Biomimetic Oxidative Coupling of Sinapyl Acetate by Silver Oxide

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Biomimetic oxidative coupling of sinapyl acetate by silver oxide: preferential formation of #-O-4 type structures Takao Kishimoto, Nana Takahashi, Masahiro Hamada, and Noriyuki Nakajima J. Agric. Food Chem., Just Accepted Manuscript • DOI: 10.1021/jf506111q • Publication Date (Web): 05 Feb 2015 Downloaded from http://pubs.acs.org on February 18, 2015

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Journal of Agricultural and Food Chemistry

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Biomimetic Oxidative Coupling of Sinapyl Acetate by Silver Oxide:

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Preferential Formation of β-O-4 Type Structures

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Takao Kishimoto,*,† Nana Takahashi, †,‡ Masahiro Hamada† and Noriyuki Nakajima†

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†

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Prefectural University, Imizu 939-0398, Japan

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‡

Bioorganic Chemistry Laboratory, Department of Biotechnology, Faculty of Engineering, Toyama

Nitto Medic Co., Ltd., Toyama 939-2366, Japan.

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Corresponding Author *Phone: +81-766-56-7500 (ext. 567). Fax: +81-766-56-2498. E-mail: [email protected]

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ABSTRACT: Biomimetic oxidations of sinapyl alcohol and sinapyl acetate were carried out with Ag2O

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to better understand the high frequency of β-O-4 structures in highly acylated natural lignins. The major

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products from the Ag2O oxidation of sinapyl alcohol were sinapyl aldehyde (14% yield), β-O-4-coupled

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dimer (32% yield) and β-β-coupled dimer (3% yield). In contrast, the Ag2O oxidation of sinapyl acetate

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produced β-O-4-coupled dimer in 66% yield. Oligomeric products with predominantly β-O-4 structures

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were also obtained in 18% yield. The yield of the β-O-4-coupled products from sinapyl acetate was

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much higher than that from sinapyl alcohol. Computational calculations based on density functional

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theory showed that the negative charge at Cβ was significantly reduced by the γ-acetyl group. The

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computational calculations suggest that the Coulombic repulsion between Cβ and O4 in sinapyl acetate

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radicals was significantly reduced by the γ-acetyl group, contributing to the preferential formation of β-

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O-4 structures from sinapyl acetate.

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KEYWORDS: β-O-4, acylated lignin, sinapyl alcohol, DFT, Coulombic repulsion

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INTRODUCTION

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Lignin is one of the major components in higher plant cell walls. There are three monomers of lignin

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(monolignols): coniferyl alcohol, 1, sinapyl alcohol, 2 and p-coumaryl alcohol, 3 (Figure 1). These

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monolignols contribute to the formation of complicated macromolecular lignins. Softwood lignins are

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predominantly produced from coniferyl alcohol (guaiacyl lignin). Hardwood lignins are made from both

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coniferyl alcohol and sinapyl alcohol (guaiacyl-syringyl lignin). Grass (Gramineae) lignins are produced

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from coniferyl alcohol, sinapyl alcohol and p-coumaryl alcohol.1 The polymerisation of these

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monolignols produces several interunit linkages in lignin, including β-O-4, β-5, β-β, 4-O-5 and 5-5

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linkages. Among the interunit linkages, the β-O-4 linkage is the most important for the biomass

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conversion because of the lability toward various treatments. The frequency of β-O-4 linkages is about

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50–60% in softwood and hardwood lignins.1 We have investigated the β-O-4 structures and synthesized

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artificial lignin polymers composed only of the β-O-4 linkages.2-4

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In addition to the typical monolignols, sinapyl acetate, 4, sinapyl p-coumarate, 5 and sinapyl p-

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hydroxybenzoate, 6 are considered to be lignin monomers (Figure 1).5 These non-traditional

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monolignols are also thought to be polymerized by peroxidase or laccase to produce γ-acylated lignins in

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many natural plants. Kenaf (Hibiscus cannabinus) lignin was found to be a highly γ-acetylated lignin

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with a high syringyl-to-guaiacyl (S/G) ratio.6 More interestingly, the β-O-4 structures predominate other

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substructures such as β-β and β-5, and the frequency of the β-O-4 structures reaches 80% in kenaf

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lignin.6 Recently, highly acylated (acetylated and/or p-coumaroylated) lignins were isolated from the

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herbaceous plants sisal (Agave sisalana), kenaf, abaca (Musa textilis) and curaua (Ananas erectifolius).7

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The structures of all these highly acylated lignins were characterized by a very high S/G ratio, the

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significant predominance of β-O-4 structures (up to 94%) and a low proportion of the traditional β-β

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structure.7 Miscanthus (Miscanthus x giganteus) native lignin is also reported to be highly acylated and

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have a high frequency of β-O-4 structures.8 These findings suggest that acylated monolignols may

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couple to produce β-O-4 structures more frequently than non-acylated monolignols. ACS Paragon Plus Environment

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In order to understand lignin biosynthesis in plant cell walls, the in vitro oxidative couplings of

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monolignols have long been investigated using horseradish peroxidase (HRP). In our previous study, the

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effect of the S/G ratio on the structures of natural and synthetic lignin was investigated.9 The presence of

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both guaiacyl and syringyl units was effective for producing higher amounts of β-O-4 structures.9 On the

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other hand, the biomimetic oxidation of acylated sinapyl alcohol using HRP has also been reported, and

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the formation of non-traditional β-β coupled dimers were found.10 However, neither the HRP/peroxide

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oxidation of sinapyl acetate nor p-hydroxybenzoate11 effectively formed β-O-4-coupled products. It was

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also reported that the HRP/peroxide oxidation of sinapyl alcohol preferentially produces β-β-coupled

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products.12 These in vitro experimental results partly conflict with the high frequency of β-O-4

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structures observed in highly acylated natural lignins.

