Reaction behavior of Cryptomeria japonica treated with pyridinium chloride–water mixture

Abstract

Cryptomeria japonica was treated with 90% pyridinium chloride ([Py]Cl) and 10% water w/w solution at 80 and 120 °C. Most hemicellulose in C. japonica was liquefied and over half the lignin in C. japonica was solubilized after treatments at 80 and 120 °C. However, cellulose was mostly insoluble at 80 °C and partially soluble at 120 °C. The crystal structure of cellulose in the cell walls was retained after treatment at 80 °C for 48 h. The degradation products from the polysaccharides were obtained in different yields. The 90% [Py]Cl and 10% water w/w solution is effective for the treatment of lignocellulosics, such as liquefaction of lignocellulosics and the production of useful low molecular weight compounds.

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1. Introduction

Fossil resources have enormously enriched our life in the world. However, exploring new resources for raw materials and energy is required to sustain our society. Biomass is an attractive alternative to fossil fuels because it is renewable, recyclable and reusable. Among biomass sources, lignocellulosics are the most abundant on the earth. They are mostly composed of three polymers: cellulose, hemicellulose and lignin. Cellulose is the predominant component in lignocellulosics, and its chemical structure is linked glucose rings through β-1,4 glycosidic bonds. Hemicelluloses are composed of monosaccharides such as glucose, mannose, xylose, galactose and arabinose. These polysaccharides can be converted to biofuels¹ and useful chemicals by chemical and biological reactions.² Lignin is a heterogeneous biopolymer synthesized from monolignols via oxidative radical coupling. It can be transformed to useful aromatic chemicals and liquid fuels through degradation or structural conversion.^3,4

These polymers constitute the cell wall in lignocellulosics and have complicated and rigid three-dimensional network structures, which causes the recalcitrance of lignocellulosics to chemical and biological treatments. Therefore, effective, efficient and economically feasible treatments of lignocellulosics are necessary to convert these components into value-added products. To overcome these difficulties, developments in the liquefaction, degradation and modification of lignocellulosics have been increasingly performed.

Highly polar solvents containing salts have been applied for the complete liquefaction of whole wood cell walls.^5,6 Studies on the treatment of lignocellulosics with ionic liquids (ILs), which are organic salts with melting points near ambient temperature, have been increasing in the past 10 years because of the advantages in high dissolving power, low vapor pressure, low toxicity and reusability. ILs have been widely used for the liquefaction of lignocellulosics and related compounds.^7–10 In addition, the reaction behavior and microscopic analyses of wood with ILs have been reported,^11–14 and many ILs have been used for the conversion of lignocellulosics into valuable compounds. Saccharides (e.g., glucose, fructose and cellulose) have been used as starting materials in the production of 5-hydroxymethylfurfural (5-HMF), furfural, 2-hydroxyacetylfuran (2-HAF), levulinic acid (Fig. 1) and other low molecular weight compounds.^15–22 However, few studies on the production of useful chemicals from wood with ILs without catalysts have been performed, and most of these studies with ILs have been conducted without water addition. An IL with water addition has three advantages for biorefining lignocellulosics, as described in a previous paper.²² First, the drying step of lignocellulosics is demanded in the industrial process because they contain water. However, IL–water method can omit this drying step. Second, the solid-state ILs at room temperature after addition of water can be used as a solvent for the treatment of lignocellulosics although the ILs cannot behave as solvents by themselves at room temperature. Third, the ILs with water addition reduce the financial cost and the potential environmental impact because the total amount of the ILs used is decreased.

Fig. 1 Chemical structures of degradation products obtained from saccharides.

