Sugar loss and enzyme inhibition due to oligosaccharide accumulation during high solids-loading enzymatic hydrolysis

Saisi Xue, Nirmal Uppugundla, Michael J. Bowman, David Cavalier, Leonardo Da Costa Sousa, Bruce E. Dale, Venkatesh Balan

    Research output: Contribution to journalArticle

    • 13 Citations

    Abstract

    Background: Accumulation of recalcitrant oligosaccharides during high-solids loading enzymatic hydrolysis of cellulosic biomass reduces biofuel yields and increases processing costs for a cellulosic biorefinery. Recalcitrant oligosaccharides in AFEX-pretreated corn stover hydrolysate accumulate to the extent of about 18-25 % of the total soluble sugars in the hydrolysate and 12-18 % of the total polysaccharides in the inlet biomass (untreated), equivalent to a yield loss of about 7-9 kg of monomeric sugars per 100 kg of inlet dry biomass (untreated). These oligosaccharides represent a yield loss and also inhibit commercial hydrolytic enzymes, with both being serious bottlenecks for economical biofuel production from cellulosic biomass. Very little is understood about the nature of these oligomers and why they are recalcitrant to commercial enzymes. This work presents a robust method for separating recalcitrant oligosaccharides from high solid loading hydrolysate in gramme quantities. Composition analysis, recalcitrance study and enzyme inhibition study were performed to understand their chemical nature. Results: Oligosaccharide accumulation occurs during high solid loading enzymatic hydrolysis of corn stover (CS) irrespective of using different pretreated corn stover (dilute acid: DA, ionic liquids: IL, and ammonia fibre expansion: AFEX). The methodology for large-scale separation of recalcitrant oligosaccharides from 25 % solids-loading AFEX-corn stover hydrolysate using charcoal fractionation and size exclusion chromatography is reported for the first time. Oligosaccharides with higher degree of polymerization (DP) were recalcitrant towards commercial enzyme mixtures [Ctec2, Htec2 and Multifect pectinase (MP)] compared to lower DP oligosaccharides. Enzyme inhibition studies using processed substrates (Avicel and xylan) showed that low DP oligosaccharides also inhibit commercial enzymes. Addition of monomeric sugars to oligosaccharides increases the inhibitory effects of oligosaccharides on commercial enzymes. Conclusion: The carbohydrate composition of the recalcitrant oligosaccharides, ratios of different DP oligomers and their distribution profiles were determined. Recalcitrance and enzyme inhibition studies help determine whether the commercial enzyme mixtures lack the enzyme activities required to completely de-polymerize the plant cell wall. Such studies clarify the reasons for oligosaccharide accumulation and contribute to strategies by which oligosaccharides can be converted into fermentable sugars and provide higher biofuel yields with less enzyme.

    Original languageEnglish (US)
    Article number195
    JournalBiotechnology for Biofuels
    Volume8
    Issue number1
    DOIs
    StatePublished - Nov 26 2015

    Profile

    Oligosaccharides
    Hydrolysis
    Carbohydrates
    Enzymes
    enzyme
    sugar
    polymerization
    maize
    biomass
    Sugars
    Polymerization
    Biomass
    Zea mays
    biofuel
    hydrolysis
    Enzyme inhibition
    Biofuels
    Enzymatic hydrolysis
    Oligomers
    Chemical analysis

    Keywords

    • AFEX-CS hydrolysate
    • Charcoal fractionation
    • Commercial enzymes
    • Enzyme inhibition
    • High solids-loading
    • Recalcitrant oligosaccharides
    • Size exclusion chromatography

    ASJC Scopus subject areas

    • Energy(all)
    • Management, Monitoring, Policy and Law
    • Biotechnology
    • Applied Microbiology and Biotechnology
    • Renewable Energy, Sustainability and the Environment

    Cite this

    Sugar loss and enzyme inhibition due to oligosaccharide accumulation during high solids-loading enzymatic hydrolysis. / Xue, Saisi; Uppugundla, Nirmal; Bowman, Michael J.; Cavalier, David; Da Costa Sousa, Leonardo; Dale, Bruce E.; Balan, Venkatesh.

    In: Biotechnology for Biofuels, Vol. 8, No. 1, 195, 26.11.2015.

    Research output: Contribution to journalArticle

    Xue, Saisi; Uppugundla, Nirmal; Bowman, Michael J.; Cavalier, David; Da Costa Sousa, Leonardo; Dale, Bruce E.; Balan, Venkatesh / Sugar loss and enzyme inhibition due to oligosaccharide accumulation during high solids-loading enzymatic hydrolysis.

    In: Biotechnology for Biofuels, Vol. 8, No. 1, 195, 26.11.2015.

