In MS, compounds are ionized, ionized molecule

fragments into smaller ions/radicals.

The positively charged fragments produced are
separated based on their nominal_mass/charge
(m/z) ratio.
Mass Spectroscopy ionization fragmentation
M M+. M+1 + M+.2+..+N1+ N2. + …..
Parent ion daughter ions, radicals, neutral

Most of the ions has z=+1; m/z = mass of the fragment.

A plot of relative abundance vs m/z of all charged
particles is presented as the MS spectrum.

Base peak Fragmentation:

M 1 + + N1 .
Molecular
ion M+. M+. M+ M1+ +N
M2+. + N2

M+. Radical ion (odd e)

N. Neutral radical (odd e)
N Neutral (even e)
Nominal mass
M+ (even e) would not break up into a radical ion….
Spectrum presented as a bar graph.

Isotope peaks:
In mass spectroscopy mass of actual fragments
generated are determined. Therefore fragments with
different isotopes are distinguished, e.g. Lead metal.

Actual signal
has peaks with
a line width.
For molecular fragments, the isotope peak abundance
is dependent on the molecular constitution and the
natural isotope abundance of the constituent elements.

1
In mass spectroscopy the masses of individual ‘ions’
are measured.

Mass and ‘abundance’ of each isotopic composition

is measured!! – not the average molecular mass.

Isotope peaks

Mass Spectrometer:

Sample Introduction
Create gas-phase ions of sample
Separate ions in space or time basis based on m/z ratio
accomplished by mass analyzers.
Detect of the quantity of ions of each m/z ratio

Cl-CH2-CH2-S-CH2-CH2-Cl

Ionization: Mass Analyzers:

Magnetic Sector Mass Analyzer (Single/Double Focusing
Electron Ionization (Electron Impact, EI) Tandem Mass Spectrometry
Electrospray Quadrupole
Matrix Assisted Laser Desorption Ionization (MALDI) Quadrupole Ion Trap
Atmospheric Pressure Chemical Ionization (APCI) Fourier-Transform Mass Spectrometry (FTMS)
Fast Atom Bombardment (FAB) Time-of-flight (TOF)
Chemical Ionization (CI)
Inductively Coupled Plasma (ICP)
Ion Detection:
Faraday Cup
Electron Multiplier
Photomultiplier Conversion Dynode

2
Electron impact ionization
Magnetic sector separation Electron impact ionization (EI)
single focusing

Mass Spectrometer
70V
- +

V
Ion optics
Ekin = zV = mv2/2
Evacuated
System 70eV – high energy electrons,
- 10-6 torr molecular ion - very energetic, low/no abundance.

Volatilized compound is ionized by electron impact.

An electron beam is generated by a accelerating the
electrons from a heated filament through an applied
voltage.
The electron energy is defined by the potential
difference between the filament and the source
housing and is usually set to 70 eV (~1.12x10-17J).
All ions are subsequently accelerated out of the ion
source by an electric field produced by the potential
difference applied to the ion source and a grounded
Electrode, V.
A 'repeller' serves to define the field within the ion
source.

A field keeps the electron beam focused across the
ion source and onto a trap (collimating magnets).
Upon impact with a 70 eV electron, the gaseous
molecule may lose one of its electrons to become a
positively charged radical ion, daughter ions, etc.
Depending on the lifetime of the excited state,
fragmentation will either take place in the ion source
giving rise to stable fragment ions, or on the way to
the detector, producing metastable ions.

Magnetic sector mass analyzer:

Each m/z beam follows it’s own path (r) for a
given B and V in the magnetic sector (60o/900).
V
B
m/z = (eB2r2)/(2V)
r
note: slits For specific V and B ions of unique m/z pass thro’
Ion source accelerates ions to a KE the magnetic sector and reaches the stationary
detector. Variations of V and/or B causes fragments
KE = ½ mv2 = zeV
of different m/z value to reach the detector.
In the magnet F = mv2 /r = Bzev,

Upon rearrangement r = mv/zeB = (2Vm/ze)1/2/B

Usually B is scanned to allow different m/z’s to
reach the detector sequentially generating the
m/z = (eB2r2)/(2V) complete mass spectrum.

3
Magnetic Sector Mass Analyzer: Double Focusing (EB)

B
+
E
-

mv 2 mv 2
zeE = and zeV=
r 2
The distribution of a given mass by way of energy
2V m
r= ; r independent of in E; focussing! distribution of kinetic energy refined.
E z

Resolving power 1.
The m/z ratio of the ions that reach the detector can
be varied by scanning either the magnetic field (B)
or the applied voltage of the ion optics (V).

i.e. by varying the voltage or magnetic field

of the magnetic-sector analyzer, the individual ion
beams are separable spatially, radius of curvature Actual signal
Is held constant. has peaks with a
line width.

