I am also having the erratic retention time in the UPLC MSMS which is used to determine pesticide residues.There is fluctuation in the retention time for all the analytes. For example for one pesticide analyte the retention times are between 9.77-9.81 minute for 6 injections from the same vial and same day, repeatability (100 ppb standard). Oven temperature is controlled.
Before I used to have very slight fluctuation ±0.02 minute in the whole batch of standards and samples. I call the service engineer. He is convincing me that it's normal. The equipment is only one year.
No one would describe that as "erratic". Retention time variation for six injections from 9.77 to 9.81 minutes would be considered normal. That would be the same as having an average of 9.79 (+/- 0.02).
The retention time were as follows: 9.79, 9.81, 9.77, 9.80, 9.77 for the 6 injections. Whereas in June 2018 the retention time for the same analyte was 10.45 for all the standards (different concentrations) and for the spiked sample it was 10.43 min.
That is a completely different issue than you initially reported by HPLC/MS/MS. If acquired on the same day, a change in retention time from 10.43 minutes to 9.79 minutes indicates a problem with your method. For scientific purposes, compare the sample and std data acquired at the same times not several months or one year apart.
Since you are comparing data from ~ ONE YEAR AGO, it is not fair to compare the data at all, only to compare samples and standards from the same instrument, same method, acquired on the same day. Are you using the same column as last year ? If so, I would not expect the results to be the same from one year ago (even a new column would not give the same exact results), esp after many injections or even storage. Too many variables and too many possible changes may have occurred. *This is why we create methods with Calibration Tables and continuously re-check the values using standards (bracketed).
The technique of chromatographic analysis is not the same as measuring the mass of a sample using a calibrated balance. It is far more complex with many variables. Going from a new to used column or fouled column could cause it. Changing the temperature could cause it. Changing the injection volume could change it. Any changes to the mobile phase (or preparation of the mobile phase) could result in such a change.
- Please refer to this article for assistance and also contact someone local with experience in chromatography to help you (as these types of problems are easiest to solve on-site); "HPLC Retention Time Drift, Change, Variability or Poor Reproducibility. Common Reasons for it"; https://hplctips.blogspot.com/2015/11/hplc-retention-time-drift-change.html
you can not compare your data with old one. If variation is within the same batch or continue run then, we can say there may me problem in machine.
But during run if your variation is +- 0.02 then it is a normal things in case of Liquid chromatography.
However, the change in Rt from 10.43 to 9.79, there are too many reasons. i.e. change in solvent quality, poor sample preparation, change in column, etc.
Karen, the poster did NOT report a change of 0.02 after one year. The change after one year was from 10.43 minutes to 9.79 which is a difference of 0.64.
I want to know if it is normal to have a variation in RT like 9.79, 9.81, 9.77, 9.80, 9.77 in a repeatability test for validation?
Madhvi, your question has been answered. What you describe appears to be normal variation.
- " No one would describe that as "erratic". Retention time variation for six injections from 9.77 to 9.81 minutes would be considered normal. That would be the same as having an average of 9.79 (+/- 0.02)."
One of the questions to ask is what variation do you see when you repeat the analysis on another day?
Thank for the answer William.
Next day with the same vial injection carried out 6 times in a batch, the RTs are 9.77, 9.88, 10.14, 9.89, 9.91 and 9.84 mins. Is these fluctuations normal. last year the RT stays very stable in the apparatus for all the run in a batch.
No, those values indicate that you need to troubleshoot the problem. Wide variation from 9.77 to 10.14 minutes is not acceptable. Is there someone at your school or company with HPLC experience who can help you identify the reasons for the variation? You need local assistance.
BTW: As noted earlier, here is a link to an article on troubleshooting retention time variations; "
HPLC Retention Time Drift, Change, Variability or Poor Reproducibility. Common Reasons for it."; https://hplctips.blogspot.com/2015/11/hplc-retention-time-drift-change.html
in my opinion, this is a normal condition. if the change in retention time only changes 2% RSD. I have a suggestion, you have to rinse the stationary phase longer, so that the stationary phase will be more stable when it will be used
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