L-Cysteine (L-Cys) and N-Isopropyl acrylamide (NIPAM)-modified silica spheres as a novel stationary phase for
hydrophilic interaction liquid chromatography (HILIC) was produced firstly by cross-linking polymerization.
Some characterizations in this article confirmed that the synthesis of Cys-Si-NIPAM is successful. Some polymer
layers can be observed through transmission electron microscopy (TEM). In addition, through nitrogen ad-
sorption porosity method, scanning electron microscopy (SEM), thermal gravimetric analysis (TGA) and other
characterization methods, we can find the significant changes after modify. It has good hydrophilic property and
higher column effect than bare silica column, Si-Cys column and Si-NIPAM column under the same conditions. It
has good separation effect for some hydrophilic analytes such as 5 nucleosides and nucleoside bases, 5 amino
acids, 4 sulfonamide drugs and 10 saccharides. The effects including column temperature, pH and organic
solvent content on chromatographic performance were studied, which proved that hydrophilic interactions can
be simultaneously existed between the stationary phase and the analytes. In addition, reproducibility and effi-
ciency of the Cys-Si-NIPAM column was also investigated, the results illustrated that the stationary phase have
passable stability (the intraday RSDs 0.08–0.44%, n = 3 and the interday RSD 0.46–3.50%, n = 3) and ideal
efficiency (plates per meter, ~45700 plates/m). In conclusion, the preparation process of this hydrophilic liquid
chromatography stationary phase is not only simple, but also can meet the basic requirements for the separation
of hydrophilic analytes.