Studies of crystallinity of Scots pine and Norway spruce cellulose

The variation in the mass fraction of crystalline cellulose (crystallinity of wood), the intrinsic crystallinity of cellulose, and the thickness of cellulose crystallites in early wood of Norway spruce [Picea abies (L.) Karst.], and Scots pine (Pinus sylvestris L.) grown in Finland were studied using wide angle X-ray scattering and nuclear magnetic resonance spectroscopy. The mass fraction of crystalline cellulose in wood increased slightly with the distance from the pith and was about 30±4% in mature wood of both species. The crystallinity of cellulose and the thickness of cellulose crystallites were almost constant for both species. The crystallinity of cellulose was 52±3% for both species and the average thickness of the cellulose crystallites was 32±1 Å and 31±1 Å for Norway spruce and Scots pine, respectively. The mass fraction of cellulose in wood, calculated from the crystallinity values, increased with the distance from the pith for both species.     

Quantitative trait locus (QTL) analysis of wood quality traits in <Emphasis Type="Italic">Eucalyptus nitens</Emphasis>

To identify the chromosomal regions affecting wood quality traits, we conducted a genome-wide quantitative trait locus (QTL) analysis of wood quality traits in Eucalyptus nitens. This information is important to exploit the full potential of the impending Eucalyptus genome sequence. A three generational mapping population consisting of 296 progeny trees was used to identify QTL associated with several wood quality traits in E. nitens. Thirty-six QTL positions for cellulose content, pulp yield, lignin content, density, and microfibril angle (MFA) were identified across different linkage groups. On linkage groups (LG)2 and 8, cellulose QTL cluster with pulp yield and extractives QTL while on LG4 and 10 cellulose and pulp yield QTLs cluster together. Similarly, on LG4, 5, and 6 QTL for lignin traits were clustered together. At two positions, QTL for MFA, a physical trait related to wood stiffness, were clustered with QTL for lignin traits. Several cell wall candidate genes were co-located to QTL positions affecting different traits. Comparative QTL analysis with Eucalyptus globulus revealed two common QTL regions for cellulose and pulp yield. The QTL positions identified in this study provide a resource for identifying wood quality genes using the impending Eucalyptus genome sequence. Candidate genes identified in this study through co-location to QTL regions may be useful in association studies.     

Morphogenesis of complex plant cell shapes: the mechanical role of crystalline cellulose in growing pollen tubes

Cellulose is the principal component of the load-bearing system in primary plant cell walls. The great resistance to tensile forces of this polysaccharide and its embedding in matrix components make the cell wall a material similar to a fiber composite. In the rapidly growing pollen tube, the amount of cellulose in the cell wall is untypically low. Therefore, we want to investigate whether the load-bearing function of cellulose is nevertheless important for the architecture of this cell. Enzymatic digestion with cellulase and inhibition of cellulose crystal formation with CGA (1-cyclohexyl-5-(2,3,4,5,6-pentafluorophenoxy)-1λ4,2,4,6-thiatriazin-3-amine) resulted in the formation of tubes with increased diameter in Solanum chacoense and Lilium orientalis when present during germination. In pre-germinated tubes, application of both agents resulted in the transient arrest of growth accompanied by the formation of an apical swelling indicating a role in the mechanical stabilization of this cellular region. Once growth resumed in the presence of cellulase, however, the cell wall in the newly formed tube showed increased amounts of pectins, possibly to compensate for the reduced amount of cellulose. Scanning electron microscopy of pollen tubes subjected to digestion of matrix polysaccharides revealed the mechanical anisotropy of the cell wall. In both Lilium and Solanum, the angle of highest stability revealed by crack formation was significantly below 45°, an indication that in the mature part of the cell cellulose may not the main stress-bearing component against turgor pressure induced tensile stress in circumferential direction.     