Abe et al. reported tetra-n-butylammonium hydroxide and tetra-n-phosphonium hydroxide ([P_4,4,4,4]OH)–water mixtures liquefied wood at room temperature.²³ The liquefaction of cedar powder (final concentration of 5.0 wt%) was completed in 50% [P_4,4,4,4]OH aqueous solution at 60 °C under mild stirring for 24 h. Additionally, [P_4,4,4,4]OH–water mixtures can isolate polysaccharides from wood, and it is easy to control the component ratio of the isolates from poplar by changing the amount of water.²⁴

We previously reported the identification of 2-HAF,²² which was used as the substrate in the total synthesis of boronolide²⁵ from C. japonica treated with various IL–water mixtures at 120 °C. The 90% [Py]Cl and 10% water w/w solution was the most effective IL–water mixture for the production of furan compounds.²² However, studies on the treatment of lignocellulosics with IL–water mixtures and the analysis of the treated lignocellulosics have not been sufficiently conducted, although they have great potential to produce valuable compounds from lignocellulosics. Therefore, understanding the reaction behavior of C. japonica by the treatment with 90% [Py]Cl and 10% water w/w solution is important for biorefining lignocellulosics.

2. Experimental

2.1 Materials

Solvents were of analytical grade and purchased from Wako Pure Chemical Industries, Ltd. (Osaka, Japan). ILs and authentic samples of degradation products shown in Fig. 1 were purchased from Tokyo Chemical Industry Co., Ltd. (Tokyo, Japan), Wako Pure Chemical Industries, Ltd., Sigma-Aldrich (St. Louis, MO, USA) and Iolitech (Ionic Liquids Technologies GmbH, Heilbronn, Germany). [Py]Cl (>98.0%) was used for the all experiments in this study. Lignocellulosic samples were prepared with almost the same procedure as that in a previous paper.²² C. japonica (Japanese cedar) was ground to a particle size between 90 and 180 μm and successively extracted with ethanol/benzene (1/2, v/v) for 6 h in a Soxhlet extractor to remove several extractives. The extracted wood powder was dried in an oven at 105 °C for 24 h and stored at room temperature under air atmosphere (see Sections 2.3 and 2.4). In addition, a C. japonica sample (5 × 5 × 5 mm) was extracted as per the above method for microscopic analyses (see Section 2.6).

2.2 Instruments

High-performance liquid chromatography (HPLC) was conducted on a Shimadzu Prominence (Kyoto, Japan) equipped with a pump (LC-20AD), a column oven (CTO-20A), a refractive index detector (RID-10A) and photodiode array (SPD-M20A). HPLC conditions were as follows: an Aminex HPX-87H column (Bio-Rad Laboratories, Inc., Hercules, CA, USA), a flowrate of 0.6 mL min⁻¹, 5 mM H₂SO₄ as eluent, and a column temperature of 45 °C; or a KS-801 sugar column (Showa Denko Co. Ltd., Tokyo, Japan), a flowrate of 1.0 mL min⁻¹, Milli-Q water as eluent, and a column temperature of 80 °C.

2.3 Treatment of C. japonica with IL–water mixtures at room temperature

ILs shown in Table 1 (6.0 g) and distilled water (0.66 g) were added to a 100 mL round-bottom flask and stirred at room temperature. A cedar powder from Section 2.1 (0.12 g) was added to the solution. After 24 h, the reaction media was filtered with a 0.45 μm membrane filter and washed with 100 mL of water. The IL–water-insoluble residue obtained was dried in an oven at 105 °C overnight and weighed to calculate the yield.

Table 1 Yields of the IL–water-insoluble residue of C. japonica treated with IL–water mixtures at room temperature for 24 h

Ionic liquids	Yield of residue (wt%)
1-Ethyl-3-methylimidazolium acetate	95.6
1-Ethyl-3-methylimidazolium chloride	92.8
1-Ethyl-3-methylimidazolium hydrogensulfate	91.3
1-Ethyl-3-methylimidazolium methanesulfonate	92.8
1-Ethyl-3-methylimidazolium p-toluenesulfonate	93.0
1-Ethylpyridinium bromide	95.9
1-Ethylpyridinium chloride	91.1
1-Ethylpyridinium iodide	93.5
1-Methylimidazolium chloride	93.0
1-Methylimidazolium hydrogensulfate	91.3
Pyridinium chloride	87.6
Pyridinium p-toluenesulfonate	91.3
Tetrabutylammonium salicylate	93.0

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2.4 Treatment of C. japonica with 90% [Py]Cl and 10% water w/w solution

[Py]Cl (6.0 g) and distilled water (0.66 g) were added to a 100 mL round-bottom flask and stirred in an oil bath at 80 or 120 °C under reflux. A cedar powder from Section 2.1 (0.12 g) was added to the solution. At the specified treatment time, 20 μL of the reaction media was dissolved in 180 μL of water to be analyzed using HPLC equipped with an Aminex HPX-87H column. To recover the [Py]Cl–water-insoluble residues, the reaction media was filtered with a 0.45 μm membrane filter. The [Py]Cl–water-insoluble residue was washed with 100 mL of water and dried in an oven at 105 °C overnight to calculate the yield. All experiments were duplicated, and the average was calculated for the yield of the degradation products in the [Py]Cl–water-soluble fractions and the [Py]Cl–water-insoluble residues.

