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LC Column Cleaning and Regeneration

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Over time, our liquid chromatography columns will accumulate some contamination, especially if we regularly analyze dirty samples. There are some steps that we can take to keep our columns clean. Further, we can do some limited column regeneration as well, which can often help restore some performance to our columns and extend their lifetime.

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Thanks for joining us for the latest Restek tip, where I’ll discuss LC column cleaning and regeneration. Column contamination from adsorption of analyte or matrix impurities can have adverse chromatographic effectssuch as split peaks, changes in selectivity, or reduced sensitivityall of which can ultimately impact quantitation. It is highly recommended to use guard columns or an UltraShield precolumn filter to protect your analytical columns. However, over time, even with this precaution, you may observe decreased column performance, increased backpressure, or other signs of column contamination.

For HPLC or UHPLC column cleaning and regeneration, flushing using a series of stronger solvent combinations is recommended with the flow in the direction shown on the column. For reversed phase columns, flush the column with a minimum of 20 column volumes of each solvent in the order shown in the table starting from 5% methanol in water to highly organic solvents and return to your original mobile phase conditions. For normal phase columns, flush with a minimum of 20 column volumes of each solvent in the order shown in the table starting from isopropyl alcohol to ethanol and return to your original mobile phase conditions. Column volume can be estimated using the formula shown here. It is highly recommended to re-equilibrate your column to your original method conditions by performing a series of injections prior to starting your batch. After column regeneration, it is important to store your LC columns with end plugs securely fastened and with your column manufacturer’s recommended solvent for either short- or long-term storage.

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