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In this study, the biomimetic oxidations of sinapyl alcohol and sinapyl acetate were performed with

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silver (I) oxide (Ag2O). The effect of the γ-acetyl group on the oxidative coupling of sinapyl alcohol was

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investigated in order to better understand the high frequency of β-O-4 structures in highly acylated

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lignins. Ag2O is often used in the biomimetic oxidation of phenols. The oxidative coupling of a

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syringyl-type phenol 7 by Ag2O was reported to produce the β-O-4-coupled dimeric product 9 in 75%

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yield (Figure 2).13 We expected that Ag2O may be more suitable than HRP/peroxide to investigate the

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formation of β-O-4 structures by the oxidative coupling of syringyl-type monolignols.

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MATERIALS AND METHODS General. All chemicals were purchased from Wako (Osaka, Japan) and used as received without any

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purification. 1H and

13

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MHz) or a Bruker AMX500 FT-NMR (500 MHz) spectrometer in CDCl3 or acetone-d6 with

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tetramethylsilane (TMS) as an internal standard. Chemical shifts (δ) and coupling constants are reported

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in δ-values (ppm) and Hz, respectively. Assignments of erythro and threo isomers were made by the

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comparison of chemical sifts with the similar lignin model compounds. ESI-MS spectra were recorded

C NMR spectra were recorded with a Bruker AVANCE II 400 FT-NMR (400


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on an 1100 LC-MSD spectrometer (Agilent Technology, Santa Clara, California). High resolution ESI-

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ToF-MS was done with a Bruker micrOTOF. Biomimetic oxidations of sinapyl alcohol and sinapyl

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acetate were carried out at least twice during the isolation process of reaction products. The isolated

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yields of the reaction products were calculated on the basis of one set of reactions described below.

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Synthesis of Sinapyl Alcohol and Sinapyl Acetate. Sinapyl alcohol, 2 and sinapyl acetate, 4 were synthesized from 2,6-dimethoxyl phenol, 10, as shown in Figure 3.14

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2-(Allyloxy)-1,3-dimethoxybenzene, 11: To a stirred solution of 2, 6-dimethoxyphenol, 10 (5 g, 32.4

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mmol) in N,N-dimethylformamide (DMF; 50 mL) under argon, allyl bromide (98%, 3.5 mL, 38.9

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mmol), potassium carbonate (6.72 g, 48.7 mmol) and tetra-n-butyl ammonium iodide (1.2 g, 3.24 mmol)

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were added at room temperature. The reaction mixture was kept at 50 °C for 3.5 h. The reaction mixture

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was filtered, diluted with ethyl acetate, washed with brine, dried over Na2SO4 and concentrated to

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dryness in vacuo. The product was purified on a silica gel column with ethyl acetate/hexane (1/4, v/v) to

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afford a syrup, 11 (6.0 g, 95%). Compound 11: 1H NMR (400 MHz, CDCl3): δ 3.85 (6H, s, OCH3), 4.52

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(2H, dt, J = 6.1, 1.1 Hz, OCH2CH=CH2), 5.18 (1H, ddt, J = 10.3, 1.6, 1.1 Hz, OCH2CH=CH2), 5.30

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(1H, dq, J = 17.2, 1.6 Hz, OCH2CH=CH2), 6.12 (1H, ddt, 17.2, 10.3, 6.1, -CH2CH=CH2), 6.58 (2H, d, J

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= 8.4 Hz, C2, 6-H), 6.99 (1H, t, J = 8.4 Hz, C1-H); 13C NMR (400 MHz, CDCl3): δ 56.1 (OCH3), 74.1

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(OCH2CH=CH2), 105.4 (C2, 6), 117.6 (OCH2CH=CH2), 123.6 (C1), 134.6 (OCH2CH=CH2), 136.9

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(C4), 153.8 (C3, 5); ESI-MS (m/z): 217 ([M+Na]+, 100); High resolution ESI-ToF-MS: calculated for

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C11H14NaO3+: 217.0835, found: 217.0839.

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4-Allyl-2,6-dimethoxyphenol, 12: Compound 11 (8 g, 41.2 mmol) was placed in a stainless steel

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autoclave and heated at 200 °C under stirring for 6 h. The product was purified on a silica gel column

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with ethyl acetate/hexane (1/4, v/v) to afford a syrup, 12 (8.0 g, 97%). Compound 12: 1H NMR (400

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MHz, CDCl3): δ 3.31 (2H, br d, J = 6.7 Hz, α), 3.86 (6H, s, OCH3), 5.04-5.12 (2H, m, γ), 5.95 (1H, ddt,

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J =17.0, 10.0, 6.7 Hz, β), 6.41 (2H, s, C2, 6-H); 13C NMR (400 MHz, CDCl3): δ 40.4 (α), 56.3 (OCH3),

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105.2 (C2, 6), 115.8 (γ), 131.1(C1), 133.0(C4), 137.7(β), 147.0 (C3, 5); ESI-MS (m/z): 217 ([M +


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Na]+); High resolution ESI-ToF-MS: calculated for C11H14O3Na+: 217.0835, found: 217.0843.

