1. Chemistry - Option A : Modern Analytical Chemistry
MASS SPECTROMETRY

2. Background and
Overview of
Mass Spectrometer

3. Determining the Structure of an Organic Compound
The analysis of the outcome of a reaction requires that we know the full structure of the products as well as the reactants
In the 19th and early 20th centuries, structures were determined by synthesis and chemical degradation that related compounds to each other
Physical methods now permit structures to be determined directly. We will examine:
mass spectrometry (MS)
infrared (IR) spectroscopy
nuclear magnetic resonance spectroscopy (NMR)
ultraviolet-visible spectroscopy (VIS)

4. Major Types of Spectroscopy
Infrared (IR) spectroscopy measures the bond vibration frequencies in a molecule and is used to determine the functional group.
Mass spectrometry (MS) fragments the molecule and measures the masses.
Nuclear magnetic resonance (NMR) spectroscopy detects signals from hydrogen atoms and can be used to distinguish isomers.
Ultraviolet (UV) spectroscopy uses electron transitions to determine bonding patterns.

5. Mass Spectrometry (MS)
Uses the interaction of electric and/or magnetic fields (i.e. eletromagnetic radiation) with matter to determine weight or mass
Measures mass, not absorption or emission of electromagnetic radiation
Measures molecular weight
Sample vaporized and subjected to bombardment by electrons that remove an electron
Creates a cation-radical
Bonds in cation radicals begin to break (fragment)
Charge to mass ratio is measured

6. MS History
Concept first put into practice by Francis Aston, a physicist working in Cambridge England in 1919
Designed to measure mass of elements (esp. isotopes)
Awarded Nobel Prize in 1922
Now one of the MOST POWERFUL ANALYTIC TOOLS IN CHEMISTRY

7. MS Principles
Different compounds can be uniquely identified by their mass
Butorphanol L-dopa Ethanol N OH HO
-CH2-
-CH2CH-NH2
COOH HO
CH3CH2OH
MW = MW = MW = 46.1

8. MS Principles Find a way to “charge” an atom or molecule (ionization)
Place charged atom or molecule in a magnetic field or subject it to an electric field and measure its speed or radius of curvature relative to its mass-to-charge ratio (mass analyzer)
Detect ions using microchannel plate

9. Mass Spec Principles
Sample + _
Ionizer
Mass Analyzer
Detector

10. Typical Mass Spectrometer

12. Mass Spectrometer

13. The Mass Spectrum
Plot mass of ions (m/z) (x-axis) versus the intensity of the signal (roughly corresponding to the number of ions) (y-axis)
Tallest peak is base peak (100%)
Other peaks listed as the % of that peak
Peak that corresponds to the unfragmented radical cation is parent peak or molecular ion (M+)

14. Typical Mass Spectrum
aspirin

15. Different Types of MS GC-MS - Gas Chromatography MS
separates volatile compounds in gas column and ID’s by mass
LC-MS - Liquid Chromatography MS
separates delicate compounds in HPLC column and ID’s by mass
MS-MS - Tandem Mass Spectrometry
separates compound fragments by magnetic field and ID’s by mass

16. Gas Chromatography
Sample (must contain stable, volatile compounds) is vaporized in a heated chamber
Column is filled with silanized (silicon-coated) calcium silicate
Column is kept hot (400 oC) in oven
Sample is pushed through column using gas pressure (He or N2)

17. Electron Impact MS of CH3OH
Molecular ion
EI Breaks up Molecules in Predictable Ways

18. Electron Impact MS of CH3Br
Isotopes can help in identifying compounds

19. The Mass Spectrum

20. Masses are graphed or tabulated according to their relative abundance.
The Mass Spectrum
Masses are graphed or tabulated according to their relative abundance.

21. MOLECULAR MASS DETERMINATION USING MASS SPECTROMETRY
Mass spectrometry is used to identify unknown or new compounds.
When a molecule is ionized it forms a MOLECULAR ION which can also undergo FRAGMENTATION or RE-ARRANGEMENT to produce particles of smaller mass.
Only particles with a positive charge will be deflected and detected.
The resulting spectrum has many peaks.
The final peak (M+) shows the molecular ion (highest m/z value) and indicates the molecular mass. The rest of the spectrum provides information about the structure.
IONIZATION
MOLECULAR ION
FRAGMENTION
RE-ARRANGEMENT
FRAGMENTION

22. highest m/z value usually corresponds to the molecular ion
THE MASS SPECTRUM
Spectra obtained for organic molecules have many peaks. Each peak is due to a particular fragment with a certain m/z value.
highest m/z value usually corresponds to the molecular ion
its position provides information about the molecular mass of a substance
the tallest peaks come from the most stable species

23. differences between major peaks
THE MASS SPECTRUM
Interpretation of thousands of spectra has shown that many classes of organic compound show characteristic fragmentation patterns due to their functional groups.
It is possible to identify the type of compound from its spectrum by looking at the ...
position of peaks
differences between major peaks

24. . THE MOLECULAR ION Abundance % m/z
In the spectrum of octane, a signal occurs at 114 due to the species C8H18+
The species due to the final signal is known as the molecular ion and is usually corresponds to the molecular mass of the compound.
Abundance %
molecular ion
114 .
m/z

