I would like to know how I can regenerate a C18 column, following the formation of distorted and/or double peaks. I try with a series of washings with water and acetonitrile (good results but for a short time)
The fast return of the problem suggests that your problem could be contaminated samples rather than ineffectiveness of your cleaning procedure. If so you should search for a clean-up step for your samples to solve the problem like preperative C18 (SPE) or protein precipitation/filtration.
Hi, the best way to use to wash the column is by using aliquots of 1-10% 3N TFA or nitric acid. then you will regenerate mostly. If you use proteins or peptides its best method. Still donot work! then heat the column to 50 °C. by this methods if the pores of C18 is filled with active charcoal, will be reopens. But take care donot use more pressure. it can damage the column packing..
The common practice of cleaning the RP column after use is to flash either with 1:1 Acetonitrile: Water or 1:1 Methanol : Water. If you continue having problems after clean up you may have contaminated guard column or your sample prep is not appropriate or your mobile phase needs sonication. Air bubbles will give you a lot of noise peaks.
Thanks! Thinking about your comments, I really believe in the need to clean up my samples. My samples are lichens and the extraction solvent is absolute ethanol... which molecule/compound can lead to the formation of distorted and/or double peaks?
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