Investigating and applying free solution capillary electrophoresis with direct UV detection to bioethanol research

Abstract

Bioethanol fermentation is an important process that is reducing the global demand on fossil fuels and remains a field of research for the foreseeable future. Carbohydrates are sourced from crops and hydrolyzed into simpler sugars then fermented into ethanol. Fermentation of sugars sourced from food crops presents a sustainability issue with a growing population. Fermentation of lignocellulosic material is more sustainable since it is sourced from non-food crops or wastes. It is comprised of a variety of both pentose and hexose sugars. The composition and ratio of these varies depending on the source of the material. Accurate analysis of the material is essential for both the monitoring of the hydrolysis and its fermentation to valuable end-products such as ethanol. Although there have been major advances in novel fermentation processes and the discovery and construction of novel microorganisms, development of methods for analysis of these complex substrates and their fermentation to ethanol has not advanced as rapidly. High Performance Liquid Chromatography (HPLC) is one of the most common techniques for the analysis of these complex substrates. The resolution of popular HPLC modes was compared and no mode was found to have complete separation of common fiber sugars. Free solution Capillary Electrophoresis / capillary zone electrophoresis (CE) is used and recognized in both research and industry as a viable technique for the separation of carbohydrates. Recent studies on the use of direct UV detection for determination of underivatized carbohydrates have shown great promise and, in this work, the technique was applied to lignocellulosic plant fiber analysis as well as monitoring its fermentation to ethanol and sugar alcohols. All resolution values with CE were higher than 0.5, in contrast to any HPLC mode investigated. The running cost of HPLC, for this application, is also much higher than CE. Determination of carbohydrates from lignocellulosic fiber by both HPLC on a cation exchange resin and by CE resulted in values 17-22 % higher with CE than HPLC. The influence of the counter-ion in the BackGround Electrolyte (BGE) was found to affect the resolution and time of the separation. 130 mM KOH was shown to be effective for a fast separation of simple mixtures and a mixture of 65 mM NaOH and 65 mM LiOH achieved a better resolution with more complex carbohydrate mixtures than the other BGE's studied. In a quantitative study on fermentation samples, CE, HPLC and High Performance Anion Exchange Chromatography (HPAEC) closely agreed within experimental error (less than 7 % difference from the average total detected amount).

Date of Award	2014
Original language	English