PcGCE is a potent elicitor of defense responses in aspen

Using microbial enzymes in transgenesis is a powerful means to introduce new functionalities in plants. Glucuronoyl esterase (GCE) is a microbial enzyme hydrolyzing the ester bond between lignin and 4-O-methyl-α-D-glucuronic acid present as a side chain of glucuronoxylan. This bond mediates lignin-carbohydrate complex (LCC) formation, considered as crucial factor of lignocellulose recalcitrance to saccharification. Previous studies showed that hybrid aspen (Populus tremula L. x tremuloides Michx.) constitutively expressing Phanerochaete carnosa Burt GCE (PcGCE) had better efficiency of cellulose-to-glucose conversion but were stunned and had lower cellulose content indicating that more studies are needed to design strategy for deployment of this enzyme in planta. Here we report that the transgenic plants exhibit premature leaf senescence, increased accumulation of calcium oxalate crystals, tyloses and necrotic lesions and have strongly activated immune defense reactions as revealed by their altered profiles of transcriptomes, metabolomes and hormones in the leaves. To elucidate if these effects are triggered by damage-associated molecular patterns (DAMPs) or by PcGCE protein perceived as a pathogen-associated molecular pattern (PAMP), we ectopically expressed in aspen an enzymatically inactive PcGCES217A. The mutated PcGCE induced similar growth retardation, leaf necrosis and premature senescence as the active one, providing evidence that PcGCE protein is recognized as PAMP. Transcriptomics analysis of young expanding leaves of 35S:PcGCE plants identified several candidates for receptors of PcGCE, which were not expressed in developing wood tissues. Grafting experiments showed that PcGCE transcripts are not cell-to-cell mobile and that leaves augment systemic responses. In agreement, expressing PcGCE in developing wood by using the wood-specific promoter (WP), avoided all off-target effects. Moreover, WP:PcGCE lines grew better than control plants providing evidence that this strategy can be used in transgenic crops dedicated for biorefinery.

Elucidation of the Structure of Lignin-Carbohydrate Complexes in Ginkgo CW-DHP by 13C-2H Dual Isotope Tracer

To elucidate the chemical linkages between lignin and carbohydrates in ginkgo cell walls, 13C-2H-enriched cell wall-dehydrogenation polymers (CW-DHP) were selectively prepared with cambial tissue from Ginkgo biloba L. by feeding D-glucose-[6-2H2], coniferin-[α-13C], and phenylalanine ammonia-lyase (PAL) inhibitor. The abundant detection of 13C and 2H confirmed that D-glucose-[6-2H2] and coniferin-[α-13C] were involved in the normal metabolism of ginkgo cambial cells that had been effectively labelled with dual isotopes. In the ginkgo CW-DHP, ketal and ether linkages were formed between the C-α of lignin side chains and carbohydrates, as revealed by solid state CP/MAS 13C-NMR differential spectroscopy. Furthermore, the DMSO/TBAH ionic liquids system was used to fractionate the ball-milled CW-DHP into three lignin-carbohydrate complex (LCC) fractions: glucan–lignin complex (GL), glucomannan–lignin complex (GML), and xylan–lignin complex (XL). The XRD determination indicated that the cellulose type I of the GL was converted into cellulose type II during the separation process. The molecular weight was in the order of Ac-GL > Ac-GML > XL. The 13C-NMR and 1H-NMR differential spectroscopy of 13C-2H-enriched GL fraction indicated that lignin was linked with cellulose C-6 by benzyl ether linkages. It was also found that there were benzyl ether linkages between the lignin side chain C-α and glucomannan C-6 in the 13C-2H-enriched GML fraction. The formation of ketal linkages between the C-α of lignin and xylan was confirmed in the 13C-2H-enriched XL fraction.

Lignin/Carbohydrate Complex Isolated from Posidonia oceanica Sea Balls (Egagropili): Characterization and Antioxidant Reinforcement of Protein-Based Films

A lignin fraction (LF) was extracted from the sea balls of Posidonia oceanica (egagropili) and extensively dialyzed and characterized by FT-IR and NMR analyses. LF resulted water soluble and exhibited a brownish-to-black color with the highest absorbance in the range of 250–400 nm, attributed to the chromophore functional groups present in the phenylpropane-based polymer. LF high-performance size exclusion chromatography analysis showed a highly represented (98.77%) species of 34.75 kDa molecular weight with a polydispersity index of 1.10 and an intrinsic viscosity of 0.15. Quantitative analysis of carbohydrates indicated that they represented 28.3% of the dry weight of the untreated egagropili fibers and 72.5% of that of LF. In particular, eight different monosaccharides were detected (fucose, arabinose, rhamnose, galactose, glucose, xylose, glucosamine and glucuronic acid), glucuronic acid (46.6%) and rhamnose (29.6%) being the most present monosaccharides in the LF. Almost all the phenol content of LF (113.85 ± 5.87 mg gallic acid eq/g of extract) was water soluble, whereas around 22% of it consisted of flavonoids and only 10% of the flavonoids consisted of anthocyanins. Therefore, LF isolated from egagropili lignocellulosic material could be defined as a water-soluble lignin/carbohydrate complex (LCC) formed by a phenol polymeric chain covalently bound to hemicellulose fragments. LCC exhibited a remarkable antioxidant activity that remained quite stable during 6 months and could be easily incorporated into a protein-based film and released from the latter overtime. These findings suggest egagropili LCC as a suitable candidate as an antioxidant additive for the reinforcement of packaging of foods with high susceptibility to be deteriorated in aerobic conditions.

