2.2.4. Purification of Peptides Using Ion Exchange Chromatography

The chromatography column enables the preparation of DEAE cellulose (diethylaminoethyl cellulose, MB110-5G) with a 4 cm thick bed. To perform our test, ethanol was used to wash before each use, using a 20 mm Tris-Hcl, and prepare a 100 mL volume, attuning pH to 8.5 after using NaOH. For the purpose of preparing the bed, counter ions (salt gradient) of 40 mL of 25 mm NaCl in 50 mm tris (pH-7.2) and 0.3% of DEAE were applied to the column. The elutes, using sodium phosphate citrate buffer (pH-7.2), were run for 4 h to collect 2 mL and collected in a beaker. Post dialysis, the crude AMPs were poured into the column without disturbing the bed and left for 20 min to settle. Next, the first eluting buffer, i.e., sodium phosphate citrate buffer (pH-7.2), was loaded into the column and used to elute the sample. The column was finally allowed to settle for 15–20 min when the elute was stored at freezing conditions.