CH437 ORGANIC STRUCTURE ANALYSIS
CLASS 5: MASS SPECTROMETRY 5
Synopsis.
Determination of ion formulas (elemental composition) from accurate mass
measurement and from isotope peak intensities. Fragmentation: molecular ions, general
fragmentation modes, metastable ions.
Interpretation of Mass Spectra: Identification of Ions by Formula Mass
The most important information that can be obtained from a mass spectrum
is the formula mass of each ion (this includes relative molecular mass from
the
molecular
ion).
This
information helps
the
chemist to devise
fragmentation pathways that may ultimately give clues about the structure of
an unknown compound.
There are always several molecular formulas possible for a particular formula
mass. For example, if the mass spectrum of an unknown compound reveals a
molecular ion at m/z = 110, the most likely molecular formulas are C 8H14, C7H10O,
C6H6O2 and C6H10N2. These all have nominal masses of 110. There are various
ways of distinguishing amongst these: sometimes information from other areas or
other kinds of spectra can eliminate one or more of these possibilities, but mass
spectrometry can be used alone, as described below.
Determination of Accurate Mass
High-resolution mass spectrometers (such as double focusing BE or FTICR
instruments) can provide mass measurements that are accurate to better than
0.0001 mass unit, making it possible to distinguish between formulas with the
same nominal mass. For example, both C5H12 and C4H8O have formula mass 72,
but the accurate masses differ beyond the decimal point: C5H12 has mass 72.0939
amu and C4H8O has mass 72.0575 amu. A high-resolution instrument can easily
distinguish between these.
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Isotope Peak Intensities
Most elements have less common heavier isotopes (e.g. hydrogen - 2H, carbon –
13C,
nitrogen –
15N,
etc). The peak representing the molecular ion results from
ionization of the molecule containing all the common isotopes (1H,
12C, 15N,
etc).
Peaks at higher m/z values than the molecular ion (usually of much lower
intensity), at M + 1 and M + 2, etc, arise from molecules containing the heavier,
less abundant isotopes.
These isotopic peaks make it possible to determine molecular formulas, without the
need to determine accurate mass. Knowing the relative natural abundance of
isotopes, it is possible to calculate the relative intensity of for each isotopic peak (M
+ 1, M + 2, etc) for a particular molecular formula. The actual calculations are not
dealt with in this course, but the data is tabulated (for example, see handout) and
can be used to match the relative intensities of the M + 1 and M + 2 peaks (etc) of
an unknown ion. An example is shown below.
100
m/z = 44
Calculated intensities of isotopic peaks
Rel. Abundance
/%
?
M
M+1
M +2
100
0.80
0.20
100
1.16
0.40
C2H4O 100
C3H8 100
1.91
0.01
3.37
0.04
N2O
CO2
1.9
44 45
m/z
!
M+1 and M+2 isotopic abundances and exact masses, for CHON ions up to a
nominal mass of 100, are given in a separate hand-out.
Elements with high abundance heavier isotopes are easily spotted in mass
spectra, by the appearance of high intensity peaks at m/z values two units apart
and with intensities matching the relative abundance of the isotopes:
2
Cl
Br
100
100
98
Relative abundance
/%
32.5
35 37
m/z
79 81
m/z
More examples of isotope cluster patterns that are commonly found in the mass
spectra of organic compounds are given below.
The lines are in the unenclosed pictures are 2 m/z units apart. In organic
molecules, carbon (and nearly always hydrogen and often oxygen) will also be
present and their isotopes must be taken into account, as in the enclosed picture,
but usually the isotopic patterns arising from combinations of Br, Cl and S can be
clearly seen in the presence of many carbon and other atoms, as in the two
examples below.
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Fragmentation
Fragmentation is a characteristic feature of many mass spectra and is pronounced
in those spectra that have been achieved by “hard” ionization (EI) of the compound,
but may also be present to a more limited extent in CI, APCI and other “soft”
ionization spectra.
Molecular Ions
Generally, organic molecules ionize by losing an electron from:
(i) the highest energy occupied molecular orbital (HOMO) of multiple-bonded or
aromatic systems.
CH3
CH3
ionization
+
CH3
CH3
CH3
CH3
.
+
.
or
CH3
CH3
CH3
4
(ii) non-bonded orbitals of heteroatoms, such as N, O or S.
..
CH3CH2CH2CH2OH
..
ionization
.
..
+
CH3CH2CH2CH2OH
or
CH3CH2CH2CH2OH
.
+
Stable (high abundance) molecular ions include those of aromatic derivatives,
whereas unstable (low abundance) molecular ions include those of highly
branched or long chain molecules and most alcohols.
