Method for isolating, from plant biomass, at least one selected from group consisting of lignin, hemicellulose, lignin-polysaccharide composite, cellulose, and hemicellulose-cellulose composite

Abstract

The present invention addresses the problem of providing a method for isolating, from a plant biomass, at least one selected from the group consisting of lignin, hemicellulose, a lignin-polysaccharide composite, cellulose, and a hemicellulose-cellulose composite. The problem is solved by a method for isolating, from a plant biomass, at least one selected from the group consisting of lignin, hemicellulose, a lignin-polysaccharide composite, cellulose, and a hemicellulose-cellulose composite, the method comprising a step (A) for bringing a solution containing an organic acid and a peroxide into contact with the plant biomass.

Description

A method for isolating at least one member selected from the group consisting of lignin, hemicellulose, a lignin-polysaccharide complex, cellulose, and a hemicellulose-cellulose complex from plant biomass

[0001] The present invention relates to a method for isolating at least one selected from the group consisting of lignin, hemicellulose, a lignin-polysaccharide complex, cellulose, and a hemicellulose-cellulose complex from plant biomass.

[0002] Lignin, along with polysaccharides (cellulose and hemicellulose), is a major component of plant cell walls. Lignin is an aromatic polymer that is abundant in nature and has attracted attention. However, currently, it is mainly obtained as a by-product of the pulp and paper manufacturing process or the bioethanol production process. However, because the isolated lignin is denatured, it is difficult to enhance its functionality.

[0003] For this reason, various methods have been proposed for isolating lignin while avoiding denaturation as much as possible (Patent Document 1).

[0004] JP 2013-241391 A

[0005] An objective of the present invention is to provide a method for isolating at least one selected from the group consisting of lignin, hemicellulose, lignin-polysaccharide complexes, cellulose, and hemicellulose-cellulose complexes from plant biomass.

[0006] The present inventors have conducted extensive research to solve the above-mentioned problems and have come up with a new method for isolating lignin, which involves extracting lignin in the reverse order of wood biosynthesis, and have found that this method can solve the above-mentioned problems. The present invention was completed through further research based on this finding and includes the following aspects.

[0007] Item 1. A method for isolating at least one species selected from the group consisting of lignin, hemicellulose, lignin-polysaccharide complexes, cellulose, and hemicellulose-cellulose complexes from plant biomass, comprising: (A) a step of contacting plant biomass with a solution containing an organic acid and a peracid.

[0008] Item 2. The method according to Item 1, wherein step (A) comprises: (A1) contacting the plant biomass with a solution containing an organic acid and a peracid at 90°C or less, and / or (A2) contacting the plant biomass with a solution containing an organic acid and a peracid at greater than 90°C and not greater than 160°C.

[0009] Item 3. The method according to Item 1 or 2, comprising obtaining at least one selected from the group consisting of lignin, hemicellulose, a lignin-polysaccharide complex, and a hemicellulose-cellulose complex from the soluble portion obtained in step (A).

[0010] Item 4. The method according to any one of Items 1 to 3, comprising obtaining at least one selected from the group consisting of cellulose, hemicellulose, and a hemicellulose-cellulose complex from the insoluble fraction obtained in step (A).

[0011] Item 5. The method according to any one of Items 1 to 4, wherein the step (A) comprises irradiating the plant biomass with microwaves or ultrasound.

[0012] Item 6. The method according to any one of Items 1 to 5, wherein the plant biomass used in step (A) is an alkali-treated plant body or a mechanically processed product thereof.

[0013] Item 7. Lignin having a β-O-4 ether structure content of 50% or more and a solubility of 2 w / v% or more in 60-80% ethanol at 25°C.

[0014] Item 8. Lignin obtainable by the method according to any one of Items 1 to 6.

[0015] Item 9. A lignin-polysaccharide complex having at least one bond selected from the group consisting of an α-ether bond between lignin and a polysaccharide, an α-ester bond between lignin and a polysaccharide, and a γ-ester bond between lignin and a polysaccharide.

[0016] Item 10. The lignin-polysaccharide complex according to Item 9, having a solubility of 2 w / v % or more in 60 to 80% ethanol at 25°C.

[0017] Item 11. A lignin-polysaccharide complex obtainable by the method according to any one of Items 1 to 6.

[0018] Item 12. Cellulose obtainable by the method according to any one of Items 1 to 6.

[0019] Item 13. A hemicellulose-cellulose composite obtainable by the method according to any one of Items 1 to 6.

[0020] Item 14. Hemicellulose obtainable by the method according to any one of Items 1 to 6.

[0021] Item 15. At least one self-assembled body selected from the group consisting of the lignin according to Item 7 or 8 and the lignin-polysaccharide complex according to any one of Items 9 to 11.

[0022] Item 16. The self-assembled body according to Item 15, which is in the form of a sheet or particles.

[0023] Item 17. A phosphor comprising the self-assembled body according to Item 15 or 16.

[0024] Item 18. The phosphor according to Item 17, which exhibits anti-Stokes fluorescence.

[0025] Item 19. An ultraviolet absorber comprising at least one selected from the group consisting of the lignin according to Item 7 or 8 and the lignin-polysaccharide complex according to any one of Items 9 to 11.

[0026] The present invention can provide a method for isolating at least one selected from the group consisting of lignin, hemicellulose, lignin-polysaccharide complexes, cellulose, and hemicellulose-cellulose complexes from plant biomass.

[0027]

[0023] Figure 1 shows a flowchart of the method of the present invention (M-APA method). Figure 2 shows a flowchart of the method of the present invention (A-APA method). Figure 3 shows the microwave irradiation conditions, power, temperature, and pressure profile for lignocellulose component separation, and the specifications of the Biotage Initiator+ 60 microwave synthesizer, with an agitation speed of 600 rpm, cooling on, and absorption high. Figure 4 shows a 2D NMR spectrum of a low-denatured, high-purity, ether-rich lignin obtained by the present invention. Figure 5 shows the analysis results of lignin isolated by the M-APA method (microwave treatment at 50°C for 10 minutes, acetic acid and peracetic acid 1%) using eucalyptus as the starting material. Figure 6 shows the partial chemical structure of lignin interunit bonds and lignin-hemicellulose covalent bonds. Figure 7 shows a 2D NMR spectrum (a-d) of a lignin-hemicellulose copolymer, and enlarged views (a'-d') of the interunit bonds and sugar regions. Lignin-hemicellulose copolymers obtained by the M-APA method from hardwoods (a, b) and softwoods (c, d) were analyzed. The circles in the figure indicate LC bonds (ether and ester types). Synchrofluorescence spectra in the near-infrared wavelength region of APA lignin derived from bagasse (sugarcane). Synchro-scan measurements were performed to detect a constant Stokes shift, i.e., the fluorescence wavelengths Em, which are 80, 90, 150, 160, and 170 nm longer than the excitation wavelength Ex, were detected (scanning the excitation wavelength). The 3D fluorescence spectrum in the near-infrared wavelength region of APA lignin from Japanese red pine (Pinus densiflora). APA lignin obtained by treating Japanese red pine wood flour with the M-APA method (microwave at 50°C for 10 min using acetic acid and 0.2 M hydrogen peroxide), was measured in an 80% acetonitrile solution (concentration: 0.1 wt%). This graph shows a comparison of synchroscan fluorescence spectra of reagent lignin (Tokyo Kasei TCI alkaline lignin, blue line) and lignin (APA lignin, red line). Fluorescence longer than the excitation wavelength (plus 90 nm) was detected (scanning the excitation wavelength). Lignin (APA lignin) exhibits characteristic fluorescence in the long wavelength region. The synchroantifluorescence spectra of a lignin solid thin film and a lignin-polysaccharide copolymer solid thin film are shown.The excitation wavelength was scanned to selectively detect anti-Stokes-shifted fluorescence shorter than the excitation wavelength. Anti-Stokes shift spectra (a-f) and control (g) are shown. The anti-Stokes shift is 50 nm below the excitation wavelength, and 40 nm for spectrum (b). Comparisons between spectra (b) and (d) and between spectra (e) and (f) reveal that anti-Stokes intensity is higher when peracetic acid is used as the oxidizing agent than hydrogen peroxide. Comparisons between spectra (b) and (c) reveal that the anti-fluorescence intensity of APA lignin is higher than that of lignin polysaccharide copolymers. The results of simultaneous differential thermal and thermogravimetry measurements of APA lignin and lignin-hemicellulose copolymers are shown. The results of simultaneous differential thermal and thermogravimetry measurements of cellulose and raw eucalyptus wood flour are shown. The UV absorption spectra of lignin and lignin-hemicellulose copolymers are shown. The vertical axis represents absorbance, and the horizontal axis represents wavelength (nm). For the legend, see Table 8. The results of solution NMR measurements of hemicellulose obtained in Test Example 7-1 are shown. The results of solid-state CP / MAS NMR measurements are shown. Spectrum a is from cellulose fiber (derived from eucalyptus wood flour) obtained in Test Example 7-1, and spectrum b is from white beech wood chips obtained in 7-2, which were decolorized with acetic acid, washed with water, and then freeze-dried. Spectrum c is from white beech wood chips obtained in 7-2, which were bleached in addition to the conditions in b, and then freeze-dried. Solid-state 13C CP / MAS spectra were measured by placing the powdered white powder in a 4 mm pencil-type sample tube (50 μL volume) using a solid-state NMR system (Varian, Inc., 400 MHz) under 15 kHz MAS conditions. Analysis was performed using a Varian VnmrJ4.2. In the solid-state 13C CP / MAS NMR spectrum, unbleached beech wood chips (Figure 13b) showed clear signals of the hemicellulose-cellulose complex, mainly carbonyl groups (172 ppm) derived from the acetyl groups and side-chain glucuronic acid of acetylated glucuronoxylan, and methyl groups (22 ppm) derived from the acetyl groups. This indicates that hemicellulose forms a strong complex structure with cellulose, while retaining a high degree of acetyl groups and side chains. The FT-IR measurement results for the white wood chips obtained in Test Example 7-2 are shown below. Items a to c are as described in Figure 13. The measurement conditions were as follows:PerkinElmer FT-IR Spectrum Two Diamond ATR, measuring white cellulose powder. 400-4000 cm. -1 , Scan number 10. This is an enlarged view of a portion of Figure 14. The circled area is a peak characteristic of hemicellulose. -1 ; acetyl group-derived C=O stretching vibration of acetylated glucuronoxylan and C=O stretching vibration of glucuronic acid, and 1243cm -1 The CO stretching vibrations of acetylated glucuronoxylan and glucuronic acid were clearly observed. These are photographs of the white wood chip obtained in Test Example 7-2 and the processed product thereof. In a, the left side shows the state of the acetic acid peroxide-treated wood chip after acetic acid decolorization, and the right side shows the wood chip before being subjected to the alkali treatment of Test Example 7. b shows the state of the wood chip on the left side of a after being decolorized, bleached, and washed with water. c shows the state of the wood chip on the left side of a after being freeze-dried.

