Characterization of lignin isolated from some nonwood available in Bangladesh

Bioresource Technology 98 (2007) 465–469

Short Communication

Characterization of lignin isolated from some nonwood available in Bangladesh M. Sarwar Jahan a

a,¤

, D.A. Nasima Chowdhury a, M. Khalidul Islam a, S.M. Iqbal Moeiz

b

Pulp and Paper Research Division, BCSIR Laboratories, Dhaka, Dr. Qudrat-E-Khuda Road, Dhaka 1205, Bangladesh b Department of Chemistry, Dhaka College, Dhaka 1205, Bangladesh Received 27 September 2005; received in revised form 3 January 2006; accepted 4 January 2006 Available online 9 March 2006

Abstract Lignins isolated from cotton stalks, jute stick and dhaincha by acidolytic dioxane were characterized using alkaline nitrobenzene oxidation, elemental analysis, methoxyl analysis and molecular weight analysis and UV, IR 1H NMR spectroscopy. The C9 formulas for cotton stalks, jute stick and dhaincha (Sesbania aculeata) lignin were C9H9.36O4.50(OCH3)1.23, C9H9.02O4.57(OCH3)1.35 and C9H8.88O4.65(OCH3)1.50, respectively. All three lignins were of the guaiacyl–syringyl type. Cotton stalks lignin contained more p-hydroxy phenyl unit than dhaincha and jute stick lignins as observed by alkaline nitrobenzene oxidation products. The -O-4 units in these nonwood lignins had predominately erythro stereochemistry type. © 2006 Elsevier Ltd. All rights reserved. Keywords: Nonwood; Syringyl unit; Guaiacyl unit; -O-4 Structure; erythro; Cotton stalks; Jute stick; Dhaincha

1. Introduction Over a last few years, forest preservation and rational use of forest and agricultural residues has received attention. The world consumption of paper, especially Wne paper is increasing, which increases demand for short Wber. Traditionally, hardwood meets the demand for short Wber of Wne paper production. The wood is being gradually replaced by nonwood. In many countries, including Bangladesh, the wood supplies available will not continue to meet the rising demand for Wber. Hence, there is to Wnd out alternative sources. Research on jute, cotton stalks and dhaincha shows that these could serve as potential sources of Wbers for papermaking. Cotton stalks is an agricultural waste, available to the extent of over 36 thousands ton per annum in Bangladesh. The stalks are rich in cellulose and close to the Wbrous structure of hardwoods. Many studies (Pandey and Shaikh, 1987; Ali et al., 2001) have showed that good quality writ*

Corresponding author. E-mail address: [email protected] (M.S. Jahan).

0960-8524/$ - see front matter © 2006 Elsevier Ltd. All rights reserved. doi:10.1016/j.biortech.2006.01.005

ing and printing paper could be produced from cotton stalks. Jute has a long historical role in the socio-economic development of Bangladesh. Once jute was known as golden Wber of Bangladesh. The export of jute and related products accounts for a signiWcant portion of total export. Jute Wber is obtained from the bark of the plant after retting whole plant and the woody residue remained as jute stick. Jute stick is used for fencing in the rural area. If jute stick could be used for making value-added products such as pulping source, farmers would be beneWted. Many studies were done on jute pulping (Jahan, 2001; Akhtaruzzaman and ShaW, 1995). Dhaincha (Sesbania aculeata) is also grown all over the Bangladesh. It is a nitrogen Wxing plant. Presently, it is used as domestic fuel and for fencing. It is characterized by short and slender thin-walled Wbers. Its chemical characteristics are comparable to hardwoods (Jahan, 2004). Dhaincha pulp also showed excellent strength properties (Jackson et al., 1997). In spite of the abundance of published work on the technological aspects of the nonwood pulping and papermaking, published works on the chemistry of nonwood materials are scarce. This work reports the isolation of

