Chromatographic system capacity

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. 

It is also apparent from Figure 20 that any property of the chromatographic system that places a limit on the maximum value of (k ) must also limit the maximum peak capacity that is attainable. One property of the system that limits the maximum value... 

The explicit form of those equations that satisfy the preliminary data criteria, must then be tested against a series of data sets that have been obtained from different chromatographic systems. As an example, such systems might involve columns packed with different size particles, employed mobile phases or solutes having different but known physical properties such as diffusivity or capacity ratios (k"). 

The peak capacity, n, of a single-column chromatographic system generating N theoretical plates is given by ... 

The limitations of one-dimensional (ID) chromatography in the analysis of complex mixtures are even more evident if a statistical method of overlap (SMO) is applied. The work of Davis and Giddings (26), and of Guiochon and co-workers (27), recently summarized by Jorgenson and co-workers (28) and Bertsch (29), showed how peak capacity is only the maximum number of mixture constituents which a chromatographic system may resolve. Because the peaks will be randomly rather than evenly distributed, it is inevitable that some will overlap. In fact, an SMO approach can be used to show how the number of resolved simple peaks (5) is related to n and the actual number of components in the mixture (m) by the following ... 

Fractionation of components into polarity groups, and their optimized separation (followed by detection) by subsequent development steps increases the separating capacity of the chromatographic system. 

It is appropriate to refer here to the development of non-suppressed ion chromatography. A simple chromatographic system for anions which uses a conductivity detector but requires no suppressor column has been described by Fritz and co-workers.28 The anions are separated on a column of macroporous anion exchange resin which has a very low capacity, so that only a very dilute solution (ca 10 4M) of an aromatic organic acid salt (e.g. sodium phthalate) is required as the eluant. The low conductance of the eluant eliminates the need for a suppressor column and the separated anions can be detected by electrical conductance. In general, however, non-suppressed ion chromatography is an order of magnitude less sensitive than the suppressed mode. 

A general approach to the problem of identification, should more definitive detectors not be available, is to change the chromatographic system , which in the case of HPLC is usually the mobile phase, and redetermine the retention parameter. The change obtained is often more characteristic of a single analyte than is the capacity factor with either of the mobile phases. 

As was reported by Soczewinski, a three-component mixture containing 5 mg of each of the ingredients can be completely separated using a 90 X 100 X 0.5 mm layer of silica [26]. This indicates that the capacity of the chromatographic system can be considerably increased by the apphcation of wider and thicker layers of adsorbent on the chromatoplate [44]. 

Many chromatographic systems show linear relationships between the logarithm of the capacity factor and the reciprocal of the column temperature (van t Hoff plots) [255,258-261]. In thermodynamic terms the interaction of the solute with the stationary phase can be described by... 

The potential of a chromatographic system to provide a certain separation can be estimated from its separation number, S SH, also referred to as the spot capacity in TLC. The separation j nund>er in TLC is defined as the number of spots that can be. completely separated (R, > 1) between R 0 and R, ° 1 (6). It is calculated in an approximate form by equation (7.16) and morej exactly by equation (7.17) with b, and b, as defined in Figure 7.4. 

Standards and blanks are the usual controls used in analytical HPLC. Standards are usually interspersed with samples to demonstrate system performance over the course of a batch run. The successful run of standards before beginning analysis demonstrates that the system is suitable to use. In this way, no samples are run until the system is working well. Typically, standards are used to calculate column plate heights, capacity factors, and relative response factors. If day-to-day variability has been established by validation, the chromatographic system can be demonstrated to be within established control limits. One characteristic of good science is that samples... 

While chromatographic peak capacity is not adequate to resolve hundreds of thousands of components, many researchers argue that MS itself is an additional separation dimension with an orthogonal selectivity (separation is based on mass-to-charge ratio). Therefore, the combined resolution of LC and MS is greater than the chromatographically defined peak capacity. The question therefore stands What is the achievable peak capacity of the 2DLC-MS/MS system ... 

As with most aspects of downstream processing, the operation of chromatographic systems is highly automated and is usually computer controlled. Whereas medium-sized process-scale chromatographic columns (e.g. 5-151 capacity) are manufactured from toughened glass or plastic, larger... 

The retention time tj is a non-linear function of the ring size n and thus allows the identification of new species of the homologous series S by interpolation. Furthermore, the logarithm of the capacity factor k = (t — (with = death time of the chromatographic system) is a linear function of the ring size, n, although two such functions are obtained see Fig. 7... 

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