Chem. Biol. 2011, 18, 1537-1549.

Molecular Structure Elucidation: Mass Spectrometry

Novriyandi Hanif
nhanif@apps.ipb.ac.id

Division of Organic Chemistry

Department of Chemistry
Bogor Agricultural University
MS/U/NYH/2019-1
 Source for Further Reading

1. Clayden, J.; Greeves, N.; Warren, S. Organic Chemistry, 2nd Edition; Oxford University
Press: New York, 2014.
2. Crews, P.; Rodriguez, J.; Jaspars, M. Organic Structure Analysis, 2nd Edition; Oxford
University Press: New York, 2009.
3. Pavia, D. L.; Lampman, G. M.; Kriz, G. S.; Vyvyan, J. R. Introduction to Spectroscopy, 4th
Edition; Brooks & Cole: California, 2009.
4. Lambert, J. B.; Shurvell, H. F.; Lightner, D. A.; Cooks, R. G. Organic Structural
Spectroscopy; Prentice Hall: New Jersey, 2001.
5. Silverstein R. M.; Webster, F. X, Kiemle, D. J. Spectrometric Identification of Organic
Compounds, 7th Edition; John Wiley & Sons, Inc: New Jersey, 2005.

MS/U/NYH/2019-2
 Basic Concepts of Mass Spectrometry (MS)
1. To create ion: a molecule is bombarded with highly energetic electron with an
appropriate method so that it will give a variety of ions → Ionization

2. To separate ion: after ionization of molecule, ions will be separated in electric

field/magnetic field on the basis of their ratio mass to charge (m/z)→ Mass
Analyzer.
3. To detect ion: qualitative and quantitative through relation of m/z and their
abundance → Detector

MS/U/NYH/2019-3
 Objective of Structure Determination via MS

1. To determine molecular weight and molecular formula of organic or

inorganic molecules. The obtained ion molecules should be able to elucidate.
Therefore, we need to undertsand characteristic of molecules and a variety
of ionization methods.
2. To rationalize/elucidate the ms data including molecular ion and fragment
peaks by proposing of fragmentation mechanisms.

Fragmentation mechanisms involve

various organic reactions and concept in Organic Chemistry

MS/U/NYH/2019-4
 Ionization

• Hard Ionization (70 ev)

(Electron Impact/EI, Field Ionization/FI)

• Semi hard Ionization

(Fast Atom Bombardment/FAB)
Increasing E of e-
• Soft Ionization
(Chemical Ionization/CI, Atmospheric-Pressure Chemical
Ionization, Electrospray Ionization/ESI, Matrix-Assisted Laser
Desorption Ionization/MALDI, Field Desorption/FD)

MS/U/NYH/2019-5
 Hard vs Soft Ionization

Hard

The
The fragment ion molecular
ion

Soft

How soft or hard ionization is giving difference implication to structure elucidation?

Keywords: molecular and fragment ions
MS/U/NYH/2019-6
 Hard vs Soft Ionization

Keywords: molecular and fragment ions

• Able to determine the correct molecular ions

• Able to elucidate the fragment ions and confirm them to the corresponding structures
MS/U/NYH/2019-7
Molecular Ions of Various Ionization

MS/U/NYH/2019-8
 Electron Impact MS EIMS
The base peak, m/z =43 M+ or M+.

MW
The fragment ion

MF

The molecular ion

M+, m/z 114

MS/U/NYH/2019-9
 Chemical Ionization MS
CIMS
[M+H]+
The pseudo molecular ion
[M+H]+
Relative Intensity

The fragment ion

MW
The real
molecular ion
(difficult to identify firmly) [M + gas reagent]+

MF
m/z
MS/U/NYH/2019-10
 Independent Exercise 1
1. The EIMS of methylcyclohexane A appears below. Locate the following information in it:
(a) The molecular ion (m/z).
(b) The base peak (m/z).
(c) The M+. − C3H7 (m/z).
83
100

55
80 A

Relative Intensity (%)

41
60

40 27

69
20
98

0
20 40 60 80
m/z
MS/U/NYH/2019-11
 Conversion of Molecular Weight of [M] to
Molecular Formula: The Rule of 13
Ion formed
in various ionization
See previous table

M+. M M/13 = n + r/13 = CnHn+r

Example:
• M+. = 92 ; M = 92 92/13 = 7 n= 7, r = 1 C7H7+1 = C7H8

When an odd amu M+. is encountered,

the first heteroatom to consider is nitrogen (N) or odd multiplies (priority).