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lignin from cotton stalks, jute stick and dhaincha by acidic dioxane solution. The isolated lignin was characterized by alkaline nitrobenzene oxidation, elemental analysis, methoxyl analysis, molecular weight analysis and UV, IR, FTIR and 1H NMR spectroscopy. 2. Experimental 2.1. Isolation of lignin Cotton stalks, jute stick and dhaincha meals (40–60 mesh) were extracted with alcohol–benzene solvent. The alcohol–benzene extract free wood meals were then reXuxed with acidic dioxane (9:1) solution for one hour in nitrogen atmosphere (N2 Xow was 50 mm/min). The concentration of HCl in dioxane solution was 0.2 N by using concentration HCl. The dioxane to wood meal ratio was 8. After this, wood meal-dioxane mixture was Wltered in Buckner funnel using Wlter no. 2. The residue was washed with dioxane solution. The Wltrate was concentrated in vacuum evaporator at 40 °C. Then conc. dioxane solution was added drop-wise to ion exchange water to precipitate lignin. Precipitate was centrifuged and washed till neutrality and dried. Dried crude lignin was dissolved in dioxane (9:1), and again precipitated in ether with constant stirring by magnetic bar. The precipitated pure lignin was dried under vacuum over P2O5 and weighed. 2.2. Alkaline nitrobenzene oxidation Alkaline nitrobenzene oxidation of cotton stalks, jute stick and dhaincha was carried out according to Mun’s modiWed method (Mun and Wi, 1991). GC analysis was conducted using a Shimatzu GC 17A gas chromatograph, equipped with Neutrabond 1 capillary column (30 m £ 0.53 mm). Conditions used were as follows: column temperature was programmed to increase from 150 to 250 °C at the rate of 5 °C/ min; injection and detection temperature were 220 and 250 °C, respectively; column Xow rate was 6 ml/min and split ratio 30. 2.3. Acetylation of lignin PuriWed lignin (100 mg) was added in 1.5 ml of dry pyridine-acetic anhydride (1:1) for 72 h. The solution was added to a 10-fold volume of ice-cold water whereupon the acetylated sample was recovered as a precipitate, which was puriWed by successive washing with water and dried under vacuum over P2O5. 2.4. Elemental analysis C and H analyses of cotton stalks, jute stick and dhaincha lignin were carried out using C, H analyzer and the oxygen was determined by diVerence. The methoxyl content in lignins was determined in accordance to Japan International Standard Methods (JIS P8013 1972).

2.5. Molecular weight The weight average (Mw) and number average (Mn) molecular weight of cotton stalks, jute stick and dhaincha acetylated lignins were determined by GPC on a Sodex KF-802.5 column. The samples were dissolved in tetrahydrofuran (THF) and 10 l was injected to the column. The column was operated at 30 °C and eluted with THF at a Xow rate of 1 ml/min. The column was calibrated using polystyrene standards. 2.6. Spectroscopy Ultraviolet: Cotton stalks, jute stick and dhaincha lignins (7–8 mg) was dissolved in 100 ml dioxane (9:1) followed by two times dilution. Then spectra were recorded using Hewlett Packard 8452A spectrophotometer. FTIR: Infra red spectra were recorded by using a Shimadzu FTIR spectrometer model 8201PC. The dried samples were embedded in KBr pellets in the concentration of about 1 mg/100 mg KBr. The spectra were recorded in the absorption band mode in the range 4000–400 cm¡1. 1 H NMR: Spectra of lignin solution (100 mg of acetylated lignin contained in 0.5 ml CDCl3) were recorded in a Bruker 400 spectrometer. Solvent was used as internal standard (7.25 ppm). For quantiWcation of protons, the signal in speciWed regions of the spectrum were integrated with respect to a spectrum-wide baseline drawn at the level of the background noise, and the results were referred to the signal for methoxyl protons, whose average number per C9 unit was established as described above. 3. Results and discussion 3.1. Elemental analysis and C9 formula The average C9 formula was calculated from the elemental analysis and methoxyl content. The number of methoxyl groups per C9 unit in cotton stalks, jute stick and dhaincha lignin were 1.23, 1.35 and 1.50. Islam and Sarkanen (1993) found methoxyl content per C9 unit of jute stick MWL 1.34, which was almost similar to dioxane lignin from jute stick. But the OCH3/C9 of enzyme-liberated lignin from jute stick was 1.41. Therefore, in this experiment dioxane lignin was considered as pure lignin like MWL. Jahan (2004) found that the tropical hardwood lignin contained lower methoxyl group (1.30–1.36/C9 unit) than that of temperate hardwood lignin (1.50/C9). The methoxyl content in nonwood lignin varied within the range of temperate to tropical hardwood lignin. The C9 formula for cotton stalks was C9H9.36O4.50(OCH3)1.23, whereas it was C9H9.02O4.57(OCH3)1.35 for jute stick and C9H8.88O4.65(OCH3)1.50 for dhaincha (results not shown). 3.2. Alkaline nitrobenzene oxidation The standard procedures for analyzing lignin by chemical degradative techniques such as alkaline nitrobenzene