2.5 Klason lignin and sugar analysis of the residues

The lignin content in the [Py]Cl–water-insoluble residue from C. japonica treated with 90% [Py]Cl and 10% water w/w solution was determined as Klason lignin and acid-soluble lignin.^26,27 The total amount of cellulose and hemicellulose in the [Py]Cl–water-insoluble residues was calculated from these lignin contents. Moreover, the filtrate of the acid hydrolysates obtained from the [Py]Cl–water-insoluble residue in the Klason lignin method was measured for the amounts of the constituent monosaccharides in the [Py]Cl–water-insoluble residue using HPLC. A KS-801 sugar or Aminex HPX-87H column was used for the analysis of the filtrate of the acid hydrolysates from the [Py]Cl–water-insoluble residue obtained by treatments at 80 and 120 °C. The ratio of cellulose and hemicellulose in the [Py]Cl–water-insoluble residue was calculated from the amounts of constituent monosaccharides. All experiments were duplicated, and the average was calculated for the ratio and yield of three components in the [Py]Cl–water-insoluble residue.

2.6 Treatment of C. japonica with 90% [Py]Cl and 10% water w/w solution for light microscopy analysis

The extracted sample (5 × 5 × 5 mm) was cut into a 15 μm-thick section with a sliding microtome (TU-213, Yamato Kohki Industrial Co., Ltd., Saitama Japan), transferred to a 20 μm-deep hemocytometer (Sunlead Glass Corp., Saitama, Japan) and dried for 2 h at 105 °C. A 200 μL sample of 90% [Py]Cl and 10% water w/w solution was dropped onto the section. The hemocytometer was closed with a glass cover, placed in a separable flask filled with 100 mL of 90% [Py]Cl and 10% water w/w solution heated to 80 °C, and then the separable flask was closed with a separable cover equipped with a reflux condenser. The sample was heated to 80 °C for different durations, and the morphological changes in the section were observed using light microscopy (BH-2, Olympus, Tokyo, Japan). Three areas (cell lumen area, cell wall area and total of cell lumen area + cell wall area) were measured for five neighboring tracheids in both early- and latewood samples using image analysis software (Motic Image Plus 2.2S). The average was calculated for each area.

3. Results and discussion

3.1 Component analysis

In our previous study, furan compounds such as, 2-HAF, 5-HMF and furfural were produced from C. japonica by treating with pyridinium- and imidazolium-based IL–water mixtures at 120 °C for 1 h.²² We expected that pyridinium- and imidazolium-based IL–water mixtures can liquefy lignocellulosics at room temperature because they strongly decompose wood components, especially sugar components.

Thirteen 90% IL and 10% water w/w solutions were used for the liquefaction of C. japonica at room temperature for 24 h (Table 1). [Py]Cl can effectively liquefy over 10 wt% of wood, whereas over 90 wt% residues were recovered by the treatment with the other twelve IL–water mixtures. However, evaluating the [Py]Cl–water-insoluble residue obtained from the treatment of C. japonica with 90% [Py]Cl and 10% water w/w solution at room temperature was not possible because of insufficient liquefaction. Therefore, we conducted component analysis of C. japonica after treatments with 90% [Py]Cl and 10% water w/w solution at 80 and 120 °C.

Fig. 2 shows the changes in the chemical composition of the [Py]Cl–water-insoluble residue from C. japonica treated with 90% [Py]Cl and 10% water w/w solution at 80 and 120 °C. The values ware obtained from Klason lignin and acid-soluble lignin in the residue after filtration of the reaction mixture.²⁶ After 24 h treatment at 80 °C and 4 h treatment at 120 °C, the yields of the [Py]Cl–water-insoluble residues were 57 and 42 wt%, respectively. In addition, the changes in the component yields of the [Py]Cl–water-insoluble residue from C. japonica after treatment with 90% [Py]Cl and 10% water w/w solution at 80 and 120 °C are exhibited in Fig. 3.

