Lignin Precipitation and Fractionation from OrganoCat Pulping to Obtain Lignin with Different Sizes and Chemical Composition.

Fractionation of lignocellulose into its three main components, lignin, hemicelluloses, and cellulose, is a common approach in modern biorefinery concepts. Whereas the valorization of hemicelluloses and cellulose sugars has been widely discussed in literature, lignin utilization is still challenging. Due to its high heterogeneity and complexity, as well as impurities from pulping, it is a challenging feedstock. However, being the most abundant source of renewable aromatics, it remains a promising resource. This work describes a fractionation procedure that aims at stepwise precipitating beech wood (Fagus sp.) lignin obtained with OrganoCat technology from a 2-methyltetrahydrofuran solution, using n-hexane and n-pentane as antisolvents. By consecutive antisolvent precipitation and filtration, lignin is fractionated and then characterized to elucidate the structure of the different fractions. This way, more defined and purified lignin fractions can be obtained. Narrowing down the complexity of lignin and separately valorizing the fractions might further increase the economic viability of biorefineries.

Antileukemic Activity of Twig Components of Caucasian Beech in Turkey.

Despite the development of a range of anti-cancer agents, cancer diagnoses are still increasing in number, remaining a leading cause of death. Anticancer drug treatment is particularly important for leukemia. We screened Turkish plants and found the unique antileukemic activity of twig components in Turkish Caucasian beech, selectively inducing apoptosis in leukemia cells. This effect is unique among some kinds of beeches, presumably related to oxidative stress. This study would lead to effective use of discarded material, i.e., twig of beech, and a new anti-leukemic drug based on large tree.

Behaviour of aluminium in forest soils with different lithology and herb vegetation cover.

The aim of this study was to determine the content, distribution and behaviour of Al in soils under beech forest with different parent rock, and to assess the role of herbaceous vegetation on soil Al behaviour. We hypothesize that the contents of elements in the soil sorption complex (Al etc.) are strongly influenced by vegetation cover. Also, low molecular mass organic acids (LMMOA) can be considered as an indicator of soil organic matter (SOM) decomposition and vegetation litter turnover. Speciation of LMMOA, nutrition content (PO43-, Ca2+, K+) and element composition in aqueous extracts were determined by means of ion chromatography and inductively coupled plasma - optical emission spectrometry (ICP-OES) respectively. Active and exchangeable pH, sorption characteristics and exchangeable Al (Alex) were determined in BaCl2 extracts by ICP-OES. Elemental composition of parent rocks was assessed by means of X-ray fluorescence spectroscopy. Herb-poor localities showed lower pH, less nutrients (PO43-, Ca2+, K+), less LMMOA, a larger stock of SOM and greater cation exchange capacity. There was also lower mobilisation of Al in organic horizons, which explains the larger pools of Al. Generally, we can conclude that LMMOA, and thus soil vegetation cover, play an important role in the Al soil cycle.

Physicochemical characteristics of the cold-pressed oil obtained from seeds of Fagus sylvatica L.

A physicochemical characteristic of the cold-pressed oil obtained from seeds of common beech (Fagus sylvatica L.) has been presented. This plant may be considered as unconventional oilseeds crops because of relatively high content of fat (27.25%). The analyzed beech seeds oil has been classified as oleic-linoleic acids oil with more than 76% percentage share of those species. Beech seeds oil contains 4.2% of gamma-linolenic acid (GLA). Unique characteristic is the high content of γ-tocopherol (75.4mg/100g) and δ-tocopherol (34.05mg/100g). γ-Tocopherol is effective scavengers of reactive nitrogen species and prevents DNA bases nitration, what makes beech seeds oil interesting raw material in the production of cosmetics. Additionally the content of carotenoids, very effective photooxidation inhibitors, is at high level in comparison with other cold-pressed oils. It was demonstrated that PCA analysis may help to determine the authenticity of oil obtained from beech seeds.

Direct lactic acid production from beech wood by transgenic white-rot fungus Phanerochaete sordida YK-624.

