Extra-column pressure

Table 3.4 shows the contribution of extra-column volume to the observed pressure drop for a variety of column sizes operating at the same linear velocity with the same connecting tubing. The smaller i.d. columns require a lower flow rate, which results in a lower extra-column pressure arising from flow through the connecting tubing. Therefore, the relative contribution of the pressure from the extra-column volume sources can be seen to diminish as the column internal diameter decreases, with 31 % of the observed pressure attributed to extra-column somces with the 4.6 mm i.d. column, and only 10% of the pressure attributed to extra-column sources for the 1 mm i.d. column. 

Correlation was found between domain size and attainable column efficiency. Column efficiency increases with the decrease in domain size, just like the efficiency of a particle-packed column is determined by particle size. Chromolith columns having ca. 2 pm through-pores and ca. 1pm skeletons show H= 10 (N= 10,000 for 10 cm column) at around optimum linear velocity of 1 mm/s, whereas a 15-cm column packed with 5 pm particles commonly shows 10,GOO-15,000 theoretical plates (7 = 10—15) (Ikegami et al., 2004). The pressure drop of a Chromolith column is typically half of the column packed with 5 pm particles. The performance of a Chromolith column was described to be similar to 7-15 pm particles in terms of pressure drop and to 3.5 1 pm particles in terms of column efficiency (Leinweber and Tallarek, 2003 Miyabe et al., 2003). Figure 7.4 shows the pressure drop and column efficiency of monolithic silica columns. A short column produces 500 (1cm column) to 2500 plates (5 cm) at high linear velocity of 10 mm/s. Small columns, especially capillary type, are sensitive to extra-column band... 

To fully utilize small particles (i.e., less than 2 pm) packed in columns greater than 50 mm in length, an HPLC system must be developed that can operate at high pressures. For the best results using columns with 1-2-pm particles, extra-column effects, dwell volumes, and detectors must be optimized for optimum performance. Additionally, commercial... 

There are a number of limitations on the use of extremes of temperature in HPLC. Clicq et al. [91] note that instrumental issues become increasingly limiting as one goes to very high temperatures and flow rates. They suggest that most separations will occur below 90°C where there are less instrumental constraints. As detailed below, column bleed can limit the selection of columns. Highspeed separations require a faster detector response than many systems allow and constrain extra column volume. This is especially true for narrow bore columns and sub-2 jam particles. In many cases, the additional speed gained above the temperature limits of commercial HPLC ovens will not be worth the additional expense and complexity required. For macromolecules, the effect of extreme pressure can also impact retention time as noted by Szabelski et al. [92]. 

The maximum operating pressure 2/ The extra column dispersion 3/ The minimum flow Rate... 

Maximum Column Inlet Pressure Extra Column Dispersion Multipath Packing Factor Longitudinal Diffusion Packing Factor Column Mobile Phase Fraction... 

It is seen that the optimum column radius for an open tubular column varies widely with inlet pressure arid the difficulty of the separation. Considering a separation of some difficulty, for example ( a = 1.02), it is seen that at an inlet pressure of lOOOp.s.f, the optimum column diameter would be about 4 micron whereas, at an inlet pressure of only 1 psi, it would be about 43 micron. The former would be quite difficult to coat with stationary phase and would demand detectors and injection systems of almost impossibly low dispersion A column of 43 micron in diameter, on the other hand, would be piactical from the point of view of both ease of coating and an acceptable system extra column dispersion. However, the lengths of such columns arid the resulting analysis times remains to be determined and may preclude their- use. 

The properties of open tubular columns shown in figures (I) to (6) indicate that the areas where such columns would have practical use is very restricted. At pressures in excess of 10 ps.i., and whatever the nature of the separation, whether simple or difficult, the optimum column diameters are so small that they would be exceedingly difficult to fabricate or coat with stationary phase. The maximum sample volumes and extra column dispersion that couid be tolerated would also be well below that physically possible at this time. At relatively low pressures, that Is at pressures less than 10 p.s.l. the diameter of the optimum column is large enough to fabricate and coat with stationary phase providing the separations required are difficult i.c. the separation ratio of the critical pair must be less than 1.03. However, even under these conditions the sample volume will be extremely small, the extra column dispersion restricted to an almost impossibly low limit and the analysis time would be very long Nevertheless, open tubular columns used for very difficult separations... 

