Stone cell formation in the pedicel of pears and apples.

MAIN CONCLUSION: For the first time, stone cells in pear and apple pedicel were studied. The lignification of the pedicel outer part was correlated with flesh, and the secondary cell wall biosynthesis genes were activated. Fruit pedicels act as bridges between the fruit and the shoot. They have secondary thickened cell walls that presumably function in mechanical support, water and nutrient transport. Stone cells are cells with a secondary cell wall thickening. In pears, yet not in apples, the stone cells affect the flesh texture. There have been few reports on stone cell formation in pear and apple pedicels; therefore, we studied these cells for the first time. The apple pedicel had few stone cells in the cortex. The formation of stone cells in pear continued until seven weeks after flowering (WAF), and the density was significantly higher than in apple. The stone cell formation degree (SFD) of pear was 3.6-7.1 times higher than that of apple. Total lignin and lignin non-condensed structure (G and S units) content in the pear pedicle outer part was 1.5-2.7 times higher than that of the apple at harvest. The SFD of the pedicel outer part had a positive correlation with the G and S units content of the flesh. The total lignin and G and S units content between flesh and the pedicel outer part were positively correlated. Correlation analysis revealed a positive relationship between fruit and pedicel formation of the stone cells. The WGCNA showed that NST3 was linked to NAC028, MYB46, CESA, POD, LAC, and VSR6. These genes were highly expressed in the outer part of the pear pedicel, while they were suppressed in that issue of the apple at 4 WAF.

Possible mechanisms for the generation of phenyl glycoside-type lignin-carbohydrate linkages in lignification with monolignol glucosides.

The existence and formation of covalent lignin-carbohydrate (LC) linkages in plant cell walls has long been a matter of debate in terms of their roles in cell wall development and biomass use. Of the various putative LC linkages proposed to date, evidence of the native existence and formation mechanism of phenyl glycoside (PG)-type LC linkages in planta is particularly scarce. The present study aimed to explore previously overlooked mechanisms for the formation of PG-type LC linkages through the incorporation of monolignol glucosides, which are possible lignin precursors, into lignin polymers during lignification. Peroxidase-catalyzed lignin polymerization of coniferyl alcohol in the presence of coniferin and syringin in vitro resulted in the generation of PG-type LC linkages in synthetic lignin polymers, possibly via nucleophilic addition onto quinone methide (QM) intermediates formed during polymerization. Biomimetic lignin polymerization of coniferin via the ß-glucosidase/peroxidase system also resulted in the generation of PG-type as well as alkyl glycoside-type LC linkages. This occurred via non-enzymatic QM-involving reactions and also via enzymatic transglycosylations involving ß-glucosidase, which was demonstrated by in-depth structural analysis of the synthetic lignins by two-dimensional NMR. We collected heteronuclear single-quantum coherence (HSQC) NMR for native cell wall fractions prepared from pine (Pinus taeda), eucalyptus (Eucalyptus camaldulensis), acacia (Acacia mangium), poplar (Populus × eurarnericana) and bamboo (Phyllostachys edulis) wood samples, which exhibited correlations, albeit at low levels, that were well matched with those of the PG-type LC linkages in synthetic lignins incorporating monolignol glucosides. Overall, our results provide a molecular basis for feasible mechanisms for the generation of PG-type LC linkages from monolignol glucosides and further substantiates their existence in planta.

Two-dimensional NMR analysis of Angiopteris evecta rhizome and improved extraction method for angiopteroside.

