Liquid Chromatography as a separation tool has made significant strides in a systematic study. The most widely used chromatographic process is reverse-phase liquid chromatography (RPLC), which uses polar mobile and hydrophobic stationary phases. However, specific polar composites are challenging to study using this approach. Normal-phase liquid chromatography (NPLC) is another separation process that uses polar stationary phase and organic eluents. When analyzing polar compounds, NPLC produces asymmetric chromatographic peak forms and low-efficiency separations. HILIC is a promising alternative approach for polar compound analysis. HILIC is a separation process that blends the stationary phases of the NPLC method with the mobile phase of the RPLC technique. Hydrophilic interaction liquid chromatography (HILIC) was used to separate small molecules, medicinal compounds, metabolites, contaminants, sugars, peptides, oligosaccharides, proteins, and amino acids, among other liquid chromatographic separations. Tiny polar composites can be efficiently separated on polar stationary phases using HILIC. The target of the study was to look at the different ways to characterize HILIC stationary phases and how they can be used to separate polar compounds in complex matrices. Since processes other than hydrophilic partitioning may exist, the features of the hydrophilic stationary phase may influence in some situations, restricting the selections of ion intensity, mobile phase construction, and buffer pH value available. Increasing our knowledge of HILIC retention activity broadens the range of liquid chromatography applications. HILIC systems bio uses are also discussed. This paper gives a broad description of how HILIC has been used in pharmaceutical testing in various sample matrices, including plasma, pharmaceutical dosage forms, environmental samples, serum, plant origin samples, and animal origin samples. This study also reflects on the most modern and a selection of papers in pharmaceutical science from 1991 to the suggestion deadline in 2020, which deal with examining various mechanisms using HILIC.