Viscosity of the mobile-phase

Capacity Ratio (first eluted peak of the Critical Pair) (k ) Capacity Ratio (first eluted peak of the Critical Pair) (k") Viscosity of the Mobile Phase (r])... 

The most important part of this type of interface, from a number of points of view, is the pinhole which, in conjunction with the pumping capacity of the mass spectrometer, controls the flow of eluate into the mass spectrometer. This flow, and therefore the properties of the spray being introduced into the mass spectrometer, is affected by a change in the viscosity of the mobile phase. The use of gradient elution has therefore to be approached with some caution as the sensitivity of the mass spectrometer can change significantly during the course of an analysis. 

Tenperature Efficiency Elevated teaperatures increase the rate of solute exchange between the stationary and aobile phases and also lower the viscosity of the mobile phase. 

It is apparent from early observations [93] that there are at least two different effects exerted by temperature on chromatographic separations. One effect is the influence on the viscosity and on the diffusion coefficient of the solute raising the temperature reduces the viscosity of the mobile phase and also increases the diffusion coefficient of the solute in both the mobile and the stationary phase. This is largely a kinetic effect, which improves the mobile phase mass transfer, and thus the chromatographic efficiency (N). The other completely different temperature effect is the influence on the selectivity factor (a), which usually decreases, as the temperature is increased (thermodynamic effect). This occurs because the partition coefficients and therefore, the Gibbs free energy difference (AG°) of the transfer of the analyte between the stationary and the mobile phase vary with temperature. 

A method that can decrease the viscosity of the mobile phase without impacting the mobile phase solvent strength (i.e., maintaining k) would therefore decrease the analysis time linearly. The next section illustrates the diffusion coefficients and viscosities, the unique relationship between them for EEL mixtures, their solvent-strength and other important properties. 

Higher colnmn temperatnres (T) lower the viscosity of the mobile phase (and thns the pressnre) and increase the diffusion coefficient of the analytes (increase N). The effects of these operating parameters are dis-cnssed farther in the next section on gradient analysis. 

The stability of zirconia and zirconia-based packings also allows the use of these columns at an elevated temperature. The key advantage of this is the reduction in analysis time due to the reduction in the viscosity of the mobile phase. In addition, the much broader temperature range also permits a fine-tuning of the selectivity of a separation compared with silica-based packings. 

From both theory and experimental evidence, raising the temperature by 10°C decreases the retention time by about 20% in isocratic chromatography and decreases the backpressure by 10% to 20% because of a reduction in the viscosity of the mobile phase. This can help to overcome the instrument limitations associated with running shorter columns packed with smaller particles, i.e., the pressure limitations of current HPLC systems. However, since the majority of reversed-phase columns available are silica-based, operating at temperatures above... 

Gradient separations can be accelerated at elevated temperature by reducing the viscosity of the mobile phase and hence the column operating pressure and by increasing the diffusion coefficients of the analytes, speeding up the mass transfer between the stationary and the mobile phase [156,157],... 

The situation described in Equation 9.1 is reversed at a reduced temperature. The overall column efficiency decreases rather dramatically for most samples, but successful separations are still practical with the correct choice of parameters. The reduced longitudinal diffusion in the first term means that the optimal flow rate shifts to lower flow rates. The increased viscosity of the mobile phase requires lower flow rates as well. While at high temperatures one often operates the HPLC at flow rates many times the optimal value, in subambient work, it is best to sacrifice speed and work close to the optimal flow rate. 

It has already been stated that the retention of a solute depends on the magnitude of the distribution coefficient of the solute between the mobile and stationary phases. Furthermore, according to Vant Hoff s Law, the distribution coefficient will vary according to the exponent of the reciprocal of the absolute temperature. In addition, the dispersion of a solute band in a column will be shown to depend on the dlffusivity of the solute In both phases, the viscosity of the mobile phase and also on the distribution coefficient of the solute, all of which vary with temperature. It follows that, for consistent results, the column must be carefully thermostated. The column and its contents have a significant heat capacity and, consequently, it is of little use trytng to thermostat the column in an air bath for satisfactory temperature control, the thermostating medium... 

Whether the optimum phase system is arrived at by a computer system, or by trial and error experiments (which are often carried out, even after computer optimization), the basic chromatographic data needed in column design will be identified. The phase system will define the separation ratio of the critical pair, the capacity ratio of the first eluted peak of the critical pair and the capacity ratio of the last eluted peak. It will also define the viscosity of the mobile phase and the diffusivity of the solute in the mobile phase. 

There remains little more for the operator to decide. Sometimes, alternative but similar solvent mixtures that have a lower viscosity or higher solute diffusivity could be selected. For example, a n-hexane/methanol mixture might be chosen as an alternative to the more viscous n-heptane/isopropyl alcohol mixture as it has similar elution properties. However, it will be shown later, that if a fully optimized column is employed the viscosity of the mobile phase does not seem to effect the column performance as it is taken into account in the optimization procedure. The operator would, under some circumstances, be free to choose less toxic or less costly solvents for example, in reverse phase chromatography the operator could select methanol/ water solvent mixtures as opposed to acetonitrile/water mixture on the basis of lower cost or less... 

It is interesting to note from equation (21) that the analysis time does not depend on the magnitude of the diffusivity of the solute in the mobile phase but only on the viscosity of the mobile phase. It does, however, increase... 

Equation (13) gives the minimum analysis time that can be obtained from an open tubular column, when separating a mixture of defined difficulty, under given chromatographic conditions. It is seen that, in a similar manner to the packed column, the analysis time is inversely proportional to the fourth power of the function (a-1) and inversely proportional to the inlet pressure. The contribution of the function of (k1), to the analysis time is not clear and can be best seen by calculation. It is also seen (perhaps a little surprisingly) that the analysis time is completely independent of the diffusivity of the solute in the mobile phase but is directly proportional to the viscosity of the mobile phase. 

The expression for the maximum permissible sample volume, given by equation (20), shows a strong dependence on the product of the solute diffusivity and the viscosity of the mobile phase It is also seen to vary as the inverse of the fourth power of foe-10 so that, for very difficult separations, where (a) is small, the sample volume will be a maximum... 

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