Gas Chromatography

In gas chromatography, the components of a vaporized sample are separated by

being distributed between a mobile gaseous phase and a liquid or a solid
stationary phase held in a column.1 In performing a gas chromatographic
separation, the sample is vaporized and injected onto the head of a
chromatographic column. Elution is brought about by the flow of an inert gaseous
mobile phase. In contrast to most other types of chromatography, the mobile phase
does not interact with molecules of the analyte. The only function of the mobile
phase is to transport the analyte through the column.

GC is currently one of the most popular methods for separating and analyzing
compounds. This is due to its high resolution, low limits of detection, speed,
accuracy and reproducibility.

GC can be applied to the separation of any compound that is either naturally

volatile (i.e., readily goes into the gas phase) or can be converted to a volatile
derivative. This makes GC useful in the separation of a number of small organic
and inorganic compounds

Two types of gas chromatography are encountered: gas-liquid chromatography

(GLC) and gas-solid chromatography (GSC). Gas-liquid chromatography finds
widespread use in all fields of science where its name is usually shortened to gas
chromatography (GC). Gas-solid chromatography is based on a solid stationary
phase in which retention of analytes occurs because of physical adsorption. Gas-
solid chromatography has limited application because
of semipermanent retention of active or polar molecules and severe tailing of
elution peaks.

Gas chromatography is a separation technique in which the components of a

sample partition between two phases:
1. The stationary phase.
2. The mobile gas phase.
According to the state of the stationary phase, gas chromatography can be
classiffed in gas-solid chromatogpraphy (GSC), where the stationary phase is a
solid, and gas-liquid chromatography (GLC) that uses a liquid
as stationary phase. GLC is to a great extent more widely used than GSC.
  Instrumentation of GC
A simple GC system consists of:
1. Gas source (with pressure and flow regulators)
2. Injector or sample application system
3. Chromatographic column (with oven for temperature control)
4. Detector & computer or recorder

Fig:Block diagram of GC
 Fig : Details of GC

Carrier gas: He (common), N2, H2

Pinlet 10-50 psig
Flow = 25-150 mL/min packed column
Flow = 1-25 mL/min open tubular column
Column: 2-50 m coiled stainless steel/glass/Teflon
Oven: 0-400 °C ~ average boiling point of sample
Accurate to <1 °C
Detectors: FID, TCD, ECD, (MS)
Process that include in GC

 Volatile liquid or gas injected through septum into heated port

 Sample rapidly evaporates and is pulled through the column with carrier gas
 Column is heated to provide sufficient vapor pressure to elute analytes
 Separated analytes flow through a heated detector for observation
  Carrier Gas System
The mobile phase gas in gas chromatography is called the carrier gas and must be
chemically inert. Helium is the most common mobile phase, although argon,
nitrogen, and hydrogen are also used. These gases are available in pressurized
tanks. Pressure regulators, gauges, and flow meters are required to control the flow
rate of the gas. main purpose of the gas in GC is to move the solutes along the
column, mobile phase is often referred to as carrier gas.

The carrier gas is of lesser importance than the stationary phase as the carrier gas
does not interact with the analyte but should have the following characteristics.

1. Compatible with the stationary phase and the sample.

2. Allows good separation of the analytes by the column.
3. Is of the highest purity.
4. Must not contain particulate matter, water or other chemical constituents.
5. Is compatible with the detector used.
6. Should be readily available.
7. Should be cheap.
8. It should not cause the risk of fire or explosion hazard.
9. Should be chemically inert.

Q: Selection criteria of carrier gas in GC.

Q:Writedown the selection criteria of mobile phase in GC.

The carrier gas supply is also associated with peripheral equipment such as:
 pressure regulators
 pressure gauges, and
 flow meters.

