Required analysis time

In addition to the 17 operators, "supertypes" may be defined for subsystems or groups ol components that recur in a system. This reduces the required analysis time and adds clarity to itic analysis of subsystems. A supertype is a collection of operators defined and used as a. single entity. 

Therefore we shall optimize the experimental conditions by looking for the minimum pressure at constant analysis time and efficiency for a given solute pair. It has been shown that this goal is accomplished when the column is operated at the optimum flowrate at which the plate height is minimum (19). The particle size and column length then depend on the plate number and the required analysis time. 

Finally we may ask the question how charts like those depicted in Figs. 5 to 8 can be used to determine whether a given column can meet some performance requirements. It is most probable that it will give either a too large or too small efficiency within a required analysis time. How can we find it out from the diagram ... 

Increasing a is generally advantageous for the separation of two peaks. If a series of solutes needs to be separated, then an increase in one of the a values may have an adverse effect on the required analysis time. The problem of how to distribute the peaks over the chromatogram will be discussed extensively in chapter 4. 

The required analysis time itself appears to be both a logical and an elegant choice for an optimization criterion. Either fne can be minimized, or, for reasons of consistency, l//ne can be maximized. The criterion of minimum required analysis time then corresponds to a constant value for C in eqn.(4.37) ... 

Figure 4.11 Calculated characteristics for optimum chromatograms (r = 1) containing 10 equally resolved peaks as a function of the separation factor S. Plotted on a logarithmic scale are the capacity factor of last peak (1 +k eqn.4.46), the required number of plates (Afne eqn.4.47), the required analysis time under conditions of constant flow rate and particle diameter (rne f>(ji eqn.4.48), and required analysis time under conditions of constant pressure drop fne p eqn.4.49). For explanation see text.

Outside the optimum range this is no longer true. If ten peaks are equally resolved (r = 1) with S values of 0.001, then according to eqn.(4.47), four million plates are required for adequate resolution. Moreover, we can see from figure 4.11 that the required analysis time is a factor of about 600 larger (under constant flow and diameter conditions) than it would be if S equalled 0.1. If S was 0.5, the analysis time would be a factor of about 200 larger than in the optimum. Hence, we may conclude that for optimization processes during which the capacity factors may be expected to vary dramatically, a time correction factor is required even when r is used as the optimization criterion. 

Required analysis times and time-corrected resolution products for the three chromatograms of figure 4.12. Constant flow rate and diameter (of open columns or particles) (i.e. constant f,d conditions) have been assumed. 

Finally, in the third stage of the process, a procedure similar to that of the second stage may be followed to reduce the resolution of abundantly resolved pairs of peaks (Rs> 2). During this stage, slopes may be increased and isothermal periods shortened, leading to a reduction of the required analysis time. Figure 6.10d shows the result obtained after an additional two chromatograms. It is seen that the analysis time has been reduced from about 43 to about 37 minutes. 

If the retention vs. composition plots of all solutes are known, then it is in principle possible to calculate the optimum program parameters for a simple, continuous gradient (figure 6.2a-d). In such a procedure an appropriate optimization criterion can be selected such that the distribution of all the peaks over the chromatogram, as well as the required analysis time, can be taken into account (see chapter 4). 

The capacity factor of the last peak (k. Once a column and flow rate have been selected for the analysis, this parameter determines the required analysis time (ta>). 

A possible fourth column in illustrated in figure 7.2 by an open circle. This 80 cm long column packed with 10 pm particles is not an attractive column in practice, because of both its length and the required analysis time. However, it should be noted that shorter columns... 

This example emphasizes the importance of balancing the column dimensions and the injection volume, with the required analysis time and sensitivity. If the main goal is the speed of analysis, then a smaller the injection volume is better if, on the other hand, the sensitivity is critical, then it is possible to use a large injection volume, but the chromatographic conditions should be adjusted in such a way that target analytes will have strong interactions with the stationary phase, and an adsorption compression effect will compensate the loss of efficiency. 

In this case, a compromise between separation efficiency and required analysis time has to be made. Flow rates between 2.0 mL/min and 2.3 mL/min have proved to be most suitable for practical applications. 

In most cases, sample preparation steps take up most of the required analysis time and, thus, contribute substantially to the analysis costs. It should also be mentioned, that each manipulation of samples can falsify the analytical result therefore, the care taken in the sample preparation directly affects the quality of the analytical result. 

The agents for bioluminescent assays are luminous bacteria, recombinant luminous organisms, luciferases and others enzymes for multienzymatic bioluminescent assays. The extremely high amplification of these luminescent systems allows rapid methods to be set up which can be applied to a very small amounts of biological samples. The sensitivity of these methods is often at the nanomolar level, on the border between conventional enzymatic and immunological methods. Moreover they are applicable to analytes present at very low concentration and when high sensitivity is not required, analysis time can be reduced to few seconds. ... 

From a practical viewpoint that disregards costs, the choice of method must be based on three fundamental aspects separation efficiency, accuracy of the analytical method, and total required analysis time. For some applications, the specificity and sensitivity (detectability) of a technique are also important. 

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