Adhesion of a <Emphasis Type="Italic">Pseudomonas putida</Emphasis> strain isolated from a paper machine to cellulose fibres

The adhesion to cellulose fibres of a strain of Pseudomonas putida isolated from a paper machine was studied under different environmental conditions. The physicochemical properties of both P. putida cells and cellulose fibres were also determined to better understand the adhesion phenomenon. Adhesion was rapid (1 min) and increased with time, cell concentration and temperature (from 25 to 40°C), indicating that bacterial adhesion to cellulose fibres is essentially governed by a physicochemical process. The P. putida cell surface was negatively charged, as shown by electrophoretic mobility measurements, and was hydrophilic due to a strong electron-donor character, as shown by the microbial adhesion to solvents method. Cellulose fibres were shown to be hydrophilic by contact angle measurements using the capillary rise method. These results suggest the importance of Lewis acid-base interactions in the adhesion process. In various ionic solutions (NaCl, KCl, CaCl₂ and MgCl₂), adhesion increased with increasing ionic strength up to 10–100 mM, indicating that, at low ionic strength, electrostatic interactions were involved in the adhesion process. An increase in the C/N ratio of the growth medium (from 5 to 90) decreased adhesion but this could not be related to changes in physicochemical properties, suggesting that other factors may be involved. In practice, temperature, ionic strength and nitrogen concentration must be taken into consideration to reduce bacterial contamination in the paper industry.     

Atomic layer deposition of titanium dioxide on cellulose acetate for enhanced hemostasis

TiO₂ films may be used to alter the wettability and hemocompatibility of cellulose materials. In this study, pure and stoichiometric TiO₂ films were grown using atomic layer deposition on both silicon and cellulose substrates. The films were grown with uniform thicknesses and with a growth rate in agreement with literature results. The TiO₂ films were shown to profoundly alter the water contact angle values of cellulose in a manner dependent upon processing characteristics. Higher amounts of protein adsorption indicated by blurry areas on images generated by scanning electron microscopy were noted on TiO₂-coated cellulose acetate than on uncoated cellulose acetate. These results suggest that atomic layer deposition is an appropriate method for improving the biological properties of hemostatic agents and other blood-contacting biomaterials.     

Enantioselective retardation of rac-propranolol from matrices containing cellulose derivatives

The dissolution characteristics of propranolol enantiomers from tablet formulations containing cellulose, or one of eight cellulose derivatives, were determined under a range of conditions. The derivatives examined were: cellulose tris(phenylcarbamate) (1), cellulose tris(2,3-dichlorophenylcarbamate) (2), cellulose tris 2,4-dichlorophenylcarbamate (3), cellulose tris(2,6-dichlorophenylcarbamate) (4), cellulose tris(2,3-dimethylphenylcarbamate) (5), cellulose tris(3,4-dichlorophenylcarbamate) (6), cellulose tris (3,5-dichlorophenylcarbamate) (7), cellulose tris(3,5-dimethylphenylcarbamate) (8). In water at 25°C, the release rates of (-)R-propranolol were generally greater than those of (-)-S-propranolol, although these differences were not always statistically significant; only compounds 5 and 8 demonstrated significant enantioselectivity. Using compound 8 in further experiments, statistically significant stereoselective dissolution of propranolol HCl was observed in buffer pH 7.4 at 25°C (intrinsic dissolution rates: 0.41 ± 0.01 mgcm²min⁻¹ for R-propranolol and 0.30 ± 0.02 mgcm²min⁻¹ for S-propranolol; P = 0.003). The cumulative amounts of enantiomers released at every time point were also found to be statistically significant (mean ratio R:S 1.25 ± 0.05). The observed low stereoselectivity of 8 with propranolol base was probably attributable to low solubility in pH 7.4 buffer, although stereoselective release did increase with time. This suggested that there is a relationship between stereoselectivity and contact time in an aqueous environment. Results also suggested that increased temperature may affect the release process as well as stereoselective interactions of 8 with individual enantiomers. To conclude, differential release of rac-propranolol from cellulose derivative matrices has been demonstrated, which supports the principle of stereoselective retardation as a potential means of stereoselective drug delivery for solid dosage forms. Chirality 9:307–312, 1997. © 1997 Wiley-Liss, Inc.     

