Study Notes: The GC Column

In GC, the column is the most important component as it is the interaction of the analyte with the columns stationary phase (and not the carrier gas) that permits separation of the analytes in the sample. There are two types of gas chromatography based on column characteristics:

Gas-Solid Chromatography (GSC) in which a solid stationary phase physically adsorbs analytes leading to retention of the analyte on the column. This GC has limited use due to long retention of active or polar molecules and severe tailing of elution peaks. Its only real application is for specific low molecular weight species and it will not be discussed further in this section. Gas-Liquid Chromatography (GLC) in which a thin layer of a liquid stationary phase is immobilised inside the column. The sample analytes partition between the gaseous mobile phase and the liquid phase. GLC has been adopted as the most useful method and is generally now called Gas Chromatography (GC) although GLC is still mentioned in older textbooks.

Columns are normally coiled so that they can fit inside the oven of a GC.

The GC column must have the following characteristics.

Be rigid and able to withstand moderate pressures (up to 50 psi [~145 kpa]). Contains a stationary phase that is temperature stable and does not chemically react with the analyte components. Contains a stationary phase that will cause the separation of the analytes of interest.

Two general types of column are used in GC (ie GLC):

Packed columns in which the inside of the column tube is densely packed with a solid support material onto which is coated a liquid stationary phase. The stationary phase forms a thin liquid film on the surface of a finely divided, inert support material (typically derived from diatomaceous earth which has a very high surface area). The mobile phase moves over and around the coated material as it travels down the column. Open tubular columns, known as capillary columns, are characterised by a narrow opening

running down the centre (a capillary) through which the mobile phase travels as it moves past the stationary phase. Typical physical characteristics of both column types are shown in the table below (columns are formed into coils to enable them to fit into the oven of a GC). Note that capillary columns are narrower and longer. Characteristic Length (m) Inside Diameter (mm) Coil Diameter (cm) 1-6 2-4 15 Packed Column Capillary Column 10 - 100 0.1 0.3 10 - 30

Packed columns are normally made from stainless steel but can be glass if a less reactive surface is desired. The material of choice for the construction of capillary columns is fused silica, a highly purified and inert material. An outside protective coating, called polyimide, affords strength and flexibility.

The most widely used columns in gas chromatography are referred to as WCOT (wall coated open tubular). These are capillary columns in which a liquid stationary phase is coated directly onto the column wall forming a thin film. 0.25 0.5 m thick. Other types of capillary columns exist with the stationary phase contained in different formats. The table below provides a further comparison of capillary (WCOT) and packed columns. For capillary columns, the greater efficiency (narrower peaks and therefore better separation) and smaller sample sizes means that they are used widely today particularly for new or updated methods. Parameter Efficiency (plates/m) Sample size (ng) Relative pressure Relative speed Chemical inertness Flexible column Capillary 2 000 4 000 10 - 75 Low Fast Best Yes Packed 500 - 1 000 10 1 000 000 High Slow Poorest No

Adapted from Skoog, D. and Leary, J. (1992), Principles of Instrumental Analysis, Fort Worth: Saunders College Publishing.

Study Notes: The GC Mobile Phase (Carrier Gas)

In GC the mobile phase is commonly called the Carrier Gas. Carrier gases must be chemically inert and include the following.

Helium (He) Nitrogen (N2) Argon (Ar) Hydrogen (H2).

The choice of the gas is often linked to the kind of detector chosen. The carrier gas supply is also associated with peripheral equipment such as:

pressure regulators pressure gauges, and flow meters.

Additionally, there are various filters and traps in the carrier gas system to remove water, oxygen, hydrocarbons and other impurities from the supply. Oxygen and moisture, for instance, can cause the stationary phase to degrade while hydrocarbons can lead to ghost peaks to appear on chromatograms. Molecular sieve traps are often used to remove moisture. The purchased gas must be of very high purity and is usually obtained from a gas cylinder. A pressure regulator on the gas cylinder as well as a pressure or flow regulator (controller) on the chromatograph controls the flow rate of the gas.

Another example of how the components of a GC come together. Inlet pressures are usually in the range from 10 - 50 psi (70 - 350 kPa), which lead to flow rates of 1 to 25 mL/min and higher depending upon the type of column being used. The flow rate is often assumed to be constant if the inlet pressure is constant but flow rates should be measured on the end of the column by such devices as soap bubble flow meters. Good separations will depend upon maintenance of the appropriate flow rate throughout the run.

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.

Compatible with the stationary phase and the sample. Allows good separation of the analytes by the column. Is of the highest purity. Must not contain particulate matter, water or other chemical constituents. Is compatible with the detector used. >> Study Notes >> The GC Mobile Phase (Carrier Gas)

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Study Notes: GC Detectors

The detector is the part of the apparatus that converts the signal from the bands eluting off the column into a readable form via a chart recorder. The characteristics of an ideal detector include the following.

Adequate sensitivity (ie sufficiently low limit of detection). Adequate linear range (the ratio of the largest to the smallest concentration within which the detector is linear). Good stability and reproducibility (two consecutive samples should have identical or very similar chromatograms). Temperature range from room temperature to at least 400C. Short response time independent of flow rate. Similar response to all analytes. Non-destructive of sample. High reliability and ease of use - virtually foolproof is the industry norm.

