Separation speed

Remarks Moderate selectivity Moderate separation speed Narrow mobile-phase selection Fow selectivity High-speed separation Most generous mobile-phase selection Fow selectivity High-speed separation Generous mobile-phase selection Best for high MW polymers Thermal gradient may be programmed for broad MW separation... 

While PLB were introduced first (14,15) more recently small PB 5-15 M diameter) have become of major interest. This is a result of the higher separation speeds found with such particles. Not only is the "stagnant" mobile phase mass transfer problem reduced, as in PLB, but solute mixing in the flowing stream is enhanced as a result of the smaller distance between the particles. The performances achieved with the small particle columns are equivalent to those obtained with capillary columns in gas chromatography (13), Examples illustrating the separation speed of such columns will be presented in the applications section of this paper. 

Upon substitution of the reduced parameters given above the separation time for a packed column and an open tubular column would be Identical if d 1.73 dp given the current limitations of open tubular column technology the column diameter cannot be reduced to the point %diere these columns can compete with packed columns for fast separations. This is illustrated by the practical txanple in Figure 6.3 (57). Ihe separation speed cannot be Increased for an open tubular column by increasing the reduced velocity since the reduced plate height is increased... 

A typical HPLC separation using a 15-cm column of 15,000 theoretical plates produces peak capacity (Giddings, 1991) of about 80-100 under isocratic conditions and up to 150 under gradient conditions in 1 h(Eq. 7.3, n peak capacity, A number of theoretical plates of a column, and fR and t retention time of the last and the first peak of the chromatogram, respectively). An increase in the number of separated peaks per unit time can be achieved by increased separation speed made possible by monolithic silica columns (Deng et al., 2002 Volmer et al., 2002). This has also been shown for peptides and proteins (Minakuchi et al., 1998 Leinweber et al., 2003). 

Capillary electrophoresis has also been combined with other analytical methods like mass spectrometry, NMR, Raman, and infrared spectroscopy in order to combine the separation speed, high resolving power and minimum sample consumption of capillary electrophoresis with the selectivity and structural information provided by the other techniques [6]. 

FIGURE 4.12 Comparison of separation speed between packed column SFC and OTGC. Conditions are the same as in Figure 4.11. (Adapted from Wu, N. et al. 2000. J. Microcol. Sep. 12 454-461. With permission.)... 

Equation 9.5 shows that for an acceptable retention factor, the diffusion coefficient and particle size of the packing are the primary variables that can be varied to affect the N/tf An increase in the diffusion coefficient or a decrease in the particle size causes an increase in N/tf If the diffusion coefficient can be increased without impacting the retention factor significantly, then significant increases in separation speed will result. To minimally impact the retention factor the solvent strength of the mobile phase must be maintained. 

Based on the high peak capacity of CE, the separation speed, and the availability of numerous chiral selectors and the simplicity of the systems, chiral CE is superior to chiral HPLC separations. This is as well reflected by the high number of publications on chiral CE in recent years. Chiral HPLC is suffering from low peak capacity (broad peaks), system stability (often normal phase systems), pressure sensitivity of columns (often cellulose-based column materials), and as a consequence long separation times. 

Efficiencies of 24,000 plates at permeation and 23,000 plates at total exclusion were measured at flow rates of 1.0 ml/minute for this column. Separation speed is quite good also, but does not represent the limit which can be attained. 

Even if relatively new, HF FIFFF has been used to separate supramicrometer particles, proteins, water-soluble polymers, and synthetic organic-soluble polymers. Particle separation in HF FIFFF has recently been improved, reaching the level of efficiency normally achieved by conventional, rectangular FIFFF channels. With these channel-optimized HF FIFFF systems, separation speed and the resolution of nanosized particles have been increased. HF FIFFF has recently been examined as a means for off-line and on-line protein characterization by using the mass spectrometry (MS) through matrix-assisted laser desorption ionization time-of-flight mass spectrometry (M ALDl-TOF MS) and electrospray ionization (ESl)-TOF MS, as specific detectors. On-line HF FIFFF and ESl-TOF MS analysis has demonstrated the viability of fractionating proteins by HF FIFFF followed by direct analysis of the protein ions in MS [38]. 

To obtain high EOF values in HP-CEC with RPC sorbents, such as C8- or C18-bonded silica, and thus to obtain enhanced selectivity and increased separation speed with peptides, it is necessary to use non-endcapped sorbents. 130,285 With n-alkylsilicas, a high... 

The mass spectrometer detectors place new demands on the HPLC system. The MS interface requires use of volatile buffers and reagents. Nanospray interfaces especially benefit from low-volume, high-resolution separations. The mass spectrometer is a fast response system and benefits from separation speeds higher than normally supplied by HPLC systems. All of these requirements have provided constraints on new development directions for HPLC systems. 

Separation speed and ease of use seem to be the primary factors driving changes in HPLC instrumentation. Resolution efficiency and stationary phase stability, especially at high pH, are the primary factors affecting current changes in column technology. 

The major reason why a reduction of the particle size or column diameter is expected to lead to an increase of separation speed (resolution power per time unit) can be found in its effect to decrease r. Separation speed is often expressed in the analytical literature in terms of the number of theoretical plates N per time unit f (for a definition of N in terms of experimental parameters see Sect. 3.1.1). For zone dispersion due to lateral non-equilibrium, the ratio N/t will be in general inversely proportional to r [20] ... 

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