title: Succinic acid production on xylose-enriched biorefinery streams by Actinobacillus succinogenes in batch fermentation.
abstract: BACKGROUND
  Co-production of chemicals from lignocellulosic biomass alongside fuels holds promise for improving the economic outlook of integrated biorefineries. In current biochemical conversion processes that use thermochemical pretreatment and enzymatic hydrolysis, fractionation of hemicellulose-derived and cellulose-derived sugar streams is possible using hydrothermal or dilute acid pretreatment (DAP), which then offers a route to parallel trains for fuel and chemical production from xylose- and glucose-enriched streams. Succinic acid (SA) is a co-product of particular interest in biorefineries because it could potentially displace petroleum-derived chemicals and polymer precursors for myriad applications. However, SA production from biomass-derived hydrolysates has not yet been fully explored or developed.
  
  
  RESULTS
  Here, we employ Actinobacillus succinogenes 130Z to produce succinate in batch fermentations from various substrates including (1) pure sugars to quantify substrate inhibition, (2) from mock hydrolysates similar to those from DAP containing single putative inhibitors, and (3) using the hydrolysate derived from two pilot-scale pretreatments: first, a mild alkaline wash (deacetylation) followed by DAP, and secondly a single DAP step, both with corn stover. These latter streams are both rich in xylose and contain different levels of inhibitors such as acetate, sugar dehydration products (furfural, 5-hydroxymethylfurfural), and lignin-derived products (ferulate, p-coumarate). In batch fermentations, we quantify succinate and co-product (acetate and formate) titers as well as succinate yields and productivities. We demonstrate yields of 0.74 g succinate/g sugars and 42.8 g/L succinate from deacetylated DAP hydrolysate, achieving maximum productivities of up to 1.27 g/L-h. Moreover, A. succinogenes is shown to detoxify furfural via reduction to furfuryl alcohol, although an initial lag in succinate production is observed when furans are present. Acetate seems to be the main inhibitor for this bacterium present in biomass hydrolysates.
  
  
  CONCLUSIONS
  Overall, these results demonstrate that biomass-derived, xylose-enriched hydrolysates result in similar yields and titers but lower productivities compared to clean sugar streams, which can likely be improved via fermentation process developments and metabolic engineering. Overall, this study comprehensively examines the behavior of A. succinogenes on xylose-enriched hydrolysates on an industrially relevant, lignocellulosic feedstock, which will pave the way for future work toward eventual SA production in an integrated biorefinery.
authors.affiliation: National Bioenergy Center, National Renewable Energy Laboratory, Golden, CO 80401 USA.
authors.forename: Davinia
authors.initials: D
authors.lastname: Salvachúa
authors.affiliation: National Bioenergy Center, National Renewable Energy Laboratory, Golden, CO 80401 USA.
authors.forename: Ali
authors.initials: A
authors.lastname: Mohagheghi
authors.affiliation: National Bioenergy Center, National Renewable Energy Laboratory, Golden, CO 80401 USA.
authors.forename: Holly
authors.initials: H
authors.lastname: Smith
authors.affiliation: Department of Chemical Engineering, University of Pretoria, Pretoria, South Africa.
authors.forename: Michael F A
authors.initials: MFA
authors.lastname: Bradfield
authors.affiliation: Department of Chemical Engineering, University of Pretoria, Pretoria, South Africa.
authors.forename: Willie
authors.initials: W
authors.lastname: Nicol
authors.affiliation: National Bioenergy Center, National Renewable Energy Laboratory, Golden, CO 80401 USA.
authors.forename: Brenna A
authors.initials: BA
authors.lastname: Black
authors.affiliation: National Bioenergy Center, National Renewable Energy Laboratory, Golden, CO 80401 USA.
authors.forename: Mary J
authors.initials: MJ
authors.lastname: Biddy
authors.affiliation: National Bioenergy Center, National Renewable Energy Laboratory, Golden, CO 80401 USA.
authors.forename: Nancy
authors.initials: N
authors.lastname: Dowe
authors.affiliation: National Bioenergy Center, National Renewable Energy Laboratory, Golden, CO 80401 USA.