When an even amu M+. is encountered,

the N atom(s) also can be considered (not priority)
MS/U/NYH/2019-12
 The Nitrogen Rule
When an odd amu M+. is encountered,
the first heteroatom to consider is nitrogen (N) or odd multiplies.

When an even amu M+. is encountered,

the N atom(s) also can be considered.

MF

m/z

Number of N
Odd Even Odd Even

MS/U/NYH/2019-13
Rule of 13: Heteroatom Equivalents
Element CH Element CH
Equivalent Equivalent
1H C 31P C2H7
12

16O CH4 32S C2H8

14N CH2 16O32S C4
16O14N C2H6 35Cl C2H11
19F CH7 79Br C6H7
28Si C2H4 127I C10H7

MS/U/NYH/2019-14
 Unsaturation Number (UN)
Possible UN Accurate
Formula Mass
C10H16 3 136.1248
C9H12O 4 136.0885
C8H8O2 5 136.0522
C7H4O3 6 136.0159
C9H14N 3.5 136.1123
C8H12N2 4 136.0998

[(2a+2) – (b-d+e)] [(2(8)+2)-(12-2)]

CaHbOcNdXe: UN = C8H12N2 = =4
2 2

MS/U/NYH/2019-15
 Independent Exercise 2
1. The mass spectrum of compound B shows an intense M+. = 108. How many molecular
formula are possible if this compound is oxygen containing? Indicate the unsaturation
number for each formula!
2. The mass spectrum of compound C shows an intense M+. = 93. How many molecular
formula are possible if this compound is oxygen and/or nitrogen containing? Indicate the
unsaturation number for each formula!
3. Molecule D is an amino acid. Un of D is 1. The EIMS of D can be seen in the following
spectrum:
86
100
3.1. Identify m/z the molecular ion and the base peak of D!
3.2. Identify the most probable molecular formula (MF) of
Relative Sensitivity (%)

80

60 30 D through the rule of 13!

74
40

20
131

0
20 40 60 80 100 120 140
m/z MS/U/NYH/2019-16
 Independent Exercise 2
4. Determine the UN of given molecules E, F, and G and confirm your calculation through
analysis of structure!

E F G

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Isotope

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 Fragmentation

ONLY Radical Cation/Radical Anion/Cations/Anions are able

to be detected by MS

MS/U/NYH/2019-19
 Fragmentation

[CH3]+.

[CH3]+.
Cation which will be detected by MS

Which cation will be the most possible detected by MS?

MS/U/NYH/2019-20
 Stevenson’s Rule
• The most probable fragmentation is the one that leaves the positive charge on the fragment with
the lowest ionization energy.
- Fragmentation processes that lead to the formation of more stable ions are favored over
processes that lead to less stable ions

• Cleavages that lead to formation of more stable carbocations are favored.

- Cation stability is more important than radical stability.

• When loss of more than one radical is possible, the largest alkyl radical will be lost
preferentially.

MS/U/NYH/2019-21
 Factor that Impact Fragmentation
• Energetic Factor
- Relative Bond Strength (BDE)

- Stability of the resulting cations or radical ions

• Kinetic Factor
- Availability of cyclic transition state
MS/U/NYH/2019-22
Stability of ions

Octet rule

Delocalization

-Conjugation/
Hyperconjugation

Electronegativity

MS/U/NYH/2019-23
 Independent Exercise 3
1. Molecule H is bombarded with 70 eV electron (EIMS) to give stable fragment ions H2 and
H3. Explain the fragmentation mechanisms of molecule H and draw the structure of radical
cation of H1!
H1