Imposes a limitation 10%

on the resolvability
of consecutive
peaks

2.

State the method of

calculation when
expressing resolving
power, and the
position of the lower
peak.

4
 Ionization:
Electrons in molecules occupy molecular orbitals
and hence acquire the energy associated with
such orbitals. To remove electrons from such
orbitals and ionize the molecule energy is required.

The energy required depends on the orbital of electron

occupation namely the HOMO. Thus the ease of
ionization will depend on the “types of electrons” in
the molecule.

The molecular ion (dominant) is formed by the

removal of the least tightly bound electron.

Chemical ionization (CI):

Interaction of the molecule M with a reactive ionized

reagent species (gaseous Bronsted acids). E.g..,
EI of methane, generates CH4+· which then reacts to
M+. nearly nonexistent.
give the Bronsted acid CH5+;

CH4+· + CH4 → CH5+ + CH3·

If M in the source has a higher proton affinity than CH4,

the protonated species MH+ will be formed by the
exothermic reaction.
M + CH5+ → MH+ + CH4
Abundant M+.
CI is a softer ionization process.

Fast Atom Bombardment Ionization:

The sample droplet is bombarded with energetic atoms (Ar, Xe) of 8-10
keV kinetic energy. Ions (e.g., Cs+) can be used as the bombarding
particle in a similar technique termed liquid secondary ion mass
spectrometry (LSIMS)
Beam collides with the sample and matrix molecules, producing positive
and negative sample-related ions that can be accelerated into the mass
spectrometer.

5
 Quadrupole Mass Analyzer (spectrometer):
Fast Atom Bombardment
Used for polar organic compounds, acidic and
basic functional groups.
Basic groups run well in positive ionization 4 parallel, polished metal rods
mode and acidic groups run well in negative
+[U+Vcos(ωt)] y
ionization mode.
FAB analytes: peptides, proteins, fatty acids, - [U+Vcosωt] z
x
organometallics, surfactants, carbohydrates,
antibiotics, and gangliosides.
Diagonal electrodes have potentials of the same sign
U= DC voltage, V=AC voltage, ω= angular velocity of
alternating voltage

Quadrupole Mass Analyzer (Spectrometer): The solution of equations of motion of ions traveling
through a QM analyzer shows that for an ion with a
Ions oscillate under the influence of the variable particular m/z to pass through, certain combinations
fields. of U and V must be obtained.

Combined DC and RF potentials on the quadrupole Varying rod voltages (scanning the spectrum):
rods passes only a selected m/z ratio (resonant ion) at
a time. All other ions acquire unstable trajectories a. vary ω while holding U and V constant
through the quadrupole mass analyzer. b. vary U and V but keep the ratio U/V fixed

The mass spectrum is obtained by varying the

If U and V are scanned such that U/V = constant,
voltages on the rods and monitoring which ions pass
then successive detection of ions of different m/z is
through the quadrupole rods.
achieved.
Quadrupole mass (QM) analyzer is a "mass filter".

Graphically, e.g. the three stability curves represent

Two functions a and q define a stable trajectory for values of U and V for which the masses m1, m2 and
which ions do not collide with the rods across a m3 have stable trajectories through the quadrupole.
range of values of U and V.
Only those mass values above the operating line
−4 zeU −2 zeV transmits.
a= 2 2 2 q= 2 2 2
m r0 ω m r0 ω
In principle QM can be operated for a range of
U and V values.
(U)
a 2U
=
q V
(V)

6
 The resolution is determined by the magnitude
of U/V ratio.

Resolution of the mass analyzer can be increased

by increasing the slope of the curve U/V = const.,
and that if U = 0 then ions of all m/z are transmitted.

Because quadrupoles operate at lower voltages, they

can be scanned at faster rates (~1000 a.m.u./s) than
magnet based Spectrometers. QMs are better
detectors for LC-MS and GC-MS implementation.

MS/MS Time-of-Flight Mass Analyzers (spectrometer):

MS1 MS2
Dissociation region
TOF measures the mass-dependent time required for
ions of different masses to move from the ion source
to the detector.

This requires that at the starting time t=0, (time ions

QqQ leaves the ion source) to be well-defined.

