Chapter 5

Mass
spectrometry
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INTRODUCTION
Generally in case of spectroscopy one phenomenon
is common “There is an interaction between electromagnetic
radiation and matter (sample).” While in case of Mass
spectrometry this phenomenon is not occurring. So some
people are not agree that Mass spectrometry is not a
spectroscopic technique.

In case of UV, IR, and NMR sample absorb the light

(electromagnetic radiation), so these are absorption
spectroscopy. While Mass is differ from them.

First requirement for Mass spectroscopy is organic

sample either in gaseous or vapor state.

Principle:

When organic molecule in the gaseous or vapor state

at a very low pressure are subjected to 70 ev. it loses the
electrons and forms a positively charged cation. This this
can be accelerated and deflected by magnetic and electrical
fields is depends on its mass, charge, velocity and deflecting
force. In these four variable charge is negligible, velocity
and deflected force are constant, so the deflection depends
only on mass. If mass is high, deflection is low, and if mass
is low, deflection is higher. i.e. deflection is higher for light
particle and lower for heavy particle.

So when organic molecule is gaseous or vapor state,

bombarded at 70 ev at reduce pressure, it loses the electron
and forms a positively charged ion. This ion can be
accelerated and deflecting by magnets depends on charge,
velocity, mass and deflecting force but charge is negligible ,
velocity and deflecting force are constant so deflection only

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depends on mass, it is higher for light particle and lower for
heavy particle.

Where,

H= applied magnetic field

R= radius of arc of deflection

V= applied accelerating voltage

= deflection force

The fragmentation pattern of compounds depends on

the characteristic of sample and it provide detail information
about molecular structure can be obtained.

Instrumentation:

Schematic diagram of a mass spectrometer.

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 1. The inlet system
2. The ion source (ionization chamber)
3. The electrostatic accelerating system
4. The ion separator
5. Ion collector/ read out
6. Vacuum system

Vacuum is necessary, so it can be in isolated type of

situation. It will be necessary to minimize the no. of
collision between ions and non-ionized molecule.

Requirement of the instrument:

1. To vaporize compounds of varying volatility

2. To produce cation
3. To separate ions
4. To detected and record them.

According to above fig. mass spectrometer possess

following parts

1. Sample inlet system

2. Ionization chamber
3. Accelerating region
4. Analyzer tube
5. Detector
6. Recorder

The mass spectrometer is used for the investigation

of any compound may vary in their types but generally
contain following part.

1. Sample inlet system:

Function: To vaporize the sample and uniformly
introduce into ionization chamber.

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 For successful ionization, all samples must
be converted in to gaseous or vaporized state, to
achieve this sample must be heated at 400 ºC, so
temperature of inlet system is above 400 ºC.
The rate of introducing sample in ionization chamber
is remain constant, for this it consist special device
of micrometer for determining the amount of sample
is introduce.
Another important point is the introduction of
sample vapor at low pressure (10-5-10-6 torr) is
necessary to minimize the number of collision
between ions and non- ionized molecule.
Sample size is 1mg to μg.

2. Ionization chamber/ ion source :

Function: to produce positively charged ion and
other different with m/e value.
For the production of positively charged ion,
ionization of sample is necessary.

To ionize the sample different techniques are

available:

a) EI (Electron impact ionization): In electron

impact system, when sample in gaseous state
bombarded at electron vault and low pressure at
10-5-10-6 at high energy provide a molecule to
emitted electron from filament and forms a
positively charged ion.
Means an electron from electron beam knocks an
electron from parent molecule to give parent ion.

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But for this high electron vault and low pressure is require.
- Due to this pyrolysis of molecule may take
place
- M + ion absent
- Complex spectrum is available

So this technique is harsh technique.

b) Field ionization: In this technique, very high

local electric field (2x108 v/cm) on tip of metal
wire pulls off e- for large molecule.
Means by creating vacuum in 2x108 v/cm for
small area and large molecule, so maximum
electron attack on this molecule, it knocks the
electron from parent molecule and forms parent
ion.

c) Chemical ionization: This ionization carried out

on the bases of molecular reaction.
In this technique as a reactant gas generally CH4,
isobutene and ammonia is used.
The reactant gases are stored at low pressure10-4
and in this tank introduce the sample.
So at low pressure reactant gas ionized and
forms ion, which on interact with neutral
molecule and forms chemically reactive species,
which interact with sample and forms positively
charged ion.

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 Advantage: M+ present, simple spectrum is
obtain

This technique is used for polar and high molecular

weight species at 10.000g

d) FAB (Fast atom bombardment source): In this

technique, by using highly energetic chemical
species, bombarded with sample.

This technique is used for polar and high molecular

weight species of 10,000 g.

