Evaluation and improvement of the qualitative characteristics of micellar electrokinetic capillary chromatography

Author

Ricky Dean Holland

Date of Award

8-1992

Degree Type

Thesis

Degree Name

Master of Science

Major

Chemistry

Major Professor

Michael J. Sepaniak

Committee Members

Chambers, Barnes

Abstract

Analytical methodologies are presented herein that evaluate and improve the qualitative characteristics of micellar electrokinetic capillary chromatography (MECC). Qualitative analysis in MECC benefits from the facile manner in which retention can be manipulated by altering mobile phase composition. By comparing solute retention times, from standard and sample solutions separated using more than one set of MECC conditions, peak identification is rendered more reliable and the problems with matrix interferences are reduced. The methodologies in this work are applied to 10 common mycotoxins. The first phase of this work consisted of determining two sets of optimized mobile phase conditions, each providing complete resolution and unique selectivity towards the mycotoxins and evaluating reproducibility. This was accomplished by investigating the effects of micelle type, pH, and organic modifier and cyclodextrin mobile phase additives on retention of the mycotoxins. Replicate separations of the mycotoxins using the two mobile phase systems were carried out in both a consecutive and alternate manner (between the two systems) to establish retention time coefficients of variations (CVs). By normalizing a solute's retention time (t_r) to an adjacent-eluting solute, retention time CVs averaging less than 1 % were obtained. Two fully-resolved chromatograms (one using each mobile system) of the 10 mycotoxins could be obtained, including system rinsing and equilibration, in less than 45 minutes. The second phase of this work consisted of applying this methodology to the qualitative analyses of 5 randomly-generated mycotoxin/interferent mixtures (unknowns). Solute identification was based on retention data, which was reported using the following procedures: [1] measuring t_r from injection point to peak maximum, [2] calculating the solute's capacity factor, [3] utilizing the t_m marker (Sudan III added to each unknown) as a normalization standard, and [4] utilizing tm and a spiked mycotoxin, that eluted roughly midpoint in the MECC elution window, as normalization standards. Identifications of the unknowns were based on elution within allotted tolerance windows. The window positions were established via the injections of standard mycotoxin mixtures, while tolerances were based on average CVs from the reproducibility study. The two systems were also used in tandem, i.e., solute identification required aggreement with both mobile phase's retention criteria. The successful identification of solutes in the unknown mixtures was found to depend on the normalization procedure employed. In particular, a normalization method that had the solute and it's normalization standard eluting in proximity was determined to be important. This identification criteria was most closely accomplished by applying method [4]. The reduction of misidentification also required the tandem utilization of both mobile phase systems. The combining of normalization method [4] and tandem mobile phase identification, allowed all toxins from the 5 samples to be identified without a single mis-identification.

Recommended Citation

Holland, Ricky Dean, "Evaluation and improvement of the qualitative characteristics of micellar electrokinetic capillary chromatography. " Master's Thesis, University of Tennessee, 1992.
https://trace.tennessee.edu/utk_gradthes/12135