Power-optimized, time-reversal pulse sequence for a robust recovery of signals from rigid segments using time domain NMR.

Time domain NMR (TD-NMR) has been widely used on the analysis of liquids or liquid components in heterogeneous materials such as food, biological tissues, synthetic and bio polymers, oil-bearing rocks, biomasses and cement-based materials. The use of TD-NMR for studying solid and soft mater has been growing in number and variety of applications, mostly for organic systems where the detection of 1H signals is highly advantageous. However, the strong 1H-1H dipolar interactions in solids make the 1H FID to decay in the same order of the dead time of most commercially available NMR probe heads. Thus, solid echoes are often used for recovering signals from solid components. In this article we reinvestigate the time-reversal solid-echo pulse sequence proposed by Rhim and Kessemeier, seeking for optimal pulse power and timing conditions that maximize its efficiency on recovering 1H signals from rigid segments. We show that under these optimized conditions, which we denote as Rhim and Kessemeier - Radiofrequency Optimized Solid-Echo (RK-ROSE), the experiment can be more efficient than its most popular counterparts Solid-Echo (SE) and mixed-Magic Sandwich Echoes (mixed-MSE). Our results also suggest that, despite the finite pulse power, with current probe technology the RK-ROSE experiment is potentially able to provide an accurate estimation of rigid components, without relying on an external calibration using multiple standard samples, as usually done in SFC analysis of the FID signal. At last, we demonstrate that RK-ROSE can be adapted as a simple filter to supress signals from mobile segments in heterogeneous materials.

Defining functional diversity for lignocellulose degradation in a microbial community using multi-omics studies.

BACKGROUND: Lignocellulose is one of the most abundant forms of fixed carbon in the biosphere. Current industrial approaches to the degradation of lignocellulose employ enzyme mixtures, usually from a single fungal species, which are only effective in hydrolyzing polysaccharides following biomass pre-treatments. While the enzymatic mechanisms of lignocellulose degradation have been characterized in detail in individual microbial species, the microbial communities that efficiently breakdown plant materials in nature are species rich and secrete a myriad of enzymes to perform "community-level" metabolism of lignocellulose. Single-species approaches are, therefore, likely to miss important aspects of lignocellulose degradation that will be central to optimizing commercial processes. RESULTS: Here, we investigated the microbial degradation of wheat straw in liquid cultures that had been inoculated with wheat straw compost. Samples taken at selected time points were subjected to multi-omics analysis with the aim of identifying new microbial mechanisms for lignocellulose degradation that could be applied in industrial pre-treatment of feedstocks. Phylogenetic composition of the community, based on sequenced bacterial and eukaryotic ribosomal genes, showed a gradual decrease in complexity and diversity over time due to microbial enrichment. Taxonomic affiliation of bacterial species showed dominance of Bacteroidetes and Proteobacteria and high relative abundance of genera Asticcacaulis, Leadbetterella and Truepera. The eukaryotic members of the community were enriched in peritrich ciliates from genus Telotrochidium that thrived in the liquid cultures compared to fungal species that were present in low abundance. A targeted metasecretome approach combined with metatranscriptomics analysis, identified 1127 proteins and showed the presence of numerous carbohydrate-active enzymes extracted from the biomass-bound fractions and from the culture supernatant. This revealed a wide array of hydrolytic cellulases, hemicellulases and carbohydrate-binding modules involved in lignocellulose degradation. The expression of these activities correlated to the changes in the biomass composition observed by FTIR and ssNMR measurements. CONCLUSIONS: A combination of mass spectrometry-based proteomics coupled with metatranscriptomics has enabled the identification of a large number of lignocellulose degrading enzymes that can now be further explored for the development of improved enzyme cocktails for the treatment of plant-based feedstocks. In addition to the expected carbohydrate-active enzymes, our studies reveal a large number of unknown proteins, some of which may play a crucial role in community-based lignocellulose degradation.

Dipolar filtered magic-sandwich-echoes as a tool for probing molecular motions using time domain NMR.

We present a simple 1H NMR approach for characterizing intermediate to fast regime molecular motions using 1H time-domain NMR at low magnetic field. The method is based on a Goldmann Shen dipolar filter (DF) followed by a Mixed Magic Sandwich Echo (MSE). The dipolar filter suppresses the signals arising from molecular segments presenting sub kHz mobility, so only signals from mobile segments are detected. Thus, the temperature dependence of the signal intensities directly evidences the onset of molecular motions with rates higher than kHz. The DF-MSE signal intensity is described by an analytical function based on the Anderson Weiss theory, from where parameters related to the molecular motion (e.g. correlation times and activation energy) can be estimated when performing experiments as function of the temperature. Furthermore, we propose the use of the Tikhonov regularization for estimating the width of the distribution of correlation times.

