质谱技术 - CAS

1 2020 12 8 .. 1 . 1 . 2 . 3 . 4 . 2 . 1 ICP-MS . 2 MALDI . 3 . 4 . 5 . 6 . 7 .. (Mass Spectrometry,MS) .. , . , , . , . ( . ) .. 1886 Goldstein . 1898 Wien . 1912 Thomson , Ne . 1917 . 1918 Arthur Dempster (Electron ionization ) . 1919 Francis Aston ( 50 , ). 1934 Stephens . Herzog Hintenberger .. 1940 (Alfred Nier, father of modern mass spectrometry ) . (235U, 238U,Manhattan Project). 1942 . 1946 Stephens (Time-of flight mass analysis). 1953 Paul Steinwedel (Quadrupole analyzers). 1954 Inghram Hayden Tandem , (MS/MS). 1955 Wiley & Mclarens . 1956 Gohlke and McLafferty - (GC/MS). Beynon (High-resolution MS). 1960 (Nier) . 1965 Hipple , (Ion Cyclotron Resonance). 1966 Munson & Field (Chemical ionization ). 1967 McLafferty & Jennings, (Tandem mass spectrometry).

2 1973 McLafferty - (LC/MS), . 1974 Comisarow Marshall (FT-ICR-MS). 1979 LC/MS . 1981 Barber (FAB MS) 2000 . 1982 LC/MS . 1984 . 1989 John. B. Fenn . Koichi Tanaka . 1990 Katta & Chait . 1993 . 1995 ( ESI MALDI . 1998 (Delay Extract Reflectron . 2000 Alexander Makarov . Glacial landforms can be dated by mass spectrometry, using radioactive isotopes such as 14C and 10Be.. Joseph J. Thomson , 1856-1940 . 1906 .. Francis W. Aston ( , 1877-1945) . 1922 . Hens G. Dohmelt ( , 1922- ) Wolfgang Paul ( , 1913- 1993), 1989 . John. B. Fenn ( , 1917-) (Koichi Tanaka, 1959- . 2002 NMR . , Joseph. J. Thomson . , , . , . , . A.. B.. C.. D.. E.. F.. G.. H.. I.. J.. Thomson . Francis W. Aston .. ( Fenn (Koichi Tanaka Kurt W thrich 1917) 1959) 1938.. NMR.)))

3 , . ESI .. DNA .. , 10 4 ~ 10 5 Pa 10 5 ~ 10 6. Pa .. , , .. Ms. LC Emess Vacuum reduces the chance of an ion colliding with other gaseous molecules or other ions. Higher the vacuum, greater the Mean Free Path. Lower the collision rate, greater the chance of detection. Introduce more ions than gas, but gas carries in the ions, therefore more gas means more ions, but more gas also means lower the vacuum. Vacuum determines Sensitivity Resolution Applied voltages Two Kinds of vacuum pumps are predominantly used in LC-MS. Mechanical or rough pump Turbomolecular (Turbo) pump Vacuum problems, forget mass spectrometry ! . , .. , .. , .. , , , . 50-70eV +. M+ e M + 2e -. M < 1%. + . A +. + . M + . B + R . C +. +. D.. Electron Impact ionization (EI).

4 Chemical ionization (CI). , Field ionization (FI), Field Desorption (FD). Fast Atom Bombardment (FAB). Matrix- assisted laser Desorption ionization (MALDI). Electrospray ionization (ESI). Atmospheric Pressure Chemical ionization (APCI). EI . (M-R2)+. (M-R1)+. (M-R3)+ M+. + + : R1. : R2. : R3. +. : R4. + + :e M .. Matrix- assisted laser Desorption ionization (MALDI). MALDI .. , . , , . , .. MALDI , , . , .. ESI . , .. ESI . Spray of charged microdroplets . 200,00. 0 ESI. 15,000. 1,000. APCI. Molecular FAB. Weight TSP. GC. Non Polar Polar .. m/z , .. (single-focusing). (double-focusing). (quardrupole mass analyzer). (ion trap). (time of flight, TOF). (Fourier transform mass spectrometer) (FT ICR). Orbitrap .. - .. , .. Electron Beam Ion Sample in Beam A C.

5 B. +. , , , , , , GC-MS, LC-MS . - . Time of Fligh MS, TOFMS . TOFMS . 1 2 zV 1/ 2. mv 2. zV v ( ). 2 m m/z , , , .. m/z , , . , . ( t ) , . , t s 10-7s . , , , , . m/z .. Time of Flight (TOF). Developed in 1950's Popularised in 1980's Advantage: no upper mass limit Disadvantage: poor resolution R = 2~5, 000. Time between ion generation & detection Approximately 10-7 seconds Very fast electronics required Very low maintenance It is just an empty tube! . Fourier Transform Mass Spectrometer, FT-MS.. , FT-NMR m/z , B . , , . m/z , .. , . ( v ) , ( r ) , . B , . m/z . FT-MS , . z , z , x , y .. FT-MS , .. FT-MS , , , .. Ion trap . ring electrode .. end cap electrode .. (m/z) .. Penning .. MS 3 . MS . MS .. Orbitrap +. Orb Tra Orbitrap it p Knight-style Kingdon trap ( ideal Kingdon trap'').