INTRODUCTION: The rhizome of Angiopteris evecta is of academic interest in Kalimantan, Indonesia, from an ethnobotanical perspective. Angiopteroside is a substance of pharmaceutical importance that is found in the rhizome of A. evecta. OBJECTIVE: The aims of this research are to improve the extraction method for angiopteroside from the rhizome, compared to that in a previous report, and to determine the yield of angiopteroside from the rhizome of A. evecta, as well as to obtain precise data for extractives from the rhizome of A. evecta by using two-dimensional NMR spectroscopy and liquid chromatography-mass spectrometry (LC-MS). METHODOLOGY: We investigated the chemical constituents of the whole rhizome by means of two-dimensional NMR (heteronuclear single quantum coherence or HSQC) spectroscopy, neutral sugar analysis using the alditol acetate method, and lignin analysis using alkaline nitrobenzene oxidation and Klason lignin methods. LC-MS revealed the purity of the angiopteroside. Antimicrobial assays were also performed for the purified angiopteroside by using a broth microdilution method. RESULTS: Angiopteroside was isolated by Soxhlet extraction with aqueous acetone followed by preparative thin-layer chromatography (eluent: 20% methanol/dichloromethane). LC-MS revealed that angiopteroside can be found in the rhizome of A. evecta in 9.9% yield, which is an extremely high yield for a plant extractive. CONCLUSION: HSQC analysis is a powerful tool for surveying compounds in plant materials, such as the whole rhizome of A. evecta. Soxhlet extraction with aqueous acetone is an effective method for extracting glycosides from plant materials.

Expansion of human γδ T cells for adoptive immunotherapy using a bisphosphonate prodrug.

Cancer immunotherapy with human γδ T cells expressing Vγ2Vδ2 T cell receptor (also termed Vγ9Vδ2) has shown promise because of their ability to recognize and kill most types of tumors in a major histocombatibility complex (MHC) -unrestricted fashion that is independent of the number of tumor mutations. In clinical trials, adoptive transfer of Vγ2Vδ2 T cells has been shown to be safe and does not require preconditioning. In this report, we describe a method for preparing highly enriched human Vγ2Vδ2 T cells using the bisphosphonate prodrug, tetrakis-pivaloyloxymethyl 2-(thiazole-2-ylamino)ethylidene-1,1-bisphosphonate (PTA). PTA stimulated the expansion of Vγ2Vδ2 cells to purities up to 99%. These levels were consistently higher than those observed after expansion with zoledronic acid, the most commonly used stimulator for clinical trials. Cell numbers also averaged more than those obtained with zoledronic acid and the expanded Vγ2Vδ2 cells exhibited high cytotoxicity against tumor cells. The high purity of Vγ2Vδ2 cells expanded by PTA increased engraftment success in immunodeficient NOG mice. Even low levels of contaminating αß T cells resulted in some mice with circulating human αß T cells rather than Vγ2Vδ2 cells. Vγ2Vδ2 cells from engrafted NOG mice upregulated CD25 and secreted tumor necrosis factor-α and interferon-γ in response to PTA-treated tumor cells. Thus, PTA expands Vγ2Vδ2 T cells to higher purity than zoledronic acid. The high purities allow the successful engraftment of immunodeficient mice without further purification and may speed up the development of allogeneic Vγ2Vδ2 T cell therapies derived from HLA-matched normal donors for patients with poor autologous Vγ2Vδ2 T cell responses.

A Single Subcutaneous Injection of Cellulose Ethers Administered Long before Infection Confers Sustained Protection against Prion Diseases in Rodents.

Prion diseases are fatal, progressive, neurodegenerative diseases caused by prion accumulation in the brain and lymphoreticular system. Here we report that a single subcutaneous injection of cellulose ethers (CEs), which are commonly used as inactive ingredients in foods and pharmaceuticals, markedly prolonged the lives of mice and hamsters intracerebrally or intraperitoneally infected with the 263K hamster prion. CEs provided sustained protection even when a single injection was given as long as one year before infection. These effects were linked with persistent residues of CEs in various tissues. More effective CEs had less macrophage uptake ratios and hydrophobic modification of CEs abolished the effectiveness. CEs were significantly effective in other prion disease animal models; however, the effects were less remarkable than those observed in the 263K prion-infected animals. The genetic background of the animal model was suggested to influence the effects of CEs. CEs did not modify prion protein expression but inhibited abnormal prion protein formation in vitro and in prion-infected cells. Although the mechanism of CEs in vivo remains to be solved, these findings suggest that they aid in elucidating disease susceptibility and preventing prion diseases.

