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Liquid-liquid phase separation (LLPS) is involved in various dynamic biological phenomena. In epithelial cells, dynamic regulation of junctional actin filaments tethered to the apical junctional complex (AJC) is critical for maintaining internal homeostasis against external perturbations; however, the role of LLPS in this process remains unknown. Here, after identifying a multifunctional actin nucleator, cordon bleu (Cobl), as an AJC-enriched microtubule-associated protein, we conducted comprehensive in vitro and in vivo analyses. We found that apical microtubules promoted LLPS of Cobl at the AJC, and Cobl actin assembly activity increased upon LLPS. Thus, microtubules spatiotemporally regulated junctional actin assembly for epithelial morphogenesis and paracellular barriers. Collectively, these findings established that LLPS of the actin nucleator Cobl mediated dynamic microtubule-actin cross-talk in junctions, which fine-tuned the epithelial barrier.

Claudins are cell-cell adhesion molecules located at the tight junctions (TJs) between cells in epithelial cell sheets. The claudin family in mammals consists of 27 four-transmembrane domain proteins. Claudins are responsible for the paracellular barrier function of TJs, and in some cases confer paracellular channel functions to the paracellular barriers of TJs. Based on recent breakthroughs in the molecular structure of claudins, the hypothetical 'antiparallel double row model' was proposed, which suggests how claudins polymerize in a linear fashion and form TJ strands with paracellular barrier and channel functions. Meanwhile, ongoing studies at the cell and tissue levels are clarifying how the paracellular barrier and/or channel functions of claudin-based TJs, which are both robust and flexible, organize various biological systems.

Apical constriction is critical for epithelial morphogenesis, including neural tube formation. Vertebrate apical constriction is induced by di‐phosphorylated myosin light chain (ppMLC)‐driven contraction of actomyosin‐based circumferential rings (CRs), also known as perijunctional actomyosin rings, around apical junctional complexes (AJCs), mainly consisting of tight junctions (TJs) and adherens junctions (AJs). Here, we revealed a ppMLC‐triggered system at TJ‐associated CRs for vertebrate apical constriction involving microtubules, LUZP1, and myosin phosphatase. We first identified LUZP1 via unbiased screening of microtubule‐associated proteins in the AJC‐enriched fraction. In cultured epithelial cells, LUZP1 was found localized at TJ‐, but not at AJ‐, associated CRs, and LUZP1 knockout resulted in apical constriction defects with a significant reduction in ppMLC levels within CRs. A series of assays revealed that ppMLC promotes the recruitment of LUZP1 to TJ‐associated CRs, where LUZP1 spatiotemporally inhibits myosin phosphatase in a microtubule‐facilitated manner. Our results uncovered a hitherto unknown microtubule‐LUZP1 association at TJ‐associated CRs that inhibits myosin phosphatase, contributing significantly to the understanding of vertebrate apical constriction., The neural tube‐closure regulator LUZP1 drives apical constriction of epithelial cells by inhibiting myosin phosphatase activity in a microtubule‐dependent manner

Background & Aims Most cholesterol gallstones have a core consisting of inorganic and/or organic calcium salts, although the mechanisms of core formation are poorly understood. We examined whether the paracellular permeability of ions at hepatic tight junctions is involved in the core formation of cholesterol gallstones, with particular interest in the role of phosphate ion, a common food additive and preservative. Methods We focused on claudin-3 (Cldn3), a paracellular barrier-forming tight junction protein whose expression in mouse liver decreases with age. Since Cldn3-knockout mice exhibited gallstone diseases, we used them to assess the causal relationship between paracellular phosphate ion permeability and the core formation of cholesterol gallstones. Results In the liver of Cldn3-knockout mice, the paracellular phosphate ion permeability through hepatic tight junctions was significantly increased, resulting in calcium phosphate core formation. Cholesterol overdose caused cholesterol gallstone disease in these mice. Conclusion We revealed that in the hepatobiliary system, Cldn3 functions as a paracellular barrier for phosphate ions, to help maintain biliary ion homeostasis. We provide in vivo evidence that elevated phosphate ion concentrations play a major role in the lifestyle- and age-related risks of developing cholesterol gallstone disease under cholesterol overdose. Lay summary Herein, we reveal a new mechanism for cholesterol gallstone formation, in which increased paracellular phosphate ion permeability across hepatobiliary epithelia causes calcium phosphate core formation and cholesterol gallstones. Thus, altered phosphate ion metabolism under cholesterol overdose plays a major role in the lifestyle- and age-related risks of developing cholesterol gallstone disease.

Genome-wide association studies and subsequent deep sequencing analysis have identified susceptible loci for inflammatory bowel diseases (IBDs) including ulcerative colitis (UC). A gene encoding RING finger protein 186 (RNF186) is located within UC-susceptible loci. However, it is unclear whether RNF186 is involved in IBD pathogenesis. Here, we show that RNF186 controls protein homeostasis in colonic epithelia and regulates intestinal inflammation. RNF186, which was highly expressed in colonic epithelia, acted as an E3 ligase mediating polyubiquitination of its substrates. Permeability of small organic molecules was augmented in the intestine of Rnf186-/- mice. Increased expression of several RNF186 substrates, such as occludin, was found in Rnf186-/- colonic epithelia. The disturbed protein homeostasis in Rnf186-/- mice correlated with enhanced endoplasmic reticulum (ER) stress in colonic epithelia and increased sensitivity to intestinal inflammation after dextran sulfate sodium (DSS) treatment. Introduction of an UC-associated Rnf186 mutation led to impaired E3 ligase activity and increased sensitivity to DSS-induced intestinal inflammation in mice. Thus, RNF186 maintains gut homeostasis by controlling ER stress in colonic epithelia.

Vinculin in the apical junctional complex maintains the paracellular barrier function specifically for ions, but not for large solutes, by buffering mechanical fluctuations., The paracellular barrier function of tight junctions (TJs) in epithelial cell sheets is robustly maintained against mechanical fluctuations, by molecular mechanisms that are poorly understood. Vinculin is an adaptor of a mechanosensory complex at the adherens junction. Here, we generated vinculin KO Eph4 epithelial cells and analyzed their confluent cell-sheet properties. We found that vinculin is dispensable for the basic TJ structural integrity and the paracellular barrier function for larger solutes. However, vinculin is indispensable for the paracellular barrier function for ions. In addition, TJs stochastically showed dynamically distorted patterns in vinculin KO cell sheets. These KO phenotypes were rescued by transfecting full-length vinculin and by relaxing the actomyosin tension with blebbistatin, a myosin II ATPase activity inhibitor. Our findings indicate that vinculin resists mechanical fluctuations to maintain the TJ paracellular barrier function for ions in epithelial cell sheets.

