In “comprehensive” two-dimensional liquid chromatography, the column effluent from the first separation system (the first dimension) is sequentially sampled by the second dimension separation system. The total analysis time is largely determined by the speed of the second dimension separation system; the most retained component must elute before the least retained component of the next second dimension separation. Optimization of multidimensional separation systems requires that one understand the relationship between system resolution and the number of second dimension samples across a first dimension peak. In this paper, we study the theoretical and experimental aspects of this sampling process. To obtain high two-dimensional resolution, each peak in the first dimension should be sampled at least three times into the second dimension when the sampling is in-phase. If the sampling is maximally out of phase, there should be at least four samples per peak for high-fidelity separation. The sensitivity of the resolution with respect to the sampling phase is discussed in detail and shown to be insignificant when four or greater samples are taken across the first dimension peak width. These results suggest optimal criteria for method development with multidimensional chromatography.

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The effect of liquid chromatographic separation on matrix-related signal suppression in electrospray ionization mass spectrometry (LC-ESI-MS) was investigated. A method incorporating on-line two-dimensional liquid chromatography mass spectrometry (LC/LC-MS) was developed to compensate for matrix effects and signal suppression in qualitative and quantitative analysis. The LC/LC-MS(MS) approach was successfully applied for single-component and multicomponent analysis in a variety of complex matrixes. It was demonstrated that matrix-related signal suppression could be induced solely by (i) column overload, (ii) matrix component-analyte coelution, or a combination of each. Application of on-line orthogonal LC/LC separations can be effective in reducing both causes of matrix-related signal suppression effects i.e., column overload and matrix-analyte coelution for a variety of LCn/MSn applications.

The electrokinetic chromatography (EKC) of a novel mixed surfactant system consisting of oppositely charged surfactants, sodium dodecyl sulfate (SDS) and n-dodecyltrimethylammonium bromide (DTAB), was investigated. The chromatographic characteristics of large liposome-like spontaneous vesicles and rodlike mixed micelles formed from the mixture were explored and compared with those of SDS micelles. Separations of a series of n-alkylphenones showed that the spontaneous vesicles provided about a 2 times wider elution window than SDS micelles. Both vesicle and mixed micelle systems were found to provide larger methylene selectivity than SDS. The different elution order of a group of nitrotoluene geometric isomers with DTAB/SDS spontaneous vesicles and SDS micelles pseudostationary phases suggested the possibility of different separation mechanisms with these two systems. Comparisons of polar group selectivity, retention, and efficiency were made between vesicles, mixed micelles, and SDS micelles. The correlation between the logarithms of the retention factors (log k') and octanol-water partition coefficients (log P(ow)) for a group of 20 neutral compounds was also studied with DTAB/SDS vesicles. Spontaneous vesicles have great potential as a pseudostationary phase in electrokinetic chromatography.