1. CHM 312 Fall 2008 Special Topics in MS Dr. Ralph Mead

2. Outline Introduction Chemical Ionization Electrospray Ioniszation (ESI) Atmopsheric Pressure Chemical Ionization (APCI) Meta-stable ions MS/MS

3. IONS are Key!!! Hard and Soft Ionization Techniques for the Organic Chemist “Hard” Ionization Electron Impact (usually no M+ but rich in fragments) “Soft” Ionization Chemical Ionization Electrospray APPI

4. Chemical Ionization “ Soft” Alternative to EI. Provides less fragments and usually M+ This is due to the ionization of the reagent gas which transfers as proton and energy to the molecule of interest ( more on this latter) Done in a regular EI source but with a reagent gas present (0.5-2 Torr compared to EI 10 -6 Torr) General Mechanism (GC/MS)

5. Methane Reagent Gas for GC/MS Methane: CH 4 + e -----> CH 4 + · + 2e ------> CH 3 + · + H CH 4 + · + CH 4 -----> CH 5 + · + CH 3 · CH 4 + · + CH 4 -----> C 2 H 5 + + H 2 + H · Acetone with Methane CI:

6. Other Reagent Gases

7. CI GC/MS Spectra (Which Gas is for you?) Simple Answer: Depends… Long Answer: Amount of excess energy imparted on [M+H] + ion upon formation depends on proton affinities of the conjugate base of the reactant ion and M. Proton affinity of conjugate base decreases (i.e. acidity of reactant gas ion increases)…fragmentation of protonated molecules increases since more energy is transferred upon formation Acidity increases from NH 4 + <C 4 H 9 + <C2H 5 +,CH 3 + <H 3 + Degree of fragmentation follows this as seen to the left

8. What about molecules that are not amenable to GC/MS? The trick is making ions in a solvent rich mobile present AND keeping the MS under vacuum

9. Electrospray Ionization Uses:  Small and Large Polar Molecules (Typically proteins and other bio-molecules)  Drugs, pesticides etc. No fragments form…quasi molecular ions…multiple charged fragments [M+ zH] z+ for basic compounds and [M – zH] z- for acidic compounds Charge formation

10. ESI Mass Spectra Positive ion ESI spectrum of protein Horse heart myoglobin Should be one peak for molecular ion but multiple charges since many hydrogens thus multiple peaks. Either positive mode or negative mode ionization Bases as in positive mode as conjugate acids Acids in negative mode as conjugate bases MW???

11. MW Determination from ESI

12. Atmospheric Pressure Chemical Ionization Compared to ESI-LC/MS can accept flow rates up to 2ml/min More suitable for smaller molecules (up to M r =1500) Spectrum are similar to ESI

13. Meta-Stable ions…Putting them to work for you

14. Continued Meta-Stable ions…Putting them to work for you Description

15. MS/MS …Putting it to work for you Instrument schematic and example

16. Continued MS/MS …Putting it to work for you QQQ example:

17. Continued MS/MS …Putting it to work for you