Column activation is necessary if the chromatographic column has dried out due to various reasons, and it needs to be wetted with the corresponding storage solvent before use to avoid potential damages. Without activation, normal peaks cannot be obtained, leading to incomplete detection or even damage to the chromatographic column. Therefore, if the filling of the column has dried out, activation is required. What are the situations that may cause the filling to dry out and require activation of the chromatographic column?
What conditions can lead to the drying of the column packing
and require activation of the chromatography column?
For new chromatographic columns, the plugs at both ends may loosen during storage and transportation, leading to the evaporation of the storage solution and drying out of the chromatographic column at both ends. In this case, activation should be carried out according to the instructions, usually for 1-4 hours (specific time should be based on the instructions). However, if the peak shape is still abnormal after a short time of activation, does it mean that the chromatographic column is damaged? No, this may be due to severe drying of the packing material, which cannot be fully wetted by short-term activation. In this case, it is necessary to extend the activation time of the chromatographic column. Generally, using the activation reagent specified in the instructions and activating it overnight at a flow rate of 0.2-0.3 mL/min can solve the problem.
Does an old chromatography column never need to be activated? Not necessarily. For chromatography columns that have been used, if they have not been used for more than a month, the storage solution may also be lost. Especially for the chromatography column that has not been removed from the instrument for more than a week, it needs to be reactivated before use.
What issues should be noted when activating a chromatography column?
● For chromatography columns that have not been used for a long time or have been newly opened, it is generally not recommended to activate them directly with the detection flow rate. It is necessary to maintain a low flow rate for a period of time before slowly increasing the flow rate. Otherwise, it is easy to damage the packing material due to the instantaneous high flow rate, causing damage to the chromatography column.
● When taking out a chromatography column from the refrigerator or when the instrument is in a high temperature state, it is necessary to let the column return to room temperature, and lower the column oven temperature to room temperature before connecting the chromatography column. Otherwise, it is easy to cause damage to the chromatography column due to the sudden temperature difference.
● If the chromatographic column needs to be activated due to abnormal peak shape during detection, before using the activation reagent, it is necessary to fully replace the solvent in the chromatographic column with a transition mobile phase, and then use the activation reagent for activation. If the reverse-phase chromatographic column detects a mobile phase containing salt, before activation, it is necessary to completely replace the salt with a mobile phase of low organic phase ratio, and then use the activation reagent for activation.
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