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[16] What do Chromatograms tell us? Retention Times are much Longer as Expected

1 Aug 2013

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Chromatograms are like fingerprints.  If you can “read” chromatograms well, you often can find a plausible cause. In this series, we will show a series of GC-chromatograms that are obtained from users and discuss some potential causes for the phenomena. Then we can move into some solutions for improvement.

You replace your column for a new one and you find a chromatogram as shown in Fig 1A.

Your original GC-chromatogram looks like Figure 1B.   Retention times seem to have increased. What can cause this difference?Blog16-fig1
Fig.1 New column shows much higher retention (A) then the original column (B) used for the method.

The retention times are 80% longer then in the normal run.  To find possible causes, one needs to know what factors influence retention time.  These are: temperature, carrier gas velocity, film thickness, stationary phase type, the column diameter and column length.

When a similar type of column has been selected, the stationary phase, column diameter, length and film thickness should be pretty similar. So the main parameter to check is oven temperature. For liquid stationary phases, like Rtx-1, the retention increases approximate a factor 2 every 15 C the temperature is reduced.  Check the oven temperature with a thermometer.(do not automatically trust the digital readouts).

If the temperature is OK, there may be a column related problem.

The retention time for solvent in the chromatograms is similar, which would indicate that the retention of the new column must be higher and the linear gas velocity is also good. Now the problem can be film thickness, column diameter or type of stationary phase.

If the elution order is similar, the type of stationary phase is probably similar. Also if an Rtx 5  is used instead of an Rtx-1, it will not make such a huge difference in absolute retention times. If a more polar phase would be present, we may see this, but we would also see some relative peak shifting.

There maybe a problem with the Internal Diameter.  For this you need to check the inlet pressure. Sometimes the method asks to set a linear velocity. If the column diameter is in reality smaller, you will need a significant higher inlet pressure to set the same linear velocity. Smaller ID columns with the same film thickness have a lower “beta”, which translates in a higher retention.

If the method requires a 0.32mm and we use a 0.25mm with a similar film thickness, this could generate the results obtained.

Maybe your new column also HAS a higher retention and the vendor made a mistake sending you a column with too thick film. To check this, run a test sample as the vendor has done, and see how the peaks elute.

If indeed a higher retention, ask for a new column (and ask why this column was passed QA).

If you still want to work with this particular column, you can adjust the oven temperature. Increasing the oven by 15C  will give you the approximate similar retention times as before.