[0028] In this specification, the expressions "contain" and "comprise" include the concepts of "contain," "comprise," "consist essentially of," and "consist only of."

[0029] 1. Isolation Method In one aspect, the present invention relates to a method for isolating at least one selected from the group consisting of lignin, hemicellulose, lignin-polysaccharide complexes, cellulose, and hemicellulose-cellulose complexes from plant biomass, the method comprising the step of (A) contacting plant biomass with a solution containing an organic acid and a peracid (also referred to herein as the "isolation method of the present invention"). This method is described below.

[0030] The isolation method of the present invention includes step (A), and therefore can produce high-quality (low-condensation) lignin with a high content of β-O-4 ether structures among all the bond types between monolignols.

[0031] The plant biomass is not particularly limited as long as it contains at least one selected from the group consisting of lignin, hemicellulose, lignin-polysaccharide complexes, cellulose, and hemicellulose-cellulose complexes, and examples thereof include the plant itself and mechanically processed products of the plant.

[0032] Examples of plant bodies include coniferous trees, hardwoods, and non-woody materials. Specific examples include cedar, pine, spruce, larch, black pine, Abies sachalinensis, Himekomatsu, yew, Japanese yew, spruce, Japanese spruce, Japanese spruce, Japanese yew, Japanese bead, fir, Japanese yew, Douglas fir, spruce, yellow cedar, Japanese cypress, Douglas fir, Sitka spruce, Radiata pine, eastern spruce, eastern white pine, western larch, western fir, and western Examples of suitable woods include conifers such as hemlock and tamarack; hardwoods such as asbestos, American black cherry, yellow poplar, walnut, birch, zelkova, sycamore, silver cherry, ash, teak, Chinese elm, Chinese maple, oak, beech, hard maple, hickory, pecan, white ash, white oak, white birch, red oak, acacia, and eucalyptus; and non-woody materials such as rice, sugarcane, wheat, corn, pineapple, oil palm, kenaf, cotton, alfalfa, timothy, bamboo, bamboo grass, and sugar beet.

[0033] Examples of mechanically processed products of plants include logs, timber, boards, solid wood, wood materials, laminated lumber, laminated veneer lumber, plywood, wood boards, particle boards, fiber boards, wood chips, particulate wood (chips, particles, wood flour, etc.), fibrous wood, compressed materials, crushed materials, etc.

[0034] From the viewpoint of improving the yield, the plant biomass is preferably wood flour. The volume average particle diameter of the wood flour is, for example, 500 μm or less, preferably 200 μm or less, more preferably 100 μm or less, even more preferably 50 μm or less, and even more preferably 20 μm or less. The lower limit of the average particle diameter is not particularly limited and can be, for example, 1 μm, 2 μm, or 5 μm. By using wood flour with a relatively small particle size, a high yield can be achieved without performing the alkali treatment described below.

[0035] Wood flour can be obtained by pulverizing plant biomass. The pulverization can be carried out according to or in accordance with a known method, for example, using various mills (e.g., ball mills, mixer mills, etc.).

[0036] From the viewpoint of improving yield, the plant biomass is preferably an alkali-treated product of a plant body or a mechanically processed product thereof. According to the present invention, by using the alkali-treated product, the target product can be efficiently obtained without pulverization (even in the case of a relatively large specific surface area, such as wood chips). The alkali-treated product can be obtained by alkali-treating a plant body or a mechanically processed product thereof. The alkali treatment loosens the plant cell wall structure (reducing intermolecular interactions and cleaving hydrogen bonds) and cleaves intramolecular ester bonds, thereby improving yield. This allows a high yield to be achieved without physical pulverization of the plant biomass, or even with only a small degree of pulverization, thereby reducing process costs. Furthermore, the yield can be improved by 1.2 to 2 times. Furthermore, the crystallinity of the cellulose fibers obtained through this alkali treatment is good.

[0037] Specifically, the alkali treatment is preferably, for example, (X) a step of contacting a plant body or a mechanically processed product thereof with an alkaline solution.

[0038] The alkaline solution is not particularly limited as long as it is an alkaline solution to the extent that the above-mentioned purpose can be achieved. For example, the alkaline solution may be a solution with a pH of 12 to 15. The pH of the solution is preferably 13 to 14. Furthermore, the alkaline solution may be, for example, a solution containing 0.3 to 2 mass % (preferably 0.7 to 1.5 mass %) of an alkali metal hydroxide (e.g., sodium hydroxide, potassium hydroxide, etc.).

[0039] The amount of alkaline solution used is not particularly limited as long as it is an amount sufficient to achieve the above-mentioned purpose. The amount of alkaline solution used may be, for example, 3 to 20 mL, preferably 5 to 12 mL, per 1 g of plant body or mechanically processed product thereof.

[0040] The manner in which the plant body or a mechanically processed product thereof is brought into contact with the alkaline solution is not particularly limited, but from the standpoint of treatment efficiency and the like, it is preferable to immerse the plant body or a mechanically processed product thereof in the alkaline solution.

[0041] The temperature of the alkaline solution is not particularly limited as long as it is a temperature that can achieve the above-mentioned object. The temperature of the alkaline solution can be, for example, 20 to 150°C, preferably 50 to 120°C, and more preferably 80 to 120°C. When heating, methods include internal heating using microwave heating and external heating using an oil bath or the like, with internal heating using microwave heating being preferred.

[0042] The treatment time in step (X) is not particularly limited as long as it is long enough to achieve the above-mentioned object, and is, for example, 5 to 120 minutes, preferably 15 to 60 minutes.

[0043] After step (X), the soluble portion and the insoluble portion are separated, and the insoluble portion can be subjected to step (A) as an alkali-treated product of the plant body or a mechanically processed product thereof.

[0044] The plant biomass may be one type alone or a combination of two or more types.

[0045] The solution containing an organic acid and a peracid (organic acid-peracid solution) is a solution that contains an organic acid and a peracid and is capable of extracting and solubilizing at least one selected from the group consisting of lignin, hemicellulose, and a lignin-polysaccharide complex, and is not particularly limited thereto.

[0046] The organic acid is not particularly limited, and examples thereof include acetic acid, formic acid, glyoxylic acid, maleic acid, propionic acid, folic acid, isobutyric acid, valeric acid, isovaleric acid, pyruvic acid, oxalic acid, malonic acid, succinic acid, glutaric acid, ketoglutaric acid, adipic acid, lactic acid, tartaric acid, fumaric acid, oxaloacetic acid, malic acid, isocitric acid, citric acid, benzoic acid, phthalic acid, isophthalic acid, terephthalic acid, hemimellitic acid, trimellitic acid, trimesic acid, mellophanic acid, prenitic acid, pyromellitic acid, mellitic acid, methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, camphorsulfonic acid, p-toluenesulfinic acid, and benzenesulfinic acid. Of these, preferred are acetic acid, formic acid, glyoxylic acid, 3-oxopropanoic acid, 2-methyl-3-oxopropanoic acid, etc., more preferred are acetic acid, formic acid, glyoxylic acid, etc., and particularly preferred is acetic acid.

[0047] The organic acid may be one kind alone or a combination of two or more kinds.

