Chromatography optimisation

Depending on the type of chromatography, optimisation can be fairly rapid. Optimisation in gas phase chromatography is easier than in liquid chromatography where the composition of the mobile phase plays a role. Computer software is available that has been specially designed to help determine the correct composition of the mobile phase. 

The migration of molecules between the stationary phase and the mobile phase is driven by random movement or diffusion, a factor which is deleterious to high resolution in all forms of chromatography. Since resolution in SEC is solely dependent on diffusion, unlike the other forms of chromatography, optimisation of stationary phase particles is important to improve mass transfer. Factors deleterious to mass transfer can be divided into three separate types those attributable to stagnant mobile phase in the pores of the particles, those caused by differential penetration of the solute molecules into the stationary phase and, finally, longitudinal diffusion between the particles (Snyder and Kirkland, 1979). 

Dry Lab Chromatography Modeling, LC Resources, Walnut Creek, CA, Spring 1994. S. M. Hitchen, HiPac Chromatography Optimisation Software, Phase Separations, Deeside, UK (1992). 

Delgado B, Perez E, Santano MC, Minguilldn C (2005) Enantiomer separation by counter-current chromatography. Optimisation and drawbacks in the use of L-proUne derivatives as chiral selectors. J Chromatogr A 1092 36 2... 

Aguilar, M. I., Hodder, A. N., and Hearn, M. T. W., High-performance liquid chromatography of amino acids, peptides, and proteins. LXV. Studies on the optimisation of the reversed-phase gradient elution of polypeptides. Evaluation of retention relationships with (3-endorphin-related polypeptides, /. Chromatogr., 327, 115, 1985. 

Temperature control is important for the accurate measurement of retention data, and has to be used with refractometer detectors (Section 2.4.5). Increasing the temperature can increase the speed of the separation, especially in exclusion chromatography, and usually increases the efficiency of the column (though the gain in efficiency can be lost if the mobile phase is not properly equilibrated). Complicated separations can often be optimised by increasing the temperature, but this is done very much on a trial and error basis, and most work in hplc is still done without temperature control. 

A. Jurado Lopez, M.D. Luque de Castro, Optimisation of Focused Microwave Digestion of Proteinaceous Binders Prior to Gas Chromatography, Talanta, 65, 1059 1062 (2005). 

Corr, L. T., Berstan, R. and Evershed, R. P. (2007b) Optimisation of derivatisation procedures for the determination of 813C values of amino acids using gas chromatography/combustion/isotope ratio mass spectrometry. Rapid Communications in Mass Spectrometry 21, 3759 3771. 

Merritt, D. A., Freeman, K. H., Ricci, M. P., Studley, S. A. and Hayes, J. M. (1995) Performance and optimisation of a combustion interface for isotope ratio monitoring gas chromatography/mass spectrometry. Analytical Chemistry 67, 2461 2473. 

The second group of recently developed ionic liquids is often referred to as task specific ionic liquids in the literature [15]. These ionic liquids are designed and optimised for the best performance in high-value-added applications. Functionalised [16], fluorinated [17], deuterated [18] and chiral ionic liquids [19] are expected to play a future role as special solvents for sophisticated synthetic applications, analytical tools (stationary or mobile phases for chromatography, matrixes for MS etc.), sensors and special electrolytes. 

Figures 4.31(c), 4.36 and 13.3 from Snyder and Kirkland, Introduction to Modern Liquid Chromatography, 2nd edn., (1979) 9.41(a), (b) and (c) from Cooper, Spectroscopic Techniques for Organic Chemists (1980) 9.46 from Millard, Quantitative Mass Spectrometry (1978) 4.17, 4.18, 4.31 (a), 4.33, 4.34(a), 4.37, 4.38, 4.43 and 4.45 from Smith, Gas and Liquid Chromatography in Analytical Chemistry (1988) figures 4.42 and 13.2 from Berridge, Techniques for the Automated Optimisation of Hplc Separations (1985) reproduced by permission of John Wiley and Sons Limited 11.1, 11.5, 11.6, 11.12, 11.13, 11.14, 11.18 and 11.19 from Wendlandt, Thermal Analysis, 3rd edn., (1986) reprinted by permission of John Wiley and Sons Inc., all rights reserved.

Ayrton J. et al., 1998. Optimisation and routine use of generic ultra-high flow-rate liquid chromatography with mass spectrometric detection for the direct online analysis of pharmaceuticals in plasma. J Chromatogr A 8282 199. 

Grushka, E., ed. Preparative-Scale Chromatography (Dekker, 1989). Reprinted from Sep. Sci. Technol. 22 (Nos. 8-10), (1987) 1791-2110. (Several articles in this volume deal with optimisation). 

Ahmed N, Hale K. 1994. A microassay for urinary phenol using capillary gas chromatography and optimised enzymic hydrolysis. Clin Chim Acta 230 201-208. 

The ethyl acetate solution of organic species from the pre-treatment scheme shown in Figure 1 is suitable for analysis by this method. In order to cover the range of common explosives several chromatography columns with different types of stationary phase are required to allow for difierent polarities and volatihties. Dimethylsiloxane, phenyl-modified dimethylsiloxane, cyanopropyl- phenyl- vinyl-modified dimethylsiloxane, and polyethylene glycol have been found to represent a useful set of stationary phases. Carefully optimised temperature programming is also needed to obtain the requisite resolution and avoid interferences [19, 20]. 

Specificity may be achieved through sample preparation, chromatographic selectivity, the selectivity of the detection method or combinations of these. It is often tempting to utilise the most selective detection method available (such as MS or MS/MS), since this can reduce the effort required in optimising the sample preparation and chromatography. However, whilst this is often the most expedient approach in early development, it may not always be suitable to transfer expensive, highly technical methods and instrumentation into a manufacturing environment if this is required. 

Having optimised the efficiency of a chromatographic separation the quality of the chromatography can be controlled by applying certain system suitability tests. One of these is the calculation of theoretical plates for a column and there are two other main parameters for assessing performance peak symmetry and the resolution between critical pairs of peaks. A third performance test, the peak purity parameter, can be applied where two-dimensional detectors such as diode or coulometric array or mass spectrometry detectors are being used. The reproducibility of peak retention times is also an important parameter for controlling performance. 

K. Jones, Optimisation procedure for the silanisation of silicas for reversed-phase high-performance liquid chromatography. I. Elimination of nonsignificant variables, J. Chromatogr., 392 (1987) 1-10. 

K. Jones, Process scale high-performance liquid chromatography. Part I An optimisation procedure to maximise column efficiency, Chromatographia, 25 (1988), 437-442. 

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