The document provides an overview of gas chromatography (GC), detailing its principles, mobile and stationary phases, instrumentation, derivatization, advantages, disadvantages, and applications. GC is a widely used technique for separating components of a mixture by partitioning them between a gaseous mobile phase and a stationary phase, with applications in fields such as pharmaceuticals, agriculture, and forensics. It highlights different types of columns, detectors, and sample preparation methods, as well as the process of derivatization to enhance analyte properties for better analysis.

1. GAS CHROMATOGRAPHY
(GC)
NAME- Hetav Patel
MASTER OF PHARMACY
DEPARTMENT OF PHARMACEUTICS
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2. Content
• Introduction
• Principle
• Mobile phase
• Stationary phase
• Instrumentation of GC
• Derivatization of GC
• Advantages
• Disadvantages
• Applications
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3. Introduction to Chromatography
• Chromatography is a procedure for resolving A multicomponent
mixture of minor or major constituents into its individual
fractions.
• Examples of different types of chromatography.
1. Paper chromatography:- this technique is a type of partition
chromatography in which the substance are distributed
between 2 liquids in which one is the stationary phase which
is held in the fibers of the paper and called the stationary
phase the other is moving liquid or developing solvent and
called the moving phase.
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4. 2. Thin layer chromatography:- thin layer chromatography is
similar to paper chromatography except that a thin layer of some
inert material such as Al2O3, MgO, SiO2 is used as the substrate
instead of paper.
3. Gas chromatography:- in gas chromatography a moving gas
phase is passed over a stationary phase and separate the
mixture components this technique is similar to the of liquid
chromatography with The only exception that in the former
moving gas is used as the mobile phase while the later it is a
liquid.
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5. Introduction
• P. Martin and A. T. James First time used the gas
chromatography technique in 1952 for separating long chain
fatty acid.
• GC is one of the most widely used chromatographic technique
which uses inert gas as MP.
• in GC, the components of sample are separated by being
partitioned between gaseous MP and solid/liquid SP.
• Types of GC:-
1. Gas liquid chromatography(GLC)
2. Gas solid chromatography (GSC)
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6. Principle
• So, partition co-efficient is the main factor for the separation.
• In gas chromatography the substance to be analyzed is
partitioned between the mobile phase and stationary phase
during the separation the sample is vaporized and carried
through the column by the mobile phase.
• the different components get separated based on their vapor
pressure and affinity for the stationary phase.
• the affinity of component towards the stationary phase is terms
as partition coefficient.(K)
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7. Principle
• GLC :- partition co-efficient
• GSC :- adsorption co-efficient
• In GSC, the analytes of the mixture distributes themselves
between the gas phase and solid adsorbent.
• The difference in the adsorptive behavior causes separation.
• In GLC, the analytes of mixture distributes themselves between
gas phase and liquid phase according to their partition co-
efficient.
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8. Mobile phase
• State :- gaseous
• It is also known as carrier gas as it is used to transport and carry
sample components through the column.
• Ideal characteristics of mobile phase :-
1. It should be inert so that it will not react with SP and sample.
2. Comfortable with detectors
3. Highly pure
4. Easily available and less expensive
5. Easy to handle
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9. Stationary phase
• For GSC:- different types of adsorbents.
• For GLC:- different types of liquid supported by solids.
• Generally, GC columns are made up of glass or metal tubing.
• GC columns are lengthy ( cm to m ) as compared to HPLC columns.
• Mainly 2 types of column
1. Packed column
2. Capillary column
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10. Stationary phase
Packed column
Capillary
column
Packed column
with solid
particles
Open tubular
column
Wall coated
open tubular
column(WCOT)
Support coated
open tubular
column(SCOT)
Porous layer
open tubular
column(PLOT)
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11. Instrumentation of GC
• Schematic diagram of GC:-
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12. • GC generally consists of following components:-
1. Gas source and regulation system
2. Sample injector system
3. Column
4. Temperature program
5. Detectors
6. Readout device
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13. 1. Gas source and regulation system
• It consist of gas cylinder and generators to produce carrier gas.
• The flow rate and pressure of gas is controlled by pressure
regulator.
• Gas must be purified by passing it through suitable adsorbent to
avoid any undesirable chemical changes to sample, SP and
detector.
• Contaminants present in MP are air and oxygen, hydrocarbons
and water vapor.
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14. • Considerations for selection of carrier gas:-
1. It should be inert.
2. It should be suitable for the detector employed and the type of
sample analyzed.
3. It should be readily available in high purity.
4. It should give best column performance consistent with required
speed of analysis.
5. It should be cheap.
6. It should not cause the risk of fire and explosion hazard.
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15. • Most widely used carrier
gases are hydrogen,
helium, nitrogen and air.
Gas Properties
Hydrogen • Dangerous to use
compared to others.
