Chromatograms Chromatograph

Column Efficiency. Under ideal conditions the profile of a solute band resembles that given by a Gaussian distribution curve (Fig. 11.1). The efficiency of a chromatographic system is expressed by the effective plate number defined from the chromatogram of a single band. 

A graph or chart of ion current (y-axis) vs. time (x-axis) is therefore a succession of peaks corresponding to components eluting from the chromatographic column. This chart is called a total km current (TIC) chromatogram. 

Development of the Chromatogram. The term development describes the process of performing a chromatographic separation. There are several ways in which separation may be made to occur, eg, frontal, displacement, and elution chromatography. Frontal chromatography uses a large quantity of sample and is usually unsuited to analytical procedures. In displacement and elution chromatography, much smaller amounts of material are used. 

Separated from retinol by column chromatography on water-deactivated alumina with hexane containing a very small percentage of acetone. Also chromatographed on TLC silica gel G, using pet ether/isopropyl ether/acetic acid/water (180 20 2 5) or pet ether/acetonitrile/acetic acid/water (190 10 1 15) to develop the chromatogram. Then recrystd from propylene at low temperature. 

In this chapter, the elution curve equation and the plate theory will be used to explain some specific features of a chromatogram, certain chromatographic operating procedures, and some specific column properties. Some of the subjects treated will be second-order effects and, therefore, the mathematics will be more complex and some of the physical systems more involved. Firstly, it will be necessary to express certain mathematical concepts, such as the elution curve equation, in an alternative form. For example, the Poisson equation for the elution curve will be put into the simpler Gaussian or Error function form. 

The curves show that the peak capacity increases with the column efficiency, which is much as one would expect, however the major factor that influences peak capacity is clearly the capacity ratio of the last eluted peak. It follows that any aspect of the chromatographic system that might limit the value of (k ) for the last peak will also limit the peak capacity. Davis and Giddings [15] have pointed out that the theoretical peak capacity is an exaggerated value of the true peak capacity. They claim that the individual (k ) values for each solute in a realistic multi-component mixture will have a statistically irregular distribution. As they very adroitly point out, the solutes in a real sample do not array themselves conveniently along the chromatogram four standard deviations apart to provide the maximum peak capacity. 

A satisfactory chromatographic analysis demands, a priori, on an adequate separation of the constituents of the sample that will permit the accurate quantitative evaluation of each component of interest. To achieve this, an appropriate phase system must be chosen so that the individual components of the mixture will be moved apart from one another in the column. In addition, their dispersion must be constrained sufficiently to allow all the solutes of interest to be eluted discretely. At this stage it is necessary to introduce the concept of the Reduced Chromatogram. 

Every chromatographic investigation begins with the preparation of the sample and the chromatographic system. This is followed by the crux of the separation process (development of the chromatogram) which is in turn followed by the visualization of the separated substances and the preservation of the chromatogram and finally by the analysis of the results. 

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