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- Posts: 1
- Joined: Fri Jul 01, 2016 11:18 am
I am working with 20 mM ammonium acetate buffer: acetonitrile (60 :40 ,v/v) on Synergi Hydro-RP (250 x 4.6 mm, 4 µm). During all runs, the pressure is constant (around 1550). At the end of my work every day I follow this washing procedure:
Composition Duration (min)
Water: acetonitrile (95: 5) 60
Water: acetonitrile (50: 50) 10
Water: acetonitrile (70: 30) 15
Acetonitrile (100) 10
Since two days, I realized gradual increase in column back pressure during washing as follows:
Composition Normal P New P
Water: acetonitrile (95: 5) 1550 1800 (flow 0.8 )
Water: acetonitrile (50: 50) 1550 1850
Water: acetonitrile (70: 30) 1280 1450
Acetonitrile (100) 650 1200
What could be the problem and how to return the column to its normal pressure?
N.B:
The buffer (pH 5.8 ) is freshly prepared, then mixed manually with acetonitrile and filtered before use. The solution remains completely clear without any precipitation till the end of work.