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GC Retention Order

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    GC Simulator Workout

    Learning Objectives

    After completing this exercise students should be able to:

    1. Predict and explain the elution order of analytes.
    2. Explain the effects of temperature programs and flow rate on resolution at the macroscopic and molecular level.
    3. Explain the effects of flow rate.
    4. Select separation conditions for a given set of molecules.

    This workout will make use of the following molecules.



    Boiling point




















    1. What are the major contributions to separation and elution order when using GC?


    1. If you were to analyze these compounds by GC, what elution order would you predict?


    1. Open gcsimulator (either by going to or using the downloaded version). Confirm that the oven is set to an isothermal elution mode at 200 0C, helium carrier gas, constant flow rate of 1 ml/min, and a split injection of 100:1. The only stationary phase option is an Agilent DB-5MS UI, which uses a non-polar stationary phase.

      Remove all of the compounds currently populating the compound list under the chromatogram. Add the compounds listed in the table on page 1. Use concentrations of 200-500 µM.

    2. Note that the compound table provides tR for each of the molecules. What is the elution order of the molecules? Does it match with your prediction? If not, consider why.








    1. Discuss the shape of the peaks and how the peak shape changes with retention time? Explain the molecular level explanation for these changes in peak shape.









    1. Vary the flow rate; 1-5 mL/min flow rates are common. How does the chromatogram change? Take note of peak shape (you may want to look at σtot, which is a measure of peak width), resolution, total analysis time and signal to noise ratio. Explain each of these changes at the molecular level.















    1. Reset the flow rate to 1 mL/min. Are the peaks well separated?



    1. What parameters could you change to improve peak shape and separation? Use the GC simulator to adjust the separation parameters to achieve baseline separation with narrow peaks for all molecules.

















    1. With these optimal parameters, what is the elution order? Explain, at a molecular level.


    1. Look at your optimal parameters and the boiling points of nonadecane and lidocaine. Explain, at a molecular level, why these compounds can be detected with your parameters.


    1. Resolution?


    Contributors and Attributions

    This page titled GC Retention Order is shared under a CC BY-NC-SA 4.0 license and was authored, remixed, and/or curated by Contributor.