Production and characterization ofPhanerochaete chrysosporium lignin peroxidases for pulp bleaching

The production of lignin peroxidase fromPhanerochaete chrysosporium was studied using immobilized mycelia in nylon-web cubes in semicontinuous fermentation using glucose pulses or ammonium tartrate pulses. Consistent enzyme production was achieved when glucose pulses were used, leading to an average activity of 253 U/L. The crude enzyme was added to eucalyptus kraft pulp before conventional and ECF bleaching sequences. Optimization of the enzymatic pretreatment led to the following operational conditions: enzyme load of 2 U/g of pulp, hydrogen peroxide addition rate of 10 ppm/h, and reaction time of 60 min. Pulp final characteristics were dependent on the chemical treatment sequence that followed enzymatic pretreatment. The chief advantage of enzymatic pretreatment was pulp viscosity preservation, which was observed in most of the experiments carried out with seven different chemical treatment sequences     

Production of recombinant bleaching enzymes from thermophilic microorganisms in fungal hosts

Cost-effective production of enzymes for industrial processes makes the appropriate selection of the host-vector expression system critical. We have developed two systems for the bulk production of bleaching enzymes from thermophiles. Kluyveromyces lactis has been developed as a secretion host employing expression vectors based on the 2μ-like plasmid pKD1 of Kluyveromyces drosophilarium. Our second system involves the filamentous fungus Trichoderma reesei. Fusion and nonfusion vectors have been constructed using the strong cellobiohydrolase 1 (cbh1) promoter. The KEX2 protease cleavage site and a 6 × HIS-tag have been incorporated to facilitate both cleavage and purification of the mature foreign proteins.     

Polyoxypregnanes from the stems of Marsdenia tenacissima

From the stems ofMarsdenia tenacissima two new polyoxypregnanes were isolated, their structures were elucidated by 1D, 2D-NMR as 11α,12β-di-O-tigloyl-tenacigenin B （1） and tenacigenoside E （2）.     

Non-alkaline bleaching of wood pulp using pulsed RF dielectric heating: comparison with pulsed microwave enhanced bleaching

Non-alkaline bleaching of thermo-mechanical pulps using 40 MHz RF dielectric capacitor heating is described. Experimental data obtained under RF irradiation are compared with non-alkaline bleaching under pulsed microwave at 2.45 GHz. Optimal processing time is determined. Effectiveness of non-alkaline bleaching of TMP in RF is shown to be close to that in microwave. Neither microwave nor RF irradiation, under non-alkaline bleaching conditions, is shown to result in detriment to the physical properties of the pulp.     

Determination of resin acids in pulp mill EOP bleaching process effluent

Resin acids are tricyclic diterpenoids which are natural constituents of the wood from conifers. They are released from the wood during the manufacture of pulp and paper. These acids are very resistant to chemical degradation and survive the pulping and also the EOP bleaching process (EOP=alkaline extraction, oxygen and peroxide, the chemicals used in the bleaching process). Resin acids were extracted from alkaline medium using liquid–liquid extraction with t-butyl methyl ether and solid phase extraction with RP C18 adsorbent and a highly porous polystyrene-divinylbenzene polymer. After conversion of the acids to their pentafluorobenzyl esters, the extracts were analysed by GC/MS using a 25 m OV17 capillary column. Recovery values for single resin acids were determined by all three extraction methods. The solid phase extraction methods were applied to the analysis of the EOP effluent from a pulp mill bleaching process. 14 different resin acids and one resin acid methyl ester have been identified in the effluent. One of these was an oxo resin acid which might well be a product of the bleaching process.     

Physico-chemical characterization of a cellulosic fraction from sugar beet pulp

The residue of sugar beet pulp from which pectin and alkaline soluble polysaccharides have been removed by microwave assisted extraction or conventional heat was treated with sodium monochloroacetate under alkaline pH to convert the residual cellulose present to carboxy methyl cellulose (CMC). Weight average molar masses ranged from about 96 to 220 × 10³ Daltons, weight average intrinsic viscosity from 1.9 to 4.1 dL/g and degree of substitution from 1.38 to 0.59. HPSEC with online molar mass detectors and Atomic Force Microscopy revealed that CMC was comprised of aggregated linear moieties in contact with spherical bodies. The linear portion was a mixture of rods and segmented rods. Some of the rods had long branches.     

