Identifying inhibitory compounds in lignocellulosic biomass hydrolysates using an exometabolomics approach

This study evaluated the individual and combined effects of inhibitory compounds formed during pretreatment of lignocellulosic biomass on the growth of Bacillus subtilis. Ten inhibitory compounds commonly present in lignocellulosic hydrolysates were evaluated, which included sugar degradation products (furfural and 5-hydroxymethylfurfural), acetic acid, and seven phenolic compounds derived from lignin (benzoic acid, vanillin, vanillic acid, ferulic acid, p-coumaric acid, 4-hydroxybenzoic acid, and syringaldehyde). For the individual inhibitors, syringaldehyde showed the most toxic effect, completely inhibiting the strain growth at 0.1 g/L. In the sequence, assays using mixtures of the inhibitory compounds at a concentration of 12.5% of their IC50 value were performed to evaluate the combined effect of the inhibitors on the strain growth. These experiments were planned according to a Plackett–Burman experimental design. Statistical analysis of the results revealed that in a mixture, benzoic acid and furfural were the most potent inhibitors affecting the growth of B. subtilis. These results contribute to a better understanding of the individual and combined effects of inhibitory compounds present in biomass hydrolysates on the microbial performance of B. subtilis. Such knowledge is important to advance the development of sustainable biomanufacturing processes using this strain cultivated in complex media produced from lignocellulosic biomass, supporting the development of efficient bio-based processes using B. subtilis.

Download Full-text

n-Butanol production from lignocellulosic biomass hydrolysates without detoxification by Clostridium tyrobutyricum Δack-adhE2 in a fibrous-bed bioreactor

Bioresource Technology ◽

10.1016/j.biortech.2019.121749 ◽

2019 ◽

Vol 289 ◽

pp. 121749 ◽

Cited By ~ 15

Author(s):

Jing Li ◽

Yinming Du ◽

Teng Bao ◽

Jie Dong ◽

Meng Lin ◽

...

Keyword(s):

Lignocellulosic Biomass ◽

Clostridium Tyrobutyricum ◽

Butanol Production ◽

Fibrous Bed Bioreactor ◽

Biomass Hydrolysates

Download Full-text

Butyric acid production from lignocellulosic biomass hydrolysates by engineered Clostridium tyrobutyricum overexpressing xylose catabolism genes for glucose and xylose co-utilization

Bioresource Technology ◽

10.1016/j.biortech.2017.03.073 ◽

2017 ◽

Vol 234 ◽

pp. 389-396 ◽

Cited By ~ 40

Author(s):

Hongxin Fu ◽

Shang-Tian Yang ◽

Minqi Wang ◽

Jufang Wang ◽

I-Ching Tang

Keyword(s):

Lignocellulosic Biomass ◽

Butyric Acid ◽

Acid Production ◽

Clostridium Tyrobutyricum ◽

Biomass Hydrolysates ◽

Butyric Acid Production ◽

Xylose Catabolism

Download Full-text

Engineering the Oleaginous Yeast Rhodosporidium toruloides for Improved Resistance Against Inhibitors in Biomass Hydrolysates

Frontiers in Bioengineering and Biotechnology ◽

10.3389/fbioe.2021.768934 ◽

2021 ◽

Vol 9 ◽

Author(s):

Liting Lyu ◽

Yadong Chu ◽

Sufang Zhang ◽

Yue Zhang ◽

Qitian Huang ◽

...

Keyword(s):

Lignocellulosic Biomass ◽

Oleaginous Yeast ◽

Lipid Production ◽

Cell Mass ◽

Peroxidase Gene ◽

The Past ◽

Yeast Strains ◽

Biomass Hydrolysates ◽

Improved Performance ◽

Inhibitor Resistance

Conversion of lignocellulosic biomass into lipids and related chemicals has attracted much attention in the past two decades, and the oleaginous yeast Rhodosporidiumtoruloides has been widely used in this area. While R. toruloides species naturally have physiological advantages in terms of substrate utilization, lipid accumulation, and inhibitor resistance, reduced lipid production and cell growth are noticed when biomass hydrolysates are used as feedstocks. To improve the robustness of R. toruloides, here, we devised engineered strains by overexpressing genes responsible for phenolic compound degradation. Specifically, gene expression cassettes of the manganese peroxidase gene (MNP) and versatile peroxidase gene (VP) were constructed and integrated into the genome of R. toruloides NP11. A series of engineered strains were evaluated for lipid production in the presence of typical phenolic inhibitors. The results showed that R. toruloides strains with proper expression of MNP or VP indeed grew faster in the presence of vanillin and 5-hydroxymethylfurfural than the parental strain. When cultivated in concentrated mode biomass hydrolysates, the strain VP18 had improved performance as the cell mass and lipid content increased by 30% and 25%, respectively. This study provides more robust oleaginous yeast strains for microbial lipid production from lignocellulosic biomass, and similar efforts may be used to devise more advanced lipid producers.

