Identification of benzoquinones in pretreated lignocellulosic feedstocks and inhibitory effects on yeast

Abstract

Pretreatment of lignocellulosic biomass under acidic conditions gives rise to by-products that inhibit fermenting microorganisms. An analytical procedure for identification of p-benzoquinone (BQ) and 2,6-dimethoxybenzoquinone (DMBQ) in pretreated biomass was developed, and the inhibitory effects of BQ and DMBQ on the yeast Saccharomyces cerevisiae were assessed. The benzoquinones were analyzed using ultra-high performance liquid chromatography-electrospray ionization-triple quadrupole-mass spectrometry after derivatization with 2,4-dinitrophenylhydrazine. Pretreatment liquids examined with regard to the presence of BQ and DMBQ originated from six different lignocellulosic feedstocks covering agricultural residues, hardwood, and softwood, and were produced through impregnation with sulfuric acid or sulfur dioxide at varying pretreatment temperature (165–204 °C) and residence time (6–20 min). BQ was detected in all six pretreatment liquids in concentrations ranging up to 6 mg/l, while DMBQ was detected in four pretreatment liquids in concentrations ranging up to 0.5 mg/l. The result indicates that benzoquinones are ubiquitous as by-products of acid pretreatment of lignocellulose, regardless of feedstock and pretreatment conditions. Fermentation experiments with BQ and DMBQ covered the concentration ranges 2 mg/l to 1 g/l and 20 mg/l to 1 g/l, respectively. Even the lowest BQ concentration tested (2 mg/l) was strongly inhibitory to yeast, while 20 mg/l DMBQ gave a slight negative effect on ethanol formation. This work shows that benzoquinones should be regarded as potent and widespread inhibitors in lignocellulosic hydrolysates, and that they warrant attention besides more well-studied inhibitory substances, such as aliphatic carboxylic acids, phenols, and furan aldehydes.

Mechanism of Tolerance to the Lignin-Derived Inhibitor p -Benzoquinone and Metabolic Modification of Biorefinery Fermentation Strains

“…Quinones were found to strongly inhibit Saccharomyces cerevisiae. Just 0.02 g/liter BQ and 0.2 g/liter DMBQ completely inhibit the cell growth of S. cerevisiae (7,13). BQ shows high toxicity to animal cells by increasing its reactive oxygen species (ROS) level, oxidizing DNA, breaking DNA double strands, and decreasing Ogg1 transcript levels (14)(15)(16).…”

“…BQ shows high toxicity to animal cells by increasing its reactive oxygen species (ROS) level, oxidizing DNA, breaking DNA double strands, and decreasing Ogg1 transcript levels (14)(15)(16). However, compared with the extensive studies on furan aldehydes, weak organic acids, and phenolic aldehydes, only a few studies have reported on microbial inhibition by quinones (7,13,17). Metabolic pathways for the conversion of quinones have not been clearly characterized, and strategies to lessen quinone inhibition in biorefinery fermentation strains are not well developed.…”

Effects of Benzoquinones on Radicles of Orobanche and Phelipanche Species

“…Humic acids can be obtained by the alkaline transformation of p -benzoquinones for a potential application in agriculture [ 23 ]. p -Benzoquinone (BQ) and 2,6-dimethoxy- p -benzoquinone (DMBQ) are by-products in pretreated lignocellulosic biomass, which inhibit microorganism fermentation [ 24 ]. BQ has been used to evaluate the effects of race and age in cattle on the redox condition of their blood plasma [ 25 ].…”

Factors Affecting Detoxification of Softwood Enzymatic Hydrolysates Using Sodium Dithionite

“…Certain groups of by-products that inhibit microorganisms have been extensively studied, and these include aliphatic carboxylic acids (such as acetic acid, formic acid, and levulinic acid), furan aldehydes (such as furfural and 5-hydroxymethylfurfural [HMF]), and phenolic and other phenylic substances [10,13,14]. More recently, small aliphatic aldehydes (such as formaldehyde and acetaldehyde) [15] and benzoquinones (such as p-benzoquinone) [16] were also found to be commonly occurring in pretreated biomass. Martín et al [17] reported that formaldehyde was the most important inhibitor of Saccharomyces cerevisiae yeast in a set of slurries of pretreated Norway spruce.…”