Fundamental Fragmentation Processes
The way in which a molecular ion breaks up (called the fragmentation pattern or
scheme) is a function of its structure and hence can be very useful in the
elucidation of structure.
There are three basic fragmentation modes:
(i) Ejection of a radical from a radical ion
(ii) Ejection of a small neutral molecule from a radical ion
These frequently occur via rearrangement.
O
+
.
O
+
.
C
C
OCH3
H
CH2
m/z = 150
-CH3OH
(32)
CH2
m/z = 118
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(iii) Ejection of a small molecule from an ion
O
+
+
C
CH3
CH3
-CO
(28)
m/z = 119
m/z = 91
A generalization of the above is known as the “even electron rule”, to which there
are very few exceptions (e.g. fragmentations of the type, m p+  mf+. + m., are
extremely rare).
Representation of Charged Species
Charged species (radical ions and ions) can be represented by generalized
structures thus:
O
C
+
.
O
+
C
OCH3
or
CH3
CH3
More "precise" structures can be used to aid rationalization of fragmentation:
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Direction of Cleavage in fragmentation Steps
At least one bond cleavage occurs in a fragmentation step, producing a charged
species (ion or radical ion) and a neutral species (radical or molecule). Generally,
the fragment that takes the charge is the one whose structure can best stabilize
that charge: that is, the fragment that has the lowest ionization energy (IE). The
ionization energies of some compounds and radicals are given in the table below.
Compound
IE
Compound
(eV)
IE
Compound IE
(eV)
(eV)
Compound
IE
(eV)
CHCH
11.4
RCOOR’
~10.2 RCONH2
~9.8
I.
10.5
CH2=CH2
10.5
n-ROH
~10.1 RCH=NH
~9.6
SH.
10.4
n-Alkanes
~10.4 RCHO
~9.8
Pyridine
9.3
CH3.
9.8
R2CHCHR’2
~10.2 CH3COCH3 9.7
RCH=NR’
~9.1
CH2=CH.
8.8
C6H12
9.9
ArCOOH
9.7
RCONR’2
~8.8
ROCO.
~8.6
Benzyne
9.7
CH2C=O
9.6
n-RNH2
~8.7
CH3O.
8.6
n-Alkenes
~9.6
R2O
~9.5
Pyrrole
8.2
Ar.
~8.1
Benzene
9.2
ArCOR
~9.4
R2NH
~8.0
CH2=CHCH2.
8.1
RCH=CHR’
~9.1
n-RSH
~9.1
ArNH2
~7.7
HCO.
8.1
1,3-Butadiene
9.1
Thiophene
8.9
n-RF
~12.5 n-R.
~8.0
ArCH3
8.9
Furan
8.9
n-RCl
~10.7 HOCH2.
7.6
Cyclohexene
8.8
ArOH
~8.5
n-RBr
~10.1 Branched
~6.7
alkyl.
– 7.5
ArCH=CH2
8.4
R2S
~8.4
n-RI
~9.2
CH3CO.
7.0
Naphthalene
8.1
ArOR
~8.2
ArCl
~9.1
ROCH2.
~6.9
CO
14.0
CH3SSCH3
7.4
ArBr
~9.0
Cyclic C7H7.
6.2
CO2
13.6
N2
15.6
F.
17.4
R2NCR’2.
~5.4
(R=H or Alkyl)
– 6.1
H2O
12.5
HCN
13.6
Cl.
13.0
H2C=O
10.9
NH3
12.5
Br.
11.8
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For example, examination of the molecule cocaine indicates that the
EI ~ 10 eV
O
CH3
OCH3
N
EI ~ 8 eV
O
IE ~ 9.5 eV
O
tertiary nitrogen group has the lowest IE and hence ionization is most likely to
occur this position. This in turn means that fragmentation will probably be based
upon this radical ion:
CH3
+
N
.
etc
In practice, where there is a reasonable choice of direction of cleavage, both will
occur, but the dominant pathway will be that which gives the more stable charged
species.
Some Empirical Rules Regarding Fragmentation
The Nitrogen Rule
This “rule” helps in the prediction of whether charged species in the fragmentation
pattern are radical ions (odd electron ions, OE+.) or ions (even electron ions EE+),
according to whether there are no nitrogen atoms (or an even number of N) or an
odd number of nitrogen atoms in the species.
If the precursor species has an even m/z value, the situation is straightforward, as
shown below:
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ionization
EEo
EE+.
even mass
even m/z
OE+.
EEo
+
even m/z
EE+
even mass
OE.
