Page 135 - Multidimensional Chromatography

Coupled-Column Liquid Chromatography 127

(number of columns connected in series), although the column pressure drop
increases linearly with the length. In practice, the operating conditions essentially
limit the number of columns that may be coupled together and therefore restricts the
total number of theoretical plates that can be generated in a reasonable time. An
alternative way to improve the number of theoretical plates is the use of the same
column in a recycling chromatographic system (55). This technique consists of a
switching-column method which is accomplished by adding a switching valve to the
fluid system to permit the eluent, or some designed portion thereof, to be directed
from the column outlet back into the column inlet. The process can then be repeated,
with each subsequent pass resulting in an increase of the separation between the
peaks. In practice, recycling increases the effective length of the column by returning
the eluted analyte to the head of the column for further separation. A schematic rep-
resentation of a conventional chromatographic recycling system is depicted in Figure
5.6. The recycling technique offers, with respect to coupled columns, the possibility
to increase the resolution by a large number of theoretical plates obtained by increas-
ing the number of sample passes through the HPLC column and maintaining the col-
umn length and pressure constant. There are two limiting factors in recycling
chromatographic methods, as follows: (i) the phenomenon of remixing of the
separated analytes, which occurs when the fastest moving peak of the sample over-
takes the slowest moving components after a certain number of cycles; (ii) the extra-
column band broading occurring in the pump, valve devices, connecting tubes, and
detector at each cycle in the detector-to-injection transfer. The influence of column
and extra-column effects on the maximum efficiency realizable in recycling

Figure 5.6 Schematic design for recycling chromatography in which the effluent is recy-
cled through the pump: 1, eluent; 2, pump; 3, injector; 4, chromatographic column; 5, detec-
tor; 6, three way valve; 7, waste; 8, T-connection.