Fig. 2 Product compositions of C. japonica treated with 90% [Py]Cl and 10% water w/w solution at (a) 80 °C and (b) 120 °C.

Fig. 3 Changes in the percentages of cellulose, hemicellulose and lignin in the [Py]Cl–water-insoluble residue from C. japonica treated with 90% [Py]Cl and 10% water w/w solution at (a) 80 °C and (b) 120 °C from the original amounts.

In the treatment at 80 °C, half of hemicellulose was liquefied within 1 h. Lignin was gradually solubilized in 90% [Py]Cl and 10% water w/w solution until 24 h and about 40 wt% of lignin remained in C. japonica. However, most cellulose was insoluble even after 24 h. In the treatment at 120 °C, most hemicellulose was liquefied, and all the hemicellulose was removed from C. japonica within 2 h. About half the lignin was rapidly liquefied at 20 min and another 10% of the lignin was gradually liquefied until 4 h. Cellulose was liquefied slower than hemicellulose and lignin. Most cellulose was not liquefied after 20 min of treatment at 120 °C, but 35% of cellulose was liquefied after 4 h of treatment. These results indicate that the [Py]Cl–water solution resembles 1-ethylpyridinium bromide without water addition in the reaction behavior of C. japonica.²⁷

3.2 Microscopic observation

We examined C. japonica in the transverse sections at 0 and 48 h of treatment with 90% [Py]Cl and 10% water w/w solution using light and polarized light microscopy (Fig. 4). In the treatment at 120 °C, the treated cells were deconstructed. Therefore, the comparison of the original cells with the treated cells was conducted at 80 °C.

Fig. 4 Light and polarized light microscopy images of transverse sections in C. japonica before (a–d) and after the treatment with 90% [Py]Cl and 10% water w/w solution for 48 h at 80 °C (e–f). (a), (b), (e) and (f) are magnified views of the earlywood. (c), (d), (g) and (h) are magnified views of the latewood.

In earlywood, significant morphological changes were not observed after 48 h (Fig. 4a and e). The cell structures after the treatment for 48 h were unchanged, although the cell walls in earlywood swelled at an early stage after the treatment. Cell walls in latewood were distorted after treatment for 48 h (Fig. 4c and g). Moreover, deconstructions of cells in latewood were observed in the treatment at 48 h (arrows in Fig. 4g).

In the polarized light microscopy images, the brightness from the birefringence of crystalline cellulose in early- and latewood was visible at 0 h (Fig. 4b and d), and they were unchanged at 48 h (Fig. 4f and h). These results indicate that the crystalline structures of cellulose in early- and latewood were preserved after the treatment with 90% [Py]Cl and 10% water w/w solution. These findings indicate the low level of liquefaction of cellulose as mentioned above.

To analyze the dissociations and distortions in early- and latewood treated with 90% [Py]Cl and 10% water w/w solution, we used image analysis software to examine the swelling behavior of cell walls in the transverse sections. The swelling degree of cell walls was evaluated by determining the cell lumen area, cell wall area and total of cell lumen area + cell wall area as defined in previous papers.^13,14

In earlywood, the cell lumen area decreased, whereas the cell wall area increased at an early stage. Therefore, the total of cell lumen area + cell wall area was unchanged at an early stage (Fig. 5). In addition, both areas in the cells were retained until 48 h, and the total of cell lumen area + cell wall area was also maintained. However, the cell wall area in latewood increased by approximately 1.5 times, although this area did not increase at an early stage. Thus, the total of cell lumen area + cell wall area increased. The differences in the morphological changes between early- and latewood by treating wood with 90% [Py]Cl and 10% water w/w solution are similar to that of a previous study using conventional ILs without water addition for wood treatment.^13,14

Fig. 5 Changes in the cell wall area, the cell lumen area and the total of the cell lumen area + the cell wall area during 90% [Py]Cl and 10% water w/w solution treatment at 80 °C.

