Hydrophilic interaction liquid chromatography (HILIC) is a powerful technique for separating polar compounds on polar stationary phases. This review discusses the characterization of HILIC stationary phases and their applications in complex matrices. HILIC employs polar stationary phases such as silica, amino, or cyano, but uses mobile phases similar to those in reversed-phase liquid chromatography (RP-LC). HILIC can analyze charged substances and is particularly useful for polar compounds that are difficult to separate using RP-LC. The retention mechanism in HILIC involves partitioning between the mobile and stationary phases, with the more hydrophilic analytes being retained on the polar stationary phase. The selection of mobile phases and additives, such as ionic compounds, can significantly affect retention. HILIC is well-suited for orthogonal and two-dimensional separations, and its orthogonality with RP-LC makes it ideal for multidimensional chromatography. HILIC has found applications in various fields, including pharmaceutical chemistry, proteomics, metabolomics, and food chemistry, due to its ability to handle polar compounds and its compatibility with mass spectrometry detection.Hydrophilic interaction liquid chromatography (HILIC) is a powerful technique for separating polar compounds on polar stationary phases. This review discusses the characterization of HILIC stationary phases and their applications in complex matrices. HILIC employs polar stationary phases such as silica, amino, or cyano, but uses mobile phases similar to those in reversed-phase liquid chromatography (RP-LC). HILIC can analyze charged substances and is particularly useful for polar compounds that are difficult to separate using RP-LC. The retention mechanism in HILIC involves partitioning between the mobile and stationary phases, with the more hydrophilic analytes being retained on the polar stationary phase. The selection of mobile phases and additives, such as ionic compounds, can significantly affect retention. HILIC is well-suited for orthogonal and two-dimensional separations, and its orthogonality with RP-LC makes it ideal for multidimensional chromatography. HILIC has found applications in various fields, including pharmaceutical chemistry, proteomics, metabolomics, and food chemistry, due to its ability to handle polar compounds and its compatibility with mass spectrometry detection.