Conversion of a model saponine to its aglycone in a microfluidic channel by an enzyme entrapped in hydrogel

In this thesis, an effective and novel method for the production of baicalein was developed by hydrolysis of baicalin. Routine chemical and free enzyme hydrolysis procedures were performed for the production of baicalein and the yields obtained, were compared for the first time with the reaction efficiencies obtained from the microfluidic systems designed in this study. For enzymatic hydrolysis studies carried out in a microfluidic platform, a total of three different microreactor systems with the different channel length, geometry and volume - one with PDMS-glass (single channel, 100 μL) and the other two with glass-silicon-glass (Type-I and Type-II, 37 μL) - were used. Helix pomatia derived beta-glucuronidase (GUS) enzyme was immobilized in different microfluidic systems operating on the principle of continuous flow by silica-based sol-gel materials. Kinetic constants such as KM and VMAX for both free and encapsulated GUS enzyme in sol-gel materials were measured using pNPG (para-nitrophenol-β-D-glucuronic acid) and determined as 1,57 mM, 56,82 µmol ml-1 min-1 and 0,40 mM, 11,88 μmol ml-1 min-1, respectively. Yields of baicalein production with 4h acidic and basic hydrolysis of baicalin resulted in 10±3% and 0,10±0,02%, respectively, together with high by-product formation. 99±1% of baicalin molecule was converted to baicalein at the end of the 4 hours of incubation with free GUS enzyme without any by-product formation. After 4 hours of process period, the production yields of baicalein in PDMS-glass, glass-silicon-glass Type-I and Type-II microchips were determined as 62±2%, 50±1% and 10±2%, respectively. Although it has been found that the conversion efficiency of baicalein obtained from microreactors is lower than the yield obtained with free enzyme, it has been revealed that the developed microreactor systems can produce effective processing alternatives in the production of baicalein by providing the possibility of recovery and continuous operation of the enzyme.