Detectors can be classified into two general types: Universal Detectors for detection of a wide range of analytes:

Flame ionisation detector (FID) (destructive). Thermal conductivity detector (TCD) (non-destructive).

Selective/Specific Detectors for detection of particular types of analytes, and include:

Electro capture detector (ECD). Photoionisation detector (PID). Flame photometric detector (FPD).

The most widely used detector is FID, which, because of its high sensitivity, makes it very useful with capillary columns (able to detect analytes in the small quantities of samples analysed by such columns). In contrast, the relatively low sensitivity of the TCD means that it is not useful for this application.

Components of the FID Detector FID can detect most organic compounds, but is insensitive to water. This means that that the technique is very useful for the analysis of aqueous solutions or water-contaminated samples. Some characteristics of detectors (notice the variation in sensitivity) are shown in the following. Detector TCD Selectivity Organics and various inorganics Organics Application Wide range of sample types including gases Wide range of sample types for example, beer alcohol analysis Trace level halogenated pesticides, food residues, pharmaceuticals & environmental samples Environmental applications (soil, water) Pesticides Sensitivity g levels

FID

ng levels

ECD

Halogens, peroxides, quinones, nitro groups Aliphatics, aromatics, phenols, olefins Phosphorus, sulfur

pg to ng levels

PID

pg to g levels

FPD

P: pg to ng levels S: pg to ng levels

g = microgram = 10-6 g ng = nanogram = 10-9 g pg = picogram = 10-12 g

Study Notes: GC - More About Columns

This study note will give an overview of various aspects that have a direct influence on the ability of a column to resolve analyte peaks. Experience, skill and knowledge are required by the GC operator to achieve good separations. There are two parameters that provide a measure of resolution Column Efficiency and Solvent Efficiency (as defined by McNair, H. and Bonelli, E., Basic Gas Chromatography (1969), Palo Alto, Varian Instrument Division). Column efficiency and solvent efficiency As with HPLC, column efficiency is measured in theoretical plates (N) and is a quantitative measure of

the extent of peak broadening the narrower the peak, the more efficient will be the column thereby leading to improved resolution. There are various factors that affect column efficiency and so when comparing columns these factors need to be stated:

Stationary phase (and how thick) Temperature Carrier gas and flow rate Particle diameter (packed column) Column pressure Column diameter.

Whereas column efficiency concerns the narrowness of peaks, solvent efficiency concerns the separation between peak maxima. This parameter can be expressed quantitatively as a ratio of retention times. The factors having most impact on solvent efficiency are the selection of the column (in effect, the stationary phase) and temperature. The following graphic illustrates the effect of increasing column efficiency or solvent efficiency.

Notice the changes in resolution and position caused by improving column efficiency or improving solvent efficiency. The stationary phase

The properties of the stationary phase include the following.

Low volatility - the boiling point of the liquid should be at least 100C above the maximum operating temperature. Thermal stability - the stationary phase should not degrade under normal operating conditions. Chemical inertness - must be non-reactive under normal operating conditions. Solvent characteristics that allow good separations of analytes (analytes will dissolve or partition in the stationary phase to different degrees enabling their separation).

Examples of stationary phases include:

Polydimethyl siloxane Polyethylene glycol Poly(dicyanoallyldimethyl) siloxane.

The choice of stationary phase is the key element for achieving good separations. Liquid phase percentage For packed columns, the liquid phase used should be enough to coat the particles of the column with a thin uniform layer. Too much (> 30%) will pool between the particles with a decrease in efficiency. The retention time is also proportional to the amount of liquid phase present so modern trends are to use low % liquid phases to produce fast analyses.

Notice that as % liquid phase decreases, band broadening, resolution and retention time is also

reduced. Column temperature An increased temperature means reduced migration time (ie shorter retention time) but decreased resolution. Generally an increase of 30C will double the rate of migration and significantly decrease resolution. Generally resolution can be improved by lowering the temperature but of course this will increase the run time.

Study Notes: GC Retention Time and Peaks

Separation of sample components on a GC column depends on the time it takes for each individual component to move along and elute from the column. Faster moving (migrating) components will be eluted first and slower moving components will be eluted later. The time it takes for an individual component to elute from the column is called the retention time. It is defined as the time from the point of sample injection to the peak maxima. It is also sometimes called migration time.

