Nicole Labbé

Comparative genome analysis of lignin biosynthesis gene families across the plant kingdom.

BackgroundAs a major component of plant cell wall, lignin plays important roles in mechanical support, water transport, and stress responses. As the main cause for the recalcitrance of plant cell wall, lignin modification has been a major task for bioenergy feedstock improvement. The study of the evolution and function of lignin biosynthesis genes thus has two-fold implications. First, the lignin biosynthesis pathway provides an excellent model to study the coordinative evolution of a biochemical pathway in plants. Second, understanding the function and evolution of lignin biosynthesis genes will guide us to develop better strategies for bioenergy feedstock improvement.ResultsWe analyzed lignin biosynthesis genes from fourteen plant species and one symbiotic fungal species. Comprehensive comparative genome analysis was carried out to study the distribution, relatedness, and family expansion of the lignin biosynthesis genes across the plant kingdom. In addition, we also analyzed the comparative synteny map between rice and sorghum to study the evolution of lignin biosynthesis genes within the Poaceae family and the chromosome evolution between the two species. Comprehensive lignin biosynthesis gene expression analysis was performed in rice, poplar and Arabidopsis. The representative data from rice indicates that different fates of gene duplications exist for lignin biosynthesis genes. In addition, we also carried out the biomass composition analysis of nine Arabidopsis mutants with both MBMS analysis and traditional wet chemistry methods. The results were analyzed together with the genomics analysis.ConclusionThe research revealed that, among the species analyzed, the complete lignin biosynthesis pathway first appeared in moss; the pathway is absent in green algae. The expansion of lignin biosynthesis gene families correlates with substrate diversity. In addition, we found that the expansion of the gene families mostly occurred after the divergence of monocots and dicots, with the exception of the C4H gene family. Gene expression analysis revealed different fates of gene duplications, largely confirming plants are tolerant to gene dosage effects. The rapid expansion of lignin biosynthesis genes indicated that the translation of transgenic lignin modification strategies from model species to bioenergy feedstock might only be successful between the closely relevant species within the same family.

Enhancing the combustible properties of bamboo by torrefaction

Bamboo has wide range of moisture content, low bulk energy density and is difficult to transport, handle, store and feed into existing combustion and gasification systems. Because of its important fuel characteristics such as low ash content, alkali index and heating value, bamboo is a promising energy crop for the future. The aim of this study was to evaluate the effects of torrefaction on the main energy properties of Bambusa vulgaris. Three different torrefaction temperatures were employed: 220, 250 and 280°C. The elemental characteristics of lignite and coal were compared to the torrefied bamboo. The characteristics of the biomass fuels tend toward those of low rank coals. Principal component analysis of FTIR data showed a clear separation between the samples by thermal treatment. The loadings plot indicated that the bamboo samples underwent chemical changes related to carbonyl groups, mostly present in hemicelluloses, and to aromatic groups present in lignin.

FT-IR imaging and pyrolysis-molecular beam mass spectrometry: new tools to investigate wood tissues

Fourier transform infrared (FT-IR) microimaging spectroscopy and pyrolysis molecular beam mass spectrometry (py-MBMS) were used as rapid analysis tools to evaluate differences in the chemical composition of 1-year-old transgenic aspens. Multivariate analysis of the spectroscopic data sets was used to compare the cell wall composition of nontransformed control to transgenic aspen plants with GRP-iaaM gene and with GRP-iaaM/35S-ACCase gene. Principal component analysis (PCA) was applied to both the FT-IR and py-MBMS spectra, which revealed sample groupings due to differences in chemical composition. Evaluating the PCA loadings allows determination of the chemical features responsible for the groupings. The FT-IR microimaging data was also used to monitor changes in the chemical composition as a function of the distance from the pith to the bark using partial least squares (PLS) analysis. The analysis shows that the changes in the composition of the xylem that occur over one annual growth ring can be monitored with FT-IR microimaging.

Compatible Ionic liquid‐cellulases system for hydrolysis of lignocellulosic biomass

Ionic liquids (ILs) have been increasingly recognized as novel solvents for dissolution and pretreatment of cellulose. However, cellulases are inactivated in the presence of ILs, even when present at low concentrations. To more fully exploit the benefits of ILs it is critical to develop a compatible IL‐cellulases system in which the IL is able to effectively solubilize and activate the lignocellulosic biomass, and the cellulases possess high stability and activity. In this study, we investigated the stability and activity of a commercially available cellulases mixture in the presence of different concentrations of 1‐ethyl‐3‐methylimidazolium acetate ([Emim][OAc]). A mixture of cellulases and β‐glucosidase (Celluclast1.5L, from Trichoderma reesei, and Novozyme188, from Aspergillus niger, respectively) retained 77% and 65% of its original activity after being pre‐incubated in 15% and 20% (w/v) IL solutions, respectively, at 50°C for 3 h. The cellulases mixture also retained high activity in 15% [Emim][OAc] to hydrolyze Avicel, a model substrate for cellulose analysis, with conversion efficiency of approximately 91%. Notably, the presence of different amounts of yellow poplar lignin did not interfere significantly with the enzymatic hydrolysis of Avicel. Using this IL‐cellulase system (15% [Emim][OAc]), the saccharification of yellow poplar biomass was also significantly improved (33%) compared to the untreated control (3%) during the first hour of enzymatic hydrolysis. Together, these findings provide compelling evidence that [Emim][OAc] was compatible with the cellulase mixture, and this compatible IL‐cellulases system is promising for efficient activation and hydrolysis of native biomass to produce biofuels and co‐products from the individual biomass components. Bioeng. 2011; 108:1042–1048.

