During the use of the chromatographic column, there will be high column pressure and poor peak shape. The main reason is the contamination of the chromatographic column packing materials, of which the packing materials are a special kind of contamination of the packing materials. Packing materials to analyze the reasons and solutions.
Performance of packing materials compaction
The packing materials of the chromatographic column are hardened. After the column is opened, it will be found that the color of the packing materials does not change, but the packing materials become flakes, blocks, and crystals.
Reason analysis and solution of packing materials compaction
1) Whether it is a new column or a column that has already been used, when using a mobile phase containing salt, first use a solution with the same composition as the mobile phase but without salt for flushing and replacement (avoid using pure water), and the recommended time is 30 above the column volume.
2) Mobile phase replacement. In the process of method development, various attempts with different buffer salt systems and different organic phase systems will be involved in most cases. In the process of adjusting and replacing the composition of the mobile phase, be sure to fully consider different types of buffers. Dissolution of salts. If you are not sure about the mutual solubility between the buffer salt used and the organic phase, you can mix the two according to different ratios, let stand for 30 minutes, and observe whether there is precipitation or floc at the bottom.
3) Gradient conditions, when it comes to buffer salt-organic phase gradient conditions, please confirm whether the buffer salt can be fully dissolved at the highest point of the organic phase in the gradient conditions. Of course, when you are not sure, you can also mix the buffer salt and the organic phase in the highest proportion of the gradient and let it stand to observe whether there is precipitation, as in the above method.
2.The preparation time of the mobile phase is too long, the mobile phase will grow bacteria, or the organic phase such as acetonitrile will cause flocculent precipitation.
Solution: The mobile phase should be prepared as needed, and the pure water mobile phase should be prepared and used for no more than 12 hours at a time. Other mobile phases containing organic phase should not be used for more than 24 hours at a time, and the shelf life of the mobile phase can also be adjusted according to the actual situation.
3.Others: samples such as gelatinous and flocculent samples cannot be removed by filtration, and they will also accumulate on the sieve plate and the head end of the column, causing the column pressure to rise.
Solution: The gelatinous and flocculent samples pass through the process, and the pretreatment is removed before entering the liquid chromatography system.