Sasikumar Elumalai

Hydroxycinnamate Conjugates as Potential Monolignol Replacements: In vitro Lignification and Cell Wall Studies with Rosmarinic Acid

The plasticity of lignin biosynthesis should permit the inclusion of new compatible phenolic monomers, such as rosmarinic acid (RA) and analogous catechol derivatives, into cell-wall lignins that are consequently less recalcitrant to biomass processing. In vitro lignin polymerization experiments revealed that RA readily underwent peroxidase-catalyzed copolymerization with monolignols and lignin oligomers to form polymers with new benzodioxane inter-unit linkages. Incorporation of RA permitted extensive depolymerization of synthetic lignins by mild alkaline hydrolysis, presumably by cleavage of ester intra-unit linkages within RA. Copolymerization of RA with monolignols into maize cell walls by in situ peroxidases significantly enhanced alkaline lignin extractability and promoted subsequent cell wall saccharification by fungal enzymes. Incorporating RA also improved cell wall saccharification by fungal enzymes and by rumen microflora even without alkaline pretreatments, possibly by modulating lignin hydrophobicity and/or limiting cell wall cross-linking. Consequently, we anticipate that bioengineering approaches for partial monolignol substitution with RA and analogous plant hydroxycinnamates would permit more efficient utilization of plant fiber for biofuels or livestock production.

Epigallocatechin gallate incorporation into lignin enhances the alkaline delignification and enzymatic saccharification of cell walls

BackgroundLignin is an integral component of the plant cell wall matrix but impedes the conversion of biomass into biofuels. The plasticity of lignin biosynthesis should permit the inclusion of new compatible phenolic monomers such as flavonoids into cell wall lignins that are consequently less recalcitrant to biomass processing. In the present study, epigallocatechin gallate (EGCG) was evaluated as a potential lignin bioengineering target for rendering biomass more amenable to processing for biofuel production.ResultsIn vitro peroxidase-catalyzed polymerization experiments revealed that both gallate and pyrogallyl (B-ring) moieties in EGCG underwent radical cross-coupling with monolignols mainly by β–O–4-type cross-coupling, producing benzodioxane units following rearomatization reactions. Biomimetic lignification of maize cell walls with a 3:1 molar ratio of monolignols and EGCG permitted extensive alkaline delignification of cell walls (72 to 92%) that far exceeded that for lignified controls (44 to 62%). Alkali-insoluble residues from EGCG-lignified walls yielded up to 34% more glucose and total sugars following enzymatic saccharification than lignified controls.ConclusionsIt was found that EGCG readily copolymerized with monolignols to become integrally cross-coupled into cell wall lignins, where it greatly enhanced alkaline delignification and subsequent enzymatic saccharification. Improved delignification may be attributed to internal trapping of quinone-methide intermediates to prevent benzyl ether cross-linking of lignin to structural polysaccharides during lignification, and to the cleavage of ester intra-unit linkages within EGCG during pretreatment. Overall, our results suggest that apoplastic deposition of EGCG for incorporation into lignin would be a promising plant genetic engineering target for improving the delignification and saccharification of biomass crops.

Thermo-chemical pretreatment of rice straw for further processing for levulinic acid production

A variety of pretreatment protocols for rice straw fiber reconstruction were evaluated under mild conditions (upto 0.2%wt. and 121°C) with the object of improving polymer susceptibility to chemical attack while preserving carbohydrate sugars for levulinic acid (LA) production. Each of the protocols tested significantly enhanced pretreatment recoveries of carbohydrate sugars and lignin, and a NaOH protocol showed the most promise, with enhanced carbohydrate preservation (upto 20% relative to the other protocols) and more effective lignin dissolution (upto 60%). Consequently, post-pretreatment fibers were evaluated for LA preparation using an existing co-solvent system consisting of HCl and THF, in addition supplementation of DMSO was attempted, in order to improve final product recovery. In contrast to pretreatment response, H2SO4 protocol fibers yielded highest LA conc. (21%wt. with 36% carbohydrate conversion efficiency) under the modest reaction conditions. Apparent spectroscopic analysis witnessed for fiber destruction and delocalization of inherent constituents during pretreatment protocols.

