Master Degree in:
Chemistry and
Advanced Chemical Methodologies
Physical Methods in Chemistry
Mass Spectrometry
Summary: acquiring a mass spectrum
Ionization
Mass Sorting (filtering)
Ion
Source
Detection
Ion
Detector
Mass Analyzer
Form ions
(charged molecules)
Sort Ions by Mass (m/z)
Detect ions
100
75
50
Inlet
•
•
•
Solid
Liquid
Vapor
25
0
1330
1340
1350
Mass Spectrum
Best Practice Guide for Generating Mass Spectra
http://www.rsc.org/images/MS2new_tcm18-102519.pdf
Interpretation of mass spectra
• Identification of the Molecular Ion
• Nitrogen rule
• Rule of 13
• Isotopic pattern
• Stevenson rule
Identification of the molecular ion
In order to derive reliable analytical information, it is therefore important to
have some criteria at hand to identify the molecular ion.
1. The molecular ion must be the ion of highest m/z in the mass spectrum
(besides
the corresponding isotopic peaks).
2. It has to be an odd-electron ion, M+•.
3. The peaks at the next lowest m/z must be explicable in terms of reasonable
losses, i.e., of common radicals or molecules. Signals at M–5 to M–14 and at
M–21 to M–25 point towards a different origin of the presumed M+•.
4. Fragment ions may not show isotopic patterns due to elements that are not
present in the presumed molecular ion.
5. No fragment ion may contain a larger number of atoms of any particular
element
than the molecular ion does
Identification of the molecular ion
The EI mass spectrum of 2,5,5-trimethyl-heptane shows no molecular ion
peak. The highest m/z peak is at m/z 113, demanding a nitrogen to be
present; otherwise it must be interpreted as a fragment ion.
• For a molecule with an odd molecular weight most of the fragments
should have an even molecular weight.
• There is an M-14
The “Nitrogen Rule”
•
Molecules containing atoms limited to C,H,O,N,S,X,P of evennumbered molecular weight contain either NO nitrogen or
an even number of N.
•
This is true for radicals as well.
•
Not true for pre-charged, e.g. quats, (rule inverts).
•
In the case of Chemical Ionization, where [M+H]+ is observed,
need to subtract 1, then apply nitrogen rule.
•
Example, if we know a compound is free of nitrogen and gives
an ion at m/z=201, then that peak cannot be the molecular
ion.
The Nitrogen Rule
NH2
A molecule with an odd
number of nitrogen atoms has
an odd molecular weight.
A molecule that contains only
C, H, and O or which has an
even number of nitrogen
atoms has an even molecular
weight.
93
NO2
O2N
NH2
183
O2N
NH2
138
Molecular Weight
One of the first pieces of information we try to obtain when
determining a molecular structure is the molecular formula.
We can gain some information about molecular formula from the
molecular weight.
Mass spectrometry makes it relatively easy to determine molecular
weights.
If necessary use low energy EI or soft ionization (CI, ESI)
Exact Molecular Weight
CH3(CH2)5CH3
Heptane
O
CH3CO
Cyclopropyl acetate
Molecular formula
C7H16
C5H8O2
Molecular weight
100
100
Exact mass
100.1253
100.0524
– Mass spectrometry can measure exact masses.
– Therefore, mass spectrometry can be used to distinguish
between molecular formulas.
Rule of 13
number of C: n = MW/13 (the digits before the decimal point)
number of H = the number of C atoms + 13*digits after the
decimal (n+r)
Once determining the number of carbons and hydrogens,
subtract
for each oxygen: 16 (1C+4H; 16 H)
for each sulfur: 32 (2C+8H; 32 H)
for each nitrogen: 14 (C+2H; 14 H)
Example
Using Exact Mass Measurements
Suppose you determined the exact mass of an ion by mass spectrometry to be
56.0377. Nominal mass 56
How can you figure out all the possible formulas that add to 56?
First use the Rule of 13
Divide the nominal mass by thirteen; the number in front of the decimal is the
number of carbons; multiply the number following the decimal by 13 and add
it to the number of carbons; this equals the number of hydrogens.
a. To add an oxygen: remove a carbon and 4 hydrogens
b. To add a nitrogen: remove a carbon and 2 hydrogens
c. To add a sulfur: remove two carbons, 6 hydrogens; or 2
oxygens
d. …
Example
Mass of 56
56/13 = 4.3077; The number of carbons is n = 4
r = 13*0. 3077 = 4; therefore the number of hydrogens is n + r = 4 + 4
=8
Therefore the hydrocarbon formula is C4H8
Other possible molecular formulas are:
C4H8 - CH4 = C3H4O
C4H8 - CH2 = C3H6N
C4H8 - 2CH4 = C2O2 ;
X
C4H8 - 2CH4 = C2S ;
C4H8 - 2CH2 = C2H4N2
C4H8 - CH4, CH2 = C2H2NO
X
C4H8 - 3CH2 = CH2N3
X
C4H8 - C = C3H20
X
C4H8 - CH4, 2CH2 = CN2O
C4H8 - 4CH2 = N4
Example
The exact mass of the ion was determined to be 56.0377 amu
Nominal mass 56
N4
CN2O
CH2N3
C2O2
C2H2NO
C2H4N2
C3H4O
C3H6N
C4H8
4*14.0031
12.00+2*14.0031+ 15.9949
…
exact mass
56.0124
56.0011
56.0249
55.9898
56.0136
56.0375
56.0262
56.0501
56.0626
Relative Abundance of Isotopes
Isotope
1H
12C
14N
16O
32S
79Br
35Cl
28Si
RA
100
100
100
100
100
100
100
100
Isotope
2H
13C
15N
17O
33S
RA
0.016
1.11
0.38
0.04
0.78
Isotope
RA
18O
0.20
4.40
98
32.5
3.35
34S
81Br
37Cl
29Si
5.1
30Si
Isotopic pattern: the molecular «ion
cluster»
The Nominal mass is
m/z of the lowest
member of the cluster.