+
odd mass
odd m/z
No N atoms
or even number
of N atoms
EE+
EEo
+
odd m/z
even mass
If the charged species has an odd m/z value, the situation is not quite as certain as
above, but a useful scheme can be drawn, as above.
ionization
EEo
EE+.
odd mass
odd m/z
OE+.
+
odd m/z
EE+
even m/z
EEo
even mass
+
OE.
odd mass
Odd number of
N atoms
In practice, a molecular ion and daughter radical ions of odd m/z value are likely to
have an odd number of nitrogen atoms in their structures.
Other Empirical Rules
1. It is impossible to lose CH2 (mass 14) from an ion, because the IE of CH2
(methylene carbene) is too high.
2. It is almost impossible to lose fragments of m/z values between 2 and 15
from ions containing C, H, (O), (N). If a mass loss of 3 occurs in the mass
spectrum of a compound, it must be accompanied by mass losses of 2 and
1. N (mass 14) is not lost from nitrogen-containing ions.
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3. Mass losses between 20 and 26 and also between 36 and 42 are hardly
ever observed.
Common Neutral Losses
Finally, some common neutral losses are given in the table below, along with
possible identities.
Loss
Possibilities
Loss
Possibilities
M–1
H.
M – 32
CH3OH
M – 15
CH3.
M – 33
HS.*
M – 16
O
N- M – 35
(rare,
S*
Cl.*
oxides); NH2.
M – 17
OH.; NH3 (rare)
M – 36
HCl*
M – 18
H2O
M – 42
CH2C=C=O
CH2=CHCH3
M – 19
F.
M – 43
CH3CO.
M – 20
HF
M – 44
CO2
M – 26
HCCH
CN.
M – 45
C2H5O.
M - 27
HCN
H2C=CH2
M – 46
NO2
C3H7.
COOH.
(nitro
compounds
M – 28
CO
M – 29
C2H5.
M – 30
NO
CH2=CH2
HCO.
M – 57
C4H9.
M – 77
C6H5. (phenyl)
(nitro M – 79
C2H5C.=O
Br.*
compounds)
CH2O
M – 31
CH3O.
M – 91
C6H5CH2. (benzyl)
M – 127
I.
* Isotope peak intensity pattern should be checked
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See handout for masses and formulas of some common fragment ions and some
common neutral fragments.
Appearance of a Mass Spectrum
The most common representation of EIMS is as a bar diagram, with % relative
abundance (or relative intensity) plotted against m/z. Two different representations
of the EI mass spectrum of diethyl 2-acetoglutarate are shown below.
The bar diagram is preferred nowadays, but the profile of ion abundances plot
enables the study of small, but significant peaks, such as those of metastable ions
(marked *).
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Metastable Ions
Metastable ions are characteristic of electron ionization: they appear as broad
peaks (usually of low abundance) at non-integral values of m/z, as seen in the BE
mass spectrum above.
Origins
Most ions that reach the detector are produced by very rapid decompositions in the
ion source: within 10-5 s of the ionization event. However, a few fragmentations
require a somewhat longer time and actually occur “in flight”, somewhere in the
field-free region between the ion source and the analyzer. These are called
metastable ions. The ability to detect these ions depends on the analyzer: the
transmission quadrupole (Q) cannot detect them, because the motions of ions in
the analyzer are not dependent on conditions that the ions experience before their
arrival: a product ion will be detected as such, irrespective of whether it was formed
in, or just outside the ion source. Magnetic sector (B) instruments, on the other
hand, are able to detect metastable ions, because when a precursor ion fragments
in the field-free region before reaching the magnetic field of the analyzer, the
fragment ion will lose some kinetic energy to the neutral fragment and so it reaches
the detector via a different trajectory to the “normal” fragment ion. See below.
precursor ion
fragment ion
*
mp
k ~ 104 - 106 s-1
Original KE of mp
mf
+
m
neutral fragment
is partitioned between
mf and m
It does not appear at mf on the m/z scale but at a lower value, m*, given by
m* = mf2/mp.
(m* is typically non-integral)
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The metastable peak is broad because of a narrow range of kinetic energies the
fragment ions actually possess. It is of low abundance because only ions that
decompose in a narrow time “window” (~10-4 – 10-6 s) are detected in this way.
Fragment ions that are produced more rapidly (<~10 -6 s) are recorded as “normal”
fragment ions, whereas ions that are produced by fragmentations in the analyzer
are not focused at a point and hence contribute only to the “background” of the
mass spectrum.
The observation of metastable ions is indicative of a process m p  mf
occurring in one step (i.e. mf is formed directly from mp) and hence is a
valuable aid to structure determination.
On the other hand, the absence of metastable ions does not necessarily mean that
mf is not formed from mp.
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