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467

Table 1 Assignment of FTIR spectra of lignin from cotton stalks, jute stick and dhaincha Peak location range (cm¡1)

Assignment

Dhaincha

Jute stick

3412–3460 3000–2842 1738–1709 1675–1655 1593–1605 1505–1515 1460–1470 1422–1430 1365–1370 1325–1330 1266–1270 1221–1230 1166 1140 1125–1128 1086 1030–1035

O–H stretching C–H stretch in methyl and methylene group CBO stretch in unconjugated ketone, carbonyl and ester groups CBO stretching in conjugated p-subst. Aryl ketones Aromatic skeleton vibrations plus CBO stretching; S > G: Gcondensed > GetheriWed Aromatic skeleton vibrations (G > S) C–H deformations (asym in –CH3 and –CH2–) Aromatic skeleton vibrations combined with C–H in plane deformations Aliphatic C–H stretching in CH3 and phen. OH Condensed S and G ring (G ring bound via position 5) G ring plus C+O stretching C–C + C–O + CBO stretching (Gcondensed > GetheriWed) Typical for HGS lignins; CBO in ester groups (conj.) Aromatic C–H in-plane deformation (typical of G unit; Gcondensed > GetheriWed) Typical of S unit; also secondary alcohol and CBO strt. C–O deformation in sec. alcohol and aliphatic ether Aromatic C–H in-plane deformation (G > S) plus C–O deform. in primary alcohols plus C–H stretching (unconjugated) –HCBCH– out of plane deformation (trans) C–H out of plane (aromatic ring) C–H out of plane in positions 2, 5 and 6 (G units) C–H out of plane in positions 2 and 6 (S units) C–H out of plane in positions 2, 5 and 6 (G units)

3425.9 2854.5 1724.2 – 1595.0 1508.2 1460.0 1421.4 – 1328.9 – 1224.7

3425.3 2939.3

3423.1 2922.0 1712.7

1595.0 1510.2 1460.0 1423.0

11596.9 1510.2 1458.1 1423.4 –

1222.8

1222.8

1120.6

1120.6

1120.6

1031.6

1033.8

1033.8

833.5

833.4

833.5

966–990 915–925 853–858 834–835 817–832

oxidation can be used to obtain information about the composition of the original lignin polymer. In the case of alkaline nitrobenzene oxidation, the three constitutive monomeric lignin units p-hydroxyphenyl, guaiacyl and syringyl produce the corresponding p-hydroxybenzaldehyde, vanillin and syringaldehyde. Alkaline nitrobenzene oxidation products showed that the syringyldehyde (S) was the predominant product, which comprised 20.2 § 1.9% in cotton stalks, 25.6 § 2.1% in jute stick and 19.3 § 1.8% in dhaincha lignin (results not shown). Vanillin (V) appeared as the second major degradation products, resulting from the noncondensed guaiacyl unit. Vanillin percentage varied from 9% to 17% in these nonwood lignins. The relative ratio of S to V was 1.2 for cotton stalks, 1.6 for jute stick and 2.1 for dhaincha. Islam and Sarkanen (1993) obtained S/V ratio 1.4 for jute stick. Jahan et al. (2006) found S/V ratio of golpata lignin 1.4. Generally, S/V ratio of temperate hardwood is higher than that of tropical hardwood (Sarkanen and Hergert, 1971). The cotton stalks gave considerable amount of p-hydroxybenzaldehyde (2.6 § 0.4%), while it was a minor amount in dhaincha (0.5 § 0.1%) (results not shown). 3.3. UV spectroscopy The cotton stalks, jute stick and dhaincha lignin had welldeWned maxima at 282 nm. The absorptivities of lignin were 12.0 for cotton stalks, 14.4 for jute stick and 14.7 l g¡1 cm¡1 for dhaincha (results not shown). Normally temperate zone hardwood lignin shows absorption coeYcient as high as 16 (Goldschmid, 1971). The shoulder at 310–320 nm was observed in all these lignins, which is characteristic of – unsaturated and -carbonyl groups. Shoulder at 320 nm in