Preparation of a Near-Infrared Ray Absorption Film from N-Phenylthiocarbamoyl Chitosan Derivative.

We recently observed that the decanoylation of N-phenylthiocarbamoyl chitosan (2) with a mixture of decanoic anhydride and pyridine at 60 °C for 24 h afforded N,N-(decanoyl)phenythiocarbamoyl-/2-isothiocynato chitosan decanoate (3b) rather than the expected product N,N-(decanoyl)phenylthiocarbamoyl chitosan decanoate (3a). This result suggested that some of the N,N-(decanoyl)phenylthiocarmbamoyl groups had been converted to isothiocyanate groups during the decanoylation process. The subsequent reaction of compound 3b with aniline gave N,N-(decanoyl)phenylthiocarbamoyl/N-phenylthiocarbamoyl chitosan decanoate (4) in high yield. A solution of compound 4 in CHCl3 was then added to a solution of copper decanoate (5) in the same solvent, and the resulting mixture was cast onto a glass plate to give a cast film. The film was annealed at 200 °C in an oven to give a greenish film, which showed good near-infrared absorption characteristic in the range of 800-2200 nm.

Selective cross-metathesis of cellobiose derivatives with amido-functionalized olefinic structures: A model study for synthesis of cellulosic diblock copolymers.

This work describes a model study for synthesis of cellulose-based block copolymers, investigating selective coupling of peracetyl ß-d-cellobiose and perethyl ß-d-cellobiose at their reducing-ends by olefin cross-metathesis (CM). Herein we explore suitable pairs of ω-alkenamides that permit selective, quantitative coupling by CM. Condensation reactions of hepta-O-acetyl-ß-d-cellobiosylamine or hepta-O-ethyl-ß-d-cellobiosylamine with acyl chlorides afforded the corresponding N-(ß-d-cellobiosyl)-ω-alkenamide derivatives with an aromatic olefin or linear olefinic structures. Among the introduced olefinic structures, CM of the undec-10-enamide (Type I olefin) and the acrylamide (Type II olefin) gave the hetero-block tetramers, N-(hepta-O-ethyl-ß-d-cellobiosyl)-N'-(hepta-O-acetyl-ß-d-cellobiosyl)-alkene-α,ω-diamides, with >98 % selectivity. Moreover, selectivity was not influenced by the cellobiose substituents when a Type I olefin with a long alkyl tether was used. Although the amide carbonyl group could chelate the ruthenium atom and reduce CM selectivity, the results indicated that such chelation is suppressed by sterically hindered pyranose rings or the long alkyl chain between the amido group and the double bond. Based on this model study, selective end-to-end coupling of tri-O-ethyl cellulose and acetylated cellobiose was accomplished, proving the concept that this model study with cellobiose derivatives is a useful signpost for selective synthesis of polysaccharide-based block copolymers.

Immunolocalization of 8-5' and 8-8' linked structures of lignin in cell walls of Chamaecyparis obtusa using monoclonal antibodies.

Mouse monoclonal antibodies were generated against dehydrodiconiferyl alcohol- or pinoresinol-p-aminohippuric acid (pAHA)-bovine serum albumin (BSA) conjugate as probes that specifically react with 8-5' or 8-8' linked structure of lignin in plant cell walls. Hybridoma clones were selected that produced antibodies that positively reacted with dehydrodiconiferyl alcohol- or pinoresinol-pAHA-BSA and negatively reacted with pAHA-BSA and guaiacylglycerol-beta-guaiacyl ether-pAHA-BSA conjugates containing 8-O-4' linkage. Eight clones were established for each antigen and one of each clone that positively reacted with wood sections was selected. The specificity of these antibodies was examined by competitive ELISA tests using various lignin dimers with different linkages. The anti-dehydrodiconiferyl alcohol antibody reacted specifically with dehydrodiconiferyl alcohol and did not react with other model compounds containing 8-O-4', 8-8', or 5-5' linkages. The anti-pinoresinol antibody reacted specifically with pinoresinol and syringaresinol and did not react with the other model compounds containing 8-O-4', 8-5', or 5-5' linkages. The antibodies also did not react with dehydrodiconiferyl alcohol acetate or pinoresinol acetate, indicating that the presence of free phenolic or aliphatic hydroxyl group was an important factor in their reactivity. In sections of Japanese cypress (Chamaecyparis obtusa), labeling by the anti-dehydrodiconiferyl alcohol antibody was found in the secondary walls of phloem fibers and in the compound middle lamellae, and secondary walls of tracheids. Weak labeling by the anti-pinoresinol antibody was found in secondary walls of phloem fibers and secondary walls and compound middle lamellae of developed tracheids. These labelings show the localization of 8-5' and 8-8' linked structure of lignin in the cell walls.

