THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity

Patri, Abhishek S.; Mohan, Ramya; Pu, Yunqiao; Yoo, Chang G.; Ragauskas, Arthur J.; Kumar, Rajeev; Kisailus, David; Cai, Charles M.; Wyman, Charles E.

doi:10.1186/s13068-021-01904-2

Title: THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity

Journal Article · Tue Mar 09 00:00:00 EST 2021 · Biotechnology for Biofuels

DOI:https://doi.org/10.1186/s13068-021-01904-2· OSTI ID:1788081

Patri, Abhishek S. ^[1]; Mohan, Ramya ^[2]; Pu, Yunqiao ^[3]; Yoo, Chang G. ^[4]; Ragauskas, Arthur J. ^[5]; Kumar, Rajeev ^[6]; Kisailus, David ^[6]; Cai, Charles M. ^[1];

^[1]

Univ. of California, Riverside, CA (United States); Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
Univ. of California, Riverside, CA (United States)
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States)
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); State Univ. of New York (SUNY), Syracuse, NY (United States)
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Univ. of Tennessee, Knoxville, TN (United States)
Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Univ. of California, Riverside, CA (United States)

Background: Conventional aqueous dilute sulfuric acid (DSA) pretreatment of lignocellulosic biomass facilitates hemicellulose solubilization and can improve subsequent enzymatic digestibility of cellulose to fermentable glucose. However, much of the lignin after DSA pretreatment either remains intact within the cell wall or readily redeposits back onto the biomass surface. This redeposited lignin has been shown to reduce enzyme activity and contribute to rapid enzyme deactivation, thus, necessitating significantly higher enzyme loadings than deemed economical for biofuel production from biomass. Results: In this study, we demonstrate how detrimental lignin redeposition on biomass surface after pretreatment can be prevented by employing Co-solvent Enhanced Lignocellulosic Fractionation (CELF) pretreatment that uses THF–water co-solvents with dilute sulfuric acid to solubilize lignin and overcome limitations of DSA pretreatment. We first find that enzymatic hydrolysis of CELF-pretreated switchgrass can sustain a high enzyme activity over incubation periods as long as 5 weeks with enzyme doses as low as 2 mg protein/g glucan to achieve 90% yield to glucose. A modified Ninhydrin-based protein assay revealed that the free-enzyme concentration in the hydrolysate liquor, related to enzyme activity, remained unchanged over long hydrolysis times. DSA-pretreated switchgrass, by contrast, had a 40% drop in free enzymes in solution during incubation, providing evidence of enzyme deactivation. Furthermore, measurements of enzyme adsorption per gram of lignin suggested that CELF prevented lignin redeposition onto the biomass surface, and the little lignin left in the solids was mostly integral to the original lignin–carbohydrate complex (LCC). Scanning electron micrographs and NMR characterization of lignin supported this observation. Conclusions: Enzymatic hydrolysis of solids from CELF pretreatment of switchgrass at low enzyme loadings was sustained for considerably longer times and reached higher conversions than for DSA solids. Analysis of solids following pretreatment and enzymatic hydrolysis showed that prolonged cellulase activity could be attributed to the limited lignin redeposition on the biomass surface making more enzymes available for hydrolysis of more accessible glucan.

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Research Organization:: Oak Ridge National Laboratory (ORNL), Oak Ridge, TN (United States)

Sponsoring Organization:: USDOE Office of Science (SC), Biological and Environmental Research (BER). Biological Systems Science Division; US Air Force Office of Scientific Research (AFOSR); US Army Research Office (ARO); National Institute of Justice

Grant/Contract Number:: AC05-00OR22725; FA9550-15-1-0009; W911NF-16-1-0208; 2016-R2-CX-0015

OSTI ID:: 1788081

Alternate ID(s):: OSTI ID: 1819514

Journal Information:: Biotechnology for Biofuels, Vol. 14, Issue 1; ISSN 1754-6834

Publisher:: BioMed CentralCopyright Statement

Country of Publication:: United States

Language:: English

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THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity Patri, Abhishek S.; Mohan, Ramya; Pu, Yunqiao https://doi.org/10.6084/m9.figshare.c.5333223 The current record is supplemented by this collection	collection	January 2021
Additional file 1 of THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity Patri, Abhishek S.; Mohan, Ramya; Pu, Yunqiao https://doi.org/10.6084/m9.figshare.14190078.v1 The current record is supplemented by this text	text	January 2021
Additional file 1 of THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity Patri, Abhishek S.; Mohan, Ramya; Pu, Yunqiao https://doi.org/10.6084/m9.figshare.14190078 The current record is supplemented by this text	text	January 2021

Title: THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity

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