Mobile phase optimum

Linear velocity of mobile phase Optimum linear velocity of mobile phase Practical linear velocity of mobile phase Reduced velocity of the mobile phase Specific retention volume... 

For forced flow separations a constant plate height independent of the solvent-front migration distance is obtained. Figure 6.3. The minimum plate height for capillary flow is always greater than the minimum for forced flow. This is an indication that the limited range of capillary flow velocities is inadequate to realize the optimum kinetic performance for the layers. At the mobile phase optimum velocity, forced flow affords more compact zones and shorter separation times compared with capillary flow. As expected the intrinsic efficiency increases with a reduction of the average particle size for the layer. 

Enantiomeric resolution depends also on the concentration and type of organic modifier (methanol or acetonitrile) in mobile phase. Optimum enantiomeric resolution occurs in a narrow range 20-30% of organic modifier and the / f values of enantiomers do not change in this region [28] (Figure 12.6). 

To increase the number of theoretical plates without increasing the length of the column, it is necessary to decrease one or more of the terms in equation 12.27 or equation 12.28. The easiest way to accomplish this is by adjusting the velocity of the mobile phase. At a low mobile-phase velocity, column efficiency is limited by longitudinal diffusion, whereas at higher velocities efficiency is limited by the two mass transfer terms. As shown in Figure 12.15 (which is interpreted in terms of equation 12.28), the optimum mobile-phase velocity corresponds to a minimum in a plot of H as a function of u. 

The analysis of cigarette smoke for 16 different polyaromatic hydrocarbons is described in this experiment. Separations are carried out using a polymeric bonded silica column with a mobile phase of 50% v/v water, 40% v/v acetonitrile, and 10% v/v tetrahydrofuran. A notable feature of this experiment is the evaluation of two means of detection. The ability to improve sensitivity by selecting the optimum excitation and emission wavelengths when using a fluorescence detector is demonstrated. A comparison of fluorescence detection with absorbance detection shows that better detection limits are obtained when using fluorescence. 

The stationary phase is selected to provide the maximum selectivity. Where possible, the retention factor is adjusted (by varying the mobile phase composition, temperature, or pressure) to an optimum value that generally falls between 2 and 10. Resolution is adversely affected when k 2, while product dilution and separation time... 

The temperature of the ineoming mobile phase will also alter the environment within tire eoluirrtr.. Some laboratories have advoeated that equilibration of the ineoming mobile phase with the eolumn temperature is essential for good peak shapes and optimum effieieney. However, a number of studies have now demonstrated that if the mobile phase is eooler than the eolumn the effieieney is improved, in some eases quite markedly. 

The mixture of acetonitrile/water (1 1, v/v) was selected as most effective mobile phase. The optimum conditions for chromatography were the velocity of mobile phase utilization - 0,6 ml/min, the wave length in spectrophotometric detector - 254 nm. The linear dependence of the height of peack in chromathography from the TM concentration was observed in the range of 1-12.0 p.g/mL. 

It is seen that by a simple curve fitting process, the individual contributions to the total variance per unit length can be easily extracted. It is also seen that there is minimum value for the HETP at a particular velocity. Thus, the maximum number of theoretical plates obtainable from a given column (the maximum efficiency) can only be obtained by operating at the optimum mobile phase velocity. 

It is seen from equation (26) that the optimum velocity is determined by the magnitude of the diffusion coefficient and is inversely related to the particle diameter. Unfortunately, in LC (where the mobile phase is a liquid as opposed to a gas), the diffusivity is four to five orders of magnitude less than in GC. Thus, to achieve comparable performance, the particle diameter must also be reduced (c./., 3-5 p)... 

Thus, for significant values of (k") (unity or greater) the optimum mobile phase velocity is controlled primarily by the ratio of the solute diffusivity to the column radius and, secondly, by the thermodynamic properties of the distribution system. However, the minimum value of (H) (and, thus, the maximum column efficiency) is determined primarily by the column radius, secondly by the thermodynamic properties of the distribution system and is independent of solute diffusivity. It follows that for all types of columns, increasing the temperature increases the diffusivity of the solute in both phases and, thus, increases the optimum flow rate and reduces the analysis time. Temperature, however, will only affect (Hmin) insomuch as it affects the magnitude of (k"). 

The conditions required to minimize tube dispersion are clearly indicated by equation (10). Firstly, as the column should be operated at its optimum mobile phase velocity and the flow rate, (0) is defined by column specifications it is not a variable that can be employed to control tube dispersion. Similarly, the diffusivity of the solute (Dm)... 