Thermoresponsive hydrogel of diblock methylcellulose: formation of ribbonlike supramolecular nanostructures by self-assembly.

This article provides detailed insight into the thermoresponsive gelation mechanism of industrially produced methylcellulose (MC), highlighting the importance of diblock structure with a hydrophobic sequence of 2,3,6-tri-O-methyl-glucopyranosyl units for this physicochemical property. We show herein, for the first time, that well-defined diblock MC self-assembles thermoresponsively into ribbonlike nanostructures in water. A cryogenic transmission electron microscopy (cryo-TEM) technique was used to detect the ribbonlike nanostructures formed by the diblock copolymers consisting of hydrophilic glucosyl or cellobiosyl and hydrophobic 2,3,6-tri-O-methyl-cellulosyl blocks, methyl ß-D-glucopyranosyl-(1→4)-2,3,6-tri-O-methyl-celluloside 1 (G-236MC, DP(n) = 10.7, DS = 2.65), and methyl ß-D-glucopyranosyl-(1→4)-ß-D-glucopyranosyl-(1→4)-2,3,6-tri-O-methyl-celluloside 2 (GG-236MC, DP(n) = 28.2, DS = 2.75). Rheological measurements revealed that the gel strength of a dispersion of GG-236MC (2, 2.0 wt %) in water at 70 °C was 3.0 times stronger than that of commercial MC SM-8000, although the molecular weight of GG-236MC (2) having M(w) = 8 × 10(3) g/mol was 50 times smaller than that of SM-8000 having M(w) = 4 × 10(5) g/mol. Cryo-TEM observation suggested that the hydrogel formation of the diblock copolymers could be attributed to the entanglement of ribbonlike nanostructures self-assembled by the diblock copolymers in water. The cryo-TEM micrograph of GG-236MC (2) at 5 °C showed rectangularly shaped nanostructures having a thickness from 11 to 24 nm, although G-236MC (1) at 20 °C showed no distinct self-assembled nanostructures. The ribbonlike nanostructures of GG-236MC (2) having a length ranging from 91 to 864 nm and a thickness from 8.5 to 27.1 nm were detected above 20 °C. Small-angle X-ray scattering measurements suggested that the ribbonlike nanostructures of GG-236MC (2) consisted of a bilayer structure with a width of ca. 40 nm. It was likely that GG-236MC (2) molecules were oriented perpendicularly to the long axis of the ribbonlike nanostructure. In addition, wide-angle X-ray scattering measurements revealed that GG-236MC (2) in its hydrogel formed the same crystalline regions as 2,3,6-tri-O-methylcellulose. The influence of the DP of diblock MC with a DS of around 2.7 on the gelation behavior will be discussed.

2-O-Methyl- and 3,6-di-O-methyl-cellulose from natural cellulose: synthesis and structure characterization.

For the first time, 2-O-methyl- (2MC) and 3,6-di-O-methyl-cellulose (36MC) were synthesized via 3-O-allyl- and 3-O-methyl-cellulose, respectively. Position 6 of 3-O-allyl- and 3-O-methyl-cellulose was protected with the 4-methoxytrityl groups. The reaction time and temperature were optimized to achieve a high regioselectivity at C-6 and to prevent the introduction of the 4-methoxytrityl group at C-2 of the polymer. It was found that the substituent at C-3 of 3-O-functionalized celluloses influenced the reactivity of the hydroxyl group at C-6. The structure was characterized by means of (1)H and (13)C NMR spectroscopy of the acetates of 2MC and 36MC. 2MC and 36MC were soluble in water and did not show thermoreversible gelation.