Scan with MS1 (only) turned on – entire MS spectrum.
Set MS1 to filter fragment of interest, dissociate further
in q collisionally and mass analyze by scanning
with MS2.
Ions are created by a pulsed method (MALDI), or
by rapid electric field switching that serves as a 'gate'
to release the ions from the ion source in a very
short time.

mv 2 L m 2Vt 2 1 m
KE = zeV = v= = 2 t=L
2 t ze L 2eV z Reflection time-of-flight mass spectrometer

7
Parent/Molecular Peak M: High Resolution MS:

An molecular ion that has not lost/gained atoms.
The nominal mass of which is calculated with the
mass numbers of the predominant isotopes of atoms.
Base peak:
Base peak is the peak from the most abundant ion,
which is often the most stable ion.
Using mass number for isotopes of atoms is
approximate. Actual mass of a given isotope deviates
this integer by a small but unique amount (∆E = ∆mc2).
Relative to 12C at 12.0000000, the isotopic mass of
16O is 15.9949146 amu., etc.
High resolution mass spectrometers that can
determine m/z values accurately to four/more
decimal places, making it possible to distinguish
different molecular formulas having the same
nominal mass.

m/z=74
Very short list. MF Unsaturation
C2H2O3 2.0
Accurate
Isotope
Mass CH2N2O2 2.0
1-H 1.007825 C6H2 6.0
2-H 2.014102
C3H3FO 2.0
12-C 12.000000000
13-C 13.0033548 C2H3FN2 2.0
14-N 14.0030740 C3H6O2 1.0
15-N 15.0001090
C2H6N2O 1.0
16-O 15.9949146
17-O 16.9991306 C4H7F 1.0
18-O 17.9991594
C4H10O 0.0
C3H10N2 0.0
http://www.chem.queensu.ca/FACILITIES/NMR/nmr/mass-spec/mstable3.htm

MF Unsaturation Exact Mass Isotope Peaks

m/z=74
C2H2O3 2.0 74.00040
The peaks from the isotopes.
CH2N2O2 2.0 74.01163
C6H2 6.0 74.01565
The intensity ratios (relative intensities) in the isotope
C3H3FO 2.0 74.01679 patterns are arising from the natural abundance of the
C2H3FN2 2.0 74.02803 isotopes, thus are valuable to ascertain the atomic
C3H6O2 1.0 74.03678 composition of ions.
C2H6N2O 1.0 74.04801
C4H7F 1.0 74.05318
M+1 peaks are primarily due the presence of 13C
in the sample.
C4H10O 0.0 74.07316
C3H10N2 0.0 74.08440
M+2, M+4, .. indicative of presence of Br, Cl, S;
MF finder (79Br:81Br = 1:1, 35Cl : 37Cl = 3:1)

8
 Isotope peak abundance depends on the molecular
constitution. Example, halogens Cl, Br.

100
0.801
4.52

Calculating isotope peak abundances (%) to confirm Parent/Molecular Peak M: an ion that has not
fragment and parent peaks. lost/gained atoms (odd electron fragment, U=integer;
reverse not true).

Parent Peak leads to molecular formula.

X=M Molecular formula leads to the structural (/partial)
features of the possible molecular structure.

Unsaturation can be calculated from the molecular

formula of the parent ion.

U = R + DB + 2TB = c − h / 2 + n / 2 + 1
c = #C & Si
E.g. h = #H & halogens
%M+1=(0.012nH+1.08nC+0.369nN+0.038nO+5.08nSi+0.801nS)100
n = #N, P, As..

Rule of Thirteen:

Used to generate possible formula for a given

molecular mass.

imax is the total number of different elements in the 1. Generate a base formula;
composition, Ni the number of atoms of element i, M r
and Vi is the valence of atom i. =n+ CnHn+r
13 13

2. Calculate the index of H deficiency, U for the

base formula.
n−r+2
U=
2

9
3. If needs to find the formula with nO of O for the Useful Links
same M;

New formula = Base formula + nO O – nOC – 4nOH

http://www.colby.edu/chemistry/NMR/NMR.html
which changes U to U+nO ;
http://www.colby.edu/chemistry/PChem/Fragment.html
4. If needs to find the formula with nN of N for the
same M,
http://www.chemcalc.org/ Main Page
MF finder
New formula = Base formula + nN N – nN C – 2nN H
and recalculate U; Fractional U’s – unlikely formula.

U < 0 is an impossible combination, indicates likely http://www.chem.uni-potsdam.de/tools/index.html

presence of O and N.

Example 1 only peaks

(M+1)/M ratio is paticularly useful to estimate the
m/z Relative abundance(normalized)
#C in the species.
64 100.0
%M + 1 65 0.9±0.2
#C ≤ × 100
%M (next slide) 66 5.0±0.5

Nitrogen Rule:
Many peaks can be ruled out as impossible simply on
the grounds of reasonable structure requirements.

