gas chromatography mass spectrometry

1. M.Sc. Forensic Science

2.  Introduction to chromatography
 Introduction to gas chromatography
 Operating principle
 Instrumentation
 Applications
 Advantages
 Limitations

3.  Chromatography –chroma + graphy - means study of
color
 A non destructive procedure for resolving a
multicomponent mixture of trace ,minor or major
constituents into its individual fractions .
 Defined as –a method of separating a mixture of
components into individual components through
equilibrium distribution between 2 phases(stationary and
mobile phase).
 First invented by M. Tswett , a botanist in 1906
 In 1930 thin layer chromatography and ion
exchange chromatography introduced .
 In 1941 , Martin and Synge introduced partition and paper
chromatography and gas chromatography in 1952 .

4.  GC- MS is a synergestic combination of two powerful
analytical techniques( hyphenated technique)-
 GC(gas chromatograph)- seperates the components of
a mixture in time(stationary phase- immobilised liquid or
solid packed in a closed tube and mobile phase- gas) .
 MS(mass spectrometer)- provides information that
aids in the structural identification of each component .
 Provide a characteristic fragmentation pattern / chemical
fingerprint
 Sample required for analysis in both the instruments is
less than 1ng
 Introduced by Synge and Martin in 1952

5.  Powerful analytical tool used to quantify known materials
, identify unknown compounds within a sample .
 Complete process involve conversion of sample into
gaseous ions , with or without fragmentation ,
characterized by their mass to charge ratios(m/z) and
relative abundances .
 Resolution – in MS , it measures the ability to
distinguish 2 peaks of slightly different mass to charge
ratio , in a mass spectrum .
a larger resolution indicates a better separation
of peaks .
resolution(R) =M/resolving power
Where, M is mass accuracy-difference between measured
and actual mass .

6. Gas Chromatograph
 Differential separation of various
components of a chemical mixture on the
basis of their affinity towards the –
stationary phase(solid or liquid) and
mobile phase (gas /vapors).
 In GC volatilization of the sample in a
heated inlet /injector takes place, thus
,separation of the components is followed
Mass spectrometer
 basic principle of MS utilizes the
nature of ions.
 By accelerating an ion (an atom or
molecule with an electrical charge) to a
certain speed, and passing it through a
magnetic field, the path of the ion will be
deflected by the magnetic field along a
circular path on a radius.
 The amount of deflection depends on –
the intensity of the magnetic
field and
the mass number of the ion.

8. Gas chromatograph Mass spectrometer
 Carrier gas –N2 or He , 1-
2ml/min
 Injector
 Oven
 Column
 Interface to the GC
 Ion source(ionization
chamber)
 Analyzer
 Pumping system
 Detector
 Working

9.  A cylinder made of steel containing carrier gas under
high pressure , connected to the injection port of the
GC column .
 The gas served as mobile phase allowed to pass at a
pressure of 40-80 psi .
 Most commonly used gases are Nitrogen(N2) ,
Helium(He) , Hydrogen(H2) , Argon(Ar) , Carbon
dioxide(CO2) .
 The choice of carrier gas depends on-
nature of the sample
type of detector used
column efficiency

10.  Amount of the sample required , depends
upon-
 nature and concentration of solutes .
 Size of the column .
 Sensitivity of the detector column .
 Generally , 0.1-50microletres for gas and liquid ,
fraction of mg for solids .

11.  Split injection
 Split less injection
 On –column
injection
 Routine method
 0.1-1% sample to column
 Remainder to waste
 All sample to column
 Best for quantitative analysis
 Only for samples in trace amount
 For samples that decompose above
boiling point-no heated injection
port .
 Column at low temperature to
condense sample in narrow band
 Heating of column starts
chromatography

13.  Important part of GC – decides separation efficiency .
 Made up of glass and stainless steel .
 Diameter = 4.8 mm , length can be from few cm to over
a 100 m
 Can be coiled bent or straight
 Most commonly used stationary phase coated in
column is methyl silicone .
3 types:-
 Packed column
 Open tubular column
 Support coated open tubular column

14.  Packed column :- prepared by packing metal or glass
tubings with granular stationary phase.
for GSC –column packed with porous polymers
For GLC –packing prepared by coating the liquid phase
over a size graded inert solid support .
 Open tubular column :- also referred as capillary or
Golay columns , made of 30-90 m long capillary tubing (
internal diameter=0.025-0.075cm) .
Inside wall coated with liquid in the form of thin and
uniform film .
 Support coated open tubular columns :-made by
depositing a micron size porous layer of support material
and then coating with a thin film of liquid phase .
Preferred for trace analysis .

15.  It is critical for system performance
 Transfers sample from the GC into the MS source without mixing
separated bands
 The compound existing in the GC is a trace components in its carrier
gas at a pressure of 760 torr.
 But the MS operates at a vacuum of about 10 (-6) to 10 (-5) torr.
This difference in pressure of 8 to 9 orders of a magnitude is a
considerable problem
Types of interfaces- jet separator
open split interface

16.  Based on diffusion principle
 Eluate from GC comes,
expands through the nozzle
at A , and shoots towards an
orifice in the wall of the
adjoining chamber on its
way to ion source.
 Across the gap between A
and B there is a tremendous
expansion of gases .
 Compounds having high
diffusivity will diffuse at
right angle(known as
effusion) .
orifice

17.  Offers a convenient
connection between GC
and MS
 Function –depends on
use of purge gas
entering
 All or only a fraction of
the elute enter the ion
source – depending in
the flow rates of –GC
column and purge gas
(H2/He)

18.  As the ions travel from the ion source through analyser to
detector , they are free in motion .
 Their path and direction of travel must be determined only
by the electric/ magnetic fields-used to separate them
according to their m/z values .
 That is why ,the MS –operated at very low pressure (10 -
3Pa or less i.e. vacuum) so that ions not collide with any
other matter and change their direction /break them apart
.
 So to maintain the required pressure , carrier gas to be
removed continually.
 The main function of the vacuum system is to
remove the entering gas so that pressure is
maintained .

19.  Electron impact/fast atom bombardment
 Chemical ionization - Negative chemical ionization
- Positive chemical ionization
 Spark source ionization
 Electro spray ionization

20. Electron ionisation Chemical ionisation

22. Used to produce ions using an electrospray in which a high
voltage is applied to a liquid to create an aerosol

23. Quadrupole analyzer
 Quadrupole analyser
 Ion trap analyser
 Ion Cyclotron Resonance
(ICR)
 Time of flight analyzer

24. Ion trap analyzer Ion cyclotron resonance (ICR)

26.  some type of electron multiplier is used, though
other detectors including Faraday cups and ion-to-
photon detectors are also used. Because the number
of ions leaving the mass analyzer at a particular instant
is typically quite small, considerable amplification is
often necessary to get a signal. Microchannel plate
detectors are commonly used in modern commercial
instruments and photomultiplier tube.

28. Mass spectrum of n-butane
Fragmentation pattern of n-butane

29.  Drug detection
 Fire investigation
 Environmental analysis(organic pollutants)
 Explosives investigation
 Identification of unknown samples
 Pesticides

30.  Separates components of a complex mixture so that
mass spectra of individual compounds can be obtained
for qualitative purpose along with quantitative
information.
 Simultaneous quantification and confirmation of target
analytes .

31.  Only compounds with vapour pressure exceeding about
10(exp10) Torr can be analyzed by GC/MS .
 Certain isomeric compounds cannot be distinguished
by MS . Eg. :- naphthalene vs. azulene