This
is
the
isotopomer that has
all the C’s as 12C, all
protons as 1H, all N’s
as 14N, etc.
A rule of thumb, made possible by knowing the isotopic abundance is
that the number of C in a formula is given by:
N= Intensity

M 1 x90.1
M 
63 mm
5
x 90.1  7,15
63
5 mm
Be careful with Br and Cl
Stevenson Rule:
1- fragment ions form by unimolecular processes
2- The most favorable unimolecular processes give rise to the most
fragment ions
3- The most probable fragmentation is the one that implies the lowest
ionization energy (more stable ions favored)
In other words:
- Cleavages that lead to more stable carbocations are favored
- The largest alkyl radical is lost preferentially
Even Electrons Rule:
.+
EE +
+
R
.+
+
N
+
EE
EE +
+
N
+
.+
OE
.+
OE
EE
OE
OE
+
R
.
.
Non-Nitrogen molecular ion
odd-electron ions occur as even mass
EM
.+
EM
.+
OM +
+
R
.+
+
N
+
OM
OM +
+
N
+
.
EE +
OM
EM
+
R
.
.
O
CH3
OE
.+
MW=152
CH3
EE +
MS-FRAG.pdf
http://www.chem.umn.edu/groups/harned/classes/8361/lectures/MSFrag.pdf
Interpretation of mass spectra
• Fragmentations of common classes of
compounds
Alkane Fragmentation
• Long chains give homologous series of m/z = 14 units
(M-15, M-29, M-43, M-57)
• Long chains rarely lose methyl radical
• Straight chain alkanes give primary carbocation
• Branched alkanes have small or absent M+
• Enhanced fragmentation at branch points
+
H3C
H3C
CH3
+
CH3
H3C
CH3
C
Obs. in Mass Spec
+
CH3
Alkanes undergo extensive fragmentation
CH3—CH2—CH2—CH2—CH2—CH2—CH2—CH2—CH2—CH3
Relative
intensity
100
43
80
Decane
57
60
71
40
85
20
0
142
99
20
40
60
80
100
120
Alkanes
CH3
easy
More stable
carbocations
will be more
abundant.
CH3
CH3CH2CH2CHCH3
m/z 86
CH3CH2CH2•
+
+
CHCH3
m/z43
+
•
easy
CH3
CH3• + CH3CH2CH2CH +
more difficult
m/z71
CH3
CH3CH2•
+
+ CH
2CHCH3
m/z57
Cycloalkanes: Loss of side chain, Loss of ethylene fragments
(M-28)
1e-
2e- +
.+
OE
N
.+
OE
Alkenes
• Fairly prominent M+
• Fragment ions of CnH2n+ and CnH2n-1+
• Terminal alkenes lose resonance-stabilized allyl cations (typical
m/z=41)
Cycloalkenes
Prominent molecular ion
Retro Diels-Alder cleavage
+
+
•
•
+
M-28
CH3
+
•
CH3
H3 C
C
H3 C
C
+
•
+
CH2
CH2
Limonene
(m/z 136)
A neutral diene A radical cation
(m/z 68)
(m/z 68)
Alkynes
•
Molecular ion readily visible
•
Terminal alkynes readily lose hydrogen atom (M-1)
•
Terminal alkynes lose propargyl cation if possible
Aromatic Hydrocarbons
• Molecular ion usually strong
• Alkylbenzenes cleave at benzylic carbon (tropylium ion formation)
Aromatic Hydrocarbons
• Alkylbenzenes with a propyl or longer group exhibit a peak at
m/z=92 due to a McLafferty rearrangement
Alcohols




Fragment easily resulting in very small or missing parent ion peak
May lose hydroxyl radical (M+ - 17) or water (M+ - 18)
Commonly lose an alkyl group (the largest) attached to the carbinol
carbon forming an oxonium ion.