Tight junction is a cell adhesion apparatus functioning as barrier and/or channel in the paracellular spaces of epithelia. Claudin is the major component of tight junction and polymerizes to form tight junction strands with various morphologies that may correlate with their functions. Here we present the crystal structure of mammalian claudin-3 at 3.6 Å resolution. The third transmembrane helix of claudin-3 is clearly bent compared with that of other subtypes. Structural analysis of additional two mutants with a single mutation representing other subtypes in the third helix indicates that this helix takes a bent or straight structure depending on the residue. The presence or absence of the helix bending changes the positions of residues related to claudin-claudin interactions and affects the morphology and adhesiveness of the tight junction strands. These results evoke a model for tight junction strand formation with different morphologies – straight or curvy strands – observed in native epithelia., The main components of tight junctions (TJ) are claudins that polymerize and form meshwork architectures called TJ strands. Here the authors present the 3.6 Å crystal structure of murine claudin-3 and show that residue P134 causes a bending of the third transmembrane helix which affects the morphology and adhesiveness of the TJ strands.

Background & Aims: Epithelial cells are joined by tight junctions (TJs) to form a cell sheet. In the stomach, epithelial cell sheet forms an essential barrier against gastric material, including gastric acid. Although the decreased expression of stomach-type claudin-18 (stCldn18), a TJ protein, is generally observed in human gastritis and gastric cancer, its pathological roles are not fully understood. We previously reported that mice lacking stCldn18 (stCldn18-/-) exhibit gastric acid leakage through TJs, which induces active gastritis at a young age. Here, we examined the gastric pathologies in mice after long-term stCldn18 deficiency. Methods: The gastric pathologies in stCldn18-/- mice were sequentially analyzed from youth to old age, and compared to those in humans. To examine the relationship between stCldn18 deficiency-induced gastric pathologies and Wnt-dependent tumorigenesis, we generated Wnt1-overexpressing stCldn18-/- mice. Results: StCldn18-/- mice developed chronic active gastritis at middle age, with expression of the chemoattractant CCL28. At old age, 20-30% of these mice developed gastric tumors with CXCL5 expression, indicative of EMT. In this process, spasmolytic polypeptide-expressing metaplasia (SPEM) cells appeared. Increased expressions of CD44-variants, TLR2, and CXCL5 indicated age-dependent changes in cell characteristics. Some features of the stCldn18-/- mouse gastric tumorigenesis resembled H pylori-infection-related human carcinogenesis. The gastric tumorigenesis was accelerated in Wnt1-overexpressing stCldn18-/- mice, indicating that Wnt is involved in the stCldn18-/- mouse gastric tumorigenesis. Conclusions: StCldn18 deficiency induced gastric tumorigenesis in mice without H pylori infection. Our findings revealed that several signaling networks, including the cytokine-, stemness-, and Wnt-signaling pathways, may be activated under the stCldn18-deficiency-induced chronic active gastritis to accelerate the gastric tumorigenesis. Keywords: Epithelial Barrier, Tight Junction, Chronic Active Gastritis, SPEM

Objective To design novel anti-inflammation treatments, it is important to recognise two distinct steps of inflammation: initiation and acceleration. In IBDs, intestinal inflammation is reported to be accelerated by dysfunction in the epithelial paracellular barrier formed by tight junctions (TJs). However, it is unclear whether changes in paracellular barrier function initiate inflammation. Some of the intestinal claudin-family proteins, which form the paracellular barrier, show aberrant expression levels and localisations in IBDs. We aimed to elucidate the role of paracellular-barrier change in initiating colonic inflammation. Design We generated intestine-specific conditional knockout mice of claudin-7 (Cldn7) , one of the predominant intestinal claudins. Results The intestine-specific Cldn7 deficiency caused colonic inflammation, even though TJ structures were still present due to other claudins. The paracellular flux (pFlux), determined by measuring the paracellular permeability across the colon epithelium, was enhanced by the Cldn7 deficiency for the small organic solute Lucifer Yellow (457 Da), but not for the larger organic solute FITC-Dextran (4400 Da). Consistent with these results, the intestine-specific claudin-7 deficiency enhanced the pFlux for N -formyl-l-methionyl-l-leucyl-l-phenylalanine (fMLP) (438 Da), a major bacterial product, to initiate colonic inflammation. Conclusions These findings suggest that specific enhancement of the pFlux for small organic solutes across the claudin-based TJs initiates colonic inflammation.

The claudins are a family of membrane proteins with at least 27 members in humans and mice. The extracellular regions of claudin proteins play essential roles in cell-cell adhesion and the paracellular barrier functions of tight junctions (TJs) in epithelial cell sheets. Furthermore, the extracellular regions of some claudins function as paracellular channels in the paracellular barrier that allow the selective passage of water, ions, and/or small organic solutes across the TJ in the extracellular space. Structural analyses have revealed a common framework of transmembrane, cytoplasmic, and extracellular regions among the claudin-based paracellular barriers and paracellular channels; however, differences in the claudins' extracellular regions, such as their charges and conformations, determine their properties. Among the biological systems that involve fluid flow and metabolism, it is noted that hepatic bile flow, renal Na+ reabsorption, and intestinal nutrient absorption are dynamically regulated via site-specific distributions of paracellular channel-forming claudins in tissue. Here, we focus on how site-specific distributions of claudin-2- and claudin-15-based paracellular channels drive their organ-specific functions in the liver, kidney, and intestine.

