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PubMed1996

Binding of the cellulose-binding domain of exoglucanase Cex from Cellulomonas fimi to insoluble microcrystalline cellulose is entropically driven.

Isothermal titration microcalorimetry is combined with solution-depletion isotherm data to analyze the thermodynamics of binding of the cellulose-binding domain (CBD) from the beta-1,4-(exo)glucanase Cex of Cellulomonas fimi to insoluble bacterial microcrystalline cellulose. Analysis of isothermal titration microcalorimetry data against two putative binding models indicates that the bacterial microcrystalline cellulose surface presents two independent classes of binding sites, with the predominant high-affinity site being characterized by a Langmuir-type Ka of 6.3 (+/-1.4) x 10(7) M-1 and the low-affinity site by a Ka of 1.1 (+/-0.6) x 10(6) M-1. CBDCex binding to either site is exothermic, but is mainly driven by a large positive change in entropy. This differs from protein binding to soluble carbohydrates, which is usually driven by a relatively large exothermic standard enthalpy change for binding. Differential heat capacity changes are large and negative, indicating that sorbent and protein dehydration effects make a dominant contribution to the driving force for binding.

Binding Sites↗

PubMed2003

Three distinct rice cellulose synthase catalytic subunit genes required for cellulose synthesis in the secondary wall.

Several brittle culm mutations of rice (Oryza sativa) causing fragility of plant tissues have been identified genetically but not characterized at a molecular level. We show here that the genes responsible for three distinct brittle mutations of rice, induced by the insertion of the endogenous retrotransposon Tos17, correspond to CesA (cellulose synthase catalytic subunit) genes, OsCesA4, OsCesA7 and OsCesA9. Three CesA genes were expressed in seedlings, culms, premature panicles, and roots but not in mature leaves, and the expression profiles were almost identical among the three genes. Cellulose contents were dramatically decreased (8.9%-25.5% of the wild-type level) in the culms of null mutants of the three genes, indicating that these genes are not functionally redundant. Consistent with these results, cell walls in the cortical fiber cells were shown to be thinner in all the mutants than in wild-type plants. Based on these observations, the structure of a cellulose-synthesizing complex involved in the synthesis of the secondary cell wall is discussed.

Amino Acid Sequence↗

PubMed1992

Ethanol production from cellobiose, amorphous cellulose, and crystalline cellulose by recombinant Klebsiella oxytoca containing chromosomally integrated Zymomonas mobilis genes for ethanol production and plasmids expressing thermostable cellulase genes from Clostridium thermocellum.

The Zymomonas mobilis genes for ethanol production have been integrated into the chromosome of Klebsiella oxytoca M5A1. The best of these constructs, strain P2, produced ethanol efficiently from cellobiose in addition to monomeric sugars. Utilization of cellobiose and cellotriose by this strain eliminated the requirement for external beta-glucosidase and reduced the amount of commercial cellulase needed to ferment Solka Floc SW40 (primarily crystalline cellulose). The addition of plasmids encoding endoglucanases from Clostridium thermocellum resulted in the intracellular accumulation of thermostable enzymes as coproducts with ethanol during fermentation. The best of these, strain P2(pCT603T) containing celD, was used to hydrolyze amorphous cellulose to cellobiose and produce ethanol in a two-stage process. Strain P2(pCT603T) was also tested in combination with commercial cellulases. Pretreatment of Solka Floc SW40 at 60 degrees C with endoglucanase D substantially reduced the amount of commercial cellulase required to ferment Solka Floc. The stimulatory effect of the endoglucanase D pretreatment may result from the hydrolysis of amorphous regions, exposing additional sites for attack by fungal cellulases. Since endoglucanase D functions as part of a complex in C. thermocellum, it is possible that this enzyme may complex with fungal enzymes or bind cellulose to produce a more open structure for hydrolysis.

Cellobiose↗

PubMed1998

Analysis of molecular size distributions of cellulose molecules during hydrolysis of cellulose by recombinant Cellulomonas fimi beta-1,4-glucanases.

