Optimization of conditions for the separation of peptide mixtures using strong cation exchange chromatography

The separation conditions for strong cation exchange chromatography were investigated and compared using tryptic digest of protein extract from yeast. The conditions included the loading amount of the sample, the type of the salt in mobile phase, the type of the acid used to adjust the pH of the mobile phase and the proportion of organic solvent added in the elution solution. The experimental results indicated that high separation efficiency can be obtained by using ammonium chloride solution as optimal mobile phase, phosphoric acid to adjust the pH of mobile phases and 30% (v/v) organic solvent in mobile phases. The results suggested that a beneficial reference was presented for the selection of separation conditions of strong cation exchange chromatography when applying in two-dimensional chromatography-mass spectrometry strategy in proteomic research.