6 Orbital traps Kingdon (1923). Proteomics. 2006 Sep;6 Suppl 2:16-21 Mass Spectrometry Reviews, 2008, 27, 661 699. Orbitrap Geometry Dr. Alexander Makarov Anal. Chem. 2000, 72, 1156-1162. Ion Motion in Orbitrap Only an axial frequency does not depend on initial energy, angle, and position of ions, so it can be used for mass analysis The axial oscillation frequency follows the formula k . m/ z w = oscillation frequency k = instrumental const. m/z = . what we want! Makarov, Anal. Chem. 2000, 72: 1156-1162. Makarov et al., Anal. Chem. 2006, 78: 2113-2120. Proteomics. 2006 Sep;6 Suppl 2:16-21. Ions of Different m/z in Orbitrap Proteomics. 2006 Sep;6 Suppl 2:16-21. J. Mass Spectrom. 2005; 40: 430 443.. , . m/z , .. EM. MCP.. , .. (sensitivity).

7 (resolution) . (Mass Range). : 1~600Da, 1~4000Da, 1~10000Da . 1~2000Da, 1~4000Da : 1~300Da : 1~600Da, 800Da : 1~2000Da : 1~10, 000Da, 50 Da (resolution) . R= M/ M, M .. R10%= aM/b M . M . a . b . 5% . FT-ICR TOF R= M/ M M M.. Reported in parts per million (ppm). (Resolution Power, RP) How close is the mass measured by the MS in relation to the theoretically calculated mass? Measured mass: 10%( 5%, 50%), Actual mass: rel. error: 545 = or 177 ppm Therefore the mass accuracy of this instrument is 177 ppm m1 m2. A: B: C: .. m1 m1. RP (1) m1 m2 . m m 2 m1. m (2) m , m . RP . m 28 .. CO 28 N2 28 C2H4 28 , RP=200, , . , . CO/C2H4: (RP)2 = ( ) = 770. N2/C2H4 : (RP)3 = ( ) = 1100. CO/N2 : (RP)1 = ( ) = 2490. 770 , CO/C2H4 . 1100 , CO/C2H4 N2/C2H4.

8 2500 , . 2000 10000 . FT-ICR-MS 2 .. , .. , . ( ), , . , , , , , .. , , . , . , , , .. , , .. GC/MS . HEWLETT 5972A Mass Selective PACKAR Detector D. DEG/MI. N. HEWLETT. PACKARD. 5890. Samples Chromatograph (GC or LC) Mass Spectrometer (MS). B. D A. A C B. D C. C. A B. D. Sample Separation Identification . API - Ion Trap Interface (Esquire-LC).. Thermo Electron Corp. 1956 George N. Waters Associate Ltd. 1958 Jim Waters Hatsopoulos MAT, 1947, Ludolf Jenckel, Bremen Vacuum Generator (VG),1962, Bernard Eastwell, Manchester Finnigan, 1967, Robert E. Finnigan, Roger Sant, , San Jose VG Micromass ,1969, Eastwell & R D Craig 1991, Acquired by Fisons Instruments 1981, Finnigan MAT. 1996, Micromass Limited MBO. 1990, Thermo Finnigan 1997, Acquired by Waters 2002, Merger of Operation, Waters Micromass VG OA-TOF hybrid instrument (1995).

9 Thermo Fisher . Orbitrap . Waters Qtof . Agilent AB Sciex . Qtof . Shimazu /Bruker MALDI-TOF, .. Bruker FT-ICR - . MRMS . 2 . 1 ICP-MS . 2 MALDI . 3 . 4 . 5 . 6 . 7 . ICP-MS .. ICP 7000K . CyTOF2 Mass Cytometer . /10184633?fr=aladdin Mass Cytometry .. MALDI-TOF . The Principles Intens.[ ]. 6000 Polymer analysis 3 7 4 0 .5 7 5. 5000 3 8 7 2 .4 7 4. 3 5 6 4 .6 6 4. 3 9 6 0 .3 9 5. 3 5 1 9 .6 7 2. 4 0 4 8 .3 1 0. 3 4 7 5 .6 8 6. 4000. 3 4 3 1 .6 8 9. 4 0 9 1 .2 4 4. 3 3 8 7 .7 0 1. 4 0 9 2 .2 4 0. 3 3 4 3 .6 8 8. 3000 4 1 8 0 .1 1 3. 3 2 9 9 .6 7 1. 3 2 5 6 .6 9 1 4 1 8 1 .1 0 4. 3 2 1 1 .6 6 6. 4 2 2 4 .0 5 2. 2000. 3 1 6 7 .6 9 0. 4 3 1 1 .8 9 7. 3 0 7 9 .6 4 7. 4 3 1 2 .9 0 0. 3 0 3 5 .6 4 1 4 3 9 9 .7 3 0. 1000. 4 4 4 3 .6 1 9. 2 9 9 1.

10 6 1 3. 4 4 4 4 .6 2 3. 2 9 0 3 .6 0 0. 4 5 7 5 .2 9 4. 2 8 1 5 .5 6 8. 2 4 1 8 .3 5 3 2 6 3 9 .4 4 2. 0. 2500 3000 3500 4000 4500 5000. m /z matrix The Application x10 5. Intens.[ ]. x10 5. Intens.[ ]. Intact protein analysis Peptide mapping analysis 20000 40000 60000 80000 100000 120000 140000 160000 180000 200000 1000 1500 2000 2500 3000. m /z m /z MALDI . The Principles Applicable Instruments Imaging Workflow . Cross-linking . CXMS Cross-linking with Mass Spectrometry .. HDX-MS .. - .. (D2O) . (D) .. (H) .. Leap . Orbitrap Fusion HSP90 N Radicicol . HDX-MS . 1 - . 2 - . 3 . 4 . HSP90N (Radicicol) HDX-MS . HSP90N HSP90N+. Radicicol 61-72 110-119 160-177. HSP90N. HSP90N+. Radicicol .. LC-MS. GC-MS.. GC-MS .. Full scan . SIM .. H .. LC-MS.