Comparison of Reversed-Phase HPLC Separation Using Carbon Dioxide and Fluoroform for Enhanced-Fluidity Liquid Mobile Phases

Abstract

Lowering the viscosity of an HPLC mobile phase using liquid CO2/organic solvent mixtures has proven to be an effective means of increasing chromatographic efficiency, shortening analysis time, and lowering the pressure drop under ambient conditions. Such mixtures easily retain the high solvating power of common organic solvents but have low viscosities that are similar to those of supercritical fluids. To overcome the disadvantages of CO2 such as acidity in aqueous solution and reactivity toward some basic analytes, liquid mixtures of CHF3/methanol/H2O are investigated in HPLC separations. Triazine herbicides were used as the test solutes for reversed-phase HPLC. While either CO2- or CHF3-containing (20 mol %) enhanced-fluidity mobile phase provided better efficiency, shorter analysis time, and lower pressure drop than the methanol/H2O mobile phase, the mobile phase containing CHF3 was superior to that with CO2 in chromatographic performance. CO2 also influences the pH of the water-containing mobile phase, which results in resolution loss for some analytes with high pKas. The addition of a 10 mM phosphate buffer improved the efficiency for all the mobile phases, either with or without CO2 or CHF3, which affirms the importance of maintaining pH for ionizable analytes in reversed-phase HPLC using either common or enhanced-fluidity liquid mobile phase. The strongest enhancement in chromatographic performance was provided by the CHF3-containing mobile phase with buffer.