Advantage:

- Ionization due to transition

- Minimum energy reaction , so minimize the
destructive
- No deviation

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3. Ion accelerating region:
Function: to accelerate the ion in a uniform velocity
and produce ionic beam.
It contain: 1) Repleter plate 2) Ion accelerating slit 3)
Ion focusing slit.
High electrostatic field about 1000 to 2000 v is
applied between two accelerating slits so when ion
pass through this region repleting plate replete to
each other to form uniform way, after this when pass
through between accelerating tube to form narrow
beam due to high voltage moving ion is formed. The
ion focusing plate focuses it. So, uniform beam of
ion is obtained.

Suppose mass of ion is M, charge e-, initial kinetic

energy is negligible.
So, ion is accelerated velocity V.
So, kinetic energy must equal to electrostatic energy.

1)
Electrostatic = ev
Repellers = which direct the ion to the accelerating
plate
Accelerating = accelerate the ion
Ion focusing = which directs the ion into uniform
beam

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4. Analyzer tube :
Function: to disperse the ionic beam in to different
frequency according to their m/e value, an analyzer
must possess following characteristic:
i) It should have a high resolution power.
ii) It should have a high rate of transmission of
ions.

There are two types of analyzer:

1. Ion- trapped mass analyzer

2. Time of flight analyzer

1. Ion- trapped mass analyzer:

Ion- trapped mass analyzer working two way for
trapping ions.
a) Three dimensional ion trap
e.g. Quadrupole mass analyzer
b) Ion- cyclotron resonance

Both analyzer trap the ions by using combine effect

of DC and RF

a) Quadrupole mass analyzer:

Principle: combination of DC and radio frequency

on the four parallel opposite charge rods can be set
to pass only a selected mass to charge ratio.
Remaining ions do not flight through this
quadrupole analyzer and will collide with these rods.
As a result never complete the path up to detector.

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Advantages:
- Classical mass spectrum obtained
- Relatively cheaper system
- Reproducibility of the result is possible

Limitations:

- Resolution is good for limited range

- Peak height is variable as a function of mass
- It is not suitable for pulsed ionization method

The selection of mass analyzer depends on

resolution. Generally at low resolution quadrupole
and lop trap working very good. Time of light and
sector instruments(magnet) work better than other
analyzer. Ion cyclotron resonance was performed
best at ultra-high resolution.

b) Ion- cyclotron resonance:

Ion moves in a circular path under the effect of
external magnetic field. The cyclotron frequency
of the ions depends on their mass, by using
following equation, one can measure the mass of
ions.

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 But angular frequency

Advantages:

- The best resolution as compare to the other

analyzer
- Well compatible for pulsed ionization as well as
MALDI
- Non-destructive ion detection

Limitations:

- Limited dynamic range

- Strictly low pressure required
- Ion- molecule reaction is possible

2. Time of flight (TOF):

Principle: a time of flight mass analyzer determine
the mass dependent time; it takes ions with various
masses to flight from the ion source to the detector.
The necessity of this analyzer is sharp starting time.

Ion leaving the ion source is

…………….1)

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 The ion velocity(v) is the length of flight path L
divided by the time
…………..2)
Put the value of V in kinetic energy equation..1)

√
Advantage:
- It is compatible with pulsed ionization or fast
ionization technique.
- Resolution is better than quadruple.
5. Detector and Recorder :
its function is to detect the ionic beam and produce a
mass spectrum expressed as a bar graph in which
relative abundance on ordinate is plated against m/z
ratio.

Ionization technique:
Several methods are available for the ionization of
sample, the most common being electron impact
technique to produce ion.

1) EI (Electron ionization): vaporized sample is

introduced under low pressure operate at
pressure of 10-5-10-6 in to a tube called „ion
source‟.
The entering molecule of sample is bombarded with
electron emitted from a hot tungsten or Rh wire and
then accelerated to an anode.

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Limitation:

1) As the sample should be in vapor state,

possibility of pyrolysis of less volatile
substance or sample increase.
2) It produce molecular ion with excess internal
energy, which leads to various fragmentation
process. Fragmentation is drastic, complex
fragmentation pattern is obtained.
3) 70 ev often destroys the molecular ion, hence
the absence of very weak molecular ion peak
does not allow the detection of molecular
weight and empirical formula.
2) Field ionization:
Field ionization is achieved by applying a very
high local electrical field of order 2x108 v/cm.
When the sample is subjected to a very strong
electric field, it pulls off an electron forming a
positive ion. It is a particular technique for the
study of large molecules that do not show a
parent ion.
3) Chemical ionization:
This technique based on ion-molecule
reaction.
In it, reaction gas like methane, isobutane, and
ammonia is introduced at high pressure (1-torr)
with the sample (10-4 torr) to be analysed by the
mass spectrometer in the ionization chamber.
When a beam of electron passes the ionization
chamber. The reaction gas undergoes ionization
to produce ion, which reacts with the neutral
molecule to form a product.

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 To the product so formed are chemical
reactive species and can interact with the sample
and forms a positive ion.
Thus methane from CH4•+ + CH3+ initially which
react with a neutral molecule.

The CH5+ is strong proton donor and thus reacts with

analyte molecule to form (M+1)
ions.

Advantage:

1) It produces molecular ion peak.

