Bioconversion of lignin into lipids via heterophic oleaginous Rhodococci

The development of a lignin-to-lipids metabolic pathway can broaden the application of certain recalcitrant lignocellulose constituents, and thereby optimize an integrated biorefinery strategy. To review, we have assessed Kraft lignin, ethanol organosolv lignin (EOL), and lignocellulosic organosolv pretreatment effluent as suitable substrates for oleaginous Rhodococcus opacus in order to develop a sustainable microbial biodiesel platform. This evaluation was based on cell sustainability as measured by cell dry weight (CDW), living cell or colony forming unit concentration (CFU/mL), optical density (OD), and percentage of total lipid accumulation as well as lipid composition. In addition to this, we characterized substrates throughout fermentation via gel permeation chromatography (GPC), HPLC, ¹H-NMR, ¹³C-NMR, and 2D-HSQC. In our first demonstration, EOL was utilized sufficiently by R. opacus DSM1069 as a sole carbon and energy source resulting in an accumulation of lipid reserves up to 4.08% CDW. In subsequent studies, we elucidated the degradation pattern by R. opacus PD630 and DSM1069 using softwood Kraft lignin. We observed degradations of select aromatic units and β-O-4 linkages in some lower molecular weight (MW) lignins during the first step, following by the further degradation of high MW lignins. Our investigation with organosolv pretreatment effluent showed that R. opacus DSM 1069 was capable of accumulating 26.99% of the CDW in single cell oils (SCOs), which were predominantly composed of oleic, palmitic, and stearic fatty acid. Upon characterization, we observed the degradation of low molecular weight components coupled with depletion of sugars, low MW lignin and distinct lignin modification.