The primary obstacle to producing renewable fuels from lignocellulosic biomass is a plant's recalcitrance to releasing sugars bound in the cell wall. From a sample set of wood cores representing 1,100 individual undomesticated Populus trichocarpa trees, 47 extreme phenotypes were selected across measured lignin content and ratio of syringyl and guaiacyl units (S/G ratio). This subset was tested for total sugar release through enzymatic hydrolysis alone as well as through combined hot-water pretreatment and enzymatic hydrolysis using a high-throughput screening method. The total amount of glucan and xylan released varied widely among samples, with total sugar yields of up to 92% of the theoretical maximum. A strong negative correlation between sugar release and lignin content was only found for pretreated samples with an S/G ratio < 2.0. For higher S/G ratios, sugar release was generally higher, and the negative influence of lignin was less pronounced. When examined separately, only glucose release was correlated with lignin content and S/G ratio in this manner, whereas xylose release depended on the S/G ratio alone. For enzymatic hydrolysis without pretreatment, sugar release increased significantly with decreasing lignin content below 20%, irrespective of the S/G ratio. Furthermore, certain samples featuring average lignin content and S/G ratios exhibited exceptional sugar release. These facts suggest that factors beyond lignin and S/G ratio influence recalcitrance to sugar release and point to a critical need for deeper understanding of cell-wall structure before plants can be rationally engineered for reduced recalcitrance and efficient biofuels production. L ignocellulosic biomass is the only sustainable resource in terms of cost, availability, and scale that can be converted into liquid fuels to reduce the prevailing role of petroleum in providing energy for the world's transportation needs (1, 2) and to decrease the emissions of fossil CO 2 that damage the world's climate (3). The primary obstacle to producing liquid transportation fuels by bioconversion methods is the release of sugars in high quantities at low costs from recalcitrant lignocellulosic biomass feedstocks (4, 5). Genetic modification of plants to make them less recalcitrant is a promising path to address this challenge on the feedstock side, but the effort would be greatly aided by improving understanding of the fundamental relationship between cell-wall composition and sugar release through pretreatment and enzymatic hydrolysis.In this paper, we focus on the influence of lignin content and the ratio of its syringyl and guaiacyl units (S/G ratio) on recalcitrance to sugar release, because these two traits were previously identified as dominant factors (6). Although it is generally perceived that low lignin contents increase the ability of cellulolytic enzymes to hydrolyze plant fibers (7-11), only a limited number of studies investigated the effect of lignin S/G ratio on sugar release through combined pretreatment and enzymatic hydrolysis. A...

An autohydrolysis pretreatment that suppresses lignin repolymerisation helps overcoming the recalcitrance of softwood for enzymatic hydrolysis of its cellulose.

Lignocellulosic biomass is uniquely suited as a sustainable feedstock for the biotechnological production of alternative fuels and chemicals. However, due to the biomass recalcitrance, the enzymatic conversion process is complex and needs to be simplified. To this end, we developed a process, which allows the consolidated bioprocessing of lignocellulose to ethanol in a single multi-species biofilm membrane reactor featuring both aerobic and anaerobic conditions necessary for the simultaneous fungal cellulolytic enzyme production and alcoholic yeast fermentation of the hydrolysis-derived sugars. The general feasibility of the concept was successfully demonstrated by producing ethanol with a 67% yield from undetoxified whole slurry dilute acid pretreated wheat straw by the combined action of Trichoderma reesei, Saccharomyces cerevisiae and Scheffersomyces stipitis. The results achieved underscore the potential of the process as a versatile cheap sugar platform for the production of fuels and chemicals based on lignocellulosic biomass by specifically compiled consortia of industrially proven robust microorganisms. Broader context Biofuels made from lignocellulosic biomass have the potential to be useful elements in the overall approach to tackle the issues of climate change and sustainable energy supply. However, this potential can only be unlocked if they are cost competitive to petroleum and starch and sucrose based biofuels. For the biochemical conversion route comprising the three main steps of physicochemical pretreatment, enzymatic hydrolysis and fermentation, the following objectives have to be addressed: (i) development of an integrated process without washing and detoxication steps, (ii) reduction of cellulase costs, (iii) improving hexose and pentose sugar co-utilization, and (iv) overcoming the plants' recalcitrance. As an alternative to employing one genetically engineered superior biocatalyst capable of both degrading biomass and producing biofuel for consolidated bioprocessing (CBP), we present in this paper a novel microbial consortium based approach to tackle issues (i) to (iii). We developed a membrane biolm reactor, which, as a unique feature, enables the coexistence of aerobic and anaerobic conditions at the same time. This allows the coexistence of the two common "workhorses" of the cellulosic ethanol industry in one system: the aerobic hydrolytic enzyme producing fungus Trichoderma reesei and the anaerobic ethanol producing yeast Saccharomyces cerevisiae.