Carbohydrate Research 346 (2011) 76–85 Contents lists available at ScienceDirect Carbohydrate Research journal homepage: www.elsevier.com/locate/carres Isolation and characterization of cellulose nanofibrils from wheat straw using steam explosion coupled with high shear homogenization Anupama Kaushik a,⇑, Mandeep Singh b a b University Institute of Chemical Engineering and Technology, Panjab University, Chandigarh, India Centre for Emerging Areas in Science & Technology, Panjab University, Chandigarh, India a r t i c l e i n f o Article history: Received 17 July 2010 Received in revised form 20 October 2010 Accepted 26 October 2010 Available online 30 October 2010 Keywords: Wheat straw Cellulose nanofibers High shear homogenizer Steam explosion AFM Thermal degradation a b s t r a c t Cellulose nanofibrils of diameter 10–50 nm were obtained from wheat straw using alkali steam explosion coupled with high shear homogenization High shear results in shearing of the fiber agglomerates resulting in uniformly dispersed nanofibrils The chemical composition of fibers at different stages were analyzed according to the ASTM standards and showed increase in a-cellulose content and decrease in lignin and hemicellulose Structural analysis of steam exploded fibers was carried out by Fourier Transform Infrared (FT-IR) spectroscopy and X-ray diffraction (XRD) Thermal stability was higher for cellulose nanofibrils as compared to wheat straw and chemically treated fibers The fiber diameter distribution was obtained using image analysis software Characterization of the fibers by AFM, TEM, and SEM showed that fiber diameter decreases with treatment and final nanofibril size was 10–15 nm FT-IR, XRD, and TGA studies confirmed the removal of hemicellulose and lignin during the chemical treatment process Ó 2010 Elsevier Ltd All rights reserved Introduction Natural fibers are abundantly present in plants such as grasses, reeds, stalks, and woody vegetation They are also referred to as cellulosic fibers due to the main chemical component cellulose, or as lignocellulosic fibers, since the fibers usually contain a natural polyphenolic polymer, lignin, in their structure These fibers are long regarded as being promising candidates for replacing conventional reinforcing fibers (e.g glass fibers) in composites for semi-structural and even structural applications The biodegradable nature of plant fibres can contribute to the formation of a healthy eco-system as well as their high performance fulfills the economic interest of industries Plant based natural fibres like sisal,1 jute,2 bamboo,3 wood4,5, and paper in their natural condition, as well as several waste cellulose products, such as shell flour, wood flour, and pulp, can be used as reinforcement materials for different thermosetting and thermoplastic polymeric matrices Cellulose nanofibers have gained importance due to their unique characteristics such as very large surface to volume ratio, high surface area, good mechanical properties including a high Young’s modulus, high tensile strength6,7 and a very low coefficient of thermal expansion,8 and formation of highly porous mesh as compared to other commercial fibers Functional hydroxyl groups in cellulose also enable chemical modifications for further applications ⇑ Corresponding author Tel.: +91 0172 2534925; Mob.: +91 9815177772 E-mail addresses: anupamasharma@pu.ac.in, anupamachem@gmail.com (A Kaushik), smandeep.virk@gmail.com (M Singh) 0008-6215/$ - see front matter Ó 2010 Elsevier Ltd All rights reserved doi:10.1016/j.carres.2010.10.020 Biocompatibility, non-toxicity, and biodegradability of cellulose nanomaterials are important properties in biochemical and biomedical applications All these features make cellulose microfibrils a very promising material for nanotechnology Plant cell walls usually consist of rigid cellulosic microfibrils embedded in soft hemicelluloses and lignin matrix It is the structural material of the fibrous cells with high levels of strength and stiffness per unit weight Cellulose has a straight carbohydrate polymer chain consisting of b-(1?4)-linked glucopyranose units and a degree of polymerization (DP) of about 10,000.9 Hydroxyl (–OH) groups in cellulose structures play a major role in governing its reactivity and physical properties Several groups have reported methods for preparing cellulosic microfibrils.10–16 These materials, however, are rather non-homogeneous and in addition to microfibrils contain larger fibril bundles and residual fiber fragments Steam explosion is one of the ways of fibril isolation, which is to be subsequently used in chemical fractionation and biotechnological conversion The steam explosion process was first introduced by Mason in 1927 to defibrate wood into fiber for broad production High pressure steaming followed by rapid decompression is called steam explosion The steam explosion process includes saturating the dry material with steam at elevated pressure and temperature followed by sudden release of pressure, during which the flash evaporation of water exerts a thermo mechanical force causing the material to rupture.17 Steam explosion in alkaline