[0048] The peracid is not particularly limited as long as it is an acid having a hydroperoxide group (—O—OH), and examples thereof include organic peroxoacids such as percarboxylic acids, persulfuric acid, percarbonic acid, perphosphoric acid, and hypoperhalogen acids. Examples of percarboxylic acids include peracetic acid, performic acid, perbenzoic acid, and metachloroperbenzoic acid, and examples of hypoperhalogen acids include hypoperchloric acid, hypoperbromic acid, and hypoperiodic acid. In addition to the above, examples of peracids include hydrogen peroxide, lithium peroxide, sodium peroxide, potassium peroxide, sodium percarbonate, urea peroxide, sodium perborate, tert-butyl hydroperoxide, cumene hydroperoxide, di-tert-butyl peroxide, dimethyldioxirane, acetone peroxide, methyl ethyl ketone peroxide, and hexamethylene triperoxide diamine. Among these, preferred are organic peroxoacids, hydrogen peroxide, sodium percarbonate, urea peroxide, sodium perborate, etc., more preferred are percarboxylic acids (particularly peracetic acid), hydrogen peroxide, etc., and particularly preferred is hydroperoxy acid.

[0049] The peracid may be one kind alone or two or more kinds in combination.

[0050] The content of the organic acid in the organic acid-peracid solution is, for example, 5 to 30 mol / L, preferably 8 to 25 mol / L, and more preferably 10 to 20 mol / L. The content of the organic acid in the organic acid-peracid solution is preferably 3 to 10 g, more preferably 4 to 8 g, and even more preferably 5 to 7 g per 1 g of plant biomass to be contacted with the organic acid-peracid solution.

[0051] The content of peracid in the organic acid-peracid solution is, for example, 0.05 to 3 mol / L, preferably 0.08 to 2.5 mol / L, and more preferably 0.1 to 2 mol / L. The content of peracid in the organic acid-peracid solution is preferably 0.03 to 0.1 g, more preferably 0.04 to 0.08 g, and even more preferably 0.05 to 0.07 g per 1 g of plant biomass to be contacted with the organic acid-peracid solution.

[0052] The manner in which the plant biomass is brought into contact with the organic acid-peracid solution is not particularly limited, but from the viewpoint of treatment efficiency and the like, it is preferable to immerse the plant biomass in the organic acid-peracid solution.

[0053] The temperature of the organic acid-peracid solution is not particularly limited, as long as it is a temperature that can extract and solubilize at least one selected from the group consisting of lignin, hemicellulose, and lignin-polysaccharide complexes. The temperature of the organic acid-peracid solution is, for example, 15 to 160°C (preferably 40 to 160°C). By using a temperature of this range, it is possible to obtain high-quality (low-condensation) lignin and a complex of the lignin and polysaccharide, which have a high content of β-O-4 ether structures among all the bond types between monolignols. In step (A), treatment at a higher temperature tends to produce a lignin-polysaccharide complex, while treatment at a lower temperature tends to produce lignin alone. For example, when the starting material is woody biomass, performing step (A) with an organic acid-peracid solution at, for example, 90°C or lower (preferably 40 to 90°C, more preferably 40 to 70°C, and even more preferably 40 to 60°C) tends to selectively produce lignin alone, while performing step (A) with an organic acid-peracid solution at, for example, higher than 90°C and lower than 160°C (preferably 100 to 160°C, more preferably 120 to 160°C) tends to produce a complex of lignin and polysaccharides. For example, when the starting material is herbaceous biomass, performing step (A) with an organic acid-peracid solution at, for example, 60°C or lower (preferably 15 to 60°C) tends to selectively produce lignin alone, while performing step (A) with an organic acid-peracid solution at, for example, higher than 60°C and lower than 150°C (preferably 70 to 150°C) tends to produce a complex of lignin and polysaccharides. When heating is performed, examples of the method include internal heating using microwaves and external heating using an oil bath, but internal heating using microwaves is preferred. Rapid internal heating using microwaves enables efficient pretreatment in a short time.

[0054] The treatment time in step (A) is not particularly limited as long as it is long enough to achieve the above-mentioned object, and is, for example, 1 to 60 minutes, preferably 1 to 30 minutes, and more preferably 5 to 20 minutes.

[0055] In step (A), it is preferable to perform microwave treatment while the biomass is immersed in the solution. This improves the lignin yield. The microwave treatment conditions can be set appropriately, but for example, a commercially available microwave synthesis device can be used to perform the treatment while stirring the solution. The microwave treatment time can be, for example, 1 to 30 minutes, and is preferably about 10 minutes. When microwave treatment is performed in the isolation method of the present invention, irradiation for a relatively short period of time is sufficient, so it can easily be constructed as a batch system or a continuous system.

[0056] In step (A), it is preferable to irradiate the plant biomass with microwaves or ultrasound, from the viewpoint of promoting penetration of the solution into the pores inside the plant cell walls and improving the yield. The irradiation time is the same as above.

[0057] Step (A) may be carried out in multiple stages while varying the conditions as necessary. For example, the solution temperature may be varied. In this case, it is preferable that the solution temperature in a particular stage is higher than that in the next stage. For example, step (A) may include (A1) contacting the plant biomass with a solution containing an organic acid and a peracid at 90°C or less, and / or (A2) contacting the plant biomass with a solution containing an organic acid and a peracid at a temperature higher than 90°C and 160°C or less. Note that the "plant biomass" in step (A2) includes the insoluble portion obtained in step (A1).

[0058] Lignin alone can be selectively extracted and solubilized in step (A1), and lignin-polysaccharide complexes can be extracted and solubilized in step (A2). Therefore, by performing step (A1) first and step (A2) in that order, lignin alone and lignin-polysaccharide complexes can be efficiently obtained.

[0059] After step (A), a separation solvent is added as needed to separate the soluble and insoluble fractions. The method for separating the soluble and insoluble fractions is not particularly limited, and methods such as decantation, spin-down, and filtration can be used. Lignin and lignin-hemicellulose copolymers are isolated using separation solvents such as dioxane, ethanol, acetone, acetonitrile, DMSO, DMF, NMI, various alcohols such as ethyl acetate, methanol, and butanol, acetic acid, water, and mixtures of these solvents. From the perspectives of extraction efficiency, selectivity, and environmental impact, organic solvents such as acetone, ethanol, and acetonitrile, as well as 10-20% v / v water-mixed organic solvents, are preferred.

[0060] Furthermore, using a water-free extraction solvent (acetic acid-peracetic acid) or separation solvent allows for the production of highly pure lignin-hemicellulose copolymers. When using a water-containing solvent, some of the hemicellulose polysaccharide component is also extracted at the same time. A catalyst that accelerates the hydrolysis reaction in the presence of an acid or alkali can also be used.

[0061] The soluble portion obtained in step (A) contains at least one selected from the group consisting of lignin, hemicellulose, and lignin-polysaccharide complexes. Therefore, the soluble portion can be obtained as is, or after undergoing other treatments as necessary, as at least one selected from the group consisting of lignin, hemicellulose, and lignin-polysaccharide complexes.

[0062] Other examples of treatments include the following: After step (A), if necessary, the water can be removed by evaporation and / or freeze-drying to obtain the target product as a dry powder. If desalting is required, the concentrated solution may be extracted with alcohol, acetone, or the like, and the solvent may be distilled off. This allows the target product to be obtained as a dry powder.

[0063] When lignin is selectively obtained, for example, lignin can be obtained from the soluble portion obtained in step (A1).

[0064] When a lignin-polysaccharide complex is selectively obtained, for example, the insoluble portion obtained in step (A1) is used as plant biomass in step (A2), and the lignin-polysaccharide complex can be obtained from the soluble portion obtained in step (A2).

[0065] In the case of obtaining hemicellulose, for example, the lignin-polysaccharide complex can be subjected to alkaline hot water extraction to obtain hemicellulose. If necessary, the hemicellulose can be purified by ethanol precipitation, ion exchange column chromatography, gel filtration, hydrophobic chromatography, or the like.

[0066] Hemicellulose can also be obtained by contacting the insoluble fraction obtained in step (A) (preferably step (A1)) with a hemicellulose extraction solution prepared by adding a peracid (the type and final concentration of which are as described in step (A) above) to an alkaline solution (the composition and the like are as described in step (X) above). The contact conditions are as described in step (A) above. The soluble fraction obtained in this step can be subjected to purification (dialysis, etc.), water removal, etc., as necessary, to obtain hemicellulose. On the other hand, cellulose (cellulose fiber) can be obtained from the insoluble fraction obtained in this step by subjecting it to bleaching, washing, etc., as necessary.

[0067] The insoluble portion obtained in step (A) contains cellulose. Therefore, the insoluble portion can be obtained as cellulose directly or, if necessary, after undergoing other treatments as described above. For example, hemicellulose can be extracted from the insoluble portion by subjecting it to alkaline hot water extraction. However, if the purpose is to obtain type I cellulose, it is preferable not to treat the insoluble portion with a high-concentration alkaline solution in order to avoid mercerizing the cellulose.