Claudin protein family members, of which there are at least 27 in humans and mice, polymerize to form tight junctions (TJs) between epithelial cells, in a tissue- and developmental stage-specific manner. Claudins have a paracellular barrier function. In addition, certain claudins function as paracellular channels for small ions and/or solutes by forming selective pores at the TJs, although the specific claudins involved and their functional mechanisms are still in question. Here we show for the first time that claudin-21, which is more highly expressed in the embryonic than the postnatal stages, acts as a paracellular channel for small cations, such as Na(+), similar to the typical channel-type claudins claudin-2 and -15. Claudin-21 also allows the paracellular passage of larger solutes. Our findings suggest that claudin-21-based TJs allow the passage of small and larger solutes by both paracellular channel-based and some additional mechanisms.

Claudins (Cldn) are essential membrane proteins of tight junctions (TJs), which form the paracellular permselective barrier. They are produced by a multi-gene family of 24 reported members in mouse and human. Based on a comprehensive search combined with phylogenetic analyses, we identified three novel claudins (claudin-25, -26, and -27). Quantitative RT-PCR revealed that the three novel claudins were expressed in a tissue- and/or developmental stage-dependent manner. Claudins-25 and -26, but not claudin-27, were immunofluorescently localized to TJs when exogenously expressed in cultured MDCK and Eph epithelial cell lines. These findings expand the claudin family to include at least 27 members.Structured summaryClaudin-25 and ZO-1 colocalize by fluorescence microscopy (View interaction)ZO-1 and Claudin-26 colocalize by fluorescence microscopy (View interaction)

Organization and place for partnership of local society and experts is called urban design center. Architecture and planning schools have its examples. This paper reports the first year of Urban Design Center Tamura (UDCT) to show how local government, local community and university collaborate to revitalize a small city.

Titanium dioxide (TiO2) powder-containing sheet composites, called TiO2 sheet, were prepared by a papermaking technique, and their photocatalytic efficiency was investigated. The TiO2 powders were homogeneously scattered over the fiber-mix networks tailored within the catalyst sheet. Under UV irradiation, the TiO2 paper could decompose p-hydroxyacetophenone (p-HAP), although the degradation efficiency by the TiO2 sheet was lower than that by the TiO2 powder. Scanning electron microscopy revealed that coverage of the TiO2 particles inside the sheet by alumina binder which was used to improve the sheet strength caused the deterioration of the photocatalytic performance. Internal addition of alumina binder made the TiO2 sheet porous and such a TiO2 sheet exhibited high photocatalytic performance equivalent to that of TiO2 powder. The porous structure of TiO2 sheet might contribute to effective transport of p-HAP to the surface of TiO2 particles inside the sheet, resulting in high degradation performance. In addition, TiO2 sheet prepared using TiO2 sol showed higher photocatalytic efficiency than TiO2 powder and it was indicated that the porous sheet structure might provide suitable conditions for TiO2 catalysis for photodecomposition.

The reactivity ratios for the aqueous free-radical copolymerization of diallyldimethylammonium chloride and N-vinylformamide were found to be 0.13 and 1.92, respectively, from a Fineman–Ross analysis of a series of batch polymerizations. Because batch polymerization could not give a uniform product in a high yield with two monomers of such different reactivities, a semibatch procedure was developed in which the more reactive N-vinylformamide was added in 10 steps over the course of the copolymerization. The poly(diallyldimethyl- ammonium chloride-co-N-vinylformamide) copolymers were hydrolyzed to give poly(diallyldimethylammonium chloride-co-vinylamine). The utility of the vinylamine/diallyldimethylammonium chloride copolymers was demonstrated by the preparation and characterization of three derivatives: (1) a copolymer with coupled dansyl groups for fluorescence detection; (2) a copolymer with coupled dabsyl groups for ultraviolet–visible detection; and (3) an ultra-high-molecular-weight (1.6 × 106 Da) poly(diallyldimethylammonium chloride) by chain extension (coupling) with glycerol diglycidyl ether. © 2007 Wiley Periodicals, Inc. J Appl Polym Sci 104: 1068–1075, 2007

To isolate lignin peroxidase (LiP), extracellular activity of this type of enzyme was assayed against several white-rot basidiomycetes. Trametes cervi72;a was found to show relatively higher lignin peroxidase (LiP) activity in extracellular culture medium. LiP from T. cervina (TCLiP) was purified using several

Protoplasts of the basidiomycete, Fomitopsis palustris (formerly Tyromyces palustris), were utilized to study a function of the fungal plasma membrane. Fungal protoplasts exhibited metabolic activities as seen with intact mycelial cells. Furthermore, the uptake of certain compounds into the protoplast cells was quantitatively observed by using non-radioactive compounds. Vanillin was converted to vanillyl alcohol and vanillic acid as major products and to protocatechuic acid and 1,2,4-trihydroxybenzene as trace products by protoplasts prepared from F. palustris. Extracellular culture medium showed no activity responsible for the redox reactions of vanillin. Only vanillic acid was detected in the intracellular fraction of protoplasts. However, the addition of disulfiram, an aldehyde dehydrogenase inhibitor, caused an intracellular accumulation of vanillin, strongly suggesting that vanillin is taken up by the cell, followed by oxidation to vanillic acid. The addition of carbonylcyanide m-chlorophenylhydrazone, which dissipates the pH gradient across the plasma membrane, inhibited the uptake of either vanillin or vanillic acid into the cell. Thus, the fungus seems to possess transporter devices for both vanillin and vanillic acid for their uptake. Since vanillyl alcohol was only observed extracellularly, the reduction of vanillin was thought to be catalyzed by a membrane system.

Biphenyl (BP), biphenylene (BN), dibenzofuran (DF), dibenzo-p-dioxin (DD), and diphenyl ether (DE) were utilized as diphenyl substrates for the lignin-degrading basidiomycete, Phanerochaete chrysosporium and its extracellular enzyme, lignin peroxidase (LiP). Among these compounds, only BN and DD were oxidized by LiP. Cyclic voltammetry measurement revealed that BN and DD possess lower redox potentials than other diphenyl substrates utilized, being accordance with a reactivity of LiP. Although the degradations of BP, DF and DE were not extracellularly initiated by LiP, they were metabolized via an intermediate formation of hydroxylated products by intracellular enzymes. Upon addition of piperonyl butoxide, a cytochrome P450 inhibitor, these hydroxylation reactions were effectively inhibited, indicating the involvement of cytochrome P450s. Thus, two individual mechanisms seem to be compulsory for P. chrysosporium to metabolize a wide variety of recalcitrant diphenyl compounds; one is the activation of the aromatic ring via LiP catalyzed one-electron oxidation forming the aryl cation radical and the other is the activation of molecular oxygen by cytochrome P450 enzymes causing hydroxylation reactions on the aromatic ring.

