Effect of heat treatment on extracellular enzymatic activities involved in beech wood degradation by Trametes versicolor

Effect of heat treatment on extracellular enzymes involved in wood degradation by Trametes versicolor was investigated. Heat-treated and untreated beech blocks were exposed to T. versicolor on malt-agar medium and extracellular enzymatic activities investigated. A strong ABTS oxidizing activity has been detected during the first stage of colonization in both cases, while cellulase activities are mainly detected in the case of untreated beech wood. Further investigations carried out on holocellulose, isolated using sodium chlorite delignification procedure and subjected to heat treatment or not, indicate that commercially available cellulases and xylanases are able to hydrolyse untreated holocellulose, while heat-treated holocellulose was not affected. All these data suggest that chemical modifications of wood components during heat treatment disturb enzymatic system involved in wood degradation.     

Histological changes in the cell wall structure during wood decay by Trametes hirsuta and Trametes versicolor in neem (Azadirachta Indica A. Juss)

ABSTRACT

Structural alterations in the wood cell walls of neem inoculated with by Trametes hirsuta and T. versicolor were studied by microscopic methods. In vitro decayed wood showed extensive weight loss of test blocks (26.7 and 41.38% by T. versicolor and T. hirsuta, respectively) at the end of 3 months. Selective delignification in the initial phase followed by simultaneous removal of lignin was evident in test blocks inoculated with both the species. The separation of middle lamellae and patches of cellulosic polysaccharides stained blue with Astra blue in the delignified region of the fiber wall during early stages indicates selective mode of decay. In contrast, the occurrence of erosion troughs with characteristic U-notch in tangential sections is a characteristic feature of simultaneous rot that was apparent after 3 months of incubation. The decay pattern occurred concomitantly in all the xylem elements irrespective of general resistance pattern shown by vessel and axial parenchyma cells. At an advance stage, both species of Trametes showed formation of erosion channels along the microfibrils angle of cellulose which is considered as characteristics of soft rot decay type. The sharing of white rot and soft rot decay pattern by both the fungi suggest a phylogenetic link between both groups of fungi.     

Degradation of lipophilic wood extractive constituents in Pinus sylvestris by the white-rot fungi Bjerkandera sp. and Trametes versicolor

The white-rot fungi Trametes versicolor and Bjerkandera spp. are among the most frequent decomposers of angiosperm wood in forest ecosystems and in wood products in service. Wood extractives have a major impact on wood properties and wood utilization. This work evaluated the ability of two white-rot fungal strains (Bjerkandera sp. strain BOS55 and T. versicolor strain LaVec94-6) to degrade the main lipophilic extractive constituents in Scots pine (Pinus sylvestris L.). The time course of wood decay and wood extractive degradation was monitored in stationary batch assays incubated for eight weeks. The strains tested eliminated high levels of total resin, 34 to 51% in two weeks. Wood triglycerides were the most readily degraded extractive components (over 93% elimination in only two weeks). Free fatty acids and resin acids, which are potential fungal inhibitors, were also rapidly decomposed by the fungal strains. Sterols were used more slowly, nonetheless, the fungal degradation of this extractive fraction ranged from 50 to 88% after four weeks. Received 19 March 1999     

Characterization of acetylated wood decayed by brown-rot and white-rot fungi

The objective of this study was to characterize the decay of acetylated wood due to brown-rot and white-rot fungi by analysis of chemical composition, X-ray measurements, and¹³C-NMR spectroscopy. The decay by brown-rot fungus became inhibited at a weight percent gain (WPG) due to acetylation of more than 10%, and the mass loss (LOSS) due to decay became zero at a WPG of about 20%. The LOSS due to white-rot fungus decreased slowly with the increase in WPG, reaching zero at a WPG of about 12%. The losses of lignin by brown-rot decay increased initially with the decrease in LOSS owing to the progressing acetylation and then decreased at a LOSS of less than 60%. Polysaccharides were more easily decomposed than lignin during the decay of acetylated wood due to brown-rot fungus. The losses of both components due to white-rot decay decreased as the LOSS decreased with progressing acetylation. The white-rot fungus tended to preferentially decompose the lignin during the decay of acetylated wood. The brown-rot fungus decomposed the cellulose in the crystalline region to a large degree when the LOSS was more than 40%, whereas the white-rot fungus decomposed the crystalline region and the noncrystalline region in acetylated wood to the same degree. The brown-rot fungus preferentially decomposed unsubstituted xylose units in acetylated wood and partly decomposed the mono-substituted xylose units. It was suggested that the mono- and disubstituted cellulose were partly decomposed by brown-rot fungus.This paper was presented at the 46th and 47th annual meetings of the Japan Wood Research Society at Kumamoto and Kochi in April 1996 and April 1997, respectively     