[0068] When lignin is selectively isolated from plant biomass in step (A1), the insoluble portion (residue) contains a hemicellulose-cellulose complex. When a material having a certain shape (e.g., wood chips) is used as the plant biomass, the hemicellulose-cellulose complex is obtained as relatively strong pieces of material. White material pieces can be obtained by decolorizing the pieces (by immersing them in an organic acid solution, washing, bleaching, etc.).

[0069] According to the present invention, a new plant biomass pretreatment method can be provided as a new paper pulp production process that requires low energy and does not produce hazardous waste liquid, and is a low-temperature, mild process that produces cleanly delignified polysaccharides and therefore produces fewer inhibitors in the recovered material that are problematic for ethanol fermentation.

[0070] 2. Lignin, lignin-polysaccharide complex, cellulose, hemicellulose-cellulose complex According to the isolation method of the present invention, it is possible to obtain lignin having at least the following characteristics: a β-O-4 ether structure content of 50% or more and a solubility of 2 w / v% or more in 60-80% ethanol at 25°C.

[0071] According to the isolation method of the present invention, it is possible to obtain a lignin-polysaccharide complex characterized by having at least one bond selected from the group consisting of an α-ether bond between lignin and polysaccharide, an α-ester bond between lignin and polysaccharide, and a γ-ester bond between lignin and polysaccharide.

[0072] According to the isolation method of the present invention, it is possible to obtain cellulose that is characterized at least by having fewer inhibitors that are problematic in ethanol fermentation.

[0073] According to the isolation method of the present invention, it is possible to obtain a hemicellulose-cellulose complex.

[0074] These will be explained below.

[0075] 2-1. Lignin In one aspect, the present invention relates to lignin having a β-O-4 ether structure content of 50% or more and a solubility of 2 w / v% or more in 60-80% ethanol at 25°C, or lignin obtainable by the isolation method of the present invention.

[0076] The term "lignin" used in the present invention refers to lignin isolated from biomass (including natural lignin). Natural lignin has a wide variety of bonding modes. Representative bonding modes of natural lignin include the following:

[0077]

[0078] The above is called a β-O-4 bond.

[0079]

[0080] The above is called a resinol (β-β) bond.

[0081]

[0082] The above is called a phenylcoumaran (β-5) bond.

[0083]

[0084] The above is called a 5-5 bond.

[0085]

[0086] The above is called dibenzodioxin (DBDO 5-5 / 4-O-β) binding.

[0087]

[0088] The above is called a 4-O-5 bond.

[0089]

[0090] The above is called β-1 bond.

[0091]

[0092] The above is called spirodienone binding.

[0093] In natural lignin, the content of β-O-4 ether structures is the highest among all bond types between monolignols, although this varies depending on the plant species. For example, in coniferous trees, the content is said to be 40-60%. In broad-leaved trees, such as eucalyptus, the content is said to be around 45%.

[0094] The lignin of the present invention has a β-O-4 ether structure content of 50% or more, preferably 60% or more, more preferably 65 to 85%, and even more preferably 70 to 80% of all bond types between monolignols.

[0095] The lignin of the present invention preferably has a β-O-4 ether structure content (%) of all bond types between monolignols, where x% is the lignin content in the raw biomass, and is preferably 1.2x% or more, more preferably 1.4x% or more, and even more preferably 1.6x% or more. The upper limit is preferably 2x%.

[0096] The content of β-O-4 ether structures among all the linkages between monolignols in lignin was measured as follows. Samples were dissolved in deuterated dimethyl sulfoxide (DMSO-d6) to a concentration of 1-10 wt% and NMR spectra were acquired using a Bruker Avance III 600 MHz NMR instrument equipped with a cryogenic probe. The content of β-O-4 ether structures was estimated from the signal volume in the heteronuclear single-quantum correlation spectroscopy (HSQC) spectrum. The quantitative values ​​obtained using the 2D NMR method were corrected according to the literature (Okamura, H. Nishimura, T. Nagata, T. Kigawa, T. Watanabe, and M. Katahira, "Accurate and molecular-size-tolerant NMR quantitation of diverse components in solution," Scientific Reports, 6, 21742. 2016).

[0097] The lignin of the present invention is characterized by its solubility in 60-80% ethanol at 25°C of 2 w / v% or more. Therefore, by using the lignin of the present invention, the self-assembled bodies described below can be easily obtained, and highly stable self-assembled bodies can be obtained. The solubility is preferably 4 w / v% or more, more preferably 5 w / v% or more, even more preferably 6 w / v% or more, and even more preferably 7 w / v% or more.

[0098] The solubility is measured as follows: The sample is stirred and dissolved in 25°C ethanol, and then separated into an ethanol-soluble portion and an ethanol-insoluble portion by solid-liquid separation. A 25°C aqueous ethanol solution (1 / 1 ethanol / water) is added to the ethanol-insoluble portion and stirred to dissolve, obtaining an aqueous ethanol-soluble portion. The aqueous ethanol-soluble portion is mixed with the ethanol-soluble portion to obtain a sample solution. The amount of aqueous ethanol added is adjusted so that the ethanol concentration of the sample solution is in the range of 60-80%. In the process of obtaining the soluble portion, the sample is dispersed and solubilized, if necessary, using an ultrasonic cleaner for 3 min at 28 kHz. The maximum sample concentration at which the sample solution becomes visually transparent is defined as the "solubility in 60-80% ethanol at 25°C."

[0099] The lignin of the present invention preferably has a constituent ratio of biphenyl-type lignin, which has a condensed structure, of 5% or less.

[0100] The lignin of the present invention preferably has a number average molecular weight (Mn) of 0.7 to 9 kDa, more preferably 1 to 5 kDa, and even more preferably 1.5 to 4 kDa.

[0101] The lignin of the present invention preferably has a weight average molecular weight (Mw) of 1 to 10 kDa, more preferably 1.5 to 8 kDa, and even more preferably 2 to 6 kDa.

[0102] In this invention, the number-average molecular weight (Mn) and weight-average molecular weight (Mw) of lignin are specifically measured as follows. The molecular weight of lignin can be determined by size exclusion chromatography (SEC), also known as gel permeation chromatography (GPC). The sample is dissolved in a solution of acetic anhydride and a base (e.g., pyridine) and reacted for 3-6 hours to obtain an acetylated product. This is then dissolved in tetrahydrofuran (THF) at a concentration of 2.0 mg / mL and separated by high-performance liquid chromatography. A Tosoh HZ-M column (150 mm x 4.6 mm id, 4 μm, three columns connected in series) is used. Analysis is performed by injecting 10 μL of sample, using tetrahydrofuran (THF) as the mobile phase at a column temperature of 40°C and a flow rate of 0.35 mL / min. A mass calibration curve (calibration curve) is prepared using standard polystyrene, Tosoh PStQuickC (molecular weights (MW) of 2,110,000, 427,000, 37,900, and 5970), pinoresinol (MW 352), and vanillin (MW 152), to determine the Mw and Mn of the sample.

[0103] The lignin of the present invention preferably has a polydispersity (Mw / Mn) of 2.5 or less, more preferably 2 or less.

[0104] 2-2. Lignin-Polysaccharide Complex In one aspect, the present invention relates to a lignin-polysaccharide complex having at least one bond selected from the group consisting of an α-ether bond between lignin and a polysaccharide, an α-ester bond between lignin and a polysaccharide, and a γ-ester bond between lignin and a polysaccharide, or a lignin-polysaccharide complex obtainable by the isolation method of the present invention.

[0105] The lignin-polysaccharide complex of the present invention refers to a lignin-polysaccharide complex in a state isolated from biomass.

[0106] Polysaccharides include, for example, xylan and mannan, as well as glucomannan, galactoglucomannan, glucuronoxylan, and arabinoglucuronoxylan. Hemicellulose is primarily composed of glucomannan in conifers, glucuronoxylan in broad-leaved trees, and arabinoxylan in herbaceous plants. The monomers mentioned above include xylose, arabinose, glucose, mannose, galactose, and uronic acid. The side chains also contain acetyl and methoxy groups. In herbaceous plants, hemicellulose includes ferulic acid and diferulic acid bound to the arabinose side chain of arabinoxylan.

[0107] The lignin-polysaccharide complex of the present invention is a lignin-polysaccharide complex that can be uniformly dispersed in a separation solvent and is characterized by a polymer structure in which lignin and polysaccharides are copolymerized through covalent bonds. The lignin-polysaccharide complex of the present invention is isolated by hydrolyzing a small portion (less than 10% of the bonds) of the hemicellulose main chain bonds in the lignin-polysaccharide complex structure in plant cell walls. By carrying out the hydrolysis reaction under mild conditions, excessive decomposition, side reactions, and degradation to lower molecular weight compounds can be suppressed.

[0108] The lignin-polysaccharide complex of the present invention is characterized by having at least one bond selected from the group consisting of an α-ether bond between lignin and polysaccharide (LC α-ether bond), an α-ester bond between lignin and polysaccharide (LC α-ester bond), and a γ-ester bond between lignin and polysaccharide (LC γ-ester bond).

[0109] In the lignin-polysaccharide complex of the present invention, the α-ether bond (LC α-ether), which is the copolymerization point between lignin and polysaccharide, is preferably concentrated at a concentration of 1.2 times or more compared to the original bond unit composition ratio of the raw material biomass.