A lactic acid (LA)-producing strain of the hyper-lignin-degrading fungus Phanerochaete sordida YK-624 with the lactate dehydrogenase-encoding gene from Bifidobacterium longum (Blldh) was constructed. When the endogenous pyruvate decarboxylase gene-knocked down and Blldh-expressing transformant was cultured with beech wood meal, the transformant was able to successively delignify and ferment the substrate. Supplementation of calcium carbonate into the culture medium, significantly increased the level of LA accumulation. Direct LA production (at 0.29g/l) from wood was confirmed, and additional inclusion of exogenous cellulase in this fermentation yielded significant further improvement in LA accumulation (up to 1.44g/l). This study provides the first report of direct production of LA by fermentation from woody biomass by a single microorganism, and indicates that transgenic white-rot fungi have a potential use for development of simple/easy applications for wood biorefinery.

Non-targeted metabolomic profile of Fagus sylvatica L. leaves using liquid chromatography with mass spectrometry and gas chromatography with mass spectrometry.

INTRODUCTION: Fagus sylvatica L. is one of the most widely distributed broad-leaved tree species in central and western Europe, important to the forest sector and an accurate biomarker of climate change. OBJECTIVE: To profile the beech leaf metabolome for future studies in order to investigate deeper into the characterisation of its metabolic response. METHODS: Leaf extracts were analysed using LC-MS by electrospray ionisation in negative mode from m/z 100-1700 and GC-MS by electron ionisation in scan mode from m/z 35-800. RESULTS: The LC-MS profile resulted in 56 compounds, of which 43 were identified and/or structurally characterised, including hydroxycinnamic acid derivatives, flavan-3-ols and proanthocyanidins, and flavonols. From a second analysis based on GC-MS, a total of 111 compounds were identified, including carbohydrates, polyalcohols, amino acids, organic acids, fatty acids, phenolic compounds, terpenoids, sterols and other related compounds. Many of the compounds identified were primary metabolites involved in major plant metabolic pathways, however, some secondary metabolites were also detected. Some of them play roles as tolerance-response osmoregulators and osmoprotectors in abiotic stress, or as anti-oxidants that reduce the effect of reactive oxygen species and promote many protective functions in plants. CONCLUSIONS: This study provides a broad and relevant insight into the metabolic status of F. sylvatica leaves, and serves as a base for future studies on physiological and molecular mechanisms involved in biotic or abiotic stress.

Atmospheric particulate deposition in temperate deciduous forest ecosystems: interactions with the canopy and nutrient inputs in two beech stands of Northeastern France.

As wood harvests are expected to increase to satisfy the need for bio-energy in Europe, quantifying atmospheric nutrient inputs in forest ecosystems is essential for forest management. Current atmospheric measurements only take into account the <0.45 µm fraction and dry deposition is generally modeled. The aims of this study were to quantify atmospheric particulate deposition (APD), the >0.45 µm fraction of atmospheric deposition, below the canopy, to study the influence of the canopy on APD, and to determine the influence of APD below canopy to nutrient input-output budgets with a focus on base cations calcium, magnesium and potassium, and phosphorus. APD was sampled every four weeks by passive collectors. We divided APD into an organic and a mineral fraction, respectively POM and MDD. MDD was divided into a soluble and a hardly soluble fraction in hydrogen peroxide, referred to as S-MDD and H-MDD, respectively. In order to better understand the influence of the canopy on APD, we studied APD in three pathways below the canopy (litterfall, stemflow and throughfall), and in open field. Our results indicated that APD in throughfall (123 ± 64 kg ha(-1)year(-1)) was significantly higher and synchronic with that in open field (33 ±9 kg ha(-1)year(-1)) in the two study sites. This concerned both POM and MDD, suggesting a large interception of APD by foliar surfaces, which is rapidly washed off by rain within four weeks. Throughfall H-MDD was the main pathway with an average of 16 ± 2 kg ha(-1)year(-1). Stemflow and litterfall were neglected. In one study site, canopy intercepted about 8 kg ha(-1)year(-1) of S-MDD. Although base cations and phosphorus inputs by APD are lower than those of <0.45 µm deposition, they contributed from 5 to 32% to atmospheric deposition and improved the nutrient budget in one of the study sites.

The cough suppressive activity of sulfated glucuronoxylan from Fagus sylvatica L.