H
H2
H1

H3
2. Molecule I is investigated by EIMS to give fragment ions I1, I2, and I3. Explain the
fragmentation mechanisms of molecule I and draw the structure of radical cation of I1 and
I2!
I1 I2

I
I3
MS/U/NYH/2019-24
 Fragmentation

Initial loss of electron

non-bonding orbital > -orbital > -orbital

• From a non-bonding orbital

MS/U/NYH/2019-25
 Fragmentation
• From a  orbital

or

or

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 Fragmentation
• From a  orbital

or or

MS/U/NYH/2019-27
 Fragmentation
• Homolytic cleavage

• Heterolytic cleavage

MS/U/NYH/2019-28
 Fragmentation
• One bond cleavage: -cleavage

R = alkyl, aryl, H
Y = heteroatom
MS/U/NYH/2019-29
 Fragmentation

• One bond cleavage: -cleavage (cont.)

R = alkyl, aryl, H
Y = heteroatom
MS/U/NYH/2019-30
 Fragmentation
• One bond cleavage: Inductive cleavage

R = alkyl, aryl, H
Y = heteroatom
MS/U/NYH/2019-31
 Fragmentation
• Two bond cleavage: Elimination

R = alkyl, aryl, H
X = heteroatom
MS/U/NYH/2019-32
 Fragmentation
• Rearrangement: McLafferty

MS/U/NYH/2019-33
 Fragmentation
• Rearrangement: Retro Diels-Alder

Abbreviated
Mechanism

Full
Mechanism

MS/U/NYH/2019-34
Electron Count & Fragmentation

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Fragmentation

MS/U/NYH/2019-36
Commonly Lost Fragments

MS/U/NYH/2019-37
Common Fragment Peaks

MS/U/NYH/2019-38
Reporting Mass Spec Data (Low Resolution)

MS/U/NYH/2019-39
Reporting Mass Spec Data (Low Resolution)

MS/U/NYH/2019-40
Reporting Mass Spec Data (High Resolution)

MS/U/NYH/2019-41
Reporting Mass Spec Data (High Resolution)

MS/U/NYH/2019-42
 Straight Chain Alkanes
EIMS

MW = 86

Could you draw the structure

of assigned fragment ions and
propose the fragmentation
mechanism?

MS/U/NYH/2019-43
 Straight Chain Alkanes

The fragment ion

The molecular ion

MS/U/NYH/2019-44
 Branched Alkanes
EIMS


MW = 86

Could you draw the structure

 of assigned fragment ions and
? propose the fragmentation
mechanism?

 

MS/U/NYH/2019-45
 Cycloalkanes
EIMS

MW = 84

Could you draw the structure

of assigned fragment ions and
propose the fragmentation
mechanism?

MS/U/NYH/2019-46
 Cycloalkanes

The fragment ion

The molecular ion

MS/U/NYH/2019-47
 Independent Exercise 4
1. The EIMS of molecule J, 2, 2, 4, 6, 6-pentamethylheptane (MW = 170) and molecule K,
ethylcyclohexane (MW = 112), are shown below. Draw the structure of assigned fragment
ions and the molecular ions! Propose the fragmentation mechanism of each molecule!

57 83
100
100 Molecule J Molecule K

Relative Sensitivity (%)

80 55
Relative Sensitivity (%)

80

60 60

99 40
40
71
20 20
131 112
170
0 0
20 40 60 80 100 120 140 160 180 20 40 60 80 100
m/z m/z

MS/U/NYH/2019-48
 Alkenes
EIMS

MW = 70

Could you draw the structure

of assigned fragment ions and
propose the fragmentation
mechanism?

MS/U/NYH/2019-49
 Alkenes

The fragment ion

The molecular ion

MS/U/NYH/2019-50
 Cyclic Alkenes
EIMS

MW = 136

Could you draw the structure

M (136) of assigned fragment ions and
propose the fragmentation
mechanism?

MS/U/NYH/2019-51
 Cyclic Alkenes

The fragment ion

The molecular ion

MS/U/NYH/2019-52
 Alkynes

EIMS

MW = 68

Could you draw the structure

of assigned fragment ions and
29 propose the fragmentation
M-39
mechanism?