Synthesis-property-performance relationships of multifunctional bacterial cellulose composites fermented in situ alkali lignin medium.

This work demonstrates a unique approach of utilizing alkali lignin (AL), as smart additive to in situ BC fermentation in which it concurrently acts as promoter to microbial growth as well as reinforcing filler for fabrication of multifunctional composites. Traditionally, BC fermentation is accompanied by inhibitor formation with sudden drop in pH leading to low yield and biomass growth. AL due to its antioxidant nature prevents formation of gluconic acid as byproduct, at ∼0.25 wt.% AL based on inhibitory byproduct kinetics. Interestingly, AL self-assembles to form primary and secondary structures in BC pores, resulting in simultaneous improvement in thermal stability as well as toughness. The BC/AL films show strong UV-blocking capacity with prolonged radical scavenging activity and preventing browning of freshly cut apples making it suitable as food packaging. Therefore, present work opens up new avenues for fabrication of high-performance BC-based composites through selection of smart materials which can simultaneously improve BC bioprocessing.

Synthesis of polysaccharide-based block copolymers via olefin cross-metathesis.

Polysaccharide-based copolymers with brush-like, graft architectures have been prepared by many investigators. In contrast, it is challenging to prepare linear polysaccharide-based block copolymers. Only a few approaches have been reported for preparation of such architectures, despite the clear application potential of renewable-based, linear block copolymers. The challenging nature of regioselective polysaccharide end-group functionalization has impeded their preparation. Herein we report a different, flexible approach to linear block copolymers, demonstrated for derivatives of renewable, natural cellulose. To illustrate this approach, trimethyl cellulose-b-poly(ethylene glycol), trimethyl cellulose-b-poly(tetrahydrofuran) and trimethyl cellulose-b-poly(lactic acid) were synthesized by a coupling strategy. Trimethyl cellulose was functionalized regiospecifically at the reducing end anomeric carbon to create an ω-unsaturated alkyl acetal by solvolysis of trimethyl cellulose with an ω-unsaturated alcohol. Subsequently, mild and versatile olefin cross-metathesis was used to couple efficiently (100% conversion) the trimethyl cellulose block with a series of acrylate derivatives, including acrylated polymer blocks. The targeted block co-polymeric structures were confirmed by nuclear magnetic resonance (NMR) spectroscopy, and the degree of polymerization was relatively well-preserved during the coupling step, as shown by size exclusion chromatography (SEC). The results confirm that this new solvolysis/cross-metathesis method is a promising approach to polysaccharide-based linear AB, and likely ABC, block copolymers.

Analysis of Fruit Lignin Content, Composition, and Linkage Types in Pear Cultivars and Related Species.

Lignin content, composition, and linkage types were investigated in pear fruit cultivars and related species. Lignin content increased during early stages and then decreased toward ripening in the core and flesh of "Gold Nijisseiki" and "Alexandrine Douillard". The lignin content was highest at harvest in Chinese quince. Only trace amounts of lignin were detected in apple flesh. The lignin content was low in Japanese pears "Ohshu", "Hosui", and "Kosui", and the noncondensed lignin index was high in flesh. The lignin type was guaiacyl-syringyl (GS) in these pears and related species. The S/G ratio at harvest varied widely (0.75-2.64) and increased during early stages and remained constant toward harvest in "Gold Nijisseiki" and "Alexandrine Douillard". "Gold Nijisseiki" and "Alexandrine Douillard" were determined to be G- and S-lignin-rich types, respectively. ß-Aryl ether, phenylcoumaran, and resinol interunit linkage types were detected among monolignol bonds, and ß-Aryl ether units were the main linkages in the pear.

Cellulosic graft copolymer: poly(methyl methacrylate) with cellulose side chains.

