Studies on holocellulose and alpha-cellulose from spruce wood using cryo-ultramicrotomy

Summary Cryo-ultramicrotomy was applied to holocellulose and alpha-cellulose from spruce wood (Picea abies Karst.) for a light and electron microscopical study of the removal of lignin during chlorite delignification and the changes in swelling during delignification and alkali extraction. The swelling state of the fibre walls during each stage of treatment was well preserved, and distinct differences could be observed. Staining with uranyl acetate brought out the fine structure of the fibre walls down to the range of elementary fibrils.Submitted by Deutsche Forschungsgemeinschaft. The assistance of Dr. D. Grosser at the light microscope and of Miss U. Schwarz with the chemical analysis is greatly appreciated.     

沙棘籽原花青素提取物抗氧化作用研究

本试验研究了沙棘籽原花青素提取物的抗氧化作用。结果表明，该提取物按100mg／kg，300mg／kg及600mg／kg给小鼠灌胃，皆可提高小鼠红细胞SOD活性和小鼠全血GSH—PX活性，并能降低小鼠肝脏LPO含量。为此，沙棘籽原花青素提取物具有抗氧化作用。     

Studies on holocellulose and alpha-cellulose from spruce wood using cryo-ultramicrotomy part 2: The influence of heavy metal salt impregnation and the dimensions of delignified cell wall layers

Summary Delignified samples of spruce wood were impregnated with various heavy metal salts and afterwards cut on a cryo-ultramicrotome. In the electron microscope a different swelling of the cell walls and different staining effects depending on the impregnated salt were observed. The thickness of the cell wall layers was determined in holocelluloses containing different amounts of residual lignin and in holocelluloses impregnated with various heavy metal salts. From the results it can be seen that the swelling state of the wall layers is determined by the competitive processes of swelling and loss of substance. Metal salt impregnations cause different swelling states of the wall layers. Some of the salts show a specific deposition within the cell wall which is interpreted as a chemical affinity to polyoses.Supported by Deutsche Forschungsgemeinschaft. The assistance of Miss U. Schwartz with the preparation is greatly appreciated.     

The ultrastructure of the middle lamella region in resin canal tissue isolated from slash pine holocellulose

Summary Microscopic and electron microscopic examination of the resin canal complex isolated from slash pine chlorite holocellulose showed it to be composed of thin-walled, unlignified tissue apparently without secondary thickening. The mature structure consisted of a central canal surrounded by three cellular layers. Epithelium forms the inner layer while the middle and outer layers consist of short (intermediate) and elongated (outer) parenchyma cells, respectively. In contrast to previous concepts, the middle lamella of these tissues is a complex structure which contains intercellular membranes and cellulosic and noncellulosic (probably galacturonan) intercell wall fibrils. Intercellular membranes encapsulate two or more cells and consist of one or more intact ancestral walls The number and total thickness of ancestral walls in the mature middle lamella vary with the number of cell divisions and the location of the cell plates within a fusiform cambium daughter cell. The development of the canal complex and of the ray tissue associated with it can be traced by studying the cell wall ultrastructure.Symbols used in Figures A Canal complex axis - Ar Artifact - C Cytoplasmic remnant - CS Canal complex surface - D Canal - E Epithelial cells - F Intercell-wall fibrils - I Intermediate cells - L Cell lumen - M Intercellular membrane - M ₁ Intercellular membrane - M ₂ Intercellular membrane - M ₃ Intercellular membrane - ML Middle lamella - O Outer cell - P Primary cell wall - PL Part of an ancestral wall partially pulled from the canal complex surface - T Thin-walled portion of an epithelial cell - x Cleavage within middle lamella - y Cleavage along middle lamella-primary wall boundary The authors thank Drs. I. H. Isenberg and L. L. Winton, members of the thesis advisory committee, for their assistance in the conduct of the research, Miss Olga Smith for assistance in electron microscopy and for drawing Figs. 1, 5, and 7, and J. D. Hankey for assistance in photomicroscopy.     