25. . THE MOLECULAR ION Abundance % m/z
The small peak (M+1) at 115 due to the natural abundance (about 1%) of carbon-13. The height of this peak relative to that for the molecular ion depends on the number of carbon atoms in the molecule. The more carbons present, the larger the M+1 peak.
Abundance %
114 .
m/z

26. . FRAGMENTATION Abundance % m/z
The rest of the spectrum provides additional information of the molecule’s structure.
Peaks appear due to characteristic fragments (e.g. 29 due to C2H5+) and differences between two peaks also indicates the loss of certain units (18 for H2O, 28 for CO). 43
Abundance % 29 57 71 85
114 .
m/z

27. FRAGMENTATION PATTERNS
ALKANES
The mass spectra of simple hydrocarbons have peaks at m/z values corresponding to the ions produced by breaking C-C bonds. Peaks can occur at ...
m/z etc CH3+ C2H5+ C3H C4H C5H C6H13+
• the stability of the carbocation formed affects its abundance
• the more stable the cation the higher the peak
• the more alkyl groups attached to the carbocation the more stable it is
most stable tertiary 3° > secondary 2° > primary 1° least stable
alkyl groups are electron releasing and stabilize the cation

28. FRAGMENTATION PATTERNS
HALOGENOALKANES
Multiple peaks occur in the molecular ion region due to different halogen isotopes.
There are two peaks for the molecular ion of C2H5Br, one for the molecule containing the isotope 79Br and the other for the one with the 81Br isotope. Because the two isotopes are of similar abundance, the peaks are of similar height.
m/z
Abundance %
molecular ion contains...79Br Br

29. FRAGMENTATION PATTERNS
Aldehydes and ketones
The position of the carbonyl group influences the fragmentation pattern because the molecular ion fragments either side of the carbonyl group. O
CH3 C C4H9
MOLECULAR ION
has m/z = 100
• +

30. FRAGMENTATION PATTERNS
Aldehydes and ketones
The position of the carbonyl group influences the fragmentation pattern because the molecular ion fragments either side of the carbonyl group. O
CH3 C C4H9
MOLECULAR ION
has m/z = 100
• + O
C4H9 C+
CH3•
Breaking the bond between the methyl group and the carbonyl group produces two possible ions, depending on how the bond breaks.
Two peaks at m/z values 15 and 85 will appear in the mass spectrum.
m/z = 85 O
C4H C•
CH3+
m/z = 15

31. FRAGMENTATION PATTERNS
Aldehydes and ketones
The position of the carbonyl group influences the fragmentation pattern because the molecular ion fragments either side of the carbonyl group. O
CH3 C C4H9
MOLECULAR ION
has m/z = 100
• + O
CH3 C+
Breaking the bond between the butyl group and the carbonyl group produces two further ions, depending on how the bond breaks.
Two peaks at m/z values 43 and 57 will appear in the mass spectrum.
C4H9•
m/z = 43 O
CH3 C•
C4H9+
m/z = 57

32. FRAGMENTATION PATTERNS
Aldehydes and ketones
The position of the carbonyl group influences the fragmentation pattern because the molecular ion fragments either side of the carbonyl group. O
CH3 C C4H9
Example;
MOLECULAR ION
has m/z = 100
• + O
C4H9 C+ O
CH3 C+
CH3•
C4H9•
m/z = 85
m/z = 43 O
C4H C• O
CH3 C•
CH3+
C4H9+
m/z = 15
m/z = 57
A further peak occurs at m/z = 72 (100-28) due to loss of CO

33. Mass Spectra of Alkanes
More stable carbocations will be more abundant.

34. Mass Spectra of Alkenes
Resonance-stabilized cations favored.

35. Mass Spectra of Alcohols
Alcohols usually lose a water molecule.
M+ may not be visible.

36. Molecules with Heteroatoms
Isotopes: present in their usual abundance.
Hydrocarbons contain 1.1% C-13, so there will be a small M+1 peak.
If Br is present, M+2 is equal to M+.
If Cl is present, M+2 is one-third of M+.
If iodine is present, peak at 127, large gap.
If N is present, M+ will be an odd number.
If S is present, M+2 will be 4% of M+.

37. Isotopic Abundance
81Br

38. Mass Spectrum with Sulfur
=>

39. Mass Spectrum with Chlorine

40. Mass Spectrum with Bromine

41. MS Applications
and
Conclusions

42. Applications
Determination or confirmation of chemical structure of drugs and drug metabolites (MS-MS)
Detection/quantitation of impurities
Detection/quantitation of drugs and their metabolites in biofluids and tissues
High throughput drug screening
Analysis of liquid mixtures (LC-MS)

43. MS vs NMR
Absorbance
aspirin
EI-MS NMR

44. MS vs. NMR MS peaks are narrower than NMR peaks
MS is much more (104 x) more sensitive than NMR (among most sensitive tools)
MS generally allows one to analyze much larger molecules (>50 kD) than NMR
MS samples are more difficult to prepare
MS is not particularly quantitative
MS instruments cost a little less than NMR