firstly make sure the problem not from the column , by the way you can make the routine wash for the whole system by the following sequence :

1-start with 100 % water(hplc grade) for 15 min with acceptable flow rate

2- 80:20 (water : ACN or methanol ) for 15 min

3- 50:50 (water : ACN or methanol ) for 15 min

4_100% (ACN or methanol ) for 15 min

if the problem not solved (reverse the uv conections and flush system with isopropanol for 5 min )

Dear Sharif.

Thanks for your answer

I did the routine wash, but I have extra peak and responce low.

Dear mahsa

Is there any leakage from the flow cell ?

second, try to replace the column ..what the result ?

Last , try to reverse the detector connections (outlet in the inlet ) then flush with isopropanol (HPLC grade) for about 5 min .. then reconnect the system in the correct direction ..