303351 Instrumental Analysis

Lecturer: !
Mass Spectrometry Somsak Sirichai
 Mass Spectrometry
What is Mass spectrometry (MS)?
An analytic method that employs ionization and mass analysis
of compounds in order to determine the mass, formula and
structure of the compound being analyzed.

In general there are four steps associated with a mass

spectroscopic experiment:
generate gas-phase molecules from analyse (solid, liquid, gas, etc)!
ionize those molecules!
separate the ions based on mass-to-charge ratio!
detect the ion beam
Molecular Mass Spectra

Figure 20-1 Mass spectrum of ethyl benzene

C6 H 5CH 2CH 3 + e C6 H 5CH 2CH 3i+ + 2e

molecular ion
 How are mass spectra produced?
1. Ions are produced in the source and are transferred into the
mass analyser
2. They are separated according to their mass/charge ratio in the !
mass analyser (e.g. Quadrupole, Ion Trap, Time of Flight)

3. Ions of the various m/z values exit the analyser and are !
counted by the detector

C6 H 5CH 2CH 3 + e C6 H 5CH 2CH 3i+ + 2e

molecular ion
 Mass Spectrometer

Figure 20-11 Components of a mass spectrometer

 1. Sample Inlet Systems
to permit introduction of a representative sample into the ion
source with minimal loss of vacuum

modern mass spectrometers are equipped with several types of

inlets to accommodate various kinds of samples; these include
batch inlets, direct probes inlets, chromatographic inlets, and
capillary electrophoretic inlets.
 Inlet system - three requires are:
(1) that sample must be in the vapour phase prior to ionisation;!
(2) that sample does not suffer thermal decomposition during
the vaporisation process;!
(3) that during admission of the sample, the pressure inside the
mass spectrometer us kept as low as possible.
 Batch Inlet Systems
the conventional (and simplest) inlet system

the sample is volatilised externally and then allowed to leak into the
evacuated ionisation region.

Figure 20-12a is a schematic of a typical system that is applicable to

gaseous and liquid samples having boiling point up to about 500o C
Figure 20-12 (a) Schematic of an external sample-introduction system
 The Direct Probe Inlet
solid and nonvolatile liquid can be introduced into the ionisation
region by means of a sample holder, or probe, which is inserted
through a vacuum lock (Figures 20-12b)

Figure 20-12(b) !
a sample probe for inserting a
sample directly into the ion source!
!
temperature-programmable
under data system control!
vaporizes small amounts of
material into MS source!
requires pure sample for
meaningful spectra!
can dirty MS rapidly!
Labor-intersive: all-manual
operation
Chromatographic Inlet System

Figure 27-14 Schematic of a typically capillary GC/MS system

 2. Ion Source
Once we have produced gas phase molecules, we can start to look at
the ionisation process

the appearance of mass spectra for a given molecular species

strongly depends on the method used for ion formation

the methods fall into two major categories: (1) gas-phase sources and
(2) desorption source (Table 20-1)
 gas-phase source: the sample is first vaporised and then ionised. !
generally restricted to ionisation of thermally stable compounds !
that have boiling point less than about 500oC

desorption source: the solid- or liquid-state sample is converted

directly into gaseous ions.!
advantages - they are applicable to nonvolatile and thermally !
unstable samples
Ion sources are also classified as being hard sources or soft sources.

Hard ionisation sources!

impart enough energy to analyte molecules to leave them in a
highly excited energy state. Relaxation then involves rupture of bonds,
producing fragment ions that have mass-to-charge ratios less than of
molecular ion.

Soft ionisation sources!

cause little fragmentation. Thus, the mass spectrum from a soft
ionisation source often consist of the molecular ion peak and only a
few, if any, other peak.
Figure 20-2 Mass spectrum of 1-decanol from (a) a hard ionisation source (electron
inpact) and (b) a soft ionisation source (chemical ionisation)
Hard- and soft- ionisation sources!
!