[0110] In biomass, lignin and polysaccharides are also bound together by non-covalent bonds. These non-covalent bonds are due to hydrophobic interactions, hydrogen bonds, electrostatic interactions, and entanglement of polymer chains. For this reason, previous studies have not provided clear evidence that lignin and polysaccharides, which are polymers with different properties, form copolymers through covalent bonds in lignin-polysaccharide complexes. Furthermore, conventional methods for preparing lignin-polysaccharide complexes involve multiple steps, resulting in low yields (less than 5%) or low purity.

[0111] Clear evidence of the copolymerization point (covalent bond) in the lignin-hemicellulose copolymer can be confirmed by signal assignment using HSQC, a two-dimensional NMR technique (Figures 3 and 4). Further clear evidence can be confirmed by continuous correlation analysis using HMBC and TOCSY-HSQC. For more details, please refer to the literature (Nishimura et al., Direct evidence for α ether linkage between lignin and carbohydrates in wood cell walls, Sci Rep 2018).

[0112] LC α ether: C–H correlation signals at the α-position of lignin (LC ether, δH / δC = 4.50 ±0.15 ppm / 80.1 ±1 ppm region) and (LC ether, δH / δC = 4.95 ±0.2 ppm / 81.5 ±1.5 ppm region), which are derived from the ether bond between the hydroxyl group of the 6-position of hexose or pentose and the α-position (benzylic position) of lignin.

[0113] LC α ester: A C–H correlation signal at the α-position of lignin (LC α ester, δH / δC = 5.88 ±0.15 ppm / 74.0 ±1.5 ppm region) derived from the ester bond between the carboxyl group at position 6 of glucuronic acid (uronic acid) and the α-position (benzylic position) of lignin.

[0114] LC γ ester: C–H correlation signal at the γ position of lignin (LC γ ester, δH / δC = 4.0 ±0.15 ppm, 4.4 ±0.15 ppm, / 64.8 ±1.5 ppm region) derived from the ester bond between the carboxy group at position 6 of glucuronic acid (uronic acid) and the γ position of lignin.

[0115] The lignin-polysaccharide complex of the present invention is characterized by its solubility of 2 w / v% or more in 60-80% ethanol at 25°C. Therefore, by using the lignin-polysaccharide complex of the present invention, the self-assembled bodies described below can be easily obtained, and highly stable self-assembled bodies can be obtained. The solubility is preferably 4 w / v% or more, more preferably 5 w / v% or more, even more preferably 6 w / v% or more, and even more preferably 7 w / v% or more.

[0116] In the lignin-polysaccharide complex of the present invention, the bond unit composition ratio of LC α ether and LC α ester, which are copolymerization points, is preferably 5% or more, more preferably 10% or more, in terms of the lignin aromatic ring ratio.

[0117] In the lignin-polysaccharide complex of the present invention, the ratio of polysaccharide to lignin is preferably 0.1 to 2, more preferably 0.3 to 1.

[0118] The lignin-polysaccharide complex of the present invention preferably has a molecular weight (Mw) of 2,000 to 15,000, more preferably 3,000 to 7,000. The lignin-polysaccharide complex of the present invention preferably has a polydispersity index (Mw / Mn) of 3.5 or less, more preferably 2.5 or less.

[0119] The method for measuring each of the above parameters is in accordance with the measurement method for lignin of the present invention or the measurement method in the Examples described below.

[0120] 2-3. Cellulose In one aspect, the present invention relates to cellulose obtainable by the isolation method of the present invention.

[0121] The cellulose of the present invention refers to cellulose in a state isolated from biomass.

[0122] The cellulose of the present invention has reduced levels of inhibitors that are problematic in ethanol fermentation.

[0123] The cellulose of the present invention can be used in various fields, for example, as cellulose fiber, pulp for papermaking, pretreatment of plant biomass, biofuel produced by saccharification and fermentation, and the like.

[0124] 2-4. Hemicellulose-Cellulose Complex In one aspect, the present invention relates to a hemicellulose-cellulose complex obtainable by the isolation method of the present invention.

[0125] The hemicellulose-cellulose complex of the present invention refers to a hemicellulose-cellulose complex in a state isolated from biomass.

[0126] The hemicellulose-cellulose composite of the present invention can be in the form of a material piece (a material piece having a certain shape and size), and can be white (see FIG. 13).

[0127] The hemicellulose-cellulose composite of the present invention can be obtained under mild conditions (treatment at 60°C or below using a low-concentration alkaline solution, an organic acid, a low-concentration peracid, or a combination of any of these), and therefore can be obtained while retaining the natural hemicellulose structure. For example, in hardwoods, the hemicellulose-cellulose composite has hemicellulose with a high degree of acetyl groups and side-chain glucuronic acid retained (see Figures 13, 14, and 15). The hemicellulose-cellulose composite of the present invention can be used in a variety of fields, such as thermal insulation materials, lightweight carbon materials, acoustics, soundproofing, and piezoelectric elements.

[0128] Furthermore, the hemicellulose-cellulose composite of the present invention can be impregnated with a resin to form a transparent material. From this perspective, in one aspect, the present invention relates to a transparent material comprising the hemicellulose-cellulose composite of the present invention and a resin. The resin impregnation method and the resin to be used can be selected according to publicly known information (e.g., JP-A-2020-515437). For example, the resin can be selected from poly(methyl methacrylate) (PMMA), epoxy, poly(glycidyl methacrylate) (PGMA), polydimethylsiloxane (PDMS), polystyrene (PS), or copolymers or mixtures thereof, all of which have a refractive index of 1.4 to 1.6 or 1.45 to 1.55.

[0129] 3. Self-Assembled Body In one aspect, the present invention relates to at least one self-assembled body selected from the group consisting of the lignin of the present invention and the lignin-polysaccharide complex of the present invention.

[0130] The shape of the self-assembled body of the present invention is not particularly limited, and can be, for example, a sheet (e.g., a coating film, a coating film, etc.) or a particle (e.g., fine particles, aggregates, etc.).

[0131] The lignin of the present invention and the lignin-polysaccharide complex of the present invention can form a self-assembled body. Self-assembly occurs easily, and the conditions for self-assembly can be appropriately set based on known conditions. For example, a sheet-like self-assembled body can be obtained by applying an aqueous solution containing at least one selected from the group consisting of the lignin of the present invention and the lignin-polysaccharide complex of the present invention onto a solid surface and evaporating the solvent from the aqueous solution.

[0132] In the above, the aqueous solution may be, for example, a mixture of water and an organic solvent such as alcohol (preferably ethanol) or acetonitrile (the separation solvents described above can be used). Specifically, for example, a mixture of ethanol and water may be used, and the mixing ratio in such a mixture is not particularly limited and can be set appropriately, but for example, the ethanol:water (volume ratio) may be 5:95 to 40:60.

[0133] The self-assembled bodies of the present invention exhibit fluorescence in the long wavelength region. Specifically, they exhibit fluorescence in the wavelength region of 400 nm to 700 nm. Furthermore, in the self-assembled bodies of the present invention, fluorescence that is significantly shifted to shorter wavelengths than the excitation wavelength, known as anti-Stokes fluorescence, is observed. Anti-Stokes fluorescence is exhibited even when a weaker light source than conventional sources, such as a xenon lamp, is used as the excitation light source. Therefore, the self-assembled bodies of the present invention can be used as phosphors (preferably phosphors that exhibit anti-Stokes fluorescence). Specifically, the self-assembled bodies of the present invention can be used as anti-Stokes fluorescent agents. In this case, for example, agents containing the self-assembled bodies of the present invention can be used as anti-Stokes fluorescent paints, inks, identification agents, and up-conversion / wavelength conversion molecular materials.

[0134] The self-assembled body of the present invention is stably dispersed in an aqueous solution, and can maintain the above-mentioned fluorescence even after drying as a solid.

[0135] 4. Ultraviolet Absorber The lignin of the present invention and the lignin-polysaccharide complex of the present invention can exhibit strong absorption of ultraviolet light, particularly UV-B and UV-C. From this perspective, in one aspect, the present invention relates to an ultraviolet absorber comprising at least one member selected from the group consisting of the lignin of the present invention and the lignin-polysaccharide complex of the present invention.

[0136] The ultraviolet absorber of the present invention can be mixed with a polymer to prevent ultraviolet degradation of the polymer, and can also be incorporated into a topical composition to protect the skin from ultraviolet rays.

[0137] The present invention will be described below with reference to examples, but the present invention is not limited to these examples, etc. Unless otherwise specified, units in the tables represent mass.

[0138] Test Example 1. Isolation of lignin, lignin-polysaccharide complexes, hemicellulose, and cellulose Lignin, lignin-polysaccharide complexes, hemicellulose, and cellulose were isolated from biomass using the M-APA method, the A-APA method, or a method for examining their conditions, and the yield and physical properties of the isolated products were measured. The isolation method of the present invention is a production method that enables highly efficient isolation of soluble lignin, lignin-polysaccharide complexes, hemicellulose, and cellulose from natural polymers in biomass, and includes a step (A) of contacting plant biomass with a solution containing an organic acid and a peracid. Specifically, the method was carried out as follows.

