Chromatography factors affecting resolution

Before we discuss the influence of pH, additives, and temperature in more detail, we need to understand another important factor for optimization of peak resolution. Peak resolution is affected by the difference in retention time between peaks and the peak shape. We have discussed the intrinsic effect of band dispersion in chromatography, which leads to a symmetrical broadening of peaks. This peak width is described in a standardized way by the plate number. In practical LC, however, perfectly symmetric peaks are rather the exception, and it is an important criterion of method optimization to remove root causes for peak distortion. Every increase in peak asymmetry negatively affects resolution (under otherwise constant conditions). It must be emphasized in this context that Eq. 2.1 in Section 2.2.1 is only valid for Gaussian peaks with perfect symmetry, while any peak distortion will lead to smaller effective resolution. There are cases where the... 

In order to estimate resolution among peaks eluted from a chromatography column, those factors that affect N must first be elucidated. By definition, a low value of Hs will result in a large number of theoretical plates for a given column length. As discussed in Chapter 11, Equation 11.20 obtained by the rate model shows the effects of axial mixing of the mobile phase fluid and mass transfer of solutes on Hs. 

Capillary electrophoresis (CE) is an emerging analytical technique for determination of catechins. The majority of CE studies involve the analysis of catechins in tea infusion, extracts as well as supplements. The three variants of CE suitable for the analysis of catechins include capillary zone electrophoresis (CZE), micellar electro-kinetic chromatography (MEKC), and microemulsion electrokinetic chromatography (MEEKC) with UV detection. In general, the resolution of MEKC was found to be superior to CZE for separation of catechins. MEEKC is a relatively new technique, and the few reports available suggest that it offers a performance similar to MEKC. CE conditions are often quite complex, and many factors, such as buffer composition, pH, presence of surfactants, and column temperature, can all affect the quality of separation and should be optimized individually. On the other hand, CE offers several advantages over HPLC. The short analysis time (<20 minutes), low running costs, and reduced use of solvents make it an attractive alternative for routine analysis of catechins. 

---