A cellulose macromonomer, N-(15-methacryloyloxypentadecanoyl)-tri-O-acetyl-beta-cellulosylamine (CTA-C15-MA (2); M(2)) with number averaged degree of polymerization (DP(n)) = 13), was copolymerized with methyl methacrylate (MMA; M(1)) to give cellulosic copolymer with CTA side chains, PMMA-g-(CTA-C15) (3-A). An absolute molecular weight determined by multiangle laser light scattering (MALS; M(w,LS)), degree of polymerization of MMA (X) and CTA-C15-MA (Y) of PMMA-g-(CTA-C15) (3-A) were determined to be M(w,LS) = 6.30 x 10(4), X = 4.14 x 10(2), and Y = 3.86. Cellulose graft copolymer with cellulose side chains, PMMA-g-(cellulose-C15) (3-H) was successfully obtained after deacetylation of the copolymer 3-A. Thermal analysis of copolymers 3-A and 3-H by means of differential scanning calorimetry (DSC) measurements revealed that a small amount of CTA and cellulose side chains affected thermal properties of the PMMA main chain.

Basic study for acyl chitosan isothiocyanates synthesis by model experiments using glucosamine derivatives.

Affecting factors to the acyl chitosan isothiocyanate synthesis by N-phenylthiocarbamoylation and the following acylation was investigated using octyl 2-amino-2-deoxy-ß-D-glucopyranoside as a model compound. It was found from the acetylation of N-phenylthiocarbamoyl glucosamine derivative with acetic anhydride/pyridine that the glucosamine isothiocyanate was formed via N,N-(acetyl)phenylthiocarbamoyl glucosamine derivative and the conversion of N,N-(acetyl)phenylthiocarbamoyl glucosamine derivative to the glucosamine isothiocyanate proceeded mainly by thermal degradation of N,N-(acetyl)phenylthiocarbamoyl groups. The reaction temperature was an important factor to the isothiocyanate synthesis. On the other hand, it was found from the acetylation of N-ethylthiocarbamoyl derivative (or N-allylthiocarbamoyl derivative) with acetic anhydride/pyridine that the glucosamine isothiocyanate was also prepared from N-ethylthiocarbamoyl glucoamine derivative (or N-allylthiocarbamoyl glucosamine derivative), but that N-phenylthiocarbamoylation was more preferable to the isothiocyanate synthesis than N-ethylthiocarbamoylation and N-allylthiocarbamoylation. Then, acylation products of N-phenylthiocarbamoyl chitosan (N,N-(hexanoyl)phenylthiocarbamoyl chitosan hexanoate or hexanoyl chitosan isothiocynate) could be controlled by the reaction temperature and reaction system.

Preparation and electro-optical properties of triphenylamine-bound chitosan derivative.

3,6-Di-O-hexanoyl-N-[4-(N,N-diphenylamino)-1-phenyl] thiocarbamoyl chitosan was prepared from 3,6-di-O-hexanoyl chitosan isothiocyanate in a 78% yield, and spin-coated films of the chitosan derivative and tris(2-phenylpyridine)iridium (Ir(ppy)3) were fabricated. Ultraviolet-visible absorption spectra and photoluminescence spectra of the films indicated efficient Förster energy transfer from the chitosan derivative to the Ir(ppy)3. An electroluminescent device using both compounds emitted green luminescence when voltage was applied. The results suggested that the regio-selectively substituted chitosan derivative could be used as a scaffold in the emitting layer of organic light emitting diode.

Thermo-reversible supramolecular hydrogels of trehalose-type diblock methylcellulose analogues.

