1. Mass Spectrometry THE MAIN USE OF MS IN ORG CHEM IS:
DETERMINE THE MOLECULAR MASS OF
ORGANIC COMPOUNDS
DETERMINE THE MOLECULAR FORMULA OF
ORGANIC COMPOUNDS

2. HOW DO WE ACHIEVE THIS?
PERSUADE THE MOLECULE TO ENTER THE VAPOR PHASE (CAN BE DIFFICULT)
PRODUCE IONS FROM THE MOLECULES THAT ENTER
THE GAS PHASE
SEPARATE THE IONS ACCORDING TO THEIR
MASS-TO-CHARGE RATIOS (m/z))
MEASURE AND RECORD THESE IONS

3. What is Mass Spec?
Analytical tool measuring molecular weight (MW) of sample
Only femtomolar concentrations required
Within an accuracy of 0.01% of total weight of sample and within 5 ppm for small organic molecules
For a Mr of 40 kDa, there is a 4 Da error
This means it can detect amino acid substitutions / post-translational modifications

4. What sort of info is returned?
Structural information can be generated
Particularly using tandem mass spectrometers
Fragment sample & analyse products
Useful for peptide & oligonucleotide sequencing
Plus identification of individual compounds in complex mixtures

5. Simplified Schematic
The analyser, detector and ionisation source are under high vacuum to allow unhindered movement of ions
Operation is under complete data system control

6. m/z spectrum example Leucine Enkephalin, Pentapeptide, YGGFL
University of Mons-Hainaut
Leucine Enkephalin, Pentapeptide, YGGFL
Expected Mr = Da, Calculated Mr = Da (dominant ions at 556.1)

7. Mass Spectrometry Ion source Mass analyzer Detector
The general operation of a mass spectrometer is:
create gas-phase ions
separate the ions in space or time based on their mass-to-charge ratio
measure the quantity of ions of each mass-to-charge ratio
The ion separation power of a mass spectrometer is described by the resolution, which is defined as: R = m / delta m, where m is the ion mass and delta m is the difference in mass between two resolvable peaks in a mass spectrum. E.g., a mass spectrometer with a resolution of 1000 can resolve an ion with a m/e of from an ion with an m/e of
Ion source
Mass analyzer
Detector
(EI=Electron Impact; CI=Chemical Ionization; SIMS=Secondary Ions Mass Spec; FAB=Fast Atom Bombardment; LDMS=Laser Desorption Mass Spec; PDMS=Plasma Desorption Mass Spec; TS=Thermospray; AS=Aerospray; ESMS=Electrospray Mass Spec).
(EB=Electrostatic-Magnetic; ion trap; Q=Quadrupole; TOF=Time of Flight.)

8. Sample Introduction & Ionisation
Direct into ionisation source or via chromatography for component separation (HPLC, GC, capillary electrophoresis)
Ionisation can be positively charged (for proteins) or negatively charged (for saccharides and oligonucleotides)

9. Ionisation methods Atmospheric Pressure Chemical Ionisation (APCI)
Chemical Ionisation (CI)
Electron Impact (EI)
Electrospray Ionisation (ESI)
Fast Atom Bombardment (FAB)
Field Desorption / Field Ionisation (FD/FI)
Matrix Assisted Laser Desorption Ionisation (MALDI)
Thermospray Ionisation

12. WHY MALDI?
Less Sensitive to Salts
Lower PRACTICAL detection limits
Easier to interpret spectra (less multiple charges)
Quick and easy
Higher mass detection
Higher Throughput (>1000 samples per hour)

20. Matrix
Needs to be involatile (most are solids at room temperature)
Needs to absorb the laser wavelength that you are using. (most cases 337 nm)
Preferably dissolves in same solvent as the sample
Typically, the matrices are acidic.

22. HOT and Cold Matrices
DHB is a cold matrix, the samples are not as likely to be fragmented, may not ionize some molecules.
Alpha-cyano dihydroxybenzoic acid is considered a hot matrix. More likely to fragment the molecules. Can produce multiply charged proteins.

43. Time-of-Flight Mass Spectrometry (TOF-MS)
A time-of-flight mass spectrometer uses the differences in transit time through a drift region to separate ions of different masses. It operates in a pulsed mode so ions must be produced or extracted in pulses. An electric field accelerates all ions into a field-free drift region with a kinetic energy of qV, where q is the ion charge and V is the applied voltage.
the kinetic energy of an ion leaving the ion source is: 
The ion velocity, v, is the length of the flight path, L , divided by the flight time, t: 

66. - Protein identification strategies Maldi-Tof MS Small proteins
Unknown proteins

75. Parent ions MS/MS Spectrum CID Non-dissociated Parent ions+ + + + F L G K + + F L G K b3 y1 F L G K + +
Parent
ions + + F L G K F L G K
CID b2 y2 + F L G K + + + + F L G K F L G K b1 y3
Non-dissociated
Parent ions
Daughter ions y1 y2 y3 b1 b2 b3 K G L F
MS/MS
Spectrum
Relative Intensity L F G K
m/z