Optimization of buffer conditions for the removal of endotoxins using Q-sepharose

Anion-exchange chromatography using Q-sepharose was used to remove endotoxins. The optimized buffer conditions of pH, buffer concentration and NaCl concentration provided four log reductions in the endotoxin level (from 5 x 10⁵ EU/ml to 10 EU/ml). In glycine-NaOH buffer (pH 8.0-10.5), the upper limit of the concentration of NaCl was 300 mM and that of glycine was 50 mM. For the Tris-HCl buffer (pH 7.5-9.0), the upper limit of the concentration of NaCl and buffer was 200 and 75 mM, respectively. In phosphate buffer (pH 6.0-8.0), the presence of phosphate significantly reduced the adsorption of endotoxins on Q-sepharose. The upper limit of the concentration of NaCl and phosphate in the phosphate buffer was 90 and 25 mM, respectively. The pH of the buffers showed very limited effects on the adsorption removal of endotoxins.