    Research output: Contribution to journalArticle

    @article{e82205f376d14062b1dc1fed3356f1dd,
    title = "Sugar loss and enzyme inhibition due to oligosaccharide accumulation during high solids-loading enzymatic hydrolysis",
    abstract = "Background: Accumulation of recalcitrant oligosaccharides during high-solids loading enzymatic hydrolysis of cellulosic biomass reduces biofuel yields and increases processing costs for a cellulosic biorefinery. Recalcitrant oligosaccharides in AFEX-pretreated corn stover hydrolysate accumulate to the extent of about 18-25 % of the total soluble sugars in the hydrolysate and 12-18 % of the total polysaccharides in the inlet biomass (untreated), equivalent to a yield loss of about 7-9 kg of monomeric sugars per 100 kg of inlet dry biomass (untreated). These oligosaccharides represent a yield loss and also inhibit commercial hydrolytic enzymes, with both being serious bottlenecks for economical biofuel production from cellulosic biomass. Very little is understood about the nature of these oligomers and why they are recalcitrant to commercial enzymes. This work presents a robust method for separating recalcitrant oligosaccharides from high solid loading hydrolysate in gramme quantities. Composition analysis, recalcitrance study and enzyme inhibition study were performed to understand their chemical nature. Results: Oligosaccharide accumulation occurs during high solid loading enzymatic hydrolysis of corn stover (CS) irrespective of using different pretreated corn stover (dilute acid: DA, ionic liquids: IL, and ammonia fibre expansion: AFEX). The methodology for large-scale separation of recalcitrant oligosaccharides from 25 % solids-loading AFEX-corn stover hydrolysate using charcoal fractionation and size exclusion chromatography is reported for the first time. Oligosaccharides with higher degree of polymerization (DP) were recalcitrant towards commercial enzyme mixtures [Ctec2, Htec2 and Multifect pectinase (MP)] compared to lower DP oligosaccharides. Enzyme inhibition studies using processed substrates (Avicel and xylan) showed that low DP oligosaccharides also inhibit commercial enzymes. Addition of monomeric sugars to oligosaccharides increases the inhibitory effects of oligosaccharides on commercial enzymes. Conclusion: The carbohydrate composition of the recalcitrant oligosaccharides, ratios of different DP oligomers and their distribution profiles were determined. Recalcitrance and enzyme inhibition studies help determine whether the commercial enzyme mixtures lack the enzyme activities required to completely de-polymerize the plant cell wall. Such studies clarify the reasons for oligosaccharide accumulation and contribute to strategies by which oligosaccharides can be converted into fermentable sugars and provide higher biofuel yields with less enzyme.",
    keywords = "AFEX-CS hydrolysate, Charcoal fractionation, Commercial enzymes, Enzyme inhibition, High solids-loading, Recalcitrant oligosaccharides, Size exclusion chromatography",
    author = "Saisi Xue and Nirmal Uppugundla and Bowman, {Michael J.} and David Cavalier and {Da Costa Sousa}, Leonardo and Dale, {Bruce E.} and Venkatesh Balan",
    year = "2015",
    month = "11",
    doi = "10.1186/s13068-015-0378-9",
    volume = "8",
    journal = "Biotechnology for Biofuels",
    issn = "1754-6834",
    publisher = "BioMed Central",
    number = "1",

    }

    TY - JOUR

    T1 - Sugar loss and enzyme inhibition due to oligosaccharide accumulation during high solids-loading enzymatic hydrolysis

    AU - Xue,Saisi

    AU - Uppugundla,Nirmal

    AU - Bowman,Michael J.

    AU - Cavalier,David

    AU - Da Costa Sousa,Leonardo

    AU - Dale,Bruce E.