A multi-species comparison of delta13C from whole wood, extractive-free wood and holocellulose

The stable carbon (C) isotope composition (delta13C) of tree rings is a powerful metric for reconstructing past physiological responses to climate variation. However, accurate measurement and interpretation are complicated by diagenesis and the translocation of compounds with distinct isotopic signatures. Isolation and analysis of cellulose minimizes these complications by eliminating variation due to biosynthetic pathways; however, isolation of cellulose is time-consuming and has no clear endpoint. A faster and better-defined analytical method is desirable. Our objectives were to determine if there is a direct relationship between the isotopic compositions of whole wood (WW), whole wood treated with solvents to remove mobile extractives (extractive-free wood; EF) and holocellulose (HC) isolated by extractive removal and subsequent bleaching. We also determined if total C concentration could explain the isotopic composition and variation among these three wood components of each sample. A set of wood samples of diverse phylogeny, anatomy and chemical composition, was examined. The mean offset or difference between HC and EF delta13C was 1.07 +/- 0.09 per thousand and the offset between HC and WW was 1.32 +/- 0.10 per thousand. Equivalence tests (with alpha = 0.05) indicated that the relationship between EF delta13C and HC delta13C had a slope significantly similar to 1 +/- 5.5%, whereas for the WW delta13C: HC delta13C relationship, the slope was significantly similar to 1 +/- 10.08%. A regression model using EF delta13C to predict HC delta13C had a slope of 0.97, which was not significantly different from unity (P = 0.264), whereas the regression for WW had a slope of 0.92 which was significantly different from unity (P = 0.0098). Carbon concentration was correlated with HC:WW offset and cellulose:EF offset (P = 0.0501 and 0.007, respectively), but neither relationship explained much of the variation (r2 = 0.12 and 0.14, respectively). We suggest that HC extraction is unnecessary for most analyses of tree-ring delta13C; a simple solvent extraction is a suitable alternative for many applications.     

Effect of pressure on organic acid production from Japanese beech treated in supercritical water

The effect of pressure as described by density was studied on organic acid production from Japanese beech (Fagus crenata) treated in supercritical water. At a reaction temperature of 380°C, the maximum yield of organic acids was 35% at a pressure of 30 MPa (density of water: 0.53 g/ml) for 1 min in a batch-type system. Furthermore, the yield of organic acids decreased with increasing reaction pressure. It was also found that fragmented products from sugars such as methylglyoxal and glycolaldehyde could be more easily converted to organic acids than dehydrated products such as furfural and 5-hydroxymethyl furfural. This result suggests that organic acids can be mainly derived from fragmented products. Part of this article was presented at the 55th Annual Meeting of the Japan Wood Research Society (2005), and at the 17th Annual Meeting of Japan Institute of Energy (2007)     

Studies on anti-diarrhoeal activity of Ficus hispida. Leaf extract in rats

Methanol extract of Ficus hispida L. showed significant inhibitory activity against castor oil-induced diarrhoea and PGE(2)-induced enteropooling in rats. It also showed a significant reduction in gastro-intestinal motility on charcoal meal test in rats. The results obtained establish the F. hispida leaf extract as an anti-diarrhoeal agent.     

Pathogenicity of Colletotrichum dematium isolated from current-year beech seedlings exhibiting damping-off

The pathogenicity of Colletotrichum dematium was evaluated as the agent causing post-emergence damping-off of current-year beech (Fagus crenata) seedlings. Current-year beech seedlings inoculated with C. dematium died within 20 days after both wound and non-wound inoculation, indicating that this fungus is pathogenic to current-year seedlings. C. dematium was frequently isolated from hypocotyls exhibiting damping-off at five different sites. The incidence of isolation was high and little variation occurred between sites and years. C. dematium was also detected in all litter samples examined although the incidence of detection differed depending on sites and/or detection methods. These results strongly indicate that C. dematium plays an important role in rapid seedling death during the early stages of beech regeneration.     

Accumulation of flavanols in yellowing beech leaves from forest decline sites

Beech leaves were sampled during two consecutive years from three sites characterized by forest decline. Both monomeric and oligomeric flavanols from green and yellowing leaves were determined quantitatively by reversed-phase, high-performance liquid chromatography (HPLC) combined with a chemical reaction detection technique (CRD). Yellowing leaves generally contained more than twice the quantity of flavanols than normal green leaves. The monomeric flavanols, epicatechin and (+)-catechin, comprised up to 80% of the total flavanol fraction. Histochemical staining with p-dimethylaminocinnamaldehyde was used to determine local deposition of flavanols, including insoluble oligomeric flavanols, in leaf tissues. The yellowing leaves stained intensely, whereas the green leaves stained lightly. Flavanol staining was strongest in the spongy parenchyma followed by the palisade cells. The upper epidermis stained more intensely than the lower epidermis. A 4-day treatment of small leaf pieces (5 x 5 mm) with 0.04 mM paraquat resulted in browning of the leaf pieces. The browning reaction was prevented when the paraquat treatment was carried out in the presence of 0.16 mM (+)-catechin, indicating an antioxidative property for this flavanol.     

