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HS-SPME-GC/MS Analysis: Fiber Conditioning and Desorption Parameters

MatteoUnisa

Hello everyone, it’s me again!

I’m currently working on HS-SPME-GC/MS analysis of essential oils, and I’ve run into a few issues with my blanks that I’d love to get your advice on.

ANALYSIS SETUP: I’m using an Agilent 8890 GC system coupled with a 5977B mass spectrometer, with a PAL autosampler and SPME tool. The fiber I’m using is the dark grey (5191-5874), and the column is an Agilent DB-WAX (122-7032), with a temperature range of 20°C-250°C (260°C).
The fiber was conditioned for 2 hours at 240°C in the inlet, following the manufacturer’s recommended range (up to 280°C), but I kept it below the maximum temperature to stay within the column’s limits.

During the two blank injections (using a 20 mL empty vial), I observed some significant peaks, some of which seem to correspond to cyclosiloxanes (which could be from the PDMS portion of the fiber). However, in the second blank, these peaks had lower intensities compared to the first.

QUESTIONS:

  • FIBER CONDITIONING:

    • If these peaks are indeed from the fiber, could this indicate that I did something wrong during the conditioning process?
    • Even though the fiber conditioning range goes up to 280°C, I chose to condition it at 240°C to stay within the column’s temperature limits. Is it correct to consider the column’s temperature range when setting the inlet temperature? I’ve come across conflicting opinions on this in some articles.

For clarity, I’m attaching the chromatogram images from the two blank injections. The first injection is shown in black, and the second one is shown in red, so you can compare the intensity of the peaks.

  • PURGE FLOW TO SPLIT VENT:

    • I’ve set the purge flow (10 ml/min) to split vent at 1 minute, but the desorption time is 5 minutes. Could this cause any issues? Or does the injection actually correspond to the total desorption time?
    • If I wanted to add a post-injection desorption, how would this be handled in this case?

  • SAMPLE SEQUENCE:

    • I’ve noticed something new now that I’m running samples in sequence. Specifically, the method is structured as follows:
      • 10 minutes of conditioning time;
      • 30 minutes of incubation time;
      • 15 minutes of extraction time;
      • 5 minutes of desorption time.

    The entire process takes about 60 minutes. Adding the 56 minutes of chromatographic run time, the total time per sample is 116 minutes. However, I’ve observed that after about 10 minutes of the chromatographic run, the fiber reinserts into the inlet. Could this be related to the GC Cycle Time, which is set to 60 minutes? Additionally, I noticed a peak appearing shortly after this event that was not present before. Has anyone experienced something similar or can explain how the GC Cycle Time might influence this?

  • VIAL CAP INTEGRITY:

    • After running a blank sample three times to evaluate the fiber and the analysis, I inspected the vial and noticed some peculiarities (at least to my eyes, though perhaps not to an expert). Specifically, I observed three small “cylinders” on the inner side of the cap (Agilent part number: 5188-2759), which I suspect were created by the insertion of the fiber holder structure. These look similar to small core drillings in the material of the cap. Could these cause any issues during the analysis? Could they also potentially damage the fiber?

    • Additionally, I noticed that two of the punctures created by the fiber holder structure remained open and clearly visible. Considering that this analysis targets volatile compounds, wouldn’t this pose a potential problem, particularly when performing duplicate analyses?


  • SAMPLE VIAL PENETRATION DEPTH (SVPD):

I have a question about the Sample Vial Penetration Depth (SVPD) setting. Does this value take the length of the fiber into account?
For example, if I set the SVPD to 40 mm and the fiber is 10 mm long, what is the actual penetration depth? Is it

"Real SVPD" = 40 mm depth + 10 mm fiber = 50 mm TOT

"Real SVPD" = 30 mm depth + 10 mm fiber = 40 mm TOT

Thank you very much again for any help and suggestions!

  • 0

    This needs to be run in SPLIT mode. Try 20:1 split. Splitless is not typically used with HS/SPME.

    A fiber conditioning station is available as an accessory to add on the PAL rail instead of conditioning it in the injection port.

    What is your injection port temperature? If it is 240° C - is that your GC final oven temperature?  Was the oven at the same temperature as the inlet while you were conditioning the fiber?  If not, anything that goes off the fiber will stop in the column and show as peaks the next run.

    What do you mean by "post-injection desorption" ?

    Repeat injections are not typically run out of the same vial for HS/SPME.  The syringe needle is just too big for the septum to always close tightly around the hole and the hole may be open or, at the very least, may leak after the first puncture. Subsequent punctures are not in the same place at all. 

    The GC cycle time should be set for the total time it takes from START run, the full GC run time, cool down, and until the GC comes ready again - one GC cycle. This could be ten minutes or more longer than the GC run time.  

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