Composite sheets composed of biodegradable polymers (BP) and paper were prepared, and their physical and biodegradable properties were investigated. The paper sheets were soaked in BP emulsions and cured at 100°C for 20 min. The wet strength of composite sheets with the same basis weight increased significantly with increasing BP content, although dry strength increased only moderately. A 0.5% addition of a common wet paper-strengthening agent, poly(amidoamine-epichlorohydrin) (PAE) resin, enhanced the wet strength, which reached 9.3 MPa, of composite sheets consisting of a BP : paper ratio of 20 : 80. Further enhancement was achieved by the addition of polyvinylamines (PVAm). The wet tensile strength of composite sheets consisting of BP and paper (20 : 80) increased by 27% with the addition of 0.2% PVAm and 0.5% PAE, whereas it was enhanced by only 3–4% with the addition of 0.7% PAE. The biodegradability of the composite sheets was somewhat retarded by the addition of PAE and PVAm. However, a 90% weight loss was observed 60 days after placing composite sheet samples in soil. A similar weight loss was observed after 45 days for composites without additives and after 30 days for base paper alone. © 2005 Wiley Periodicals, Inc. J Appl Polym Sci 96: 861–866, 2005

Activation of lignin peroxidase (LIP) in an organic solvent by reversed micelles was investigated. Bis(2-ethylhexyl)sulfosuccinate sodium salt (AOT) was used as a surfactant to form a reversed micelle. Lyophilized LIP from an optimized aqueous solution exhibited no enzymatic activity in any organic solvents examined in this study; however, LIP was catalytically active by being entrapped in the AOT reversed micellar solution. LIP activity in the reversed micelle was enhanced by optimizing either the preparation or the operation conditions, such as water content and pH in water pools of the reversed micelle and the reaction temperature. Stable activity was obtained in isooctane because of the stability of the reversed micelle. The optimal pH was 5 in the reversed micellar system, which shifted from pH 3 in the aqueous solution. The degradation reaction of several environmental pollutants was attempted using LIP hosted in the AOT reversed micelle. Degradation achieved after a 1-h reaction reached 81%, 50%, and 22% for p-nonylphenol, bisphenol A, and 2,4-dichlorophenol, respectively. This is the first report on the utilization of LIP in organic media.

A novel paper-based material containing titanium dioxide (TiO(2)) photocatalyst was successfully prepared by a papermaking technique with the internal addition of inorganic fibers on which TiO(2) particles were supported. Photodegradation performance of acetaldehyde gas, an indoor pollutant, and the durability of the TiO(2)-containing papers were investigated under UV irradiation. Ceramic fiber suspension and polydiallyldimethylammonium chloride as a cationic flocculant were mixed, followed by the addition of TiO(2) suspension and anionic polyacrylamide. Subsequently, the inorganic mixture was poured into a pulp suspension, and TiO(2) handsheets then prepared by a papermaking method. The tensile strength of TiO(2)-containing paper without a ceramic carrier decreased by more than 30% after 240-h UV irradiation (2 mW/cm(2)), although the strength of the TiO(2) sheet with ceramic fibers remained reasonably stable. The efficiency of acetaldehyde decomposition by the TiO(2) paper containing an inorganic carrier was nearly equal to that of the carrier-free TiO(2) paper. Scanning electron microscopic observation suggested that most TiO(2) particles were predominantly supported on the inorganic fiber matrix, and were mostly out of contact with organic pulp fibers. The TiO(2) paper with an inorganic carrier demonstrated both excellent photocatalytic performance and durability, which before had been mutually incompatible for organic materials containing TiO(2) photocatalyst. The two-stage mixing procedure for TiO(2) sheet-making is promising for the simple manufacture of high performance paper with photocatalytic ability.

Two types of powders, titanium dioxide (TiO 2 ) photocatalyst and zeolite adsorbent, were made into a paper-like composite by a papermaking technique using a dual polyelectrolyte system. The photodegradation behavior of bisphenol A (2,2-bis(4-hydroxyphenyl)propane, BPA) in water was investigated under UV irradiation. It was found that TiO 2 sheets could photocatalytically decompose BPA in an aqueous medium. Composite TiO 2 –zeolite sheets, however, demonstrated a higher efficiency for BPA removal from water than did zeolite-free TiO 2 sheets owing to the synergistic effect obtained through the combined use of TiO 2 and zeolite, as reported in our previous study. Furthermore, the total organic carbon (TOC) content of the system was reduced, evidently through the use of composite sheets, rather than a TiO 2 powder suspension or zeolite-free TiO 2 sheets. Trace amounts of intermediates resulting from BPA degradation were detected in the water treated with the composite sheets. In general, various intermediates are formed during the TiO 2 photodecomposition of organic compounds, which can potentially cause a secondary pollution because the degradation products sometimes turn out to be more hazardous than the parent compound. The composite TiO 2 –zeolite sheets prepared in the present study helped to overcome this problem by allowing the elimination of BPA without the release of hazardous intermediates into water: most of the intermediates were temporarily captured by the zeolite adsorbent contained in the composite sheets, and eventually decomposed through TiO 2 photocatalysis. Paper-like TiO 2 –zeolite composites clearly offer great advantages in efficacy and safety for the aqueous decomposition of BPA.