An investigation of mercerization in decayed oak wood by a white rot fungus (Lentinula edodes)

The crystal transformation of cellulose I to cellulose II during alkali swelling was investigated in decayed oak wood that was used for shiitake mushroom cultivation and the results were compared with those of sound wood using X-ray diffraction analysis and ultraviolet microscopy. During mercerization, the sapwood cellulose of decayed wood was easily transformed into Na-cellulose I and then Na-cellulose I was easily converted into cellulose II after washing and drying. The sapwood cellulose of sound wood was converted more slowly to Na-cellulose I and very little Na-cellulose was converted to cellulose II. Na-cellulose I of sound wood can be reconverted to cellulose I during washing and drying. Therefore, it could be concluded that lignin prevented the alkali swelling of wood cellulose and the transformation from cellulose I to cellulose II. The decay of crystalline cellulose might cause an increase in the susceptibility of alkali swelling, so that the degree of mercerization may be also affected.     

Cytochemical localization of cellulases in decayed and nondecayed wood

Summary Enzyme assay showed that the commercial cellulase from Trichoderma reesei degraded several polysaccharide substrates; highest activity was on xylan. Transmission electron micrographs showed that the T. reesei cellulase degraded nondecayed wood extensively; the attack always progressed from the point of contact into the cell wall. Cytochemically prepared wood that had been decayed by Poria placenta showed uniform distribution of electron-dense particles throughout the walls; the same results were observed with added T. reesei cellulase. In wood decayed by Ganoderma applanatum without or with added T. reesei cellulase, the progress of degradation was similar to that in nondecayed wood. Negatively stained T. reesei cellulase had 3–7 nm diameter for the smallest particles; the smallest diamaters of electron-dense particles in wood ranged from 3–7 nm. This suggests that the electron-dense particles probably are cellulase molecules.     

Alkaline degradation of spruce and beech wood

Summary The alkaline delignification of spruce and beech wood is accomplished in three phases. The first phase results in a decrease of yield down to 83...80%, the second one in a decrease from 83...80% to 64...61% and the third one in a yeld drop from 64...61% to 50...46%. The amount of polysaccharides removed in the first phase was 22%, in the second phase 5...6% and in the third phase 10% of the total polysaccharides in the original wood. The amount of lignin removed in the first phase of delignification was 9...11% in the second phase 53...54% and in the third phase 28% of the total lignin in the original wood. The polysaccharides extracted in the second and third phase amounted to 70...78% of the polysaccharides removed in the first delignification phase. For an entire characterization of the delignification reaction not only the course of lignin removal is of importance but also the course of the polysaccharide extraction. The results show that for a two-stage alkali-oxygen-cooking technology the optimum yield after initial partial alkaline delignification lies in the range of 64...61%.     

Analysis of beech wood fatty acids by supercritical acetone extraction

Summary Soxhlet and supercritical fluid extracts of freshly cut oriental beech (Fagus orientalis) have been obtained using acetone. While the yield of the Soxhlet extract was 2.54% the yield of the supercritical acetone extract obtained at 240°C and 6.0–6.5 MPa was 9.55% (dry wood basis). The fatty acids present in the extracts were separated by chemical and Chromatographic methods and analysed by combined gas chromatography and mass spectrometry. Linoleic acid was the major fatty acid in both extracts.     

Non-destructive monitoring of early stages of white rot by Trametes versicolor in Fraxinus excelsior

? Non-destructive detection of fungal decay in living trees is relevant for forest management of valuable species, hazard tree assessment, and research in forest pathology. A variety of tomographic methods, based on stress wave timing, radioactive radiation, or electrical resistivity have been used to detect decay in standing trees non-destructively. But apart from mobile gamma ray computed tomography (Habermehl and Ridder, 1993) which is virtually unavailable, the detection of incipient stages of decay is still not possible.

