Advances in Biochemical Engineering. Biotechnology Modern

This assessment issues the hot chromatographic technique – capillary electrochromatography(CEC) – that's lately receiving awesome awareness. the foundations of this system basedon a mix of electroosmotic move and analyte-stationary section interactions, CEC instrumentation,capillary column expertise, separation stipulations, and examples of a varietyof functions are mentioned intimately.

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Unfortunately, on-column detection is required in CEC to avoid excessive peak broadening and loss of the high intrinsic efficiency of this method. Therefore, the capillary diameter is the only light pass length available for detection. This situation dramatically impairs the sensitivity of the UV detection. Several attempts to address this weakness such as the high sensitivity Z-cell developed by Agilent Technologies did not meet with a broad acceptance. The very sensitive laser induced fluorescence is another option; however, only a small fraction of molecules fluoresce while a tedious functionalization must precede the detection of others.

Besides all these constrains, the cost of the production process is an important factor, which consequently forces the separation scientist to be effective with regard to the scaleup of the separation processes and to use reliable, fast means of controlling them. Thus liquid chromatographic supports should meet the following requirements [2, 4–6]: – fast and efficient separation in order to decrease losses due to biomolecule degradation – high, flow unaffected capacity – good, flow unaffected resolution – low backpressure – easy and fast scale-up and scale-down potential – meet all safety regulations, especially regarding leachables and sanitation – the same material may be used for analytical and preparative purposes – easy to handle and operate – stable even if harsh conditions are applied during sanitation – high batch-to-batch reproducibility Chromatographic columns packed with conventional porous particles meet these requirements only to a limited extent.

The first two groups, Nakanishi and co-workers [32] and Hjerten [33] were trying to replace the standard (long) chromatographic column with the new type of continuous support allowing convective mass transport. e. As a result the theory of short column layer was developed, which is based on the fact that in gradient chromatography of proteins and other biomacromolecules, a critical distance X0 exists at which the separation of zones is at a maximum and band spreading is at a minimum. With step gradients and high elution velocity the column length may be reduced to the level of membrane thickness [35] as will be shown later in this chapter.

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