Combined sodium hydroxide and ammonium hydroxide pretreatment of post-biogas digestion dairy manure fiber for cost effective cellulosic bioethanol production

BackgroundThe current higher manufacturing cost of biofuels production from lignocellulosics hinders the commercial process development. Although many approaches for reducing the manufacturing cost of cellulosic biofuels may be considered, the use of less expensive feedstocks may represent the largest impact. In the present study, we investigated the use of a low cost feedstock: post-biogas digestion dairy manure fiber. We used an innovative pretreatment procedure that combines dilute sodium hydroxide with supplementary aqueous ammonia, with the goal of releasing fermentable sugar for ethanol fermentation.ResultsPost-biogas digestion manure fiber were found to contain 41.1% total carbohydrates, 29.4% lignin, 13.7% ash, and 11.7% extractives on dry basis. Chemical treatment were applied using varying amounts of NaOH and NH3 (2-10% loadings of each alkali on dry solids) at mild conditions of 100°C for 5 min, which led to a reduction in lignin of 16-40%. Increasing treatment severity conditions to 121°C for 60 min improved delignification to 17-67%, but also solubilized significant amounts of the carbohydrates. A modified severity parameter model was used to determine the delignification efficiency of manure fiber during alkaline pretreatment. The linear model well predicted the experimental values of fiber delignification for all pretreatment methods (R2 > 0.94). Enzymatic digestion of the treated fibers attained 15-50% saccharification for the low severity treatment, whereas the high severity treatment achieved up to 2-fold higher saccharification. Pretreatment with NaOH alone at a variety of concentrations and temperatures provide low delignification levels of only 5 − 21% and low saccharification yields of 3 − 8%, whereas pretreatment with the combination of NaOH and NH3 improved delignification levels and saccharification yields 2–3.5 higher than pretreatment with NH3 alone. Additionally, the combined NaOH and NH3 pretreatment led to noticeable changes in fiber morphology as determined by SEM and CrI measurements.ConclusionsWe show that combined alkaline treatment by NaOH and NH3 improves the delignification and enzymatic digestibility of anaerobically digested manure fibers. Although pretreatment leads to acceptable saccharification for this low-cost feedstock, the high chemical consumption costs of the process likely will require recovery and reuse of the treatment chemicals, prior to this process being economically feasibility.

Integrated two-stage chemically processing of rice straw cellulose to butyl levulinate

A two-stage reaction system was developed to synthesize butyl levulinate (BL), a derivative chemical of levulinic acid, from agricultural residue (rice straw). A single reactor was employed during the first processing stage for the conversion of rice straw cellulose to levulinic acid (LA) in a novel co-solvent system consisting of dilute phosphoric acid and tetrahydrofuran. The highest yield of 10.8% wt. LA concentration (i.e., ∼42% of theoretical LA yield) with intermediate residuals concentration of 1.5% wt. glucose and 0.5% wt. 5-hydroxymethylfurfural (5-HMF) on dry weight basis of biomass was obtained at modest reaction conditions. During subsequent esterification reaction, approximately 7.8% wt. BL yield (at 89% conversion yield) was achieved from the solvent extracted precipitate containing majorly LA and residual 5-HMF in the presence of 0.5M sulfuric acid using n-butanol. Based on comparative esterification results obtained using commercial chemicals (LA and 5-HMF), apparently 5-HMF exhibited ∼8% wt. BL yield through direct synthesis in the presence of sulfuric acid using n-butanol under the same specified reaction conditions. Alongside, effectiveness of co-solvent treatment on rice straw for potential fermentable sugar release (glucose) was investigated by subjecting the respective post-reaction solid residues to enzymatic digestion using cellulase and yielded highest of 11% wt. per wt. solids (27% wt. glucose conversion efficiency), amongst solid residues underwent different processing conditions.