Four beta-1,4-glucanases (cellulases) of the cellulolytic bacterium Cellulomonas fimi were purified from Escherichia coli cells transformed with recombinant plasmids. Previous analyses using soluble substrates had suggested that CenA and CenC were endoglucanases while CbhA and CbhB resembled the exo-acting cellobiohydrolases produced by cellulolytic fungi. Analysis of molecular size distributions during cellulose hydrolysis by the individual enzymes confirmed these preliminary findings and provided further evidence that endoglucanase CenC has a more processive hydrolytic activity than CenA. The significant differences between the size distributions obtained during hydrolysis of bacterial microcrystalline cellulose and acid-swollen cellulose can be explained in terms of the accessibility of beta-1,4-glucan chains to enzyme attack. Endoglucanases and cellobiohydrolases were much more easily distinguished when the acid-swollen substrate was used.

Actinomycetales↗

PubMed2006

Cellulose nanocrystal-filled carboxymethyl cellulose nanocomposites.

Polymer nanocomposites are one of the important application areas for nanotechnology. Naturally derived organic nanophase materials are of special interest in the case of polymer nanocomposites. Carboxymethyl cellulose is a polyelectrolyte derived from natural materials. It has been extensively studied as a hydrogel polymer. Methods to modify the mechanical properties of gels and films made from CMC are of interest in our lab and in the commercial marketplace. The effect of nano-sized fillers on the properties of CMC-based composites is of interest in the development of novel or improved applications for hydrogel polymers in general and CMC in particular. This project investigated cellulose nanocrystals (CNXLs) as a filler in CMC and compared the effects to microcrystalline cellulose (MCC). The composite material was composed of CMC, MCC or CNXL, with glycerin as a plasticizer. CNXL and MCC concentrations ranged from 5% to 30%. Glycerin concentrations were kept constant at 10%. CNXLs improved the strength and stiffness of the resulting composite compared to MCC. In addition, a simple heat treatment was found to render the nanocomposite water resistant.

Absorption↗

PubMed2003

Properties of microcrystalline cellulose and powder cellulose after extrusion/spheronization as studied by fourier transform Raman spectroscopy and environmental scanning electron microscopy.

In this study, the effect of powder cellulose (PC) and 2 types of microcrystalline cellulose (MCC 101 and MCC 301) on pellet properties produced by an extrusion/spheronization process was investigated. The different investigated types of cellulose displayed different behavior during the extrusion/spheronization process. Pure PC was unsuitable for extrusion, because too much water was required and the added water was partly squeezed during the extrusion process. In contrast, MCC 101 and MCC 301 were extrudable at a wide range of water content, but the quality of the resulting products varied. In the extrusion/spheronization process, MCC 101 was the best substance, with easy handling and acceptable product properties. The properties of the extrudates and pellets were determined by Fourier transform (FT) Raman spectroscopy and environmental scanning electron microscopy (ESEM). FT-Raman spectroscopy was able to distinguish between the original substances and also between the wet and dried extrudates. The particle sizes of the raw material and of the extrudates were determined by ESEM without additional preparation. For MCC, the size of the resulting particles within the extrudate or pellet was smaller. However, in the extrudates of PC, changes in particle size could not be observed.

Cellulose↗

PubMedDate not supplied

Use of cellulose ether containing excipients with microcrystalline cellulose for the production of pellets containing metformin hydrochloride by the process of extrusion-spheronization.

The project is concerned mainly with the evaluation of two cellulose ether containing excipients, Aquacoat WG and Avicel 955 MCC for the improved extrusion-spheronization of metformin hydrochoride. Factorially designed experiments subject to statistical analyses were employed and products obtained were evaluated by sieve, packing density and image analysis, scanning electron microscopy and dissolution testing at pH 6.8. Aquacoat WG did not improve the ease of spheronization of drug mixes containing microcrystalline cellulose wetted with the optimum level of water. However, Avicel 955 MCC, which is a new experimental excipient containing 95% microcrystalline cellulose and 5% methylcellulose, did aid ease of spheronization facilitating acceptable yield of good spheres with high drug loadings (70%). Avicel 955 MCC containing drug mixes were more tolerant to minor alterations in level of hydration and yielded spheres which showed a small retardation of drug release despite the very high solubility of metformin hydrochloride.

