Solvent mismatch

Another injection-related effect that can diminish the separation performance is the diluent effect, also known as solvent mismatch. This occurs when the elution strength of the sample solvent is greater than the starting mobile-phase strength. The retention of the analyte on the stationary phase is less in the small plug of sample solvent than it is in the surrounding bulk... 

Band tailing causes inferior resolution and reduced precision. Thus conditions resulting in tailing or asymmetric peaks should be avoided. Peak asymmetry or band tailing can arise from several sources partially plugged column frits, void(s) in the column, buildup of sample components and impurities on the column inlet following multiple sample injections, sample overload, solvent mismatch with reference to the sample, chemical or nonspecific interactions (e.g., silanol effects), contamination by heavy metals, and excess void volume in the HPLC system. 

FIGURE 1.8 System peaks, (a) A commonly observed void volume refractive index pulse. It is indicative of a solvent mismatch, (b) A system peak that elutes a er the analyte of... 

Mobile phase/sample solvent mismatch is a common problem. As seen in the methanol example in Figure 1.16, peak shape can be grossly affected. Papers have been published dealing with the effect that tte injection solvent has on peak shape in acetonitrile mobile phases as well. Hoffman et al. [887] used benzyl alcohol, tryptophan, cimetidine, and phenylalanine as test solutes. A Cig column was used... 

Another effect of the mobile phase/sample solvent mismatch is a change in the sensitiviy (AR/AC) of the method for an analyte. As an example, Perlman and Kirschbaum [889] prepared a series of solutes (e.g., captopril, nadolol, o-nitroani-line, triamcinolone acetate, methylparaben) in neat solvents acetonitrile, methanol, DMSO, and dichloromethane. These solutions were then injected onto a C,g or I enyl column (2 = 214 run or 270 run) and eluted with 50/50 methanol/water or 38.8/1.1/960 ethanol/water/dichloromethane mobile phases. Significant differences in the peak areas resulted for some but not all analytes. Deterioration of peak shapes was also common. Prediction of these changes was nearly impossible. For example, o-nitoaniline (in methanol) exhibited an increased peak area in methanol/water, whereas p-nitroaniline was unaffected. An awareness of the unexpected and unpredictable effects the sample solvent has on both the quantitative results and the overall separation is critical when developing a method. 

The primary considerations in filtration are the pore size and composition of the filters [39]. The choices are based on the specific analyte and sample matrix. As mentioned previously, filters of 0.45 pm should be used prior to injection. The composition of the membrane must be compatible with both the sample and the solvent. Mismatch with a solvent can result in disintegration of the filter or leaching of some of its components. Mismatching with a sample can result in adsorption for example, proteins may bind to either very hydrophobic or very polar surfaces. 

Other applications that utilize different types of reversed-phase columns in both dimensions have been advocated by Carr (Stoll et al., 2006) for metabolomics work in small-molecule separations. These stationary phases include a pentafluorophenyl-propyl stationary phase in the first dimension and a carbon-coated zirconia material stationary phase in the second dimension. A common mistake in 2D method development is to mismatch the solvent system the two solvent systems must be miscible as discussed below. 

In CSGE, mildly denaturing solvents in an appropriate buffer can accentuate conformational changes produced by single-base mismatches in heteroduplexed DNA. This increases the differences in electrophoretic mobility between heteroduplex and homoduplex. 

Another application would be to minimize the swelling of a cross-linked elastomer in contact with a solvent. In this case, of course, one would be looking for a polymer giving the largest mismatch with the solubility parameters of the solvent to which the elastomer was to be exposed. 

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