• Has better thermal
conductivity and lower
density
• It may reacts with
unsaturated compounds
and that may cause
fire and explosive
hazards.
Helium • less dangerous but it is
expensive.
Nitrogen • It is inexpensive but has
less sensitivity.
Air • Only used when oxygen
in air is useful in
separation.
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16. 2. Sample injection system
• Liquid sample are generally introduced by hypodermic syringe
through self-sealing rubber septum into a small chamber and
the chamber is heated to cause flash evaporation.
• If the sample is solid, it must be dissolved in volatile liquids or
may be introduced directly if they can be liquefied.
• According to the nature of sample and column use different
methods of sample injection are employed.
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17. 3. Column
• It can be constructed of glass or metal tubing for analytical work
it has 4.8 millimeter diameter.
• It can be of any length from few centimeters to over 100 meter
and can be coiled, bent or straight.
• Maily two types of column used in GC.
I. Packed column
II. Capillary column
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18. I. Packed column
• SP is packed in glass, SS or nickel tubing to produce packed
column.
• Length:- 3 meter
• Inner diameter:- 1.6-9.5 mm
• Pore diameter:- 2-9 mm
• Particle size:- 100/120 mesh
• The columns are packed with absorbent like silica gel.
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19. II. Capillary column
• Made up of fused silica which is a very high purity glass.
• The degree of cross linking is higher than ordinary glass so that it
has higher strength.
• Outer wall is coated with polyimides to protect it from scratches.
• Internal diameter:- 1 mm or less.
Packed columns with solid particles:- also called as micro packed
column.
Open tubular column:- made up of SS, copper or glass.
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20. • There are three subtypes of the open tubular column:-
1. Wall coated open tubular columns (WCOT):-
• The wall is directly coated with the stationary phase at a film
thickness of 0.05-3 µm.
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21. 2. supported coated open tubular column(SCOT):-
• This column contain an absorbent layer of very fine solid
particle coated with the liquid phase.
• This column can hold more liquid phase and have a higher
sample capacity than the thin film of early wall coated open
tubular columns.
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22. 3. porous layer open tubular column(PLOT):-
• Porous layer is coated on inner wall of column.
• PLOT column contains A porous layer of solid adsorbents such
as alumina.
• It is well suited for the analysis of light, fixed gases and other
volatile compounds.
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23. 4. Temperature regulator
• It plays a crucial role in GC.
• In this system, a lower temperature is selected initially to
resolve early peaks and then temperature is increase to
push out the higher components.
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24. 5. Detectors
• All the detectors monitor the GLC column effluent by measuring the
changes in the composition arising from the variations in the eluted
compound.
• When the carrier gas alone is passing the give a zero signal.
• e.g.
i. thermal conductivity detector (TCD)
ii. Flame ionization detector (FID)
iii. Electron capture detector (ECD)
iv. Photo ionization detector (PID)
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25. i. Thermal conductivity detector (TCD)
• Principle:- based on thermal
conductivity of a gas which
controls the temperature and
thus resistance of wire.
• Construction:-
o Filament is made up of tungsten
or tungsten ally and coated with
gold.
o Filament is placed inside the
cavity of a metal block.
o DC current is supply is there to
heat the filament.
o The standard TCD consists of 4
filament placed within one metal
block.
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26. • Working of TCD:-
The filament is
heated by constant
DC current.
Due to resistence of
filament, it will be
heated at
constant temperatue.
Sample component
enter into detector.
It will decrease
thermal conductivity
so filament
retains more heat
Leads to rise in
temperature and
resistance too.
The change in
resistance is
measured.
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27. ii. Flame ionization detector (FID)
• Principle:- based on electrical
conductivity of glass if the
electrons are present in a gas
stream it become conductive and
the change of conductivity is
recorded in terms of electrical
current.
• Constuction:-
o Flame jet:- it is a capillary jet that
can produce a flame.
o Collector electrode:- just place
above the tip of the flame.
• There are inlets for hydrogen
gas, air and column effluent.
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28. • Working of FID:-
Column effluents and
hydrogen gas is enters
through inlet.
Hydrogen gas is mixed
with sample and passes
through the jet.
It is mixed with air and
burned in the flame.
When ionisable
materials present in the
sample, it will reach the
flame and burned to
form ions and electrons.
This will raise the
current flow which then
recorded.
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29. iii. Electron capture detector (ECD)
• Principle:- based on electron
affinity of compound which in
turn produce change in
current.
• Construction:-
oCathode(-):- radioactive
electrode. Tritium or nickel-63
oAnode(+):- collector electrode
which collects electrones.
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30. • Working of ECD:-
The radio isotope (-) emits high energy electrons continuously.
Electrons bombard
the carrier gas
produce thermal
electrons and
radicals.
Voltage is applied as
a sequence of narrow
pulses of the electron
capture cell.