3.3 Degradation products in the [Py]Cl–water-soluble fractions from C. japonica

The [Py]Cl–water-soluble fractions from C. japonica after treatment with 90% [Py]Cl and 10% water w/w solution at 80 and 120 °C were analyzed using HPLC. Table 2 shows the maximum yields of degradation products and the treatment times at the maximum yield. The monosaccharides are the total yield of glucose, mannose and arabinose.²⁷ Furan compounds and organic acids are converted from monosaccharides after hydrolysis of oligo- and polysaccharides in wood samples after treatment with ILs as previously described.^11,27 As for lignin degradation products in the [Py]Cl–water-soluble fractions from C. japonica, the yield of vanillin was less than 0.1% even after treatment at 120 °C (data not shown).

Table 2 Maximum yields of the degradation products in the [Py]Cl–water-soluble fractions from C. japonica treated with 90% [Py]Cl and 10% water w/w solution at 80 and 120 °C

Products	Treatment at 80 °C		Treatment at 120 °C
	Yield^a (%)	Time	Yield^a (%)	Time
a Per wood weight.
Monosaccharides	8.51	8 h	3.98	5 min
5-HMF	3.37	24 h	2.00	30 min
Furfural	0.74	24 h	1.32	30 min
2-HAF	0.53	24 h	2.69	30 min
Acetic acid	1.37	24 h	1.69	4 h
Levulinic acid	0.28	24 h	1.83	4 h

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In the treatment at 80 °C, the yield of monosaccharides at 8 h was 8.51%, which was the highest among the degradation products. The maximum yields of 5-HMF, furfural, 2-HAF, acetic acid and levulinic acid were recorded at 24 h. These results indicate that 80 °C is an insufficient temperature to complete the conversion of saccharides to furan compounds and organic acids. The treatment of C. japonica at 120 °C showed different results. Polysaccharides in C. japonica were rapidly hydrolyzed to monosaccharides, and they reached their maximum at 5 min. Furthermore, the monosaccharides obtained were rapidly transformed to furan compounds, and the yields of the furan compounds were maximum around 30 min. After 4 h treatment, acetic acid and levulinic acid reached their maximum yields.

5-HMF is stable in 90% [Py]Cl and 10% water w/w solution at 80 °C because levulinic acid, which is converted from 5-HMF, was produced in a low yield even after 24 h of treatment. However, 5-HMF was rapidly converted into levulinic acid in the treatment at 120 °C because the maximum yield of 5-HMF was reached at 30 min. These results show that the yields and treatment times for maximum yields of the degradation products from polysaccharides in C. japonica were remarkably different between 80 and 120 °C treatments with 90% [Py]Cl and 10% water w/w solution.

4. Conclusion

In this study, the reaction behavior of C. japonica with 90% [Py]Cl and 10% water w/w solution was investigated. This solution was chosen because it was the most effective IL–water mixture for liquefaction of C. japonica from thirteen 90% ILs and 10% water w/w solutions at room temperature as listed in Table 1. Hemicellulose was liquefied, followed by the liquefaction of lignin, whereas most of the cellulose was insoluble in the treatments of C. japonica with 90% [Py]Cl and 10% water w/w solution at 80 and 120 °C. In the microscopic analyses, the crystalline structure of cellulose in C. japonica was retained after the treatment for 48 h at 80 °C. A part of the cells in latewood was deconstructed, although significant morphological changes in earlywood were not observed. Degradation products were detected in the [Py]Cl–water-soluble fractions after the treatments of C. japonica with 90% [Py]Cl and 10% water w/w solution at 80 and 120 °C. The maximum yields and treatment times at the maximum yield of the degradation products are different between 80 and 120 °C treatments.

We conclude that the 90% [Py]Cl and 10% water w/w solution is exceedingly effective for the treatment of lignocellulosics, such as liquefaction of lignocellulosics and the production of useful low molecular weight compounds.

Acknowledgements

This study was supported by the Science and Technology Research Promotion Program for Agriculture, Forestry, Fisheries and Food Industry (No. 26052A) from the Ministry of Agriculture, Forestry and Fisheries of Japan.

Notes and references

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