Cellulose↗

PubMed1992

Permeability of cellulosic and non-cellulosic membranes to endotoxin subunits and cytokine production during in-vitro haemodialysis.

The possibility of endotoxin transfer across haemodialysis membranes remains a controversial issue. Additional concern has arisen because of the recent introduction in clinical practice of highly permeable, synthetic dialysis membranes and of bacteria-contaminated bicarbonate concentrate with potential short-term and long-term hazards for haemodialysis (HD) patients. Therefore, we performed experiments in an in-vitro dialysis recirculation system using three different types of HD membranes, namely standard regenerated cellulose (Cuprophan, CU), polyacrylonitrile AN-69 (PAN), and polysulphone F-60 (PS). When radiolabelled lipopolysaccharide (125I M-LPS) from E. coli, together with 10 micrograms/ml unlabelled LPS, was added to the recirculating solution in the dialysis compartment, radioactivity could be detected in the blood compartment after 15 min and increased progressively with time up to respectively 6.7% (CU), 10.3% (PAN), and 10.3% (PS) of initial activity on the dialysate side. The addition of albumin to the solution on the blood side led to a decreased permeability of radioactivity (7.3% vs 10.3%), compared to the absence of albumin (tested only for PS membrane). Furthermore, 73% of 125I M-LPS transferred across the PS membrane in the presence of albumin was TCA-precipitable. In contrast, free iodine (Na 125I) incubated in an albumin-containing solution did not precipitate with albumin after the addition of TCA (precipitation of only 0.6%). Moreover, kinetics of transmembranous transfer of Na-125I were strikingly different from that of 125I M-LPS. Analysis by the method of sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) of the blood side solution, after LPS addition in the dialysis solution and 30 min of back-filtration, revealed the presence of several silver-stainable and autoradiographic bands of low-molecular-weight range, probably LPS fragments. Finally, the presence of LPS in the dialysate compartment led to a moderate increase in interleukin 1 (IL-1) and tumour necrosis factor alpha (TNF) concentrations in plasma as well as in monocyte culture supernatants after isolation from recirculating normal human whole blood exposed to CU, PAN, or PS membrane. In conclusion, our study provides evidence for the permeation of low-molecular-weight LPS subunits across cellulosic and non-cellulosic HD membranes. The clinical significance, if any, of such a transfer has, however, still to be demonstrated.

Cellulose↗

PubMed1992

[The use of bead cellulose for controlled drug liberation. 5. Binding of benzocaine as a model drug to dialdehyde-bead cellulose and its in vitro liberation].

In the case of ionic binding of drugs to bead cellulose and its derivatives there was no pronounced retardation obvious. Because of this lack benzocaine as a model drug was covalent bound to dialdehyde bead cellulose by an azomethine bond in analogy with enzyme immobilisation methods. The rate of liberation in phosphate buffer was low and incomplete compared with the dissolution rate of pure benzocaine under the same conditions. A further decrease of liberation rate was obtained by reduction of the azomethine to the amine bond. Retardation of drug liberation strictly speaking was not achieved. The main amount of liberated drug was liberated during the first 20 min, but after 6 h the rate was still under 50% of the drug available. The physical properties of the beads like spherical shape and porosity are not significantly influenced by oxidation of cellulose and by loading with drug.

Benzocaine↗

PubMed1982

[Enzymatic hydrolysis of cellulose. Inactivation and stabilization of the enzymes of the cellulose complex].

