Separation of Acids, Bases, Amino Acids, and Neutral Compounds using Mixed-Mode Chromatography

This application demonstrates the use of  Zodiac HST OG, a mixed-mode HPLC column, for the effective separation of a wide variety of chemical classes — including acids (e.g., mandelic acid), bases (e.g., pyridine, benzylamine), amino acids (e.g., tyrosine, phenylalanine), and neutral compounds (e.g., toluene, benzonitrile).

The figure illustrates how ion-exchange and reversed-phase mechanisms work synergistically to resolve compounds that vary in polarity and ionic character. Two different mobile phase compositions (with varying MeCN and TFA concentrations) showcase how selectivity and retention can be altered.

• Red chromatogram (MeCN 30%, TFA 0.2%): Stronger ion-exchange interaction, more retention for basic compounds.

• Blue chromatogram (MeCN 70%, TFA 0.05%): Higher organic content, reduced polarity interaction, better separation of neutral and acidic compounds.

This method is useful in pharmaceutical analysis, metabolite profiling, amino acid quantification, and chemical quality control.