Individual Carrier Gas

He:
 Most common and compatible with most detectors
 Better resolution (smaller plate heights)
 Solutes diffuse rapidly  smaller mass transfer term
N2:
 Lower detection limit for a flame ionization detector
  Lower resolution and solute diffusion rates
H2:
 Fastest separations
 Can catalytically react with unsaturated compounds on metal surfaces
 Can not be used with mass spectrometers Forms explosive mixtures with air
 Better resolution (smaller plate heights)
 Solutes diffuse rapidly  smaller mass transfer term

Diffusion coefficients follow: H2 > He > N2

Flow rate increases: N2 < He < H2
Carrier Gas or Mobile phase does not affect solute retention, but does affect:
1.) Desired efficiency for the GC System
- low molecular weight gases (He, H2)  larger diffusion coefficients
- low molecular weight gases  faster, more efficient separations

2.) Stability of column and solutes

- H2 or O2 can react with functional groups on solutes and stationary
phase or with surfaces of the injector, connections and detector
3.) Response of the detector
- thermal conductor requires H2 or He
- other detectors require specific carrier gas

Choice of Carrier Gas: Advantages and Disadvantages

 Sample Injection system

The volumes injected in GC are routinely in the range of 0.5–2 ml. The
most commonly used type of syringe; the usual syringe volumes are 5 and
10 ml. Injection generally occurs through a re-sealable rubber septum. The
injector port is held at 150–250 degree depending on the volatility of the
sample and direct injection of 0.1–10 ml of sample is made onto the head of
the column.
 1. “Sandwich” Injection

Separate sample with air bubbles and solvent

 Air bubble prevents depletion of most volatile compounds before sample

injection is complete (barrier between oven and sample during injection)
 Solvent is used to pushes out sample, but bubble prevents mixing
 Final air bubble pushes out solvent
 Gas-tight syringe is required for gas samples
 Injection volume is typically 0.1-2 mL
Injection port

 Inject rapidly ( < 1s) through septum into evaporation zone

 Injector temperature is kept high (350oC) for fast evaporation
 Rapid gas flows carries sample to mixing chamber for complete vaporization
and complete mixing before entering column

2. Split Injection /Splitless injection

i. Delivers only 0.2-2% of sample to the column
1. Split ratio of 50:1 to 600:1 (sample discarded)
ii. For samples where analytes of interest are >0.1% of sample
1. Best resolution is obtained with smaller amount of
sample
2. ≤ 1 mL with ≤ 1 ng of each compound (0.5 mL of gas
volume)
iii. Not quantitative, split not constant
 Splitless Injection
“Solvent trapping” significantly improves the performance of splitless injections
 Initial lower temperature of column during injection keeps larger volume
into a narrow band
 Chromatography is initiated by raising column temperature
 Cold trapping – condense solutes in narrow band at the beginning of column
by using an initial temperature 150oC below boiling points of solutes of
interest

3.On-column Injection
 Delivers ~100% of sample to the column
 For samples that decompose above their boiling points
 Solution injected directly on column
- Warming column initiates chromatography

 Chromatographic column/Column Configurations

and Column Ovens
Gas chromatographic columns are usually between 1 and 100 meters long.
The columns in gas chromatography are of two general types:
1. Packed columns
2. Capillary(open-tubular) columns.
In the past, the vast majority of gas chromatographic analyses used packed
columns. For most current applications, packed columns have been replaced
by more efficient and faster capillary columns.
 There are two main types of supports used in GC:
Packed columns
 large sample capacity
 preparative work
 Greater sample capacity
 Broader peaks, longer retention times and less resolution
- Improve resolution by using small, uniform particle sizes

 The major advantage and use is for large-scale or preparative purification

 Industrial scale purification maybe in the kilogram or greater range
A packed column is dense and evenly packed by solid support. The solid support
usually has a liquid stationary phase bonded to it. The solid support allows for the
liquid stationary phase to be exposed to the maximum amount of the mobile phase.
The solid support and stationary phase must be inert at high temperatures and
allow for the mobile phase to be evenly distributed as it moves through the column.

The packed columns are shorter in length and wider in diameter than the open
tubular columns. The diameter of a packed column is usually between 2 and 4 mm.
Packed columns are typically 1 to 5 meters long and also form a helical shape.The
packing particles typically have a diameter of 100 to 250 micrometers. Micro-
packed columns are packed capillary tubes and are packed with the same material
as larger packed columns. The most common stationary phase used for packed
columns is diatomaceous earth (diatomite). Diatomite is made up of diatom
(single-celled algae) skeletons. The skeletons are composed of mostly silica, and
small quantities of alumina and metallic oxides. Other popular stationary phases
are pure silica (SiO2) and alumina (Al2O3). Alumina is great for separating
aromatic hydrocarbons

Capillary (open-tubular) columns

 Commonly used in GC
 Higher resolution, shorter analysis time, and greater sensitivity
 Low sample capacity
 Increasing Resolution
- Narrow columns  Increase resolution

- Resolution is proportional to N , where N increases directly with column

length

 higher efficiency
 smaller sample size
When the stationary phase is uniformly distributed on the interior surface of
column it is called an open tubular (capillary) column.

