Design protocol, column

The information contained in the three data bases provides the necessary information required to design the optimum column. In addition, once the column has been designed, and its properties defined, a complementary set of Analytical Specifications can also be calculated. Thus, the design protocol contains three data bases. Performance Criteria, Elective Variables and Instrument Constraints. 

These data bases will provide, first, the column specifications and, second, the analytical specifications. A diagram representing the overall design protocol is shown diagramatically in Figure 1. 

The Design Protocol contains three different sources of data which will be termed the column design Data Bases. The needs of the analyst will constitute the first data base which will be given the title Performance Criteria. The performance criteria must state explicitly, in numerical terms, the quality of the separation that is required in order to achieve an accurate analysis. 

The column design protocol, therefore, consists of three data bases, performance criteria, elective variables and instruments constraints. These data bases will provide, firstly, the column specifications and finally, the analytical specifications. A diagram representing the overall design protocol is shown in figure (1). The four different components of the column design protocol will now be discussed in detail. 

It is obvious that such a protocol would not be employed to design a column for a single analysis or even for a few dozen analyses. The optimization procedure entails a considerable amount of work and therefore, would only be justified for a routine analysis that was repetitive and would be carried... 

Degradation of the sample and related problems, such as the concentration effect and anomalous flow, are the more important problems in the fractionation of UHMM polymers. The critical point in the characterization of the UHMM polymers is the fractionation in the SEC columns. For a successful fractionation of UHMM macromolecules, one must use specifically designed SEC columns with large particle sizes and ultralarge pore sizes. Furthermore, many aspects of the experimental protocol, such as flow rate and sample concentration, which is not critical in the usual molar mass range, become determining with UHMM polymers. A successful characterization of UHMM polymers requires optimization of the experimental protocol. Each step of the experimental protocol should be performed methodically to achieve the absence of sample degradation and reliable results. 

In a similar manner to the design process for packed columns, the physical characteristics and the performance specifications can be calculated theoretically for open tubular columns. The same protocol will be observed and again, the procedure involves the use of a number of equations that have been previously derived and/or discussed. However, it will be seen that as a result of the geometric simplicity of the open tubular column, there are no packing factors and no multi-path term and so the equations that result are far less complex and easier to manipulate and to understand. 

There have been very few method development processes proposed for 2DLC. One study (Schoenmakers et al., 2006) is titled A protocol for designing comprehensive two-dimensional liquid chromatography separation systems. This study advocates that one initially chooses the first-dimension maximum acceptable analysis time, the first-dimension maximum workable pressure drop, and the smallest first-dimension column diameter. The first two variables are then used to construct a Poppe plot (Poppe, 1997)—pronounced Pop-puh (Eksteen, 2007). 

In addition, analysts can create a multimodal SPE column by stacking several discs with differing bonded phase in the disc holder. With the proper chemical design, such multimodal systems can provide customized separation protocols. The stacking order of the discs depends on the particular physicochemical characteristics of the analysis under way. The general rule is to place the disc with the more selective extraction mechanism on top of the less selective adsorbent. 

In designing an assay for an enzyme, it is often necessary to introduce a termination step into the protocol (see Chapter 1). This is often done when protein is present in the incubation mixture at a concentration that would dog the column. There are a variety of ways to accomplish this termination process. Ideally, it would not be necessary to add reagents that might otherwise clog the column or alter its performance. For example, consider the changes that occur in the incubation mixture when the reaction is terminated by add. The addition of trichloroacetic acid (TCA) will reduce the pH of the incubation... 

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