Xianzhi Meng

Recent advances in understanding the role of cellulose accessibility in enzymatic hydrolysis of lignocellulosic substrates.

Cellulose accessibility has been proposed as a key factor in the efficient bio-conversion of lignocellulosic biomass to fermentable sugars. Factors affecting cellulose accessibility can be divided into direct factors that refer to accessible surface area of cellulose, and indirect factors referring to chemical composition such as lignin/hemicellulose content, and biomass structure-relevant factors (i.e. particle size, porosity). An overview of the current pretreatment technologies special focus on the major mode of action to increase cellulose accessibility as well as multiple techniques that could be used to assess the cellulose accessibility are presented in this review. The appropriate determination of cellulose accessibility before and after pretreatment can assist to understand the effectiveness of a particular pretreatment in overcoming lignocellulosic recalcitrance to improve substrate enzymatic digestibility.

Effect of lignin content on changes occurring in poplar cellulose ultrastructure during dilute acid pretreatment

BackgroundObtaining a better understanding of the complex mechanisms occurring during lignocellulosic deconstruction is critical to the continued growth of renewable biofuel production. A key step in bioethanol production is thermochemical pretreatment to reduce plant cell wall recalcitrance for downstream processes. Previous studies of dilute acid pretreatment (DAP) have shown significant changes in cellulose ultrastructure that occur during pretreatment, but there is still a substantial knowledge gap with respect to the influence of lignin on these cellulose ultrastructural changes. This study was designed to assess how the presence of lignin influences DAP-induced changes in cellulose ultrastructure, which might ultimately have large implications with respect to enzymatic deconstruction efforts.ResultsNative, untreated hybrid poplar (Populus trichocarpa x Populus deltoids) samples and a partially delignified poplar sample (facilitated by acidic sodium chlorite pulping) were separately pretreated with dilute sulfuric acid (0.10 M) at 160°C for 15 minutes and 35 minutes, respectively . Following extensive characterization, the partially delignified biomass displayed more significant changes in cellulose ultrastructure following DAP than the native untreated biomass. With respect to the native untreated poplar, delignified poplar after DAP (in which approximately 40% lignin removal occurred) experienced: increased cellulose accessibility indicated by increased Simons’ stain (orange dye) adsorption from 21.8 to 72.5 mg/g, decreased cellulose weight-average degree of polymerization (DPw) from 3087 to 294 units, and increased cellulose crystallite size from 2.9 to 4.2 nm. These changes following DAP ultimately increased enzymatic sugar yield from 10 to 80%.ConclusionsOverall, the results indicate a strong influence of lignin content on cellulose ultrastructural changes occurring during DAP. With the reduction of lignin content during DAP, the enlargement of cellulose microfibril dimensions and crystallite size becomes more apparent. Further, this enlargement of cellulose microfibril dimensions is attributed to specific processes, including the co-crystallization of crystalline cellulose driven by irreversible inter-chain hydrogen bonding (similar to hornification) and/or cellulose annealing that converts amorphous cellulose to paracrystalline and crystalline cellulose. Essentially, lignin acts as a barrier to prevent cellulose crystallinity increase and cellulose fibril coalescence during DAP.

The Effect of Alkaline Pretreatment Methods on Cellulose Structure and Accessibility

The effects of different alkaline pretreatments on cellulose structural features and accessibility are compared and correlated with the enzymatic hydrolysis of Populus. The pretreatments are shown to modify polysaccharides and lignin content to enhance the accessibility for cellulase enzymes. The highest increase in the cellulose accessibility was observed in dilute sodium hydroxide, followed by methods using ammonia soaking and lime (Ca(OH)2 ). The biggest increase of cellulose accessibility occurs during the first 10 min of pretreatment, with further increases at a slower rate as severity increases. Low temperature ammonia soaking at longer residence times dissolved a major portion of hemicellulose and exhibited higher cellulose accessibility than high temperature soaking. Moreover, the most significant reduction of degree of polymerization (DP) occurred for dilute sodium hydroxide (NaOH) and ammonia pretreated Populus samples. The study thus identifies important cellulose structural features and relevant parameters related to biomass recalcitrance.

