INTRODUCTION

The Gas chromatography consists of two types

ie., gas liquid chromatography(GLC) and gas
solid chromatography(GSC). In both the cases
gas is the mobile phase and either solid or liquid
is used as the stationary phase.
GSC: As the number of stationary phases
available are limited this technique is not widely
used. The principle of separation involved is
adsorption.
GLC: This is the widely used technique in which
partition is the principle of separation
Instrumentation : The instruments in gas
chromatography can be listed as follows:
 Carrier gas
 Flow regulators and flow meters
 Injection devices
 Columns
 Temperature control devices
 Detectors
 Recorders and integrators
Carrier Gas: The efficiency of chromatographic
separation depends on the choice of carrier gas.
The gases which are widely used for this
purpose are hydrogen, helium, nitrogen and
argon.
Hydrogen: It is mostly used in thermal
conductivity and flame ionisation detector
Merits: Better thermal conductivity
Low density
Demerits: Inflammable
It reacts with unsaturated compounds
• Helium: It is proven good when used with
thermal conductivity detector
Merits: Good thermal conductivity
Demerits: Expensive.

Nitrogen: It has reduced sensitivity but

inexpensive.
• Flow regulators and flow meters: Flow
regulators are very much necessary to deliver
gas with uniform pressure and flow rate, as
they are stored at high pressure.
Flow meters are used for the measurement of the
flow of the carrier gas. The instruments used
are
 Rotameter
 Soap bulb meter
Rotameter: It is precalibrated instrument which
is placed before the inlet of the column made
of ordinary glass tube baring a float held on to
a spring
Soap bulb meter: It is similar to that of
rotameter but, a soap bulb indicates the flow
rate
• Gas enters through an inlet of glass tube at the
bottom. Soap solution is stored in a rubber
bulb. The bulb is pressed gently a drop of the
soap solution is converted into bubble by the
pressure of carrier gas and travel up words.
Injection devices : A solid, liquid or a gaseous
substance can be introduced into the column.
Valves are used to introduce gases
Loops or septums are used for the liquids. The
loops or septums are made of high quality
rubbers attached to instrument
The rubber used is of silicon rubber which can
withstand high temperature of preheating and
also repeated injections over a period of time

Column: These are very important part of the

instruments and made of glass and stainless
steel. The stainless steel are advantageous in
one way that there is no fear of fragility but, it
have got the tendency to react with some type
of samples and in such cases glass columns are
used.
Columns are classified as :
Depending on the usage:
 Analytical column: They are of 1-1.5mts and
3-6mm diameter. They are packed columns
available in both glass and stainless steel. It
allows small quantities of samples to be
loaded.
 Preparative column: They are larger than
analytical columns as large amount of sample
is loaded. They range from 3-6mts length and
6-9mm external diameter.
Depending on its nature:
Packed column: Columns are available
commercially in packed manner. These
columns are available from low range polarity
to high range of polarity.
Open tubular column or golay column or
capillary column: These are long capillary
tubings of 30-90mts length and internal
diameter of 0.025-0.075mm. They are coil
shaped made up of stainless steel
The stationary phase is coated with thin
film of 0.5- 1.0µ on the inner walls
Merits: Offer least resistant to carrier gas (so,
more efficient than packed columns, offering
more resistant to carrier gas)
Demerits: More sample cannot be loaded
SCOT(Support coated open tubular column):
These are made by depositing a micron size
porous layer of support material on the inner
wall of the capillary column and then coated
with thin film of liquid phase. This is an
improved version of golay column
Merits: Low resistant to carrier gas.
More sample loading capacity.
Detectors: These more important in the gas
chromatography instruments. They are the
heart of apparatus. A detector has a property to
detect a pure carrier gas and an eluted
component.
Ideal characteristics of a detector
 It should be applicable to wide range of
samples
 Rapid response
 High sensitivity towards small concentrations
 Less response to low concentrations and more
response to high concentrations (linearity)
 Temperature, flow rate or carrier gas
characteristics should not effect the response of
the detector
 Non destructive to the sample in case of a
preparative work
 Maintenance easy and simple
 Inexpensive
The commonly used detectors are:
 Thermal conductivity detector or
Katharometer(TCD)
 Flame ionisation detector(FID)
 Argon Ionisation detector (AID)
 Electron capture detector(ECD)
Thermal conductivity detector or
Katharometer(TCD) : The principle is based
upon thermal conductivity difference between
carrier gas and that of component. This
detector has two platinum wires of uniform
dimensions which form part of wheatstone
bridge. Through one of them, pure carrier gas
always flows through and through the other,
the effluents of the column passes. The two
platinum wires are heated electrically and
hence assume equilibrium conditions of
temperature and electrical resistance. When pure
carrier gas passes through both of them, there is
no difference in temperature or resistance and
hence a baseline is recorded. When a component
emerges from the column, it alters thermal
conductivity and resistance of the wire. Hence this
produces a difference in resistance and so
conductivity between wires, which is amplified
and recorded as a signal.
Examples:
H2 He N2 methane Hexane