Open tubular columns are longer, smaller in diameter, and more efficient than
packed columns. Open tubular columns have less flow resistance which allows for
them to be longer and have a lot of theoretical plates. Capillary columns are
between 3 and 100 meters long and form a helical shape.

The most common stationary phases used for open tubular columns are
polysiloxanes. Polysiloxanes are silicon atoms which have attached oxygen and R
groups. The R groups can vary, which makes polysiloxanes very versatile .

There are three types of open tubular columns:

1. Wall-coated (WCOT)
2. Support-coated (SCOT)
3. Porous-layer (PLOT).

WCOT is the most popular type of open tubular column. The wall coated open
tubular column consists of a capillary tube with its interior surface coated in a tiny
layer of stationary phase. The most common type of wall coated open tubular
column used is fused-silica, because it is stronger, inert, reliable, easy to use, and
flexible. Fused silica capillary tubes are made from purified silica that has a small
quantity of metal oxides dispersed throughout the silica. The fused silica column
also has a layer of polyimide on the outside of the column, which makes the
column flexible and extends the life of the column. Wall-coated open tubular
columns can also be made out of plastic, glass, stainless steel, aluminum, or
copper. A support-coated open tubular column has a thin layer (approximately 30
µm) of liquid support matter. This type of open tubular column has a greater
amount of stationary phase than the wall coated column, so it can handle a larger
quantity of sample.
A porous-layer open tubular (PLOT) column is very similar to a support-coated
open tubular column. The only difference between the two types of columns is that
a PLOT does not have a liquid stationary phase. PLOT columns are used for gas
solid chromatography. PLOT columns have a solid layer of carbon, molecular
sieves, cyclodextrins, inorganic oxides, or porous polymers, coating the inner wall
of the column. PLOT columns can be up to 100 meters long. The inner diameter of
a PLOT column is between 0.25 and 0.53 mm. The stationary phase coating is
between 5 and 50 micrometers thick.
 Q;Why do open tubular column yield greater solute resolution than packed
columns?

 Open tubular column have a lower resistance to gas flow, so

longer columns can be used without increasing solute retention
times.
 Open tubular columns eliminate the multiple path term from the
van deemter equation.
 Open tubular columns have greater sample capacity.

Capillary vs. Packed Columns

Capillary Columns Packed Columns
– Higher R – Low R
– Smaller H ; high N – Larger H, low N
– fast – Slow
– Greater sensitivity – Greater sample capacity
– analytical – Lower cost
– Smaller sample capacity – More rugged
– Higher cost/column – preparative
– Columns fragile

Column type Packed column Capillary column

History First type of GC column Modern technology.
used Today most
GC applications are
developed
using capillary columns

Composition Packed with silica Not packed with

particles onto particulate ma-
which the stationary terial. Made of
phase is chemically
coated. treated fused silica
covered with
thin, uniform liquid
phase films.
Eficiency Low high
Outside diameter 2-4mm 0.4mm
 Column length 2-4 meter 15-30 meter
Advantages Lower cost , larger Faster , better for
sample complex mixtures
Table 1: A summary of the differerences between a packed and a capillary column.

 GC Detectors

A device used to detect each component of the sample as it appears at the exit of
the packed column. The intensity or area of the signal indicates the quantity of the
eluted component. The detector responds to a physicochemical property of the
analyte, amplifies this response and generates an electronic signal for the data
system to produce a chromatogram.