Equations (4) and (5) predict that the optimum linear velocity should be linearly related to the diffusivity of the solute in the mobile phase, whereas the minimum value of the HETP should be constant and independent of the solute diffusivity. This, of course, will only be true for solutes eluted at the same (k ). It is seen, from Table 1, that (by appropriate adjustment of the concentration of ethyl acetate) the values of both (k ) and (k e) have been kept approximately constant for all the mobile phase... 

The basically correct equation appears to be that of Giddings but, over the range of mobile phase velocities normally employed i.e., velocities in the neighborhood of the optimum velocity), the Van Deem ter equation is the simplest and most appropriate to use. 

The optimum mobile phase velocity will also be determined in the above calculations together with the minimum radius to achieve minimum solvent consumption and maximum mass sensitivity. The column specifications and operating conditions are summarized in Table 4. 

Thus as (y) will always be greater than unity, the resistance to mass transfer term in the mobile phase will be, at a minimum, about forty times greater than that in the stationary phase. Consequently, the contribution from the resistance to mass transfer in the stationary phase to the overall variance per unit length of the column, relative to that in the mobile phase, can be ignored. It is now possible to obtain a new expression for the optimum particle diameter (dp(opt)) by eliminating the resistance to mass transfer function for the liquid phase from equation (14). 

The optimum flow rate is obviously the product of the fraction of the cross-sectional area occupied by the mobile phase and the optimum mobile phase velocity, i.e.,... 

The efficiency obtained from an open tubular column can be increased by reducing the column radius, which, in turn will allow the column length to be decreased and, thus, a shorter analysis time can be realized. However, the smaller diameter column will require more pressure to achieve the optimum velocity and thus the reduction of column diameter can only be continued until the maximum available inlet pressure is needed to achieve the optimum mobile phase velocity. 

The expression for the optimum mobile phase velocity is given by equation (28) in chapter 12 and is as follows. 

The optimum mobile phase velocity for an open tubular GC column is given in chapter 13, equation (14). Reiterating this equation,... 

The use of both sub- and supercritical fluids as eluents yields mobile phases with increased diffusivity and decreased viscosity relative to liquid eluents [23]. These properties enhance chromatographic efficiency and improve resolution. Higher efficiency in SFC shifts the optimum flowrate to higher values so that analysis time can be reduced without compromising resolution [12]. The low viscosity of the eluent also reduces the pressure-drop across the chromatographic column and facilitates the... 

If the mixture to be separated contains fairly polar materials, the silica may need to be deactivated by a more polar solvent such as ethyl acetate, propanol or even methanol. As already discussed, polar solutes are avidly adsorbed by silica gel and thus the optimum concentration is likely to be low, e.g. l-4%v/v and consequently, a little difficult to control in a reproducible manner. Ethyl acetate is the most useful moderator as it is significantly less polar than propanol or methanol and thus, more controllable, but unfortunately adsorbs in the UV range and can only be used in the mobile phase at concentrations up to about 5%v/v. Above this concentration the mobile phase may be opaque to the detector and thus, the solutes will not be discernible against the background adsorption of the mobile phase. If a detector such as the refractive index detector is employed then there is no restriction on the concentration of the moderator. Propanol and methanol are transparent in the UV so their presence does not effect the performance of a UV detector. However, their polarity is much greater than that of ethyl acetate and thus, the adjustment of the optimum moderator concentration is more difficult and not easy to reproduce accurately. For more polar mixtures it is better to explore the possibility of a reverse phase (which will be discussed shortly) than attempt to utilize silica gel out of the range of solutes for which it is appropriate. 

The newcomer to chromatography, faced with a hitherto unknown sample, would do well to start with a C8 silica based reverse phase and an acetonitrile water mixture as a mobile phase and carry out a gradient elution from 100% water to 100% acetonitrile. From the results, the nature and the complexity of the sample can be evaluated and a more optimum phase system can be inferred. 

The curve exhibits a minimum, which means that there is an optimum mobile phase velocity at which the column will give the minimum HETP and consequently a maximum efficiency. In practice this usually means that reducing the flow rate of a column will increase the efficiency and thus the resolution. In doing so, however, the analysis time will also be increased. As seen in figure 5, however, there is a limit to this procedure, as reducing the column flow rate so that the mobile phase velocity falls below the optimum will result in an increase in the HETP and thus a decrease in column efficiency. 

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