M mass of solute to be separated N number of effective theoretical plates P pressure Q flow rate R resolution S peak capacity Sm specific heat of mobile phase Ss specific heat of adsorbent Sg specific heat of detector cell walls V volume in conventional units Vo system dead volume Vr retention volume V r corrected retention volume Vm volume of mobile phase in the column Vs volume of stationary phase in the column Ve extra column volume... 

It is clear from table 7.2 that in terms of extra-column dispersion a wide-bore capillary column requires instrumentation similar to that used for the packed column. However, the capillary column provides eight times as many plates (in a fifteen-fold analysis time). Conventional capillary columns require a reduction in the dispersion by about an order of magnitude, whereas narrow-bore columns require a further reduction by a factor of about 100. This, combined with the high pressures required, puts narrow-bore columns out of reach for current GC instruments. 

An ideal interface should not cause extra-column peak broadening. Historical interfaces include the moving belt and the thermospray. Common interfaces are electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCl). Several special interfaces include the particle beam—a pioneering technique that is still used because it is the only one that can provide electron ionization mass spectra. Others are continuous fiow fast atom bombardment (CF-FAB), atmospheric pressure photon ionization (APPI), and matrix-assisted laser desorption ionization (M ALDl). The two most common interfaces, ESI and APCI, were discovered in the late 1980s and involve an atmospheric pressure ionization (API) step. Both are soft ionization techniques that cause little or no fragmentation hence a fingerprint for qualitative identification is usually not apparent. 

LPRlNT Maxlmum Column Inlet Pressure LPRINT"Extra Column Dispersion LPRINT Multipath Packing Factor LPRINT longitudinal Diffusion Packing Factor LPRINT"Column Mobile Phase Fraction LPRINT... 

Use preparative materials for the adsorbent and, if possible, semi-preparative columns. Always check consistency of the results with those of the initial tests (pressure drop, fe ). Use one plant for all measurements and try to keep extra column effects low. 

During scale-up of the separation from laboratory to pilot to industrial scale, the pressure profile remains similar if the linear velocity, the column length, the particle size, the void fraction, the gradient characteristics, the load volume of feed, the load concentration, pressure drop in the extra-column tubings, the temperature of the feed and that of the mobile phase are all held constant. In practice, the pressure profile remains similar on scale-up when these details are well matched in the laboratory and in the plant. Often, however, the challenge is that, for operational reasons, these parameters cannot be maintained constant during the scale-up. Thus, additional experiments may be required in the laboratory to match the conditions in the pilot plant or in the plant. 

A pressure-optimized column has L = 2.2 cm, dp = 6.9 )Xm and A/ = 2.3 bar Shorter columns are preferred for simple problems with a separation factor of ca. 1.2. Both time and solvent are saved by using columns 3-5 cm in length (also available commercially), but the injection and extra-column volumes and the detector time constant must be kept small in order not to deteriorate the separation performance. 

V = (g p X u)ID. For a constant v it is therefore necessary to work with a constant product 6 p X u. Thereby the pressure increases markedly. In Figure 7.3 the particle diameter was decreased from 10 jam to 3 jam. For an identical number of theoretical plates it was possible to reduce the column length to 30% of the original value (note that all the extra-column volumes of the HPLC instrument need to be adapted ). Despite the shorter column the pressure increases by a factor of ten when the condition... 

The eluate may be passed alternately through the two columns up to 20 times. The advantage of this system is that extra-column volumes are smaller than with the first method and the pump volume can be chosen at random. However, the detector cell must be pressure-tight. 

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