Chromatography Forum

It is quite popular to run fast HPLC using high flow rate with smaller particle size column. My question is how about the regeration of the small particle column when you have to overload the sample to get 0.1% peak quantified. I still prefer to use conventional 5 u, 4.6 mm x 15 cm column at 1 mL/min for the impurity/degradant determination. The regeneration of the column in the gradient run is fine and we can continuously run over 40 hours with consistent results. Anyone has experience of using say 3 u, short column to analyze small peaks (with overloading sample)? How long the run could last without problem. Thank you.

I do not see a problem that could be associated with the use of smaller columns. In our ecperience the results are equivalent.

You would scale your injection volume such that the impurity peak is the same height for the fast column as for the standard column. The overload will be proportional.

A couple practical things: Reduce as much of the gradient delay volume in your system as possible when converting to a fast method. Use a detector with a fast data collection rate (scaled to the speed of the chromatogram). In fact the detector may be the rate-limiting step in your system.

I have used UPLC(Ultra fast liquid chromatography), their particle size is 1.7 micron, diameter 2.1mm. as you decrease the diameter sensitivity will increase so no need to inject more microlitre sample you shall have to inject less microlitre sample.and very low flow you can use such as 0.25ml/min because instrument has capacity to bear 15000psi pressure.as particle size is low separation is faster.so run time will be shorter.
AMITAVA GHOSH.