MS/U/NYH/2019-53
 Alkynes
The fragment ion

The molecular ion

MS/U/NYH/2019-54
 Aromatic Hydrocarbons
EIMS

MW = 92

Could you draw the structure

of assigned fragment ions and
propose the fragmentation
mechanism?

MS/U/NYH/2019-55
 Aromatic Hydrocarbons

The fragment ion

The molecular ion

MS/U/NYH/2019-56
 Independent Exercise 5
1. Molecule L, ethyl benzene, has a mass spectrum as follows:
MS (EI, 75 eV): m/z 106 (M+, 40%), 91 (100%), 65 (15%), 39 (8%).
Write reasonable structures for the molecular and the fragment ions!
2. Molecule M, butyl benzene, has a mass spectrum as follows:
MS (EI, 75 eV): m/z 134 (M+, 30%), 92 (60%), 91 (100%)
Write a reasonable structure for the fragment ion at m/z 92 and what is the name of
rearrangement!
3. Molecule N, o-xylene, has a mass spectrum as follows:
MS (EI, 75 eV): m/z 106 (M+, 60%), 105 (30%), 91 (100%)
Propose the fragmentation mechanism and assign the structure of molecular and
fragment ions!
4. Molecule O, norbornene, has a mass spectrum as follows:
MS (EI, 75 eV): m/z 94 (M+, 12%), 66 (100%)
Propose the fragmentation mechanism and assign the structure of molecular and
fragment ions!
MS/U/NYH/2019-57
 Acyclic Alcohols
EIMS

MW = 88

Could you draw the structure

of assigned fragment ions and
propose the fragmentation
mechanism?

MS/U/NYH/2019-58
 Acyclic Alcohols

The fragment ion

The molecular ion

MS/U/NYH/2019-59
 Cyclic Alcohols
EIMS

57

MW = 100

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-60
 Cyclic Alcohols

The fragment ion

The molecular ion

MS/U/NYH/2019-61
 Benzyl Alcohols
EIMS

MW = 108

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-62
 Benzyl Alcohols

The fragment ion

The molecular ion

MS/U/NYH/2019-63
 Phenols
EIMS

MW = 94

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-64
Phenols

The fragment ion

The molecular ion

MS/U/NYH/2019-65
 Independent Exercise 6
1. Elucidate the structure and molecular formula of stable fragment ion m/z = 95 when
molecule P is bombarded with EIMS!

P
2. Elucidate the structure and molecular formula of stable fragment ion m/z = 112 when
molecule Q is bombarded with EIMS!

Q
3. How to distinguish two isomers R and S through EIMS. Give evidence by elucidating their
fragments ions!

R S MS/U/NYH/2019-66
 Independent Exercise 6
4. Molecule T shows an intense fragment ion at m/z 104. The structure of fragment ion is
shown below. Propose the structure of T that has m/z 122 !

5. Propose the fragmentation mechanism of molecule U which shows the following EIMS data:
MS (EI, 75 eV): m/z 60 (M+, 10.3%), 31 (100%), 59 (15.9%), 42 (12.4%).

U
MS/U/NYH/2019-67
 Ethers
EIMS

MW = 130

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-68
 Ethers

The fragment ion

The molecular ion

MS/U/NYH/2019-69
 Aryl Ethers
EIMS

MW = 108

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-70
 Aryl Ethers

The fragment ion

The molecular ion

MS/U/NYH/2019-71
 Amines
EIMS

MW = 87

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-72
 Amines

The fragment ion

The molecular ion

MS/U/NYH/2019-73
 Cyclic Amines
EIMS

MW = 85

Could you draw the

structure of assigned
42 fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-74
 Cyclic Amines

The fragment ion

The molecular ion

MS/U/NYH/2019-75
 Independent Exercise 7
1. Propose the fragmentation mechanism of molecule V and draw strcucture of the following
molecular fragment ions! MS (EI, 75 eV): m/z 85 (M+, 9.3%), 84 (60.2%), 57 (80.3%), 42 (50.2.4%).