2) As the fragmentation is reduced, it produces a
spectrum which one quite simple.

4) FAB (fast atom bombardment):

Chemical ionization technique still requires
molecules that are moderately volatile and
thermally stable. This excludes many compounds
of biological interest and importance such as
peptides and carbohydrates. The most reset
technique FAB overcomes. These problems
polar molecules like peptide with molecular
weight 10,000 can be analyzed.

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In this method, Xe or Ar atoms of high
translational energy are produced by the
following sequence.
I) Xe atoms are ionised by bombardment of
e- beam it to form Xe cation radical.

II) They are accelerated to 6-10 K.ev. to give

radical cation of high translational energy
Xe+

III) Xenon radical cation is then allowed to

interact with neutral xenon atoms due to
which xenon radical cation passes its
energy to neutral atoms hence fast
moving Xe atoms are produced.

IV) The resultant fast atoms are directed onto

the sample held in a liquid matrix on a
metal target. The liquid matrix is
commonly glycerol.

Advantage:
1) Ionization is caused by the transition
of energy; it minimizes destruction of
molecular ion which can alert for 20-
30 min.

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 2) This method is most effective for a
relatively polar substance which is
Non- volatile or thermally unstable.
3) Derivatization not required.

The positive ions produced by the

interaction of the molecular ion with e- beam are forced
through the first accelerating slit by a weak electrostatic
field.

So the main components of ion accelerating region are:

1) Ion repellers
2) Ion accelerating plate
3) Ion focusing plate
A potential of 1000-2000v is used to
accelerate the ions through a slit system. This produces a
narrow beam of the fast moving ion. Mass of ion is M, the
charge is e- initial kinetic energy is negligible. Now ions are
accelerated to a velocity (v), hence their kinetic energy must
be equal to their electrostatic energy, accelerated in passing
through the applied volt (v).

………………. (1)

Function of mass analyzer:

Its function to disperse the ionic beam according to their
m/e value.
The ion produced may separately according to their mass to
charge ratio(m/e) using the magnetic and electrostatic field.

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The fast moving ions enter the analyzer tube. The analyzer
tube is an evacuated (10-7 to 10-8 torr) curved metal tube
through which ion beam passes from source.
In the analyzer tube, the ions are subjected to a uniform
magnetic field(H), which is generated by an electromagnet
and perpendicular to the direction of the ionic beam.
In the magnetic field, the ions are deflected along a circular
path of radius (r).

i.e.

Or

…………….. (2)

This relationship reveals that in the case of an ion with

a given mass to charge ratio (m/e).
The radius of deflection can be increased by
decreasing magnetic field (H) allow only ions of
certain suitable m/e values to pass through the
collector slit and enter into the collector.

Detection of the presence of the isotopes of the elements

and the recognition of molecular ion peak:
Mass spectra often contain peak of
significant intensity that is attributed to the presence of
isotopes. Two of these are called the M+1 and M+2 peaks,
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where M is the mass of the parent ion and 1 or 2 represent
mass increases due to heavier isotopes of the ½ mass unit.
most elements occur naturally as several isotopes and
generally, the lighter of these predominates while the
heavier one occurs to a lesser extent.
The isotopic compositions of some common elements are as
shown under.

Element M+ M+1 M+2

Hydrogen 1H 99.98% 2
H 0.016
Carbon 12C 98.42% 13
H 1.08
Nitrogen 14N 99.62% 15
C 0.38
Oxygen 16O 99.76% 17
O 0.04 18
O 0-
2
Sulphur 32S 94.825 33
S 0.74 34
S 4-
40
36 37
Chlorine 67.5% Cl - S 32-
35
Cl 5
81
Bromine 50.5% - Br 49-
79
Br 5
Iodine127I 100% - -

Isotopic composition of element is constant:

150 (M) 100%
151 (M+1) 10.2%
152 (M+2) 0.88%
C7H10N4 9.25-0.38
C8H8NO2 9.23-0.78
C8H10N2O 9.61-0.61
C8H12N2 9.98-0.45
C9H10O2 9.96-0.82
C9H12NO 10.34-0.68
C9H14N2 10.71-0.5

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 Fragmentation value of the compound shows according
to Nitrogen as well as Ring rule.

1. Nitrogen rule:
Nitrogen rule states that the compound containing an even
no.of nitrogen atoms (including O) will give a molecular ion
with an even mass number,
For e.g. the following ion gives their molecular ion at given
mass number.

CH4 16
CH3OH 32
CClF3 104
C6H5OH 94
NH2-NH2 32
C5H5N2 94

An odd number of nitrogen atoms

causes the molecular ion to be at odd mass number. e.g.
NH3 m/z = 17 and C2H5NH2 = 45
It is important to note that the
relationship applies to all ions and not only to the molecular
ions.
2. Ring rule:
Elemental composition of any fragmentation peak can be
established by using the following formula.

Where X= No. of carbon

Y= Monovalent ion
Z= Trivalent ion

For example C5H5N+

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