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(E)-3-(4-hydroxy-3,5-dimethoxyphenyl)allyl acetate, 4: To a stirred solution of AgNO3 (2.1 g, 12.4

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mmol) in H2O (12.4 mL), aqueous NaOH (0.54 g, 13.5 mmol) solution in H2O (4.9 mL) was added

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dropwise at room temperature. The reaction mixture was kept for 5 min; it was then filtered and dried

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under vacuum to afford Ag2O. To a solution of compound 12 (1.0g, 5.15 mmol) in benzene (20 mL), all

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of the freshly prepared Ag2O was added at room temperature. The reaction mixture was kept for 30 min

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with vigorous stirring. The reaction mixture in benzene was filtered through Celite into a flask

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containing acetic acid (25 mL) and sodium acetate (2 g) at room temperature. The reaction mixture was

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kept for 30 min. The reaction mixture was filtered and concentrated to dryness in vacuo. The product

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was purified on a silica gel column with ethyl acetate/hexane (1/1, v/v) to afford a syrup (0.9 g, 69%).

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1

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γ), 6.16 (1H, dt, J = 15.8, 6.6 Hz, β), 6.57 (1H, br d, J = 15.8 Hz, α), 6.64 (2H, s, C2,6-H); 13C NMR

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(400 MHz, CDCl3): δ 21.2 (COCH3), 56.4 (OCH3), 65.4 (γ), 103.6 (C2, 6), 121.3 (β), 127.9 (C1), 134.8

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(α), 135.2 (C4), 147.3 (C3, 5), 171.1 COCH3); High resolution ESI-TOF-MS: calculated for

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C13H16O5Na+: 275.0890, found: 275.0899.

H NMR (400 MHz, CDCl3): 2.10 (3H, s, COCH3), 3.91 (6H, s, OCH3), 4.71 (2H, dd, J = 6.6, 1.1 Hz,

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Sinapyl alcohol, 2: To a stirred suspension of lithium aluminum hydride (61 mg, 1.61 mmol) in dry

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THF (10 mL) at −10 °C under argon, a solution of compound 4 (200 mg, 0.79 mmol) in dry THF (1.5

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mL) was added dropwise. The solution was kept at the same temperature for 1.5 h. To the reaction

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mixture, ethyl acetate (0.5 mL) was added to stop the reaction, and wet THF (0.5 mL), cold water (4

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mL) and CH2Cl2 (3 mL) were then added. The water phase was neutralized with a 10% KHSO4 solution

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and extracted with CH2Cl2. The organic layer was dried with Na2SO4 and concentrated to 3 mL under

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vacuum. Hexane was added to the solution, and sinapyl alcohol, 2 was crystallized in the cold to afford

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light yellow crystals (76 mg, 46%). 1H NMR (400 MHz, CDCl3): δ 3.90 (6H, s, OCH3), 4.31 (2H, br s,

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γ), 6.24 (1H, dt, J = 15.8, 5.8 Hz, β), 6.52 (1H, br d, J = 15.8 Hz, α), 6.63 (2H, s, C2, 6-H); 13C NMR

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(400 MHz, CDCl3): δ 56.3 (OCH3), 63.8 (γ), 103.3 (C2, 6), 126.6 (β), 128.2 (C1), 131.5 (α), 134.8 (C4),


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147.1 (C3, 5).

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Oxidative Coupling of Sinapyl Alcohol. To a stirred solution of compound 2 (100 mg, 0.476 mmol)

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in 1,4-dioxane (3 mL) under argon, freshly prepared Ag2O (111 mg, 0.479 mmol) was added. The

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reaction mixture was kept for 1 h under vigorous stirring in the dark. The reaction mixture in 1,4-

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dioxane was filtered through Celite into a flask containing conc. HCl (0.35 mL), H2O (20 mL) and THF

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(10 mL) at room temperature. The reaction mixture was kept under stirring for 1.5 h and then

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neutralized by adding NaHCO3. The reaction mixture was diluted with ethyl acetate, washed with brine,

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dried over Na2SO4 and concentrated to dryness in vacuo. The product was purified on a silica gel

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column with ethyl acetate/hexane (3/1, v/v) to afford five fractions. Sinapyl aldehyde, 13 (14 mg, 14%)

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was obtained from fraction No. 1. Both the erythro and threo isomers of syringyl glycerol-β-sinapyl

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alcohol, 14 (34 mg, 32%) and syringaresinol, 15 (< 3 mg, 3%) were obtained from fraction No. 5 by thin

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layer chromatography (TLC) with methanol/CH2Cl2 (1/19, v/v). Sinapyl aldehyde, 13: 1H NMR (500

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MHz, acetone-d6): δ 3.92 (6H, s, OCH3), 6.69 (2H, dd, J = 15.8, 7.7 Hz, β), 7.08 (2H, s, C2,6-H),

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7.55(1H, d, J = 15.8 Hz, α), 9.64 (1H, d, J = 7.7 Hz, γ). Syringyl glycerol-β-sinapyl alcohol, 14–the

