Principles and Practice of Chromatography - Chromatography Applications > Liquid Chromatography Applications > Separation of Growth Regulators > Page 87

Separation of Growth Regulators

An example of the use of a C18 reverse phase column that separates substances purely on the basis of dispersive interactions is shown in figure 47. The packing is incorporated in a short column 3.3 cm long, 4.6 mm in diameter and packed with particles 3 mm in diameter. The example of its use is in the separation of mixture of growth regulators


The efficiency expected from such a column when operated at the optimum velocity would be about 5,500 theoretical plates. This is not a particularly high efficiency and, as a consequence, the separation relied heavily on the phases selected and the gradient employed. The separation was achieved by a complex mixture of competitive interactions, ionic and dispersive interactions between the solutes and the stationary phase and ionic, polar and dispersive between the solutes and the mobile phase. The gradient started with an initial solvent mixture of 1% acetic acid and 1 mM tetrabutyl ammonium phosphate buffered to a pH of 2.8. Initially the tetrabutyl ammonium salt would be adsorbed strongly on the reverse phase and thus acted as an adsorbed ion exchanger. During the program, acetonitrile was added to the solvent and initially this increased the dispersive interactions between the solute and the mobile phase.


Courtesy of Supelco Inc.


Figure 47 The Separation of a Mixture of Growth Regulators on a C18 Reverse Phase C18 Column