Gas Chromatography (GC) is the most widely used method for determination of fatty acid composition in biological fluids. A urine sample was submitted for analysis using the following method: Capillary 30 m x 0.25 mm i.d, x 0.25 um Column Column Temperature 60°C for 2 min 10°C/min to 200°C 5°C/min to 240°C Hold at 240°C for 7min Detection System Flame ionisation detector Injection System Splitsplit less injection (a) With reference to the method above, explain how an isothermal column temperature method is different to a temperature programmed column temperature method (b) Explain how a split/split less injection system works and how it varies from using an on column injection system. (c) When selecting which capillary column to use for separation in GC, explain the FOUR characteristics of the capillary column that must be considered. For each one, describe how it effects the separation in terms of efficiency and resolution. (d) In the method above, a flame ionisation detection system is employed. Describe how this detection system produces a signal when analysing a mixture. Solution A. Isothermal column temperature method : In this method, temperature almost remains constant throughout the method. Only one component can be detected in this method. Where as in Temperature programmed method, temperature varies at different times inorder to detect different components in the mixture. The given method in the problem is temperature programmed column temperature method... B. Splitless injection : Sample will be vaporized in the injector and all of the sample is swept onto the column by the carrier gas. Sample spends lot of time in the injector. It is best for trace concentrations of high boiling point. Where as in split injection, only a fraction of the sample enters into the column. On-Column injection is widely used in packed column and less in capillary columns. Sample deposited directly on the column.. It is very good for thermally unstable compounds.. C. I. Length of the capillary column. Increased length leads to longer separation times. II. Stationary phase thickness, resolution increases with increased thickness of the stationary phase.. III. Capillary column diameter, less the diameter of the column more the resolution. IV. Nature of the column, polar analytes require polar stationery phase. Non polar analytes require non polar stationary phase. D. When the sample is injected into the GC, immediately the mixture separated into components of pure substances and those will be mixed with the carrier gas. Ionized analytes emit photons of distinct wavelengths that generates a proportional analogue signal.. .

1. Gas Chromatography (GC) is the most widely used method for determination of fatty acid
composition in biological fluids. A urine sample was submitted for analysis using the following
method: Capillary 30 m x 0.25 mm i.d, x 0.25 um Column Column Temperature 60°C for 2
min 10°C/min to 200°C 5°C/min to 240°C Hold at 240°C for 7min Detection System
Flame ionisation detector Injection System Splitsplit less injection (a) With reference to the
method above, explain how an isothermal column temperature method is different to a
temperature programmed column temperature method (b) Explain how a split/split less injection
system works and how it varies from using an on column injection system. (c) When selecting
which capillary column to use for separation in GC, explain the FOUR characteristics of the
capillary column that must be considered. For each one, describe how it effects the separation in
terms of efficiency and resolution. (d) In the method above, a flame ionisation detection system
is employed. Describe how this detection system produces a signal when analysing a mixture.
Solution
A. Isothermal column temperature method : In this method, temperature almost remains constant
throughout the method. Only one component can be detected in this method.
Where as in Temperature programmed method, temperature varies at different times inorder to
detect different components in the mixture. The given method in the problem is temperature
programmed column temperature method...
B. Splitless injection : Sample will be vaporized in the injector and all of the sample is swept
onto the column by the carrier gas. Sample spends lot of time in the injector. It is best for trace
concentrations of high boiling point.
Where as in split injection, only a fraction of the sample enters into the column.
On-Column injection is widely used in packed column and less in capillary columns. Sample
deposited directly on the column.. It is very good for thermally unstable compounds..
C. I. Length of the capillary column. Increased length leads to longer separation times.
II. Stationary phase thickness, resolution increases with increased thickness of the stationary
phase..

2. III. Capillary column diameter, less the diameter of the column more the resolution.
IV. Nature of the column, polar analytes require polar stationery phase. Non polar analytes
require non polar stationary phase.
D. When the sample is injected into the GC, immediately the mixture separated into components
of pure substances and those will be mixed with the carrier gas. Ionized analytes emit photons of
distinct wavelengths that generates a proportional analogue signal..