32.7 33.9 5.2 6.5 3.0

Merits:
• It is applicable in most of the compounds.
• Linearity is good.
• As the sample is not destroyed it is used in
preparative scale.
• Inexpensive, easy maintenance and simple.
Demerits: Low sensitivity
It is affected by temperature and flowrate.
No absolute response it is only relative
Not applicable to biological samples
Flame ionisation detector: These are based on
the electrical conductivity of carrier gas. These
act as insulators at normal temperature and
pressure. But, they are conductive if ions are
present.
The hydrogen carrier gas or mixed with
nitrogen or argon is used to reach the tip of the
burner made up of platinum capillary, which acts
as cathode(electrode). The anode (electrode)
made up of silver gauze placed above the burner
tip. When pure gas flows alone no current flows
as there is no ionisation.
When the component is given from the column,
ions are produced due to ionisation by the
thermal energy of the flame which causes the
potential difference and causes a flow of current
which is amplified and recorded as signal.
Merits:
Micro gram samples are detected as it is
highly sensitive and also low background noise.
It is not affected by the changes in the flow
rate of carrier gas and water vapour.
It responses to most of the organic
compounds
Argon ionisation detector: This type depends
on the excitation of atoms to a metastable
state, using radioactive energy. This is done by
α and β particles obtained by radium-D and
90
Sr or titanium respectively. The argon atoms
are ionised by the these high energy particles
and are excited to metastable state. These
molecules colloid with the effluent molecules
and ionises them. These ions when they reach
the detector will cause an increase in current.
Thus the compounds can be detected.
Merit: It shows response to most of the organic
compounds.
Sensitivity is very high
Demerits: It shows only relative response
Linearity is poor
Water affects the sensitivity and it is reduced by
the halogens
Response is temperature dependent of the
detector usually less then 240⁰C, 1000V voltage
or less are necessary.
Electron capture detector: It consists of two
electrodes with column effluent passing
between them. One electrode emits electrons as
it is treated with radioactive isotope which as it
decays. This leads to production of secondary
electrons which are collected by the anode,
when 20V potential is applied between them .
All the secondary electrons are collected by
positively polarised electrode when carrier gas
alone flows. so, the steady base line is recorded.
Effluent molecules with affinity for electrons
capture these electrons as the pass through the
electrodes.
Hence, the steady state current is reduced. This
difference is amplified and recorded as output
signal.
The carrier gas used in this type depends
on the electron affinity of the compounds
analysed. In case of high affinity compounds
argon and in case of low affinity compounds
nitrogen, hydrogen helium or carbon dioxide
can be used as carrier gas.
Merits: Highly sensitive ie., nanogram quantities
are detected.
Demerits: High electron affinity compounds are
only detected
It cannot detected halogenated compounds,
pesticides etc.
Recorders: These are used to record the
response of the detector after amplification.
They also record the base line and all peaks
along with time.
Merits: Retention time of all the peaks is found
out.
Demerits:Area of individual peaks is not known
Integrators: These are improved type then the
recorders with some data processing
capabilities. They record the individual peaks
with retention time, height, and width of peaks,
peak area, percentage of area etc. These give us
more information on the peaks than the
recorders.
 Factors affecting the choice of carrier
gas:
• Inertness: The carrier gas used should be
highly inert and should not show any affect on
the compound to be analysed
• Suitable to detector : The carrier gas used
should be suitable to the detector. eg: Helium
is used only with the thermal conductivity
detector (TCD) and not with others.
 High purity: The carrier gas used should be
pure form as its impurities may interfere with
the peaks.
 Easily available: The carrier gas used should
be easily available because any scarcity may
prolong the timed of research and also it may
effect the stability of the samples prepared.
 Cheap: the carrier gas should always be
inexpensive.
 Risk: the carrier gas used should always have
less risk of explosion and fire hazards.
 Performance: It should give the best column
performance.
TEMPERATURE PROGRAMMING IN G.C

