After method development, I find that a lot of chemists will
delve into method validation studies like accuracy, precision, etc. but leave
robustness for the end. However, I think it is better to do robustness first.
The reason for this is that the results can help you further refine the method in
ways that you may not have considered during the method development stage.
Let’s
say for example that you do your method development have a perfectly
satisfactory separation where your critical peak pair is baseline-resolved. You
move on to validation and go through all the necessary studies (e.g. accuracy,
linearity, repeatability, intermediate precision, LOD, LOQ, etc.). Then in the
robustness studies you find that alteration of one of the method variables
actually leads to a superior separation. This was what I found when I varied
the TFA concentration. I chose a 0.1% concentration originally during the
method development stage but hadn’t given much thought to it at the time. Then
in robustness studies, I found that the critical peak pair, which had been just
baseline-resolved with 0.1% TFA, became vastly better separated upon a decrease
to 0.05%. Before proceeding with the rest of the validation, I could use the
0.05% concentration in my method.
I hope
this information helps you as much as it helped me. I am always learning new
tips and tricks to make chromatographic analytical techniques more streamlined
and efficient.
