Empirical correlation of retention factor of mononucleotides to buffer concentration in RP-HPLC

The retention factors of ionic samples with the types and concentrations of buffers were considered in RP-HPLC. The samples were the five mononucleotides and the buffers were acetic acid, sodium phosphate monobasic, potassium phosphate monobasic, and ammonium phosphate monobasic. The mobile phase was composed of water and 5 vol.-% methanol with the different concentrations of buffers. The three empirical correlations were suggested to predict retention factors of the ionized samples based on the assumption that the factor was proportional to the concentration of buffers. From the regression analysis of the correlation equations, the concentration of acetic acid is square root proportional to the retention factor, while the concentrations of other monobasic buffers are almost linearly proportional.