Notice that retention time is the period from injection (start) to the top of the eluting peak. Also note the area under the third peak. Retention time is affected by all of those factors that have an impact on resolution of peaks as discussed previously. Two factors are the rate of flow of carrier gas through the column and temperature. Carrier gas rate Increasing carrier gas rate decreases the retention time this can be thought of sweeping analytes out of the column at a faster rate. Conversely, decreasing the carrier gas rate increases the retention time. Column temperature Increasing column temperature decreases the retention time this can be thought of as reducing the analytes solubility in the stationary phase. Conversely, decreasing the column temperature increases the retention time. The peak and its shape/size The retention time is measured from the peak maxima, but what is a peak? The peak area represents the amount of electronic charge (for an FID detector) being measured by the detector over a given period of time. As the component comes off the column, the component is detected via an electrical output from the detector. This output is directly related to the amount of component coming off the column. The component comes off the column as a band ie not all at once but over a short period of time depending upon the amount of analyte being detected. This results in the component having a

characteristic distribution (eg tall sharp or flat squat peak) rather than as a single line. In general, the longer the retention time the broader and flatter the peak (band broadening effect)

Study Notes: GC Troubleshooting

The technique of GC is inherently complex with many pieces of equipment interacting to analyse a sample and to produce a chromatogram as a result. In such a system the potential for things to go wrong is necessarily high if operators do not know what they are doing or are less than careful in their approach to the work. But even for experienced and meticulous technicians things can and do go wrong! This is where the ability to troubleshoot is important. The following table contains examples of errors/malfunctions and the likely appearance of a chromatogram under these circumstances. These examples will be useful when you work with the virtual GC. Symptom 1. No Peaks. Possible cause No carrier gas. Injector temperature too cold sample condensing. Syringe is blocked. Septum is damaged. FID flame out. 2. Poor sensitivity with normal retention time. Attenuation too high. Checks Check carrier gas supply. Check injector temperature versus that stated in method. Replace syringe. Replace septum. Check air/hydrogen supply and re-ignite flame. Check attenuation and compare it too that stated in the method. Check filament temperature and current and compare too method.

TCD filament temperature too low.

3. Poor sensitivity with an increase Carrier gas flow rate too in the retention time. low.

Check arrier glow flow rate pressure. Replace septum.

Injector septum continuously leaking.

4. Negative peaks.

Polarity switch in wrong position (TCD).

Check test method and polarity position.

5. High background signal (noise). Contaminated column or excessive bleed from the column. Carrier gas flow rate too high. Carrier gas flow leak.

Re-condition column.

Check carrier gas pressure.

Check carrier gas pressure.

Hydrogen flow rate too high Check pressure of Hydrogen or too low (FID). supply. Air flow too high or too low Check pressure of Air supply. (FID). 6. Tailing Peaks. Column oven temperature too low. Check oven temperature to that stated in the test method.

7. Leading peaks.

Sample condensed in the system.

Ensure injector and detector temperatures are correct.

8. Unresolved peaks

Column oven temperature too high. Carrier gas flow too high.

Check test method and adjust oven temperature accordingly. Thoroughly flush syringe and repeat injection.

9.Extra Peaks.

Residual material from previous sample.

Allow sufficient time for the previous sample injections to be eluted.

Dirty Sample. Contamination syringe. Thoroughly flush syringe and repeat injection.

10. Sudden drop-off of a normal peak.

Loss of hyrogen or air.

Check hydrogen and air pressures. Check carrier gas pressure to that stated in the method.

Carrier gas flow rate too high.

It may be useful for you to print a copy of this for troubleshooting activities. Note that there may be other errors/malfunctions that may occur. Resources and Training Room >> Study Notes >> The GC Column

Study Notes: More about the FID and TCD

Flame Ionisation Detector (FID) In an FID, carrier gas exiting from the end of the column is mixed with hydrogen and burned in air. Ions formed in the flame from combustion of the analytes enter the gap between two electrodes and reduce the gap resistance. This causes an increased current flow across the electrodes whereupon a signal is sent to the chart recorder to print out the chromatogram.

The higher the concentration of ions, the greater the current flow. That is, the amount of ions formed is directly related to the concentration of the component. The role of hydrogen and air is therefore to support the flame. Helium is commonly used as the carrier gas with FID as it provides best sensitivity and resolution. An FID also requires make up gas in order to sweep analytes exiting the column into the detection zone. Nitrogen or helium are normally the gases of choice. The ratio of hydrogen to air is important for keeping the flame alight and providing optimum response. The ratio should be between 8 and 12% with typical flows being around 30 mL/min and 300 mL/min respectively. The make up gas is typically set at about 30 mL/min.

Notice that increasing airflow will increase the FID response up to a maximum. Hydrogen has an opitmal flow rate with reduced response on either side. Flow rates should be monitored for each chromatographic run. A soap bubble flow meter is commonly used for this purpose. Gases are usually supplied from high purity, high-pressure cylinders piped to the GC. Thermal Conductivity Detector (TCD) The TCD is based on the principle that a hot body will lose heat at a rate which is dependent upon the composition of the surrounding gas. This heat loss is used as a measure of the gas composition. Heat is transferred by conduction when the gas molecules strike a heated filament. The greater the number of gas molecule collisions with the filament, the greater the rate of heat loss, which is measured and converted into an electrical signal that is sent to the chart recorder. The chart recorder prints out a chromatogram representing the components in the sample. It should also be noted that different gases have different thermal conductivity so the choice of the carrier gas and the sample composition may be considerations in the choice of this detector. The smaller the gas molecule, the higher its conductivity will be.

The filament is the heart of the TCD. Increasing the filament current to suit particular separations will increase the filament temperature but higher filament temperatures shorten the filament life so a balance between detector efficiency and filament life is usually determined