(409d) Transgenic Energycane As an Alternative Feedstock for Biodiesel Production: An Industrially Relevant Study

Lignocellulosic biomass has abundant renewable carbon that can be converted conveniently to biofuels such as bioethanol, biodiesel, and bio-jet fuel. Non-transgenic bioenergy crops have shown great potential for providing cellulosic sugars for bioethanol and high-value bioproduct production. To further increase the energy density of plants, high biomass bioenergy crops such as energycane, sugarcane, and sorghum, are being genetically modified to sequestrate carbon in the vegetative tissues and divert the carbon flux to synthesize and accumulate triacylglyceride (TAG) molecules (Luo et al., 2022; Parajuli et al., 2020; Vanhercke et al., 2019). These transgenic crops cater to both cellulosic sugars and vegetative lipids. Simulations based on lab-scale experiments with transgenic crop suggests that these transgenic crops have the potential to replace oilseeds as feedstocks for biodiesel production (Clark and Schwender, 2022; Huang et al., 2016; Maitra et al., 2022b). Recently, energycane has been successfully engineered to accumulate 30-fold higher TAG molecules and double the amount of total fatty acids in green leaves as compared to their wild-type variety. The hyperaccumulation of TAG molecules was achieved by overexpression of the lipogenic factors DGAT1-2 (Diacylglycerol acyltransferase1-2) and OLE1 (Oleosin1) along with the suppression of SDP1 (SUGAR-DEPENDENT1) and TGD1 (Luo et al., 2022). The amount of total fatty acid and TAG of the transgenic lines varied from 3.56 to 4.96 % and 0.11 to 1.52% of leaf dry weight, respectively (Luo et al., 2022).

However, since lipids in the vegetative tissues of transgenic energycane are present in lower amounts and are in a complex form (Maitra et al., 2021; Parajuli et al., 2020), appropriate and efficient bioprocessing technologies are needed to deconstruct the lignocellulosic structure and recover vegetative lipids without degenerating along with cellulosic sugars from the feedstock. Chemical-free hydrothermal pretreatment has been shown to maintain the lipid profile during processing and enrich biomass residues with vegetative lipids that can be recovered at the end of the process (Maitra et al., 2022b; Maitra et al., 2022a). The initial characterization of transgenic energycane and process optimization at the lab scale has been established (Luo et al., 2022; Maitra et al., 2022b; Maitra and Singh, 2021).

We are presenting the proof-of-concept of bioprocessing transgenic energycane at a pilot scale to demonstrate its potential commercialization as an alternative renewable feedstock for biodiesel production. The transgenic energycane plant was segregated into stems, green leaves, and brown leaves in the field during harvesting. Juice was extracted from stems. Pilot-scale continuous hydrothermal pretreatment reactor was used to pretreat bagasse at 50% solids loading at 190 ºC for 10 minutes followed by disk milling. The same pretreatment conditions were used to pretreat green and brown leaves at the lab scale. Chemical-free pretreatment had no adverse effect on the percent recovery and composition of total fatty acids and TAG fatty acids. After pretreatment, a major fraction of vegetative lipids remained in the biomass residues which was recovered at the back end. The lipid recovery efficiency of the total process for untreated biomass was calculated to be 75.9% which showed approximately 17% improvement for pretreated biomass residues (88.7%). In addition, enzymatic saccharification of pretreated biomass residues recovered > 90% of cellulosic sugars. Based on the average biomass yield of transgenic energycane to be 45 tons/ha (Chiluwal et al., 2018; Knoll et al., 2021; Yang et al., 2018), the calculated potential and actual lipid yields of transgenic energycane were 0.75 tons/ha and 0.66 tons/ha, respectively. The experimental results of the study show that transgenic energycane has the potential to surpass the oil yield of soybean if the metabolic burden due to the expression of transgenes does not have a considerable effect on the biomass yield of the transgenic variety.

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