Maintain knowledge and experience in liquid chromatography

First, the mixed solvent must be filtered before it can be mixed. If it is mixed and filtered, it can only be used organically, and it must not be used in water. Since the water-based material is cellulose, the organic phase dissolves a little more or less cellulose, causing pollution.
Second, the standard sample or sample is best dissolved with the active phase (excluding special circumstances), can avoid a lot of trouble.
Third, to warm the column, the purpose of temperature increase, the department is to increase the solubility of the solute, but more importantly, reduce the viscosity of the solvent, thereby improving the peak shape and resolution. Note: Increasing the temperature will reduce the retention time, which also affects the resolution. The temperature change has a great influence on the width of the band, and the height of the plate is affected by the temperature, depending on the type of chromatography used. Always reduce the height of the tray.
Fourth, the column temperature of the C18 column should not exceed 40 degrees Celsius, otherwise the fitness phase will easily fall off.
5. Injector use, except for washing with water each time, after a period of time, it is best to use methanol or acetonitrile repeatedly to pick up and clean.
6. When using HPLC for analysis, the retention time sometimes drifts, sometimes it changes rapidly. What are the reasons? How to solve it?
A: About the drift topic:
1. The temperature control is not good. The solution is to use a constant temperature device to keep the column temperature constant 2. The active phase changes. The solution is to prevent the active phase from evaporating and reacting. 3. The column is not well balanced, and the column needs to be longer. Balance 7. Questions about rapid change:
1. The flow rate changes. The solution is to reset the flow rate so that it is not disordered. 2. There are bubbles in the pump, and the bubbles can be driven out by the operation of exhaust gas.
3. The activities are different. The solution is to change the active phase or make the active phase mix properly in the control room. 8. What is the cause of peaking or peaking in the liquid chromatography?
1. The screen plate is blocked or the column fails. The solution is to reverse the column, replace the sieve plate or replace the column.
2. There is interference peak. The solution is to use a longer column, change the active phase or replace the selective column. 9. What is the main reason for nonlinear shunt when splitting with capillary chromatography?
1. The temperature of the injector is too low, the vaporization of the sample is incomplete, and a grading split occurs.
2. The temperature of the injector is too high, some components may be thermally decomposed, and some samples may be catalytically decomposed, or the sample part may be adsorbed on the inner surface of the injector.
3. The sample is not mixed or even mixed before the split point.
4, the system's injection pad, column joints and other places leak.
Ten, when doing liquid chromatography analysis, the column pressure is not chaotic, what is the reason? How to solve?
A: The reasons may be:
1. There is air in the pump. The solution is to remove the air from the pump and degas the solvent.
2. The proportional valve fails and the proportional valve can be replaced.
3. The pump seal is damaged and the gasket can be replaced.
4, the bubble in the solvent, the solution is to degas the solvent, if necessary, change the degassing method;
5, the system leak detection, find the leak point, seal it.
6. Gradient elution, where pressure fluctuations are normal.
11. How to prevent tar-like pollutants from entering the capillary column when doing PGS/MS analysis?
A: Polymers, especially those containing nitrogen, sulfur and halogens, often have tar-like substances. In order to prevent the tar-like substance from entering the capillary column and causing pollution, the column function can be lowered. The protective pre-column can be used, that is, a pre-column is connected between the cracker and the capillary column, and the tar-like substance is retained by controlling the temperature of the pre-column. In the pre-column, the pre-fill can be placed in the GC gasification chamber, which can easily control the temperature and reduce the dead volume of the system. Of course, the pre-column packing needs to be replaced frequently.
12. I recently replaced another ODS column of the same grade. Although the separation is still possible, the retention time cannot be reproduced. Why?
A: This is because the analyte may have the ability to form hydrogen. Liquid Chromatographs Although the manufacturing technology of fillers has greatly improved over the past few years, the concentration of silanol groups on the surface of ODS fillers varies from manufacturer to manufacturer. It is precisely these silanol groups that may interact with the sample. Therefore, the relative retention time of the components in the uniform analyte on different grades of ODS columns may be different. Adding a small amount of competitor, such as triethylamine (TEA), to the active phase will saturate the bonding ability of the silanol groups, thereby ensuring good reproducibility of relative retention times on different grades of columns.
13. What is the service life of high temperature capillary columns?
A: The life of the capillary column depends on the function of the column itself, depending on the use, such as the temperature, sample state, injection volume, etc. If the sample is clean within the temperature range, the column is not In the case of pollution, the life of the column is generally between 2 and 3 years.

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