{"id":87712,"date":"2020-03-06T21:42:00","date_gmt":"2020-03-06T21:42:00","guid":{"rendered":"https:\/\/discover.restek.com\/?p=87712"},"modified":"2026-02-20T16:31:49","modified_gmt":"2026-02-20T16:31:49","slug":"discovering-analyte-breakdown-on-column","status":"publish","type":"post","link":"https:\/\/discover.restek.com\/zh-hans\/blogs-de\/gnbl5614\/discovering-analyte-breakdown-on-column","title":{"rendered":"Discovering Analyte Breakdown On-Column"},"content":{"rendered":"\n

I completed my winter internship at Restek and my main task was to expand the Pro-EZGC libraries. So I created a library for Nitrosamines on the Rxi-1301Sil MS and while acquiring data we noticed a foot on a couple of my peaks.<\/p>\n\n\n\n

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Figure 1:<\/strong> On-column breakdown can be observed for the late eluting nitrosamine compounds as indicated by the foot at the leading edge of the peak.<\/p>\n\n\n\n

I attempted to translate the simulation to a column with the same phase but with a thicker film. Since the Rxi-624Sil MS and Rxi-1301Sil MS are the same stationary phase I modeled a run with the Rxi-1301Sil MS Library using the column dimensions of an Rxi-624 Sil MS. As I ran the conditions provided by EZGC we noticed a strange rise of the baseline.<\/p>\n\n\n\n

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Figure 2:<\/strong> Rise in the baseline is observed which is most likely caused by analyte breakdown on-column. Breakdown components remain in the stationary phase and are observed as carryover.<\/p>\n\n\n\n

One of my colleagues immediately suggested that the nitrosamines could break down as a function of elution temperature. In order to test this theory I prepared three different run conditions with different elution temperatures for the late eluting nitrosamines on the Rxi-624 Sil MS:<\/p>\n\n\n\n

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Figure 3:<\/strong> Adjusting the final hold changes the elution temperature of the last three compounds. If the compound response drops as elution temperature increases it will support our theory of on-column breakdown.<\/p>\n\n\n\n

These are the compounds I used (sorted by elution order):<\/p>\n\n\n\n

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Figure 4:<\/strong> List of nitrosamines found in the chromatogram where the last three compounds are labeled as 1 2 and 3.<\/p>\n\n\n\n

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Figure 5:<\/strong> Three chromatograms illustrating the three different programs with increasing elution temperature of the last three nitrosamines. Notice the significant increase in breakdown.<\/p>\n\n\n\n

I labeled the last three peaks; the ones that break down appear to have the most activity on-column. Thanks to the MS I was still able to determine the retention times and it became evident that the breakdown increases with higher elution temperature. Was this a general problem with these compounds on this phase or was this specific to the Restek column?<\/p>\n\n\n\n

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Figure 6:<\/strong> Chromatograms with three different oven conditions. The last three nitrosamines have increasing elution temperatures from top to bottom. These chromatograms were performed on a competitor column to verify the on-column breakdown is most likely related to the cyano stationary phase and not specific to a manufacturer.<\/p>\n\n\n\n

We only had a 60m competitor column available so it’s not a direct comparison however we see similar breakdown. We can tell that the nitrosamines break down on-column on this phase in general. But if you use a film that is thin enough and run the compounds with a low elution temperature it will minimize on-column breakdown. Using an Rxi-1301Sil MS is still faster than using a “5-type” phase but you will have to enter your nitrosamines into ProEZGC and find out for yourself.<\/p>\n\n\n\n

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