Insights into the effect of dilute acid, hot water or alkaline pretreatment on the cellulose accessible surface area and the overall porosity of Populus

Pretreatment is known to make biomass more reactive to cellulase by altering the chemical compositions as well as physical structures of biomass. Simons’ staining technique along with mercury porosimetry was applied on the acid, neutral, and alkaline pretreated materials to measure the accessible surface area of cellulose and pore size distribution of Populus. The results indicated that acid pretreatment is much more effective than water and alkaline pretreatment in terms of cellulose accessibility increase. Further investigation suggests that lignin does not dictate cellulose accessibility to the extent that hemicellulose does, but it does restrict xylan accessibility which in turn controls the access of cellulase to cellulose. The most interesting finding is that severe acid pretreatment significantly decreases the average pore size, i.e. 90% average size decrease could be observed after 60 min dilute acid pretreatment at 160 °C; however, the nano-pore space formed between the coated microfibrils increased after pretreatment, especially with the acid pretreatment, suggesting that this particular type of biomass porosity is probably the most fundamental barrier to effective enzymatic hydrolysis.

Thermo-responsive and fluorescent cellulose nanocrystals grafted with polymer brushes

Cellulose nanocrystals (CNCs) grafted with fluorescent and thermo-responsive poly(N-isopropylacrylamide) (PNIPAAM) brushes were prepared via surface-initiated activators generated by electron transfer for atom transfer radical polymerization (SI-AGET-ATRP) in the CH3OH–H2O mixing solvent with different volume ratios. The successful grafting was supported by Fourier transform infrared (FTIR) and nuclear magnetic resonance (NMR) measurements. Gravimetric analysis plus 1H NMR and gel permeation chromatography (GPC) measurements showed that there was an increase in the monomer conversion and molecular weight of polymer brushes with increasing H2O proportion of the solvent system. The variation trend of graft length was further evidenced by the gradual change of decomposition and glass transition temperatures of the surface-grafted CNCs. A large scale of chain transfer occurred on the surface of CNCs in view of the minute quantity of free polymers generated by a sacrificial initiator. Free polymers cannot be used as a substitute to characterize grafted polymers in terms of the big difference between their molecular weights. The obtained surface-grafted CNCs showed thermo-enhanced fluorescence owing to the thermal-driven chain dehydration of the grafted PNIPAAM brushes.

Characterization of products from hydrothermal carbonization of pine

This study aims to reveal the structural features and reaction pathways for solid-liquid products from hydrothermal carbonization of Loblolly pine, where the solid products can be used as catalysts, adsorbents and electrode materials while liquid products can be treated yielding fuels and platform chemicals. Results revealed when treated at 240°C, cellulose and hemicellulose were degraded, in part, to 5-hydroxy-methyl furfural and furfural which were further transformed to aromatic structures via ring opening and Diels Alder reactions. Lignin degradation and formation of carbon-carbon bonds, forming aromatic motifs in the presence of furanic compounds connected via aliphatic bridges, ether or condensation reactions. After hydrothermal treatment, condensed aromatic carbon materials with methoxy groups were recovered with high fixed carbon content and HHV. The recovered liquid products are lignin-like value-added chemicals consisting of furfural and polyaromatic structure with alkanes and carboxyl, their total hydroxyl group content decreased when increasing reaction time.

Conversion of corn stover alkaline pre-treatment waste streams into biodiesel via Rhodococci

The bioconversion of second-generation cellulosic ethanol waste streams into biodiesel via oleaginous bacteria is a novel optimization strategy for biorefineries with substantial potential for rapid development. In this study, one- and two-stage alkali/alkali-peroxide pretreatment waste streams of corn stover were separately implemented as feedstocks in 96 h batch reactor fermentations with wild-type Rhodococcus opacus PD 630, R. opacus DSM 1069, and R. jostii DSM 44719T. Here we show using 31P-NMR, HPAEC-PAD, and SEC analyses, that the more rigorous and chemically-efficient two-stage chemical pretreatment effluent provided higher concentrations of solubilized glucose and lower molecular weight (∼70–300 g mol−1) lignin degradation products thereby enabling improved cellular density, viability, and oleaginicity in each respective strain. The most significant yields were by R. opacus PD 630, which converted 6.2% of organic content with a maximal total lipid production of 1.3 g L−1 and accumulated 42.1% in oils based on cell dry weight after 48 h.