Improved levulinic acid production from agri-residue biomass in biphasic solvent system through synergistic catalytic effect of acid and products

In this study, levulinic acid (LA) was produced from rice straw biomass in co-solvent biphasic reactor system consisting of hydrochloric acid and dichloromethane organic solvent. The modified protocol achieved a 15% wt LA yield through the synergistic effect of acid and acidic products (auto-catalysis) and the designed system allowed facile recovery of LA to the organic phase. Further purification of the resulting extractant was achieved through traditional column chromatography, which yielded a high purity LA product while recovering ∼85% wt. Upon charcoal treatment of the resultant fraction generated an industrial grade target molecule of ∼99% purity with ∼95% wt recovery. The system allows the solvent to be easily recovered, in excess of 90%, which was shown to be able to be recycled up to 5 runs without significant loss of final product concentrations. Overall, this system points to a method to significantly reduce manufacturing cost during large-scale LA preparation.

Advances in Transformation of Lignocellulosic Biomass to Carbohydrate-Derived Fuel Precursors

Cellulosebased, second-generation biofuels have been considered as an alternative fuel source to compensate for depleting fossil fuel reserves. Considering compounds that may be obtained from lignocellulose during biorefining, furfural, 5-hydroxymethylfurfural, and levulinic acid are among the most promising building blocks for energy fuels preparation via chemical or biological synthesis reactions and are thus described as platform chemicals. In this chapter, we present a review on advances made over the traditional strategies for the preparation of these fuel precursors from biomass. The recalcitrant nature of biomass, caused primarily by cellulose crystallinity and nonreactive lignin, hampers the successful commercialization of these valuable by-product chemicals. To date, different processes and production schemes have been adapted to improve product yields and lower production costs and include examples such as supermolecular structure modification of cellulose for improved saccharification using solvents or selective removal or displacement of biomass constituents such as lignin to improve enzyme mobility. Additionally, schemes have included direct conversion of biomass including forestry and secondary agricultural residues to platform chemicals using novel catalysts and reaction systems such biphasic or extractive distillation. Unfortunately, the details of most biomass chemical and biochemical reactions are still unclear, due to their complex nature, hindering improvements to the process. Thus, continued research and development is needed to further understand the biomass component characteristics, the overall cell wall, interrelationships between fractional components, transformation of component during reaction, and competing degradation reactions. This research is critical to enable natural lignocellulosic materials utilization to value-added chemicals during the production of fuels.

Sustainable Production of Chemicals and Energy Fuel Precursors from Lignocellulosic Fractions

From time immemorial, bioprocessing of lignocelluloses via chemical catalysis has been an impressive methodology of numerous value added commodities and energy fuel precursors (drop-in-fuel) synthesis. The most common technique for biomass fragmentation is catalytic hydrolysis using various acid catalysts covering inorganic or organic liquid acids as well as solid acids (heterogeneous). Most research in the past decade has been focused on cost-effective production of such biomass derived commodities with the aim of their commercialization. Till date, in order to improve final product yields and minimize production costs, various improvised production schemes have been developed like pretreatment methods for improved saccharification and displacement and/or reconstruction of recalcitrant biomass constituents, such as lignin to improve accessibility, employing multi-functional catalysts to promote single stage transformations, continuous extraction of desired product by use of specific solvents to improve product stability as well as to inhibit by-product formation, integration of physical processes for example microwave and ultrasonic irradiation resulting in decreased residence time, etc. With these technological advancements, researchers have overcome substantial limitations associated with lignocellulose transformation including mass-transfer hindrances and expensive downstream processing; as a result a wide array of commercially important chemicals and fuel precursors have been synthesised. The chapter provides an account of value addition to biomass via chemical catalysis of cellulosic, hemicellulosic and lignin fractions towards product chemicals synthesis.