If the column pressure suddenly changed from one run to another, using the exact same mobile phase composition and same flow rate, then you probably have plugged up some of the stationary phase inside. - Remove the pre-column, flush and place somewhere safe for now.  Using just the main column, flush with 90/10 (ACN/Water) for 20 minutes and record the backpressure seen. Next, you will want to flush the column with a solution that is STRONGER than your mobile phase AND fully dissolves any potential samples that were injected. This is needed to "flush" them off the support. You could use ACN with water, that has some formic acid in it (0.1%), first (90% ACN / 10% Water) and also a a solution with Methanol and water (plus acid), 90 % MeOH / 10% Water too. These are both very safe to use on any RP column. *Additionally, if permitted by the column manufacture (most are OK) you can reverse the direction of the column to backflush it. This helps to flush material off from the column inlet frit and head (where things tend to clog first). Start with these easy steps and record the backpressure seen with each test (so you can determine if there is any change).

Most importantly of all, you must find out why your column was fouled. Are you filtering ALL samples before injecting them? You should be. Are you using the highest quality solutions, additives and solvents with your HPLC? HPLC's require the use of high resistance water (Mega Ohm filtered and pure) and HPLC grade solvents only. Are you samples fully dissolved in solution before you place them in vials (or inject them)? If you use any salt buffers, are you flushing the HPLC system down with water and some MeOH at the end of the day (never leave the system in buffer)? These basic areas must be addressed when using an HPLC system. Perhaps one of them caused the initial problem?

Here is a link to an article that may also help, "Two Common HPLC Problems and their Causes (Sudden changes to either the HPLC Backpressure or Peak Shape) ".

http://hplctips.blogspot.com/2013/12/two-common-hplc-problems-and-their.html

If you are using Salt in your mobile phase, try giving warm water washing. 1 liter of warm (60-70C) water with 2-3 drops ortho phosphoric acid would help.

Also, there might be some choke up in the peek tubing. Cut 0.5-1 cm of peek tubing from its end towards the column entrance and check the pressure.

Check the pressure in your flow cell by disconnecting the connector. If the difference between system pressure before and after disconnecting the connector is more than 1000 psi, then the flow cell is choked with some particulate matter. So, connect inlet tubing in outlet and keep inlet of flow cell open. give flow of 0.1 ml/min for 3-4 mins. Particulate matter will come out from inlet and system pressure will get normal. 

pls check...

First thing first, are you doing Reverse phase chromatography? Which kind of samples you usually run in that system?

Apparently seems clogging somewhere in the liquid path.

Start simple, have you a way to remove the precolumn? Try it and see if the pressure drops, if so change or regenerate the precolumn.

Check if the inlet frits (the filters inside the bottles) are clogged, if unsure change it and try again.

If not try to disconnect the column and the precolumn, if the the pressure remain high the problem is upstream the column. In this case leave the column disconnected and try to flush the system with a mixture of 95%water/5 % ACN Without any salts or buffers for some time and see if the pressure drop. If not  see if in the lab there is someone who could maintain the valves and the pistons, if not call the assistance.

If it a problem downstream the column, put the column in place and disconnect the detector, if the pressure drops the problem is in the detector, in this case the resolution depends on which detector you have.

If not is a clogged column problem, 

Since you already washed the system with ACN mixture and isopropanol, I suggest to run a mixture of 90% water / 10% ACN at 1ml/min, usually it should run for at least 10 column volume but I suggest at least 30 min and see if it there is any improvements. 

Yes it is  reverse phase . The system without the column has no pressure so i guess its the column or the pre-column. The pre-column has been washed properly and USE for 20 minutes with MeOH at first and then with2- isopropanol so i think its okey. The option for  90 % water run have not been done yet so i will try it! My sample was Salvia in 70% MeOH. Thank you both for your advice !

You said the system without the column have no pressure so the eluent flow freely, right? In this case this mean the  block is in the precolumn-column-detector part and I suggest to try the 90% wash in all the system.

If the column pressure suddenly changed from one run to another, using the exact same mobile phase composition and same flow rate, then you probably have plugged up some of the stationary phase inside. - Remove the pre-column, flush and place somewhere safe for now.  Using just the main column, flush with 90/10 (ACN/Water) for 20 minutes and record the backpressure seen. Next, you will want to flush the column with a solution that is STRONGER than your mobile phase AND fully dissolves any potential samples that were injected. This is needed to "flush" them off the support. You could use ACN with water, that has some formic acid in it (0.1%), first (90% ACN / 10% Water) and also a a solution with Methanol and water (plus acid), 90 % MeOH / 10% Water too. These are both very safe to use on any RP column. *Additionally, if permitted by the column manufacture (most are OK) you can reverse the direction of the column to backflush it. This helps to flush material off from the column inlet frit and head (where things tend to clog first). Start with these easy steps and record the backpressure seen with each test (so you can determine if there is any change).

Most importantly of all, you must find out why your column was fouled. Are you filtering ALL samples before injecting them? You should be. Are you using the highest quality solutions, additives and solvents with your HPLC? HPLC's require the use of high resistance water (Mega Ohm filtered and pure) and HPLC grade solvents only. Are you samples fully dissolved in solution before you place them in vials (or inject them)? If you use any salt buffers, are you flushing the HPLC system down with water and some MeOH at the end of the day (never leave the system in buffer)? These basic areas must be addressed when using an HPLC system. Perhaps one of them caused the initial problem?

Here is a link to an article that may also help, "Two Common HPLC Problems and their Causes (Sudden changes to either the HPLC Backpressure or Peak Shape) ".

http://hplctips.blogspot.com/2013/12/two-common-hplc-problems-and-their.html

This is a common problem in HPLC. 

Washing the system and column with appropriate solvents only can resolve this problem.

Nothing more than this.

Regards,

Bujji Reddy.