Production, purification, and biochemical characterization of two beta-glucosidases from Sclerotinia sclerotiorum

The filamentous fungus Sclerotinia sclerotiorum produces beta-glucosidases in liquid culture with a variety of carbon sources, including cellulose (filter paper), xylan, barley straw, oat meal, and xylose. Analysis by native polyacrylamide gel electrophoresis (PAGE) followed by an activity staining with the specific chromogenic substrate, 5-bromo 4-chloro 3-indolyl beta-1,4 glucoside (X-glu) showed that two extracellular beta-glucosidases, designated as beta-glu1 and beta-glu2, were in the filter paper culture filtrate. Only one enzyme designated as beta-glu x was revealed by the same method in the xylose culture filtrate. Beta-glu1 and beta-glu2 were purified to homogeneity. The purification procedure consist of a common step of anion-exchange chromatography on DEAE-Sepharose CL6B, both high-performance liquid chromatography (HPLC) anion-exchange and gel filtration columns for beta-glu1 and only HPLC gel filtration for beta-glu2. Beta-glu1 has a molecular mass of 196 kDa and 96.5 kDa, as estimated by gel filtration and sodium dodecyl sulfate (SDS)-PAGE, respectively, suggesting that the native enzyme may consist of two identical subunits. The same analysis showed that beta-glu2 is a monomeric protein with an apparent molecular mass of about 76.5 kDa. Beta-glu1 and beta-glu2 hydrolyses PNPGlc and cellobiose, with apparent Km values respectively for PNPGlc and cellobiose of 0.1 and 1.9 mM for beta-glu1 and 2.8 and 8 mM for beta-glu2. Both enzymes exhibit the same temperature and pH optima for PNPGlc hydrolysis (60 degrees C and pH 5.0). beta-glu1 was stable over a pH range of 3-8 and kept 50% of its activity after 30 min of heating at 60 degrees C without substrate. It was further characterized by studying the effect of some cations and various reagents on its activity.     

Fourier transform Raman spectroscopic studies of a novel wood pulp bleaching system

The use of near-infrared (NIR) Fourier transform (FT) Raman spectroscopy for the study of lignocellulosic materials is discussed. An application utilizing NIR FT-Raman spectroscopy to study a novel chlorine-free process for the bleaching of wood pulps is presented in detail. The new process, still under development, entails the oxidation of residual lignin in wood pulps by vanadium-substituted polyoxometalates, and reoxidation of the reduced polyoxometalates by chlorine-free oxidants such as air, dioxygen, peroxides or ozone. Results from FT-Raman measurements of polyoxometalate-treated pulps are compared with those from chemical, spectroscopic and optical techniques commonly used in the pulp and paper industry.     

Biobleaching - An ecofriendly and environmental benign pulp bleaching technique: A review

The review summarizes the use of ligninolytic enzymes in bleaching processes in paper industries. The awareness of society regarding pollution problems demands an effort to develop novel and less polluting processes for paper production. In this regard, there is an emerging interest in various applications of microbes and enzymes. Previously, appropriate enzymes were unavailable, apart from partial use in modification of starch for application in paper coatings. Continued exploration has led to the improvement in enzymes that offer remarkable profits to paper industries.     

Production, purification, and biochemical characterization of two β-glucosidases from Sclerotinia sclerotiorum

The filamentous fungus Sclerotinia sclerotiorum prudces ß-glucosidases in liquid culture with a variety of carbon sources, including cellulose (filter paper), xylan, barley straw, oat meal, and xylose. Analysis by native polyacrylamide gel electrophoresis (PAGE) followed by an activity staining with the specific chromogenic substrate, 5-bromo 4-chloro 3-indolyl ß-1,4 glucoside (X-glu) showed that two extracellular β-glucosidases, designated as ß-glul1 and \-glu2, were in the filter paper culture filtrate. Only one enzyme designated as ß-glus was revealed by the same method in the xylose culture filtrate. ß-glu1 and ß-glu2 were purified to homogeneity. The purification procedure consist of a common step of anion-exchange chromatography on DEAE-Sepharose CL6B, both high-performance liquid chromatography (HPLC) anion-exchange and gel filtration columns for ß-glu1 and only HPLC gel filtration for ß-glu2. ß-glu1 has a molecular mass of 196 kDa and 96.5 kDa, as estimated by gel filtration and sodium dodecyl sulfate (SDS)-PAGE, respectively, suggesting that the native enzyme may consist of two identical subunits. The same analysis showed that ß-glu2 is a monomeric protein with an apparent molecular mass of about 76.5 kDa. ß-glu1 and ß-glu2 hydrolyses PNPG1c and cellobiose, with apparent K _m values respectively for PNPGlc and cellobiose of 0.1 and 1.9 mM for ß-glu1 and 2.8 and 8 mM for ß-glu2. Both enzymes exhibit the same temperature and pH optima for PNPG1c hydrolysis (60°C and pH 5.0). ß-glu1 was stable over a pH range of 3–8 and kept 50% of its activity after 30 min of heating at 60°C without substrate. It was further characterized by studying the effect of some cations and various reagents on its activity.     

