HPLC Common Problems And Solutions – Pressure

Problem 1: No pressure display

Reason:

Usually the one-way valve is blocked.

Solution:

1) Carefully disassemble the inlet check valve and outlet check valve, then place them in a glass beaker with purified water added in advance, ultrasonicate the purified water for more than 10 minutes (the purified water should preferably be heated to about 50°C in advance), and then replace the methanol with ultrasonic waves. Wash for 15 minutes.

2) Install the check valve on the pump (note that all check valves are oriented upwards). If the above methods cannot solve the problem, the sealing ring problem or the wear of the one-way valve, the ball is not smooth, etc. should be considered, and the damaged parts need to be replaced.

Problem 2: The pressure fluctuates greatly

Reason:

The gem ball and the ball seat of the check valve are not well sealed, and the mobile phase has a backflow phenomenon.

Solution:

1) Carefully disassemble the inlet check valve and outlet check valve, then place them in a glass beaker with purified water added in advance, ultrasonicate the purified water for more than 10 minutes (the purified water should preferably be heated to about 50°C in advance), and then replace the methanol with ultrasonic waves. Wash for 15 minutes.

2) Install the check valve on the pump (note that all check valves are oriented upwards). If the above methods cannot solve the problem, the sealing surface problem or the wear of the one-way valve, the ball is not smooth, etc. should be considered, and the damaged parts need to be replaced.

Problem 3: The pressure is too high

Reason:

It is usually due to blockage in the flow path.

Solution:

1) First disconnect the inlet of the vacuum pump. At this time, the PEEK tube is filled with liquid, so that the PEEK tube is lower than the solvent bottle to see if the liquid drips freely. If the liquid does not drip or drips slowly, the solvent filter head is blocked. Treatment method: soak in 30% nitric acid for half an hour, then rinse with ultrapure water. If the liquid drips freely and the solvent filter is OK, check again.

2) Open the Purge valve so that the mobile phase does not pass through the column. If the pressure does not drop significantly, the filter head is blocked. Treatment method: Remove the filter head from the flow path and sonicate with 10% isopropanol for half an hour. If the pressure drops below 100PSI (6.7 Bar), the filter head is OK and further inspection is required.

3) Remove the outlet end of the chromatographic column. If the pressure does not drop, the column is blocked. Treatment method: If the buffer salt is blocked, flush it with 95% water until the pressure is normal. If some strongly retained substances cause blockage, flush with a stronger mobile phase than the current mobile phase until the pressure is normal. If the flushing pressure does not drop for a long time according to the above method, you can consider connecting the inlet and outlet of the column to the instrument in reverse, and flushing the column with the mobile phase.

Problem 4: The pressure is too low

Reason:

Usually due to system leakage, just tighten it.

Solution:

Air has entered the pump, open the Purge valve, flush with a flow rate of 3-5mL/min, if not, use a special syringe to suck out the air bubbles at the emptying valve by external force.