Gas Chromatography Flashcards
what is chromatography
- Separation process that is achieved by distributing the components of a mixture between 2 phases
what are the two phases
o A stationary phase – chemical partitioning
o A mobile phase – contains the analytes
- Mobile phase flow will drive separation
- Analytes interact different with the stationary phase
o Analytes favouring the mobile phase will take short time to elute
o Analytes favouring the stationary phase will take longer time to elute
separation fundamentals
- The stationary phase is usually fixed in a column
- Mobile phase
GC- inert gas such as N2, He or H2
LC- water mixed with organic solvent
how is separation achieved
2 component mixture
- Onto stationary phase (polar)
- Continuous flow of mobile phase (varying polarity)
- Called norm phase chromatography
- More polar the component the slow it will travel through the column
Components have different affinities for the phases
Greater affinity for stationary phase (more polar)
- Spends more time in that phase not moving
- Less time moving in mobile phase
- Moves through system slowly
Components have different affinities for the phases
Less affinity for stationary phase (less polar)
- Spends less time in that phase
- More time moving in mobile phase
- Moves through system quickly
chromatographic parameters
- Resolution Rs
- Retention (capacity) factor, K
- Selectivity factor (α)
- Efficiency- number of theoretical plates (N)
resolution
see powerpoint for resolution equation
Gas chromatography
Sometimes referred as GLC – Gas-Liquid-Chromatography
- Mobile phase is a gas (usually nitrogen or helium)
- Stationary phase is a liquid (very high boiling point)
Separates volatile organic compounds (VOC)
Analyte in gas phase (suitable for thermostable, non polar compounds
Gas chromatography instrument
see powerpoint
mobile phase
- Three main gases (nitrogen, hydrogen and helium, helium most frequently used)
- Carrier gases drives the analyte forward through the column
- Must be chemically inert
gas choice- van deter plots
- You want a high linear flow in GC so that the separation is quick
- The higher the linear flow, the higher the back pressure so want a gas with low viscosity
- Also there is less time for retention of analytes (less efficient due to high theoretical plate height)
- Carrier gas choice and linear velocity significantly af fect column separation efficiency (illustrate using van Deemter plots)
SEE POWERPOINT FOR GRAPH
The optimum linear velocity for each gas is at the lowest point on the curve, where plate height H is minimised and efficiency is maximised
- Nitrogen provides the best efficiency
Steepness of its van Deemter plot on each side of optimum means that small changes in linear velocity can result in large negative changes in efficiency - Helium has a wider range for optimal linear velocity, but offers slightly less efficiency
Only a small decrease in efficiency when velocity changes slightly - Hydrogen has the shortest analysis times and the widest range of average linear velocity over which high efficiency is obtained
Gas inlet
- Gas is fed from cylinders through supply piping to the instrument
- It is usual to filter gases to ensure high gas purity
- Gas supply may be regulated at the bench to ensure an appropriate supply pressure
sample inlet- injector
- Heats injected liquid samples to gas phase
- Temperature of the sample inlet is usually about 500C higher than the b.pt of the least volatile component of the sample
- Many inlet types exist including
Split/splitless
two sample inlet modes
- Split: proportion of the analyte/solvent gas passes onto the column, most exits through the split outlet
- Splitless: all analyte/solvent gas enters the column
splitless injection
- In splitless mode the split vent is closed during the first part of injections
- All sample goes into the column (much higher detection limits)
- Splitless is used for low concentration samples
- Must be careful with solvent peak
split injection
- In split mode the split vent is opened during the first part of the injection
- A fraction of the sample enters the column (usually ratios such as 1:10, 1:20, 1:50, 1:100 and leads to average detection limits
- Primarily used for non-trace analysis of volatile samples
two types of GC columns
packed
capillary
packed GC columns
- Finley divided, solid support materials coated with liquid stationary phase
- Made of glass or stainless steel
- 1.5 – 10m in length
- Internal diameter of 2-4mm
- Large sample capacity – used for preparative work
capillary/ open tubular column
- Internal diameter of a few tenths of a mm
- 10-80m in length
- Film thickness of 0.1- 5 micrometers
- High efficiency
- Small sample size
- Used for analytical application
- WCOT stationary phase include unreactive silicones, saturated hydrocarbons, esters and amines
WCOT
wall coated open tubular
PLOT
porous layer open tubular
SCOT
support coated open tubular
GC oven settings
- The column is contained in a thermostatic oven controllable to 0.1 degrees C (separation of the analytes depends on vapour pressure which depends on temperature)
- Separation can be improved by adjusting column temperature
temperature oven settings
- When a single temperature is used – isothermal (isocratic)
- A changing temp profile is called a temperature gradient
- Temp changes can be finely controlled
- Can be increased (or decreased) either quickly or slowly
(Useful when compounds have a wide range of boiling points) - Adjustments are made to
Increase separation
Increase resolution
Decrease run time
low temp in oven settings
For a range of compounds of the same type
but with a wide range of boiling points;
at a low temp all are separated but higher
boiling ones are broad and take longer to
elute.
higher temp in oven
At a higher temp, above boiling point, all will
be vapour but lower boiling ones will not be
separated.
temp gradient in oven
A temp gradient allows all to be separated
and resolved – as T rises higher boiling
components will vaporise and be separated.
All within a reasonably short time.
types of GC detector
- The following are common types of GC detectors
• Flame ionisation detector (FID)
• Thermal conductivity detector (TCD)
• Electron capture detector (ECD)
• Nitrogen-phosphorus detected (NPD)
• Mass spectrometers (MS) – covered in another lecture - The choice of detector will depend on the analyte and what the purpose of the analysis is
Flame ionisation detector
- Ignition of effluent from column
- Organic compounds burning the flame produce ions and electrons
- Conduct electricity through the flame
- A large electrical potential is applied at the burner tip and a collector electrode is located above the flame
- Quantities of the analyte down to µg-level can be detected
- Good linearity
- Signal is proportionate to concentration
analysis of BAC using GC-FID
• Blood alcohol concentration (BAC) corresponds directly to the level of impairment of an intoxicated driver
• Breathalyser and field sobriety test provide subjective indication of impairment
• Any court requires quantitation of ethanol content
– The most widely run test in toxicology labs
• Due to the large number of samples and their relative short hold times there is a need for rapid and accurate tests
• Headspace GC is widely used
• This combined with a FID is the most common set-up
• Also one of the practicals for this module
GC detector comparison
see powerpoint
advantages of GC
fast analysis
high efficiency- leading to high resolution
sensitive detectors
high quantitative accuracy
requires small samples
rugged and reliable techniques
disadvantages of GC
limited to volatile samples
not suitable for samples that degrade at elevated temperatures
not suited to preparative chromatography
requires MS detector for analyse structural elucidation
most non-MS detectors are destructive