The thermal inactivation of the individual cellulolytic components (endoglucanase, EC 3.2.1.4; exoglucosidase, EC 3.2.1.74; cellobiase, EC 3.2.1.21) from the fungi Trichoderma reesei, T. viride, T. lignorum an Aspergillus foetidus has been studied without resolution of the cellulase complexes. The kinetics of the thermal inactivation follow the first order for cellobiase of Asp. foetidus alone and show a more complex picture which is typical for a number of isoenzymes of different thermal stabilities for other cellulolytic components of all the cellulases under study. It was shown that selective elimination of acid proteinase from the cellulase preparations by affinity chromatography did not affect the time course of the thermal inactivation. Covalent attachment of the cellulases to porous glass and to some soluble high polymer supports only resulted in slight stabilizing effects (200-250%). Some polymer effectors (e. g. polyethylene glycols with Mr of 4000 and 40,000 and maltodextrins), as well as the end products of enzymatic hydrolysis of cellulose (i. e. glucose) did not affect the thermal stability of the cellulases under study. In some cases cellulose itself (cotton, CM-cellulose) produced a thermostabilizing effect (3-4-fold) on cellulolytic enzymes.

Aspergillus↗

PubMed1981

The best coating material for hemoperfusion: comparation of cellulose nitrate with cellulose acetate and derivatives.

In this study the clearance characteristics of the two most widely used membrane systems (cellulose nitrate and cellulose acetate) were compared in order to obtain the best coating material for hemoperfusion. The columns containing uncoated or coated charcoal granules were tested using creatinine, uric acid and vitamin B-12. The platelet loss were also analyzed using bovine blood. The membrane thickness was kept constant (0.5% by weight) in all membrane systems. Due to the simple coating procedure, cellulose nitrate coating, which also has good biocompatibility and high removal rates for toxins, was found to be preferable.

Adsorption↗

PubMed1980

[Synthesis and properties of carrier-bound enzymes. XI. Fixation of trypsin to various cellulose derivatives. Comparison of the kinetic properties of the trypsin-cellulose complexes].

Trypsin was covalently bound to carboxymethyl cellulose (CMC) azide and dialdehyde cellulose (DAC). Thereby the maximum protein binding capacity of CMC (420 mg/g) is 28 times that of DAC (15 mg/g). The high protein binding capacity of CMC is explained by a change in structure, i. e. by surface increase of the original cellulose powder due to chemical modification. By activation with sodium bisulfite solution we achieved an increase in protein binding capacity of DAC to values similar to those of CMC. The value of Vmax for all trypsin-DAC-complexes is about 38% of that of the free enzyme. With increasing protein content (from 1 to 12 mg/g) Km rises continuously from 0,14 to 0.36 mM. An analogous kinetic behaviour was found for trypsin-CMC-complexes only up to a protein content of 100 mg/g. Offering larger quantities of trypsin the enzyme is immobilized in an active form, so that all trypsin-CMC-complexes from 100 mg/g upwards have the same specific activity and the same Km; on the other hand the value of Vmax for the immobilized trypsin decreases to 17% of that for the free enzyme.

Cellulose↗

PubMed1997

Trichoderma reesei cellobiohydrolase I with an endoglucanase cellulose-binding domain: action on bacterial microcrystalline cellulose.

Cellulolytic enzymes consist of distinct catalytic and cellulose-binding domains (CBDs). The presence of a CBD improves the binding and activity of cellulases on insoluble substrates but has no influence on their activities on soluble substrates. Structural and biochemical studies of a fungal CBD from Trichoderma reesei cellobiohydrolase I have revealed a wedge shaped structure with a flat cellulose binding surface containing three essential tyrosine residues. The face of the wedge is strictly conserved in all fungal CBDs while many differences occur on the other face of the wedge. Here we have studied the importance of these differences on the function of the T. reesei CBHI by replacing its CBD by a homologous CBD from the endoglucanase, EGI. Our data shows that, apart from slightly improved affinity of the hybrid enzyme, the domain exchange does not significantly influence the function of CBHI.

Amino Acid Sequence↗

PubMed1981

[HbA2 evaluation: comparison between microchromatography on a DEAE cellulose column and conventional cellulose acetate electrophoresis].

