Say I am analysing a sample containing analyte A and B with the retention time of each at 5 minutes and 11 minutes. If my target compound is analyte A, can I shorten the run time to 7 minutes or do I need to set to 15 minutes to let both the analytes to completely elute? Will there be a risk of carry over if I shorten run time to 7 minutes?
good day! you can reduce the analysis time to 7 minutes, but it is better to extend the analysis to 12 minutes to wash off the remaining analytes from the column
If you provide details of the analysis, perhaps you can reduce the time of analysis while both analytes exit the column. Column? Flow rate? Gradient or isocratic conditions?
Kanapathy Semaselaun Asked: "sample containing analyte A and B with the retention time of each at 5 minutes and 11 minutes. Will there be a risk of carry over if I shorten run time to 7 minutes?" -
The answer is YES. Do not do that. Shortening the runtime, before all of the samples have eluted off the column may indeed result in carry over, contamination and/or a method that is now invalid. Why are you in a hurry? BEST OPTION: You could optimize the method to retain/elute all compounds properly, but in less overall time (i.e. use a gradient and change the % composition). HPLC Method optimization may include adjusting/changing: the flow rate, mobile phase composition, temperature, column dimensions, detector settings and so on. These changes are fundamental to good HPLC method development and should be addressed early on.
- Alternatively, you could program a change to the mobile phase conditions AFTER the compound of interest elutes off the column and passes through the detector to waste, BUT in doing so put yourself at risk for MISSING something important (other peaks) if your "sample(s)" change ( never assume things stay the same ). *It is for this reason that we would never make such a change in an actual analysis method, as doing so would invalidate it (it would not follow good chromatography fundamentals and not be accurate).
- A properly developed HPLC method will retain all compounds present in the sample, then detect and elute them ALL off the column. The column is then washed, with a solution that is stronger than the original mobile phase, re-equilibrated and ready for use again. This procedure reduces the chances of any carry-over, column fouling or contamination and improves reliability and accuracy.
However, if you are just running samples for training/experiments to learn about the technique (e.g. school class), then you could change the analysis condition to flush the remaining sample(s) off the column to start a new equilibration.
You can increase the flowrate for several minutes after last compound of interest elutes. This should flush the other crap off your column before the next injection.
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