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Russell Rouseff

mass spectrometry. FOS 6355 Summer 2005. Russell Rouseff. What is Mass Spectroscopy Analytical Chemistry Technique Used to identify and quantify unknown compounds Can also elucidate structure and chemical properties Sensitive technique As little as 10 -12 g can be analyzed

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Russell Rouseff

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  1. mass spectrometry FOS 6355 Summer 2005 Russell Rouseff

  2. What is Mass Spectroscopy • Analytical Chemistry Technique • Used to identify and quantify unknown compounds • Can also elucidate structure and chemical properties • Sensitive technique • As little as 10 -12 g can be analyzed • Identification based on molecular fragmentation patterns • fragments are separated by their m/z ratio

  3. 1899 - Early Mass Spectrometry 1943 - Double focusing analyzer 1946 - Time of Flight MS 1947 - Preparative Mass Spectrometry 1953 - Quadrupole MS 1956 - GC/MS 1956 - Identifying Organic Compounds 1966 - Chemical Ionization 1966 - Peptide Sequencing 1968 - Atmospheric Pressure Ionization 1978 - GC-C-IRMS 1980 - Inductively Coupled Argon Plasma 1990 - Protein Structure 1993 - Protein Mass Mapping/Fingerprinting 1996 - MS of a virus 1996 - First proton transfer MS MS Time lines 1885 1905 1925 1945 1965 1985 2005

  4. Mass Spectroscopy Applications • Detect and identify the use of steroids in athletes • Monitor the breath of patients by anesthesiologists during surgery • Determine the composition of molecular species found in space • Determine whether honey is adulterated with corn syrup • Monitor fermentation processes for the biotechnology industry • Detect dioxins in contaminated fish • Establish the elemental composition of semiconductor materials • Perform forensic analysis – arson identification • Determine exact atomic mass and isotope abundance

  5. Objectives • How did mass spectrometry originate? • What is a mass spectrometer? • What are the characteristics of a mass spectrum? • How is the sample introduced into the mass spectrometer? • How are the molecules and fragment ions produced in the ion source? • How does the analyzer work? • How does the detector work? • What are the roles of computers in mass spectrometry?

  6. Objectives • How can mass spectrometric data be used for structure analysis? • What other techniques are used to produce ions? • How large a molecule can be analyzed? • How is mass spectrometry used with GC,LC, and other separation techniques? • How is mass spectrometry used for quantitative analysis? • What is mass spectrometry/mass spectrometry (MS/MS)? • What is isotope ratio mass spectrometry?

  7. Vacuum pumps Ion Formation Ion Sorting Ion Detection source analyzer ion detection data system Data Handling Sample Introduction Data Output Mass spectrum

  8. Sample Introduction • Neutral molecules in gaseous state • Can be either liquid, solid or gas under STP • must volatilize under vacuum • must be pure • often coupled with GC, HPLC or CE • but can use solids probe if pure

  9. Ionization Techniques • Electron Impact, EI • Chemical Ionization, CI • Fast Atom Bombardment, FAB • Atmospheric Pressure Chemical Ionization, APCI • Proton Transfer Reaction

  10. Electron Impact

  11. - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - Ion Formation (EI) Electron collector Ion focusing lenses - repeller - - + - - + - - - - - - + + + + Sample entrance + - - - - + + - - - filament

  12. CO2+ Molecular ion Base ion O+ CO+ C+

  13. Contains C, H, O

  14. Sample Ionization • Electron Impact, EI, most common • 70 eV standard ionization energy • M + e- => M+ + 2e- • But sometime 70 eV too powerful

  15. EI may overfragment

  16. Chemical Ionization

  17. Chemical Ionization • A softer ionization technique • Use ionized molecules to transfer protons (+) • M + CH5+ => (M+H)+ + CH4 • Chemical Ionization gases • methane • isobutane

  18. Chemical Ionization • Produces spectra with little fragmentation • However, no standard spectral libraries • Spectra tends to be instrument specific • reagent gas pressures, ionization efficiencies • Must generate own standards

  19. Desorption Ionization

  20. Large Molecule Ionization Can routinely analyze molecules of 10,000 Da or more Upper limit used to be 2,000 Da

  21. APCI • A form of chemical ionization • Used for LC/MS interfaces • Allows for removal of solvent before vacuum

  22. http://www.chm.bris.ac.uk/ms/theory/apci-ionisation.html

  23. Proton Transfer Mass Spectrometery glow discharge H30+ + M => (M + H) = H2O M+ H30+ M+ M H30+ Europhysics News (2004) Vol. 35 No. 6

  24. Proton Transfer Mass Spectrometery Produces spectra with little ionization, quantify using M+ Detects aroma compounds at ppb levels Europhysics News (2004) Vol. 35 No. 6

  25. Mass Analyzers

  26. Mass Analyzer Types • Magnetic Sector • Quadrupole mass analyzers • Ion trap • Time of flight

  27. Magnetic Sector Mass Spectrometers e- Molecular Source Ionization chamber H0 Magnetic Field Exit slit detector

  28. Magnetic Sector Disadvantages • Not well-suited for pulsed ionization methods • Very High cost • Large size • High maintence costs

  29. quadrupole mass analyzer detector resonant ion nonresonant ion quadrupole rods source Mass filters focusing lens

  30. Quadrupole Advantages • Classical mass spectra • Good reproducibility • Relatively small/ compact • Relatively low-cost systems • Can improve sensitivity 100x with SIM

  31. Quadrupole limitations • Limited resolution • Peak heights variable as a function of mass (mass discrimination). • Peak height vs. mass response must be 'tuned'. • Not well suited for pulsed ionization methods

  32. Ion Trap Advantages • High sensitivity • Multi-stage mass spectrometry, MSn • Compact mass analyzer

  33. Ion Trap Limitations • Requires careful quantitation • Limited dynamic range • Subject to space charge effects and ion molecule reactions • No sensitivity gain using SIM

  34. Time of Flight MS

  35. MS/MS

  36. General Mass Spectrometry Sites • ASMS- American Society for Mass Spectrometry • http://www.asms.org/whatisms/edu_resources.html • Excellent educational MS resource with many MS related links. • Cambridge University WWW Mass Spectrometry Serverhttp://www-methods.ch.cam.ac.uk/meth/ms/theory/index.html • Comprehensive site with information on ionization processes, mass analyzers, and other mass spectrometry techniques. • Organic Chemistry Online • http://chipo.chem.uic.edu/web1/ocol/spec/MS.htm • Introductory information about analysis of organic compounds by electron impact mass spectrometry, tables of common fragment losses. • University of Leeds:  Introduction to Mass Spectrometryhttp://www.astbury.leeds.ac.uk/Facil/MStut/mstutorial.htm • Introductory site focusing on the analysis of biochemical samples. • Information on Specific Techniques

  37. Misc. Mass Spectrometry Sites Sheffield ChemPuter: Isotope Patterns Calculator http://www.shef.ac.uk/chemistry/chemputer/isotopes.html Site predicts isotope pattern based on molecular formula. UCSF Protein Prospector http://prospector.ucsf.edu/ Site includes useful programs for analyzing the mass spectra of proteins and peptides. Proteomics tools for mining sequence databases in conjunction with Mass Spectrometry experiments

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