Bark phenolics of American beech (Fagus grandifolia) in relation to the beech bark disease1

The amount of total extractable phenols was determined for bark sections obtained from behind cankers naturally induced by Nectria coccinea var. faginata and from behind mechanically inflicted wounds on stems of American beech. Healthy bark from susceptible trees was found to contain a similar level of phenolics as bark from trees determined to be resistant to the beech bark disease. Six months after wounding, wound-altered bark from susceptible trees was found to be lower in phenols than woundaltered bark from resistant trees. Inoculation of wounds with N. coccinea var. faginata resulted in decreased phenolic levels in bark sections nearest the wound surface, and increased phenolic levels in sections nearest the vascular cambium, several mm distant. Phenolic levels in injured or infected bark appear to follow similar patterns as those resulting from injury or infection of xylem tissues.     

Studies on antidepressant and antinociceptive effects of ethyl acetate extract from Piper laetispicum and structure-activity relationship of its amide alkaloids

Piper laetispicum C.DC. (Piperaceae), is an endemic climbing, glabrous plant distributed in the southern part of China. A novel alkaloid amide, Laetispicine, from it has been proven to possess antidepressant activity. In this present study, antidepressant and antinociceptive effects of the ethyl acetate extract (EAE) of P. laetispicum have been studied in forced swimming, open field, acetic acid writhing and formalin tests in KM mice. A significantly antidepressant-like effect was showing at doses of higher than 60 mg/kg, which was not due to an increase in locomotive activity. The EAE also presented an analgesic effect, in our studies. At lower doses (30 mg/kg) the antinociceptive effect was likely mediated via peripheral inflammation and changes in central processing, and at higher doses (120 mg/kg) that was due to both central and peripheral pathways. We also quantitatively analyzed the major components of EAE by HPLC and approached the structure–activity relationship between structure of amide alkaloids and its antidepressant activities. The antidepressant effective components of EAE might be Leatispiamide A and Laetispicine. In their molecular structures, the isolated double bond from benzene ring and conjugated double bond located at 2–3 and 4–5 were necessary for its antidepressant activity.     

Fate of nitrogen released from 15N-labeled litter in European beech forests

The decomposition and fate of 15N-labeled beech litter was monitored in three European beech (Fagus sylvatica L.) forests (Aubure, France; Ebrach, Germany; and Collelongo, Italy) for 3 years. Circular plots around single beech trees were isolated from roots of neighboring trees by soil trenching, and annual litterfall was replaced by 15N-labeled litter. Nitrogen was continuously released from the decomposing litter. However, over a 2-year period, this release was balanced by the incorporation of exogenous N. Released N accumulated mainly at the soil surface and in the topsoil. Microbial biomass remained almost constant during the experiment at all sites except for considerably lower values at Ebrach. The 15N enrichment of the microbial biomass increased strongly during the first year and then remained stable. The 15N released from the decomposing litter was rapidly detected in roots and leaves of the beech trees, increasing regularly and linearly over the course of the experiment. The uptake of litter-released 15N by the trees was reduced under conditions that reduced tree growth. Under these conditions, leaves and fine roots were the dominant N sinks, and little N was allocated to other plant parts. By contrast, N uptake and N allocation from leaves to stem and bark tissues increased when tree growth was enhanced. Budgets for 15N showed that 2 to 4% of litter-released N was incorporated into the trees, about 35% remained in the litter and about 50% reached the topsoil.     

