Ruyu Li

Sodium hydroxide pretreatment of genetically modified switchgrass for improved enzymatic release of sugars.

Overcoming biomass recalcitrance to bioconversion is crucial for cellulosic biofuels commercialization. In this study, Alamo switchgrass (Panicum virgatum L.) was genetically transformed to suppress the expression of 4-coumarate-CoA ligase (4CL). The transgenic plants were determined to have lignin content reductions of up to 5.8%. The ratios of acid soluble lignin (ASL) to acid insoluble lignin (AIL) and syringyl/guaiacyl (S/G) in transgenic plants were 21.4-64.3% and 11.8-164.5%, respectively, higher than those of conventional biomass. Both conventional and transgenic plants were pretreated with 0.5%, 1%, and 2% (w/v) NaOH for 15, 30, and 60min at 121°C, followed by enzymatic hydrolysis with commercial cellulases and xylanases. At the optimal conditions, the glucan and xylan conversion efficiency in the best transgenic plants were 16% and 18% higher than the conventional plant, respectively. The results show that down-regulation of 4CL gene promoted enzymatic hydrolysis of plant cell walls following a mild alkali pretreatment.

Dilute sulfuric acid pretreatment of transgenic switchgrass for sugar production.

Conventional Alamo switchgrass and its transgenic counterparts with reduced/modified lignin were subjected to dilute sulfuric acid pretreatment for improved sugar production. At 150 °C, the effects of acid concentration (0.75%, 1%, 1.25%) and residence time (5, 10, 20, 30 min) on sugar productions in pretreatment and enzymatic hydrolysis were investigated, with the optimal pretreatment conditions determined for each switchgrass genotype based on total sugar yield and the amounts of sugar degradation products generated during the pretreatment. The results show that genetic engineering, although did not cause an appreciable lignin reduction, resulted in a substantial increase in the ratio of acid soluble lignin:acid insoluble lignin, which led to considerably increased sugar productions in both pretreatment and enzymatic hydrolysis. At an elevated threshold concentration of combined 5-hydroxyfuranmethal and furfural (2.0 g/L), the overall carbohydrate conversions of conventional switchgrass and its transgenic counterparts, 10/9-40 and 11/5-47, reached 75.9%, 82.6%, and 82.2%, respectively.

CO2-Enrichment and photosynthetic photon flux affect the growth of in vitro-cultured apple plantlets

Micro-cuttings (shoots with two small leaves) of cultivar M9 apple were cultured in-vitro for 40 d under CO2-enriched and non-enriched (i.e., ambient air) conditions, and at a PPF of 40 or 100 μmol m-2 s-1 Afterward, shoot length, number of leaves, leaf area, chlorophyll content, shoot and root fresh weights, and % survival were recorded. Those plant-lets grown under CO2- and PPF-enriched treatments were healthy and vigorous, and showed higher values for their growth parameters. In contrast, those grown without supplemental CO2 or PPF often showed hyperhydricity. We also demonstrated that CO2 enrichment and a relatively high PPF during in-vitro culture promoted normal photosynthesis and growth after ex-vitro transplantation.

Effect of Dilute Alkali Pretreatment on Sugar Release from Transgenic Switchgrass

The recalcitrance of lignocellulosic materials to biochemical conversion is a major hurdle for cost-effective production of cellulosic sugars that can be processed into fuels and valuable chemicals. In this study, Alamo switchgrass (Panicum virgatum L.) was genetically transformed to suppress the expression of 4-coumarate-CoA ligase (4CL). The transgenic plants were determined to have lignin content reductions of up to 5.8%. The ratios of acid soluble lignin (ASL) to acid insoluble lignin (AIL) in transgenic plants were 21.4-64.3% higher than those of conventional biomass. Both conventional and transgenic plants were pretreated with alkalis at mild temperatures: lime at 50°C and the combination of lime and NaOH at 21°C, followed by enzymatic hydrolysis with commercial cellulases and xylanases. At the recommended conditions (0.1 g/g raw biomass and 12 h) for lime pretreatment at 50°C, the glucan and xylan conversions of transgenic switchgrass were 12% and 10%, respectively, higher than those of conventional plant. These increases were reduced to 7% and 8% for glucan and xylan conversions, respectively, when the best conditions (0.025 g lime/g raw biomass, 0.1 g NaOH/g raw biomass, and 6 h) for combined alkali pretreatment at 21°C were employed. The results show that down-regulation of 4CL gene promoted enzymatic hydrolysis of plant cell walls following mild alkaline pretreatments.

Dilute Sulfuric Acid Pretreatment of Genetically-Engineered Switchgrass for Ethanol Production

Switchgrass is a promising feedstock for bioethanol production due to its high biomass yield, potentially low requirements for agricultural inputs and positive environmental impacts. Genetic modification of conventional switchgrass by down-regulating lignin biosynthetic genes provides an option for improving biomass conversion efficiency. In this study, genetically modified switchgrass varieties were pretreated using dilute sulfuric acid at different combinations of acid concentration (0.75, 1, and 1.25 %, w/v) and residence time (5, 10, 20, and 30 min) to further break down the intrinsic complex structure of lignocellulosic biomass, and the yield of total sugar including xylose in prehydrolyzate and glucose in hydrolyzate was used to evaluate modification and pretreatment effectiveness. Preliminary results show that genetic modification caused the increased ratio of acid soluble lignin : acid insoluble lignin, which might be associated with the improved sugar production of some genetically modified switchgrass varieties. Increasing acid concentration or residence time didn’t necessarily result in better sugar production due to the increased degradation of monomeric sugars. After pretreatment using 1% sulfuric acid for 20 minutes, the total sugar production of genetically modified switchgrass Al10/9-40 reached 508 mg, which was 22% higher than the maximum yield obtained from the conversional switchgrass.