Two-year field analysis of reduced recalcitrance transgenic switchgrass.

Switchgrass (Panicum virgatum L.) is a leading candidate for a dedicated lignocellulosic biofuel feedstock owing to its high biomass production, wide adaptation and low agronomic input requirements. Lignin in cell walls of switchgrass, and other lignocellulosic feedstocks, severely limits the accessibility of cell wall carbohydrates to enzymatic breakdown into fermentable sugars and subsequently biofuels. Low-lignin transgenic switchgrass plants produced by the down-regulation of caffeic acid O-methyltransferase (COMT), a lignin biosynthetic enzyme, were analysed in the field for two growing seasons. COMT transcript abundance, lignin content and the syringyl/guaiacyl lignin monomer ratio were consistently lower in the COMT-down-regulated plants throughout the duration of the field trial. In general, analyses with fully established plants harvested during the second growing season produced results that were similar to those observed in previous greenhouse studies with these plants. Sugar release was improved by up to 34% and ethanol yield by up to 28% in the transgenic lines relative to controls. Additionally, these results were obtained using senesced plant material harvested at the end of the growing season, compared with the young, green tissue that was used in the greenhouse experiments. Another important finding was that transgenic plants were not more susceptible to rust (Puccinia emaculata). The results of this study suggest that lignin down-regulation in switchgrass can confer real-world improvements in biofuel yield without negative consequences to biomass yield or disease susceptibility.

Rapid Assessment of Lignin Content and Structure in Switchgrass (Panicum virgatum L.) Grown Under Different Environmental Conditions

Switchgrass (Panicum virgatum L.) is a candidate feedstock in bioenergy, and plant breeding and molecular genetic strategies are being used to improve germplasm. In order to assess these subsequent modifications, baseline biomass compositional data are needed in a relevant variety of environments. In this study, switchgrass cv. Alamo was grown in the field, greenhouse, and growth chamber and harvested into individual leaf and stem tissue components. These components were analyzed with pyrolysis vapor analysis using molecular beam mass spectrometry, Fourier transform infrared, and standard wet chemistry methods to characterize and compare the composition among the different growth environments. The details of lignin content, S/G ratios, and degree of cross-linked lignin are discussed. Multivariate approaches such as projection to latent structures regression found a very strong correlation between the lignin content obtained by standard wet chemistry methods and the two high throughput techniques employed to rapidly assess lignin in potential switchgrass candidates. The models were tested on unknown samples and verified by wet chemistry. The similar lignin content found by the two methods shows that both approaches are capable of determining lignin content in biomass in a matter of minutes.

Activation of lignocellulosic biomass by ionic liquid for biorefinery fractionation.

Fractionation of lignocellulosic biomass is an attractive solution to develop an economically viable biorefinery by providing a saccharide fraction to produce fuels and a lignin stream that can be converted into high value products such as carbon fibers. In this study, the analysis of ionic liquid-activated biomass demonstrates that in addition of decreasing crystallinity, the selected ILs (1-butyl-3-methylimidazolium acetate, 1-butyl-3-methylimidazolium chloride and 1-ethyl-3-methylimidazolium acetate) deacetylate Yellow poplar under mild conditions (dissolution at 60-80 °C), and lower the degradation temperature of each biomass polymeric component, thereby reducing the recalcitrance of biomass. Among the three tested ILs, 1-ethyl-3-methylimidazolium acetate performed the best, providing a strong linear relationship between the level of deacetylation and the rate of enzymatic saccharification for Yellow poplar.

Extraction of information from laser-induced breakdown spectroscopy spectral data by multivariate analysis

Laser-induced breakdown spectroscopy (LIBS) is being proposed more and more as a high-throughput technology to assess the elemental composition of materials. When a specific element is of interest, semiquantification is possible by building a calibration model using the emission line intensity of this element for known samples. However, a unique element has usually more than one emission line, and there are many examples where several emission lines used in combination give dramatically better results than any of the individual variables used alone. With a multivariate approach, models can be constructed that take into account all the emission lines related to a specific element; therefore more robust models can be developed. In this work, chemometric methods such as principal component analysis and partial least squares are proposed to resolve and extract useful information from the LIBS spectral data collected on biological materials.

Increasing the revenue from lignocellulosic biomass: Maximizing feedstock utilization

Replacing petroleum by biomass can be economically feasible by generating revenue from the three primary biomass constituents. The production of renewable chemicals and biofuels must be cost- and performance- competitive with petroleum-derived equivalents to be widely accepted by markets and society. We propose a biomass conversion strategy that maximizes the conversion of lignocellulosic biomass (up to 80% of the biomass to useful products) into high-value products that can be commercialized, providing the opportunity for successful translation to an economically viable commercial process. Our fractionation method preserves the value of all three primary components: (i) cellulose, which is converted into dissolving pulp for fibers and chemicals production; (ii) hemicellulose, which is converted into furfural (a building block chemical); and (iii) lignin, which is converted into carbon products (carbon foam, fibers, or battery anodes), together producing revenues of more than

Enhanced discrimination and calibration of biomass NIR spectral data using non-linear kernel methods.

500 per dry metric ton of biomass. Once de-risked, our technology can be extended to produce other renewable chemicals and biofuels.