Plate number enhancement

Owing to the EK bias in favor of faster migrating species in stack injection, the signal enhancement ranged from 31 to 8. However, stack injection produced less resolution, i.e., fewer plate numbers (N), as compared to those obtained in pinched injection. In addition, RSD (n = 6) in peak areas for stack, non-stacked, and pinched injection are 2.1%, 1.4%, and 0.75%, respectively [346]. 

When packed into chromatography columns, TRIM beads imprinted with Boc-L-Phe were shown to have column efficiencies and separation abilities superior to ground and sieved bulk material [5]. The theoretical plate number was approximately double that obtained with conventional crushed polymer under the same conditions and the resolution of a racemate was also slightly enhanced. The difference, however, was not that great considering the additional preparation time and effort involved. 

The separation characteristics of a considerable variety of other TLC supports were also tested using different dye mixtures (magnesia, polyamide, silylated silica, octadecyl-bonded silica, carboxymethyl cellulose, zeolite, etc.) however, these supports have not been frequently applied in practical TLC of this class of compounds. Optimization procedures such as the prisma and the simplex methods have also found application in the TLC analysis of synthetic dyes. It was established that six red synthetic dyes (C.I. 15580 C.I. 15585 C.I. 15630 C.I. 15800 C.I. 15880 C.I. 15865) can be fully separated on silica high-performance TLC (HPTLC) layers in a three-solvent system calculated by the optimization models. The theoretical plate number and the consequent separation capacity of traditional TLC can be considerably enhanced by using supports of lower particle size (about 5 fim) and a narrower particle size distribution. The application of these HPTLC layers for the analysis of basic and cationic synthetic dyes has also been reviewed. The advantages of overpressured (or forced flow) TLC include improved separation efficiency, lower detection limit, and lower solvent consumption, and they have also been exploited in the analysis of synthetic dyes. 

Anything that increases the column efficiency N, the column selectivity a. or the retention factor k will enhance the separation power of the column. Packed columns are characterized by low plate numbers and PCGC is therefore a low-resolution technique. The lower efficiency is compensated by the high. selectivity a of the stationary phase, and this is the main reason why so many different stationary pha.ses have been developed for PCGC. Capillary columns on the other hand have very high plate numbers and, therefore, the number of stationary phases can be restricted because the selectivity is less important. In fact, most separation problems can be handled with four basic. stationary pha.ses and half a dozen tailor-made stationary phases. Other important features of capillary columns are their inertness and compatibility with spectroscopic detectors. In the framework of this discussion, emphasis is, therefore, on capillary columns. 

From this relationship it follows that thin-layer efficiency (plate number N) and composition of mobile phases (monitored through K2lK and Rj) can be optimized separately. Enhancement of thin-layer performance in terms of raising N will be the subject of Section VI, while the approaches aiming to optimize the composition of mobile phases will be discussed below. 

Enhancement of thin-layer performance is basically understood as elevation of the theoretical plate number N for a given type of chromatographic plate. The quantity N was defined in Section III.A... 

One would obtain a quite small improvement in the resolution as a result of a reasonable extension of the separahon time (k value of 6). Therefore, the enhancement of the interaction (e.g., the increase of the water content of the eluent) is a common, easily implemented, but mostly quite inefficient procedure for routinely improving resolution. If, however, the plate number could be improved to 15,000, for example (smaller particles, injection tricks, etc.), in the case of a k value of 4, then the resolution would improve to 1.17. 

This step encompasses procedures that are usually performed within the scope of an optimization. In doing so, mostly the interactions between the sample and the stationary phase are changed. The intention is a change of the retention factor k (mostly enhancement), but ideally also of the separation factor Ct. Otherwise, at constant interaction strength ( chemistry constant and therefore k and a as well), one attempts to enhance the plate number or in the case of a miniaturization to prevent dilution or to enhance the relative mass sensitivity. Using a trial-and-error procedure, one needs 1-2 weeks. Using a systematic procedure, aided perhaps by an optimization program, the time can be reduced measurably see Part 4 for chapters on computer-aided optimization. 

---