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chemical sifts and coupling constants were determined for erythro and threo mixtures: 1H NMR (500

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MHz, acetone-d6): erythro: δ 3.41 (1H, γ), 3.81 (OCH3), 3.85 (1H, γ), 3.90 (OCH3), 4.18 (1H, m, β),

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4.24 (2H, br d, J = 5.2 Hz, γ′), 4.37 (1H, α-OH), 4.99 (1H, α), 6,39 (2H, dt, J = 15.9, 5.2 Hz, β′), 6.57

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(1H, br d, J = 15.9 Hz, α′), 6.72 (2H, s, C2,6-H or C2′,6′-H), 6.82 (2H, s, C2,6-H or C2′,6′-H); threo:

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3.33 (1H, γ), 3.67 (1H, γ), 3.80 (OCH3), 3.92 (OCH3), 3.95 (1H, m, β), 4.99 (1H, α), 6,39 (2H, dt, J =

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15.9, 5.2 Hz, β′), 6.56 (1H, br d, J = 15.9 Hz, α′), 6.77 (2H, s, C2,6-H or C2′,6′-H), 6.82 (2H, s, C2,6-H

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or C2′,6′-H). 13C-NMR (400 MHz, acetone-d6): erythro/threo: 56.7 (OCH3), 61.0(γ), 63.2 (γ′), 73.7(α),

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74.3(α), 88.2(β), 89.9(β), 104.6, 104.7, 104.9, 105.6, 129.9, 131.0, 132.8, 132.5, 134.8, 136.2, 148.5,

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154.1, 154.3; High resolution ESI-TOF-MS: m/z (M + Na)+: calculated for C22H28O9Na+: 459.1626,

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found: 459.1647. Syringresinol, 15: 1H NMR (500 MHz, acetone-d6): δ 3.10 (2H, m, β) 3.82-3.85(2H,

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m, γ), 3.84 (6H, s, OCH3), 4.23 (2H, m, γ), 4.67 (2H, d, J = 3.8 Hz, α), 6.69 (4H, s, C2,6-H), 13C-NMR ACS Paragon Plus Environment

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(500 MHz, acetone-d6): 55.3 (β), 56.6 (OCH3), 72.3 (γ), 86.8 (α), 104.4 (C2, 6), 133.2 (C1), 136.2 (C4),

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148.7 (C3, 5); High resolution ESI-TOF-MS: m/z (M + Na)+: calculated for C22H26O8Na+: 441.1520,

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found: 441.1565.

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Oxidative Coupling of Sinapyl Acetate. To a stirred solution of compound 4 (100 mg, 0.396

172

mmol) in 1,4-dioxane (3 mL) under argon, freshly prepared Ag2O (100 mg, 0.43 mmol) was added. The

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reaction mixture was kept under vigorous stirring for 20 min in the dark. The reaction mixture in 1,4-

174

dioxane was filtered through Celite into a flask containing concentrated HCl (0.35 mL), H2O (20 mL)

175

and THF (10 mL) at room temperature. The reaction mixture was kept for 30 min under stirring and was

176

then neutralized by adding NaHCO3. The reaction mixture was diluted with ethyl acetate, washed with

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brine, dried over Na2SO4 and concentrated to dryness in vacuo. The product was purified on a silica gel

178

column with ethyl acetate/hexane (2/1, v/v) to afford five fractions. Fraction No. 4 (18 mg, 18%) was

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mainly composed of β-O-4 oligomeric products, as was confirmed by the HSQC NMR spectra. The β-

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O-4-coupled dimers erythro-16 (50 mg, 48%) and threo-16 (19 mg, 18%) were obtained from fractions

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No. 2 and 3, respectively, by TLC with ethyl acetate/hexane/benzene (10/5/1, v/v/v).

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erythro-16: 1H NMR (500 MHz, acetone-d6): δ 1.87 (3H, s, COCH3), 2.04 (3H, s, COCH3), 3.81 (6H,

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s, OCH3), 3.91 (6H, s, OCH3), 4.09 (1H, dd, J = 11.8, 3,3 Hz, γ), 4.36 (1H, m, γ), 4.53 (1H, m, β), 4.70

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(2H, dd, J = 6.3, 1.0 Hz, γ′), 4.91 (1H, m, α), 6.36 (1H, dt, J = 15.9, 6.3 Hz, β′), 6.66 (1H, dt, J = 15.9

185

Hz, α′), 6.69 (2H, s, C2,6-H or C2′,6′-H), 6.86 (2H, s, C2,6-H or C2′,6′-H).

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acetone-d6): δ 20.8 (COCH3), 20.9 (COCH3), 56.7 (OCH3), 63.5 (γ), 65.4 (γ′), 73.1 (α), 84.5 (β), 104.6

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(C2, 6 or C2′, 6′), 104.9 (C2, 6 or C2′, 6′), 124.4 (β′), 131.6 (C1, C1′, C4 or C4′), 133.5 (C1, C1′, C4 or

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C4′), 134.4 (α′), 136.0(C1, C1′, C4 or C4′), 136.5 (C1, C1′, C4 or C4′), 148.6 (C3, 5 or C3′, 5′), 154.5

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(C3, 5 or C3′, 5′), 170.9 (COCH3); High resolution ESI-TOF-MS: m/z (M + Na)+: calculated for

190

C26H32O11Na+: 543.1837, found: 543.1811.

