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Understanding the recognition mechanisms occurring between histidine\u2010based ligands and nucleic acids enables more efficient purification of a DNA vaccine, as the binding and elution conditions can be adjusted in order to enhance the purification performance. Decreasing pH to slightly acidic conditions increases the positive charge of histidine ligand, what influences the type of interaction between chromatographic support and analytes. This was proven in this work, where hydrophobic effects established in the presence of ammonium sulfate were affected at pH 5.0 in comparison to pH 8.0, while electrostatic and cation\u2010\u03c0 interactions were intensified. Histidine ligand at pH 5.0 interacts with phosphate groups or aromatic rings of plasmid DNA. Due to different responses of RNA and pDNA on mobile phase changes, the elution order between RNA and pDNA was changed with mobile phase pH decrease from 8.0 to 5.0. The phenomenon was more evident with L\u2010histidine ligand due to more hydrophilic character, leading to an improved selectivity of L\u2010histidine\u2010modified chromatographic monolith, allowing the product recovery with 99% of purity (RNA removal). With the 1\u2010benzyl\u2010 L\u2010histidine ligand, stronger and less selective interactions with the nucleic acids were observed due to the additional hydrophobicity associated with the phenyl aromatic ring. Optimization of sample displacement chromatography parameters (especially (NH<jats:sub>4<\/jats:sub>)<jats:sub>2<\/jats:sub>SO<jats:sub>4<\/jats:sub> concentration) at slightly acidic pH enabled excellent isolation of pDNA, by the removal of RNA in a negative mode, with binding capacities above 1.5\u00a0mg pDNA per\u00a0mL of chromatographic support.<\/jats:p>","DOI":"10.1002\/elps.201700147","type":"journal-article","created":{"date-parts":[[2017,7,6]],"date-time":"2017-07-06T13:26:38Z","timestamp":1499347598000},"page":"2975-2980","source":"Crossref","is-referenced-by-count":8,"title":["Chromatographic HPV\u201016 E6\/E7 plasmid vaccine purification employing L\u2010histidine and 1\u2010benzyl\u2010L\u2010histidine affinity ligands"],"prefix":"10.1002","volume":"38","author":[{"given":"L\u00facia F. 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