Quantitation Strategies

[GCgirl26]

Hello,

Thank you for reading this post. I would like to reach out to the seasoned chromatographers out here for advice.

I began using GC-MS six months ago with methods designed by the person who held my position before me. I can look at all the Agilent documents and troubleshooting articles I want, but there is no substitute for experience.

I am looking to expand on quantitation possibilities of flavor compounds in horseradish. Currently, I can quantitatively measure the most abundant and potent flavor chemical. I am interested in compounds that are less abundant, but visible in the chromatogram.

One of the compounds of interest appears as a wide hump of a peak toward the end of the run time. The gradient is a constant rate of 10 degrees/min with a bake off at the end. My first thought is to steepen the temperature gradient right before the RT of the peak I seek to refine. It is the last peak to come off. Also, the peak is sometimes larger or smaller in chromatograms of an identical sample. Does this mean affinity to the column could make it more difficult to analyze this compound?

Are there any strategies given the scenario that jump out to you experienced chromatographers?

Thank you,

[Consumer Products Guy]

Do you hold the column at high temperature a decent amount of time after that last peak elutes to clean out prior to the n ext injection? You can actually turn most MS detectors off during that.

Also, does that wide peak have same MS spectra through its elution time, so only one material?

[GCgirl26]

Hello,

Thanks for your reply. Yes the method incorporates several minutes holding at 200 C, 50 C under the max for my column (DB-WAX). Yes it is pure Phenylethyl Isothiocyanate that is eluting. I used to do flash chromatography to purify products of organic synthesis. I would manipulate the polarity gradient in pretty specific ways sometimes, and it worked well. My instinct is to increase the rate of heat/time right before the RT of this peak to try to sharpen it. I fear that it has an affinity for the column that might make it difficult to analyze though. I'm going to give the strategy mentioned a try today. I was hoping anyone else would help me to brainstorm other strategies.

Best,

[James_Ball]

If you start that increased ramp try to start it about 20C before the peak elutes if you can, that will help more than doing it just before it elutes, since you want to speed it through the entire column not just the last couple meters.

Another thing to watch for is if that analyte is being trapped and not eluting on the first injection. If you inject a sample then a blank, do you still see it? If not you are ok, if you do then it is being trapped at the end of the run and eluting on the second, which could explain the differences in peak abundance for replicate injections.

If it is a polarity problem you may want to try a column that is in the middle of the polarity range as a compromise between the polar and nonpolar analytes that are in the mixture since you are at the very polar end of the scale with the wax column.

[GCgirl26]

James-

Thank you for your reply. That is all useful information.

Best,