• Preheaters: These are used to convert the

sample into vapour form and then mixed with
mobile phase or carrier gas. These are present
along with the injection devices and when the
liquid sample is injected they convert it into
vapour form
Thermostatically controlled oven: Principle of
separation is partition. Partition Co-efficient is
the ratio of concentration of solute distributed
between two immiscible liquids. Temperature
regulation of the column is highly necessary as
the partition coefficient and solubility of solute
depends on it which is highly essential for
efficient separation. So, both the column and
injection devises should be maintained at a
particular temperature.
There are two types of programming available in
G.C for temperature regulation.
Isothermal programming: The temperature
throughout the process of separation is maintained
same.
Linear programming: In this process the oven is
heated over a period of time linearly. In this initially
150⁰C to 200⁰C at the end of separation with an
increase in temperature at the rate of 5⁰C/minute.
This type of linear programming is employed when
there is a mixture of high and low boiling point
compounds. This method is efficient for separation
of such complex mixtures.
Pyrolysis and derivatization in G.C: It is a
method of chemical analysis in which the sample
is heated to decomposition to produce smaller
molecules that are separated by gas
chromatography.
Pyrolysis is the thermal decomposition of
materials in an inert atmosphere or a vacuum.
The sample is put into direct contact with
platinum wire or quartz sample tube and heated
rapidly to 600 to 1000⁰C. There are three
different heating techniques.
They are
a) Isothermal furnace
b) Inductive heating
c) Resistive heating
Using platinum filaments
Derivatisation in G.C: It is the process of
improving the process of separation or
detection by detector.
It is of two types
 Pre column derivatisation.
 Post column derivatisation.
Pre column derivatisation: It increases the
volatility and thermostability of the
derivatives. Improved separation and less
tailing is seen.
The conditions in which this technique is followed:
 In case of less volatile componenets
 Thermolabile or heat sensitive compounds
 To reduce tailing
 To improve separation factor
Post column derivatisation: This is used to improve
the response shown by the detector. The
components may not be detected unless the
derivatisation is done. These are converted in such
a way the their ionisation or ability towards
electrons is increased. This is an “On-Line”
detection technique where the flowrate is neither
APPLICATIONS:
Qualitative analysis: This is done by comparing the
retention time of sample and standard, if there is
any deviation then the sample and standard are not
of the same compound.
Checking the purity of a compound: It is done by
comparing the standard and sample chromatogram
and if any additional peaks are present it is due to
the impurities
Presence of impurity: This can be seen by the
presence of additional peaks when compared with
a standard and sample. The percentage of
Quantitative analysis :It is used to determine the
quantity of a component by several methods
a) Direct comparison method: The quantity of the
sample can be determined by comparing the peak
areas which are injected separately.
Area of the peak= Peak height X Width of the peak at
half height
A1/A2 = αW1/W2
Where A1 and A2 are peak area of sample and
standard
W1 and W2 are weight or concentration of sample
and standard
Α is the response factor
b) Calibration curve method: In calibration
curve method, standardization of varying
concentration are used to determine their peak
areas. A graph of peak area Vs concentration of
the drug is plotted, from the peak area of
unknown sample, by intrapolation, the
concentration of the sample can be
determined. The errors if any are minimised.
c) Internal standard method: Compound with
similar retention characteristics is used. A
known concentration of the internal standard is
added to the standard solution and sample
solution whose concentration is not known.
The chromatogram is recorded and the peak area
ratio of standard and internal standard is
determined. By using the peak area ratio of
sample and internal standard, the concentration
of unknown solution is determined. This
advantageous when more extraction steps are
involved and the sample matrix is complex
Multiple analysis or Determination of mixture
of drugs: It is similar to that quantification of
single drug , multiple component analysis is
also done. The quantity of the each component
is determined by any one of the above method
Isolation and identification of drugs or
metabolites in urine, plasma, serum etc can be
carried out.
Isolation and identification of mixture of
components like acids, plant extracts volatile
oils, etc.