[0139] Test Example 1-1 Preparation of Finely Pulverized Wood Flour Fine powder (finely pulverized wood flour) with a volume average particle size of 10 μm (1-100 μm) was prepared from wood flour of broad-leaved trees (eucalyptus or beech), coniferous trees (red pine, cedar, or cypress), or herbaceous plants (bagasse (sugar cane) or bamboo) using a ball mill or bead mill method.

[0140] An example of a preparation method using a ball mill is shown below. A planetary ball mill (P-6, Fritsch) was used. 2 g of wood flour and 100 g of 3 mm diameter zirconia beads were placed in an 80 cc agate container and sealed in a twin overpot. After evacuating the overpot for 20 minutes, inert nitrogen gas was injected. One cycle consisted of 1 minute of milling followed by a 1 minute 20 second pause, and the milling was carried out for 180 cycles at 550 rpm (a total of 3 hours). These conditions allowed for the production of finely powdered wood flour while suppressing oxidation and heat-induced degradation during milling.

[0141] Test Example 1-2. Preparation of wood flour In two pots, left and right, of a mixer mill (MM301, manufactured by Retsch), 1 g each of wood flour or wood chips (10 mm x 10 mm x 1 mm) from a broad-leaved tree (eucalyptus or beech), a conifer (red pine, cedar, or cypress), or a herbaceous plant (bagasse (sugar cane) or bamboo) was placed and ground for a grinding time of 5 minutes at a frequency of 1 / 15 seconds, yielding a powder (wood flour) with a volume average particle size of 0.1 mm.

[0142] Test Example 1-3. M-APA (Milling-Acid-PerAcid) Method The starting material was the finely ground wood flour (average particle size 10 μm) obtained in Test Example 1-1. 2 g of finely ground wood flour and 10 mL of an acetic acid-peracetic acid solution (composition: 17.4 M acetic acid (99%), 0.14 M peracetic acid (slightly less than 1%), 0.01 M hydrogen peroxide, and trace amounts of water) were placed in an initiator vial and microwaved for 10 min using a Biotage Initiator+ 60 microwave synthesizer at 50°C, initial power 400 W, and stirring speed 600 rpm. After microwave treatment, the soluble fraction was separated by adding 10 mL of separation solvent (acetone) and spinning down (RCF 8000 xg, 5 min). The insoluble fraction was extracted with 10 mL of separation solvent (acetone) and washed three times, and then combined with the soluble fraction. This soluble lignin is called acid peroxyacid lignin (APA lignin).

[0143] The insoluble fraction was subjected to the second extraction procedure, but the microwave heating conditions were changed to 140°C. The soluble fraction was identified as lignin-hemicellulose copolymer. The insoluble fraction was obtained as a cellulose-based copolymer.

[0144] Test Example 1-4. A-APA Method (Alkali-Acid-Peracid Method) The wood flour (average particle size 0.1 mm) obtained in Test Example 1-2 was used as the starting material. A 0.25 M sodium hydroxide solution was prepared (1N diluted 4 times). 2.4 mL of 0.25 M sodium hydroxide solution was added to 0.3 g of wood flour, and microwave extraction was performed (100°C, 30 min). The required amount of alkali is approximately 1 mmol of NaOH per 1 g of wood flour.

[0145] After microwave treatment, the soluble and insoluble fractions can be easily separated by decantation, spin-down, or filtration. An additional three rounds of acetone (separation solvent) extraction were performed and combined with the soluble fraction. The insoluble fraction was extracted using the same extraction method as the M-APA method in Test Example 1-3 to obtain soluble lignin: acid peroxide lignin (APA lignin) and lignin-hemicellulose copolymer. The insoluble fraction was composed primarily of cellulose.

[0146] <Test Example 1-5. Conditions> An investigation was carried out using other organic acids (formic acid, glyoxylic acid) instead of acetic acid. Furthermore, an investigation was carried out using other peracids (hydrogen peroxide) instead of peracetic acid. When other peracids (hydrogen peroxide) were used, the composition was acetic acid: 12 M, and hydrogen peroxide concentration: the concentrations shown in the table below.

[0147] The contact temperature between plant biomass or insoluble matter and organic acid + peracid was varied from room temperature to 180°C. In addition, the heating method used was an external heating method using an oil bath, instead of an internal heating method using microwaves.

[0148] Test Example 1-6: Analysis and Measurement Methods Lignin, lignin-polysaccharide complexes, and cellulose were analyzed and measured by the following methods.

[0149] <Test Example 1-6-1. Molecular Weight (Mn, Mw, and Polydispersity) Measurement Method> Molecular weight was determined by size exclusion chromatography (SEC), also known as gel permeation chromatography (GPC). The sample was dissolved in a solution of acetic anhydride and a base (e.g., pyridine) and reacted for 3-6 hours to obtain the acetylated product. This was dissolved in tetrahydrofuran (THF) at a concentration of 2.0 mg / mL and separated by high-performance liquid chromatography (Shimadzu Corporation, high-pressure gradient SEC analysis system). A Tosoh HZ-M column (150 mm x 4.6 mm id, 4 μm, three columns connected in series) was used. Analysis was performed by injecting 10 μL of sample, at a column temperature of 40°C, using tetrahydrofuran (THF) as the mobile phase at a flow rate of 0.35 mL / min. Mass calibration curves were constructed using standard polystyrene, Tosoh PStQuickC (molecular weights (MW) of 2,110,000, 427,000, 37,900, and 5970), pinoresinol (MW 352), and vanillin (MW 152), to determine the weight-average molecular weight (Mw) and number-average molecular weight (Mn) of the samples. Polydispersity was calculated as Mw / Mn.

[0150] <Test Example 1-6-2. Calculation of the Composition Ratio of Molecular Bonding Units> Samples were dissolved in deuterated dimethyl sulfoxide (DMSO-d6) at 1-10 wt% and NMR spectra were acquired using a Bruker Avance III 600 MHz NMR instrument equipped with a cryogenic probe. The composition ratio of lignin interunit bonding units was calculated from the signal volume in the HSQC (heteronuclear single-quantum correlation spectroscopy) spectrum. The composition ratio was calculated as the ratio per 100 lignin aromatic rings. The content of ether-type bonding units was estimated from the sum of the integrals of the C-H correlation signals at the β-position (δH / δC = 4.0 ± 0.2 ppm / 85.6 ± 1.2 ppm, δH / δC = 4.3 ± 0.2 ppm / 83.0 ± 0.7 ppm). Quantitation in the two-dimensional NMR method was performed by correcting using the TAF method described in the literature (Okamura, H. Nishimura, T. Nagata, T. Kigawa, T. Watanabe and M. Katahira, Accurate and molecular-size-tolerant NMR quantitation of diverse components in solution, Scientific Reports, 6, 21742. 2016), and accurate content was calculated.

[0151] The integration region may shift slightly depending on the sample and measurement environment, so the signal integration region is adjusted appropriately during analysis (this also applies to the other measurement methods described in this specification).

[0152] Test Example 1-6-3. Calculation method for the composition ratio of lignin and polysaccharide The sum of the integral values ​​of the C—H correlation signals at the anomeric position (1st position) of the polysaccharide was calculated by the NMR method described above, and after correction by the TAF method, the ratio was calculated as a ratio per 100 lignin aromatic rings.

[0153] Test Example 1-6-4: Calculation of the composition ratio of lignin-polysaccharide copolymerization points As described above, the sum of the integrals of the C-H correlation signals (LC ether, δH / δC = 4.50 ± 0.15 ppm / 80.1 ± 1 ppm region) and (LC ester, δH / δC = 5.88 ± 0.15 ppm / 74.0 ± 1.5 ppm region) derived from LC α ether and LC α ester bond units was calculated using 2D HSQC NMR. After correction using the TAF method, the ratio was calculated as a ratio per 100 lignin aromatic rings. Although the A-APA method hydrolyzes some of the ester bonds by alkali treatment, it is also possible to obtain LC α ester-type lignin-hemicellulose copolymers.

[0154] <Test Example 1-6-5. Method for calculating yield from raw biomass> After checking the pH of each obtained component using a pH meter or pH test paper or peracid test paper (potassium iodide starch paper), neutralization and peracid quenching were performed as necessary. Neutralization was performed by adding 1N HCl or 1N NaOH in small amounts. Peracid quenching was performed by adding 1M aqueous sodium sulfite solution dropwise. The organic solvent was removed using an evaporator. A dry powder was then obtained by freeze-drying. If salts were generated by neutralization, the concentrated solution after organic solvent removal was re-extracted with a separation solvent (acetone) for desalting. The solvent was then removed again using an evaporator, and a dry powder was obtained by freeze-drying. Each obtained component was weighed, and the yield per raw biomass was calculated.

[0155] <Results> The molecular structure, molecular weight, and yield of the obtained polymer are desirable under the following conditions. The results are shown in Tables 1 to 7.

[0156] Table 1 shows the peracid conditions in the two-stage microwave extraction reaction of the M-APA method and the yields of each polymeric lignocellulose component (weight %, raw material biomass ratio, raw material is eucalyptus).