The PLLA and PDLLA samples consisted of three types of carbon: C-C (C-H), C-O and 0=C-O, which corresponded to their theoretical composition. However, the XPS analysis revealed that the surfaces of the PLA film highly resistant to acid hydrolysis were overspread with unknown constituents consisting of nitrogen and unoxidized carbon, not being announced officially. The AFM analysis offered a clear distinction in the surface deterioration of the PLA samples; and the alkaline treatment excavated numerous holes on the PLA surfaces. These results indicated that the commercial plastics are chemically modified on their surfaces to improve the physical properties for practical use, resulting in being deprived of biodegradability and eco-compatibility. It was also suggested that the enlargement in the specific surface area, which may occur at the early stage on PLA degradation, possibly accelerated the PLA decomposition rate. A new conception in the material design for paper-based composites was proposed; a local alkaline circumstance expected by alkaline fillers, e.g., CaO available in papermaking processes would

Chemical and biological degradation behavior of commercial polylactide (PLA) film, poly (L-lactide) (PLLA) and poly (DL-lactide) (PDLLA) was investigated by gravimetric analysis, X-ray diffractometry, differential scanning calorimetry and Fourier transform Raman (FT-Raman) spectroscopy. The PLA film exhibited an extremely high resistance to acid hydrolysis by hydrochloric acid, although alkaline treatment with a dilute sodium hydroxide brought about the quick decomposition of PLAs, not being reflected in their crystallinity. The X-ray and FT-Raman results implied the immediate decomposition occurring only on the PLA surfaces. These phenomena were observed both for crystalline PLLA and amorphous PDLLA; however, at temperature above their glass transition points, the PDLLA was greater deteriorated than the PLLA under acidic conditions, whereas in the alkaline system the tendency was reversed. On the biological degradation, the PLA filrn remained almost unchanged even after buried in soil for six months, as supported by various analytical results. Possibly, the PLA=related products commercially available have lower biodegradability than expected, and thus it was indicated that a strategic material design, e.g., the utilization of paper-based ecomaterials whose practicality and biodegradability can be compatible with each other, is of significance for high performance composite-manufacturing wlth biodegradable plastics.

Microperoxidase-11 (MP-11) derived from horse heart cytochrome c was utilized as a peroxidative catalyst. Mn II -oxidation activity of MP-11 was demonstrated for the first time. Although the activity of MP-11 was low compared to manganese peroxidase (MnP) from Phanerochaete chrysosporium in aqueous system, it can be utilized in water-miscible organic solvents if at least 5% water exists. Mn II -oxidation by MP-11 was optimized at higher pH compared to MnP, probably because of the lack of distal His acting as a proton acceptor during the catalytic cycle. Finally, the depolymerization of lignin was also achieved in 90% methanol using MP-11 in the presence of H 2 O 2 and Mn II .

Compared with conventional acidic rosin emulsion size, neutral rosin size has good sizing efficiency under neutral–alkaline conditions of paper manufacture because its emulsion particles are stable to the action of hydroxide ions. Therefore, the sizing efficiency of emulsion-type rosin sizes, i.e. their ability to increase resistance to fluid penetration, was considered to be related with their particulate structure. The supramolecular structure of the neutral glycerin rosin ester–rosin size microparticles and acidic rosin size microparticles were investigated by using differential scanning calorimetry (DSC), Fourier transform infrared (FT-IR) absorption spectroscopy, and electron spectroscopy for chemical analysis (ESCA) techniques in this study. The results indicated that the neutral glycerin rosin ester–rosin size is quite distinct from the acidic rosin sizes in their particulate supramolecular structure. A glycerol dirosinate bilayer existed in the particle structure is hypothesized for the neutral glycerin rosin ester–rosin size microparticles. The results also suggest that the glycerol dirosinate bilayer distributes on the outermost layer of the neutral rosin size microparticles, covering the rosin acid molecules, so as to prevent the dissociation of rosin by hydroxide ions under neutral–alkaline conditions. Consequently, the difference in the supramolecular structure of the rosin sizes particles resulted in their different stabilities and retention characteristics.

Zeolite sheets with different uniformities were prepared using a papermaking technique with different dual-polymer retention systems. The sheet uniformity was studied by light-transmission image analysis to calculate a formation index (FI). The lower the value of FI, the better the uniformity of the zeolite sheet. In this report, the relationship between the sheet adsorptivity with respect to acetaldehyde, one of the typical volatile organic compounds (VOCs), and the FI of the zeolite sheets was studied. The maximum adsorption capacity of the zeolite sheet depended only on the amount of zeolite in the sheet. However, the better the uniformity of zeolite sheet, the greater was the adsorptive efficiency. In particular, a uniform distribution of zeolite in the sheets led to a remarkable increase in the initial rate of acetaldehyde adsorption. Consequently, sheet uniformity is one of the most important factors determining the performance of zeolite sheets.

Electrochemical analysis of lignin peroxidase (LiP) was performed using a pyrolytic graphite electrode coated with peroxidase-embedded tributylmethyl phosphonium chloride membrane. The formal redox potential of ferric/ferrous couples of LiP was −126 mV (versus SHE), which was comparable with that of manganese peroxidase (MnP) and horseradish peroxidase (HRP). Yet, only LiP is capable of oxidizing non-phenolic substrates with a high redox potential. Since with decreasing pH, the redox potential increased, an incredibly low pH optimum of LiP as peroxidase at 3.0 or lower was proposed as the clue to explain LiP mechanisms. A low pH might be the key for LiP to possess a high redox potential. The pKa values for the distal His in peroxidases were calculated using redox data and the Nernst equation, to be 5.8 for LiP, 4.7 for MnP, and 3.8 for HRP. A high pKa value of the distal His might be crucial for LiP compound II to uptake a proton from the solvent. As a result, LiP is able to complete its catalytic cycle during the oxidation of non-proton-donating substrates. In compensation, LiP has diminished its reactivity toward hydrogen peroxide.

Titanium dioxide (TiO2) photocatalyst and zeolite adsorbent were made into a paper-like composite by a papermaking technique using pulp and ceramic fibers as sheet matrix. The photocatalytic performance for the degradation of bisphenol A (BPA) dissolved in water was investigated under UV irradiation. The TiO2 sheet prepared was easier to handle than the original TiO2 powders in aqueous media. The TiO2 sheet could decompose the BPA under UV irradiation, although at a lower degradation efficiency than the TiO2 suspension. The TiO2-free zeolite sheet could not remove the BPA from water completely because of its adsorption equilibrium. Furthermore, the composite TiO2-zeolite sheets exhibited a higher efficiency for BPA removal than the zeolite-free TiO2 sheets, the efficiency of the former being equivalent to that of the TiO2 suspension. The enhancement in removal efficiency was not attributed to the simple adsorption of BPA on zeolite but rather to the synergistic effect obtained through the combined use of TiO2 photocatalyst and zeolite adsorbent in the paper-like composite sheet, which is believed to accelerate the BPA photodegradation in water.

