ENZYMATIC SYNTHESIS OF XYLITOL

Xylitol is a 5-carbon sugar classified as a sugar alcohol or polyol that is derived from plant xylose. It is used in the food and pharmaceutical industries as an alternative sweetener. The benefits of xylitol over other sweeteners is that it does not react with amino acids in the Maillard reaction, has a sweetening power equivalent to sucrose with 40% less calories, is anticariogenic and undergoes insulin-independent metabolism Xylitol is currently used in chewing gum, oral hygiene products, and confectionaries. Xylitol is found in small amounts in some fruits, vegetables, algae and mushrooms, but not at a level for commercial extraction. The current commercial manufacturing method consists of chemical hydrogenation of xylose extracted from hardwood trees. This research will explore the enzymatic synthesis of xylitol from corn cob biomass using immobilized cell lysates from yeast cells housed in reactors. The goal is to use yeast cell lysates and not use genetically modified organisms or microbial fermentation systems.Hypothesis:Xylitol can be produced enzymatically by immobilized yeast cell lysate.Objectives:1. Screen yeast cell lysates grown under optimal conditions in batch systems for xylitol synthesis from xylose. These strains have been shown previously to synthesize xylitol from xylose. At least 5 different yeast strains will be used. Xylitol production will be determined spectrophotometrically. Determine xylitol synthesis as influenced by co-factor (NADH and NADPH) requirement.2. Select two yeast strains and immobilize cell lysates for xylitol synthesis from xylose in a reactor format. Immobilize cell lysates and use in a reactor format to synthesize xylitol from xylose with predetermined co-factor. Xylitol production will be determined spectrophotometrically and confirmed via HPLC analysis. Determine the efficiency (yield) and half-lives of the bioreactors.3. Investigate methods to degrade the hemicellulose fraction of corn cobs. Methods will include the use of dilute acids, heat, and pressure. Degradation will be determined via HPLC analysis.4. Investigate the use of corn cob hemicellulose hydrolysate as a substrate for the immobilized yeast lysate reactors to synthesize xylitol. Determine the efficiency and half-lives of the bioreactors.5. Explore the use of formate dehydrogenase to regenerate the co-factor in a batch system. Formate dehydrogenase will be added to the solution after xylitol synthesis. The generation of NAD(P)H will be determined spectrophotometrically.Time Line:This project should be completed in 5 years. It is expected that objective 1 will take 1 year to complete. Objective 2 will be initiated in year 2 will also take one year to complete. Objective 3 can be initiated in year 1 to be ready for use in year 3. Objective 4 can be initiated in year 3 and will take 2 years to complete. Objective 5 will be completed in the last or fifth year.