Hemicellulose polysaccharides represent a large group of natural renewable polymers, however, their application potency is still low. In our study a hardwood 4-O-methylglucuronoxylan was isolated by alkali peroxide extraction of Fagus sylvatica sawdust and modified into sulfated water soluble derivative (MGXS). Highly sulfated MGXS was characterized by HPLC, FTIR and NMR spectroscopies, and tested in vivo on chemically induced cough reflex and smooth muscles reactivity. Farmacological tests revealed an interesting antitussive activity of MGXS. Comparative tests with drug commonly used in a clinical practice revealed that antitussive activity of MGXS was lower than that of opioid receptor agonist codeine, the strongest antitussive drug. Furthermore, the specific reactivity of airways smooth muscle was not significantly affected by MGXS, indicating thus that the polymer is not involved in the bronchodilation process.

New substrates and activity of Phanerochaete chrysosporium Omega glutathione transferases.

Omega glutathione transferases (GSTO) constitute a family of proteins with variable distribution throughout living organisms. It is notably expanded in several fungi and particularly in the wood-degrading fungus Phanerochaete chrysosporium, raising questions concerning the function(s) and potential redundancy of these enzymes. Within the fungal families, GSTOs have been poorly studied and their functions remain rather sketchy. In this study, we have used fluorescent compounds as activity reporters to identify putative ligands. Experiments using 5-chloromethylfluorescein diacetate as a tool combined with mass analyses showed that GSTOs are able to cleave ester bonds. Using this property, we developed a specific activity-based profiling method for identifying ligands of PcGSTO3 and PcGSTO4. The results suggest that GSTOs could be involved in the catabolism of toxic compounds like tetralone derivatives. Biochemical investigations demonstrated that these enzymes are able to catalyze deglutathionylation reactions thanks to the presence of a catalytic cysteine residue. To access the physiological function of these enzymes and notably during the wood interaction, recombinant proteins have been immobilized on CNBr Sepharose and challenged with beech wood extracts. Coupled with GC-MS experiments this ligand fishing method allowed to identify terpenes as potential substrates of Omega GST suggesting a physiological role during the wood-fungus interactions.

Triterpenes as α-glucosidase inhibitors from Fagus hayatae.

Triterpenoids, 1-3, 8 and 9, along with 24 known compounds were isolated from leaves and twigs of Fagus hayatae. Of these, compound 1, 1,10-seco-3ß,10α,23-trihydroxyolean-12-ene-1,28-dioic acid 1,23-lactone, possesses a hitherto unknown 1,10-seco-oleanane skeleton. In addition, 2,3-seco-20(29)-lupene-2,3-dioic acid (16), previously described as a synthetic product, is now established as a plant natural product; the neolignan-9'-O-rhamnoside 19 is also characterized herein. Their structures were deduced mainly by 1D and 2D NMR spectroscopic analyses. Seven of these compounds possess moderate inhibitory activity against α-glucosidase type IV (Bacillus stearothermophilus).

[Genoprotective activities of the oils from leaves and fruits of Fagus orientalis Lipsky].

The antimutagenic activities of the oils obtained from leaves and fruits of Fagus orientalis have been shown in experiments with spontaneous and mutagen- and ageing-induced variability. The aberrations of chromosomes in the meristematic cells of the Allium cepa L., Vicia faba L., Triticum aestivum L., and marrow cells of Vistar rats as well as Arabidopsis thaliana gene mutations have been mobilized as experimental tests.

Aqueous leaf extracts display endocrine activities in vitro and disrupt sexual differentiation of male Xenopus laevis tadpoles in vivo.

