Raymond P. W. Scott

Distribution of a solute between two phases: the basic theory and its application to the prediction of chromatographic retention

Abstract A simple theory that explains the distribution of a solute between two phases is put forward and experimentally validated employing both previously published data and results from unambiguous liquid—liquid distribution experiments. The theory is tehn extended to liquid—liquidand liquid—solid distribution systems where association takes place between the components of the liquid phases. It is shown that under such circumstances a binary mixture becomes. in fact, a ternary system where the third component comprises the associated solvent. The properties of the ternary system are shown to be accounted for by the basic theory and, further, the theory can be employed to predict distribution behaviour in liquid—liquid systems where association occurs. It is shown that the theory can also be used to predict solute retention in reversed-phase liquid chromatography.

Peak dispersion and mobile phase velocity in liquid chromatography: the pertinent relationship for porous silica

The equations of Van Deemter, Giddings, Huber, Knox and Horvath for the height equivalent to a theoretical plate (H) are tested against over 25 data sets of experimental values of H and the mobile phase linear velocity (u) obtained for columns packed with silica gel. Each data set contains at least 10 complementary pairs of H and u values and furthermore, each H and u value was taken as the mean of at least three replicate measurements, thus, involving a total of over 750 individual and precise measurements of H and u. The maximum standard error for any set of replicate measurements was 2%. The data were obtained for silica gels having four different particle diameters, for six solvent mixtures and nine different solutes. It is shown that over the velocity range of 0.02–1 cm/sec, the Van Deemter equation accurately predicts the experimentally determined relationship between H and u. Consequently, under normal operating conditions in liquid chromatography, employing silica gel as the stationary phase Van Deemter equation can be employed with confidence in column design.

Effect of pressure on solute diffusivity, solvent viscosity and column temperature in liquid chromatography

Abstract The interrelationship between column pressure, solvent viscosity, solute diffusivity and column temperature is complex. Any increase in inlet pressure to provide a higher flow-rate and consequently a faster analysis increases solvent viscosity and column temperature and decreases solute diffusivity. However, a higher column temperature resulting from increased operating pressure reduces the solvent viscosity and increases the solute diffusivity, thereby masking the direct effect of pressure on these variables. Ipso facto, the net effect of pressure and temperature on solute diffusivity for an unthermostated column can be relatively small; consequently, the effect of pressure on column efficiency and column resolution can be minimal for unthermostated columns. However, the effect of this temperature increase on solute retention is very significant for unthermostated columns and leads to a 5% change in the value of k′ for a solute by merely changing the flow-rate from 0.5 to 5 ml/min. Hence, as the heat generated in the column is directly related to the flow-rate and further as the heat transfer through the packed bed of the column is very poor, the use of well thermostated small-bore columns could be mandatory for the precise measurement of solute retention.

Use of chromatographic data to determine the molecular weight of a solute eluted from a liquid chromatographic column

The diffusivities of 69 different compounds in a n-hexane—ethyl acetate solvent mixture have been determined and a precise relationship between solute diffusivity, molecular weight and molar volume established. The band dispersion for each of the same solutes has also been measured employing a liquid chromatographic column designed to emphasize the resistance-to-mass transfer factor and minimize thermal effects resulting from the use of high pressures and high mobile phase velocities. The effect of the capacity ratio of a solute on the resistance-to-mass transfer factor is determined and the relationship between the bandwidth of a solute, its diffusivity and its molecular weight established. A procedure is outlined for the determination of the molecular weight of a solute from the measurement of its bandwidth, when eluted from a liquid chromatographic column, within an error of 13% for 90% of the solutes examined providing the density of the solute lies between 0.85 and 1.25 g/ml.

Low-dispersion connecting tubes for liquid chromatography systems

Abstract The dispersion that takes place in coiled and serpentine tubes of different dimensions is reported. The advantage of using serpentine tubes suitably encased as connecting tubes in liquid chromatography is demonstrated. Such connecting tubes have been shown to have a variance contribution of less than 0.05 μl 2 /cm of linear length and a pressure drop of less than 0.05 MPa/cm at a flow-rate of 3 ml/min.