Forsskamide,a new ceramide from aerial parts of Forsskaolea tenacissima Linn.

Although the various folk medicine uses and the biological activity of Forsskaolea tenacissima L., few chemical constituents of this plant have been reported, this provoked us to make our study. Forsskamide, a new ceramide was isolated from aerial parts of F. tenacissima L. (Urticaceae). The chemical structure was established by different spectroscopic methods (¹H, ¹³C-NMR, HMBC, HSQC, ROESY, FAB-MS and HR-FAB-MS). Forsskamide showed a moderate cytotoxic activity by (MTT) method against human colorectal carcinoma cell line (HCT-116) with IC₅₀ 33.25 μM in comparison with 5-fluorouracil IC₅₀ 26.42 μM. While, it did not show any activity against human hepatocarcinaoma cell line (HepG-2).     

Rheological characterization of high concentrated MFC gel from kenaf unbleached pulp

Microfibrillated Cellulose (MFC) was isolated from unbleached kraft pulp derived from kenaf bast fiber. MFC gels with different concentrations were manufactured from pulp with varying initial consistencies. The MFC was diluted to a consistency of around one percent using distilled water and was further homogenized by passing through a microfluidizer. In order to gain an understanding of the relative changes in behavior of the resulting MFC gels, their rheological properties were characterized. Results show that all of the gels exhibit a shear-thinning behavior. It was also determined that the rheological characteristics improved with increasing gel concentration, which was achieved by using higher pulp suspension consistencies. Diluted MFC that was derived from highly concentrated MFC had more variable modulus under the same strain and frequency compared to poorly concentrated MFC. But such a strong effect was not observed for viscosity. Additionally, the value of G′, ranging from 76 to 5325 Pa under the studied concentrations, was found to be fourfold the value of G″. In the low frequency range, G′ was almost independent of frequency, but was dependent on gel consistency with a coefficient of 3, indicating that MFC gels are elastic. These results show that it is possible to produce MFC gels with good rheological properties from high consistency kenaf pulp suspensions.     

Thermal characterization of benzylcellulose derivatives prepared from bleached pinus kraft pulp

The thermal characterization (DSC and TG) of benzylcellulose derivatives prepared from the benzylation of bleached Pinus Kraft pulp is described in this paper. The objective of this study was to examine the changes in glass transition temperature (T _g) and the thermal stability of the benzylated product as a function of the benzylation extent (degree of substitution). The DSC analysis showed that the benzylcelluloses can display glass transition temperature at two different regions and that thermal stability is slightly higher than that of the parent cellulose. This revised version was published online in August 2006 with corrections to the Cover Date.     

Gas chromatographic determination of pollutants in the chlorination and caustic extraction stage effluent from the bleaching of a bamboo pulp

The gas chromatographic detection and quantitative determination of various chlorophenolics as well as resin and fatty acids have been carried out in the chlorination and caustic extraction stage effluents generated in the laboratory by bleaching a bamboo pulp. A number of chlorinated phenols, catechols, guaiacols, syringaldehydes and resin acids as well as non-chlorinated saturated and unsaturated fatty acids together with resin acids have been detected. The concentration of various compounds detected have also been compared with the reported (96)LC(50) values.     