medium results in the hydrolysis of hemicellulose within the fiber, and the resulting sugars can be subsequently washed out in water, leaving a residue of R-cellulose and lignin.18 It also leads to a cleavage of A Kaushik, M Singh / Carbohydrate Research 346 (2011) 76–85 hemicellulose–lignin bonds The reaction results in an increased water solubilization of hemicelluloses and in an increased solubility of lignin in alkaline or organic solvents, leaving the cellulose as a solid residue with a reduced degree of polymerization.17 The advantages of steam explosion include a significantly lower environmental impact, low energy consumption, lower capital investment, and less hazardous process chemicals In this study, an effort was made to isolate cellulose nanofibrils from wheat straw using steam treatment with subsequent explosive defibrillation Steam explosion in alkaline medium followed by hydrochloric acid treatment and high shear homogenization was found to be effective in the depolymerization and defibrillation of the fiber to produce cellulose nanofibrils With this method, the substrate, that is, wheat straw, is loaded into a pressure vessel and heated by steam injection for a defined time-temperature period At the end of this period of heat treatment, the pressure drop is suddenly made A rapidly opening valve was used so that, after treatment, the contents of the reaction vessel were almost instantaneously depressurized and discharged through a nozzle Alkaline medium has been used for the explosive treatment This is followed by bleaching and acidic treatment as well as high shear homogenization leading to breaking of agglomerates These cellulose nanofibers obtained were characterized by AFM, TEM, SEM, FT-IR, XRD, and TGA Experimental 2.1 Material Wheat straws were obtained from neighboring fields They were thoroughly washed to remove any extraneous impurities and dried before use This is extremely low cost material, which is used for cattle feed Other chemicals used in the experiments, sodium hydroxide (NaOH), hydrochloric acid (HCl) and hydrogen peroxide (H2O2) were supplied by Merck India Pvt Ltd 2.2 Extraction of cellulose nanofibrils from wheat straw 2.2.1 Chemical treatment 2.2.1.1 Preparation of steam exploded fibers The preparation was done in two steps First, wheat straw fibers of length around 2–5 cm were soaked in 2% solution of NaOH for overnight and then treated in 10–12% NaOH solution (solid to liquid ratio around 1:8) in an autoclave at pressure around 20 bars for h at 200 ± °C The first treatment removed excessive impurities from the surface of the fibers and resulted in swelling of fibers thereby making further treatments easy Second, treatment removed excessive amount of lignin from the fibers The obtained wheat straw pulp was then washed several times in distilled water till it was free of alkali 2.2.1.2 Preparation of steam exploded bleached fibers Alkaline treated pulp was then soaked in 8% solution of H2O2 (v/v) and kept overnight to remove any residual lignin and hemi cellulose that may have been present 2.2.1.3 Treatment of steam exploded bleached banana fibers in acidic medium Bleached pulp was treated with 10% HCl (1 N) solution and kept in Branson 2510 E-DTH ultrasonicator at temperature around 60 ± °C for h Finally the fibers were taken out and washed several times with distilled water in order to neutralize the final pH and then dried Hydrochloric acid has been used for hydrolysis instead of sulfuric acid, which is a common choice as it assists in dispersion and separation of nanofibrils due to introduction of sulfate ester groups randomly on the surface resulting 77 in nonflocculating suspensions For composite applications, the sulfate groups are problematic due to decreased thermal stability after drying, precluding typical polymer melt processing Battista and Smith19 discovered that formation of stable suspensions can also be achieved by hydrolysis of cellulose using hydrochloric acid followed by mechanical disintegration 2.2.2 High shear treatment of chemically treated fibers Finally, the fibers were suspended in water and continuously stirred with a Fluko FA25 high shear homogenizer for 15 The high shearing action breaks down the fiber agglomerates and results in nanofibrils 2.3 Characterization of cellulose nanofibrils 2.3.1 Morphology of cellulose nanofibrils AFM imaging was used to characterize the dimensions and homogeneity of cellulose nanofibrils obtained after chemical and mechanical treatment AFM was done in tapping mode wherein the oscillation frequency was constant and the changes in amplitude monitored During each oscillation cycle the tip is briefly in contact with the surface and the interaction forces between the sample and the tip cause a reduction in amplitude The images were scanned in tapping mode in air using silicon cantilevers (Bioscope II AFM, VEECO) and the drive frequency of the cantilever was about 200–300 kHz with scan rate of 0.5–3 Hz (usually around Hz) The sample size taken is 10 lm  10 lm (Fig 2a), 1.5 lm  1.5 lm (Fig 2b), lm  lm (Fig 2c), and lm  lm (Fig 2d) while the z scale is 100 nm No image processing except flattening