Monolithic columns flow rate through

In more demanding separations that require higher plate counts, specially designed rapid analysis columns packed with very high efficiency 2 to 3 /.an porous particles are available from several manufacturers. In addition, monolithic columns with improved flow-through characteristics are also commercially available. Figure 13.4 depicts a comparison of inlet pressure and flow rate for 4.6 mm inner diameter x 50, 100, and 150 mm columns packed with 5 /an particles. 

Commercially available monolithic columns are based either on silica or organic polymer and are generally characterized as a polymeric skeleton with macropores, with a diameter of approximately 2 pm, and mesopores, with a diameter of approximately 13 nm. The role of the macropores (through-pores) is to provide channels with high compounds permeability, which permits the use of higher flow rates with respect to columns based on conventional particle size, and an extended surface area, which is comparable to conventional columns packed with 3 pm particles. 

Because of its open, rigid structure, solvent flows through the monolithic column with relatively little resistance. The same pressure required to obtain a flow rate of 1 ml./min with 3.5-pm spherical particles provides a flow rate of 9 mL/min in the monolithic column. At 9 mL/min, the plate height in the monolithic column is only 50% greater than the minimum plate height observed at 2 mL/min. 

Instead of packed columns, monolithic (continuous bed), analytical, or capillary columns in the form of a rod with flow-through pores offer high porosity and improved permeability. Silica-based monolithic columns are generally prepared by gelation of a silica sol to a continuous sol-gel network, onto which a Cjg or another stationary phase is subsequently chemically bonded. Such columns provide comparable efficiency and sample capacity as conventional columns packed with 5-pm particle materials, but have three to five times lower flow resistance, thereby allowing higher flow rates and fast HPLC analyses. Rigid polyacrylamide, polyacrylate, polymethacrylate, or polystyrene monolithic columns are prepared by in sim polymerization. 

Extremely rapid RP separations of proteins have been demonstrated on poly(styrene-co-divinylbenzene) monolithic columns. A fast gradient and a flow rate of lOmL/min were used to separate live model proteins in less than 20 s on a 50 X 4.6-mm ID column [53].The same type of monolithic structure has also been prepared in capillary columns of 200 pm ID and showed excellent performance in the identification of proteins by LC-MS through peptide mass fingerprinting and accurate intact molecular mass determination [24]. 

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