Abstract
A C 4 column (InertsilWP300 C 4) was developed for a protein analysis in reversed-phase chromatography. Polar groups were embedded on this phase. And this column was evaluated with low concentration of trifluoroacetic acid (TFA) and formic acid. Test C 4 phases that controlled carbon loading and commercially available columns were compared with this embedded C 4 column. The test column with higher carbon loading showed good separations of proteins at low concentrations of TFA, although hydrophobic proteins were broadened. On the other hand, embedded polar C 4 packings showed ideal retention behavior for proteins, which is independent of ion-pairing effects of additives in mobile phase. The reason was concluded to be the shielding effects of polar groups on the sorbents, which reduce silanophilic interactions with polar groups of proteins.
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