It will allows the
collection of the
thermal electron in
the collector
electrode.
When carrier gas is
passed, it will create
standing current or
baseline current.
When sample
is passed, it
reacts with thermal
electron and produce
-ve ions.
Leads to decrease in
thermal electron.
Thus, decrease in
current which is
then recorded and
analyzed.
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31. iv. Photo ionization detector
• UV radiation is mainly used to ionize the sample components.
• This produces the ions are collected at an electrode of positive
charge.
• The change is current is measured.
• The ionization potential of the compound should be lower than
the 1 amp energy to give response.
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32. GC derivatization
• Definition:- derivatization is the process of chemically modifying
a compound to produce a new compound which has properties
that are suitable for analysis using gas chromatography.
• The goal is to form such a derivative which is less polar or more
volatile which is more readily analyzed by GC.
• Types of derivatization:-
oThey were generally three types of reaction:-
1. Silylation
2. Acylation
3. alkylation
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33. Silylation:-
• Silylation produces silyi derivatives which are more volatile, less
stable and more thermally stable.
• It is nucleophilic attack(SN2) reaction.
• In this reaction, active hydrogens are replaced
with Trimethyl silane.
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34. • Care should be taken as silylation reaction:-
a. Silylation reagents will react with water and alcohol first so
make sure that sample and solvent both are dry.
b. Solvent should be as pure as possible to eliminate excessive
peeks.
c. Silylating agents should not enter up to column.
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35. 2. Acylation:-
• Acylation reduces polarity of amino, hydroxyl and thiol groups.
• Acylating reagents target highly polar and
multifunctional compounds. (e.g. carbohydrates, amino acids)
• Acylating reagents:-
a. Acyl anhydride
b. Acyl halide
c. Acyl amide
• CARE:- the anhydride and halide reagents produces acid by-
products, which must be removes before GC analysis.
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36. 3. Alkylation:-
• It decreases molecular polarity by replacing active hydrogen
with an alkyl group.
• It target compounds with acidic hydrogen, such as phenols and
carboxylic acid.
• These reagents make ester, ether, alkyl, amines and
alkyl amides.
• Generally, used to convert organic acids into esters.
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37. Advantages
• It is a reliable technique and provide rapid analysis.
• it utilizes sensitive detectors.
• it is highly efficient and leads to high resolution.
• it requires small samples.(<1 ml)
• it provides high quantitative accuracy.
• it is well established technique with extensive literature and
applications.
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38. Disadvantages
• It is limited to volatile samples.
• it is not suitable for Thermolabile samples.
• it is not suited to preparative chromatography.
• it requires Ms. detector for structural elucidation of the analytes.
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39. Applications
• GC is an instrumental technique which is used in various fields
as a powerful tool of analysis.
1. Qualitative analysis:-
• GC is used for qualitative analysis in separation and
identification of mixtures by comparing the retention time of
sample with standards.
• It is also used to check purity of compounds in comparison with
standards.
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40. 2. Quantitative analysis:-
• GC performs quantitative analysis by calculating the area of peaks
from chromatogram.
• Area under elution peak = quantity of detected compound.
• It can be done by following method:-
I. Direct compression method
II. calibration curve method
III. international standard method
3. In agriculture:-
• It is a useful tool for the identification and determination of
pesticides.
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41. 4. In pharmaceutical analysis:-
• Used for drug analysis.
• Useful for the analysis of …
I. Antibiotics= gentamycin, neomycin
II. Anti-viral drugs= amantadine
III. Anti-cancer drugs= 5-flouriuracil
IV. Hypnotics and sedatives= barbiturates
5. In food industry:-
• For separation and identification of lipid, carbohydrates, proteins, flavors,
colorants and preservatives.
• Identification of vitamins, steroids and trace elements can be done.
• Used in determining free cholesterol in milk fat.
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42. 6. In forensic science:-
• For determination of drugs in blood sample mainly in sports activity.
• biological components such as blood plasma, serum and urine sample can be
analyzed for drug content.
7. In petroleum industry:-
• In analysis of crude petroleum products fraction, gasoline, waxes, LPG etc.
• Also use to separate hydrocarbons present in petroleum.
8. In biochemical and clinical:-
• Applicable to body components of all type.
• e.g. Blood gases, estrogen, vanillin etc.
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43. 9. In air monitoring:-
• Used for the determination of volume of organic compounds
in air such as Xylene, Toluene, Camene.
10. In cosmetics and perfume fields:-
• To determine the composition of various cosmetics.
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44. References
• Gurdeep r. Chatwal, sham k. Anand, instrumental method of
chemical analysis, Himalaya publishing house,2.673-2.708
• F. James holler, Stanley R. Crouch, Skoog west fundamentals
of analytical chemistry, 9th edition,887-909
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45. THANK YOU
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