Page 136 - Multidimensional Chromatography

128 Multidimensional Chromatography

chromatography has been treated theoretically and mathematical models have been
developed for the description of specific recycling techniques (56–60). Recycling is
a chromatographic technique mainly used in preparative LC to increase the effective
separation power (selectivity and efficiency), while avoiding the expense of purchas-
ing and operating longer columns and additional column sections. The preparative
recycling system can be improved by peak shaving (61). In this method, the recy-
cling valve is used during each cycle and switched to direct unwanted components to
waste, to collect portions of pure components of interest from peak fronts and tails,
and to recycle the remainder of incompletely separated sample mixtures back
through the column. An example of the advantages of a recycle chromatographic
method with peak shaving to separate closely related compounds with an value
near to 1 is shown in Figure 5.7. In this figure, two polymethoxyflavones were sepa-
rated to obtain pure compounds which were used as HPLC standards to construct
calibration graphs, employing cumarin as the internal standard, for their quantitative
evaluation in sweet orange and mandarin essential oils (62). Polymethoxyflavones
were isolated by a recycling system combined with peak shaving, having a configu-
ration similar to that shown in Figure 5.6. With the HPLC previously in the recycle
mode, the essential oils were fractionated on a glass column (300 60 mm id.)
filled with silica gel, using light petroleum/ethyl acetate (80:20 vol/vol) as the
mobile phase at a flow rate of 1.0 ml/min (62). As illustrated in Figure 5.7, in order to

Figure 5.7 Separation of tangeretin and heptamethoxyflavon by recycle HPLC: R, recycle;
C, collected; 1, tangeretin; 2, heptamethoxyflavon. Reprinted from Essenze Derivati
Agrumari, 63, L. Mondello et al., ‘Isolamento di polimetossiflavoni dagli olii essenziali di
arancia dolce e di mandarino mediante cromatografia su colonna e HPLC semipreparativa con
riciclo’, pp. 395–406, 1993, with permission from Essenze Derivati Agrumari.