Liquid chromatography column design

Abstract A liquid chromatography column design protocol is described utilizing three data bases which are defined as performance criteria, instrument constraints and elective variables. The optimum column length, column radius and particle size of the packing to provide minimum analyses time can be calculated from the information contained in these three data bases. An explicit equation is also derived to permit the optimum particle diameter to be calculated. It is shown that if the inlet pressure is limited, small particles are only suitable for use in short columns for simple separations. Conversely, difficult separations can only be achieved with larger particles packed in long columns. In liquid chromatographic analyses, operating at 6000 p.s.i. as opposed to 4000 p.s.i. results in a proportional reduction in analysis time (about 30%). It follows that a maximum inlet pressure of 4000 p.s.i. appears to be quite adequate and is to be recommended for general liquid chromatographic analysis. The optimum k′ value of the first solute of the critical pair of a complex mixture can range between 2 and 6; furthermore the optimization procedure compensates for changes in diffusivity by corresponding changes in optimum particle diameter and optimum column length. Consequently the numerical value of solute diffusivity is not critical. The quality of the packing remains important even for fully optimized columns and consequently, the best packing procedures should always be employed.

Peak dispersion in a liquid chromatography-atomic-absorption spectrometry system

The dispersion characteristics of a flame atomic-absorption spectrophotometer, when used in conjunction with a chromatographic column and with different interfaces, were examined. It was found that the AA unit alone produces excessive peak dispersion (variance 43 µl2) and consequently, when used in conjunction with modern high-performance liquid chromatographic columns, their capabilities cannot be fully realised. The use of low-dispersion serpentine tubing, however, does permit some of the advantages of such columns to be obtained, provided the solutes are eluted at k′ values in excess of 3. It is found that the polyethylene tubing commonly used as an interface between the LC and the AA units has a variance contribution of 1.7 µl2 cm–1, which, for a practical length of 50 cm, is 83.5 µl2 and consequently far too high for the efficient use of the LC-AAS system. Should modern atomic-absorption spectrophotometers be designed to provide sufficiently low dispersion so that they can be used with modern high-performance liquid chromatographic columns, then the major contribution to solute dispersion will reside in the interface and, consequently, the use of low-dispersion serpentine tubing will become essential.

Chromatography Coldhn Design

Publisher Summary This chapter discusses the chromatography column design. The heart of the chromatograph is the column where the separation takes place. Two processes must occur simultaneously and progressively during the development of the chromatogram in order to achieve a separation. These two processes proceed more or less independently of one another. Firstly, the individual solutes are moved spatially apart as a result of the different molecular interactions that take place between the molecules of the two phases and those of the solutes. Secondly, by careful column design, the migrating solute bands are kept sufficiently narrow such that each band is eluted discretely. Unless exclusion processes are being employed to aid in the separation, the movement of the peaks apart in the column is solely controlled by the phase system selected.

A multifunctional liquid chromatographic detector

A liquid chromatographic detector is described that monitors chromatographic column eluents simultaneously by UV absorption at 254 nm, fluorescence by excitation at 254 nm and electrical conductivity. The response of the detector has a linear range of over three orders of magnitude for each detecting function. The absolute sensitivities, defined as that concentration of solute that provides a signal to noise ratio of two, are 1.7 × 10–7 g ml–1 of toluene, 2.5 × 10–8 g ml–1 of dansylisoleucine and 5 × 10–8 g ml–1 of sodium chloride for JV absorption, fluorescence and electrical conductivity, respectively. The detector employs low-dispersion, serpentine tubing for column detector connections and has a cell volume of 2.2 µl. The dispersion of the detector has a standard deviation of only 2.8 µl at a flow-rate of 2 ml min–1 and is this suitable for use with modern high-speed 3-µm columns or high mass sensitivity, low solvent consumption, small bore columns. In addition to providing a simple basic detector with increased flexibility and versatility, simultaneous monitoring on all three functions can provide added confirmation of solute identification.