Profiling of cellulose content in Indian seaweed species

Cellulose contents were estimated in 12 seaweed samples belonging to different families e.g. red, brown and green, growing in Indian waters. Each cellulose sample was fractionated to yield alpha (α) and beta (β) celluloses. Characterization was done using various analytical tools and results were validated by comparison with those of the cellulose obtained from Whatman filter paper No. 4. The greatest yields of cellulose (crude), α- and β-cellulose were obtained from Gelidiella acerosa (13.65%), Chamaedoris auriculata (9.0%) and G. acerosa (3.10%). G. acerosa was also found to contain relatively high amount of α-cellulose (8.19%). The lowest cellulose contents were recorded from Kappaphycus alvarezii (2.00%) and Sarconema scinaioides (2.1%), while the latter contained the lowest α-, and β-celluloses (1.0% and 0.30%, respectively). It appears that agarophytic and alginophytic algae contain high cellulose and α-cellulose contents, while the carrageenophyte contains low cellulose. The brown algae, in general contain high cellulose as well as α- and β-celluloses.     

Almost Pure Iα Cellulose in the Cell Wall of Glaucocystis

Crystalline features of cellulose microfibrils in the cell walls of Glaucocystis (Glaucophyta) were studied by combined spectroscopy and diffraction techniques, and the results were compared with those of Oocystis (Chlorophyta). Although these algae are grouped into two different classes, by the composition of their chloroplasts for instance, their cell walls are quite similar in size and morphology. The most striking features of their cellulose crystallites are that they have the highest cellulose I_α contents reported to date. In particular, the I_α fraction of cellulose from Glaucocystis was found to be as high as 90% from ¹³C NMR analysis. The mode of preferential orientation of cellulose crystallites in their cell walls is also interesting; equatorial 0.53-nm lattice planes were oriented parallel to the cell surface in the case of Glaucocystis, while the 0.62-nm planes were parallel to the Oocystis cell surface. Such a structural variation provides another link to the evolution of cellulose structure, biosynthesis, and its biocrystallization mechanism.     

Role of outer membrane components in the nonspecific binding ofEscherichia coli to cellulose

The involvement of lipopolysaccharide and outer membrane proteins in the binding ofEscherichia coli to cellulose was investigated. Cellulose binding was assayed in defined strains with or without O-antigenic polysaccharide and in mutants with defects in lipopolysaccharide core synthesis. Binding was also tested in strains lacking major outer membrane proteins. Optimal cellulose binding was exhibited by rough strains and was reduced to various extents in the presence of different O-antigens. Core defects also reduced but did not abolish binding to cellulose. Reduced binding was also found in mutants lacking OmpC protein, but OmpC/OmpA double mutants orompB mutants lacking OmpC and OmpF were not affected. Mutants with reduced cellulose binding were also isolated directly through selection of nonbinding populations after chromatography on cellulose columns. Each of the independent isolates derived fromE. coli K12 with reduced cellulose binding had multiple mutations, with additional phenotypic changes such as phage resistance, increased sensitivity to bile salts, or altered patterns of outer membrane proteins. These results suggest that no single receptor that could be altered by mutation was responsible for the binding ofE. coli to cellulose. Rather, the nonspecific binding of cellulose was more likely to be due to interaction with, or the combined activity of, several integral outer membrane components that could be masked by O-antigen.     

Synthesis of 20α- and 20β-Amino-3,5-pregnadiene Derivatives and Antimicrobial Activity

Cellulose production by Acetobacter strains is enhanced by the addition of a small amount of cellulose to the production culture. The effect of an endo-β-1, 4-glucanase from Bacillus subtilis on the cellulose production by Acebohacter xylinum BPR2001 was examined by adding various amounts of the purified glucanase to the culture. The addition of a small amount of this glucanase enhanced cellulose production. Furthermore, it reduced the amount of a polysaccharide called acetan produced. However, an active-site mutant enzyme of the glucanase, which showed no enzyme activity but still had cellulose-binding ability, had no effect on cellulose production. It was concluded, therefore, that the endoglucanase activity itself, but not the cellulose-binding ability, was essential for the enhancement of cellulose production. The structural properties of the cellulose produced in the presence of the endoglucanase were found to be almost identical to those of native bacterial cellulose.     