13

C-NMR (400 MHz,

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threo-16: 1H NMR (500 MHz, acetone-d6): δ 1.96 (3H, s, COCH3), 2.04 (3H, s, COCH3), 3,79 (6H,

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s, OCH3), 3.89 (6H, s, OCH3), 3.93 (1H, dd, J = 11.9, 4.2 Hz, γ), 4.22 (1H, m, β), 4.33 (1H, dd, J = 11.9, ACS Paragon Plus Environment

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3.3 Hz, γ), 4.69 (2H, dd, J = 6.2, 1.0 Hz, γ′), 4.91 (1H, m, α), 6.35 (1H, dt, J = 15.9, 6.2 Hz, β′), 6.65

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(1H, d, J = 15.9 Hz, α′), 6.73 (2H, s, C2,6-H or C2′,6′-H), 6.85 (2H, s, C2,6-H or C2′,6′-H).

195

(400 MHz, acetone-d6): δ 20.8 (COCH3), 20.9 (COCH3), 56.6 (OCH3), 56.7 (OCH3), 64.8 (γ), 65.3 (γ′),

196

74.7 (α), 86.7 (β), 104.9 (C2, 6 or C2′, 6′), 105.4 (C2, 6 or C2′, 6′), 124.4 (β′), 132.1 (C1, C1′, C4 or

197

C4′), 133.4 (C1, C1′, C4 or C4′), 134.3 (α′), 136.4 (C1, C1′, C4 or C4′), 137.8 (C1, C1′, C4 or C4′),

198

148.5 (C3, 5 or C3′, 5′), 154.0 (C3, 5 or C3′, 5′), 170.8 (COCH3), 170.9 (COCH3); High resolution ESI-

199

TOF-MS: m/z (M + Na)+: calculated for C26H32O11Na+: 543.1837, found: 543.1820.

200

Computational

Calculations

of

Sinapyl

Acetate

and

Sinapyl

Alcohol

13

C NMR

Radicals.

201

Computational calculations were conducted with Spartan ’08 Win (Wavefunction, Inc., USA).

202

Conformational analyses of the radicals were performed with the standard conformer search program in

203

the Spartan ’08 using PM3 semi-empirical calculations. The conformers obtained were further

204

optimized at the UB3LYP/6-31G* level of theory to identify the lowest energy conformers. Single point

205

energy calculations of the conformers were performed at the same level to obtain electrostatic charges

206

and spin densities based on natural population analysis. Solvent effects were added to the energy

207

calculations using the POSTSOLVENT command based on the SM8 model in Spartan ’08.

208 209

RESULTS AND DISCUSION

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Synthesis of Sinapyl Acetate and Sinapyl Alcohol. Sinapyl acetate, 4 and sinapyl alcohol, 2 were

211

synthesized from 2,6-dimethoxy phenol, 10 by the reported procedure (Figure 3).14 The allylation of

212

compound 10 was performed with allyl bromide and potassium carbonate in dimethyl formaldehyde

213

(DMF) to afford compound 11 in 99% yield. The Claisen rearrangement of compound 11 in a sealed

214

tube under argon at 200 °C gave compound 12 in 99% yield.15 Compound 12 was treated with freshly

215

prepared Ag2O in benzene, and the suspension was filtered through Celite to give a vinyl quinone

216

methide intermediate. The benzene solution of the formed vinyl quinone methide was poured into acetic

217

acid/sodium acetate to give sinapyl acetate, 4 in 69% yield.14 The acetyl group in compound 4 was

218

removed by lithium aluminum hydride to give sinapyl alcohol, 2 in 46% yield. ACS Paragon Plus Environment

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Oxidative Coupling of Sinapyl Alcohol and Sinapyl Acetate by Silver Oxide. The oxidative

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coupling of lignin-related monomers by Ag2O was first carried out by Zanarotti.13 The oxidation of a

221

syringyl-type monomer, 2,6-dimethoxy-4-(prop-1-en-1-yl)phenol, 7, in dry benzene or in dry methylene

222

chloride quantitatively yielded quinone methide 8, which was further hydrated with aqueous acid to

223

produce the β-O-4-coupled dimeric product 9 (C-O-coupled dimer) in 75% yield (Figure 2).13 Later, this

224

procedure was applied to the oxidative coupling of a guaiacyl-type monomer, coniferyl alcohol.16 The

225

reaction products from coniferyl alcohol were different than those from the syringyl-type monomer. In

226

methylene chloride, the β-5-coupled dimer was formed as a major product in 50% yield, whereas in a

227

two phase methylene chloride/water (1:1, v/v) solution, the β-β-coupled dimer was the major product in

228

25% yield.16 Both the β-5 and β-β dimers were C-C-coupled products.

