Common Problems and Solutions of High Performance Liquid Chromatograph

Common problems and solutions

As a high-precision instrument, high-performance liquid chromatography can easily cause problems if it is not used correctly during use. The most common problems are column pressure problems, drift problems, and peak shape abnormalities.

2.1 Column Pressure Problems The column pressure problem is a place that requires close attention during the use of high-performance liquid chromatography. The stability of the column pressure is closely related to the quality of the chromatogram peak shape, column efficiency, separation effect, and retention time. The so-called column pressure stabilization does not mean that the pressure value is stable at a constant value, but it means that the pressure fluctuation range is between 50 PSI. (When gradient elution is used, steady and slow changes of column pressure are allowed). Pressure is too high, too low are all column pressure problems.

2.1.1 High pressure This is the most common problem in the use of high-performance liquids. It refers to sudden increase in pressure, which is generally due to blockage in the flow path. At this point, we should check in sections.

(1). First disconnect the inlet of the vacuum pump. At this time, the PEEK tube is full of liquid, so that the PEEK tube is lower than the solvent bottle to see if the liquid is allowed to drip freely. If the liquid does not drip or drip slowly, the solvent filter plug is blocked. Treatment: Soak in 30% nitric acid for half an hour and rinse with ultra-pure water. If the liquid drips freely, the solvent filter head is normal and inspected;

(2). Open the Purge valve so that the mobile phase does not pass through the column. If the pressure does not drop significantly, filter the white plug. Treatment method: Remove the filtered white head and soak it with 10% isopropyl alcohol for half an hour. If the pressure drops below 100 PSI, filtering the white head is normal and checking;

(3). Remove the outlet end of the column. If the pressure does not drop, the column is blocked. Treatment method: If buffer salt is blocked, flush with 95% water to normal pressure. If it is some strongly retained material that causes blockage, it should be flushed to a normal pressure with a mobile phase that is stronger than the current mobile phase. If the flushing pressure is not reduced by the above method for a long time, then the inlet and outlet of the column may be connected to the instrument and the column may be flushed with the mobile phase. At this time, if the column pressure does not drop, only the column inlet sieve is replaced, but once it is not operated, it is easy to cause the column efficiency to decrease, so use it as little as possible.

2.1.1 Pressure is too low Pressure is generally due to system leakage. Treatment method: Look for the interfaces, especially the ends of the column, and tighten the leak. Of course, there is another reason that the air is pumped into the pump. However, the pressure is often unstable at this time. It may cause the pump to fail to suck liquid. Treatment method: Open the Purge valve and flush it with a flow rate of 3 to 5 ml/min. If it does not work, use a special syringe to suck out the air bubbles at the emptying valve.