was made Samples were fixed on metal discs with double-sided adhesive tape To eliminate external vibration noise, the microscope was placed on an active vibrationdamping table The diameter and length of cellulose nanofibers (CNFs) extracted from wheat straw were examined by using transmission electron microscope (TEM) model Hitachi-2100 Images were taken at 80 kV accelerating voltage A drop of a dilute aqueous cellulose nanofiber suspension was deposited on the carbon-coated grids and allowed to dry at room temperature Scanning electron microscope model JSM JEOL-6490 was used for microstructural analysis of cellulose microfibers obtained after steam explosion Samples were mounted on a metal stub and platinum coated by using sputter coating technique for 20 s to make them conducting Images of fibers were taken at 20 kV accelerating voltage at different magnifications 2.3.2 Chemical characterization of the nanofibers Chemical composition of fibers was estimated according to the following ASTM procedures: a-cellulose (ASTM D1103-55T), lignin (ASTM D1106-56) and holocellulose (ASTM D1104-56) The standard deviations were calculated by conducting several replicate measurements for each sample The a-cellulose, hemicelluloses and lignin content were calculated by Eqs 1–3 as follows: weight of oven dry a À cellulose residue  100 ð1Þ W ÂP    weight of oven dry holocellulose residue  100 À A Hemicellulose percentage ¼ W ÂP a-cellulose percentage ðAÞ ¼ ð2Þ weight of oven dry lignin residue Lignin percentage ¼  100 W ÂP ð3Þ where W is the weight of the original oven dry fiber sample, and P is the proportion of moisture-free content 78 A Kaushik, M Singh / Carbohydrate Research 346 (2011) 76–85 2.3.3 Fourier Transform Infrared (FTIR) spectroscopy FT-IR analysis of raw as well as chemically treated wheat straw fibers was done in order to obtain composition of the fibers before and after treatment A Perkin Elmer RX infrared spectrophotometer was used to obtain spectra Fibers were ground and mixed with KBr (sample/KBr ratio, 1/99) to prepare pastilles FT-IR spectra were recorded in a spectral range of 4000–450 cmÀ1 with a resolution of cmÀ1, taking four scans for each sample 2.3.4 X-ray diffraction X-ray diffraction (WAXRD) profiles of wheat straw fibers before and after chemical process were collected in order to examine the crystallinity of the samples The samples were taken in powdered form and analyzed by using a Philips X’Pert Pro X-ray diffractometer system The radiation was Cu Ka (k = 1.54060 Å) with 40 kV voltage and 40 mA intensity Crystallinity of cellulose in pulp samples was calculated from diffraction intensity data The major diffraction planes of cellulose  1, 0 2, and are present at 14.8°, 16.7°, namely 1, 1, 20.7°, 22.5°, and 34.6° 2h angle, respectively.20 The crystallinity index was obtained using the Eq 421: Crystallinity index ¼ 100  I0 À IAmorph I0 ð4Þ where I0 is the maximum intensity of the (0 2) lattice diffraction and IAmorph is the intensity diffraction at 18° 2h degrees The calculation of the X-ray crystallinity order index was also performed using Eq The crystallinity index was calculated from the fraction of the ratio of the (0 2) to the sum of (1 1), (0 1) and (0 2) refraction areas: Crystallinity order index ¼ A0 A1 1 þ A1 þ A0 ð5Þ The Eqs and were used for the amorphous fraction in terms of intensity and area to calculate the crystallinity index The selected position that was assigned to the amorphous fraction was the diffraction angle at around 18° 2h degree The crystallite size or the thickness of crystal in a direction perpendicular to its Miller plane was estimated using the Scherrer equation This is a method based on the width of the diffraction pattern in the X-ray reflected crystalline region t hkl ¼ Kk bhkl cos h ð6Þ In Eq 6, thkl is the thickness of crystallites at the (hkl) plane of diffraction, k is an X-ray wavelength (k = 0.1542 nm for Cu Ka), h is the Bragg angle of the reflection, bhkl is the pure integral of width of the reflection at half maximum height, and K is the Scherrer constant that falls in the range 0.87–1.0.22 The crystallite size of cellulose nanofibrils was determined by using the diffraction pattern obtained from 0 lattice plane 2.3.5 Thermal characterization Thermogravimetric analysis was undertaken to compare the degradation characteristics of the chemically treated fibers with the untreated ones The thermal stability of each sample was determined using a thermogravimetric analyzer (TGA) of type PERKIN ELMER STA-6000 It has weighing capacity up to 1500 mg with resolution around 0.1 lg Samples were taken in a very small quantity (in milligram) in a sample cup made of alumina and having maximum capacity around 180 lL All the tests were performed in nitrogen environment and at heating rate of 10 °C/min from room temperature to 600 °C 2.3.6 Fiber diameter measurements Fiber diameter measurements of the wheat straw fibers after chemical and mechanical treatment were undertaken using a UTHSCSA Image Tool image analyzer program (IT version 3) TEM images of the cellulose nanofibers were used to measure the diameters The images were loaded into the