Optimization of the Evaluation in Chromatography

Kuss, H.-J. Optimization of the Evaluation in Chromatography. In Kromidas, S., Eds HPLC Made to Measure. Wiley-VCH Weinheim, 2006 105-147. 

The development of displacement separations has historically been an empirical process and even though chromatographic theory may guide the selection of operating conditions the final stage must involve experimental validation. Typically, several experiments will be carried out at or near the conditions determined by the theory. The final stage in the procedure is either experimental or numerical optimization of the displacement process to produce optimal yields, purities and productivities. At this point, the relative efficacy of selective and conventional displacement chromatography can also be evaluated. 

The first concern in the selection of the sample preparation solvent is to optimize recovery. However, a secondary consideration is the sample solvent s effect on the analysis. This is true whether the analytical technique is ultraviolet spectroscopy (UV), high-performance liquid chromatography (HPLC), or gas chromatography (GC). The method development sequence can be described as (a) development of the chromatographic separation, (b) development of the sample preparation method, and then (c) evaluation and optimization of the interaction of the sample preparation with the instrumental method. 

One important factor influencing the resolution of the column is the velocity of the gas flow. This value must be chosen optimally for every column or column system used in gas chromatography. Too high a velocity of the gas in the column can lead to very bad separation of the compounds in the column. This means that it is possible for many components of the mixture to exit the column at the same time, thus making the evaluation of the results cumbersome and prone with error or practically impossible. Too low a velocity of the gas in the column leads to the formation of adsorption-desorption equilibrium, resulting in a poor resolution of the column as well. The gas flow in any modern gas chromatograph is usually controlled by a special controller called the mass flow controller. These devices are very exact. However, they need to be calibrated. There are three most popular devices to measure the gas flow and to calibrate the mass flow controller. 

Esteban, R.E. Martinez-Castro, L and Sanz, J. Evaluation and optimization of the automatic thermal desorption method in the gas chromatographic determination of plant volatile compounds. Journal of Chromatography 1993, 657, 155-164. 

Method development and optimization are started with review of the currently available methods within the company or in literature. Available methods are used as a starting point and evaluated against the method requirements set in the method definition. If necessary the method is optimized or redeveloped in order to fulfill the requirements. DOE tools (response surface design) are preferentially applied to obtain the best optimal conditions in terms of robustness. Application of DOE methodology is not new in chromatography and DOE is frequently applied also for enantiomeric separations in Especially in... 

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