Molecule of even nominal mass must contain zero or

even number of N atoms. An odd numbered nominal
http://www.colby.edu/chemistry/PChem/Fragment.html
mass requires an odd number of N.
MF finder

Example 2 Organic compound

m/z Relative abundance Normalized
58 12.0 100.0
59 0.5±0.2 4.2±1.7
60 0.0±0.2 0.0±0.2

43 100.0
44 3.3±0.3
45 0.0±0.2

.
.
http://www.colby.edu/chemistry/PChem/Fragment.html
MF finder Aromatic parent ions – large abundance.

10
Fragmentation: Fragmentation pattern:
Fragmentation leads to smaller ions by the cleaving of
parts of the molecule. From the fragment losses the parent peak may be
predicted by working back.
Unreasonable losses from molecular ion:

M - [3-14] and M – [21-26] are unreasonable losses. n-decane

(next slide)

Reasonable losses from molecular ion

Neutral fragments expelled by simple cleavage
OE+• → EE+ + OE•
Neutral fragments expelled by multi-centered fragments
OE+• → OE+• + EE -29

See handout

Notation: two electron movement

+.

57

29

cleavage here one electron movement

Unique isotope peak patterns are useful for

Unique isotope the analysis.
peak patterns
are useful for
the analysis.

11
m/z=100 Fragmentation:

Nominal mass of parent ion containing C, H, O, S, Si,

P and halogens is even.
odd e m/z ion

(homolytic) cleavage
? R' R+.
even m/z, no N
- neutral species, e.g. CO, water, ..

even e m/z ion

Structural isomers differentiation

Fig. E3
M = odd, 17 one N Nominal mass
17
M+1 0.4% no C 14 – 1N
3 3H
Consistent with 1N
M use table
M+1 NH3

H
+N H
H
Fig. E3

PE1 PE1
Nominal mass
M = even
16
M+1 1.1% 1C 12 – 1C
4 4H

M CH4
M+1

12
PE2 PE2

0.9
1 O present
21.1

100.0 2 H present
0.06

0.2
M+1, M+2 peaks confirm H2O.

M+1 no C

Data:Contains 3 elements, one is F.

PE3 Nominal mass
34
PE3 12 – 1 C
22
19 - 1 F
3 3H

CH3F
H
H F Further 34-15=19
M+1 1C H
15

H 3C
29 note: reasonable
43 losses
57 CH 3
71
Fragment intensities depend on the stability of the
ion and the probability of formation.

13
 43 Electron repelling methyl group
H3C
CH3 stabilizes the carbocation.
57
H3C CH3

29
43
57
CH3
71

CH3
Stabilization of carbocations:
H3C CH3 Four ways to form carbocation
CH3 1. Alkenes frequently undergo fragmentations that yield allylic cations

57

2. Carbon-carbon bonds next to an atom with an unshared electron pair usually

break readily because the resulting cation is resonance stabilized.

Z =N, 0, or S; R may also be H.

3. Carbon-carbon bonds next to the carbonyl group of an aldehyde or ketone

break readily because resonance-stabilized ions called acylium ions 4. Alkyl-substituted benzenes undergo loss of a hydrogen atom or methyl group
are produced. to yield the relatively stable tropylium ion. This fragmentation gives a
prominent peak (sometimes the base peak) at m/e 91.

14
 INTENSITY PE7
m/e (AS PERCENT OF BASE PEAK)
5. Substituted benzenes also lose their substituent and yield a phenyl 14 8.0 73
cation at m/e 77. 15 38.6 36 - 3C from M+1
18 16.3 37
28 39.7 14- 1N odd M
29 23.4 23
Recalculating
42 46.6
intensities to 16- 1O from M+2
Y =halogen,-NO2.-Keto group,-R. etc.
43 10.7 normalize 7 7H
44 100.0 (base) C3H7NO 1.0
73 86.1 M+. 100
3.2 3.72
C3H7NO
74
75 0.2 0.23 Use this resource

Mass spectrum for

Problem E. 7.
FIG. E.8

INTENSITY
m/e (AS PERCENT OF BASE PEAK) C3H7NO
14 8.0
Confirming MF No halogens
15 38.6
C3H7NO 3.81 0.25 73.0528 o 1 Probably 1 O; (M+2)
16.3
18 4 C ; M+1
28 39.7 OH
H3C
29 23.4 N Nominal mass
N H3C
42 46.6 73-31 72
O
43 10.7 42 31 48 – 4 C
NH2
44 100.0 (base) 24
73 86.1 M+. 100 H3C
O
44 16 - 1 O
74 3.2 3.72
8 8H
75 0.2 0.23
Mass spectrum for Calculation; C4H8O 4.56 0.28 72.0575 o 1
Problem E. 7.
FIG. E.8 Use this resource