software and diameters of the fibers were measured using a two point measuring analysis The scale of the software was calibrated using the scale bars on each TEM image Approximately, 300 measurements were taken to obtain each fiber diameter distribution 2.3.7 Degree of polymerization Viscosity of the cellulosic preparations after steam explosion, bleaching, and acid treatment was determined by British Standard Method for determination of limiting viscosity number of cellulose in dilute solution: Part 1: Cupric–ethylenediamine method (BS 6306: Part 1: 1982) The viscosity average Degree of Polymerization (DOP) of the cellulose samples was estimated by DOP0.90 = 1.65 [g] Molecular weight of the cellulosic preparations was then calculated from their DOP by multiplying with 162, the molecular weight of an anhydroglucose.23 Results and discussion 3.1 Morphology and chemical characterization The alkaline steam explosion results in structural as well as chemical changes on fiber surfaces SEM pictures of the wheat straw after steam explosion were taken to investigate the structure of these fibers The SEM micrographs are shown in Figure 1a–d These clearly show individual fibers after the removal of hemicelluloses, lignin, and pectin after chemical treatment, which are the cementing materials around the fiber-bundles It is clear from the pictures that the average diameters of the fibers are about 10– 15 lm, which is lower than the average size of fiber bundles before chemical treatment Reduction in particle size because of the dissolution of the hemicelluloses and lignin is clearly supported by chemical analysis and FT-IR data as given in Tables and Figure shows the phase images of AFM of cellulose nanofibrils at different magnifications Figure 1a and b are phase images and Figure 1c and d are topographical images of the nanofibers extracted The results reveal that fiber diameter has reduced to the nanometer range after chemical and mechanical treatment From the AFM photographs, it is clear that the fibers are found to be slightly agglomerated The surface of the fibers is found to be smooth from the AFM images in contrast to rough surface of microfibrils as seen in SEM images Figure 3(i) shows TEM images of the cellulose nanofibers after the chemical and mechanical treatments Mechanical treatment resulted in defibrillation of the cellulose nanofibers from the cell wall and TEM images reveal the separation of these nanofibers from the microsizes fiber bundles The diameter of the fibers is found in the range of 10–60 nm A tendency of agglomeration could also be observed from TEM It is not clear whether this was due to drying of the suspension onto the carbon film covering the carbon grids or if it reflected the state of the suspension The average diameter was calculated from the electron micrographs using digital image analysis software, UTHSCSA Most of the particles were found in the diameter range of 30–40 nm Figure 3(ii) shows the distribution of nano-fiber diameter after final treatment Only 3% of fibers have diameter >70 nm Almost 64% of fibers have diameter between 30–50 nm, 19% of the fibers have a diameter less than 30 nm Table shows the chemical composition of raw, alkaline steam exploded, bleached fibers and acidic treated fibers Raw fiber has A Kaushik, M Singh / Carbohydrate Research 346 (2011) 76–85 79 Figure SEM image of the steam-exploded micro-structured cellulose fibers at different magnifications Table Chemical composition of untreated and steam exploded and chemically treated wheat straw Material Percentage of acellulose Percentage of hemi cellulose Percentage of lignin Untreated wheat straw Alkaline steam exploded Acid treated fibers Cellulose nanofibers 45.70 ± 0.18 37.12 ± 0.9 17.43 ± 2.1 65.29 ± 2.51 22.22 ± 1.12 10.27 ± 1.67 75.28 ± 2.37 12.34 ± 1.18 8.12 ± 1.35 86.38 ± 3.12 8.13 ± 0.8 6.34 ± 1.25 Table Bragg angle, fiber diameter, crystallinity index and crystallinity order index Fiber type Bragg angle (°) Fiber diameter (nm) (Eq 6) Crystallinity index (%) (Eq 4) Crystallinity order index (Eq 5) WS CTWS CNF 11.259 11.23 11.35 — 5.23 3.91 54.42 66.60 79.87 57.43 71.33 80.05 in alkaline medium, the hemicelluloses and lignin components present in the raw fiber will dissolve out Yamashiki et al.24 proposed an explanation for the solubility of steam exploded cellulose in NaOH solution, suggesting that during the steam explosion there is a partial breakdown of the intramolecular hydrogen bond at the C-3 and C-6 positions of the glucopyranose unit and this results in significant variations in the network and strength of the hydrogen bonds of the cellulose hydroxyls However, the complete removal of these components does not take place During the explosion, some changes occur in the arrangement of macromolecular chains Xiao et al.25 proposed that during steam explosion, the hemicellulose is partially hydrolyzed and the lignin is depolymerized, giving rise to sugars and phenolic compounds that are soluble in water The hydrolyses of glycosidic linkages in hemicellulose and the ether linkages in lignin are catalyzed by acetic acid formed at high temperature from acetyl groups present