GAS CHROMATOGRAPHY

MASS SPECTROMETRY
(GC-MS)
 Introduction to GC MS
Gas chromatography–mass spectrometry (GC MS) is an analytical
method that combines the features of gas-chromatography and mass
spectrometry to identify different substances within a test sample.
Gas chromatography is a technique capable of separating, detecting
and partially characterizing the organic compounds particularly
when present in small quantity.
Mass spectroscopy provides some definite structural information
from in small quantity.
The separation and identification of the components of complex
natural and synthetic mixture are achieved more quickly than any
other technique with less sample
1. Pneumatic controls 6. Ion Source
2. Injector 7. Mass Analyser
3. Oven 8. Detector
4. Column 9. Vacuum System
5. Interface 10. Control Electronics
 Introduction to GC
• Chromatography is a physical method of separation in which the
components to be separated are distributed between two phases. One of
which is stationary (stationary phase) while the other (the mobile phase)
moves in a definite direction.
•Elution chromatography is a procedure in which the mobile phase is
continuously passed through or along the chromatographic bed and the
sample is fed into the system as a finite slug. EX: Gas Liquid
Chromatography(GC)
•Gas chromatography is a separation method in which the components
of a sample partition between two phases one of these phases is a
stationary bed with a large surface area, and the other is a gas which
percolates through the stationary bed.
•Gas chromatography differs from other forms of chromatography in
that the mobile phase is a gas and the components are separated as
vapours.
•It is thus used to separate and detect small molecular weight
compounds in the gas phase.
•The sample is either a gas or a liquid that is vaporized in the injection
port. The mobile phase for gas chromatography is a carrier gas, typically
helium because of its low molecular weight and being chemically inert.
•The pressure is applied and the mobile phase moves the analyte
through the column. The separation is accomplished using a column
coated with a stationary phase.
In GC the main principle of separation is partition.

The equilibrium for gas chromatography is partitioning, and the

components of the sample will partition (i.e. distribute) between the two
phases: the stationary phase and the mobile phase. Compounds that have
a greater affinity for the stationary phase spend more time in the column
and thus elute later and have a longer retention time (Rt) than samples
that have a higher affinity for the mobile phase.
Affinity for the stationary phase is driven mainly by intermolecular
interactions and the polarity of the stationary phase can be chosen to
maximize interactions and thus the separation.
Ideal peaks are Gaussian distributions and symmetrical, because of the
random nature of the analyte interactions with the column.
The separation is hence accomplished by partitioning the sample between the
gas and a thin layer of a non-volatile liquid held on a solid support. A sample
containing the solutes is injected into a heated block where it is immediately
vaporized and swept as a plug of vapour by the carrier gas stream into the column
inlet. The solutes are adsorbed by the stationary phase and then desorbed by a
fresh carrier gas.
 The process is repeated in each plate as the sample is moved toward the outlet.
Each solute will travel at its own rate through the column. Their bands will
separate into distinct zones depending on the partition coefficients, and band
spreading.
 The solutes are eluted one after another in the increasing order of their kd, and
enter into a detector attached to the exit end of the column. Here they register a
series of signals resulting from concentration changes and rates of elution on the
recorder as a plot of time versus the composition of carrier gas stream.
The appearance time, height, width, and area of these peaks can be measured to
yield quantitative data.
Gas chromatography is mainly composed of the following parts:
Carrier gas in a high-pressure cylinder with attendant pressure regulators and
flow meters
Helium, N2, H, Argon are used as carrier gases.Helium is preferred for thermal
conductivity detectors because of its high thermal conductivity relative to that of
most organic vapors.
N2 is preferable when a large consumption of carrier gas is employed.
Flow rate is adjusted by means of a needle valve mounted on the base of the
flow meter and controlled by capillary restrictors. The operating efficiency of the
gas chromatograph is directly dependant on the maintenance of constant gas
flow.
Sample injection system
Liquid samples are injected by a micro syringe with a needle inserted through a
self-scaling, silicon-rubber septum into a heated metal block by a resistance
heater. Gaseous samples are injected by a gas-tight syringe or through a by-pass
The separation column
The heart of the gas chromatography is the column which is made of
metals bent in U shape or coiled into an open spiral or a flat pancake
shape.
Copper is useful up to 2500
Swege lock fittings make column insertion easy.
Several sizes of columns are used depending upon the requirements.
Liquid phases
An infinite variety of liquid phases are available limited only by their
volatility, thermal stability and ability to wet the support.
No single phase will serve for all separation problems at all temperatures.
Non-Polar – Parafin, squalane, silicone greases, apiezon L, silicone gum
rubber. These materials separate the components in order of their boiling
points.
Intermediate Polarity – These materials contain a polar or polarizable
group on a long non-polar skeleton which can dissolve both polar and non-
polar solutes. For example. diethyl hexyl phthalate is used for the
separation of high boiling alcohols.
Polar – Carbowaxes – Liquid phases with a large proportion of polar
groups. Separation of polar and non-polar substances.
Hydrogen bonding – Polar liquid phases with high hydrogen bonding e.g.
Glycol.
Specific purpose phases – Relying on a chemical reaction with solute to
achieve separations. e.g AgNO3 in glycol separates unsaturated
hydrocarbons.
Supports
The structure and surface characteristics of the support materials are
important parameters, which determine the efficiency of the support and the
degree of separation respectively.
The support should be inert but capable of immobilizing a large volume of
liquid phase as a thin film over its surface.
The surface area should be large to ensure the rapid attainment of
equilibrium between stationary and mobile phases.
Support should be strong enough to resist breakdown in handling and be
capable of packed into a uniform bed.
Diatomaceous earth, kieselguhr treated with Na 2CO 3 for 900 0 C causes
the particle fusion into coarser aggregates.
Glass beads with a low surface area and low porosity can be used to coat
up to 3% stationary phases.
Porous polymer beads differing in the degree of cross-linking of styrene
with alkyl-vinyl benzene are also used which are stable up to 2500
GC Detectors
1. Pneumatic controls 6. Ion Source
2. Injector 7. Mass Analyser
3. Oven 8. Detector
4. Column 9. Vacuum System
5. Interface 10. Control Electronics
Image of GC-MS
 Injector
A GC syringe penetrates a septum to inject sample into
the vaporization camber
Instant vaporization of the sample, 280 C
Carrier gas transports the sample into the head of the
column
Purge valve controls the fraction of sample that enters the
column
Columns