Targeted metatranscriptomics of compost-derived consortia reveals a GH11 exerting an unusual exo-1,4-ß-xylanase activity.

BACKGROUND: Using globally abundant crop residues as a carbon source for energy generation and renewable chemicals production stand out as a promising solution to reduce current dependency on fossil fuels. In nature, such as in compost habitats, microbial communities efficiently degrade the available plant biomass using a diverse set of synergistic enzymes. However, deconstruction of lignocellulose remains a challenge for industry due to recalcitrant nature of the substrate and the inefficiency of the enzyme systems available, making the economic production of lignocellulosic biofuels difficult. Metatranscriptomic studies of microbial communities can unveil the metabolic functions employed by lignocellulolytic consortia and identify novel biocatalysts that could improve industrial lignocellulose conversion. RESULTS: In this study, a microbial community from compost was grown in minimal medium with sugarcane bagasse sugarcane bagasse as the sole carbon source. Solid-state nuclear magnetic resonance was used to monitor lignocellulose degradation; analysis of metatranscriptomic data led to the selection and functional characterization of several target genes, revealing the first glycoside hydrolase from Carbohydrate Active Enzyme family 11 with exo-1,4-ß-xylanase activity. The xylanase crystal structure was resolved at 1.76 Å revealing the structural basis of exo-xylanase activity. Supplementation of a commercial cellulolytic enzyme cocktail with the xylanase showed improvement in Avicel hydrolysis in the presence of inhibitory xylooligomers. CONCLUSIONS: This study demonstrated that composting microbiomes continue to be an excellent source of biotechnologically important enzymes by unveiling the diversity of enzymes involved in in situ lignocellulose degradation.

Folding of xylan onto cellulose fibrils in plant cell walls revealed by solid-state NMR.

Exploitation of plant lignocellulosic biomass is hampered by our ignorance of the molecular basis for its properties such as strength and digestibility. Xylan, the most prevalent non-cellulosic polysaccharide, binds to cellulose microfibrils. The nature of this interaction remains unclear, despite its importance. Here we show that the majority of xylan, which forms a threefold helical screw in solution, flattens into a twofold helical screw ribbon to bind intimately to cellulose microfibrils in the cell wall. 13C solid-state magic-angle spinning (MAS) nuclear magnetic resonance (NMR) spectroscopy, supported by in silico predictions of chemical shifts, shows both two- and threefold screw xylan conformations are present in fresh Arabidopsis stems. The twofold screw xylan is spatially close to cellulose, and has similar rigidity to the cellulose microfibrils, but reverts to the threefold screw conformation in the cellulose-deficient irx3 mutant. The discovery that induced polysaccharide conformation underlies cell wall assembly provides new principles to understand biomass properties.

Observation of Time-Invariant Coherence in a Nuclear Magnetic Resonance Quantum Simulator.

The ability to live in coherent superpositions is a signature trait of quantum systems and constitutes an irreplaceable resource for quantum-enhanced technologies. However, decoherence effects usually destroy quantum superpositions. It was recently predicted that, in a composite quantum system exposed to dephasing noise, quantum coherence in a transversal reference basis can stay protected for an indefinite time. This can occur for a class of quantum states independently of the measure used to quantify coherence, and it requires no control on the system during the dynamics. Here, such an invariant coherence phenomenon is observed experimentally in two different setups based on nuclear magnetic resonance at room temperature, realizing an effective quantum simulator of two- and four-qubit spin systems. Our study further reveals a novel interplay between coherence and various forms of correlations, and it highlights the natural resilience of quantum effects in complex systems.

Evaluating the composition and processing potential of novel sources of Brazilian biomass for sustainable biorenewables production.

