Hiroshi Kamitakahara

Syntheses and Comparison of 2,6-Di-O-methyl Celluloses from Natural and Synthetic Celluloses

2,6-Di-O-methylcellulose was prepared from natural and synthetic celluloses. Natural cellulose was converted to 2,6-di-O-thexyldimethylsilylcellulose, then to 3-mono-O-allyl-2,6-di-O-methylcellulose, and finally into 2,6-di-O-methylcellulose. Alternatively, 2,6 di-O-methylcellulose was synthesized from the synthetic cellulose derivative 3-mono-O-benzyl-2,6-di-O-pivaloylcellulose by depivaloylation and methylation to give 3-mono-O-benzyl-2,6-di-O-methylcellulose, which was debenzylated to yield the dimethyl ether. Both types of 2,6-di-O-methylcellulose are insoluble in water and common organic solvents. The structures of all cellulose derivatives were determined by NMR.

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.

Vibrational property changes of spruce wood by impregnation with watersoluble extractives of pernambuco (Guilandina echinata Spreng.) II: structural analysis of extractive components

Heartwood of pernambuco (Guilandina echinata Spreng, synCaesalpinia echinata Lam.), which has been used as material for violin bows, was extracted by soaking in water, and the obtained extractives were analyzed. The main components of the extractives were identified to be protosappanin B and brazilin. In particular, protosappanin B occupied about 40% of the pernambuco extractives. The loss tangent (tanδ) of spruce wood impregnated with protosappanin B decreased markedly, the same as that of specimens impregnated with extractives before being purified. It is expected that protosappanin B can make a contribution to the decrease in tanδ due to impregnation with extractives.

Cellular distribution of coniferin in differentiating xylem of Chamaecyparis obtusa as revealed by Raman microscopy

Abstract Cellular distribution of coniferin in differentiating xylem of Japanese cypress (Chamaecyparis obtusa) was analyzed by Raman microscopy. Small blocks were collected from differentiating xylem, frozen, cut on their surface with a sliding microtome, and then freeze-dried. Scanning electron microscopy showed numerous needle-like deposits in the tracheid lumina from the beginning of the S1 layer formation to the S2 layer-forming stage. The Raman spectrum of the deposits in the tracheid lumen was similar to that of coniferin. The presence of coniferin in a water extract from differentiating xylem was confirmed by matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy and 1H- and 13C-nuclear magnetic resonance spectra. Differential Raman spectra taken from samples before and after washing with water and dehydration in an ethanol showed that developing secondary walls contained coniferin during the S2 layer-forming stage and also after S3 layer formation. In contrast, coniferin was detected in the cell corner middle lamella during the S2 layer-forming stage, and the differential spectra were different from that of coniferin after S3 layer formation. The differential spectrum in this stage was similar to that of a dehydrogenation polymer of coniferyl alcohol prepared by the “zulauf” method (bulk polymerization). These results suggest that free lignin oligomers of the type bulk polymerizate might exist in the cell corner middle lamella during the S3 layer-forming stage and can be removed from specimens during washing and dehydration. The results can be interpreted in a way that no such oligomer exists in the secondary wall during the same stage owing to endwise addition of monolignols (in analogy to a “zutropf” polymerization).

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.

Studies on the Dehydrogenative Polymerizations of Monolignol β-glycosides. Part 3: Horseradish Peroxidase-Catalyzed Polymerizations of Triandrin and Isosyringin

Abstract Horseradish peroxidase–catalyzed dehydrogenative polymerizations of the p-hydroxyphenyl monolignol glucoside (triandrin (1P)) and the syringyl monolignol glucoside (isosyringin (1S)) resulted in the formation of water-soluble lignin-like polymers (DHPs). The polymerization of 1P gave highly polymerized DHPs in high yields as did previously reported polymerization of the guaiacyl monolignol glucoside (isoconiferin (1G)). It was shown that the hydrophilic D-glucose units of 1G and 1P contribute to a marked increase in the molecular weights of the resulting DHPs. On the other hand, the homogeneous phase polymerization of 1S, similar to the polymerization of sinapyl alcohol, gave DHPs with extremely low molecular masses in poor yields. Structural characterization indicated that the DHPs from 1P and 1S were lignin-like polymers containing glucosidic units on their sidechains. It was also confirmed that D-glucosyl units introduced onto the γ-position of monolignols do not significantly affect the electrochemical oxidizability and the kinetics of the HRP-catalyzed initial monomer consumption.

