Development and evaluation of new reversed phase packing materials for ultra-high pressure liquid chromatography

The capabilities of ultra-high pressure liquid chromatography (UHPLC) were enhanced by the development of new stationary phase support materials. A porous organosilica hybrid material was developed through collaboration with Waters Corp. (Milford, MA). The porous material offers much greater surface area for stationary phase bonding, leading to columns with greater retentivity and sample loading capacity. This material was evaluated on the basis of chromatographic efficiency, mechanical strength, and loading capacity. 1.5-mum particles demonstrated plate heights as low as 2.5 mum. These particles were mechanically strong enough to handle run pressures in excess of 60,000 psi. They also demonstrated over 30-fold greater sample loading capacity than 1.0-mum nonporous silica particles.;Nonporous silica particles were synthesized for an investigation of the packing structure of UHPLC columns packed with monodisperse nonporous silica particles. To perform this experiment, 0.9-mum and 1.1-mum nonporous silica particles were synthesized and bonded with octadecylsilane stationary phase. The particles were packed separately into 50-mum i.d. capillary columns, then mixed and packed together. The performance of the mixed-particle columns was compared to that of the monodisperse columns.;In order to compare the performance of the porous particles to that of nonporous particles it was necessary to determine the diffusion coefficients of the test compounds. A capillary-based, stopped-flow method was developed that provided better than 1% relative standard deviation for the diffusion coefficients of four test compounds in two different mobile phase compositions. The method was then modified to allow the open-tube capillary to be pressurized during the pause in the flow. Diffusion coefficients for hydroquinone were measured at pressures up to 60,000 psi in 10% acetonitrile/90% water (V/V) and 50% acetonitrile/50% water (V/V). The diffusion coefficient was found to increase slightly with pressures up to 30,000 psi then remain relatively constant up to 60,000 psi in 10/90 acetonitrile/water. In 50/50 acetonitrile/water the diffusion coefficient of hydroquinone decreased linearly with pressure to a value approximately 35% lower at 60,000 psi than the value at atmospheric pressure.