    AU - Balan,Venkatesh

    PY - 2015/11/26

    Y1 - 2015/11/26

    N2 - Background: Accumulation of recalcitrant oligosaccharides during high-solids loading enzymatic hydrolysis of cellulosic biomass reduces biofuel yields and increases processing costs for a cellulosic biorefinery. Recalcitrant oligosaccharides in AFEX-pretreated corn stover hydrolysate accumulate to the extent of about 18-25 % of the total soluble sugars in the hydrolysate and 12-18 % of the total polysaccharides in the inlet biomass (untreated), equivalent to a yield loss of about 7-9 kg of monomeric sugars per 100 kg of inlet dry biomass (untreated). These oligosaccharides represent a yield loss and also inhibit commercial hydrolytic enzymes, with both being serious bottlenecks for economical biofuel production from cellulosic biomass. Very little is understood about the nature of these oligomers and why they are recalcitrant to commercial enzymes. This work presents a robust method for separating recalcitrant oligosaccharides from high solid loading hydrolysate in gramme quantities. Composition analysis, recalcitrance study and enzyme inhibition study were performed to understand their chemical nature. Results: Oligosaccharide accumulation occurs during high solid loading enzymatic hydrolysis of corn stover (CS) irrespective of using different pretreated corn stover (dilute acid: DA, ionic liquids: IL, and ammonia fibre expansion: AFEX). The methodology for large-scale separation of recalcitrant oligosaccharides from 25 % solids-loading AFEX-corn stover hydrolysate using charcoal fractionation and size exclusion chromatography is reported for the first time. Oligosaccharides with higher degree of polymerization (DP) were recalcitrant towards commercial enzyme mixtures [Ctec2, Htec2 and Multifect pectinase (MP)] compared to lower DP oligosaccharides. Enzyme inhibition studies using processed substrates (Avicel and xylan) showed that low DP oligosaccharides also inhibit commercial enzymes. Addition of monomeric sugars to oligosaccharides increases the inhibitory effects of oligosaccharides on commercial enzymes. Conclusion: The carbohydrate composition of the recalcitrant oligosaccharides, ratios of different DP oligomers and their distribution profiles were determined. Recalcitrance and enzyme inhibition studies help determine whether the commercial enzyme mixtures lack the enzyme activities required to completely de-polymerize the plant cell wall. Such studies clarify the reasons for oligosaccharide accumulation and contribute to strategies by which oligosaccharides can be converted into fermentable sugars and provide higher biofuel yields with less enzyme.

    AB - Background: Accumulation of recalcitrant oligosaccharides during high-solids loading enzymatic hydrolysis of cellulosic biomass reduces biofuel yields and increases processing costs for a cellulosic biorefinery. Recalcitrant oligosaccharides in AFEX-pretreated corn stover hydrolysate accumulate to the extent of about 18-25 % of the total soluble sugars in the hydrolysate and 12-18 % of the total polysaccharides in the inlet biomass (untreated), equivalent to a yield loss of about 7-9 kg of monomeric sugars per 100 kg of inlet dry biomass (untreated). These oligosaccharides represent a yield loss and also inhibit commercial hydrolytic enzymes, with both being serious bottlenecks for economical biofuel production from cellulosic biomass. Very little is understood about the nature of these oligomers and why they are recalcitrant to commercial enzymes. This work presents a robust method for separating recalcitrant oligosaccharides from high solid loading hydrolysate in gramme quantities. Composition analysis, recalcitrance study and enzyme inhibition study were performed to understand their chemical nature. Results: Oligosaccharide accumulation occurs during high solid loading enzymatic hydrolysis of corn stover (CS) irrespective of using different pretreated corn stover (dilute acid: DA, ionic liquids: IL, and ammonia fibre expansion: AFEX). The methodology for large-scale separation of recalcitrant oligosaccharides from 25 % solids-loading AFEX-corn stover hydrolysate using charcoal fractionation and size exclusion chromatography is reported for the first time. Oligosaccharides with higher degree of polymerization (DP) were recalcitrant towards commercial enzyme mixtures [Ctec2, Htec2 and Multifect pectinase (MP)] compared to lower DP oligosaccharides. Enzyme inhibition studies using processed substrates (Avicel and xylan) showed that low DP oligosaccharides also inhibit commercial enzymes. Addition of monomeric sugars to oligosaccharides increases the inhibitory effects of oligosaccharides on commercial enzymes. Conclusion: The carbohydrate composition of the recalcitrant oligosaccharides, ratios of different DP oligomers and their distribution profiles were determined. Recalcitrance and enzyme inhibition studies help determine whether the commercial enzyme mixtures lack the enzyme activities required to completely de-polymerize the plant cell wall. Such studies clarify the reasons for oligosaccharide accumulation and contribute to strategies by which oligosaccharides can be converted into fermentable sugars and provide higher biofuel yields with less enzyme.

    KW - AFEX-CS hydrolysate

    KW - Charcoal fractionation

    KW - Commercial enzymes

    KW - Enzyme inhibition

    KW - High solids-loading

    KW - Recalcitrant oligosaccharides

    KW - Size exclusion chromatography

    UR - http://www.scopus.com/inward/record.url?scp=84948414044&partnerID=8YFLogxK

    UR - http://www.scopus.com/inward/citedby.url?scp=84948414044&partnerID=8YFLogxK

    U2 - 10.1186/s13068-015-0378-9

    DO - 10.1186/s13068-015-0378-9

    M3 - Article

    VL - 8

    JO - Biotechnology for Biofuels

    T2 - Biotechnology for Biofuels

    JF - Biotechnology for Biofuels

    SN - 1754-6834

    IS - 1

    M1 - 195

    ER -