolution (continued)n Resolution varies with (m/z)-1n Lower resolution with higher massn Needs high magnetic field strength for good resolution (3-9T)11Mass Limit due to Trap Size(Ideal and Theoretical)n Example: mupper= 5.89 MDa (B0 = 7.0T, singly charged thermal ion, 1” cross-sectional radius)n mupper= 5.598x1011 z2B0r2max/Tn Upper mass limit when IC radius of thermal ion reaches radiusof trapRealistic Mass Limitn With trapping potentialn mcritical= 1.20607x107 zB20a2/(Vtrapα)n If m/z > mcriticaln Ion cyclotron motion isn’t stablen Ion spirals out12Realistic Mass Limitn Example: mcritical/z = 274,000n B0=7.0T, Vtrap=1volt, n cubic trap, a=2.54cm, α=2.77373 (defined by trap shape)How many ions do you want?n Ions can escape axiallyn Retain by increasing Vtrapn Too many ions?n Space charge, ion repulsion ~10,000 ionsn Mass peak broadening, peak coalescence13Internal Ion Sourcesn Leak vapor and ionize internallyn Electron Ionization (EI) - electron beam through center of Penning trapn Chemical Ionization (CI) n Photionization (PI) – laser or arc lamp directed through center of trapn Techniques good for: ion-molecule rxnpathways, ion structure & kinetics, compound identificationExternal Ion Sourcesn Ion source may interact with magnetic fieldn Non volatile analytesn Low Pressure: Laser desorption or MALDIn High Pressure (> 1 torr)n Electrospray Ionization (ESI)n Cluster Sourcesn High Pressure Sources14Chromatographic Couplingn First interface was GC, 1986n Effluent ionized by EIn Liquid Chromatography (LC) – 1991n Capillary Electrophoresis (CE) – 1993n Both electrospray ionization (ESI)n Technique good for biopolymer analysisMS/MS (MSn)n Tandem MS n 1) Ionization and detection of primary ionsn 2) Dissociation (fragmentation) of ionsn Through collisions with neutral speciesn 3) Mass analysis of resulting secondary ions15MS/MS (MSn) (continued)n Techniques for dissociating ions:n Collision-activated dissociation (CAD)n Surface-induced dissociation (SID)n Ultra-violet photodissociation (UVPD)n Multiphoton IR photodissociation (IRMPD)n Blackbody IR dissociation (BIRD)Advantage of High Magnetic FieldLinear Relationship Quadratic Relationship16Fourier Transform AspectsFrequency can be measured much more accuratelyn Resolution, Speed increase x10,000n Mass Range x500n Sensitivity in attomole range (10-18)n Advantage of fixed magnetic fieldApplications 1 - Elemental Compositionn Resolves ~500 singly charged ion masses in 90-300 Da rangen ppm accuracyn Several species at each nominal massn Resolution, m/∆m50% ≈ 100,000172 – Detection Limit for Biological Analysisn Image current detection, ~100 ions per m/z n Less sensitive than somen With ESI can detect single ions (large, high charge) n Sensitivity still be very highn With CE, sub attomole detectionn Unique: Ultrahigh resolution, non-destructive3 – High Mass Applicationsn Electrospray Ionization allows large molecules to acquire many charges –favorable for FT-ICR detectionn Unit resolution achieved on 2 different proteins of 112 kDa each183 – High Mass Applications(continued)Novel combinationsn Combining FT-IRC MS with ion-mobility and TOFn Collisional cross-sectionn V/z ration Ion-molecule mechanisms19FT-ICR MS & Ion Mobility/TOFRev Sci Inst 71(11): 4078-4086 Nov 2000National High Magnetic Field Lab20FT-ICR MS Conclusionn Advantages:n Extremely high resolutionn High speed, large mass, good sensitivityn Disadvantages:n Very expensiven Largen High magnetic fieldn Not best sensitivitySpecial Thanksn Alan G. Marshall Professor of Chemistry and Director, National High Magnetic Field Laboratory ICR Program21Journal Sourcesn Mass Spectrometry Reviews, 1998, 17, 1-35n Biotechnology and Bioengineering (Combinatorial Chemistry), Vol. 71, No 2, 2000/2001n Review of Scientific Instruments, 71 (11): 4078-4086, Nov 2000n European Journal of Mass Spectrometry, 8, 169-176 (2002)Web Sourcesn Alan Marshal Faculty Research Pagen http://www.chem.fsu.edu/faculty/marshall.htmn IonSpec IRMPD Pagen http://www.ionspec.com/irmpd.htmln Influence of Trapping Condition Efficiency of IRMPD in Quadrupole Ion Trapn