Chemical characterization and ultrastructure study of pulp fibers

Understanding the ultrastructure and chemical characterization of pulp fibers is highly important in utilizing wood as a raw material in a wide scope of applications, such as forest biomass-based biorefineries and low-cost renewable materials. The observation of the ultrastructure is not possible without advanced microscopy and spectroscopy techniques. Therefore, this study focuses on exploring the ultrastructure of pulp fibers with helium ion microscopy (HIM) and scanning electron microscopy (SEM). For the analysis of chemical characterization in the pulp fibers, Raman spectroscopy, Fourier transform infrared spectroscopy (FT-IR), and X-ray photoelectron spectroscopy (XPS) were performed. For these studies, the pulp fiber samples were obtained mainly from three different wood species, i.e. spruce, birch and eucalyptus. They were received in the never dried state and dried with a critical point drier (CPD) to minimize pore collapse. The spectroscopy results showed a strong signal from crystalline cellulose and confirmed the absence of lignin after Kraft pulping and bleaching. However, with XPS about 2% of lignin was detected in eucalyptus pulp. The results obtained with the microscopy techniques are compared and indicating the nanofibril size, shape, surface roughness as well as their orientation in pulp fibers. To our knowledge, this is the first time that HIM is applied to study the ultrastructure of pulp fibers and compared against more conventional microscopy and spectroscopy techniques. The main differences between HIM and SEM were found to be related to the focusing and magnification. The individual nano- and microfibrils as well as their bundles were more easily visible with HIM than with SEM. Also, with HIM it was possible to get the total area in focus at once which was not the case with SEM. The increased understanding of the ultrastructure and chemical composition of wood pulp enhance the development of novel wood-based products and processes for their manufacture.     

Production and characterization of anisotropic particles from biodegradable materials

In recent years, production and characterization of anisotropic particles has become of interest in a wide range of scientific fields including polymer chemistry, drug delivery, electronics, energy, and nanotechnology. In this work, we demonstrate a novel formulation for production of anisotropic particles via an internal phase separation of biodegradable components. Specifically, binary mixtures of biodegradable polymers poly(lactic-co-glycolic acid), polycaprolactone, and biodegradable lipid Precirol (glyceryl palmitostearate) were dissolved in dichloromethane, emulsified, and prepared into anisotropic particles using a modified solvent evaporation technique. During the slow evaporation process the components self-assembled into anisotropic particles with distinct morphologies. Polymer/polymer formulations resulted in compartmentalized anisotropic heterodimer particles, while polymer/lipid combinations yielded "ice cream cone" shaped particles. It was found that addition of certain active pharmaceuticals resulted in an altered, pox-like segregation at the particle surface of polymer/polymer formulations. The anisotropic nature of the particles was subsequently characterized using optical microscopy, scanning electron microscopy, zeta potential, electrophoresis, and X-ray diffraction. Successful formulations presented here may potentially be employed as multicompartmental drug carriers with staggered drug release rates or alternatively as a colloidal excipient for an arsenal of pharmaceutical applications.     

Application of an amperometric sensor to in-line monitoring of pulp bleaching with chlorine dioxide

Chlorine dioxide is replacing chlorine as the active compound in pulp bleaching in order to reduce the amount of chlorine used in the process and hence also in the waste waters. In bleaching with chlorine dioxide part of the effective bleaching chemical is usually chlorite. The electrochemistry of chlorine dioxide and chlorite at solid electrodes was studied by cyclic voltammetry at different pH values. The observed voltammograms indicated that reduction of chlorine dioxide gives chlorite and oxidation of chlorite gives chlorine dioxide. Both voltammograms were well developed, indicating a reversible process. Both platinum and glassy carbon were used as the working electrode. The dependence of the limiting current of chlorine dioxide and chlorite on pH was studied at both electrodes. The method was tested in the chlorine dioxide bleaching stage D1 in a typical bleaching process. A good correlation was found between the concentrations of chlorine dioxide and chlorite measured by the in-line amperometric method and a standard titrimetric method.     

Mass spectra of chlorinated aromatics formed in pulp bleaching: I—chlorinated catechols

A systematic study on the electron impact mass spectra of all nine chlorinated catechols in presented. Metastable ion analysis was used to elucidate the fragmentation pathways. The influence of the position of the chloro substituents can be used to distinguish the structural isomers. In this respect the most characteristic fragment ions are [M? CHl]⁺˙, [M? HCOOH]⁺˙, [M? COCl]⁺, [M? HCl? CO]⁺˙, [M? CHOCl]⁺˙ and [M? HCl? HCl]⁺˙.     

Mass spectra of chlorinated aromatics formed in pulp bleaching 2—Chlorinated guaiacols

The fragmentation of chlorinated guaiacols (2-methoxyphenols) on electron impact has been studied. The most common fragmentation processes are interpreted and in some cases the small differences between spectra of positional isomers are explained. In addition to the well-known alkyl-oxygen fission (loss of methyl radical), metastable ion studies and deuterium labelling have indicated several new fragmentation pathways. The most characteristic are the formation of [M? CH₃? HCl]⁺ and [M? CH₃? Cl]^+· ions. In general, however, the spectra of positional isomers are shown to be very similar.