? Wood moisture and electrolyte content influence the electric resistivity of wood. Both are changed by fungal decay. Therefore electric resistivity tomography (ERT) should detect decay in its early stages. Then it could be used to monitor the spatial and temporal progress of degradation.

? We infected four Fraxinus excelsior trees with Trametes versicolor using wooden dowels and measured two-dimensional electric resistivity tomograms 3, 10, 13 and 21 months after infection. Immediately after the last electric resistivity measurement trees were felled for further analyses of stem cross-sections. Wood moisture content and raw density had significantly increased in infected areas, but dry density had not significantly changed after 21 months. Areas of very low electric resistivity around the infected wounds correlated very well with infected wood in the stem cross-sections. Increasing areas of low electric resistivity around the infected wounds during consecutive measurements indicate increasing areas of infected wood.

? We conclude that the growth of white rot by Trametes versicolor can be monitored with electric resistivity tomography (ERT) beginning from incipient stages, even before wood density decreases. ERT could therefore be a powerful research tool for decay dynamics as well as a method for diagnosing wood decay in forestry and arboriculture.

     

优化彩绒革盖菌产漆酶条件及染料脱色研究

对一株漆酶高产白腐菌———彩绒革盖菌的培养和产酶条件,包括培养基初始pH值、金属离子Cu2 和Mn2 的含量,以及诱导剂的种类、添加量和添加时间等相关因素进行了优化。结果发现该菌种适宜在pH值3.5~5.7环境下生长并合成漆酶,培养基中添加缓冲液可以获得更高的酶活,但不加则有较高的酶产率;铜和锰可以显著提高漆酶活力,当以0.1 mmol/L 2,5-二甲基苯胺为诱导剂时,培养基中宜含0.008~0.08 mmol/L Cu2 ,0.01 mmol/L左右的Mn2 。在5种常用漆酶诱导剂中,2,5-二甲基苯胺的效果最佳,且应在菌种进入对数生长期时加入效果最好,其最适添加浓度为0.4 mmol/L,此时最适产酶Cu2 浓度为0.4 mmol/L。研究显示漆酶在30和40℃下保持6 h后残余酶活分别为原来的95%和80%;60℃条件下,该酶保温1 h后的残余活力仅为原来的14%。该酶的最适反应温度是65℃,最适反应pH值为2.4~3。对20种常用染料进行的脱色研究显示,在pH值5、40℃条件下,该粗漆酶液可以直接作用于13种工业染料。     

The chemistry of decayed aspen wood and perspectives on its utilization

Summary The objectives of this research were to investigate the proportion of decayed wood in mature aspen stems, its chemical composition and its potential utility as a fuel or as a substrate for conversion to fine chemicals as part of an integrated utilization scheme. Three sound and ten decayed aspen stems were sampled from a boreal forest site. Stem analysis indicate that on average, 20% of the merchantable stem volume was in advanced decay and that considerable sound wood recovery was possible. Wood specific gravity and chemical composition were determined. The holocellulose content (volumetric basis) in advanced decayed wood was reduced by 67%. Thermal analysis of the wood using a differential scanning calorimeter provided graphical evidence of a different sequence of events occurring during the combustion of decayed wood and a resulting heat content per unit weight that was 40% higher than that of sound aspen wood. A higher degree of enzymatic hydrolysis was attainable with white-rotted aspen wood. Approximately 62% of the theoretical glucose yield was obtained from decayed aspen wood after alkali-peroxide pretreatment followed by a 12 hour hydrolysis using technical grade enzymes. The above information is used to elucidate future opportunities for wood recovery and energy production from decayed wood resources.The authors would like to thank the Ontario Ministry of Natural Resources, Kirkland Lake for their cooperation; and for the technical assistance by Sally Krigstin, John Leigh, Samir Konar, Ganesh Deka and Doug Charles. We would also like to thank Dr. Morris Wayman, University of Toronto, for his advice and inspiration. We are especially grateful to the following persons and companies for supplying us with enzymes and their associated technical literature: Mr. John Bayard of Van Waters & Rogers Ltee, Canada, representing NOVO Industri, Denmark; Mr. Ian Hodge representing the Miles Biotechnology Group, Canada; and Dr. Gunther Eckert, B.A.S.F., Germany for obtaining and forwarding the products and information of Rohm GmbH, Germany. This work has been partially funded by the Edward Johnson Fellowship, University of Toronto     