229

On the other hand, the oxidative coupling of sinapyl alcohol, 2, a typical syringyl-type monomer, by

230

Ag2O has not been reported. Alternatively, FeCl3 has been used for the oxidative coupling of sinapyl

231

alcohol, 2 in 1,4-dioxane, which gave the β-O-4-coupled dimer 14 in 28% isolated yield.17 In this

232

investigation, sinapyl alcohol, 2 was treated with Ag2O in 1,4-dioxane, and the resultant quinone

233

methide was further hydrated with aqueous hydrochloric acid (HCl)/tetrahydrofuran (THF) solution.18

234

The Ag2O oxidation of sinapyl alcohol, 2 produced sinapyl aldehyde, 13 in 14% yield, the β-O-4-

235

coupled dimer 14 in 32% yield, the β-β-coupled dimer (syringaresinol, 15) in 3% yield and other

236

unidentified oligomeric products (Figure 4). The yield of the β-O-4-coupled dimer from the Ag2O

237

oxidation of sinapyl alcohol was a little higher than the yield reported for FeCl3 oxidation. The oxidative

238

coupling of phenols by Ag2O has been reported to preferentially form C-O-coupled products, whereas

239

FeCl3 leads to C-C-coupled products.19

240

We examined the Ag2O-mediated oxidative coupling of the γ-acetylated monomer sinapyl acetate, 4 to

241

elucidate the effect of the γ-acetyl group on the oxidative coupling of sinapyl alcohol. The Ag2O

242

oxidation of sinapyl acetate, 4 in benzene preferentially formed erythro and threo isomers of the β-O-4-

243

coupled dimer, 16 in 39% yield. In 1,4-dioxane, the selectivity for β-O-4-coupled products became even ACS Paragon Plus Environment

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244

higher. The yield of the erythro and threo isomers of the β-O-4-coupled dimer was 66% (Figure 4).

245

Oligomeric products with predominantly β-O-4 structures (Figure 5) were also obtained in 18% yield.

246

However, non-traditional tetrahydrofuran-type β-β-coupled products10 were not detected by the NMR

247

analysis. These results showed that the Ag2O-mediated oxidative coupling of sinapyl acetate

248

preferentially formed β-O-4 linkages. The yield of the β-O-4-coupled products from sinapyl acetate was

249

much higher than that from sinapyl alcohol. The oxidation of the hydroxyl group at the γ-position was

250

completely prevented by the acetyl group in sinapyl acetate, which partially contributed to the high yield

251

of β-O-4 products from sinapyl acetate.

252

Computational calculations of atomic spin densities and charges of sinapyl radicals. In order to

253

further understand the effect of the acetyl group at the γ-position in sinapyl acetate, computational

254

calculations based on density functional theory (DFT) were performed. The atomic spin densities of

255

phenoxy radicals derived from sinapyl acetate and sinapyl alcohol are shown in Table 1. The spin

256

densities show that the unpaired electrons are mainly located at C1, C3, C5, Cβ and O4 in both sinapyl

257

acetate and sinapyl alcohol radicals. The spin densities of these radicals are the highest at O4, which is

258

agreement with the reported data for sinapyl alcohol radical.20 The C1, C3 and C5 sites are not

259

considered to be important coupling sites in sinapyl alcohol radicals. Thus, the most reactive coupling

260

sites are Cβ and O4 in both sinapyl acetate and sinapyl alcohol radicals. The radical coupling at Cβ and

261

Cβ produces β-β linkages, while coupling at Cβ and O4 produces β-O-4 linkages. The spin densities of

262

carbon and oxygen atoms in sinapyl acetate and sinapyl alcohol radicals are quite similar to each other.

263

The acetyl group at the γ-position did not significantly affect the localization of the unpaired electron in

264

the sinapyl alcohol radical. These results indicate that the modification of the localization of the

265

unpaired electron cannot explain the preferential formation of β-O-4-coupled products by the oxidative

266

coupling of sinapyl acetate.

267

The charge density at each atom in monolignol radicals is also important for the coupling reaction of

268

the radicals.20 The electrostatic charges of all carbon and oxygen atoms in sinapyl acetate and sinapyl ACS Paragon Plus Environment

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269

alcohol radicals are shown in Table 2. The solvent effects of water and 1,4-dioxane were also added to

270

the calculation of the electrostatic charges. These data show that Cβ and O4 were negatively charged in

271

the sinapyl alcohol and sinapyl acetate radicals. As the two radicals approach each other, their negative

272

charges result in Coulombic repulsion between the Cβ and O4 positions.20 The data show that the

273

negative charge at the Oγ position in the sinapyl acetate radical was much smaller than that in the

274

sinapyl alcohol radical. This result can be reasonably explained by the fact that acyl groups, such as the

275

acetyl group, are electron withdrawing groups. It is interesting to note that the electron withdrawing

276

effects of the acetyl group are active even at the Cβ position in the sinapyl acetate radical. The negative

277

charge at Cβ in the sinapyl acetate radical was significantly smaller than that in the sinapyl alcohol

278

radical. This tendency was more obvious when solvent effects were considered in the calculations. The

279

differences in the electrostatic charges at Cβ between the sinapyl acetate and sinapyl alcohol radicals

280

were 0.033 e in 1,4-dioxane and 0.052 e in water. These results suggest that the Coulombic repulsion

281

between the Cβ and O4 positions was significantly reduced by the γ-acetyl group (Figure 6), which

282

resulted in the high yield of β-O-4-coupled products from the oxidative coupling of sinapyl acetate.

283

In conclusion, compared to the biomimetic oxidative coupling of sinapyl alcohol, the biomimetic

284

oxidative coupling of sinapyl acetate by silver oxide preferentially formed β-O-4-coupled products.