in hemicellulose (autohydrolysis) At the end of the process, the steam was suddenly released providing additional mechanical defibrillation The cellulose is de-polymerized and defibrillated resulting in crystalline nanofibrils The removal of hemicelluloses and lignin in chemically treated fibers is confirmed in FT-IR results also 3.2 Fourier Transform infrared (FT-IR) spectroscopy the highest percentage of hemicellulose and lignin and the lowest percentage of a-cellulose as compared to treated fibers The acellulose content increases from 45.7% in raw fiber to 86.38% in finally treated fiber Similarly, hemicelluloses content decreases from 37.12% to 8.13% and lignin decreases from 17.43% to 6.34% When the steam explosion process is done, there is a decrease in the hemicellulose and lignin component present in the wheat fiber This shows that during steam explosion, substantial breakdown of the lignocellulosic structure, partial hydrolysis of the hemicellulosic fraction, and depolymerization of the lignin components have occurred.17 When the raw fiber is subjected to steam explosion Untreated and chemically treated wheat straw fibers were analyzed using FT-IR to examine the changes occurring in the chemical constituents of fibers before and after the chemical treatments Figure shows the FT-IR spectrum of the raw wheat straw fibers and chemically treated wheat straw fibers The peaks in area 3369 cmÀ1 correspond to O–H stretching band, that is, due to vibrations of the hydrogen bonded hydroxyl group25–27 The peaks at 2922 cmÀ1 are due to the aliphatic saturated C–H stretching vibration in lignin polysaccharides (cellulose and hemicelluloses) The hydrophilic tendency of raw fibers and chemically treated wheat straw fibers is reflected in the broad absorption band in 80 A Kaushik, M Singh / Carbohydrate Research 346 (2011) 76–85 Figure AFM images of cellulose nanofibrils (tapping mode): (a) 10 lm  10 lm, (b) 1.5 lm  1.5 lm phase images, (c) lm  lm topographical image and (d) lm  lm topographical image the 3700–3100 cmÀ1 region, which is related to the –OH groups present in their main components Peak at 1734 cmÀ1 in the untreated wheat straw is attributed to either the acetyl and uronic ester groups of the hemicelluloses or the ester linkage of carboxylic group of the ferulic and p-coumeric acids of lignin and/or hemicelluloses.17,26,28 It can be seen in Figure that this peak is almost absent in the spectra of the chemically treated fibers, which indicates the near cleavage of these ester bonds The peak at 1652 cmÀ1 may be due to the bending mode of the absorbed water and some contributions from carboxylate groups.26 The aromatic C@C stretch from aromatic ring of lignin gives two peaks at 1510 and 1426 cmÀ1 that can be observed in untreated wheat straw fibers.17,28,29 The peak at 1510 cmÀ1 has almost vanished and the intensity of peak at 1426 cmÀ1 has significantly decreased in chemically treated fibers attributing to partial removal of lignin The peaks at 1373 cmÀ1 represent C–H asymmetric deformation The intensity of the peak at 1258 cmÀ1 has sharply decreased after chemical treatment indicating the removal of hemicelluloses The region of 1200–1059 cmÀ1 represents the C–O stretch band and deformation bands in cellulose, lignin and residual hemicelluloses.26 The increase of band at 897 cmÀ1 in chemically treated wheat straw fibers indicates the typical structure of cellulose (due to b-glycosidic linkages of glucose ring of cellulose).30 3.3 X-ray diffraction WAXRD analyses of untreated and treated fibers were done in order to study the crystalline behavior of the fibers and to assess the relationship between structure and properties of fiber Cellulose show crystalline nature while lignin is amorphous in nature As a result, the crystallinity of the fibers should improve after removal of the lignin Based on this idea X-ray diffraction of powdered samples of the untreated wheat straw fibers, chemically treated fibers, and mechanically high shear treated fibers was carried out so as to examine the changes occurring in the crystalline nature of the wheat straw fibers after the chemical treatment Figure 5(i) shows the XRD profiles of the untreated wheat straw fibers, chemically treated fibers, and cellulose nanofibrils after mechanical shear treatment As discussed earlier by many authors, lignocellulosic fibers are composed of three major components namely cellulose, hemicellulose, and lignin Crystalline microfibrils of cellulose are surrounded by amorphous hemicellulose and the whole is embedded in the matrix of lignin Crystalline structure of cellulose and hemicellulose exhibits variability in both structure and constitution In native cellulose the length of the crystallites can be 100–250 nm with average cross-sections of 3–10 nm Chemical and mechanical treatments affect the crystallite size as well as the crystallinity of cellulose From Figure 5(i) it is clear that finally treated cellulose nanofibrils show crystalline nature The peak intensity at 2h = 22.6 corresponding to 0 lattice plane increases with the chemical treatment It further increases sharply with high shear mechanical treatment