Many different detector types exist and the choice is based mainly on application,
analyte chemistry and required sensitivity – also on whether quantitative or
qualitative data is required.
Detector choices include:
 Flame Ionization (FID)
 Electron Capture (ECD)
 Flame Photometric (FPD)
 Nitrogen Phosphorous (NPD)
 Thermal Conductivity (TCD)
 Mass Spectrometer (MS)

Characteristics of the Ideal Detector

The ideal detector for gas chromatography has the following characteristics:
 Adequate sensitivity. In general, the sensitivities of present-day detectors lie
in the range of 10–8 to 10–15 g solute/s.
 Good stability and reproducibility.
 A linear response to solutes that extends over several orders of magnitude.
 A temperature range from room temperature to at least 4008C.
 A short response time that is independent of flow rate.
 High reliability and ease of use. To the greatest extent possible, the detector
shouldbe foolproof in the hands of inexperienced operators.
 Similarity in response toward all solutes or, alternatively, a highly
 predictable and
 selective response toward one or more classes of solutes.
 Nondestructive of sample.

 Flame Ionization Detector (FID)

 Most common detector for GC

 In an FID, effluent from the column is directed into a small air-hydrogen
flame. Most carbon atoms (except C=O) produce radicals (CHO+) in the
flame: CH + O→ CHO+ + e-
 Electrons are used to neutralize the CHO+ atoms and the ions are collected
at an electrode to create a current to be measured. This current is
proportional to the number of molecules present.
 The ionization of carbon compounds in the FID is not fully understood,
although the number of ions produced is roughly proportional to the number
of reduced carbon atoms in the flame.
 universal” detector capable of measuring the presence of almost any organic
and many inorganic compound
Advantages:
1.universal detector for organics
2.does not respond to common inorganic compounds
3.mobile phase impurities not detected
4.carrier gases not detected
5.limit of detection: FID is 1000x better than TCD
6.linear and dynamic range better than TCD

Disadvantage:
1. destructive detector
  Thermal Conductivity Detector (TCD)
 One of the earliest detectors of GC

 The device contains an electrically heated source whose

temperature at constant electrical power depends on the thermal
conductivity of the surrounding gas.
 Twin detectors are usually used, one being located ahead of the
sample-injection chamber and the other immediately beyond
the column. The bridge (Wheatstone bridge) circuit is arranged
so that the thermal conductivity of the carrier gas is canceled
 The thermal conductivities of helium and hydrogen (commonly
used carrier gases for TCD) are roughly 6~10 times greater than
those of most organic compounds. Thus, even small amounts of
organic species cause relatively large decreases in the thermal
conductivity of the column effluent, which results in a marked
rise in the temperature of the detector.

Advantages:
Simplicity, large linear dynamic range, nondestructive

Disadvantages:
Low sensitivity (precludes their use with WCOT columns with small amounts of
sample)
  Electron Capture Detector (ECD)

 Radioactive decay-based detector

 Selective for compounds containing electronegative atoms, such as

halogens, peroxides, quinones, and nitro groups
 The sample effluent from a column is passed over a radioactive β
emitter, usually 63Ni. An electron from the emitter causes ionization
of the carrier gas (often N2) and the production of a burst of electrons.
 In the absence of organic species, a constant standing current between
a pair of electrode results from this ionization process. The current
decreases significantly in the presence of organic molecules
containing electron negative functional groups that tend to capture
electrons.

 Ionization of carrier gases: N2 + β- → N2+ + e- Ar2 + β- → Ar2+ +

e-
Advantages:
 useful for environmental testing detection of chlorinated pesticides or
herbicides; polynuclear aromatic carcinogens, organometallic compounds
 selective for halogen- (I, Br, Cl, F), nitro-, and sulfur-containing compounds
 detects polynuclear aromatic compounds, anhydrides and conjugated
carbonyl compounds

Disadvantages:
 could be affected by the flow rate
  Mass spectrometry
A common combination is gas chromatography-mass spectrometry (GC/MS or
GC-MS). In this technique, a gas chromatograph is used to separate different
compounds. This stream of separated compounds is fed online into the ion source,
a metallic filament to which voltage is applied. This filament emits electrons which
ionize the compounds. The ions can then further fragment, yielding predictable
patterns. Intact ions and fragments pass into the mass spectrometer's analyzer and
are eventually detected.