both are useful for analysis. The many peaks in a hard-source

spectrum provide useful information about the kinds of functional
groups and thus structural information about analytes. !
!
Soft-source spectra are useful because they supply accurate
information about the molecular mass of the analyte molecule or
molecules.
 The Electron-Impact Source

ions from mass analysis were produced by electron impact. In this

process, the sample is brought to a temperature high enough to
produce a molecular vapour,, which is then ionised by bombarding the
resulting molecules with a beam of energetic electrons.!
!
Figure 20-3 is a schematic of a basic electron-impact ion source.
Figure 20-3 An electron-impact ion source
Figure 20-3 is a schematic of a basic electron-impact ion source.
Electrons are emitted from a heated tungsten or rhenium filament and
accelerated by applying approximately 70 V between filament and the
anode. As shown in the figure, the paths of electrons and molecules are
at the right angles and intersect near the center of the source., where
collision and ionisation occur.
 EI source: advantages vs. disadvantages
convenient to use and produce it may be difficult to measure
high ion currents , thus giving relative molecular masses for
good sensitivities. ! some molecules!
! it is difficult to distinguish
extensive fragmentation and between isomers!
resulting large number of peaks is some compounds may undergo
also advantage because it often thermal decomposition prior to
makes unambiguous ionisation or be very prone to
identification of analyte possible. fragmentation after ionisation
because of the temperature
required for vaporisation !
applicable only to analytes
having molecular masses smaller
than about 103 Da
 Chemical Ionisation Source
most modern mass spectrometers are designed so that electron-
impact ionisation and chemical ionisation can be carried out
interchangeably. Such sources are called EI-CI sources.

In CI source, gaseous atoms of the sample (from either a batch inlet

or a heated probe) are ionised by collision with ions produced by
electron bombardment of an excess of a reagent gas (or buffer gas).

Buffer gas often methane (CH4) is added in a large excess (about

x100 fold) over the analyte. The electron beam ionises the methane,
gas phase reactions produce molecular ions such as CH5+ which
collide with analyte molecules (M) to gently produce the analyte
ions (MH+)
 Chemical Ionisation Source
can perform CI in same unit with EI except for much higher pressure!
!
must provide additional pumping speed to move extra buffer gas
out of ioniser so as to not degrade vacuum in analyser

Methane reacts with high-energy electrons to give several ions such as

CH4+, CH3+ and CH2+. The first two predominate and represent about
90% of the reaction products.

CH 4 + + CH 4 CH 5 + + CH 3
+ +
CH 3 + CH 4 C2 H 5 + H 2
Step 1: Formation of Reagent Ions!
Primary step by electron impact
+
CH 4 + e CH + 2e

4
+

CH + H 2
2
+

CH + H
3

Secondary step by Ion-Molecule Reactions:

+
+
CH + CH 4 CH + CH 3 i
4 5
+
+
CH + CH 4 C2 H + CH 2
3 5
+
C2 H + CH 4 C3 H 5+ + 2H 2

5
Step 2: Ionisation of sample molecule!
Protonation/adduct formation

+ +
M + CH MH
5
m/z = M + 1
+ +
M + C2 H M C2 H 5
5
m/z = M + 29
+ +
M + C3 H M C3 H 5
5
m/z = M + 41

Hydride abstraction
+
+
M + CH M M + CH 4 + H 2
5
i

Commom for large straight chain aliphatic molecules

 Field Ionisation Sources
In field ionisation source, ions are
formed under the influence of a
large electric field (108 V/cm)!
!
when a large potential difference
(10-20 kV) is applied across an
object fashioned to come to an
atomically sharp point, the
electric field is HUGE (108 V/cm).
These fields, across a single
molecule, have enough potential
Figure 20-5 Photomicrograph of a
to pull an electron directly from
carbon microneedle emitter (Anode of
the molecule FI source).
 Field Ionisation Sources
Advantages:!
! ! the field ionisation spectrum is relative simple (uncomplicated), !
with an easily distinguished (M+1)+

Disadvantages:!
! ! a limitation to field ionisation is its sensitivity, which is at least
an order of magnitude less than that of EI sources; maximum currents
are on the order of 10-11 A - low sensitivity and resolution
 Desorption sources
The ionisation methods discussed so far require that the ionisation
agents act on gaseous samples. Such methods are not applicable to
nonvolatile or thermally unstable samples.