1o alcohol usually has prominent peak at m/z = 31 corresponding to
H2C=OH+
Alcohols
43
31
Ethers
•
•
•
•
•
a-cleavage
Inductive (C-O) cleavage (requires stable cation to lose)
Ion rearrangement
M+ weak
Peaks at m/z = 43, 59, 73 etc. M-31, M-45, M-59 etc.
a-cleavage
inductive-cleavage
43
57
29
73
59
87
116
Carbonyl Compounds
•
•
•
•
•
a-cleavage
b-cleavage
McLafferty rearrangement
Aldehydes: M+ weak. M-1; m/z=29; M-29; M-43; m/z=44
Ketones: M+ strong. M-15; M-29; M-43 etc.; m/z=43; m/z=58,72,86 etc.
m/z=42, 83
a-cleavage
Note that an a cleavage
of an aldehyde could
produce a peak at M – 1
by eliminating H atom.
This is useful in
distinguishing between
aldehydes and ketones.
O
+
+
•
O
a-c l e a v a g e
m / z 1 28
+
m / z 43
CH3 •
+
•
O
+
m / z 1 13
b-cleavage
McLafferty
H
O
+
•
M cLaf f e r t y
r ea r r ange m e nt
M ol e cul a r i on
m / z 11 4
H
+
O
+
•
m / z 58
CH3CH2CH2CH2CH2+
resulting from b cleavage.
CH3CO+
resulting from
a cleavage.
CH3CH2CH2CH2CH2CH2CO+
resulting from a cleavage.
Carboxylic Acids
•
•
•
•
•
•
•
M+ weak
a-cleavage to give the ion [CO2H]+ of m/z 45.
McLafferty rearrangement if gH present.
Loss of 44 is the loss of CO2
Loss of water, especially in CI
Fragments at M-17, M-45
Fragments CnH2n-1O2 (especially long chain acids)
Carboxylic Acids
H
McLa ffe rty
re a rra ng e m e n t
+
•
O
OH
a-cle a v a g e
OH
Mo le cu la r io n
m /z 8 8
•
+
O= C-O- H
+
+
•
O
OH
Mo le cula r io n
m /z 8 8
O
H
m /z 6 0
+
m /z 4 5
C3H5O2 m/z=73
Molecular ion
m/z 102
Esters
H
M+ weak
a-cleavage and
McLafferty rearrangement
Methyl esters: M-31
+
•
O
OCH3
a-cleavage
Molecular ion
m/z 102
O
+
m/z 71
O
OCH3
Molecular ion
m/z 102
+
McLafferty
rearrangement
H
+
• OCH 3
OCH3
m/z 59
+
•
O
OCH3
m/z 74
H
+
OCH3
m/z 59
+
•
O
OCH3
m/z 74
O
+
+
+
•
McLafferty
rearrangement
OCH3
Molecular ion
m/z 102
H
+
+
•
O
O
+
Amines
M+ weak or absent
aliphatic amines: a-cleavage
Fragment at m/z=30
Amides
M+ usually present
Primary amides: peak at m/z=44
McLafferty rearrangement if the acyl moiety is long enough
CH3CH2CH2CONH2
59
44
Alkyl Chlorides and Bromides
Strong M+2 peak (M/M+2 - for Cl, 3:1; for Br 1:1)
Loss of Br
Loss of HCl, loss of Cl
Cyclic ions
a-cleavage
No bromine
Bromine
43
85
135
137
164 166
Common Neutral Losses of Diagnostic Value - 1
15
CH3
Alkyl branching if intense peak, otherwise neglect
16
O
Nitroaromatic, oxime, sulfoxide
16
NH2
RCONH2
18
H2O
Alcohol, (ketone, aldehyde, less common)
20
HF
Alkyl fluoride
26
C2H2
Aromatic hydrocarbon
27
HCN
ArCN, N-heterocylic compounds, ArNH2 rarely
27
C2H3
Ethyl ester (low abundance)
28
CO
Quinones, some phenols
28
C2H4
n-Propyl ketones, ethyl esters, ArOC2H5
29
C2H5
Ethyl ketones, Ar - n-C3H7 compounds
30
CH2O
Aromatic methyl esters
31,32
CH3O,CH3OH
Methyl esters of carboxylic acids
33,34
SH, H2S
RSH
Common Neutral Losses of Diagnostic Value - 2
41
C3H5
Propyl ester
42
C3H6
n-butyl ketone
CH2CO
RCOCH3, ArOCOCH3, ArNHCOCH3
43
C3H7
RCOC3H7, Ar-n-C4H9 compounds
44
CO2
Anhydrides, esters
45
COOH
RCOOH
OC2H5
Ethyl esters of carboxylic acids
46
NO2
Aromatic nitrocompounds
48
SO
Aromatic sulfoxide
55
C4H7
Butyl ester of carboxylic acid
56
C4H8
RCOC5H11, ArOC4H9, Ar-C5H11 (n- or i-)
57
C4H9
RCOC4H9
C2H5CO
RCOC2H5
CH3COOH
Acetate
60
Example
74
43
87
59
99 (M-31)
29
31
M (130)
Example
86
58
30
42
44
101/13=7.76923 0.76923x13=10
mass odd, probably one N, C6H15N
C7H17 not possible
101
119
methyl 2-methylbenzoate
91
150
and
methyl 3-methylbenzoate
65
39
119
91
118
39
65
150