Cross section Columns

 Coupling of GC to MS
For Coupling the interface b/w the GC&MS plays an important role in
the overall efficiency of the instrument.
Types of interfaces:
Capillary direct interface:
Today most GC-MS systems use capillary columns &
fused silica tubing permits an inert, high efficiency, direct
transfer between the 2 systems.
Flow rates is 5ml/min.

Jet separator (packed column):

The separator consist of two glass tubes aligned with a
Small distance between them.
Carrier gas entering from the GC column is pumped away
by a separate vacuumed system.
The larger sample molecules maintain their momentum
&pass preferentially in to the second capillary.
Sample enrichment occurs & the initial atmospheric
pressure is reduced.
Watson & Biermann effusion separator:
 It consists of a sintered glass tube .
 The carrier usually Helium, passes preferentially
through the sintered glass tube & the effluent in
concentrated by a factor of up to 100.
 The gas flow rates in the order of 20-60ml/min.
The most common form of ionization is EI.
Electrons are produced by tungsten filament.
These electrons accelerated towards the ion source chamber.
The electrons require an energy equal to the voltage B/W the
filament & ion source chamber.
70 ev is commonly used.
A proportion of electron beam will strike the electron trap
producing trap current.
A permanent magnet is positioned across the ion chamber to produce a
magnetic flux in parallel to the electron beam.
A (+)ve ion repelle voltage & (-)ve ion excitation voltage works to gather
to produce an electric field in the source chamber.
 Such that ions leaves through ion exit slit.
The ions are directed through the various focusing & centring lenses
are focused on to the source exit slit.
In CI a reagent gas methane or ammonia or isobutene
are introduced into the mass spectrometer.
The reagent gas will interact with the electron to produce
radical electrons.
EG;
R + e -----------> R+ + 2e
 Negative ion chemical ionization(NICI):

In NICI a r reagent gas is used & the electrons collide with it so
that their energies are reduced to 10Ev.
Molecules with a high affinity for electrons are able to
capture these low energy thermal electrons.
This is known as NICI but it does not involved in the
formation of a chemical adduct.
 Mass Analyzers
• Mass Analyzers :
A mass analyzer is the component of the mass spectrometer that takes
ionized masses and separates them based on charge to mass ratios and
outputs them to the detector where they are detected and later
converted to a digital output.

Following type of mass analyzer used for LC/MS

• Quadrupole
• Time-of-flight
• Ion trap
• Fourier transform-ion cyclotron resonance
(FT-ICR or FT-MS)
1) Quadrupole :
The quadrupole consists of four parallel metal rods. Each opposing rod
pair is connected together electrically, and a radio frequency (RF)
voltage with a DC offset voltage is applied between one pair of rods and
the other. Ions travel down the quadrupole between the rods. Only ions
of a certain mass-to-charge ratio will reach the detector for a given ratio
of voltages, other ions have unstable trajectories and will collide with
the rods. This permits selection of an ion with a particular m/z.
Quadrupole tend to be the simplest and least expensive mass analyzers.
Quadrupole mass analyzers can operate in two modes:
• Scanning (scan) mode.
• Selected ion monitoring (SIM) mode
•In scan mode, the mass analyzer monitors a range of mass-to
charge ratios.
• In SIM mode, the mass analyzer monitors only a few
mass-to-charge ratios.
SIM mode is significantly more sensitive than scan mode but
provides information about fewer ions. Scan mode is typically
used for qualitative analysis or for quantitation when all analyte
masses are not known in advance. SIM mode is used for
quantitation and monitoring of target compounds
 Multiple quadrupole
A linear series of three quadrupoles is known as a 
triple quadrupole mass spectrometer. The first (Q1) and third (Q3)