Cleavage of Two bonds:

O 1. Alcohols frequently show a prominent peak at M - 18.
This corresponds to the loss of a molecule of water.
H3C H3C
CH3
O

OH
H3C
H3C
CH2
OH

2. Cycloalkenes can undergo a retro-Diels-Alder reaction that produces

72-44=28 loss of 28 from M to form base peak an alkene and an alkadienyl radical cation.

No straight forward cleavage possible; probably

a rearrangement occurs before cleavage.

15
 The compound is butanal. m/z = 44 arises from the McLafferty
rearrangement.
3. Carbonyl compounds with a hydrogen on their -/-carbon undergo a
fragmentation called the McLafferty rearrangement.

m/z=29 arises from acylium ion.

Y may be R, H, OR, OH, etc.

In addition to these reactions, we frequently find peaks in mass spectra that
result from the elimination of other small stable neutral molecules, for
example, N2, NH3, CO,HCN, H2S, alcohols, and alkenes.

M+=116
present

MS of which compound?
O
O

Isopropyl butyl ether Propyl butyl ether

+ +.
O O
+
O
73

101 101 Chromatography – Mass

73 Spectroscopy
+
+ +. O
O O

87
87
73

16
GC-MS, LC_MS are hyphenated analytical techniques. Chromatograph
Output
Two techniques are combined to form a single method Data
Output

for analyzing mixtures of chemicals. Chromatography

separates the components and MS detects and Inlet
Data
System
characterizes each of the components. Combination
of the techniques allows both qualitative and
quantitative evaluation of analytical samples.
Ion Mass Ion

The MS spectrometer can be highly selective for

Source Analyzer Detector

QM
analyte of interest.
Electrospray All fragment ions/or
API single ion detected and
Vacuum
Pumps
(total) ion current
determined

Each analyte’s MS produced Total (reconstructed) ion current vs time

http://www.gmu.edu/departments/SRIF/tutorial/gcd/gc-ms2.htm

GC,LC/MS takes data in three dimensions

simultaneously, recording the number of ions
created along with their masses over time.

This information is generally represented by

examining the total ion chromatogram (TIC) t
MS – analyte 1
and ‘slicing’ along the third dimension (m/z) of a MS – analyte 2
given peak to look at the mass spectrum at a MS – analyte 3
chosen time.

t
3D data set

17
MS requires high vacuum ~10-3 – 10-6 torr. The
eluate from a chromatographic system has much
more carrier (gas/eluent) than the analyte. If eluate
from the chromatograph is allowed to enter directly
into the MS it would overwhelm the vacuum system
(especially in LC – liquid vaporizes into a large
volume).

To alleviate the strain on the system, different

strategies are employed at the sample
introduction/ionization stage.

Electrospray interface:

Coaxial flow

Strong electric field

+ nebulization;
ions vaporize

For positive ion MS

Low collisional dissociation; minimal fragmentation. Atmospheric Pressure Chemical Ionization API-CI
Positive ions accelerate
and collide with N2 molecules;
few fragments form. Changing
skimmer voltage to larger –ve
would produce more fragmentation.

Coaxial flow

Electrons formed at corona, injects into aerosol.

This technique produces single charged ions.

Little fragmentation, less structural information.

18
 Selected ion monitoring possible with the quadrupole
N 2 + e → N 2+. + 2e mass analyzer by appropriate tuning.

N 2+. + 2 N 2 → N 4+. + N 2 Usually total ion current monitored ion current from all
+. +. ions from an analyte is constructed.
N + H 2 O → 2 N 2 + H 2O
4

H 2O +. + H 2O → OH . + H 3O +
H 3O + + nH 2O → H 3O + ( H 2O )n
H 3O + ( H 2O ) n + M → MH + + (n + 1) H 2O

Selectivity and S/N ratio increased markedly with

QqQ triple quad arrangement.
Selected ion chromatogram
QM filter tuned to select the ion

Quadrupole Ion Trap Mass Analyzer.

19
 MALDI MALDI

mv 2 L m 2Vt 2 1 m
KE = zeV = v= = 2 t=L
2 t ze L 2eV z

Lasers are used to deliver a focused high density of

monochromatic radiation to a sample target, which is
vaporized and ionized.

The yield of ions is often increased by using a

secondary ion source or a matrix.

20