Mechanism of initial rapid rate retardation in cellobiohydrolase catalyzed cellulose hydrolysis

Despite intensive research, the mechanism of the rapid retardation in the rates of cellobiohydrolase (CBH) catalyzed cellulose hydrolysis is still not clear. Interpretation of the hydrolysis data has been complicated by the inability to measure the catalytic constants for CBH‐s acting on cellulose. We developed a method for measuring the observed catalytic constant (k^obs) for CBH catalyzed cellulose hydrolysis. It relies on in situ measurement of the concentration of CBH with the active site occupied by the cellulose chain. For that we followed the specific inhibition of the hydrolysis of para‐nitrophenyl‐β‐D ‐lactoside by cellulose. The method was applied to CBH‐s TrCel7A from Trichoderma reesei and PcCel7D from Phanerochaete chrysosporium and their isolated catalytic domains. Bacterial microcrystalline cellulose, Avicel, amorphous cellulose, and lignocellulose were used as substrates. A rapid decrease of k^obs in time was observed on all substrates. The k^obs values for PcCel7D were about 1.5 times higher than those for TrCel7A. In case of both TrCel7A and PcCel7D, the k^obs values for catalytic domains were similar to those for intact enzymes. A model where CBH action is limited by the average length of obstacle‐free way on cellulose chain is proposed. Once formed, productive CBH–cellulose complex proceeds with a constant rate determined by the true catalytic constant. After encountering an obstacle CBH will “get stuck” and the rate of further cellulose hydrolysis will be governed by the dissociation rate constant (k_off), which is low for processive CBH‐s. Biotechnol. Bioeng. 2010;106: 871–883. © 2010 Wiley Periodicals, Inc.     

Characterization of cellulose degrading bacteria from the larval gut of the white grub beetle Lepidiota mansueta (Coleoptera: Scarabaeidae)

The goal of this study is to identify and characterize the cellulose degrading microorganisms in the larval gut of the white grub beetle, Lepidiota mansueta. Thirty bacterial strains were isolated and tested for cellulolytic activity using soluble carboxymethyl cellulose (CMC) degrading assays. Of these strains, five (FGB1, FB2, MB1, MB2, and HB1) degrade cellulose. Cellulolytic activity was determined based on formation of clear zone and cellulolytic index on CMC plate media. The highest cellulolytic index (2.14) was found in FGB1. Partial 16S rDNA sequencing, morphological, and biochemical tests were used to identify and characterize the five isolates, all Citrobacter sp. (Enterobacteriaceae). This study identifies new cellulose degrading microorganisms from the larval gut of L. mansueta. The significance of identifying these strains lies in possible application in cellulose degradation.     

Cellulose production from<Emphasis Type="Italic">Gluconobacter oxydans</Emphasis> TQ-B2

Gluconobacter oxydans that produces the cellulose was isolated. In order to confirm the chemical features of cellulose, various spectrophtometeric analysis were carried out using electron microscopy, X-ray diffractogram, and CP/MAS¹³C NMR. The purified cellulose was found to be identical to that ofAcetobacter xylinum. For effective production of cellulose, the various carbon and nitrogen sources, mixture of calcium and magnesium ions, and biotin concentration were investigated in flask cultures. Among the various carbon sources, glucose and sucrose were found to be best for the production of cellulose, with maximum concentration of 2.41 g/L obtained when a mixture of 10 g/L of each glucose and sucrose were used. With regard to the nitrogen sources, when 20 g/L of yeast extract was used, the maximum concentration of bacterial cellulose was reached. The concentration of cellulose was increased with mixture of 2 mM of each Ca²⁺ and Mg²⁺. The optimum biotin concentration for the production of cellulose was in the range of 15 to 20 mg/L. At higher biotin concentration (25–35 mg/L), the bacterial cellulose production was lower.     

Comparative studies on hydrothermal pretreatment and enzymatic saccharification of leaves and internodes of alamo switchgrass

Hydrothermal pretreatment was performed on the leaves and internodes portions of Alamo switchgrass, Panicum virgatum L., to enhance the digestibility of cellulose towards cellulase. It was observed that extractives free leaves portion provided 18.1% lower pretreatment gravimetrical yield and 33.8% greater cellulose-to-glucose yield than internodes portion. The degree of polymerization (DP) and ultrastructure of cellulose were determined by gel-permeation chromatography and solid-state cross polarization/magic angle spinning ¹³C NMR experiments. The results suggested that hydrothermal pretreatment hydrolyzed amorphous cellulose and yielded a product enriched in paracrystalline cellulose. Furthermore, the DP of cellulose was reduced to one third of the origin value after hydrothermal pretreatment. The resulting biomass after pretreatment for leaves and internodes has similar cellulose ultrastructure and chemical profiles. The results of the enzymatic hydrolysis studies of cellulose suggest that the reduced DP of cellulose of pretreated switchgrass was an important factor influencing the enhanced digestibility of pretreated switchgrass.     