Study of the kinetic regularities of the grain extrusion process in the production of highly digestible feed with protected protein for cattle

The problem of developing of highly digestible compound feeds with protected protein for cattle is due to the specifics of the gastric tract of cows and the peculiarities of protein assimilation. Due to the importance of preliminary moisture-heat treatment of grain for the subsequent course of the extrusion process, it was carried out by steam at a pressure of 0.6 МРа before moistening the feed to a humidity of 17–20% and heating to a temperature of 70–80° C. It is established that heat treatment has a significant effect on the carbohydrate complex of grain: heating it at high temperatures causes the destruction of starch, accompanied by the formation of easily soluble carbohydrates, which has a positive effect on the digestibility of feed. The kinetic regularities of the processes of moisture-heat treatment, grinding and extrusion of grain in the production of highly digestible feed with protected protein for cattle were studied. The degree of dextrinization and the digestibility of starch increased with the heating temperature of corn and its mixtures with wheat up to 100–110° C, when the performance of the extruder was 300–320 kg/h, the digestibility of starch extruded corn and grain mixture is increased to 85 and 68 mg of glucose per 1 g of the product (hereinafter mg/g), respectively. For wheat, this indicator is lower and, accordingly, is 50 mg/g. When heated during the extrusion of corn to a temperature of up to 120–140° C, the digestibility of starch was 100–110 mg/g, and for the grain mixture – 80–83 mg/g. At this temperature, the digestibility of the starch of extruded wheat corresponded to 60–65 mg/g. The optimal moisture content of feed in the process of extrusion for the purpose of forming pellets is 18%. Studies of the extrusion effect on the carbohydrate complex of processed feed have shown that the destruction of starch in the extruded product increases. Thus, the content of soluble carbohydrates increases by 27–32%, and the digestibility of starch increases twice in extruded feed compared to unprocessed.

INVESTIGATION OF THE COMPOSITION OF POLYPHENOLIC SUBSTANCES OF THE JUICE FROM ARTICHOKE TUBERS

Currently, the growth of diseases with diabetes, metabolic disorders, and obesity increases the demand for preventive and functional products. Currently available technologies for the production of preventive products provide for the replacement of sugar with sugar substitutes or its complete absence, which naturally reduces the nutritional and energy value of the products produced. One of the main promising areas of the processing industry is the production of new products and sugar substitutes based on non-traditional types of plant raw materials that have a rich carbohydrate complex in their composition. These types of raw materials among vegetable crops include jerusalem artichoke. Currently, jerusalem artichoke juice is of particular interest for use in the production of soft drinks, as it has a rich chemical composition, which makes Jerusalem artichoke indispensable in dietary nutrition, in the preparation of highly effective medicines. In this article, the chemical composition of tubers and juice-semi-finished products from jerusalem artichoke is investigated. The results of the studies showed that the bulk of the dry substances in jerusalem artichoke tubers are carbohydrates, most of which are represented by fructosides. In the semi-finished juice passes: oligosaccharides 22.54 %, monosaccharides 1.61%, as well as part of the structural polysaccharides-fiber 0.07 %, pectin substances 1.84 %. The change in the fractional composition of the polyphenolic substances of the pulp and semi - finished juice was studied when the pulp was kept for 30 minutes. When developing the technology for the production of juices and beverages based on jerusalem artichoke, it was found that the polyphenolic substances of jerusalem artichoke tubers have a great influence on the technological properties of raw materials, the quality and nutritional value of the finished product. It was found that the enzymatic process of oxidative transformation of polyphenols occurs as much as possible in the first 5-10 minutes.

On the Use of Molecular Dynamics Simulations for Elucidating Fine Structural, Physico-Chemical and Thermomechanical Properties of Lignocellulosic Systems: Historical and Future Perspectives

The use of Molecular Dynamics (MD) simulations for predicting subtle structural, thermomechanical and related characteristics of lignocellulosic systems is studied. A historical perspective and the current state of the art are discussed. The use of parameterised MD force fields, scaling up simulations via high performance computing and intrinsic molecular mechanisms influencing the mechanical, thermal and chemical characteristics of lignocellulosic systems and how these can be predicted and modelled using MD is shown. Individual discussions on the MD simulations of the lignin, cellulose, lignin-carbohydrate complex (LCC) and how MD can elucidate the role of water on the surface and microstructural characteristics of these lignocellulosic systems is shown. In addition, the use of MD for unearthing molecular mechanisms behind lignin-enzyme interactions during precipitation processes and the deforming/structure weakening brought about by cellulosic interactions in some lignocellulosic systems is both predicted and quantified. MD results from relatively smaller systems comprised of several hundred to a few thousand atoms and massive multi-million atom systems are both discussed. The versatility and effectiveness of MD based on its ability to provide viable predictions from both smaller and massive starting systems is presented in detail.