Hydrophilic Interaction Chromatography - Anionic EHILIC

Anionic EHILIC

By analogy to the above, one can use an anion exchange (positively charged) column surface chemistry to reduce the influence on retention of cationic (positively charged) functional groups for a set of analytes, such as when selectively isolating phosphorylated peptides or sulfated polysaccharide molecules. Use of a pH between 1 and 2 pH units will reduce the polarity of two of the three ionizable oxygens of the phosphate group, and thus will allow easy desorption from the (oppositely charged) surface chemistry. It will also reduce the influence of negatively charged carboxyls in the analytes, since they will be protonated at this low a pH value, and thus contribute less overall polarity to the molecule. Any common, positively charged amino groups will be repelled from the column surface chemistry and thus these conditions enhance the role of the phosphate's polarity (as well as other neutral polar groups) in the separation.

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