[0157]

[0158] Table 2 shows the yields (wt %, ratio to raw material biomass) of each polymeric lignocellulose component of various biomass in the A-APA method two-stage microwave extraction reaction.

[0159]

[0160] Table 3 shows the peracid conditions in the two-stage microwave extraction reaction of the M-APA method and the molecular weights of each component of high molecular weight lignocellulose (acetylated GPC analysis values).

[0161]

[0162] Table 4 shows the yields (wt %, ratio to raw material biomass) of each polymeric lignocellulose component of various biomass in the two-stage microwave extraction reaction of the M-APA method.

[0163]

[0164] Solvent: Acetic acid (containing 1% peracetic acid, 0.14 mol / L) Microwave extraction conditions: 50°C, 10 min (first stage) to obtain APA lignin soluble fraction, 140°C, 10 min (second stage) to obtain lignin hemicellulose copolymer. Table 5 shows the yields (wt%, ratio to raw biomass) of each polymeric lignocellulose component of various biomass in the A-APA method one-stage microwave extraction reaction.

[0165]

[0166] Table 6 shows the molecular weights (acetylated GPC analysis values) of each component of high molecular weight lignocellulose obtained from various biomass materials by the A-APA method.

[0167]

[0168] As a result of examining the conditions, it was found that when the starting material is woody biomass, a contact temperature of 40 to 90°C with organic acid and peracid is suitable for obtaining soluble lignin, and a contact temperature of 100 to 160°C is suitable for obtaining lignin-hemicellulose copolymer.Furthermore, as a result of examining the conditions, it was found that when the starting material is herbaceous biomass, a contact temperature of room temperature to 60°C is suitable for obtaining soluble lignin, and a contact temperature of 70 to 150°C is suitable for obtaining lignin-hemicellulose copolymer.

[0169] As a result of examining the conditions, it was found that the heating time by microwave irradiation is 5-30 min, preferably 10 min. The net irradiation time is short. See Figure 2 for the microwave profile, temperature, and pressure data.

[0170] As a result of examining the conditions, it was found that microwave heating had higher yield and selectivity than external heating under conditions of low temperature, short time, and low energy input. In other words, the microwave effects of local heating, internal heating, electron transfer, and drug penetration promotion were confirmed.

[0171] As a result of examining the conditions, lignin could be isolated using all of the organic acids, acetic acid, formic acid, and glyoxylic acid, but acetic acid was the most suitable from the standpoints of yield, purity, and industrial application.

[0172] The NMR results for Test Examples 1-6-2 and 1-6-3 are shown in Figure 3. Figure 3 shows the analysis results for lignin isolated by the M-APA method (microwave treatment at 50°C for 10 minutes, acetic acid and peracetic acid 1% conditions) using eucalyptus as the starting material. The proportion of lignocellulose interunit bonds was quantified using TAF-NMR, and the results showed that 99% of the signals were derived from lignin and 1% from polysaccharides (hemicellulose). The breakdown of lignin interunit bonds was 81% ether-type β-O-4, 5% β-5, 11% β-β, 1% dibenzodioxocine, and 1% other. See Figure 4 for partial chemical structures of lignin interunit bonds and lignin-hemicellulose covalent bonds.

[0173] The NMR results for Test Example 1-6-4 are shown in Figure 5. Figure 5 shows the analysis results for lignin-hemicellulose copolymers isolated by the M-APA method (microwave treatment at 50°C for 10 minutes, acetic acid peracetic acid 1%) using each plant as the starting material. LC bonds (ether and ester types), shown by circles in the figure, are the main branching structures present in proportions equal to or greater than the β-5 and β-β bonds, which are the main interunit bonds of lignin. The isolated lignin-hemicellulose copolymers are characterized by the presence of LCα-ester type bonds in particular.

[0174] Test Example 2. Preparation of self-assembled bodies Self-assembled bodies were prepared using lignin isolated by the M-APA method (microwave treatment at 50°C for 10 minutes, acetic acid-peracetic acid 1% or acetic acid-0.2M hydrogen peroxide conditions) using eucalyptus or bamboo as the starting material, or lignin-polysaccharide copolymer isolated by the M-APA method (microwave treatment at 140°C for 10 minutes, acetic acid-peracetic acid 1% or acetic acid-0.2M hydrogen peroxide conditions) using eucalyptus as the starting material. Specifically, the procedure was as follows.

[0175] A DMSO solution with a sample concentration of 1-2 wt% was prepared and diluted to 0.01-0.02 wt% with a mixture (1:1 or 2:1) of a separating solvent (acetonitrile or ethanol) and water (or buffer solution). 0.1-0.2 mL of the sample solution was applied to a glass slide or cover glass, and the solvent was evaporated under atmospheric pressure to obtain a thin film of self-assembled aggregates. Even under the above conditions, a high-concentration self-assembled aggregate was obtained by adding an equal volume of water (or buffer solution) to a DMSO solution with a sample concentration of 1 wt% and applying it.

[0176] Next, the optical properties of the obtained self-assembled bodies (lignin solid thin films or lignin-polysaccharide copolymer solid thin films) and a comparison object (lignin obtained by the APA method (APA lignin)) were analyzed. Specifically, the optical properties were analyzed as follows.

[0177] The following was used for the measurements.

[0178] Ultraviolet-visible absorption spectrum: UV-2700 (Shimadzu Corporation) Fluorescence spectrum: Spectrofluorometer RF-6000 (Shimadzu Corporation) Fluorescence microscope measurement: All-in-one fluorescence microscope BZ-X810 (Keyence Corporation) Analysis was performed using the analysis software provided with each device.

[0179] UV-visible absorption spectrum: black quartz cell for spectroscopic measurement (optical path length: 10 mm), fluorescence measurement: fully transparent synthetic quartz cell (optical path length: 10 mm) and Starna triangular quartz cell SF3 were used. For solid samples, a solid measurement unit was used, and the sample was applied to a cover glass and sandwiched between quartz plates for measurement.

[0180] The 3D fluorescence spectrum was measured at an excitation wavelength of 310-800 nm with a data interval of 5 nm, an emission wavelength of 350-900 nm with a data interval of 1 nm, a scan speed of 12,000 nm / min, and an ultraviolet cut filter U310 attached to the excitation side to reduce the effects of scattered light and multi-order light. The bandwidth was 5 nm on the excitation side and 15 nm on the emission side, and the sensitivity was set to Low.

[0181] Synchroscan spectroscopy is a scanning mode in which the excitation and fluorescence spectrometers of a spectrofluorometer are scanned simultaneously, with a fixed wavelength interval between them. Measurements were performed by varying the wavelength interval between the excitation and fluorescence light in the range of -90 nm to 260 nm. When the wavelength interval is negative, fluorescence with shorter wavelengths than the excitation light is detected. In this case, fluorescence due to the anti-Stokes shift can be selectively detected. The scan was set to cover the fluorescence wavelength range of 350-800 nm, with a data interval of 0.1 nm, a scan speed of 600 nm / min, a bandwidth of 15 nm on the excitation side and 20 nm on the emission side, and a sensitivity setting of Low.

[0182] The UV-visible absorption spectrum was measured using a double monochromator UV-2700. A wavelength scan (200-900 nm) was performed.

[0183] The results are shown in Figures 6 to 9. Figure 9 shows that the lignin solid thin film or the lignin-polysaccharide copolymer solid thin film exhibits anti-Stokes fluorescence.

[0184] Test Example 3. Differential thermal and thermogravimetric analysis Simultaneous differential thermal and thermogravimetric analysis was performed on APA lignin, lignin hemicellulose copolymer, and cellulose obtained by the APA method, along with their raw material, eucalyptus wood flour. The Shimadzu DTG-60 instrument was used. Approximately 10 mg of each powder sample was placed in a platinum cell, and after 15 minutes of nitrogen substitution at 30°C in a nitrogen atmosphere, the temperature was raised to 1000°C at a rate of 20°C / min and measurements were performed. The results are shown in Figures 10-1 and 10-2.

[0185] Test Example 4: Solubility Measurement The solubility of lignin and lignin-hemicellulose copolymers obtained by the method of Test Example 1 or a method similar thereto was measured in 60-80% ethanol at 25°C. Details of the samples measured are shown in Table 7. The abbreviations in Table 7 are as follows: PA: Peracetic acid (peracetic acid), HP: Hydrogen Peroxide, 50: MW50 ACTN, microwave-treated at 50°C, high-quality polymeric lignin, 140: MW50-MW140 ACTN, residue after microwave-treated at 50°C, second-stage treatment at 140°C, lignin-hemicellulose copolymer, A50: Acetic acid-peracetic acid system, HP, PA 1% AA, Mw50 ACN, high-quality polymeric lignin treated at 50°C, H50: Acetic acid-hydrogen peroxide system, HP, 1M hydrogen peroxide, Mw50 ACN, high-quality polymeric lignin treated at 50°C.