BACKGROUND: The search for promising and renewable sources of carbohydrates for the production of biofuels and other biorenewables has been stimulated by an increase in global energy demand in the face of growing concern over greenhouse gas emissions and fuel security. In particular, interest has focused on non-food lignocellulosic biomass as a potential source of abundant and sustainable feedstock for biorefineries. Here we investigate the potential of three Brazilian grasses (Panicum maximum, Pennisetum purpureum and Brachiaria brizantha), as well as bark residues from the harvesting of two commercial Eucalyptus clones (E. grandis and E. grandis x urophylla) for biofuel production, and compare these to sugarcane bagasse. The effects of hot water, acid, alkaline and sulfite pretreatments (at increasing temperatures) on the chemical composition, morphology and saccharification yields of these different biomass types were evaluated. RESULTS: The average yield (per hectare), availability and general composition of all five biomasses were compared. Compositional analyses indicate a high level of hemicellulose and lignin removal in all grass varieties (including sugarcane bagasse) after acid and alkaline pretreatment with increasing temperatures, whilst the biomasses pretreated with hot water or sulfite showed little variation from the control. For all biomasses, higher cellulose enrichment resulted from treatment with sodium hydroxide at 130°C. At 180°C, a decrease in cellulose content was observed, which is associated with high amorphous cellulose removal and 5-hydroxymethyl-furaldehyde production. Morphological analysis showed the effects of different pretreatments on the biomass surface, revealing a high production of microfibrillated cellulose on grass surfaces, after treatment with 1% sodium hydroxide at 130°C for 30 minutes. This may explain the higher hydrolysis yields resulting from these pretreatments, since these cellulosic nanoparticles can be easily accessed and cleaved by cellulases. CONCLUSION: Our results show the potential of three Brazilian grasses with high productivity yields as valuable sources of carbohydrates for ethanol production and other biomaterials. Sodium hydroxide at 130°C was found to be the most effective pretreatment for enhanced saccharification yields. It was also efficient in the production of microfibrillated cellulose on grass surfaces, thereby revealing their potential as a source of natural fillers used for bionanocomposites production.

Effects of pretreatment on morphology, chemical composition and enzymatic digestibility of eucalyptus bark: a potentially valuable source of fermentable sugars for biofuel production - part 1.

BACKGROUND: In recent years, the growing demand for biofuels has encouraged the search for different sources of underutilized lignocellulosic feedstocks that are available in sufficient abundance to be used for sustainable biofuel production. Much attention has been focused on biomass from grass. However, large amounts of timber residues such as eucalyptus bark are available and represent a potential source for conversion to bioethanol. In the present paper, we investigate the effects of a delignification process with increasing sodium hydroxide concentrations, preceded or not by diluted acid, on the bark of two eucalyptus clones: Eucalyptus grandis (EG) and the hybrid, E. grandis x urophylla (HGU). The enzymatic digestibility and total cellulose conversion were measured, along with the effect on the composition of the solid and the liquor fractions. Barks were also assessed using Fourier-transform infrared spectroscopy (FTIR), solid-state nuclear magnetic resonance (NMR), X-Ray diffraction, and scanning electron microscopy (SEM). RESULTS: Compositional analysis revealed an increase in the cellulose content, reaching around 81% and 76% of glucose for HGU and EG, respectively, using a two-step treatment with HCl 1%, followed by 4% NaOH. Lignin removal was 84% (HGU) and 79% (EG), while the hemicellulose removal was 95% and 97% for HGU and EG, respectively. However, when we applied a one-step treatment, with 4% NaOH, higher hydrolysis efficiencies were found after 48 h for both clones, reaching almost 100% for HGU and 80% for EG, in spite of the lower lignin and hemicellulose removal. Total cellulose conversion increased from 5% and 7% to around 65% for HGU and 59% for EG. NMR and FTIR provided important insight into the lignin and hemicellulose removal and SEM studies shed light on the cell-wall unstructuring after pretreatment and lignin migration and precipitation on the fibers surface, which explain the different hydrolysis rates found for the clones. CONCLUSION: Our results show that the single step alkaline pretreatment improves the enzymatic digestibility of Eucalyptus bark. Furthermore, the chemical and physical methods combined in this study provide a better comprehension of the pretreatment effects on cell-wall and the factors that influence enzymatic digestibility of this forest residue.

Quantum information processing by nuclear magnetic resonance on quadrupolar nuclei.

Nuclear magnetic resonance is viewed as an important technique for the implementation of many quantum information algorithms and protocols. Although the most straightforward approach is to use the two-level system composed of spin 1/2 nuclei as qubits, quadrupolar nuclei, which possess a spin greater than 1/2, are being used as an alternative. In this study, we show some unique features of quadrupolar systems for quantum information processing, with an emphasis on the ability to execute efficient quantum state tomography (QST) using only global rotations of the spin system, whose performance is shown in detail. By preparing suitable states and implementing logical operations by numerically optimized pulses together with the QST method, we follow the stepwise execution of Grover's algorithm. We also review some work in the literature concerning the relaxation of pseudo-pure states in spin 3/2 systems as well as its modelling in both the Redfield and Kraus formalisms. These data are used to discuss differences in the behaviour of the quantum correlations observed for two-qubit systems implemented by spin 1/2 and quadrupolar spin 3/2 systems, also presented in the literature. The possibilities and advantages of using nuclear quadrupole resonance experiments for quantum information processing are also discussed.

