Purification of Peptides and Proteins Using Monolithic Silica Disk-Packed Spin Columns

抄録

Sample pretreatment is an important step in chromatographic analysis of peptides and proteins. In this study, we investigated the purification of peptides and proteins by using monolithic silica disk-packed spin columns modified with octadecylsilyl moieties. First, digested bovine serum albumin (BSA) peptide samples were purified using the spin columns to examine the purification of peptides. The spin columns removed polar compounds from the samples and reduced the salt concentration. When adrenomedullin, a vasodilating peptide, was spiked into a control serum sample, the addition of trifluoroacetic acid was necessary to remove the pigment derived from the serum while maintaining high recovery. Next, protein purification was investigated and its recovery was evaluated using samples containing ribonuclease A, insulin, lysozyme, BSA, and myoglobin. When ethanol was used as the organic solvent, almost 100% recovery was achieved for all proteins, except BSA. Furthermore, we investigated the quantity of protein loaded onto the spin column. Insulin was more retained on the spin column than BSA. Insulin, a small molecule, can easily penetrate the pores of the stationary phase and thus is easily retained in the stationary phase of the spin column. These results indicate that this method is suitable for the purification of peptides and proteins using spin columns.