Question: I am injecting cell extract samples on my HPLC instrument and I find that the back pressure increases after each run. Any ideas on what the problem might be?

Answer: There are a number of possible causes.

First, always be sure to sonicate and filter your mobile phase before introducing to the LC system especially when using MP additives especially when dissolved solids such as ammonium formate are used.

Second, since the problem may not necessarily be from the mobile phase the pressure could be from particulates in the sample which end up in the column. In this case of using cell extracts, you could try modifying your sample prep to include more thorough removal of particulates by using an appropriate syringe filter. You may also want to consider a disposable inline pre-column filter. In order to remove particulate matter that does get on your column, try back flushing the column but check with the column manufacturer first as not all columns can be back flushed.

Finally, the source of the pressure may not be in the column or mobile phase but could be coming from the pump. You might try disconnecting the column, running 5mL / minute, and see if you observe a significant pressure with no column attached. In this case you would have a problem with the pump and need to have it repaired.