Stationary phases peak capacity

A 0.53-mm-i.d. column offers the best of both worlds, because it combines the attributes of a fused-silica capillary column with the high sample capacity and ease of use of a packed column. For many applications, analytical methods developed with a packed column can easily be transferred to a megabore column with the appropriate stationary phase. Peaks generated with a megabore column typically are sharper and exhibit less tailing compared to those with a packed column. Redistribution of the stationary phase can occur at the inlet of a packed column with large injections of solvent and leave an exposure of silanol sites on a diatomaceous earth support. With a cross-linked phase in a megabore... 

To minimize the multiple path and mass transfer contributions to plate height (equations 12.23 and 12.26), the packing material should be of as small a diameter as is practical and loaded with a thin film of stationary phase (equation 12.25). Compared with capillary columns, which are discussed in the next section, packed columns can handle larger amounts of sample. Samples of 0.1-10 )J,L are routinely analyzed with a packed column. Column efficiencies are typically several hundred to 2000 plates/m, providing columns with 3000-10,000 theoretical plates. Assuming Wiax/Wiin is approximately 50, a packed column with 10,000 theoretical plates has a peak capacity (equation 12.18) of... 

In common with all multidimensional separations, two-dimensional GC has a requirement that target analytes are subjected to two or more mutually independent separation steps and that the components remain separated until completion of the overall procedure. Essentially, the effluent from a primary column is reanalysed by a second column of differing stationary phase selectivity. Since often enhancing the peak capacity of the analytical system is the main goal of the coupling, it is the relationship between the peak capacities of the individual dimensions that is crucial. Giddings (2) outlined the concepts of peak capacity product and it is this function that results in such powerful two-dimensional GC separations. 

A theoretical model whereby maximum peak capacity could be achieved by the use of 3-D planar chromatographic separation was proposed by Guiochon and coworkers (23-27). Unfortunately, until now, because of technical problems, this idea could not be realized in practice. Very recently, however, a special stationary phase, namely Empore silica TLC sheets, has now become available for realization of 3-D PC. This stationary phase, developed as a new separation medium for planar chromatography, contains silica entrapped in an inert matrix of polytetrafluoroethy-lene (PTFE) microfibrils. It has been established that the separating power is only ca. 60% of that of conventional TLC (28) this has been attributed to the very slow solvent migration velocity resulting from capillary action. 

The peak capacity is not pertinent as the separation was developed by a solvent program. The expected efficiency of the column when operated at the optimum velocity would be about 5,500 theoretical plates. This is not a particularly high efficiency and so the separation depended heavily on the phases selected and the gradient employed. The separation was achieved by a complex mixture of ionic and dispersive interactions between the solutes and the stationary phase and ionic, polar and dispersive forces between the solutes and the mobile phase. The initial solvent was a 1% acetic acid and 1 mM tetrabutyl ammonium phosphate buffered to a pH of 2.8. Initially the tetrabutyl ammonium salt would be adsorbed strongly on the reverse phase and thus acted as an adsorbed ion exchanger. During the program, acetonitrile was added to the solvent and initially this increased the dispersive interactions between the solute and the mobile phase. 

Trying to determine which column is ideal for a specific analysis can be difficult with over 1000 different columns on the market [74]. A proper choice implies a definition of parameters such as column material, stationary phase (polarity), i.d., film thickness and column length. Guides to column selection are available [74,75]. The most important consideration is the stationary phase. When selecting an i.d., sample concentration and instrumentation must be considered. If the concentration of the sample exceeds the column s capacity, then loss of resolution, poor reproducibility and peak distortion will result. Film thickness has a direct effect on retention and the elution temperature for each sample compound. Longer columns provide more resolving probe, increase analysis times and cost. 

As yet, the number of applications is limited but is likely to grow as instrumentation, mostly based on existing CE systems, and columns are improved and the theory of CEC develops. Current examples include mixtures of polyaromatic hydrocarbons, peptides, proteins, DNA fragments, pharmaceuticals and dyes. Chiral separations are possible using chiral stationary phases or by the addition of cyclodextrins to the buffer (p. 179). In theory, the very high efficiencies attainable in CEC mean high peak capacities and therefore the possibility of separating complex mixtures of hundreds of... 

Modem technologies provide many techniques for expanding the throughput of an analytical laboratory. The task that needs to be accomplished and the possible drawbacks should be carefully considered. Optimized LC equipment can utilize columns packed with much smaller stationary phase particles to achieve significant reductions in gradient time while still achieving the same or even better peak capacities than conventional methods. 

Stationary phase, mobile phase, and column length should yield sufficient sample and peak capacity. 

The series of regioisomeric amines 48-50, methamphetamine (29) and phentermine (31), can be identified in forensic screening analyses by RP-HPLC-UVD (254/280 nm dual accessory) using a Cis stationary phase and a mobile phase buffered at pH 3.0. The capacity factors and retention times increase in the order 48 < 49 < 29 < 31 < 50. Other methods for identifying these compounds failed for example, the base peak in MS is m/z = 58 for all five compounds, corresponding to a loss of a benzyl group from the molecular peak also their IR and UVV spectra are too similar to be useful for this... 

In the development and optimization of a comprehensive LCxLC method, many parameters have to be taken in acconnt in order to accomplish snccessfnl separations. First of all, selectivity of the columns used in the two dimensions must be different to get maximum gain in peak capacity of the 2D system. For the experimental setup, column dimensions and stationary phases, particle sizes, mobile-phase compositions, flow rates, and second-dimension injection volumes should be carefully selected. The main challenges are related to the efficient coupling of columns and the preservation of mobile phase/column compatibility. 

Comprehensive 2D liquid chromatography is emerging as a new powerfnl technique for the separation of complex samples because of increased peak capacity, selectivity, and resolution in comparison to single-dimensional HPLC. 2D LC x LC systems essentially represent programming of stationary phases. Comprehensive LC x LC techniqne represents specific 2D mode, where all sample componnds eluting from the first dimension are snbjected to separation in the second dimension [167]. The whole effluent from the first dimension is transferred into the second-dimension... 

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