 The GC eluate is a mixture separated into segmentsof pure substances with

each analyte segment mixedwith the mobile phase.
 These are introduced into a ion source of a MS,blasted with electrons, which
cause them to break intopieces and turn into positively charged molecular
ionsand fragmented ions (ion source).
 Ions are directed to travel thro‟ a „filter‟ where the ions,based on „masses‟
are filtered and „detected‟. The filter continuously scans through the range of
masses as the stream of ions come from the ion source.

Other GC Detectors

 Photoionization Detector
aromatic hydrocarbons organosulfur/organophosphorous

 Atomic Emission Detector

element-selective detector

 Flame Photometric Detector

sulfur and phosphorous containing compounds
 Advantages of GC
 Due to its high efficiency, GC allows the separation of the components of
complex mixtures in a reasonable time.
 Accurate quantitation (usually sharp reproducible peaks are obtained)
 Mature technique with many applications notes available for users.
 Multiple detectors with high sensitivity (ppb) are available, which can also
be used in series with a mass spectrometer since MS is a non-destructive
technique.
 Fast analysis
 High efficiency – leading to high resolution
 Sensitive detectors (ppb)
  Non-destructive – enabling coupling to Mass Spectrometers (MS) - an
instrument that measures the masses of individual molecules that have been
converted into ions, i.e. molecules that have been electrically charged
 High quantitative accuracy (<1% RSD typical)
 Requires small samples (<1 mL)
 Rugged and reliable techniques
 Well established with extensive literature and applications

 Disadvantages of GC
 Limited to thermally stable and volatile compounds.
 Most GC detectors are destructive, except for MS.
 Limited to volatile samples
 Not suitable for samples that degrade at elevated temperatures (thermally
labile)
 Not suited to preparative chromatography
 Requires MS detector for analyte structural elucidation (characterization)
 Most non-MS detectors are destructive

 Typical GC Applications.

 Pharmaceutical
In the pharmaceutical industry GC is used to analyze residual solvents in both raw
materials (drug substance) and finished products (drug product).
Biopharmaceutical applications include urine drug screens for barbiturates and
underivatized drugs and for ethylene oxide in sterilized products such as sutures.
 Food/Flavors/Fragrances
The food industry uses GC for a wide variety of applications including quality
testing and solvents testing. The Flavors and Fragrances industries use GC for
quality testing and fingerprinting of fragrances for characterization.

 Petrochemical
GC applications include natural gas analysis or refineries, gasoline characterization
and fraction quantitation, aromatics in benzene, etc. Geochemical applications
include mapping of oil reserves and tracing of reservoirs etc.
 Chemical/Industrial
Chemical / Industrial uses include determination of product content, determination
of purity, monitoring production processes, etc. GCs are used to detect organic
acids, alcohols, amines, esters, and solvents
 Environmental
Environmental GC applications include detection of pollutants such as pesticides,
fungicides, herbicides, purgeable aromatics, etc. Industrial environmental
protection applications include stack and waste emissions as well as water
discharges.

Table 2: Relative advantages and disadvantages of GC versus HPLC

GC HPLC
Sample must be volatile or Volatility is not important, however
derivatized previous to GC analysis solubility in the mobile phase
. becomes critical for the analysis
Most analytes have a molecular There is no upper molecular weight
weight (MW) below 500 Da (due to limit as far as the sample can be
volatility issues) dissolved in the appropriate mobile
phase
Can be coupled to MS. Several mass Methods must be adapted before
spectral li- using an MS detector (non-volatile
braries are available if using electron buffers cannot be used)
ionization

Can be coupled to several detectors For some detectors the solvent must
depending on the application be an issue. When changing detectors
some methods will require prior
modification

References:
1. [David_G_Watson]_Pharmaceutical_analysis__a_textb(BookZZ.org)
2. simulab.ltt.com.au/5/Laboratory/StudyNotes/index.htm
3. Douglas A. Skoog, Donald M. West, F. James Holler, Stanley R.
Crouch-Fundame
4. http://www.chromedia.org
5. https://www.slideshare.net/faizanakram77/gas-chromatography-gc-
by-faizan-akram
6. principles-of-gas-chromatography-2
7. http://chemdata.nist.gov/4

Md.Nazmul Islam (ABIR)

#151330 (2ND Batch)
Contact: 01766388247
Abir140044@gmail.com
Department of Pharmacy
Pabna University of Science and Technology