Production of Diol-Modified Column Packing Material and Its Use in Novel HPLC Method for Urea Analysis in Dialysis Process

Dialysis is a widely used and effective method for removing urea from protein solutions, particularly following protein denaturation or solubilization. Due to its small molecular weight (~60 Da), urea can pass through semipermeable membranes, while larger protein molecules are retained. Therefore, the performance of dialyzers in urea removal is critically important. Dialyzer performance is commonly evaluated according to the method specified in the ISO 8637-1 standard, which is based on the measurement of urea concentrations in aqueous solutions before and after dialysis. In this study, a novel diol-modified silica, typically used as a column packing material in HPLC, was synthesized and applied as a new column. Using this column, a new HPLC method combined with a refractive index detector (RID) was developed for the sensitive and accurate determination of urea in aqueous sample solutions. This study is the first to report quantitative urea analysis using a diol-modified column packing material. Method accuracy was evaluated through standard addition-recovery experiments. The method showed recovery values between 99 and 110%, a limit of detection (LOD) of 0.66 mg/L, a limit of quantification (LOQ) of 2.0 mg/L, and a relative standard deviation (RSD) of 4.4%.