Download Full-text

Does Acid Addition Improve Liquid Hot Water Pretreatment of Lignocellulosic Biomass towards Biohydrogen and Biogas Production?

Sustainability ◽

10.3390/su12218935 ◽

2020 ◽

Vol 12 (21) ◽

pp. 8935 ◽

Cited By ~ 1

Author(s):

George Dimitrellos ◽

Gerasimos Lyberatos ◽

Georgia Antonopoulou

Keyword(s):

Lignocellulosic Biomass ◽

Biogas Production ◽

Hot Water ◽

Liquid Hot Water ◽

Acid Addition ◽

Water Pretreatment ◽

Inhibitory Compounds ◽

Production Of Hydrogen ◽

Potential Inhibitors ◽

Hot Water Pretreatment

The effect of liquid hot water (LHW) pretreatment with or without acid addition (A-LHW) on the production of hydrogen—through dark fermentation (DF)—and methane—through anaerobic digestion (AD)—using three different lignocellulosic biomass types (sunflower straw (SS), grass lawn (GL), and poplar sawdust (PS)) was investigated. Both pretreatment methods led to hemicellulose degradation, but A-LHW resulted in the release of more potential inhibitors (furans and acids) than the LHW pretreatment. Biological hydrogen production (BHP) of the cellulose-rich solid fractions obtained after LHW and A-LHW pretreatment was enhanced compared to the untreated substrates. Due to the release of inhibitory compounds, LHW pretreatment led to higher biochemical methane potential (BMP) than A-LHW pretreatment when both separated fractions (liquid and solid) obtained after pretreatments were used for AD. The recovered energy in the form of methane with LHW pretreatment was 8.4, 12.5, and 7.5 MJ/kg total solids (TS) for SS, GL, and PS, respectively.

Download Full-text

A novel AST2 mutation generated upon whole-genome transformation of Saccharomyces cerevisiae confers high tolerance to 5-Hydroxymethylfurfural (HMF) and other inhibitors

PLoS Genetics ◽

10.1371/journal.pgen.1009826 ◽

2021 ◽

Vol 17 (10) ◽

pp. e1009826

Author(s):

Gert Vanmarcke ◽

Quinten Deparis ◽

Ward Vanthienen ◽

Arne Peetermans ◽

Maria R. Foulquié-Moreno ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Lignocellulosic Biomass ◽

Causative Mutation ◽

Small Scale ◽

Whole Genome ◽

Biomass Hydrolysis ◽

Inhibitor Tolerance ◽

Cell Factories ◽

Biomass Hydrolysates

Development of cell factories for conversion of lignocellulosic biomass hydrolysates into biofuels or bio-based chemicals faces major challenges, including the presence of inhibitory chemicals derived from biomass hydrolysis or pretreatment. Extensive screening of 2526 Saccharomyces cerevisiae strains and 17 non-conventional yeast species identified a Candida glabrata strain as the most 5-hydroxymethylfurfural (HMF) tolerant. Whole-genome (WG) transformation of the second-generation industrial S. cerevisiae strain MD4 with genomic DNA from C. glabrata, but not from non-tolerant strains, allowed selection of stable transformants in the presence of HMF. Transformant GVM0 showed the highest HMF tolerance for growth on plates and in small-scale fermentations. Comparison of the WG sequence of MD4 and GVM1, a diploid segregant of GVM0 with similarly high HMF tolerance, surprisingly revealed only nine non-synonymous SNPs, of which none were present in the C. glabrata genome. Reciprocal hemizygosity analysis in diploid strain GVM1 revealed AST2N406I as the only causative mutation. This novel SNP improved tolerance to HMF, furfural and other inhibitors, when introduced in different yeast genetic backgrounds and both in synthetic media and lignocellulose hydrolysates. It stimulated disappearance of HMF and furfural from the medium and enhanced in vitro furfural NADH-dependent reducing activity. The corresponding mutation present in AST1 (i.e. AST1D405I) the paralog gene of AST2, also improved inhibitor tolerance but only in combination with AST2N406I and in presence of high inhibitor concentrations. Our work provides a powerful genetic tool to improve yeast inhibitor tolerance in lignocellulosic biomass hydrolysates and other inhibitor-rich industrial media, and it has revealed for the first time a clear function for Ast2 and Ast1 in inhibitor tolerance.

Download Full-text