Chemically treated fibers show a narrow peak at 26.5°, which may be due to heavy loading of chemicals Higher crystallinity is due to more efficient removal of noncellulosic polysaccharides and dissolution of amorphous zones The A Kaushik, M Singh / Carbohydrate Research 346 (2011) 76–85 81 Figure 3(i) TEM images of cellulose nanofibrils at different magnifications: (a) 50,000Â, (b) 15,000Â, (c) 80,000 and (d) 40,000 results also confirm that hydrolysis takes place preferentially in the amorphous region as acids dissolve the amorphous regions while crystalline regions are more stable toward chemical attack This increase of crystallinity after acid treatment has been reported by several authors.17,28,31 To evaluate the crystallinity order index, peak separation of each X-ray diffraction was conducted for each plane The operation used a non-linear multi-peak fitting function of OriginPro software The Pseudo-Voigt type I equation was chosen There are five variables in this equation; peak offset (y), peak amplitude or area (A), peak center (xc), peak width (w) and peak shape factor (mu) After an equation and data from WAXD were assigned, the fitting was initiated by entering number of replicas equalent to the number of peaks À The fitting was performed with several iterations until the optimum fit result was obtained This was indicated by no further decline of v2 value and R2 approached greater than 0.99 Fitted data for untreated wheat straw fiber as an example have been shown in Figure 5(ii) Table gives the value of Bragg angle, crystallite size (t), crystallinity index (CI), and crystallinity order index (COI) for untreated, chemically treated, and mechanically treated fibers The crystallinity increases from 54.42% for untreated fibers to 79.87% for cellulose nanofibrils and crystallinity order index increases from 57.43% to 80.05% for cellulose nanofibrils The increase in 82 A Kaushik, M Singh / Carbohydrate Research 346 (2011) 76–85 40 35 25 B frequency (percent) 30 20 15 10 0-20 20-30 30-40 40-50 50-60 60-70 >70 particle diameter (nm) Figure 3(ii) Diameter distribution of wheat straw fiber after chemical treatment (obtained from TEM analysis) crystallinity could also be attributed to the facts that steam at high temperatures reorganizes the amorphous and paracrystalline cellulose regions It releases strains from native cellulose that arise during the crystallization phase of cellulose biosynthesis and the interaction of the cellulose with hemicellulose and lignin in cell wall formation.17 It is evident from the results that with chemical and mechanical treatment value of b increases and thus there is a decrease in fiber diameter 3.4 Thermal characterization As we aim to reinforce the particles in thermoplastic starch based polymers to make completely biocompatible polymers, the thermal properties of these untreated and treated fibers is important in order to gauge their applicability for biocomposites, in which processing temperature for thermoplastic polymers rises above 200 °C The TGA results for wheat straw fibers at different stages are shown in Figure 6(i) These results clearly show that the thermal stability of the wheat straw fibers increases after chemical treatments and it further increases after high shear mechanical treatments The degradation temperature increased after chemical treatment This was probably because more non-cellulosic material was removed and the high degree of structural order was retained This revealed a relationship between structure and the thermal degradation of cellulose A greater crystalline structure required a higher degradation temperature.32 However, both noncellulosic components and the crystalline order of cellulose played an important role in thermal degradation of the fibers.33 Different amounts of the residues are obtained from the fibers remaining after 600 °C heating for untreated, treated and nanofibers Maximum residue was obtained in untreated wheat straw fibers and relatively small amount in nanofibers after chemimechanical treatment It can be concluded that higher temperature of thermal decomposition and lesser residual mass of the fibers obtained after chemi-mechanical treatment have been related to partial removal of hemicelluloses and lignin from the fibers and higher crystallinity of the cellulose These results are consistent with results obtained from crystallinity and FT-IR measurements Figure 6(ii) shows DTG for untreated, chemically treated, and mechanically treated fibers For untreated wheat straw fibers the first decomposition shoulder peak at about 236 °C is attributed to thermal depolymerisation of hemicelluloses or pectin in an inert atmosphere (mass loss 6.3%)34; (2) the major second decomposition peak at about 321.86 °C is attributed to cellulose decomposition (mass loss 27.81%); (3) the small tail peak at 343 °C (mass loss 30%) may be attributed to degradation of lignin35; (4) a very small peak at about 433 °C is for oxidative degradation of the charred residue In chemically treated fibers the first peak disappears but the second peak exists while in nanofibers, DTG shows a sharp peak only at 337.58 °C indicating decomposition