Cotton stalks

1328.9 –

cotton stalk lignin was stronger than jute stick and dhaincha lignin. This result was consistent with the results of nitrobenzene oxidation products (results not shown). 3.4. FTIR spectra To elucidate the structure of lignin, and to investigate the diVerences in the structure of the dioxane lignin isolated from cotton stalks, jute stick and dhaincha, FTIR spectra were recorded and the assignment made by Faix (1991) were as given in Table 1. The band at 1710 cm¡1 was assigned to the carbonyl stretching-unconjugated ketones and carbonyl group was observed in cotton stalks, jute stick and dhaincha. The band at 1600 cm¡1 was assigned to the aromatic skeletal vibrations, 1510 cm¡1 assigned to the aromatic skeletal vibrations coupled with C–H in plane deformations, 1460 cm¡1 assigned to C–H deformations (asymmetric in methyl, methylene and methoxyl group). All spectra were showing absorbance near 1330 cm¡1 (syringyl), which was typical for hardwood lignin and shoulder at 1270 cm¡1 (guaiacyl). But the intensity of absorbance at 1330 cm¡1 was lower in case of cotton stalks, which suggested of lower syringyl unit than dhaincha and Table 2 The weight average (Mw) and number average (Mn) molecular weight and polydispersity (Mw/Mn) of dioxane lignin isolated from cotton stalks, jute stick and dhaincha Sample

Mw

Mn

Mw/Mn

Cotton stalks Jute stick Dhaincha

15,453 18,978 18,654

3263 3435 3325

4.7 5.5 5.6

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Table 3 Assignments of signals and protons per C9 structural unit in the 1H NMR spectra of acetylated lignins of cotton stalks, jute stick and dhaincha Range ppm

Main assignments

Proton per C9 unit Cotton stalks

7.25–6.80 6.80–6.25 6.25–5.75 5.75–5.24 5.20–4.90 4.90–4.30 4.30–4.00 4.00–3.48 2.50–2.22 2.22–1.60

Aromatic proton in guaiacyl units Aromatic proton in syringyl units H of -O-4 and -1 structures H of -5 structure H of xylan residue H and H of -O-4 structures H of – structures H of xylan residue H of methoxyl groups H of aromatic acetates H of aliphatic acetates

Total proton per C9 structural unit

jute stick lignin. The strong intensities of the band at 1122 cm¡1 were associated with syringyl structure of the lignin molecule present in cotton stalks, jute stick and dhaincha. The bands at 1225, 1034 and shoulder at 1156 cm¡1 was associated with guaiacyl units in lignin molecules, which indicated the presence of both guaiacyl and syringyl unit in the lignin molecule. 3.5. Molecular weight The weight average (Mw) and number average (Mn) molecular weight, and polydispersity of cotton stalks, jute stick and dhaincha lignin were computed from their chromatograms (Table 2). The data showed that the cotton stalks lignin had Mw 15,453, which was lower than dhaincha and jute stick lignin. The highest Mw reported for analytical lignin were 77,000 for an enzymatically isolated MWL of Eastern hemlock (Tsuga canadensis), and 85,000 for a dioxane spruce lignin fraction (Fengel and Wegener, 1984). The polydispersity of cotton stalks, jute stick and dhaincha lignin was 5.6, 5.5 and 4.7, respectively.