Factors influencing sizing response were investigated by the combined analysis of X-ray photoelectron spectroscopy (XPS) and pyrolysis gas chromatography (PyGC). Fatty acids (FAs) such as stearic acid (SA) and palmitic acid (PA) were used as models of rosin size, and the handsheets prepared beforehand with aluminum sulfate (alum) were soaked in the FAs⁄chloroform solution. Nearly no sizing effect appeared in the FAs-sized sheets without alum. The sizing degrees of the sized sheets increased with an increase in the internal alum addition level, although the size content determined by PyGC was almost constant regardless of the amount of alum added or a variety of FAs as sizing agents. However, the sizing response of the sheets sized by SA⁄PA mixture was quite higher than that of SA- or PA-sized sheets. High resolution XPS analysis revealed that both of internal alum addition and blending FAs brought about an increase in the atomic ratio of carbon originating from the size components present on the pulp fibers. As based on the XPS analytical principle that the detectable depth of photoelectrons is quite shallow (‹ 10 nm), it was possibly suggested that one of the factors affecting the sizing response was the subtle size aggregation over 10 nm in diameter on the sheet surfaces. A creative analysis of XPS⁄PyGC comparative quantification offered a new aspect in sizing mechanisms that the delicate behavior of size aggregation must govern the sizing response.

Sheets with a photocatalytic action were prepared using a papermaking technique. These sheets included single TiO2-sheet (Ti) and composite TiO2 sheets with A type zeolite (Ti-ZeA) and Y type zeolite (Ti-ZeY). A sheet (ZeA-Ti sol) containing A type zeolite treated with TiO2 sol was also prepared. Nitrogen monoxide (NO) was photocatalytically decomposed continuously when these sheets were used under UV irradiation. The subsequent photooxidation of NO to NO2 and of NO2 to HNO3 was observed. The NO x removal efficiencies of the sheets increased in the following order; Ti-ZeY < Ti < Ti-ZeA < ZeA-Ti sol. The reaction mechanism is discussed on the basis of the combined effects of the photocatalytic ability of TiO2 and the function of zeolites to adsorb NO and retain the products.

Microperoxidase-11 (MP-11), which is a heme-containing undecapeptide derived from horse heart cytochrome c, was utilized as a peroxidative catalyst in the system of bis(2-ethylhexyl)sulphosuccinate sodium salt (AOT)-reversed micelles to oxidatively decolorize water-insoluble dyes in isooctane. MP-11 entrapped in the reversed micelles exhibited a peroxidative activity in the presence of H2O2, effectively catalyzing the oxidative decolorization of an azo dye Solvent Yellow 7 and an anthraquinone dye Solvent Blue 11 in the hydrophobic organic solvent. H2O2-derived heme bleaching was successfully suppressed by replacing H2O2 with an organic hydroperoxide, tert-butyl hydroperoxide (t-BuOOH), thus sustaining a stable decolorization activity in the system. To optimize the reversed micellar system, the effect of the pH and the molar ratio of H2O/AOT hydration degree (Wo) was examined, indicating that MP-11 exhibited a maximal decolorization activity at pH 8–10 with the Wo value of 20.

A number of fluorescently labeled polymers have already been prepared. Since these polymers have usually been synthesized by copolymerization, the distribution of fluorescent groups will in such a case be heterogeneous because monomers always have different reactivity ratios. In this article, the preparation of fluorescently labeled cationic polyacrylamides (C-PAM) with homogeneous distributions of both fluorescent and cationic groups is described. Four fluorescently labeled C-PAM with isoindol, pyrene, anthracene, and dansyl groups were prepared in the homogeneous reaction system. These polymers were water soluble. Their wavelengths for maximum emission were in order of the above 380, 382 (smaller peak at 400), 435, and 540 nm, which thus covered a wide wavelength range. It is noteworthy that C-PAM with pyrene and anthracene groups have little fluorescence intensity at 540 nm, which is the maximum emission wavelength for C-PAM with dansyl group, making an accurate determination of the two polymers in the same solution possible. Even at the low degree of labeling, the detection limit of polymers is very low (approximately 1 ppm). © 2002 Wiley Periodicals, Inc. J Appl Polym Sci 86: 672–675, 2002

A near-infrared Fourier transform Raman (NIR-FTR) spectroscopic technique was utilized to determine the chemical structure of lignin in a woody matrix. In the NIR-FTR spectra of coniferaldehyde and coniferyl alcohol, the Raman bands for the carbonyl group and the alpha, beta unsaturated bond were detected at 1620 and 1660 cm(-1), respectively. These peaks were also found in the NIR-FTR spectra of chemically synthesized lignins, isolated lignin from conifer wood, and conifer wood meal. Upon the reduction of carbonyl groups in the lignin samples and wood meal, the band at 1620 cm(-1) disappeared; on the other hand, the band at 1660 cm(-1) remained unchanged. However, upon the oxidation of reduced lignin at the benzyl hydroxyl group using dicyanodichrolobenzoquinone, the band at 1620 cm(-1) clearly appeared, strongly suggesting that the band at 1620 cm(-1) can be assigned as a carbonyl marker band. The hydrogenation reaction optimized for the reduction of the unsaturated bond in lignin caused the disappearance of the band at 1660 cm(-1), indicating that the band at 1660 cm(-1) is an alpha, beta unsaturated bond marker band. The change in carbonyl content during the wood decay process was also shown to be monitored using the Raman intensity of the carbonyl marker band. It was indicated that the NIR-FTR spectroscopic techniques were suitable analytical method for a rapid and nondestructive analysis of wood samples.

The oxidation of bisphenol A [BPA: 2,2-bis(4-hydroxyphenyl)propane] catalyzed in organic solvents by laccase (polyphenoloxidase, EC 1.10.3.2) entrapped in a reversed micellar (RM) system was investigated. The laccase/RM system effectively catalyzed the oxidation reaction in isooctane, while laccase, lyophilized from an aqueous buffer solution in which its activity was optimized, exhibited no catalytic activity in nonaqueous media. To optimize the preparation and reaction conditions for the laccase/RM system, we examined the effects of pH in the water pools of RM, the concentration of laccase, and the degree of surfactant hydration ( W o ) on the laccase activity in organic media. The laccase/RM system showed a strong pH-dependency in organic media. Its optimum activity was obtained when the laccase/RM was prepared at pH=5. Interestingly, native laccase in an aqueous buffer solution exhibited an optimal activity at pH=3. The effect of a cosolubilized mediator (1-hydroxybenzotriazole, HBT) on the reaction was also investigated. BPA was effectively oxidized by the laccase/RM system in the absence of a mediator. On the other hand, the addition of 1-hydroxybenzotriazole strongly enhanced the catalytic performance of the laccase/RM system against the oxidation of various chlorophenols in organic media.

