Method development for high performance liquid chromatography: novel organic modifiers and column packing conditions
Neutral eluents are proved to be applicable for the separations of organic acids and bases on reversed phase liquid chromatographic columns when an appropriate organic modifier is present. Straight chain alcohols, diols and amino alcohols are excellent candidates. The modifier can be incorporated into the eluent at low percentage or used to pre-treat the column. The function of the modifiers on the separations is to coat the surface of the stationary phase and make it less hydrophobic;Novel surfactant additives were used in mobile phases to improve the separations of various organic compounds. Compared with separations obtained without additives, shorter retention times and sharper peaks are obtained. The reason for the improvement appears to be a stronger solution-phase interaction between analyte and discrete additive molecules. The effect of additives is akin to gradient elution. Solvent strength and selectivity can be varied by controlling the type and concentration of the additive. Binding constants between solute and surfactant additives were calculated by relating capacity factor to surfactant concentration;An even greater reduction in k' resulted from the use of binary additive systems, particularly those containing a cationic and an anionic surfactant. It is believed that an ion-pair is formed that is more effective in forming solution complexes with organic analytes than either additive alone. Other binary additive systems included mixtures of a neutral and a charged additive or a mixture of two anionic surfactant additives. A ternary mixture of additives was the most effective of all in modifying the chromatographic behavior of organic solutes in HPLC;Study of HPLC column packing conditions shows that the choice of slurry/packing solvent is critical to obtain a high efficiency column. Lower packing pressure yields columns with better performance when an appropriate solvent was chosen. A simple resin sizing method gives resin with narrower size range and thus better chromatographic columns.