Problem-Solving Approach to Chromatography in the Biochemistry Lab

Students are given mixtures of colored compounds and three chromatographic matricies (cation exchange, anion exchange, and gel filtration). In the first of two lab periods students are given two mixtures of three components each. They run each of these samples on each of the three columns. Before the next lab period, each student is expected to design a multi-column procedure for separating a five-component mixture using the knowledge gained from the first phase of the experiment. In the second lab period, students perform the procedure they have devised. This experiment gives students experience in linked multiple chromatographic steps that are typically used in protein purification. Colored compounds are used instead of proteins to eliminate the need for reading fractions in a spectrophotometer and to allow students to watch the separations happen in real time . Since the physical and chemical properties (size, charge, etc.) of the compounds involved are well defined, students are able to correlate these properties with the chromatographic behavior of the compound. This experiment also provides an occasion for students to exercise their problem-solving skills