285

Computational calculations showed that the γ-acetyl group significantly decreased the negative charge at

286

Cβ in the sinapyl acetate radical. The Coulombic repulsion between Cβ and O4 seems to be reduced

287

when two sinapyl acetate radicals approach each other. These results suggest that acylated monolignols

288

preferentially couple at Cβ and O4, resulting in the high frequency of β-O-4 structures in highly acylated

289

lignins.

290 291 292 293

ABBREVIATIONS USED DFT, density functional theory; HRP, horseradish peroxidase; HSQC, heteronuclear single quantum coherence. ACS Paragon Plus Environment

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294 295

AUTHOR INFORMATION

296

Corresponding Author

297

*Phone: +81-766-56-7500 (ext. 567). Fax: +81-766-56-2498. E-mail: [email protected]

298

Funding

299

A part of this work was supported by JSPS KAKENHI Grant Number 24658156.

300 301

ACKNOWLEDGMENTS

302

We are grateful to Ms. Yuka Nonaka and Ms. Junko Kawabata, graduates of the Department of

303

Biotechnology, Toyama Prefectural University, for their contributions to the preliminary experiments of

304

this investigation.

305 306

REFERENCES

307

(1) Sjöström, E, Lignin. In Wood Chemistry: Fundamentals and applications; Academic Press: San

308

Diego, 1993, pp. 71-89.

309

(2) Kishimoto, T.; Uraki, Y.; Ubukata, M. Easy synthesis of β-O-4 type lignin related polymers. Org.

310

Biomol. Chem., 2005, 3, 1067-1073.

311

(3) Kishimoto, T.; Uraki, Y.; Ubukata, M. Chemical synthesis of β-O-4 type artificial lignin. Org.

312

Biomol. Chem., 2006, 4, 1343-1347.

313

(4) Kishimoto, T.; Uraki, Y.; Ubukata, M. Synthesis of β-O-4-type artificial lignin polymers and their

314

analysis by NMR spectroscopy. Org. Biomol. Chem., 2008, 6, 2982-2987.

315

(5) Ralph, J.; Landucci, L. L., NMR of lignins. In Lignin and Lignans: Advances in Chemistry; Heitner,

316

C., Dimmel, D. R., Schmidt, J. A., Eds.; CRC Press: New York, 2010, pp. 137-244.

317

(6) Ralph, J., An unusual lignin from kenaf. J. Nat. Prod., 1996, 59, 341-342. ACS Paragon Plus Environment

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(7) del Río, J.; Rencoret, J.; Marques, G.; Gutiérrez, A.; Ibarra, D.; Santos, J.; Jiménez-Barbero, J.;

319

Zhang, L.; Martínez, Á. Highly acylated (acetylated and/or p-coumaroylated) native lignins from diverse

320

herbaceous plants. J. Agric. Food Chem., 2008, 56, 9525-9534.

321

(8) Villaverde, J. J.; Li, J.; Ek, M.; Ligero, P.; de Vega, A. Native lignin structure of Miscanthus x

322

giganteus and its changes during acetic and formic acid fractionation, J. Agric. Food Chem., 2009, 57,

323

6262-6270.

324

(9) Kishimoto, T.; Chiba, W.; Saito, K.; Fukushima, K.; Uraki, Y.; Ubukata, M. Influence of syringyl

325

to guaiacyl ratio on the structure of natural and synthetic lignins. J. Agric. Food Chem., 2010, 58, 895-

326

901.

327 328

(10) Lu, F.; Ralph, J., Novel tetrahydrofuran structures derived from β–β-coupling reactions involving sinapyl acetate in Kenaf lignins. Org. Biomol. Chem., 2008, 6, 3681-3694.

329

(11) Lu, F.; Ralph, J.; Morreel, K.; Messens, E.; Boerjan, W. Preparation and relevance of a cross-

330

coupling product between sinapyl alcohol and sinapyl p-hydroxybenzoate. Org. Biomol. Chem., 2004, 2,

331

2888-2890.

332

(12) Tanahashi, M.; Takeuchi, H.; Higuchi, T., Dehydrogenative polymerization of 3,5-disubstituted

333

p-Coumaryl Alcohols. Wood Res., 1976, 61, 44-53.

334

(13) Zanarotti, A. Synthesis and reactivity of lignin model quinone methide. Biomimetic synthesis of

335

8.O.4′ neolignan. J. Chem. Res., Synop., 1983, 306-307.

336

(14) Zanarotti, A. Preparation and reactivity of 2,6-dimethoxy-4-allyliden-2,5-cyclohexadien-1-one

337

(vinyl quinone methide). A novel synthesis of sinapyl alcohol. Tetrahedron Letters, 1982, 23, 3815-

338

3818.

339 340

(15) Jing, X.; Gu, W.; Bie, P.; Ren, X.; Pan, X. Total synthesis of (±)-eusiderin K and (±)-eusiderin J. Synth. Commun., 2001, 31, 861-867. ACS Paragon Plus Environment

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341 342


(16) Quideau, S.; Ralph, J. A biomimetic route to lignin model compound via silver (I) oxide oxidation. 1. Synthesis of dilignols and non-cyclic benzyl aryl ethers. Holzforschung, 1994, 48, 12-22.

343

(17) Alam, S.; Suzuki, T.; Katayama, T. Formation of optically active erythro-syringylglycerol-8-O-

344

4′-(sinapyl alcohol) ethers from achiral monolignols with enzyme preparations of higher plant. Middle-

345

East J. Sci. Res., 2010, 6, 340-345.