A number of desorption ionisation methods have been developed for

dealing with this type of sample. !
advantages: enable mass spectra to be obtained for thermally
delicate biochemical species and species having molecular masses of
greater than 100,000 Da.
 Electrospray Ionisation (ESI)
one of the most important techniques for analysing biomolecule
such as polypeptide, proteins, and oligonucleotide, having
molecular weights of 100,000 Da or more

produce multiply charged ions based on ion evaporation process

Figure 20-9 Apparatus for electrospray ionisation
 Fast Atom Bombardment Sources
FAB sources, aslo called liquid secondary-ion sources !
!
concept: if a beam of fast moving neutral atoms are directed onto a
metal plate coated with a sample, then much of the high kinetic
energy of the atoms is transferred to the sample molecules on
impact. This energy ca be dissipated in various ways, some of
which lead to ionisation of the sample. !
!
The bombarding atoms are usually rare gases, either xenon or
argon. In order for them to achieve a high kinetic energy, atoms of
the gas are first ionised and these ions are then pass through an
electric field.
 3. Mass Analyzers
At the heart of a mass spectrometer is the mass analyser system.
The selection is always based upon a mass-to-charge ratio, rather
than on absolute mass.!
!
ideally, the mass analyser should be capable of distinguishing minute
mass differences. In addition, the analyser should allow passage of a
sufficient number of ions to yield readily measurable ion current!
!
The principal devices for mass selection are!
! ! Magnetic sector Analyzer!
! ! Double-Focusing Spectrometers!
! ! Quadrupole Mass Spectrometers!
! ! Time-of-Flight (TOF) Mass Analyzers!
! ! Ion-Trap Analyzers
 m
Resolution (R) of Mass Spectrometers R=
m

where m is the mass difference between two adjacent peaks that are
just resolved and m is the nominal mass of the first peak (the mean
mass of the two peaks is sometimes used instead)

Example 20-3 !
What resolution is needed to separate the ion C2H4+ and CH2N+, with
masses of 28.0313 and 28.0187, respectively?
Magnetic Sector
A charges particles
(an ion) experiences a
force that bends its
path when moving
through a magnetic
field. The balance
between magnetic
force and centripetal
force brings an ion of
a particular m/z to
entrance slit of a
detector.
 Magnetic Sector Equations
The magnetic force Fm is given by Fm = Bzev
2
mv
This is balanced by the centripetal force Fc Fc =
r
These are equated and solved for the velocity
2
mv Bzer
Bzev = v=
r m
Substitute into expression for kinetic energy. Solve for m/z
2
1 2 1 Bzer m B2r 2e
Ek = zev = mv = m =
2 2 m z 2V
Double-Focusing Spectrometers
A single magnetic
sector instruments
resolution is limited by
spread of translational
energy of the ions
coming from the source.
A double focusing
instrument uses an
electrostatic field to
narrow the energy
spread before the ions
enter the magnetic
sector. Resolutions of
105 are achievable with
these instruments.
Quadrupole Mass Spectrometers
Ion-Trap MS
This device is able to trap and
hold ions in a space charge
region with their masses
extending over a range of
several thousand Da.
Adjusting the end cap fields to
a particular frequency can
drive a narrow Da range into
the detector. !
has a high ion yield for species.!
dynamic concetration range is
low. Ions spends lots of time in
the trap and ion-ion interaction
can alter the fragmentation
pattern.
 Tandem Mass Spectrometry
Usually referred to as MS/MS. Here one take a mass spectrum of a
mass spectrum. A particular ion fragment isolated in the first stage is
directed to the second phase for further fragmentation and dispersion. !
!
There are three common types:!
! EBEB: !! Double Double Focusing. Two double focusing ! ! !
! ! ! ! magnetic sector stages.!
! QQQ: !! A triple Quad. Three quadrupoles sections in series.!
! Q-TOF: ! A quadrupole stage preceding a TOF analyzer
Figure 20-23 Schematic of a triple
quadrupole mass spectrometer
 TOF
Ions accelerated to same energy. Mass difference mean different
velocities. There travel a long path (1-2 m) and arrive at the detector at
different times. Time of arrival calculates m/z.
 2
m 2Vt m
= 2
t = L
z L 2zV