of crystalline cellulose Thermal decomposition parameters for untreated, chemically treated, and mechanically treated fibers were determined from the TG, DTG, and second time derivatives, D2TG, curves at a heating rate of 10 °C/min as an example (Fig 6(iii)) using method given by Groni et al.36 The extrapolated onset temperature of decomposi- spl.code:CNF spl.code:Treated spl.code:WS 0.9 0.8 Transmittance 0.7 0.6 0.5 0.4 0.3 0.2 0.1 3500 3000 2500 2000 1500 1000 500 Wavenumbers [1/cm] Figure FT-IR spectra of wheat straw fibers (untreated and chemically treated and cellulose nanofibrils) 83 A Kaushik, M Singh / Carbohydrate Research 346 (2011) 76–85 2000 002 1800 Intensity (counts) 1600 1400 1200 101 1000 800 021 101 600 040 400 10 15 20 25 30 35 40 45 50 55 Diffraction angle (2θ) Figure 5(i) XRD spectra of different fibers: (a) untreated wheat straw, (b) chemically treated wheat straw, and (c) cellulose nanofibrils tion, To, was obtained by extrapolating the slope of the DTG curve in correspondence with the first local maximum in D2TG curve and down to the zero level of the DTG axis The peak temperature, Tp, was determined by DTG peak where the maximum decomposition rate was obtained The percentage weight loss corresponding to peak temperature is symbolized with MTp The final, tailing region indicated the end of cellulose decomposition Further reactions continued decomposition of lignin and tar or char was obtained from main components decomposition.34 The shift temperature, Ts, is defined here by extrapolating the slope of DTG curve corresponding to the local minimum in D2TG curve in this region and down to the zero level of DTG axis The percentage weight loss corresponding to Ts is marked as MTs The residual solid mass fraction percentage left at the final temperature, 600 °C is marked as MT600 The decomposition characteristics of wheat straw, chemically treated, and mechanically treated fibers are summarized in Table Figure 5(ii) Wide angle X-ray diffractogram and fitted data of untreated wheat straw (as an example) The parameter To indicates an onset decomposition temperature, it being 239.5 °C for wheat straw, 276.2 °C for chemically treated straw, and 283.2 °C for cellulose nanofibrils obtained after mechanical high shear treatment Weight loss in this period (referred to MTo) was observed around 7% for all fibers The parameter Tp presents the maximum decomposition rate of the fibers and it lies in a range of 321 to 345 °C The weight loss at this point is indicated by parameter MTp From peak to shift temperature, fibers had a rapid degradation in a narrow temperature range All the fibers completed almost 70% or higher weight loss at 356–372 °C as shown by MTs and Ts, respectively The residual weight was maximum for wheat straw which (22%) followed by chemically treated fibers (18%) Minimum residual weight (11%) was obtained after mechanical high shear treatment This is because of absence of non-organic components in the cellulose nanofibrils as confirmed by FT-IR and XRD analysis Figure 6(i) TG curves: (a) finally mechanical treated fibers, (b) after acidic treatment, and (c) wheat straw 84 A Kaushik, M Singh / Carbohydrate Research 346 (2011) 76–85 Figure 6(ii) DTG curves: (a) finally mechanical treated fibers, (b) after acidic treatment, and (c) wheat straw Figure 6(iii) Determination of decomposition characteristic parameters of wheat straw fibers as an example at a heating rate of 10 °C/min 3.5 Degree of polymerization The values for viscosity average degree of polymerization (DOP) and the corresponding molecular weight of steam-exploded wheat straw in alkaline medium, bleached pulp, acid treated bleached pulp, and cellulose nanofibrils were calculated from the intrinsic viscosity of its solution in 0.5 M CED, according to Robert and Adrian23 method For alkaline steam exploded wheat straw, intrinsic viscosity was 720.2 mL/g, viscosity average DOP was 2609.72, and molecular weight (Mw) was 422,775 For steam exploded bleached fibers viscosity reduced to 420.3 mL/g, viscosity average DOP was 1434.54, and molecular weight (Mw) was 232,395 The 85 A Kaushik, M Singh / Carbohydrate Research 346 (2011) 76–85 Table Decomposition characteristics of wheat straw, chemically treated fibers and nanofibrils Fiber type To (°C) Tp (°C) Ts (°C) Weight loss at To (MTo) (%) Weight loss at Tp (MTp) (%) Weight loss at Ts (MTs) (%) %age residue at T6 0 MT6 0 (%) Ts À To (°C) MTs À MTo (%) WS CTWS CNF 239.5 276.2 283.2 321.86 345.71 337.58 356.2 372.3 360.3 6.54 7.35 6.876 27.81 49.88 23.49 63.72 69.94 74.12 78.42 82.45 89.01 116.7 96.1 77.1 57.18 62.59 67.24 immersion of lignocellulosic fibers in dilute alkaline medium facilitates the adhesive nature of the fiber surface by removing natural and artificial impurities, and causes separation of structural linkages between lignin and carbohydrate and the disruption of lignin structure The mechanism of bleaching involves oxidation of lignin, which leads to lignin dissolution and its degradation These processes are accompanied by loss of cellulose as evident in decreasing molecular weight and degree of polymerization Acid treatment hydrolyzed the