1. Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao Lecture 2 A Glimpse of Mass Spectrometry Began from 1920s: Aston, Dempster, Thompson MS is often an ideal starting point in organic structure analysis EI-MS of benzamide C 7 H 7 NO M.W. 121

2. What is Mass Spectrometry? A technique used to determine the masses of atoms or molecules in which electrical charges are placed on them and the resulting ions are separated by their mass-to-charge ratios (m/z). A Mass Spectrometer is an instrument that can separate charged atoms or molecules according to their mass-to-charge ratio. Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao

3. Presentation of Data A Low-Resolution EI-MS of n-pentane. Each m/z value is the nominal molecular mass of the fragment (z = +1)

4. C 9 H 14 C 7 H 10 N 2 C 8 H 10 OC7H6O2C7H6O2 C 4 H 10 O 4 C 4 H 10 S Exact mass 122.1096122.0845122.0732122.0368122.0579122.0225 Exact mol. masses can be measured by using a High-Resolution Mass Spectrometer (HR-MS) Species with a nominal mol. mass of 122 amu Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao The fate of a sample molecule in a mass spectrometer Ionization—Molecular Ion (M +· ) Fragmentation and rearrangement—(fragment ions, radicals, neutral species)

5. IonizationAnalyzerDetector Data system Sample introduction Major components of an MS instrument Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao Principles and Instrumentation of MS

6. Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao Schematic of an electron-impact mass spectrometer with a single magnetic sector analyzer

7. Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao + + + + + + + + + + + +- Acceleration V Magnetic field B A single-focusing, 180º sector mass analyzer + m/z 185 173 185 m/z = 185 305 173 Detector A mass spectrum Analyzer tube 100% Base peak m z = B2r2B2r2 2V2V How does it work?

8. Resolution 250.1933 250.1807 m/z Two adjacent peaks with almost equal intensity height valley < 10% 250.1933 R = 250.1933-250.1807 = 20,000 High-resolution mass spectrum (HRMS) can be used to measure exact masses C 16 H 26 O 2 C 15 H 24 NO 2 Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao

9. Why HRMS? Exact mass H 1.0078522C 12.0000000 N 14.0080744O 15.99491502 CH 2 =CH 2 Nominal mass is 28 CO Nominal mass is 28 Exact mass 2  12.0000000 = 24.000000 4  1.00782522 = 4.03130 28.03130 Exact mass 1  12.0000000 = 12.000000 1  15.99491502 = 15.99491502 27.994915 Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao

10. e Anode Cathode M e e e + · Electron Impact/Ionization (EI) EI is a “hard” ionization method One electron is removed to form singly charged molecular ion, M +· M +· carries excess internal energy and tends to fragment Give reproducible spectra but limit to MW < 1,000 Da Pressure < 10 -5 Torr ionization efficiency is high electron energies ~ 20-70 eV Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao

11. M Chemical ionization (CI) Anode Cathode e R + Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao

12. Chemical ionization (CI) CI is especially useful (“soft” ionization) technique when no molecular ion is observed in EI mass spectrum, and also in the case of confirming the mass to charge ratio of the molecular ion. Reagent gas is first ionized by EI. Sample molecules are ionized by interaction with reagent gas ions. Reagent gas : sample molecules ~ 100 : 1 Positive and negative ions are formed in the CI process. However, depending on the instrument setup, only positive ions or negative ions are recorded in the spectrum. Pressure = 0.1 – 2 Torr Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao

13. Chemical ionization (CI) Positive ion mode: G +· + MM +· + Gcharge transfer G-H + + M M-H + + Gproton transfer TiCl 2 + + M(M + TiCl 2 ) + adduct formation Negative ion mode: [G-H] - + M [M-H] - + Gproton transfer M + eM -· electron capture Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao

14. Desorption ionization (DI) Fast atom bombardment (FAB) = sample molecule = solvent molecule Xe Sample solution + + + Ion cluster Multiple desolvation steps + + + + Primary incident beam Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao

15. Desorption ionization (DI) Matrix assisted laser desorption/ionization (MALDI) laser + substrate Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao

16. Electrospray ionization (ESI) needle + ++ + ++ + + + + + + + + + + + + + + + + + Drying gas Single ion in droplet theory: assume that increased charge density, due to solvent evaporation, causes large droplets to divide into smaller droplets eventually leading to single ions. Ion evaporation theory: assumes the increased charge density that results from solvent evaporation causes Coulombic repulsion to overcome the liquid’s surface tension, resulting in a release of ions from droplet surfaces. Sample solution Voltage ~3 – 5 kV Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao

17. Mass analyzer s N + r Magnetic sector Time of flight (TOF) detector V Field free region Acceleration region m/z = 1.93 t 2 V/d 2 d Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao

18. Quadrupole mass filter + + - - ++ Ion trap Ion cyclotron resonance (ICR) Mass Spectrometry Chem 3500 Mass Spectrometry Y. Zhao