346 347

(18) Hawkes, G. E.; Smith, C. Z.; Utley, H. P.; Chum, H. Key structural feature of acetone-soluble phenol-pulping lignins by 1H and 13C NMR spectroscopy. Holzforschung, 1986, 40 (Suppl.), 115-123.

348

(19) Cotelle, P.; Vezin, H. The reaction of methyl isoferulate with FeCl3 or Ag2O–hypothesis on the

349

biosynthesis of lithospermic acids and related nor and neolignans. Tetrahedron Lett., 2003, 44, 3289-

350

3292.

351 352

(20) Shigematsu, M.; Masamoto, H. Solvent effects on the electronic state of monolignol radicals as predicted by molecular orbital calculations. J. Wood Sci., 2008, 54, 308-301.

353 354 355 356 357 358 359 360 361 362 363 364 ACS Paragon Plus Environment

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365 366

FIGURE CAPTIONS

367 368

Figure 1. Traditional monolignols 1-3, and non-traditional monolignols 4-6.

369

Figure 2. Ag2O-mediated oxidative coupling of a syringyl-type monomer by Zanarotti.13

370

Figure 3. Synthesis of sinapyl acetate and sinapyl alcohol. Reaction conditions: (a) allyl bromide,

371

K2CO3, n-Bu4NI, DMF; (b) neat, under Ar, 200 °C; (c) Ag2O, benzene, and then acetic acid/sodium

372

acetate; (d) LiAlH4, THF, −10 °C.

373

Figure 4. Ag2O-mediated oxidative coupling of sinapyl alcohol and sinapyl acetate.

374

Figure 5. HSQC NMR spectrum of oligomeric products formed form the Ag2O-mediated oxidative

375

coupling of sinapyl acetate.

376

Figure 6. Reduced negative charges by acetyl group result in the high yield of β-O-4 products.

377 378 379 380 381 382 383 384 385 386 387 388 389 ACS Paragon Plus Environment

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Table 1. Calculated spin densities of sinapyl alcohol and sinapyl acetate radicals.

Sinapyl alcohol

Sinapyl acetate

radical

radical

C1

0.2216

0.2216

C2

−0.1007

−0.1028

C3

0.1802

0.1824

C4

0.0337

0.0310

C5

0.1532

0.1549

C6

−0.0933

−0.0957

Cα

−0.0841

−0.0845

Cβ

0.1797

0.1767

Cγ

−0.0078

−0.0080

O3

0.0750

0.0781

O4

0.3597

0.3611

O5

0.0607

0.0634

Oγ

0.0003

0.0001


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Table 2. Calculated electrostatic charges of carbon and oxygen atoms in sinapyl alcohol and sinapyl acetate radicals Sinapyl alcohol radical

Sinapyl acetate radical

Solvent

None

1,4-dioxane

Water

None

1,4-dioxane

Water

C1

0.3958

0.3983

0.4162

0.3714

0.3839

0.4017

C2

−0.5513

−0.5480

−0.5579

−0.5422

−0.5479

−0.5550

C3

0.3014

0.2962

0.2941

0.3028

0.3102

0.3040

C4

0.2888

0.2880

0.2802

0.2851

0.2775

0.2688

C5

0.2811

0.2881

0.2876

0.2930

0.3062

0.3091

C6

−0.5268

−0.5376

−0.5503

−0.5313

−0.5461

−0.5599

Cα

−0.2147

−0.2133

−0.2294

−0.1869

−0.1974

−0.2184

Cβ

−0.1883

−0.1953

−0.1656

−0.1714

−0.1619

−0.1194

Cγ

0.1344

0.1579

0.1521

0.0289

0.0067

−0.0271

O3

−0.1972

−0.1913

−0.2022

−0.1896

−0.1908

−0.2024

O4

−0.4326

−0.4313

−0.4664

−0.4272

−0.4244

−0.4605

O5

−0.2160

−0.2162

−0.2279

−0.2128

−0.2133

−0.2256

Oγ

−0.5779

−0.5824

−0.6039

−0.3887

−0.3847

−0.3834


18

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HO HO

HO

HO

O

HO

O

O

OMe

MeO

O

MeO

OMe

O

OMe

O

MeO

OMe

OH

OH

OH

OH

OH

1

2

3

4

5

MeO

OMe OH

6

Figure 1


19


Page 20 of 25

Figure 2


20

Page 21 of 25


Figure 3


21


Page 22 of 25

OMe OH HO

O

H

OH

HO MeO

O OMe

HO

O

1) Ag2O

OMe

MeO O

MeO

OMe

2) aq/HCl MeO

OH

OMe OH

2

MeO

14%

13

OMe OH

HO

32%

14

OMe

15

3%

OAc AcO

AcO MeO HO

1) Ag2O

O OMe

2) aq/HCl MeO

OMe

MeO

OMe

OH

OH

4

16

66%

Figure 4


22

Page 23 of 25


OCH3

β-O-4(γ) Sinapyl acetate (γ)

β-O-4(α)

β-O-4(β)

Figure 5


23


Page 24 of 25

Figure 6


24

Page 25 of 25


Graphical Abstract


25

Biomimetic Oxidative Coupling of Sinapyl Acetate by Silver Oxide

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