Impact of water scarcity on spruce and beech forests

One of the greatest threats posed by ongoing climate change may be regarded the drought caused by changes in precipitation distribution. The aim of presented study was to characterize reactions to dry conditions and conditions without drought stress on gross primary production (GPP) and net ecosystem production (NEP) of spruce and beech forests, as these two species dominate within the European continent. Daily courses of GPP and NEP of these two species were evaluated in relation to an expected decrease in CO2 uptake during dry days. The occurrence of CO2 uptake hysteresis in daily production was also investigated. Our study was performed at Bily Kriz(spruce) and Stitna(beech) mountain forest sites during 2010–2012 period. We applied eddy covariance technique for the estimation of carbon fluxes, vapor pressure deficit and precipitation characteristics together with the SoilClim model for the determination of drought conditions, and the inverse of the Penman–Monteith equation to compute canopy conductance. Significant differences were found in response to reduced water supply for both species. Spruce reacts by closing its stomata before noon and maintaining a reduced photosynthetic activity for the rest of the day, while beech keeps its stomata open as long as possible and slightly reduces photosynthetic activity evenly throughout the entire day. In the spruce forest, we found substantial hysteresis in the light response curve of GPP. In the beech forest, the shape of this curve was different: evening values exceeded morning values.     

Bud dormancy in beech (Fagus sylvatica L.). Effect of chilling and photoperiod on dormancy release of beech seedlings

Two-year-old Fagus sylvatica L. seedlings were subjected to natural winter chilling or were overwintered in a heated greenhouse. Plants were then grown in controlled environment chambers with photoperiods of 9 or 13 h. Renewal of bud growth was found to be mainly determined by winter chilling. There was a slight interaction between chilling and photoperiod. Sprouting of apical buds took two to three times as long in unchilled plants as in chilled plants. Shoot elongation was influenced by chilling and was also greater in the 13-h photoperiod than in the 9-h photoperiod, but this may have been due at least in part to the higher irradiance. Chilling resulted in rapid dormancy loss and changed the growth pattern from basitonal to acrotonal.     

Cytotoxic activity of the root extract from Myoschilos oblongum

The crude methanolic root extract of Myoschilos oblongum exhibited a significant cytotoxic activity against PZ-HPV-7 human prostate cells. Furthermore, two esters of docosanol were isolated from the CH(2)Cl(2) extract.     

Estrogenic activity of the dichloromethane extract from Pueraria mirifica

Pueraria mirifica and its extracts are widely used as the ingredient(s) in many rejuvenating products. Up to now, the extract of P. mirifica roots that has been used in most studies, is the alcoholic extract. In the present study, we investigated the estrogenic activity using uterotropic and MCF-7 cell proliferation models of the dichloromethane extract as well as the water extract which was obtained from partitioning the ethanolic extract. The results indicated that among the three extracts, i.e. the ethanolic extract (PM1), the water extract (PM2) and dichloromethane extract (PM3), PM3 exhibited the most potent estrogenic activity in both models, followed by PM1. The extracts produced uterotropic activity associated with the increase of water content while uterotropic activity of 17beta-estradiol was related to the increase of muscle mass. The two isoflavonoids, genistein and daidzein, were not the major active phytoestrogens involving the estrogenic activity of these extracts.     

Methane production from organic acids obtained by supercritical water treatment of Japanese beech

Japanese beech (Fagus crenata) wood was treated in supercritical water at 380°C/30 MPa and 380°C/100 MPa. The hydrolysate (water-soluble portion) was found to contain the fragmented and dehydrated compounds of sugars and organic acids. Although organic acids are expected to be utilized for methane fermentation, the effects of the fragmented and dehydrated compounds of sugars on methane production are not known. The objective of this study is, therefore, to elucidate the potential of supercritical water treatment as a new pretreatment for methane production by evaluating the methane fermentability of the hydrolysate. From the methane fermentation tests for those model compounds with digested sludge, it is found that methane was produced not from the fragmented and dehydrated compounds but from the organic acids. The yield of methane from the hydrolysate obtained at 380°C/30 MPa was higher than that from the hydrolysate obtained at 380°C/100 MPa because the former contains more organic acids than the latter. The maximum yield of methane was seven times greater compared to the untreated wood, indicating that the supercritical water treatment is effective for enhancing the productivity of methane from wood.     

Chemical changes during the production of thermo-treated beech wood

Thermal treatments of beech wood with different temperature loads on the wood cause characteristic changes in the chemical composition. The determination of specific changes was carried out by means of suitable methods, both wet chemical and instrumental analyses. It could be confirmed that in addition to the degradation of polyoses, lignin, known as the thermally most stable compound, shows significant thermal alterations too.