Even if different methods were employed in Hb A2 level detection, so far the borderline values between normal and pathological "range" depend on the different laboratories and techniques. The Authors report here an investigation carried out on Hb A2 levels using two different methods: the chromatographic technique by DEAE cellulose column, and the cellulose acetate electrophoresis. The investigation regards 46 normal and 50 beta-thalassemia obligate carrier samples. The results demonstrate that chromatographic technique provides a lowest misclassification rate with a greater reliability. Therefore the use of the microchromatographic procedure for beta-thalassemia screening is recommended.

Chromatography, DEAE-Cellulose↗

PubMed1991

Thixotropy of different concentrations of microcrystalline cellulose:sodium carboxymethyl cellulose gels.

An extensive study is made of the rheologic behavior of microcrystalline cellulose:sodium carboxymethyl cellulose gels at six different concentrations (between 1 and 2.5% by weight). The thixotropy of these gels was analyzed in terms of agitation time, the duration of storage, concentration, and temperature; a semiempirical function was thus obtained providing thixotropic area as a function of these variables. The instantaneous variation rate in thixotropic area was defined, providing a better understanding of the rate of structural breakdown. Also, it is shown that after 5 min of agitation, the variation rate becomes negligible (approximately 0.7% of the initial value); this result is practically independent of the remaining variables.

Carboxymethylcellulose Sodium↗

PubMed1993

A new bacterial cellulose substrate for mammalian cell culture. A new bacterial cellulose substrate.

A new substrate for mammalian cell culture was developed using a cellulose membrane produced by Acetobacter aceti. Modification of the ionic charge of the membrane and adsorption of collagen to it promoted cellular adhesion to the membrane surface. The growth of eight kinds of cells on the membrane, was comparable to that achieved in plastic Petri dishes. The membrane was tested for use in the production of recombinant Erythroid Differentiation Factor (EDF)/activin A using genetically engineered Chinese hamster ovary cells. Both the viability of the cells and production of EDF/activin A were maintained for about 1 month, while cultures on plastic dishes lasted only 12 days. It was considered that the mechanism of improved cell viability was related to the ultrastructure of the cellulose membrane.

Acetobacter↗

PubMed1992

Improvement of blood compatibility on cellulose dialysis membrane. 2. Blood compatibility of phospholipid polymer grafted cellulose membrane.

The blood compatibility of a cellulose haemodialysis membrane whose surface was grafted with a methacrylate having a phospholipid polar group, 2-methacryloyloxyethyl phosphorylcholine, was evaluated with attention to platelet adhesion to the membrane surface and complement activation induced by the membrane. When the original cellulose membrane came in contact with platelet-rich plasma for 30 min, numerous platelets adhered to the surface and aggregated. On the other hand, the membrane grafted with 2-methacryloyloxyethyl phosphorylcholine effectively suppressed platelet adhesion and activation. This effect became more pronounced with increasing surface distribution. Especially, the 2-methacryloyloxyethyl phosphorylcholine grafted membranes, whose distribution exceeded 0.27, completely inhibited platelet adhesion, even when the contact time was 180 min. Moreover, the complement activation was also reduced with increased 2-methacryloyloxyethyl phosphorylcholine distribution on the surface of the membrane.

Animals↗

PubMed1976

Relation of certain infrared bands to conformational changes of cellulose and cellulose oligosaccharides.

The i.r. spectra of D-glucose and cellulose oligosaccharides up to cellopentaose have been compared with those of cellulose at various temperatures between that of liguid nitrogen and approximately 250 degrees. Significant changes in frequency and intensity of the bands at approximately 3400 cm -1 were observed. The a 1372 cm -1/a2900 cm -1 ratio for each carbohydrate studied decreased gradually as the temperature was increased above ambient. The change of the band intensities at 1429 and 893 cm -1 with temperature was also investigated. The observed spectral changes are assumed to be associated with changes of hydrogen bonding.

Cellulose↗