Jute stick

0.97 0.74 0.35 0.56 1.53 –

Dhaincha

0.69 0.73 0.30 0.30

0.64 0.86 0.33 0.25

1.22 –

1.51 –

3.69 1.27 4.47

4.05 0.78 3.47

4.50 0.96 3.75

13.06

13.77

13.04

syringyl units ( 6.6). Dhaincha and jute stick lignins showed stronger peak in syringyl units region ( 6.6) than guaiacyl units region ( 6.9). But cotton stalks lignin had higher guaiacyl peak than syringyl peak. NMR integration showed that cotton stalks lignin contained 0.97 + 074 D 1.71 aromatic proton per C9 units, while these were 0.69 + 0.73 D 1.42 for jute stick 0.64 + 0.86 D 1.50 for dhaincha lignin, respectively (Table 3). This result was in good agreement with the results of alkaline nitrobenzene oxidation products. -O-4 Structure: The aryl glycerol -O-4 aryl ether linkage constituted the main intermonomeric connection in lignin. NMR spectra of cotton stalks, jute stick and dhaincha lignin showed that the structural element might contain both erythro and threo conWgurations due to the presence of proton at the C- position of the side chain. The erythro protons (H) have stronger peak at  6.01 than the peak for thero form at  6.09 in cotton stalks, jute stick and dhaincha lignin. It has been reported that the angiosperms lignin contains higher erythro form in -O-4 units than thero form (Akiyama et al., 2003). 4. Conclusions

1

3.6. H NMR Table 3 lists the position of signal assigned by Lundquist (1992) and their proton number per C9 unit calculated from the integration of NMR spectra. Hydroxyl group: The number of free aliphatic and phenolic hydroxyl groups per C9 unit were determined from the corresponding acetate signals. The number of proton per C9 unit of phenolic hydroxyl group of cotton stalks, jute stick and dhaincha lignin were 0.42, 0.26 and 0.32, respectively. The proton of aliphatic hydroxyl group per C9 unit was 1.49, 1.16 and 1.25 for cotton stalks, jute stick and dhaincha, respectively. This result was consistent with the result of molecular weight (Table 2). It would expect less aromatic and aliphatic OH for high molecular weight lignin. Aromatic protons: Cotton stalks, jute stick and dhaincha lignin spectra showed two peaks in the aromatic proton region, which corresponded to guaiacyl units ( 6.9) and

It was concluded that dioxane lignin extracted from nonwood, viz. cotton stalks, jute stick and dhaincha was syringyl–guaiacyl type. The methoxyl content of these lignins was similar to hardwood lignin. The weight average molecular weight (Mw) was comparable to temperate hardwood and the -O-4 bond in cotton stalks, jute stick and dhaincha lignin was erythro-ether type. References Akhtaruzzaman, A.F.M., ShaW, M., 1995. Pulping of jute. Tappi J. 78 (2), 106–112. Akiyama, T., Matsumoto, Y., Okuyama, T., Meshitsuka, G., 2003. Ratio of erythro and thero form of -O-4 structure in tension wood lignin. Phytochemistry 64, 1157–1162. Ali, M., Byrd, M.M., Jameel, H., 2001. Soda-AQ pulping of cotton stalks. In: Tappi Pulping Conference Proceedings (CD version). Faix, O., 1991. ClassiWcation of lignins from diVerent botanical origins by FTIR spectroscopy. Holzforschung 45 (Suppl.), 21–27.

M.S. Jahan et al. / Bioresource Technology 98 (2007) 465–469 Fengel, D., Wegener, G., 1984. Wood: Chemistry, Ultrastructure, Reaction. Walta de Gruyter, Berlin, pp. 150. Goldschmid, 1971. Ultraviolet spectra. In: Sarkaned, K.V., Ludwig, C.H. (Eds.), Lignins: Occurrence, Formation, Structure and Reactions. Wiley Interscience, New York, pp. 241–266. Islam, A., Sarkanen, K.V., 1993. The isolation and characterization of the lignin of jute (Corchorus capsularis). Holzforschung 47, 123–132. Jahan, M.S., 2001. Evaluation of additive in soda pulping of jute. Tappi 84 (8), 1–11. Jackson, M., Markila, L., Calmell, R. Mokvist, A., 1997. Sesbania bispinosa—a new source of raw material for high yield pulping. In: 1997 Pulping Conference Proceedings, pp. 1–15. Jahan, M.S., 2004. Characteristics of milled wood lignins isolated from diVerent ages of Nalita Wood (Trema orientalis). Post-doctoral report. Chonbuk National university, Chonju, Korea.

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