Studies of the compositions of humic acids from Amazonian Dark Earth soils.

The compositions of humic acids (HAs) isolated from cultivated and forested "Terra Preta de Indio" or Amazonian Dark Earth soils (anthropogenic soils) were compared with those from adjacent non-anthropogenic soils (control soils) using elemental and thermogravimetric analyses, and a variety of solid-state nuclear magnetic resonance techniques. The thermogravimetric index, which indicates the molecular thermal resistance, was greater for the anthropogenic soils than for the control soils suggesting polycyclic aromatic components in the former. The cultivated anthropogenic soils were more enriched in C and depleted in H than the anthropogenic soils under forest, as the result of the selective degradation of aliphatic structures and the possible enrichment of H-deficient condensed aromatic structures. The combination of variable amplitude cross-polarization (VACP) and chemical shift anisotropy with total suppression of spinning sidebands experiments with composite pi pulses could be used to quantify the aromaticity of the HAs from the anthropogenic soils. From principal component analysis, using the VACP spectra, it was possible to separate the different constituents of the HAs, such as the carboxylated aromatic structures, from the anthropogenic soils and plant derived compounds. The data show that the HAs from anthropogenic soils have high contents of aryl and ionisable oxygenated functional groups, and the major functionalities from adjacent control soils are oxygenated functional groups from labile structures (carbohydrates, peptides, and with evidence for lignin structures). The anthropogenic soils HAs can be considered to be more recalcitrant, and with more stable reactive functional groups which may, in part, explain their more sustainable fertility due to the organic matter contribution to the soil cation exchange capacity.

Characterisation of black carbon-rich samples by (13)C solid-state nuclear magnetic resonance.

There are difficulties in quantifying and characterising the organic matter (OM) in soils that contain significant amounts of partially oxidised char or charcoal materials. The anthropogenic black carbon (BC), such as that found in the Terra Preta de Indio soils of the Amazon region, is a good example of the OM that is difficult to analyse in such soils. (13)C direct polarisation/magic angle spinning (DP/MAS) at high MAS frequency, (1)H-(13)C cross polarisation (CP)/MAS with total suppression of spinning sidebands (TOSS), and chemical shift anisotropy (CSA) filter nuclear magnetic resonance techniques have been applied successfully for quantifying the different components of OM. However, because pyrogenic materials present strong local magnetic susceptibility heterogeneities, the use of CSA-filter and TOSS make the pulse sequences very sensitive to imperfections in the pi pulses. In this study, the DP/MAS pulse sequence was replaced by a CP with a radio frequency ramp--variable amplitude CP (VACP)--VACP/MAS pulse sequence, and composite pi pulses were used in the CSA-filter and TOSS pulse sequences. In that way, the component functionalities in a humic acid from a BC soil were successfully determined. The spectrometer time needed was greatly decreased by employing this VACP/MAS technique. This development provides an accurate method for characterising BC-rich samples from different origins.

A solid-state NMR study of the fast and slow dynamics of collagen fibrils at varying hydration levels.

We report solid-state NMR investigations of the effect of temperature and hydration on the molecular mobility of collagen isolated from bovine achilles tendon. (13)C cross-polarization magic angle spinning (MAS) experiments were performed on samples at natural abundance, using NMR methods that detect motionally averaged dipolar interactions and chemical shift anisotropies and also slow reorientational processes. Fast motions with correlation times much shorter than 40 micro s scale dipolar couplings and chemical shift anisotropies of the carbon sites in collagen. These motionally averaged anisotropic interactions provide a measure of the amplitudes of the segmental motions expressed by a molecular order parameter. The data reveal that increasing hydration has a much stronger effect on the amplitude of the molecular processes than increasing temperature. In particular, the Cgamma carbons of the hydroxyproline residues exhibit a strong dependence of the amplitude of motion on the hydration level. This could be correlated with the effect of hydration on the hydrogen bonding structure in collagen, for which this residue is known to play a crucial role. The applicability of 1D MAS exchange experiments to investigate motions on the millisecond time-scale is discussed and first results are presented. Slow motions with correlation times of the order of milliseconds have also been detected for hydrated collagen.