authors.forename: Gregg T
authors.initials: GT
authors.lastname: Beckham
keywords: Actinobacillus succinogenes; Biochemicals; Biofuels; Biorefinery; Fermentation; Pretreatment; Succinic acid
references.citation: Microb Cell Fact. 2013 Sep 17;12:80
references.pmid: 24044876
references.citation: Appl Biochem Biotechnol. 2003 Spring;105 -108:69-85
references.pmid: 12721476
references.citation: Chem Rev. 2007 Jun;107(6):2411-502
references.pmid: 17535020
references.citation: Appl Biochem Biotechnol. 2002 Spring;98-100:327-40
references.pmid: 12018260
references.citation: Bioresour Technol. 2009 Apr;100(8):2425-9
references.pmid: 19128958
references.citation: Biotechnol Biofuels. 2014 Apr 05;7(1):51
references.pmid: 24708699
references.citation: J Microbiol Biotechnol. 2009 Feb;19(2):167-71
references.pmid: 19307766
references.citation: Appl Biochem Biotechnol. 2010 Jan;160(2):477-85
references.pmid: 18830824
references.citation: Annu Rev Chem Biomol Eng. 2011;2:121-45
references.pmid: 22432613
references.citation: J Biosci Bioeng. 1999;87(2):169-74
references.pmid: 16232445
references.citation: Biotechnol Biofuels. 2012 Feb 27;5:8
references.pmid: 22369467
references.citation: Bioprocess Biosyst Eng. 2003 Nov;26(1):63-7
references.pmid: 14530958
references.citation: Int J Syst Bacteriol. 1999 Jan;49 Pt 1:207-16
references.pmid: 10028265
references.citation: Bioresour Technol. 2002 Jan;81(1):33-44
references.pmid: 11708754
references.citation: Bioprocess Biosyst Eng. 2013 Nov;36(11):1779-85
references.pmid: 23649828
references.citation: Bioresour Technol. 2013 May;135:574-7
references.pmid: 23010211
references.citation: BMC Genomics. 2010 Nov 30;11:680
references.pmid: 21118570
references.citation: Appl Environ Microbiol. 2002 Apr;68(4):1715-27
references.pmid: 11916689
references.citation: Biotechnol Biofuels. 2013 Jul 09;6(1):99
references.pmid: 23837621
references.citation: Bioresour Technol. 2010 Jan;101(2):674-8
references.pmid: 19766484
references.citation: J Ind Microbiol Biotechnol. 2010 Oct;37(10):1033-40
references.pmid: 20532948
references.citation: Appl Microbiol Biotechnol. 2007 Sep;76(4):727-40
references.pmid: 17609945
references.citation: FEMS Microbiol Lett. 2004 Jul 15;236(2):163-73
references.pmid: 15251193
references.citation: J Ind Microbiol Biotechnol. 2011 Aug;38(8):1001-11
references.pmid: 20882312
references.citation: Proc Natl Acad Sci U S A. 2014 Aug 19;111(33):12013-8
references.pmid: 25092344
references.citation: J Microbiol Biotechnol. 2009 Nov;19(11):1379-84
references.pmid: 19996691
references.citation: Curr Microbiol. 2010 Jan;60(1):71-7
references.pmid: 19777303
references.citation: Microb Cell Fact. 2014 Aug 19;13:111
references.pmid: 25259880
references.citation: Science. 2014 May 16;344(6185):1246843
references.pmid: 24833396
references.citation: Bioresour Technol. 2015;187:37-42
references.pmid: 25836372
references.citation: Appl Biochem Biotechnol. 2009 May;155(1-3):418-28
references.pmid: 19142588
references.citation: Methods Mol Biol. 2009;581:125-43
references.pmid: 19768621
references.citation: Biotechnol Bioeng. 2010 Oct 15;107(3):430-40
references.pmid: 20552667
references.citation: Appl Microbiol Biotechnol. 2014 Sep;98 (17 ):7379-86
references.pmid: 24816619
references.citation: Bioresour Technol. 2013 Sep;143:405-12
references.pmid: 23819977
references.citation: Bioresour Technol. 2008 Apr;99(6):1736-42
references.pmid: 17532626
references.citation: Metab Eng. 2013 Nov;20:1-8
references.pmid: 23876411
references.citation: Biotechnol Prog. 1999 Oct 1;15(5):777-793
references.pmid: 10514248
references.citation: Bioresour Technol. 2011 May;102(10):6147-52
references.pmid: 21470857
references.citation: Appl Microbiol Biotechnol. 2004 Nov;65(6):664-70
references.pmid: 15205933
references.citation: Biotechnol Biofuels. 2015 Nov 14;8:181
references.pmid: 26581168
references.citation: Appl Environ Microbiol. 2006 Mar;72(3):1939-48
references.pmid: 16517641
references.citation: Curr Opin Biotechnol. 2014 Dec;30:190-7
references.pmid: 25118136
references.citation: Biotechnol Bioeng. 2008 Apr 1;99(5):1140-53
references.pmid: 17972330
references.citation: Chem Soc Rev. 2012 Feb 21;41(4):1538-58
references.pmid: 21909591
references.citation: Bioresour Technol. 2005 Apr;96(6):673-86
references.pmid: 15588770