On-Line Monitoring of Enzymatic Bioprocesses by Microdialysis Sampling, Anion Exchange Chromatography, and Integrated Pulsed Electrochemical Detection

Abstract

Microdialysis sampling coupled to column liquid chromatography with integrated pulsed electrochemical detection (IPED) has been shown to be a hyphenation of techniques well suited for the analysis of oligomeric carbohydrates in a continuously changing matrix due to biological activity (Torto et al, 1995). Microdialysis provides a simultaneous sampling and sample clean-up step. Proper choice of a microdialysis membrane with known characteristics, e.g. molecular mass cut-off, porosity and sterilisability ensures enhanced performance of the technique in a crude medium, as it does not perturb the reaction under investigation. Only small amounts of the hydrolysis products (carbohydrates) are removed. Carbohydrates are separated in their enolate form at high pH, eliminating the need for pre- or post-column derivatisation. The chromatographic separation facilitates data evaluation, as carbohydrates are oxidised at the same potential during detection by IPED (Johnson et al, 1992). The purpose of this investigation was to develop an analytical system that could be used to sty dy a liquefaction step during the hydrolysis of wheat starch in a fermentation process where glucose is the substrate (see Chapter 8). System development was carried out using soluble starch according to Zulkowsky.