Monday, March 4, 2013
INTRODUCTION
Prior to the invention of gas chromatography by James and Martin (Biochem J.,
1952, 50, 679), the separation of close-boiling volatile liquids was at best extremely
difficult. Gas chromatography is a surprisingly simple technique with great versatility,
and is now a given in the analytical chemist's arsenal for the separation and analysis of
volatile mixtures.
The procedure involves vaporizing a sample and sweeping it through a column
with a moving stream of gas termed the mobile phase or the carrier gas. The gases are
commonly supplied by compressed gas cylinders. The sample is introduced into the
injection port. The most common type of analysis involves the injection of 1 to 3
microliters of a liquid sample into a heated inlet, either manually or by an automated
injection device. The injection port is interfaced to the column where the actual
separation takes place. In most cases capillary columns are used to obtain the best
possible separation. The downside to capillary columns is that they have a limited
"capacity". In other words not much sample can be separated at a time. Therefore a split
injection port is often used to allow only a fraction of the injected volume of sample onto
the capillary column. The capillary column's inner walls are coated with either a porous
solid or a viscous liquid material. This inner coating will interact with different solute
molecules to different extents. Those molecules which interact more strongly with the
stationary phase spend on average a higher percentage of their time associated with the
stationary phase than those solutes which do not interact strongly. Those compounds
which do not often associate with the stationary phase pass more quickly through the
column than those compounds which have strong interactions with the stationary phase,
and a separation of the components in the mixture is achieved. Since the compounds
have different mobilities, they exit the column at different times; i.e., they have different
retention times, tR. The retention time is the time between injection and detection. There
are numerous detectors which can be used in gas chromatography. It is a device that
senses the presence of components different from the carrier gas (mobile phase) and
converts that information to an electrical signal. For qualitative identification one must
rely on matching retention times of known compounds with the retention times of
components in the unknown mixture. It is important to remember that any changes in
operating conditions will affect the retention time which will affect the accuracy of
identification. Thus GC is most often used when one is performing a target compound
analysis, where one has a good idea of the compounds present in a mixture so reference
standards can be used for determining retention times. For a sample of largely unknown
composition qualitative identification can be determined by gas chromatography-mass
spectrometry. A mass spectrum of any or all peaks in the chromatogram is compared
with spectra contained in spectral libraries on the system's computer. Quantitative analysis is possible, but given the small amounts of sample that are
injected onto the capillary column, without an automatic injector these volumes are
impossible to reproduce. It is thus recommended that the method of internal standards be
used for quantitative analysis (Skoog/Holler/Nieman, 5th Edition, pp. 18).
Gas chromatographs are really rather simple. The apparatus consists of a
pressurized tank of carrier gas, usually He, a pressure regulator to control the flow rate of
the gas through the chromatograph, a sample inlet, the column, a detector with associated
electronics, some kind of interface to the outside world such as a recorder, and a flow
meter to measure the flow
rate of carrier gas. Chromatographs also provide heating for the column, the sample
inlet, and the detector. The temperatures of these three components can usually be
controlled independently.
Good separation of a given pair of compounds by gas chromatography depends on
the choice of column (which has already been done for you) and on the efficiency of the
overall GC system. The relative position of the various components in the sample on the
chromatogram is affected by a solute-solvent type of interaction with the column
substrate. Column efficiency is concerned with the broadening of an initially compact
band of solutes as it passes through the column. The broadening is a result of column
design and of column operating conditions.
Problems with separation occurs at primarily three places:
1. Sample injection. The sample should be injected all at once into the column and
should be vaporized immediately. For this reason, good GC's have heated sample
chambers. In other words, the sample should enter the packed column as a plug of gas.
The size of the sample should also be small so as not to "overload" the column.
2. Column Characteristics. Assuming you have chosen an appropriate column that
affords selectivity, flow rate is very important. If the flow of carrier gas were shut off, the
band would eventually diffuse throughout the column. Even with carrier gas flowing,
this longitudinal diffusion still takes place forward and backward, and results in band
broadening. This undesirable diffusion depends upon the time that the band remains in
the column, and therefore on the flow rate of carrier gas (at a given column temperature,
column, and type of sample). If the flow rate is increased, the effect of diffusional
broadening is diminished. At higher flow rates however, a second factor creeps in which
increases the broadening; partition equilibrium between the mobile and stationary phase
is no longer maintained. Therefore, there exists a flow rate for maximum column
efficiency. The problem is thus how to find a flow rate which minimizes the peak
broadening due to the mass-transfer rate and longitudinal diffusion simultaneously. This
is a qualitative description of the van Deemter equation (discussed in Chapter 26 of
Skoog/Holler/Nieman, 5th Edition, pp. 681-687). 3. Detector Volume. This you have no control over unless you are designing your own
instrument. Once the band issues from the column, it should go immediately into a
detector of small volume.
For samples with a broad boiling range, it is often desirable to employ
temperature programming, whereby the column temperature is increased continuously or
in steps as the separation proceeds. This is the answer, in gas chromatography, to what is
called The General Elution Problem. This is treated quite nicely in your
Skoog/Holler/Nieman text in Chapter 26 for chromatographic separations in general, pp.
689-693. Basically, the isothermal analysis of wide boiling range mixtures frequently
leads to a very wide spread in retention times. The longer the retention time, the broader
the peak, so for those components which take a long time to elute, detector sensitivity is
diminished and analysis times can be very long.. With temperature programming,
successively eluted substances experience increasingly higher average temperatures and
so emerge from a column more rapidly than they would under isothermal conditions. So
long as one does not experience peak overlap (i.e. resolution remains tolerable),
temperature programming gives a superior separation.
General
• Separation and possible identification of all types of volatile organic compounds
• Qualitative and quantitative determination of volatile compounds in mixtures
Common Specific Applications
• Raw materials, intermediates and final product analyses in manufacturing
industries
• Environmental, forensic, pharmaceutical, clinical/medical applications
Limitations
• Analyte must be volatile, so not conducive for analysis of large molecular
compounds such as proteins and polymers
Complementary or Related Techniques
• Liquid chromatography provides analyses of non-volatile analytes such as
proteins and polymers.
• Supercritical fluid chromatography provides analyses of volatile, non-volatile and
thermally labile compounds • Capillary electrophoresis provides superior analyses in many
biological/pharmaceutical applications
• Ion Chromatography provides analyses of ionic compounds, as does capillary
electrophoresis
References used to devise this web page:
1. "Handbook of Instrumental Techniques for Analytical Chemistry" Frank Settle,
Editor: "Gas Chromatography", M.J. Van Sant, Prentice Hall, 1997, pp. 125-146.
2. "Principles of Instrumental Analysis", 5th Edition. Skoog, Holler, Nieman, Saunders
College Publishing, 1998, pp. 673-724.
Additional references:
1. "Modern Practice of Gas Chromatography" R.L. Grob, 3rd Edition, New York, Wiley,
1995.
GC Links
General treatment of chromatography including an overview of plate and rate theory:
http://ull.chemistry.uakron.edu/analytical/Chromatography/
Tutorials on chromatography and molecular spectroscopy:
http://www.shu.ac.uk/schools/sci/chem/tutorials/