V
2. A cationic surfactant W shown below exhibits a peak in the FD mass spectrum at m/z 304.
Collision activated dissociation on m/z 304 gives the fragmentation pattern as follow: m/z 58
(80%), 91 (98%), 134 (35%), 212 (20%), 304 (100%). Draw the structure of each fragment ions
and draw the complete structure of W!

W
MS/U/NYH/2019-76
 Aliphatic Aldehydes
EIMS

MW = 86

Could you draw the

structure of assigned
fragment ions and propose
57 the fragmentation
mechanism?

MS/U/NYH/2019-77
 Aliphatic Aldehydes

The fragment ion

The molecular ion

MS/U/NYH/2019-78
 Aliphatic Ketones
EIMS

MW = 100

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?
57
15

MS/U/NYH/2019-79
Ketones

The fragment ion

The molecular ion

MS/U/NYH/2019-80
 Cyclic Ketones
EIMS

MW = 98

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?
83

MS/U/NYH/2019-81
Cyclic Ketones

The fragment ion

The molecular ion

MS/U/NYH/2019-82
 Aliphatic Carboxylic Acid
EIMS

MW = 88

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-83
 Aliphatic Carboxylic Acid

e-

The fragment ion

The molecular ion

MS/U/NYH/2019-84
 Esters
EIMS

MW = 102

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-85
 Esters

The fragment ion

The molecular ion

MS/U/NYH/2019-86
 Esters
EIMS

MW = 150

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-87
 Esters

The fragment ion

The molecular ion

MS/U/NYH/2019-88
 Amides
EIMS

MW = 101

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-89
 Amides

The fragment ion

The molecular ion

MS/U/NYH/2019-90
 Independent Exercise 8
1. Propose the fragmentation mechanism of molecule X and draw structure of the following
molecular fragment ions! MS (EI, 75 eV): m/z 86 (100%), 74 (46%), 30 (60%)!

2. Propose the fragmentation mechanism of molecule Y and draw structure of the following
molecular fragment ion! MS (EI, 75 eV): m/z 133 (95%)!

Y
MS/U/NYH/2019-91
 Independent Exercise 8
3. A molecule Z contains a carboxylic acid functional group and posesses UN = 5 shown an
intense an EIMS fragment ion at m/z 132 corresponding to the structure of fragment ion as
follows:

m/z = 132
Deduce the structure of Z!
4. EIMS of molecule AB shows a fragement ion at m/z 255. Could you draw the structure of the
fragment ion at m/z 255?

AB
MS/U/NYH/2019-92
 Independent Exercise 8
5. Propose the fragmentation mechanism of molecule AC and draw strcucture of the following
molecular fragment ions! MS (EI, 75 eV): m/z 72 (M+, 22.1%), 57 (8.0%), 43 (100%), 29 (18.8%),
15 (6.6%).

AC
6. The molecule AD after EIMS shows fragment ions at m/z 43, 44, 59. What is the structure of the
fragment ions?

AD

MS/U/NYH/2019-93
 Nitriles
EIMS

MW = 97

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-94
 Nitriles

The fragment ion

The molecular ion

MS/U/NYH/2019-95
 Nitro Compounds
EIMS

MW = 89

Could you draw the

structure of assigned
fragment ions and propose
the fragmentation
mechanism?

MS/U/NYH/2019-96
 Nitro Compounds

The fragment ion

The molecular ion

MS/U/NYH/2019-97
 Alkyl Halides
EIMS

MW = 134

MS/U/NYH/2019-98
Alkyl Halides

The fragment ion

The molecular ion

MS/U/NYH/2019-99
 Independent Exercise 9
1. The molecule AE after EIMS shows the fragment ions at m/z 41, 43. What is the structure of the
fragment ions?

AE

2. The molecule AF shows EIMS fragmentation m/z 85, 93/95, 135/137. What is the structure of
the fragment ions?

AF
3. The molecule AG shows an intense EIMS peak at m/z 93. Draw the structure of the fragment
ion!

AG
MS/U/NYH/2019-100