Abstract
The document is a white paper on Hydrophilic Interaction Liquid Chromatography (HILIC) analysis method development. HILIC is a type of chromatography that uses an organic/aqueous mobile phase and a polar stationary phase. In HILIC, water is a strong solvent, and unlike in Reversed Phase Liquid Chromatography (RPLC), increasing the proportion of water in the mobile phase reduces the retention time of the analyte. The paper discusses when to consider HILIC analysis methods, the advantages of HILIC, and the challenges often encountered due to the lack of understanding of HILIC mechanisms compared to RPLC. It also provides a systematic flowchart for intelligent solutions for HILIC analysis method development, which includes a three-step approach for chromatography analysis method development. The first step involves gathering as much information as possible about the analyte (e.g., pKa, log P, log D). The second step involves analyzing the sample under different pH conditions using three HILIC columns in either isocratic or gradient mode to identify the suitable column/pH combination for the analyte. The third step involves optimizing the separation by investigating other parameters such as temperature and ionic strength, and assessing the robustness of the method. The paper emphasizes that the selection of the appropriate stationary/mobile phase combination, based on the differences between the HILIC stationary phases and the mobile phase pH, can provide high selectivity in the analysis. This step-by-step approach can help users develop an efficient analysis method.