A cDNA encoding cytochrome P450 oxidoreductase (CPR) from the lignin-degrading basidiomycete Coriolus versicolor was identified using RT-PCR. The full-length cDNA consisted of 2,484 nucleotides with a poly(A) tail, and contained an open reading frame. The G+C content of the cDNA isolated was 60%. A deduced protein contained 730 amino acid residues with a calculated molecular weight of 80.7 kDa. The conserved amino acid residues involved in functional domains such as FAD-, FMN-, and NADPH-binding domains, were all found in the deduced protein. A phylogenetic analysis demonstrated that C. versicolor CPR is significantly similar to CPR of the basidiomycete Phanerochaete chrysosporium and that they share the same major branch in the fungal cluster. A recombinant CPR protein was expressed using a pET/ Escherichia coli system. The recombinant CPR protein migrated at 81 kDa on SDS polyacrylamide gel electrophoresis. It exhibited an NADPH-dependent cytochrome c reducing activity.

Benzaldehyde and its metabolic intermediates were effectively degraded by the brown-rot basidiomycetes Tyromyces palustris and Gloeophyllum trabeum. The pathway of benzaldehyde degradation was elucidated by the identification of fungal metabolites produced upon the addition of benzaldehyde and its metabolic intermediates. The oxidation and reduction occurred simultaneously, forming benzyl alcohol and benzoic acid as major products. Hydroxylation reactions, which seemed to be a key step, occurred on benzaldehyde and benzoic acid, but not on benzyl alcohol, to form corresponding 4-hydroxyl and 3,4-dihydroxyl derivatives. 1-Formyl derivatives were oxidized to 1-carboxyl derivatives at several metabolic stages. All of these reactions resulted in the formation of 3,4-dihydroxybenzoic acid. This was further metabolized via the decarboxylation reaction to yield 1,2,4-trihydroxybenzene, which may be susceptible to the ring-fission reaction. Ring-U-14C-labelled benzaldehyde and benzoic acid were effectively mineralized, clearly indicating that the brown-rot basidiomycetes are capable of metabolizing certain aromatic compounds to CO2 and H2O, despite the fact that brown-rot fungi cannot degrade polymeric lignin. Inhibitor experiments, using hydroxyl radical scavengers, catalase and cytochrome P450 inhibitors, strongly suggested that the aromatic hydroxylation reactions found in the brown-rot fungi are catalysed by intracellular enzyme(s), but not by Fenton-reaction-derived hydroxyl radicals.

Possible binding sites of lignin peroxidase (LiP) for veratryl alcohol (VA) were investigated by determining the reactivity of three different chemically modified LiPs against VA when acting (i) as a reducing substrate, (ii) as a rescuing reagent for the rapid conversion of LiPIII∗ back to native LiP, and (iii) as an enzyme-bound redox mediator. The enzyme was chemically modified to alter its surface properties by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) in the presence and the absence of 2-aminoethanesulfonic acid to introduce sulfo and N-acylurea groups instead of carboxylic groups, respectively. LiP was also modified by N-bromosuccinimide (NBS) to yield Trp-modified enzyme. The spectral characteristics and compound I formation rates of modified LiPs were identical to those of unmodified LiP. The activities for VA oxidation by modified LiPs were significantly reduced but with almost unchanged pH dependencies. Several other substrates including phenolic, anionic, and polymeric substrates were also utilized to characterize the activity of the modified enzymes. Kinetic analysis of these reactions strongly suggests that LiP has at least two substrate oxidation sites, one is Trp 171 for VA oxidation and the other is for anionic substrate oxidation. For VA-supported oxidation of ferric cytochrome c (Cc3+), the VA binding site was estimated to be the same as that of VA oxidation. The reactivities of chemically modified LiPs with excess H2O2 were investigated, indicating the formation of compound III∗ species. Compound III∗ species of EDC–LiP and unmodified LiP were converted back to ferric enzymes by adding VA. However, there was a reduced recovery of ferric NBS–LiP, suggesting that the VA binding site for VA-derived rapid reversion of compound III∗ is located at a different site from that for VA oxidation.

2 The catalytic conversion of dibenzothiophenen (DBT) to DBT-5-oxide was examined using microperoxidase-11 (MP-11) as well as a series of heme peroxidases such as horseradish, lignin and manganese peroxidases. MP-11 is a heme-containing oligopeptide obtained by enzymatic hydrolysis of horse heart cytochrome c. Among the catalysts examined, only MP-11 effectively oxidizes DBT to its oxide with an expense of hydrogen peroxide. The maximal oxidation rate was obtained in the aqueous media containing 30% methanol. During MP-11 catalyzed oxidation of DBT to DBT-5-oxide by phenylperacetic acid, phenylacetic acid was formed but not benzyl alcohol. This observation strongly suggested that MP-11 first reacts with peroxide to form compound I species. DBT oxidation by the MP-11/H2O2 system was then carried out either with H218O2 or under 18O2 atmosphere, indicating that the oxygen atom of DBT-5-oxide was derived from hydrogen peroxide but not from molecular oxygen or water. These results indicated that MP-11 oxidized DBT via the peroxygenation reaction, where the oxygen was transferred from the ferryl-oxy complex of the heme to DBT forming DBT-5-oxide.