Studies on the dehydrogenative polymerizations of monolignol β-glycosides. Part 2: Horseradish peroxidase- catalyzed dehydrogenative polymerization of isoconiferin

Abstract Dehydrogenative polymerization of isoconiferin (IC; coniferyl alcohol γ-O-β-D-glucopyranoside) catalyzed by horseradish peroxidase (HRP) was carried out. The polymerization of IC proceeded in a homogeneous system, resulting in a water-soluble dehydrogenation polymer (IC-DHP). The degree of polymerization (DP) of IC-DHP was significantly higher than that of a standard dehydrogenative polymer (CA-DHP) obtained from coniferyl alcohol (CA) in a heterogeneous system. Under optimum conditions, the DP of IC-DHP was 44 (M n=1.5×104), whereas that for CA-DHP was only 11 (M n=3.0×103, as acetate). Spectroscopic analyses confirmed that IC-DHP has a lignin-like structure containing D-glucose moieties attached to the lignin side-chains. The D-glucose unit introduced into γ-O position of CA essentially influenced the water solubility and molecular mass of the resulting DHP.

Synthesis of β-O -4 type oligomeric lignin model compound by the nucleophilic addition of carbanion to the aldehyde group

A synthetic method for obtaining lignin oligomer that contains only the β-O-4 structure is described in detail. This method consists of three reaction steps: (1) the synthesis of t-butoxycarbonylmethyl vanillin (2), (2) the nucleophilic addition oligomerization of compound 2, and (3) the reduction of the oligomeric β-hydroxyl ester. In the first step, compound 2 was synthesized from vanillin in 96.8% yield. In the second step, compound 2 was oligomerized with commercial lithium diisopropylamide (LDA) to obtain oligomeric β-hydroxyl ester (3) in 87.2% yield; the repeating units of this oligomer were joined only by β-O-4 linkages as confirmed by nuclear magnetic resonance (NMR) spectroscopy. In the third step, the oligomeric β-hydroxyl ester (3) was reduced with LiAlH4 to give compound 4 in 42.4% yield. On the basis of NMR, matrix-assisted laser desorption ionization time-of-flight mass spectrometry, and gel permeation chromatography analyses of compound 4, it was concluded that compound 4 was an oligomeric lignin model compound containing only β-O-4 interunit linkages. The number average degree of polymerization (DPn) of obtained compound 4 was about 7.0 (Mw/Mn = 1.42). Using this oligomeric lignin model compound, conventional degradation and analytical methods will give new information.

Chemical structure elucidation of total lignins in woods. Part II: Analysis of a fraction of residual wood left after MWL isolation and solubilized in lithium chloride/N,N-dimethylacetamide

Abstract The residual wood meal left after extraction of milled wood lignin (MWL) was extracted with lithium chloride/N,N-dimethylacetamide, which is a well-known cellulose solvent, to afford a soluble fraction (cellulose-lignin fraction; CL) in 36.7% yield. The UV elution curve of CL acetate has the same profile as its refractive index (RI) elution curve. After partial degradation of CL by cellulase, the UV elution curve of CL acetate shifted to the low-molecular-mass region in a similar fashion as its RI elution curve. These results indicate that the lignin in CL (CL lignin) is chemically bonded to cellulose. On the other hand, half of the CL lignin was removed by xylanase treatment. It was concluded that approximately half of the CL lignin existed as a lignin-cellulose-xylan complex.

Fractionation and characterization of lignin-carbohydrate complexes (LCCs) of Eucalyptus globulus in residues left after MWL isolation. Part II: Analyses of xylan-lignin fraction (X-L)

Abstract The residual wood meal left after milled wood lignin (MWL) isolation [milled wood residue (MWR)] of 5-year-old Eucalyptus globulus was fractionated to afford a xylan-lignin fraction (X-L) in 2.9% yield (based on MWR) by the method reported previously. X-L was further fractionated with the lignin solvent 1,4-dioxane/water (9:1, v/v) to give a soluble fraction (XL-F1; 24.0%) and an insoluble fraction (XL-F1-residue; 74.6%; both yields based on X-L). XL-F1-residue was further extracted with the good xylan solvent dimethyl sulfoxide and the soluble fraction was termed XL-F2 (43.0%; based on the XL-F1-residue). XL-F1 was mainly composed of lignin with a small amount of xylan and it is similar to purified MWL, whereas XL-F2 was mainly composed of xylan with some amount of lignin and it is similar to a fraction that was prepared by the extraction of crude MWL with acetic acid [lignin-carbohydrate complex (LCC)-AcOH]. The two-dimensional heteronuclear single quantum coherence nuclear magnetic resonance spectra of XL-F1 and XL-F2 were interpreted that the former has α-ether-type lignin-carbohydrate (LC) linkages and the latter might have LC linkages of the phenyl glycoside type, which are different from those in LCC-AcOH.