[0186]

[0187] The solubility was measured as follows. The sample was dissolved in 25°C ethanol by stirring, and then separated into ethanol-soluble and ethanol-insoluble portions by solid-liquid separation. A 25°C aqueous ethanol solution (1 / 1 ethanol / water) was added to the ethanol-insoluble portion and stirred to dissolve, obtaining the aqueous ethanol-soluble portion. The aqueous ethanol-soluble portion was mixed with the ethanol-soluble portion to obtain a sample solution. The amount of aqueous ethanol added was adjusted so that the ethanol concentration of the sample solution was in the range of 60-80%. In the process of obtaining the soluble portion, dispersion and solubilization were performed using an ultrasonic cleaner for 3 min at 28 kHz, as necessary. The maximum sample concentration at which the sample solution became visually transparent was defined as the "solubility in 60-80% ethanol at 25°C."

[0188] The results are shown in Table 8. It was found that the lignin and lignin-hemicellulose copolymer obtained by the method of the present invention exhibited good ethanol solubility.

[0189]

[0190] Test Example 5. Measurement of UV absorbance The UV absorbance was measured for the lignin and lignin-hemicellulose copolymer used in Test Example 4. Specifically, a sample was dissolved in ethanol to a concentration of 0.005 w / v %, and the UV absorption spectrum of the resulting solution (transparent) was measured using a spectrophotometer (Shimadzu Corporation, UV-2700) (optical path length 1 cm, quartz cell).

[0191] The results are shown in Figure 11. It was found that the lignin and lignin-hemicellulose copolymer obtained by the method of the present invention exhibit good ultraviolet absorption properties.

[0192] Test Example 6: Isolation of lignin from wood chips Lignin was isolated by the A-APA method using beech sapwood (size: 10 mm × 10 mm × 2 mm, 1.0±0.1 g) as wood chips. Specifically, the procedure was as follows.

[0193] Wood pieces were immersed in 10.0 mL of alkaline solution (0.25 M NaOH, 0.25 M KOH, 0.1 M NaOH, or 0.1 M KOH) and subjected to ultrasonic irradiation for 15 minutes (Honda Electronics ultrasonic cleaner, W-113 multimode, 28 kHz, 45 kHz, 100 kHz). After 1 hour, the alkaline solution was filtered off. After washing, the wood pieces were immersed in 10 mL of acetic acid peroxide solution (prepared by adding 1 mL of 30% H2O2 to 9 mL of acetic acid, final hydrogen peroxide concentration 3% v / v) and subjected to ultrasonic irradiation for 30 minutes (Honda Electronics ultrasonic cleaner, W-113 multimode, 28 kHz, 45 kHz, 100 kHz). The immersion solution was recovered, and the recovered acetic acid peroxide solution was concentrated by evaporation and freeze-dried to obtain lignin powder. The yield was 3%.

[0194] Test Example 7: Isolation of lignin and white wood chips (hemicellulose-cellulose complex) from wood chips. Using beech sapwood (size: 10 mm x 10 mm x 2 mm, 1.0±0.1 g) from hardwood and cedar sapwood (size: 10 mm x 10 mm x 2 mm, 0.7±0.1 g) from softwood, lignin and hemicellulose-cellulose complex were isolated by the A-APA method. Specifically, the procedure was as follows.

[0195] The wood pieces were immersed in 10.0 mL of alkaline solution (0.25 M NaOH, 0.25 M KOH, 0.1 M NaOH, or 0.1 M KOH) and ultrasonicated for 30 minutes (Honda Electronics ultrasonic cleaner, W-113, multimode, 28 kHz, 45 kHz, 100 kHz). The water in the ultrasonic cleaner was replaced (the same amount of water added was drained) to maintain the solution temperature at approximately 50 °C. After 1 hour, the alkaline solution was filtered. The resulting wood pieces were then immersed in 10 mL of acetic acid peroxide solution (prepared by adding 1.0 mL of 30% H2O2 to 9 mL of acetic acid, with a final hydrogen peroxide concentration of 3% v / v) and ultrasonicated for 30 minutes (Honda Electronics ultrasonic cleaner, W-113, 28 kHz). After 10 hours, the soaking solution was recovered, and the pieces were again immersed in 10 mL of acetic acid peroxide solution and ultrasonicated for 30 minutes. This process was repeated twice, and the collected acetic acid peroxide solutions were combined and concentrated in a draft (using a Biochromato Convenience Evaporator), then redissolved in 1.5 mL of 80% dioxane v / v solution and freeze-dried. This yielded a white lignin powder. The yield was 8%.

[0196] 7-1) After the acetic acid peroxide treatment, the wood pieces were washed with pure water and then immersed in 10 mL of 0.1 M NaOH solution. 0.1 g of sodium percarbonate (Sigma-Aldrich, Na2CO3 1.5H2O2, CAS15630-89-4) was added, and ultrasonic waves were applied for 10 minutes (Honda Electronics ultrasonic cleaner, W-113 multimode 28 kHz, 45 kHz, 100 kHz). This was repeated three times at two-hour intervals. The immersion solution was dialyzed and then freeze-dried to obtain glucuronoxylan (4-O-Methyl-D-glucurono-D-xylan) as hemicellulose from the hardwood beech sapwood (Figure 12). The obtained product was 1 H- and 1 H- 13The residue was identified by C HSQC NMR analysis. Cellulose fibers were obtained by bleaching the residue, washing with pure water, and drying. Bleaching was carried out by adding sodium hypochlorite (5 v / v%, 5 mL) and stirring at 30°C, 1000 rpm, and 1 h (using an Eppendorf ThermoMixer C). The yield relative to the wood chips was 37%.

[0197] 7-2) Acetic acid was added to the acetic acid peroxide-treated wood chips, and decolorization progressed by continuing the immersion. After three days, the chips were washed with pure water and freeze-dried to obtain white wood chips. The white wood chips were hard and were pulverized with a cutter, after which FT-IR and solid-state CP / MAS NMR measurements were performed. A comparison was made with the cellulose obtained in 7-1. The results are shown in Figures 14 and 15. As a result, the white wood chips obtained in 7-2 (photographs shown in Figures 16a and 16c) were found to be a cellulose-hemicellulose complex. The yield relative to the wood chips was 68%.

[0198] In addition, white wood pieces were ultrasonically treated in a NaOH solution containing sodium percarbonate and washed in the same manner as in 7-1. The resulting wood pieces are shown in Figure 16b.

Claims

1. (A) contacting a solution containing an organic acid and a peracid with a plant biomass The present invention relates to a method for isolating at least one selected from the group consisting of lignin, hemicellulose, a lignin-polysaccharide complex, cellulose, and a hemicellulose-cellulose complex from plant biomass, the method comprising the steps of:

2. The step (A), (A1) contacting a plant biomass with a solution containing an organic acid and a peracid at 90° C. or less; and / or (A2) A step of contacting a plant biomass with a solution containing an organic acid and a peracid at a temperature higher than 90°C and lower than 160°C. The method of claim 1 , comprising:

3. The method according to claim 1 or 2, comprising obtaining at least one selected from the group consisting of lignin, hemicellulose, a lignin-polysaccharide complex, and a hemicellulose-cellulose complex from the soluble portion obtained in the step (A).

4. The method according to any one of claims 1 to 3, comprising obtaining at least one selected from the group consisting of cellulose, hemicellulose, and a hemicellulose-cellulose complex from the insoluble portion obtained in the step (A).

5. The method according to any one of claims 1 to 4, comprising irradiating the plant biomass with microwaves or ultrasound in the step (A).

6. The method according to any one of claims 1 to 5, wherein the plant biomass used in the step (A) is an alkali-treated plant body or a mechanically processed product thereof.

7. Lignin having a β-O-4 ether structure content of 50% or more and a solubility in 60-80% ethanol at 25°C of 2 w / v% or more.

8. Lignin obtainable by the method according to any one of claims 1 to 6.

9. A lignin-polysaccharide complex having at least one bond selected from the group consisting of an α-ether bond between lignin and a polysaccharide, an α-ester bond between lignin and a polysaccharide, and a γ-ester bond between lignin and a polysaccharide.

10. The lignin-polysaccharide complex according to claim 9, having a solubility of 2 w / v% or more in 60 to 80% ethanol at 25°C.

11. A lignin-polysaccharide complex obtainable by the method according to any one of claims 1 to 6.

12. Cellulose obtainable by the method according to any one of claims 1 to 6.

13. A hemicellulose-cellulose complex obtainable by the method according to any one of claims 1 to 6.

14. Hemicellulose obtainable by the method according to any one of claims 1 to 6.

15. At least one self-assembled body selected from the group consisting of the lignin according to claim 7 or 8 and the lignin-polysaccharide complex according to any one of claims 9 to 11.

16. The self-assembled body according to claim 15 , which is in the form of a sheet or particles.

17. A phosphor comprising the self-assembled body according to claim 15 or 16.

18. 18. The phosphor of claim 17, which exhibits anti-Stokes fluorescence.

19. An ultraviolet absorber comprising at least one selected from the group consisting of the lignin according to claim 7 or 8 and the lignin-polysaccharide complex according to any one of claims 9 to 11.

20. A lignin or lignin-polysaccharide complex, wherein a self-assembled body formed from the lignin or lignin-polysaccharide complex is capable of exhibiting anti-Stokes fluorescence.