The occurrence of natural substances acting as endocrine disrupting compounds (EDC) in the environment is to date poorly understood. Therefore, (anti)androgenic and (anti)estrogenic activities of three different aqueous leaf extracts (beech, reed and oak) were analyzed in vitro using yeast androgen and estrogen screen. The most potent extract was selected for in vivo exposure of Xenopus laevis tadpoles to analyze the potential effects on development and reproductive biology of amphibians. Tadpoles were exposed from stage 48 to stage 66 (end of metamorphosis) to aqueous oak leaf extracts covering natural occurring environmental concentrations of dissolved organic carbon. Gene expression analyses of selected genes of the hypothalamus-pituitary-gonad and of the hypothalamus-pituitary-thyroid axis as well as histological investigation of gonads and thyroid glands were used to evaluate endocrine disrupting effects on the reproductive biology and development. Female tadpoles remained unaffected by the exposure whereas males showed severe significant histological alterations of testes at the two highest oak leaf extract concentrations demonstrated by the occurrence of lacunae and oogonia. In addition, a significant elevation of luteinizing hormone beta mRNA expression with increasing extract concentration in male tadpoles indicates an involvement of hypothalamus-pituitary-gonad axis mainly via antiandrogenic activity. These results suggest that antiandrogenic EDC of oak leaf extract are responsible for inducing the observed effects in male tadpoles. The present study demonstrates for the first time that in surface waters, natural occurring oak leaf compounds at environmentally relevant concentrations display antiandrogenic activities and have considerable effects on the endocrine system of anurans affecting sexual differentiation of male tadpoles.

Protective effect of lignophenol derivative from beech (Fagus crenata Blume) on copper- and zinc-mediated cell death in PC12 cells.

Lignophenol, prepared using a phase-separation system, is a derivative of lignin, which is one of the components in the plant cell wall, and possesses high phenolic function, high stability and antioxidant properties. However, little is known about the beneficial effect of lignophenol. In this study, we investigated the protective effect of lignophenol from the beech tree (Fagus crenata Blume) on copper- and zinc-mediated apoptosis in PC12 cells by using DNA fragmentation and TUNEL assays. In DNA fragmentation assays, the DNA ladder patterns in the PC12 cells treated with 200 microM Cu and 200 microM Zn were enhanced, whereas the DNA ladder pattern was hardly observed in these cells treated with 20 mM lignophenol. In the TUNEL assay, TUNEL signals increased significantly in the untreated PC12 cells exposed to 200 microM Cu compared with the control. In contrast, the degree of apoptosis in the 20 mM lignophenol-treated cells was significantly lower than in the untreated cells, indicating that lignophenol inhibited Cu-induced apoptotic cell death in PC 12 cells. In the 200 microM Zn-exposed group, the degree of apoptosis in the 20 mM lignophenol-treated cells was also low compared with the untreated cells. In conclusion, these results suggest that lignophenol plays a role in protecting against Cu- and Zn-mediated PC12 apoptotic cell death.

Effects of simultaneous ozone exposure and nitrogen loads on carbohydrate concentrations, biomass, growth, and nutrient concentrations of young beech trees (Fagus sylvatica).

Beech seedlings were grown under different nitrogen fertilisation regimes (0, 20, 40, and 80 kg Nha(-1)yr(-1)) for three years and were fumigated with either charcoal-filtered (F) or ambient air (O3). Nitrogen fertilisation increased leaf necroses, aphid infestations, and nutrient ratios in the leaves (N:P and N:K), as a result of decreased phosphorus and potassium concentrations. For plant growth, biomass accumulation, and starch concentrations, a positive nitrogen effect was found, but only for fertilisations of up to 40 kg Nha(-1) yr(-1). The highest nitrogen load, however, reduced leaf area, leaf water content, growth, biomass accumulation, and starch concentrations, whereas soluble carbohydrate concentrations were enhanced. The ozone fumigation resulted in reduced leaf area, leaf water content, shoot growth, root biomass accumulation, and decreased starch, phosphorus, and potassium concentrations, increasing the N:P and N:K ratios. A combined effect of the two pollutants was detected for the leaf area and the shoot elongation, where ozone fumigation amplified the nitrogen effects.

Characterization of low molecular weight organic acids from beech wood treated in supercritical water.

Japanese beech (Fagus crenata Blume), its cell wall components, and model compounds were treated by supercritical water (380 degrees C, 100 MPa) for 5 s using a batch-type reactor to investigate the production behavior of low molecular weight organic acids. It was found that cellulose and hemicellulose were decomposed to formic acid, pyruvic acid, glycolic acid, acetic acid, and lactic acid, whereas lignin was barely decomposed to such organic acids under the given conditions. However, after prolonged treatment (380 degrees C, 100 MPa, 4 min) of lignin, some organic acids were recovered owing perhaps to the decomposition of the propyl side chain of lignin. It was additionally revealed that the predominant organic acid recovered was acetic acid, which might be derived from the acetyl group of hemicellulose in Japanese beech.