Fiber charge characteristics of pulp suspensions containing aluminum sulfate were investigated with relation to adsorption behavior of aluminum components on the pulp fibers by streaming potential measurement using a particle charge detector, X-ray photoelectron spectroscopy, and X-ray fluorescence analysis. When aluminum sulfate was added to a pulp suspension prepared using deionized water, a streaming potential of the suspension went from negative to slightly negative according to the adsorption of aluminum components on the pulp fibers. Subsequent addition of a dilute NaOH solution to the suspension drastically cationized the fibers in the pH range of around 5 by predominant and homogeneous adsorption of cationic aluminum complexes on the fiber surfaces. However, the aluminum flocs that formed heterogeneously on the fiber surfaces at higher pH by further alkali addition made nearly no contribution to cationization of the fibers, although the abundant aluminum components were retained in the pulp sheets. Therefore, only aluminum cations adsorbed uniformly on the fiber surfaces perform well to control the charge properties of the pulp fibers at the wet end; and the preferential aluminum adsorption behavior on the fiber surfaces, by utilizing the required amounts of hydroxyl ions, probably accounts for the effective cationization of the fibers under acidic to neutral papermaking conditions.

Using a reverse-transcription polymerase chain reaction (RT-PCR) technique, cytochrome P450 genes were cloned from the lignin-degrading basidiomycete, Coriolus versicolor. One possible P450 gene was identified, which consisted of 1,672 nucleotides and a poly(A) tail and encoded a deduced protein containing 449 amino acids. The deduced amino acid sequence revealed the presence of the P450 heme-binding motif, strongly suggesting that this protein belongs to the P450 superfamily, then designated CYP512A1. The deduced protein showed sequential similarity to other known P450s from several micro-organisms, such as Aspergillus terreus, Gibberella fujikuroi, and Neurospora crassa, with 30–35% identity. Since the identity of the amino acid sequence was less than 40% with any other P450s, this protein was suggested to be the first member of a new family of cytochrome P450. In addition, a differential display RT-PCR analysis showed the expression of the other P450 genes, which were up-regulated by the addition of dibenzothiophene and 4-methyldibenzothiophene-5-oxide. Using the 5′ rapid amplification of cDNA ends method, a 520-nucleotide sequence, including the P450 motif-coding region, was determined for one clone. The deduced protein showed high similarity to CYP512A1 but less than 40% identity with P450s from other organisms. A chemical stress-responsive expression of P450 is suggested for the first time in basidiomycetes.

Composite titanium dioxide (TiO2)-zeolite sheets were prepared using a papermaking technique and were tested with regard to their ability to remove acetaldehyde, which is often used as a typical volatile organic compound. It was demonstrated that the composite TiO2-zeolite sheet could remove acetaldehyde much more effectively under UV irradiation than a sheet of TiO2 alone. Acetaldehyde removal by a composite TiO2-zeolite sheet with a TiO2/zeolite ratio of 1 : 4 was particularly effective under the conditions adopted in this work although the composite sheet contained only one fifth of the amount of TiO2 in the TiO2 sheet. Additionally, the combination of TiO2 and zeolite was a useful method to solve the disadvantage of adsorbents, which were the decrease of the performance when the material was saturated or equilibrated with the adsorbates. Consequently, a composite TiO2-zeolite sheet can be utilized as a high functional material to prevent environmental pollution.

The complete degradation of Yperite (bis(2-chloroethyl) sulfide), a chemical warfare agent, was achieved by two basidiomycetous cultures. Two distinct metabolic pathways were detected in each fungus during degradation of Yperite. The major path involved a non-enzymatic hydrolysis to generate thiodiglycol. In the minor path, the sulfide bond was cleaved prior to the hydrolytic dechlorination reaction, yielding chloroethanol and chloromercaptoethane, both of which were then metabolized completely.

A copolymer of glycidyl methacrylate and styrene (PGS) was synthesized by emulsion polymerization. The effect of the polymer on paper dry- and wet-strength improvement was investigated. The polymer contains phenyl groups that provide hydrophobic character to fibers so as to improve paper wet performance, and glycidyl groups that react with nucleophilic groups in fibers and other polymers to increase paper strength. Results showed that other polymers, such as polyallylamine, polyacrylic acid, and modified polyacrylamide, could be used together with PGS for paper-strength improvement. Furthermore, three kinds of partially carboxymethylated pulps (CMP) were prepared and used for PGS treatment. Compared with original pulp sheet, the sheets containing CMPs were strengthened greatly both in dry and in wet strength after PGS treatment. It was confirmed that higher CMP content got higher strength improvement at the same addition level of PGS. The results showed that carboxyl groups in pulp participate in the reaction with PGS and the cocrosslinkings between fibers and polymers were formed for improving paper strength. © 2001 John Wiley & Sons, Inc. J Appl Polym Sci 80: 334–339, 2001

Zeolite is generally used in the form of beads or pellets in various industrial fields. These types of use, however, have some serious disadvantages such as a pressure loss in the reaction tower or a powdering of the zeolite through beads rubbing against each other. Therefore, in this study, the zeolite sheets were prepared by using a papermaking technique intending the manufacture of the honeycomb structure which improved these defects to a great extent. Typical composites of sheetmaking stuffs were pulp (9%), zeolite (61.5%), kaolin (18%) and glass fiber (11.5%). When 0.5% polydiallyldimethylammonium chloride (cationic retention aid) and 0.5%–1.0% anionic polyacrylamide (anionic retention aid) with high molecular weight (4.0 × 106) on these solids were added, the retention of inorganic compounds was higher than 90%. The drainage time was adequately short. These results suggest that the method can be applied to the industrial production.

The surface analyses of the papers sized with a rosin emulsion (RE) and five typical polyamines, polyallylamine (PAAm), polyvinylamine (PVAm), polyethyleneimine (PEI), poly(diallyldimethylammonium chloride (PDADMAC), and poly(dimethylamino ethyl methacrylate) (PDMAEMA), were performed using a X-ray photoelectron spectrometer. The results show that the chemical states of the polyamines on the outermost surfaces of the papers changed significantly during the neutral rosin sizing. The five polyamines could react with carboxyl groups of the fibers to form urethane linkages at 400.0 ± 0.1 eV, which resulted in absorption of the polyamines onto the fibers. Only PAAm and PVAm could combine with RE to form urethane linkages at 400.7–0.1 eV, which were related deeply to sizing effectiveness. The polyamines distributed mainly on the surfaces of the fibers. The concentrations of PAAm and PVAm on the surfaces of the sized papers were much larger than those of PEI, PDMAEMA, and PDADMAC. A larger sizing degree was related to a larger C1 component at 285.0 eV (CC) in the C1s XPS spectra of the sized papers. © 2001 John Wiley & Sons, Inc. J Appl Polym Sci 80: 2185–2190, 2001