Effects of organosolv and ammonia pretreatments on lignin properties and its inhibition for enzymatic hydrolysis

Lignin offers structural support and protection for plant cell walls; however, it also contributes to biomass recalcitrance and the costs of biofuel production via the biological pathway. Organosolv and ammonia pretreatments have been developed to reduce biomass recalcitrance and improve sugar release performance during enzymatic hydrolysis. It is believed that lignin properties are related to its inhibition on enzymatic hydrolysis; therefore, understanding the characteristics of lignin is a key for effective biomass conversion to biofuels. In this study, an organosolv pretreatment using 60% ethanol with 1.25% H2SO4 significantly deconstructed poplar lignin and reduced its molecular weights due to the cleavage of lignin inter-unit linkages. The organosolv pretreatment increased the contents of phenolic OH units and the lignin residue showed a high cellulase maximum adsorption capacity. Ammonia pretreatment with 5% ammonium hydroxide was not as effective as organosolv pretreatment on lignin deconstruction. Organosolv lignin residue had lower lignin S/G ratio than the untreated one. Compared to the organosolv lignin residue and untreated lignin, ammonia lignin residue had a higher cellulase adsorption affinity. In addition, the effects of lignin on cellulose hydrolysis was investigated and the results suggested that the presence of lignin with cellulose substrates reduced cellulose hydrolysis, and its inhibitory effect was primarily determined by the lignin properties after each pretreatment. The organosolv pretreatment resulted in a slightly lower cellulase binding strength (249.7 mL g−1) on poplar lignin than that on untreated samples (261.1 mL g−1), while ammonia lignin residue showed a higher cellulase binding strength (402.8 mL g−1) and had more significant inhibition effect on cellulose hydrolysis. These results demonstrated that the binding strength significantly affected the lignin-derived inhibition on enzymatic hydrolysis of cellulose in the cellulose-lignin mixtures.

How chip size impacts steam pretreatment effectiveness for biological conversion of poplar wood into fermentable sugars

BackgroundWoody biomass is highly recalcitrant to enzymatic sugar release and often requires significant size reduction and severe pretreatments to achieve economically viable sugar yields in biological production of sustainable fuels and chemicals. However, because mechanical size reduction of woody biomass can consume significant amounts of energy, it is desirable to minimize size reduction and instead pretreat larger wood chips prior to biological conversion. To date, however, most laboratory research has been performed on materials that are significantly smaller than applicable in a commercial setting. As a result, there is a limited understanding of the effects that larger biomass particle size has on the effectiveness of steam explosion pretreatment and subsequent enzymatic hydrolysis of wood chips.ResultsTo address these concerns, novel downscaled analysis and high throughput pretreatment and hydrolysis (HTPH) were applied to examine whether differences exist in the composition and digestibility within a single pretreated wood chip due to heterogeneous pretreatment across its thickness. Heat transfer modeling, Simons’ stain testing, magnetic resonance imaging (MRI), and scanning electron microscopy (SEM) were applied to probe the effects of pretreatment within and between pretreated wood samples to shed light on potential causes of variation, pointing to enzyme accessibility (i.e., pore size) distribution being a key factor dictating enzyme digestibility in these samples. Application of these techniques demonstrated that the effectiveness of pretreatment of Populus tremuloides can vary substantially over the chip thickness at short pretreatment times, resulting in spatial digestibility effects and overall lower sugar yields in subsequent enzymatic hydrolysis.ConclusionsThese results indicate that rapid decompression pretreatments (e.g., steam explosion) that specifically alter accessibility at lower temperature conditions are well suited for larger wood chips due to the non-uniformity in temperature and digestibility profiles that can result from high temperature and short pretreatment times. Furthermore, this study also demonstrated that wood chips were hydrated primarily through the natural pore structure during pretreatment, suggesting that preserving the natural grain and transport systems in wood during storage and chipping processes could likely promote pretreatment efficacy and uniformity.

Insights of biomass recalcitrance in natural Populus trichocarpa variants for biomass conversion

Populus has been investigated as a promising biomass feedstock for alternative fuels and chemicals. Physicochemical characteristics and genomic information of biomass feedstocks are among the essential information that can help not only advance our understanding of biomass recalcitrance but also in its efficient utilization. Herein, the recalcitrance of natural Populus variants was elucidated in three aspects: (1) sugar release, (2) physicochemical properties, and (3) relative variation of gene expression within natural poplar variants. The sugar release performance of natural Populus variants was evaluated with their correlation with biomass recalcitrance-related characteristics. Among the physicochemical properties of poplar, the lignin content, lignin molecular weight, lignin S/G ratio, and cellulose accessibility were found to correlate with sugar release. The results demonstrated that the lignin content was negatively correlated with sugar release, whereas the lignin molecular weight, lignin S/G ratio, and cellulose accessibility were positively associated with poplar sugar release. The trend of differential gene expression of each variant also supports the characterization results and their effects on biomass conversion.