SYSTEMATIC APPROACHES TO HPLC METHOD DEVELOPMENT FOR REVERSED-PHASE SEPARATION

Several approaches to HPLC method development have been reported in th e literature, many of which rely on computer software to minimize the total number of experimental runs. The major decision in method develo pment is how best to vary band spacing for improved separation of a sa mple. The present report summarizes the present status of computer-ass isted reversed-phase HPLC method development, and illustrates two poss ible approaches in more detail: (a) optimization of mobile phase compo sition for mixtures of acetonitrile, methanol and buffer; (b) optimiza tion of separation temperature and either gradient steepness or isocra tic %B.