Column switching, resolution

A practical method for enhancing the peak capacity, and thus the resolution of analytes in multicomponent complex mixtures, can be achieved by changing the mode of the separation during the chromatographic analysis, employing a column switching system in order to optimize a separation. 

Y. Oda, N. Asakawa, T. Kajima, Y. Yoshida and T. Sato, On-line determination and resolution of verapamil enantiomers by high-performance liquid chromatography with column-switching , Pharm. Res. 8 997-1001 (1991). 

E. A. Hogendoom, G. R. van der Hoff and P. van Zoonen, Automated sample clean-up and fractionation of organochlorine pesticides and polychlorinated biphenyls in human milk using NP-HPEC with column-switching , J. High Resolut. Chromatogr. 12 784-789 (1989). 

Oda, Y., Asakawa, N., Kajima, T., Yoshida, Y., Sato, T. (1991). Column-switching high-performance liquid chromatography for online simultaneous determination and resolution of enantiomers of verapamil and its metabolites. Pharm. Res. 8, 997-1001. 

Column selector valves can be added on as accessories to allow column switching for multi-dimensional chromatography (to increase the resolution of very complex samples such as in proteomics) or for automatic column selection (up to six columns) to facilitate methods development using different columns. 

MDGC is useful for separating compounds of an essential oil using two columns in line with different polarities. Through column-switching techniques, selected impure compounds in the first column can be diverted to the second column to ensure their complete separation. If the second column is chiral, then enantiomers potentially can be separated. The selected chiral stationary phase affects the resolution and separation drastically [73]. 

Another exciting development in multidimensional separation involves coupling of LC to GC or even to CE. In these coupling systems, LC can play a role as a high-resolution procedure for preliminary separation and/or trace enrichment. LC-GC can thereby be carried out in the column-switching and in the precolumn-analytical column mode. Although on-line LC-GC has not yet advanced widely into routine analysis, this powerful technique holds promise for the future. 

HPLC with column switching and mass spectrometry was applied to the online determination and resolution of the enantiomers of donepezil HC1 in plasma [38]. This system employs two avidin columns and fast atom bombardment-mass spectrometry (FAB-MS). A plasma sample was injected directly into an avidin trapping column (10 mm x 4.0 mm i.d.). The plasma protein was washed out from the trapping column immediately while donepezil HC1 was retained. After the column-switching procedure, donepezil HC1 was separated enantioselectivity in an avidin analytical column. The separated donepezil HC1 enantiomers were specifically detected by FAB-MS without interference from metabolites of donepezil HC1 and plasma constituents. The limit of quantification for each enantiomer of donepezil HC1 in plasma was 1.0 ng/ml and the intra-and inter-assay RSDs for the method were less than 5.2%. The assay was validated for enantioselective pharmacokinetic studies in the dog. 

A number of alternative techniques have been developed to tackle the general elution problem, such as flow and temperature programming, and especially column switching techniques. Coupled column chromatography involves the use of three and sometimes four different stationary phases. The sample is introduced to the primary column for a preliminary separation and then the various polar fractions are switched to the appropriate secondary columns and then to the detector. Coupled column chromatography provides enhanced resolution, and as isocratic elution is used the technique is particularly useful for the routine assay of samples with a wide range of capacity factor values. Further details on these various techniques may be found in a review by Koenigbauer and Majors [96]. 

The term column switching in liquid chromatography is used if two or more columns are connected to form a network. Column switching is a technique that changes the direction of flow of the mobile phase by valves, so the effluent from a primary column passes to a secondary column for a defined period of time as illustrated in Fig. 2. The objectives of column switching are to increase the chromatographic resolution and selectivity, to enrich trace amounts of sample, to protect sensitive detectors such as electrochemical detectors from contamination by coextractives, to prevent destabilization of the chromatographic equilibrium of the column by coextractives, and to achieve further objectives or a combination of... 

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