THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity
Abstract 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 remai...
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| Format: | Article |
| Language: | English |
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BMC
2021-03-01
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| Series: | Biotechnology for Biofuels |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s13068-021-01904-2 |
| _version_ | 1818553903336128512 |
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| author | Abhishek S. Patri Ramya Mohan Yunqiao Pu Chang G. Yoo Arthur J. Ragauskas Rajeev Kumar David Kisailus Charles M. Cai Charles E. Wyman |
| author_facet | Abhishek S. Patri Ramya Mohan Yunqiao Pu Chang G. Yoo Arthur J. Ragauskas Rajeev Kumar David Kisailus Charles M. Cai Charles E. Wyman |
| author_sort | Abhishek S. Patri |
| collection | DOAJ |
| description | Abstract 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. |
| first_indexed | 2024-12-12T09:31:54Z |
| format | Article |
| id | doaj.art-68b5c8f46de24a4eaadb212745841c46 |
| institution | Directory Open Access Journal |
| issn | 1754-6834 |
| language | English |
| last_indexed | 2024-12-12T09:31:54Z |
| publishDate | 2021-03-01 |
| publisher | BMC |
| record_format | Article |
| series | Biotechnology for Biofuels |
| spelling | doaj.art-68b5c8f46de24a4eaadb212745841c462022-12-22T00:28:49ZengBMCBiotechnology for Biofuels1754-68342021-03-0114111310.1186/s13068-021-01904-2THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activityAbhishek S. Patri0Ramya Mohan1Yunqiao Pu2Chang G. Yoo3Arthur J. Ragauskas4Rajeev Kumar5David Kisailus6Charles M. Cai7Charles E. Wyman8Department of Chemical and Environmental Engineering, Bourns College of Engineering, University of California, RiversideMaterials Science & Engineering Program, Bourns College of Engineering, University of California, RiversideCenter for Bioenergy Innovation (CBI), Oak Ridge National Laboratory (ORNL)Center for Bioenergy Innovation (CBI), Oak Ridge National Laboratory (ORNL)Center for Bioenergy Innovation (CBI), Oak Ridge National Laboratory (ORNL)BioEnergy Science Center (BESC), Oak Ridge National Laboratory (ORNL)Department of Chemical and Environmental Engineering, Bourns College of Engineering, University of California, RiversideDepartment of Chemical and Environmental Engineering, Bourns College of Engineering, University of California, RiversideDepartment of Chemical and Environmental Engineering, Bourns College of Engineering, University of California, RiversideAbstract 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.https://doi.org/10.1186/s13068-021-01904-2BiomassPretreatmentDilute acidTetrahydrofuranLigninEnzyme |
| spellingShingle | Abhishek S. Patri Ramya Mohan Yunqiao Pu Chang G. Yoo Arthur J. Ragauskas Rajeev Kumar David Kisailus Charles M. Cai Charles E. Wyman THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity Biotechnology for Biofuels Biomass Pretreatment Dilute acid Tetrahydrofuran Lignin Enzyme |
| title | THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity |
| title_full | THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity |
| title_fullStr | THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity |
| title_full_unstemmed | THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity |
| title_short | THF co-solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity |
| title_sort | thf co solvent pretreatment prevents lignin redeposition from interfering with enzymes yielding prolonged cellulase activity |
| topic | Biomass Pretreatment Dilute acid Tetrahydrofuran Lignin Enzyme |
| url | https://doi.org/10.1186/s13068-021-01904-2 |
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