This paper describes the design and synthesis of new trehalose-type diblock methylcellulose analogues with nonionic, cationic, and anionic cellobiosyl segments, namely 1-(tri-O-methyl-cellulosyl)-4-[ß-d-glucopyranosyl-(1→4)-ß-d-glucopyranosyloxymethyl]-1H-1,2,3-triazole (1), 1-(tri-O-methyl-cellulosyl)-4-[(6-amino-6-deoxy-ß-d-glucopyranosyl)-(1→4)- 6-amino-6-deoxy-ß-d-glucopyranosyloxymethyl]-1H-1,2,3-triazole (2), and 4-(tri-O-methyl-cellulosyloxymethyl)-1-[ß-d-glucopyranuronosyl-(1→4)-ß-d-glucopyranuronosyl]-1H-1,2,3-triazole (3), respectively. Aqueous solutions of all of the 1,2,3-triazole-linked diblock methylcellulose analogues possessed higher surface activities than that of industrially produced methylcellulose and exhibited lower critical solution temperatures, that allowed the formation of thermoresponsive supramolecular hydrogels at close to human body temperature. Supramolecular structures of thermo-reversible hydrogels based on compounds 1, 2, and 3 were investigated by means of scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Detailed structure-property-function relationships of compounds 1, 2, and 3 were discussed. Not only nonionic hydrophilic segment but also ionic hydrophilic segments of diblock methylcellulose analogues were valid for the formation of thermo-reversible supramolecular hydrogels based on end-functionalized methylcellulose.

The application of microwave heating to the synthesis of 6-amino-6-deoxycellulose.

Microwave heating was applied to the reactions involved in the synthesis of 6-amino-6-deoxycellulose, 4. These included, cellulose solubilization, bromination at C-6, displacement of bromine with azide ion, and reduction of the azido group to an amino group. Compared to conventional heating, this approach had the advantages of shortening reaction times and retaining the degree of polymerization of 4.

Pyrene conjugation and spectroscopic analysis of hydroxypropyl methylcellulose compounds successfully demonstrated a local dielectric difference associated with in vivo anti-prion activity.

Our previous study on prion-infected rodents revealed that hydroxypropyl methylcellulose compounds (HPMCs) with different molecular weights but similar composition and degree of substitution have different levels of long-lasting anti-prion activity. In this study, we searched these HPMCs for a parameter specifically associated with in vivo anti-prion activity by analyzing in vitro chemical properties and in vivo tissue distributions. Infrared spectroscopic and thermal analyses revealed no differences among HPMCs, whereas pyrene conjugation and spectroscopic analysis revealed that the fluorescence intensity ratio of peak III/peak I correlated with anti-prion activity. This correlation was more clearly demonstrated in the anti-prion activity of the 1-year pre-infection treatment than that of the immediate post-infection treatment. In addition, the intensity ratio of peak III/peak I negatively correlated with the macrophage uptake level of HPMCs in our previous study. However, the in vivo distribution pattern was apparently not associated with anti-prion activity and was different in the representative tissues. These findings suggest that pyrene conjugation and spectroscopic analysis are powerful methods to successfully demonstrate local dielectric differences in HPMCs and provide a feasible parameter denoting the long-lasting anti-prion activity of HPMCs in vivo.

Distribution of coniferin in differentiating normal and compression woods using MALDI mass spectrometric imaging coupled with osmium tetroxide vapor treatment.

Matrix-assisted laser desorption/ionization mass spectrometric imaging (MALDI-MSI) was employed to detect monolignol glucosides in differentiating normal and compression woods of two Japanese softwoods, Chamaecyparis obtusa and Cryptomeria japonica Comparison of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry collision-induced dissociation fragmentation analysis and structural time-of-flight (MALDI-TOF CID-FAST) spectra between coniferin and differentiating xylem also confirmed the presence of coniferin in differentiating xylem. However, as matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and MALDI-TOF CID-FAST spectra of sucrose were similar to those of coniferin, it was difficult to distinguish the distribution of coniferin and sucrose using MALDI-MSI and collision-induced dissociation measurement only. To solve this problem, osmium tetroxide vapor was applied to sections of differentiating xylem. This vapor treatment caused peak shifts corresponding to the introduction of two hydroxyl groups to the C=C double bond in coniferin. The treatment did not cause a peak shift for sucrose, and therefore was effective in distinguishing coniferin and sucrose. Thus, it was found that MALDI-MSI combined with osmium tetroxide vapor treatment is a useful method to detect coniferin in differentiating xylem.