The rate of cellulose increase is highly related to cotton fibre strength and is significantly determined by its genetic background and boll period temperature

This experiment was conducted to study the relationship between the increase in cellulose content in developing cotton bolls and their final cotton fibre strength. The rate of cellulose increase over time was estimated using logistical regression, and the logistic equation parameters were then used to compare different cotton cultivars in different temperature environments. The increase in cellulose content followed a typical “S” curve, with the boll period time divided into slow-fast-slow stages. In different cultivars, the final fibre strength was closely related to the characters of the fast cellulose content increasing stage, negatively related to the maximal cellulose increasing rate (P < 0.05), and positively related to the duration of the fast cellulose content increasing stage (P < 0.01). In the same cultivar, low temperature reduced the maximal cellulose increasing rate and prolonged the duration of the fast cellulose increasing stage. The results indicate that, in diverse genetic background, long-lasting and tempered cellulose growth during the rapid cellulose increasing stage is of significant benefit to high strength fibre development. For closely related cotton cultivars, decreasing the maximal cellulose increasing rate and the termination of rapid cellulose increasing stage reduced fibre strength that often occurs when temperatures are low.     

Cross‐species functional complementation of cellulose synthase during the development of cellular slime molds

Cellulose is a major and important component of the extracellular matrix during the development of Dictyostelium discoideum. Upon starvation, solitary amoebae of D. discoideum gather and form fruiting bodies in which cells differentiate into stalk cells and spores. The stalk tubes and walls of spores and stalk cells are made of cellulose. In the genus Acytostelium, however, all cells are destined to become spores and the stalks comprise only a cellulose tube, suggesting species‐specific regulation of cellulose synthesis. In this study, we cloned a putative cellulose synthase gene (cesA) of Acytostelium subglobosum and performed comparative analyses with the D. discoideum cellulose synthase gene (dcsA). Although the deduced amino acid sequences were highly conserved between cesA and dcsA, the numbers of transmembrane spans preceding the catalytic domain were dissimilar; 2 and 3, respectively. Since ectopic expression of cesA in dcsA^?null cells failed to restore the developmental defects of the mutant, we constructed a series of chimerical genes for complementation analyses and found that the catalytic domain of cesA was functional in D. discoideum cells if the preceding transmembrane region was swapped with dcsA. The non‐functional products that contained the cesA‐derived transmembrane region were localized to lysosomes. These results indicate that the transmembrane region of cellulose synthase is essential for proper accumulation of cellulose during the development of D. discoideum and that its differential localization in A. subglobosum may be related to the characteristic morphogenesis in this species.     

Fibrobacter succinogenes S85 ferments ball-milled cellulose as fast as cellobiose until cellulose surface area is limiting

Fibrobacter succinogenes S85 grew rapidly on cellobiose (0.31 h⁻¹) and the absolute rate of increase in fermentation acids was 0.68 h⁻¹. Cultures that were provided with ball-milled cellulose initially produced fermentation acids and microbial protein as fast as those provided with cellobiose, but the absolute cellulose digestion rate eventually declined. If the inoculum size was increased, the kinetics decayed from first to zero order (with respect to cells) even sooner, but in each case the absolute rate declined after only 20 to 30% of the cellulose had been fermented. Congo red binding indicated that the cellulose surface area of individual cellulose particles was not decreasing, and the transition of ball-milled cellulose digestion corresponded with the appearance of unbound cells in the culture supernatant. When bound cells from partially digested cellulose were removed and the cellulose was re-incubated with a fresh inoculum, the initial absolute fermentation rate was as high as the one observed for undigested cellulose and cellobiose. Based on these results, cellulose digestion by F. succinogenes S85 appears to be constrained by cellulose surface area rather than cellulase activity per se. Received: 19 January 2000 / Received revision: 18 April 2000 / Accepted: 1 May 2000