High pressure in the HPLC system

Hello,

 

for the quantification of amino acid, I am using a C18 column. Mobile phase is 40mM phosphate puffer pH 7.8 (channel A) and acetonitrile/ methanol/ water 45:45:10 (channel B). . The method worked really well, but recently I have problems with too high pressure in the HPLC system (Agilent 1260). I have disconnected my column from the system, and the pressure remained high. I would assume that I have salt precipitation in the system. Therefore, I have used warm water to flush the system without the coulumn. The pressure is still very high.

 

Thanks

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  • If you have used a full bottle of warm water and the pressure remains high, I would suggest trying to find out where the pressure is coming from and then probably change some easy parts to figure out if you can solve it that way.

     

    Easiest way to go about it is to remove the column (which you seem to have done already) and then you can open the purge valve on the pump. If the pressure is high with purge valve open, it means maybe your frit is the issue or something before the purge valve.

    After that you can check if the sampler is the issue by closing the purge valve and changing the valve to bypass. That way the flow does not go to the needle, seat and sample loop. If your pressure drop significantly, then you might have something clog in 1 or multiple of those. So you can try to put the valve in mainpass again and disconnect the sample loop from the needle. If the pressure drops, you can suspect something is going on with the needle or the needle seat. If the pressure stays high, it means it is between the sample loop and the valve. So then you can try changing sample loop and see if it fixes it, or you can also open the valve and look at your rotor seal.

Reply
  • If you have used a full bottle of warm water and the pressure remains high, I would suggest trying to find out where the pressure is coming from and then probably change some easy parts to figure out if you can solve it that way.

     

    Easiest way to go about it is to remove the column (which you seem to have done already) and then you can open the purge valve on the pump. If the pressure is high with purge valve open, it means maybe your frit is the issue or something before the purge valve.

    After that you can check if the sampler is the issue by closing the purge valve and changing the valve to bypass. That way the flow does not go to the needle, seat and sample loop. If your pressure drop significantly, then you might have something clog in 1 or multiple of those. So you can try to put the valve in mainpass again and disconnect the sample loop from the needle. If the pressure drops, you can suspect something is going on with the needle or the needle seat. If the pressure stays high, it means it is between the sample loop and the valve. So then you can try changing sample loop and see if it fixes it, or you can also open the valve and look at your rotor seal.

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