Welcome to the May 2012 issue of the chromatography newsletter. This issue the topic is Achieving Good Peak Shape in Splitless Injection.

Dr Gerard Sharp...

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Achieving Good Peak Shape in Splitless Injections


GC Splitless injection mode is necessary for trace analysis when maximum sensitivity is required. In this mode, the split valve is closed on injection resulting in the vaporized sample flowing directly into the capillary column. Whilst maximum sensitivity is achieved, it does come at a cost. Splitless is more challenging than split mode. The inlet liner flow in splitless mode is the same as the capillary column flow resulting in a long sample transfer time into the column. To achieve nice sharp peaks, there are three main techniques available:

  • Thermal Focusing
  • The Solvent Effect
  • Pulsed Splitless Mode


Let's examine how each of these work but first we shall review why we get naturally broad peaks in splitless mode.


Why Do We Get Broad Peaks?


When the sample in injected into the hot inlet, both the solvent and the sample components will vaporize. This results in filling almost the whole inlet liner quickly with vapor. The low carrier gas flow through the liner then begins to slowly transfer the liner contents into the column. The higher the liner and column flow, the faster the vapor will be transferred into the column. Because the vapor initially spreads out and fills almost the whole of the inlet liner, the slow transfer of the sample vapor into the column would result in an unacceptably wide initial sample band and therefore broad chromatographic peaks. In fact a typical liner volume is in the order of 1mL. So transferring the sample vapor at a rate of 1mL / minute through the liner would result in an initial sample band of 1 minute. The chromatographic peaks would even be wider than this as the sample components spread out further as the make their way down the column towards the detector. You wouldn’t see a one minute wide peak as it would be lost in the noise of the baseline. The peak widths of the early eluting peaks should be about 3 seconds wide.


Thermal Focusing


Thermal Focusing (also known as cold trapping) is one of the main techniques to concentrate or focus the wide sample band from the inlet into a narrow sample band at the start of the column. In this animation, when the oven temperature is set low, the long stream of molecules enters the column and then condenses into the stationary phase. It is like someone has closed a gate preventing the molecules moving further down the column. The gate we use is temperature.

The molecules won’t focus right at the start of the column because there will be a thermal gradient due to some heat from the inlet. For example the inlet temperature might be 250C and the initial oven temperature set to 50C. So the first part of the column will experience a higher temperature due to heat transfer from the inlet. The insulation between the inlet and the oven is very important because of this. If this insulation is removed, in splitless mode your chromatography can change from having sharp peaks to ugly broad peaks very quickly.
Even if the oven temperature is kept low, there are still some molecules that will vaporize out of the stationary phase and then continue to move down the column. This is because even if the initial oven temperature is lower than the boiling point of the sample component, there will still be some molecules in the vapor phase due to the vapor pressure of the component. Therefore if the initial oven temperature time is over extended, this can lead to weaker focusing and broader peaks.

The initial oven temperature is kept low for as long as the sample continues to exit the inlet. This is typically about a minute. After this time, the oven temperature is ramped and the molecules begin their journey in and out of the stationary phase towards the detector.


Solvent Effect


A second peak focusing effect is applicable for the volatile components in the sample. The solvent effect involves setting the initial oven temperature 10-20C below the boiling point of the solvent. When the solvent vapor cloud enters the column, the vapor condenses in the stationary phase over the first few meters of the column. The less volatile components in the sample follow and redissolve in this condensed band of solvent. The initial oven temperature is held constant for the length of the sample transfer from the inlet liner to the column – typically one minute. After this time, the oven is temperature programmed and this causes the solvent band to slowly vaporize. As the solvent band shrinks, the sample components concentrate in the reducing band until finally the solvent is completely vaporized and continues to move down the column. The sample components, now as a nice focused band, then vaporize with the increasing oven temperature and begin their journey down the column.


Pulsed Splitless


The electronic pressure control of modern GCs allows the change of inlet pressure to be time programmed. The inlet pressure is set to optimize resolution by optimizing the carrier gas velocity. An optimum inlet pressure for the best resolution is not always the best for sample introduction. Pulsed splitless gives us the capability to set an optimum inlet pressure to get the sample components into the column as quickly as possible and then returns to the optimum inlet pressure for resolution as the sample components continue to move down the column. Here is how it works.

If the optimum inlet pressure for best resolution is 10psi, then just before the sample is injected, the pressure is increased to say 40 psi and left at this higher pressure for a time for example 30 seconds. This results in a higher column flow but importantly, a higher inlet liner flow. The higher inlet liner flow results in faster sample transfer to the column and therefore sharper peaks. After 30 seconds the inlet pressure is returned to the optimum 10 psi.
There are other benefits using pulsed splitless injection. Because the sample doesn’t spend as long in the inlet liner, this mode minimizes breakdown which might occur for thermally labile compounds such as some pesticides or drugs.
Also, in pulsed splitless mode, the higher initial pressure suppresses vapor expansion of the sample and helps minimize the chance of the vapor spilling out of the top of the liner. A higher pressure can also facilitate a higher injection volume. An interactive animation showing pulsed splitless in action is available on the web site for members.



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