quadrupoles act as mass filters, and the middle (q2) quadrupole is

employed as a collision cell. This collision cell is an RF-only quadrupole
(non-mass filtering) using Ar, He, or N2 gas (~10−3 Torr, ~30 eV) for

collision induced dissociation of selected parent ion(s) from Q 1.

Subsequent fragments are passed through to Q3 where they may be

filtered or fully scanned.
This process allows for the study of fragments that are useful in
structural elucidation by tandem mass spectrometry. For example, the
Q1 may be set to 'filter' for a drug ion of known mass, which is

fragmented in q2. The third quadrupole (Q3) can then be set to scan the
entire m/z range, giving information on the intensities of the fragments.
Triple quadrupole mass spectrometer
Quadrupole Mass
Analyzer
 2) Time-of-flight
In a time-of-flight (TOF) mass analyzer, a uniform electromagnetic
force is applied to all ions at the same time, causing them to
accelerate down a flight tube. Lighter ions travel faster and arrive at
the detector first, so the mass-to-charge ratios of the ions are
determined by their arrival times. Time-of-flight mass analyzers have
a wide mass range and can be very accurate in their mass
measurements.
Time of flight mass analyzer
 3) Ion trap
An ion trap mass analyzer consists of a circular ring
electrode plus two end caps that together form a chamber. Ions
entering the chamber are “trapped” there by electromagnetic
fields. Another field can be applied to selectively eject ions from
the trap. Ion traps have the advantage of being able to perform
multiple stages of mass spectrometry without additional mass
Analyzers
4) Fourier transform-ion cyclotron resonance
(FT-ICR)
An FT-ICR mass analyzer (also called FT-MS) is
another type of trapping analyzer. Ions
entering a chamber are trapped in circular
orbits by powerful electrical and magnetic
fields. When excited by a radio-frequency (RF)
electrical field, the ions generate a time
dependent current. This current is converted
by Fourier transform into orbital frequencies of
the ions which correspond to their mass-to
charge ratios
 Detectors :
once the ions have passed the mass analyser they have to be detected
and transformed into a usable signal. The detector is an important
element generating secondary electrons, which are further amplified, or
by inducing a current (generated by moving charges).

Type of DETECTORS
Photo graphic plates
Faraday cup
Channel electron multipliers
Electron multiplier
Scintillation detectors
Photo graphic plates:
It is used as it is capable of higher
resolution and speeder than
electronic devices. i.e. it can detect ions
of all the masses and provide
a reverse geometry analyzer.
Faraday Cup detector
Faraday cups are perhaps the oldest and most simple detectors used
in mass spectrometers.  At a very base level, a Faraday cup is a piece
of metal that resides in the mass spectrometer's vacuum chamber
and is connected to the instrument's electronics.  Electric fields are
utilized to push ions into the piece of metal.  When ions strike the
metal, electrons flow through the circuit to meet the ions and
neutralize them at the Faraday cup's surface.  This current can be
measured and amplified by the instrument's electronics.  The amount
of current is proportional to the number of ions hitting the Faraday
cup.
Electron multipliers:
In this the current can be measured so accurately by just one ion
strikes the detector can be measured i.e. when an ion strikes the
surface of electron multiplier two electron are ejected. This process
continues until the end of electro multiplier end is reached and
electric current is analyzed and recorded with electron multiplier
surface. Equation describe is 2n
Where n= no of collisions with electron multiplier surface.
 APPLICATIONS
Elucidation of the structure of organic & biological molecules.
 Impurity profiling of pharmaceuticals.
 Identification of components in thin layer & paper chromatograms.
 Identification of drugs of abuse & metabolites of drugs of abuse in
blood, urine & saliva.
 Testing for the presence of the drugs in blood in race horses & in
Olympic athletic (in forensic GC-MS).
 Analyzer of aerosol particles.
 Determination of pesticide residues in food.
 Polymer characterization (pyrolysis methods combined GCMS).
 Drug monitoring & toxicology studies.
Synthesis of Propanolol
 Other Applications
GC-MS is becoming the tool of choice for tracking organic pollutants in the
environment.
GC-MS can analyze the particles from a human body in ord to help link a
criminal to a crime.
GC-MS especially useful here as samples often contain very complex
matrices &results used in court.
GC-MS used for detection of illegal narcotics & may eventually supplant
drug-sniffing dogs.
It’s also commonly used in forensic toxicology to find drugs
&poisons in biological specimens of victims .