Visualization of solute migration in chromatographic columns: Quantitation of the concentration in a migrating zone

The concentration distribution across a zone of iodine migrating along a column made of glass, packed with C18-bonded silica, and eluted with carbon tetrachloride was derived from a quantitative analysis of the photographs of the zone. The photographs were scanned and turned into digital images. The intensity distributions obtained from the measurement of the grayscale intensity were converted into concentration profiles using a calibration method. This procedure is illustrated and suitable corrections are introduced to account for the transverse variation of the optical path length, as a result of using a cylindrical detector cell (the column itself), and for the refraction of light due to the differences between the refraction indices of the glass wall and the liquids involved. An error analysis is also reported. It shows that the method can reliably produce results with a precision of a few percent, allowing on-column evaluation of column performance and the derivation of the radial distributions of the column efficiency, the migration velocity of the zone, and the sample distribution at the head of the column.