Preparative Column Design

Such limitations that are imposed on the preparative column design is inevitable. There will be practical limits to the column length, the analysis time and the column diameter. There will also be more subtle limitations,... 

PREPARATIVE COLUMN DESIGN EQUATIONS Optimum Particle Diameter... 

A two-column electron capture gas chromatographic system, as described under Apparatus, is used for the analysis. Take 5.0-/ liter aliquots of the concentrated extract and inject them into the gas chromatograph. The injections are made on two different types of specially prepared columns, designated as column A and column B (see section on Experimental Results). 

Although, apparently simple, equation (1) has some very significant implications on preparative column design. It is clear from equation (1) that increasing radius and length of the column increases both the maximum sample volume and the maximum sample mass. However, increasing the column length will also increase the columns efficiency (unless the particle diameter is also increased). 

It appears that the equation introduced by Van Deemter is still the simplest and the most reliable for use in general column design. Nevertheless, all the equations helped to further understand the processes that occur in the column. In particular, in addition to describing dispersion, the Kennedy and Knox equation can also be employed to assess the efficiency of the packing procedure used in the preparation of a chromatography column. 

Prepare a sieve-plate column design for the chlorobenzene distillation and make dimensioned sketches showing details of the plate layout including the weir and the downcomer. 

In previous chapters, liquid chromatography column theory has been developed to explain solute retention, band dispersion, column properties and optimum column design for columns that are to be used for purely analytical purposes. The theories considered so far, have assumed that solute concentrations approach (for all practical purposes) infinite dilution, and, as a consequence, all isotherms are linear. In the specific design of the optimum preparative column for a particular preparative separation, initially, the same assumptions will be made. 

As in the previous chapters on column design, the characteristics of many of the equations discussed in this chapter will be examined employing realistic chromatographic conditions and the typical conditions chosen for a preparative column are given in table 1. 

It is seen from figure 2 that the optimum column length ranges from over 500 meters to a fraction of a millimeter. It is also obvious that there must be further limitations placed on the design of the column to ensure its practical use. A preparative column less than 5 cm in length would be very difficult to pack as would a preparative column that had a length greater... 

Alternatively, a terminal tag such as polyhistidine can be added to the target protein to improve the efficiency of the purification procedure [24]. An affinity column designed to recognize polyhistidine could be used to isolate the tagged protein.The polyhistidine is then cleaved and the mixture dialyzed. While this approach is useful for small-scale preparations, whether it can be used for large pharmaceutical scale preparations remains to be seen. 

The same factor is used for the scale-up of the sample load. The translation of an analytical method to a preparative system is obviously dependent on the sorbent characteristics of the preparative column. Several reviews on preparative methods, column-packing techniques, theory, and equipment design have been published [132-135,145]. Specific examples of the application of preparative procedures are listed in Table 5.3. 

Column design and preparation incorporated previously described methods reported in the literature (39). Two different adsorbents were employed a 100/120 mesh crosslinked styrene/ divinylbenzene resin (Polypak P-Waters Associates) and a Woelm aniontropic activity grade alumina. These adsorbents were packed in 300 and 94 cm. stainless steel columns having a 1 mm. internal diameter. Pressure drop across the adsorbent bed was kept to a minimum (<0.02 atm.) by using a heated pressure reduction valve at the end of the column. Typical linear flow velocities through the columns were in the range of 0.27-2.17 cm/sec. 

---