Anatomical characterization of decayed wood in standing light red meranti and identification of the fungi isolated from the decayed area

To further our understanding of wood decay in living light red meranti (Shorea smithiana) trees, microscopic characteristics of the cell and cell wall degradations of S. smithiana wood in the presence of the decay fungi, the identity of the causal fungi, and the decay potential and pattern by an isolated fungus were investigated. Cell wall degradations, including cell wall thinning, bore holes formation, rounded pit erosion, and eroded channel opening were clearly observed under light and scanning electron microscopy. In transverse view, many large voids resulting from a coalition of degraded wood tissue appeared in the decayed canker zone. All these observations suggest the well-known simultaneous decay pattern caused by white-rot fungi. By phylogenetic analysis based on the sequences of internal transcribed spacer region of ribosomal DNA, a basidiomycete fungus isolated from the decayed wood was identified as Schizophyllum commune. The degradation caused by this fungus on sound S. smithiana wood in an in situ laboratory decay test was classified as the early stage of simultaneous decay, and showed a similar pattern to that observed in the wood samples naturally decayed.     

Anatomical changes in the cell-wall structure of Leucaena leucocephala (Lam.) de Wit as caused by the decay fungi Trametes versicolor and Trametes hirsuta

The structural changes in the cell wall and delignification pattern caused by Trametes versicolor and Trametes hirsuta in the sap wood of Leucaena leucocephala were examined by light and confocal laser scanning microscopy. The in vitro decay test was conducted for 12 weeks. Both species of Trametes used in this study caused anatomical characteristics specific to simultaneous white rot. Formation of boreholes, erosion troughs, erosion channels with U-shaped notches in tangential sections and thinning of cell walls were evident in the wood inoculated with each of the fungal species. Cell separation due to removal of middle lamellae occurred at the early stages of infection (30 d) with T. versicolor. In contrast, middle lamellae remained intact in wood inoculated with T. hirsuta and showed cell separation due to degradation of the outer layer of the secondary wall and degradation of the middle lamellae observed only in severely decayed wood at late stages. Confocal microscopy revealed the delignification pattern particularly from cell corners and vessel walls at an advanced stage of decay, indicating strong ligninolytic activity of both species in the sapwood of L. leucocephala.     

Microstructural and microchemical studies of wood of Swietenia mahagoni Linn. decayed by Fomes durissimus Lloyd

Histopathological studies of wood of Swietenia mahagoni Linn. decayed by Fomes durissimus Lloyd reveal the presence of mycelia in all the wood clements and considerable thinning of cell walls and tissue disintegration due to utilization of both lignin and ccllulosc at advanccd stages of decay. Microchcmical tcsts and quantitative analysis of lignin nnd ccllulosc corroborated the results of histopathological studies indicating that the fungus mainly consumes thc lignin along with some amount of cellulose.     

Determining moisture-dependent elastic characteristics of beech wood by means of ultrasonic waves

The present study investigates the influence of moisture content on the elastic characteristics of beech wood (Fagus sylvatica L.) by means of ultrasonic waves. A set of elastic engineering parameters (i.e. three Young’s moduli, three shear moduli and six Poisson’s ratios) is determined at four specific moisture contents. The results reveal the significant influence of the moisture content on the elastic behaviour of beech wood. With the exception of some Poisson’s ratios, the engineering parameters decrease with increasing moisture content, indicating a decline in stiffness at higher moisture contents. At the same time, wood anisotropy, displayed by the two-dimensional representation of the velocity surface, remains almost unchanged. The results prove that the ultrasonic technique is suitable for determining the elastic moduli. However, non-diagonal terms of the stiffness matrix must be considered when calculating the Young’s moduli. This is shown experimentally by comparing the ultrasonic Young’s moduli calculated without, and allowing for, the non-diagonal terms. While the ultrasonic technique is found to be reliable to measure the elastic moduli, based on the measured values, its eligibility to measure the Poisson’s ratios remains uncertain.     

