(221c) Lipid Production and Biofuel Yield from Wood Hydrolysates Using Oleaginous Yeast “Cutaneotrichosporon Curvatus”

The United Nations' sustainable development goals seven and thirteen focus on improving the global energy mix and promoting net-zero global greenhouse gas emissions by 2050. Producing sustainable aviation fuels (SAF) is one of the energy applications that are most difficult to wean away from fossil fuels. Non-edible lignocellulosic biomass is the most abundant renewable bioresource in the ecosphere and can be used to generate SAF. It comprises polysaccharides (cellulose and hemicellulose) and a natural polymer (lignin).

One approach to producing SAF from sustainable lignocellulose is thermal deoxygenation (TDO). This process converts cellulose-derived levulinic acid to high-energy-density hydrocarbon liquids, which can be hydrotreated into molecules suitable for jet fuel applications. A current challenge of this conversion process includes reducing the amount of pentosan sugars feeding into the initial hydrolysis/dehydration producing the levulinic acid. In the strong acid environment of the hydrolysis/dehydration process, pentosan sugars convert to resinous materials that glaze over the equipment surfaces. One approach to reducing pentosan sugars in the feed stream is adding a pre-conditioning step that enables the preemptive removal of some hemicellulose sugars. These sugars could then be converted into other biochemical or biofuel products.

Current studies in microbial biofuel production have gained attention due to the deteriorating amount of fossil fuels. In this study, we explore the use of fermentable sugars produced from dilute sulfuric acid pre-conditioning of white pine wood chips for the production of biofuel precursors. Oleaginous yeast is suggested as a viable lipid producer to promote a more sustainable biofuel industry. Using oleaginous yeasts to consume unwanted sugars has various advantages, such as a fast growth rate, high lipid accumulation that does not compete with human food, ease of handling, and no environmental or climate effects. Studies have shown that Cutaneotrichosporon Oleaginosus has the highest lipid content and biofuel yield among other yeast strains. We explore sustainable and efficient renewable sources such as wood pre-hydrolysates for biofuel production.

Methods: White pine chips were used as the starting biomass material. Mild acid (0.25% H2SO4) pre-conditioning of the wood chips was carried out at Idaho National Lab using a ton-per-day plug screw feeder with a reactor operating at different operating conditions, including different feed rates, particle sizes, temperatures, and feedstock concentrations. The pre-hydrolysate was analyzed using HPLC (HPX-87H and HPX-87P columns) and contained glucose, xylose, mannose, arabinose and galactose, and some degradation compounds. Batch fermentation was used to culture the oleaginous yeast strain, C. Oleaginosus, using wood hydrolysate as the principal substrate with additional defined nutrients. Production and accumulation of microbial lipids in the cells are enabled by nitrogen-limited stagnant growth in the presence of abundant sugars. Degradation compounds such as hydroxymethylfurfural (HMF), acetic acid, and furfural can significantly impact the growth of microorganisms, reducing their metabolic rate and limiting their lipid yield. Detoxification of hydrolysates using over-liming and activated carbon adsorption reduced this hazard. Hydrothermolysis (HTL) processing of the whole oleaginous yeast or their extracted lipids will be performed with resulting products characterized by GC-MS for applicability to biofuel applications.

Results: HPLC analysis of the pine pre-hydrolysis showed that most solubilized sugars were monomer sugars, and several degradation products were also present. Glucose and mannose were the sugars most abundant in the pre-hydrolysate. Growth of the oleaginous yeast C. Oleaginosus showed consumption of all the types of sugars, though glucose and mannose were the most sugars completely consumed. Glucose was the most rapidly consumed sugar. The effects of over-liming and activated carbon adsorption on wood hydrolysate were investigated, with activated carbon appearing more effective at reducing identified contaminants. Acid hydrolysis and solvent extraction of the disrupted cells and lipid recovery will be investigated. HTL of the whole cells and extracted lipids will be carried out, and the final products will be analyzed via GC-MS.

Implications: The TDO process with hydrotreatment has been shown to be effective at converting cellulose to SAF. Work from this project seeks to improve the operability of the acid hydrolysis/dehydration reaction that initiated the TDO process and increase the conversion of raw materials through fermentation of byproduct sugars and conversion of microbial lipids via HTL.