traces of hemicelluloses and lignin remaining after the bleaching phase by breaking down the polysaccharides to simple sugars and hence released cellulose fibers The combined acid steam treatments effectively reduce the long micro fibril chains to nanodimensions by maximum explosion of pressurized steam into the interfibrilar region.17 The viscoisity, DOP, and Mw further decrease with acidic treatment reaching a value of viscosity = 123.2 mL/g, DOP = 366.90, and Mw = 59,438 Finally the fibers were subjected to high shear mechanical treatment The high shearing action breaks down the fiber agglomerates and results in nanofibrils The viscoisity of cellulose nanofibrils obtained after high shear treatment reduced to 92.5 mL/g with DOP of 266.9 and Mw of 43,250 Conclusions In this study cellulose nanofibrils were isolated from wheat straw using alkaline steam explosion followed by chemical and high shear treatment The major constituent of these fibers was found to be cellulose TEM and AFM images confirmed fiber diameter of 30–50 nm FT-IR and XRD studies evidenced about dissolution of lignin and hemicellulose with chemical treatment of the fibers, which resulted in improved thermal stability The cellulose nanofibers were most stable to heat treatment followed by chemically treated fibers and untreated wheat straw fibers Degree of polymerization decreased from 2609.7 for steam exploded wheat straw to 266.9 for cellulose nanofibers Acknowledgments We gratefully acknowledge financial support rendered by All India Council of Technical Education (AICTE), India and University Grants Commission (UGC), India for the work References Siqueira, G.; Bras, J.; Dufresne, A Biomacromolecules 2009, 10, 425–432 Soykeabkaew, N.; Supaphol, P.; Rujiravanit, R Carbohydr Polym 2004, 58, 53– 63 Okubo, K.; Fuji, T.; Thostenson, E T Composites Part A—Appl S 2009, 40, 469– 475 Abe, K.; Iwamoto, S.; Yano, H Biomacromolecules 2007, 8, 3276–3278 Ahola, S.; Salmi, J.; Johansson, L.-S.; Laine, J.; Ãsterberg, M Biomacromolecules 2008, 9, 1273–1282 Hitoshi, T.; Akira, A Key Eng Mat 2007, 334–335, 389–392 Said Azizi Samir, M A.; Alloin, F.; Dufresne, A Biomacromolecules 2005, 6, 612– 626 Nishino, T.; Matsuda, I.; Hirao, K Macromolecules 2004, 37, 7683–7687 Fengel, D.; Wegner, G Wood: Chemistry Ultrastructure Reactions; Walter de Gruyter: Berlin, 1989 10 Nakagaito, A.N.; Yano, H American Chemical Society: Nanocomposites Based on Cellulose Microfibril Cellulose Nanocomposites; Washington, DC, 2009; pp 151– 168 11 Nakagaito, A N.; Yano, H Appl Phys A 2004, 78, 547–552 12 Nakagaito, A N.; Yano, H Appl Phys A 2005, 80, 155–159 13 Turbak, A F.; Snyder, F W.; Sandberg, K R J Appl Polym Sci.: Appl Polym Symp 1983, 37, 813–827 14 Andresen, M.; Johansson, L.; Tanem, B S.; Stenius, P Cellulose 2006, 13, 665– 677 15 Stenstad, P.; Andresen, M.; Tanem, B S.; Stenius, P Cellulose 2008, 15, 35–45 16 Liu, R.; Yu, H.; Huang, Y Cellulose 2005, 12, 25–34 17 Cherian, B M.; Pothan, L A.; Nguyen-Chung, T.; Mennig, G.; Kottaisamy, M.; Thomas, S J Agric Food Chem 2008, 56, 5617–5627 18 Purl, V P.; Mamers, H Biotech Bioeng 1983, 25, 3149–3162 19 Battista, O A.; Smith, P A Ind Eng Chem 1962, 54, 20–29 20 Structure of Cellulose and its Relation to Properties of Cellulose Fibers; Krassig, H., Ed.; Ellis Horwood: Chichester, U.K., 1985 21 Segal, L.; Creely, J J.; Martin, A E., Jr.; Conrad, C M Text Res J 1959, 29, 786– 794 22 Bodor, G Structural Investigation of Polymers; Ellis Horwood: New York, 1991 23 Robert, E.; Adrian, F A W J Appl Polym Sci 1989, 37, 2331–2340 24 Yamashiki, T.; Matsui, T.; Saitoh, M.; Okajima, K.; Kamide, K.; Sawada, T Brit Polym J 1990, 22, 12–128 25 Xiao, B.; Sun, X F.; Run, C S Polym Degrad Stab 2001, 74, 307–319 26 Sun, X F.; Xu, F.; Sun, R C.; Fowler, P.; Baird, M S Carbohydr Res 2005, 340, 97–106 27 Sain, M.; Panthapulakkal, S Ind Crop Prod 2006, 23, 1–8 28 Alemdar, A.; Sain, M Bioresour Technol 2008, 99, 1664–1671 29 Waleed, K E.-Z.; Maha, M I Polym Adv Technol 2003, 14, 623–631 30 Gañán, P.; Cruz, J.; Garbizu, S.; Arbelaiz, A.; Mondragon, I J Appl Polym Sci.: Appl Polym Symp 2004, 94, 1489–1495 31 Tang, L C.; Hon, D N S.; Pan, S H.; Zhu, Y Q.; Wang, Z.; Wang, Z Z J Appl Polym Sci 1996, 59, 483–488 32 Ouajai, S.; Shanks, R A Polym Degrad Stabil 2005, 89, 327–335 33 Yang, P.; Kokot, S J Appl Polym Sci 1996, 60, 1137–1146 34 Yao, F.; Wu, Q.; Lei, Y.; Guo, W.; Xu, Y Polym Degrad Stabil 2008, 93, 90–98 35 Antal, M J.; Varhegyi, G Ind Eng Chem Res 1995, 34, 703–717 36 Gronli, M G.; Varhegyi, G.; Blasi, C D Ind Eng Chem Res 2002, 41, 4201–4208  ... composition of untreated and steam exploded and chemically treated wheat straw Material Percentage of acellulose Percentage of hemi cellulose Percentage of lignin Untreated wheat straw Alkaline... region of 1200–1059 cmÀ1 represents the C–O stretch band and deformation bands in cellulose, lignin and residual hemicelluloses.26 The increase of band at 897 cmÀ1 in chemically treated wheat straw. .. untreated wheat straw is attributed to either the acetyl and uronic ester groups of the hemicelluloses or the ester linkage of carboxylic group of the ferulic and p-coumeric acids of lignin and/ or