Evaluation of thermally modified beech and spruce wood and their properties by FT-NIR spectroscopy

Quality assessment of thermally modified spruce (Picea abies (L.) Karst) and beech (Fagus sylvatica L.) wood and of the corresponding reference samples was carried out by means of non-destructive FT-NIR spectroscopic measurements and PLS regression. Oven-dry and basic density as well as MOE and MOR determined by 3-point bending tests were evaluated. The focus was put on specimens produced from material that had been thermally modified in an industrial scale kiln. Modelling results range from poor to very good. The results of the spectra taken from the spruce samples resulted in better prediction results than the spectra of the beech samples. This could be due to different proveniences or variation in the industrial modification process. The results indicate that FT-NIR surface measurements of sound thermally modified wood samples could be applied to evaluate several characteristics before and after the modification process. The method could be used for screening during pre-sorting of thermally modified wood.     

Measurements of fungal wood decay on Scots pine and beech by means of X-ray microdensitometry

In this work, we present a fast and promising method to evaluate the natural durability of wood based on X-ray microdensitometry. Tested on beech and Scots pine wood samples, our findings show that this methodology and the traditional EN standards methodology based on mass loss are strongly correlated. X-ray methodology is less time consuming (we can detect the effectiveness of the attack within 5–6 weeks) and less expensive (very cheap plastic Petri dishes instead of the expensive glass Kolle flasks); moreover, the proposed method allows to thoroughly examine the phases and the kinetics of the fungal attack, and to investigate the spatial repartition of the attack within the samples due to the low thickness of the sample.     

Effect of severe thermal treatment on spruce and beech wood lignins

? The structure, proportion and mode of assembly of lignin, celluloses and hemicelluloses have marked effects on the reaction mechanisms during thermal treatment and therefore have a strong influence on the quality of the final product. The effect of treatment conditions, including severe conditions (up to 553 K) and treatment duration (up to 8 h) on the structure of native spruce and beech lignins was studied.

? Lignin content was determined by the Klason method and lignin structure was evaluated by thioacidolysis.

? The results highlighted the strong reactivity of the native spruce and beech lignins towards severe heat treatments. The distinct susceptibility of syringyl (S) and guaiacyl (G) units towards thermal treatment is confirmed by comparing the data for beech and spruce samples. The most severe treatment of spruce wood (280 °C) induced a dramatic enrichment in lignin content together with the almost complete disappearance of G lignin units, whereas a more moderate treatment substantially changed lignin structure by degradation reactions that affect the p-hydroxyphenyl (H) and G lignin units similarly.

? Thioacidolysis revealed that the thermal treatment induces the appearance of vinyl ether structures in spruce lignins. The decreased yield of the G and S thioacidolysis monomers reflects the progressive disappearance of G and S lignin units only involved in β-O-4 bonds and the formation of condensed linkages in proportions related to treatment severity. In severe conditions, β-O-4 linked S units are more degraded than their G homologues.

     

Assessment of resonance wood quality by comparing its physical and histological properties

The quality of wood used for music instrument making (resonance wood) is determined by assessing six physical properties: density, modulus of elasticity, sound velocity, radiation ratio, emission ratio, and loudness index. This can easily be done by means of measurements of the resonance frequency and the corresponding damping factor. The method described here is based on vibrational analyses, adapted from standard non-destructive testing of solid material, so as to provide information both for scientific studies and for violin making. The above six properties were assessed in samples of resonance wood of different quality and in normal (control) wood of Norway spruce and sycamore. The differences observed between the samples correlated with anatomical or histological characteristics of the wood. A sample of best-quality Norway spruce resonance wood showed a high radiation ratio in the axial direction, which correlated with the presence of small wood cells with thin cell walls. In “curly maple”, a high sound velocity in the radial direction correlated with the presence of broad xylem rays. The influence of external factors like wood moisture content or the geometry of the system is discussed within the context of the present study.