Effects of Clostridium beijerinckii and Medium Modifications on Acetone-Butanol-Ethanol Production From Switchgrass.

The presence of lignocellulose-derived microbial inhibitory compounds (LDMICs) in lignocellulosic biomass (LB) hydrolysates is a barrier to efficient conversion of LB hydrolysates to fuels and chemicals by fermenting microorganisms. Results from this study provide convincing evidence regarding the effectiveness of metabolically engineered C. beijerinckii NCIMB 8052 for the fermentation of LB-derived hydrolysates to acetone-butanol-ethanol (ABE). The engineered microbial strain (C. beijerinckii_SDR) was produced by the integration of an additional copy of a short-chain dehydrogenase/reductase (SDR) gene (Cbei_3904) into the chromosome of C. beijerinckii NCIMB 8052 wildtype, where it is controlled by the constitutive thiolase promoter. The C. beijerinckii_SDR and C. beijerinckii NCIMB 8052 wildtype were used for comparative fermentation of non-detoxified and detoxified hydrothermolysis-pretreated switchgrass hydrolysates (SHs) with and without (NH4)2CO3 supplementation. In the absence of (NH4)2CO3, fermentation of non-detoxified SH with C. beijerinckii_SDR resulted in the production of 3.13- and 2.25-fold greater quantities of butanol (11.21 g/L) and total ABE (20.24 g/L), respectively, than the 3.58 g/L butanol and 8.98 g/L ABE produced by C. beijerinckii_wildtype. When the non-detoxified SH was supplemented with (NH4)2CO3, concentrations were similar for butanol (9.5 compared with 9.2 g/L) and ABE (14.2 compared with 13.5 g/L) produced by C. beijerinckii_SDR and C. beijerinckii_wildtype, respectively. Furthermore, when C. beijerinckii_SDR and C. beijerinckii_wildtype were cultured in detoxified SH medium, C. beijerinckii_SDR produced 1.11- and 1.18-fold greater quantities of butanol and ABE, respectively, than when there was culturing with C. beijerinckii_wildtype. When the combined results of the present study are considered, conclusions are that the microbial strain and medium modifications of the fermentation milieu resulted in greater production of fuels and chemicals from non-detoxified LB hydrolysates.

Viable strategies for enhancing acetone-butanol-ethanol production from non-detoxified switchgrass hydrolysates.

A process engineering strategy was investigated towards developing a viable scheme for effective conversion of hydrothermolysis pretreated non-detoxified switchgrass hydrolysates (SH) to acetone butanol ethanol (ABE) using a metabolically engineered strain of Clostridium beijerinckii NCIMB 8052, C. beijerinckii_AKR. The engineered strain was modified by homologous integration into the chromosome and constitutive expression of Cbei_3974, which encodes an aldo-keto reductase. Intermittent feeding strategy was employed in which fermentation was initiated with 30% of the SH and the remaining 70% SH was added when the optical density (OD600nm) of C. beijerinckii attained 0.5. The ABE (14.9 g/L) produced from non-detoxified SH by the inhibitor-tolerant C. beijerinckii_AKR was comparable to the P2-glucose control medium (14.7 g/L). Using intermittent feeding, wildtype and C. beijerinckii_AKR produced similar amounts of ABE (about 17.5 g/L). This shows that intermittent feeding strategy and C. beijerinckii_AKR enhanced ABE fermentation and eliminated the need for SH detoxification prior to fermentation.

Feasibility of using biochar as buffer and mineral nutrients replacement for acetone-butanol-ethanol production from non-detoxified switchgrass hydrolysate.

Biochar can be an inexpensive pH buffer and source of mineral and trace metal nutrients in acetone-butanol-ethanol (ABE) fermentation. This study evaluated the feasibility of replacing expensive 4-morpholineethanesulfonic acid (MES) P2 buffer and mineral nutrients with biochar made from switchgrass (SGBC), forage sorghum (FSBC), redcedar (RCBC) and poultry litter (PLBC) for ABE fermentation. Fermentations using Clostridium beijerinckii ATCC 51743 in glucose and non-detoxified switchgrass hydrolysate media were performed at 35 °C in 250 mL bottles for 72 h. Medium containing buffer and minerals without biochar was the control. Similar ABE production (about 18.0 g/L) in glucose media with SGBC, FSBC and RCBC and control was measured. However in non-detoxified switchgrass hydrolysate medium, SGBC, RCBC and PLBC produced more ABE (about 18.5 g/L) than the control (10.1 g/L). This demonstrates that biochar is an effective buffer and mineral supplement for ABE production from lignocellulosic biomass without costly detoxification process.