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Bioresource Technology 100 (2009) 6398–6406

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Quantitative analysis of sugars in wood hydrolyzates with 1H NMR during
the autohydrolysis of hardwoods
Ashutosh Mittal a,1, Gary M. Scott a, Thomas E. Amidon a, David J. Kiemle b, Arthur J. Stipanovic b,*

Department of Paper and Bioprocess Engineering, SUNY College of Environmental Science and Forestry, 1 Forestry Drive, Syracuse, NY 13210, United States
Department of Chemistry, SUNY College of Environmental Science and Forestry, 1 Forestry Drive, Syracuse, NY 13210, United States

a r t i c l e

i n f o

Article history:
Received 6 February 2008
Received in revised form 18 June 2009
Accepted 27 June 2009
Available online 11 August 2009
H NMR spectroscopy

a b s t r a c t
The focus of this work was to determine the utility of 1H NMR spectroscopy in the quantification of sugars
resulting from the solubilization of hemicelluloses during the autohydrolysis of hardwoods and the use of
this technique to evaluate the kinetics of this process over a range of temperatures and times. Yields of
residual xylan, xylooligomers, xylose, glucose, and the degraded products of sugars, i.e., furfural and HMF
(5-hydroxymethyl furfural), were determined. The monosaccharide and oligomer contents were quantified with a recently developed high resolution 1H NMR spectroscopic analysis. This method provided precise measurement of the residual xylan and cellulose remaining in the extracted wood samples and
xylose and glucose in the hydrolyzates. NMR was found to exhibit good repeatability and provided carbohydrate compositional results comparable to published methods for sugar maple and aspen woods.
Ó 2009 Elsevier Ltd. All rights reserved.

1. Introduction
Lignocellulosic or woody biomass is an important renewable resource that offers an environmentally attractive alternative to
petroleum in the production of fuels, chemicals and polymeric
materials. Lignocellulosic feedstocks, however, have a complex
composite structure and their efficient utilization requires separation of their main polymeric components including cellulose,
hemicellulose and lignin with maximum yield and purity of each
fraction. Due to the recalcitrance of woody biomass, efficient fractionation into individual components typically requires a pretreatment step. At our institution, we focus considerable attention on
biorefining technologies wherein the hemicellulose fraction of
woody biomass is selectively extracted using a benign, hot water
pretreatment process before the cellulose fraction is isolated for
paper or ethanol production. In our case, the objective of pretreatment is to maximize the yield of hemicellulose-based sugars, predominately xylose and its oligomers, and to prepare the pretreated
residue for enzymatic hydrolysis (Fan et al., 1982). Other commonly employed pretreatment techniques for woody biomass include physical treatments such as grinding, milling and steam
explosion or techniques involving the addition of chemicals (Overend and Chornet, 1987; Sun and Cheng, 2002). The objectives of

* Corresponding author. Tel.: +1 315 470 6860; fax: +1 315 470 4729.
E-mail addresses: (A. Mittal), (A.J.
Present address: NREL, 1617 Cole Blvd., Golden, CO 80401, United States. Tel.: +1
303 384 6136; fax: +1 303 384 5572.
0960-8524/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.

this study are: (1) to introduce a new NMR-based technique to
quantify the sugars present in solid wood and wood hydrolyzates
and, (2) to use this technique to study the kinetics of wood
For the hardwood feedstocks used in this study, the hot water
pretreatment process is especially effective at solubilizing hemicellulose and cleaving native acetate groups from hemicellulose molecules thereby lowering pH and enhancing the potential for
‘‘autohydrolysis”, a reaction that reduces the polymer to fermentable sugars. Benefits associated with autohydrolysis as a pretreatment are both economical and environmental (Garrote et al.,
1999a,b). Hydrolyzates obtained in this fashion contain residual
polymeric xylan, xylooligomers and xylose formed due to the
hydrolysis of xylan, which is the main fraction of hardwood hemicellulose (Timell, 1967). Xylooligomers and xylose can be converted into various products, which have potential utilization in
food, feed, pharmaceutical, and agricultural industries (Garrote
et al., 1999a, 2004; Parajo et al., 2004). Hardwood hydrolyzates
also contain lower amounts of other hemicellulose-derived sugars
including mannose, galactose, rhamnose and arabinose.
The ability to accurately quantify the solubilized carbohydrates
(i.e., xylooligomers and xylose) in the hydrolyzate is an important
parameter in determining the efficacy of the pretreatment method
and estimating the potential yield of the total sugars that can be
derived from a specific feedstock. Typically, sugars in the hydrolyzates are quantified chromatographically using HPLC or a related
technique, High Performance Anion Exchange Chromatography
with Pulsed Amperometric Detection (Çöpür et al., 2003; Sjostrom
and Alen, 1999) or they can be derivatized for GC-analysis (Çöpür

2000).. Table 3 Operational conditions used during autohydrolysis experiments. the heating-up time for wood chips was reduced to about 3–5 min and the beginning of the experiment was considered as the beginning of the isothermal stage. The hydrolyzate samples obtained after the autohydrolysis process were analyzed for pH at room temperature and about 25 g of hydrolyzate sample was used for total solids determination for each reaction condition.% and this would improve the mass balance closure reported below. Analysis of raw and extracted wood Sugar analysis of both the raw wood and residual wood samples after hot water extraction samples was performed by 1H NMR spectroscopy using a method described by Kiemle et al.12 0. Raw material Particle size (cm) Temperature (°C) Time (h) LSR (g/g) Sugar maple Aspen 2. In this study.. It was observed that during this heating-up period about 5–6% of the reaction material was solubilized. However. Sugar Avg. 2003.14 0.5  2. (2003). 2004).4 2. This sugar is initially attached to about 10% of the xylose residues in hardwood hemicellulose (Sjostrom and Alen. Previous work has shown that GlucA is underestimated by the NMR protocol due.6 0.0 24.0 3. A detailed discussion of the quantification algorithm employed for the NMR protocol was presented elsewhere (Kiemle et al. perhaps.1 5. Tenkanen et al. after drying.1 2.% concentration. To summarize. Mittal et al. 1999). Although an accurate quantification procedure for this sugar is still under development in our laboratory.5 3. the proton resonances associated with the C1-a and C1-b anomeric protons are integrated and summed for each sugar providing an estimate of its relative molar concentration in a mixture of sugars.0 9. the data presented herein do not attempt to quantify GlucA.05 a Average of 5 runs on the same sample.4 51. 2002. Table 1 Sugar quantification algorithm (Kiemle et al.. 1999. The treated wood chips. Data were acquired and processed in TOPSPIN v1.5  2. It should be also noted that the NMR method is capable of detecting signals associated with 4-O-methyl-glucuronic acid (GlucA) at 4. For a synthetic mixture of sugars of known concentration. 1995.07 0. 1998) to approximate the intensity of the second anomeric proton. were milled to a particle size to pass a 30-mesh screen.5 cm3).3 ppm chemical shift (Kiemle et al.4 0.0  0. to lactone formation (Marsh. the reaction was terminated by discharging the liquor through a heat exchanger.5 152–175 160 0–6 0–3 6 6 .0 1. The dynamic range of the resonances of interest was improved by pre-saturating the residual water peak.7 2. Sugar % in synthetic mixturea % Determined by NMR algorithma Difference (%) Glucose Xylose Mannose Galactose Arabinose Rhamnose 50. The aspen chips were obtained from the Department of Paper and Bioprocess Engineering.3 9..24 0.0  0. The main operational variables under which the autohydrolysis experiments were conducted are presented in Table 3 for a constant liquid/solid ratio (LSR). The air-dried chips were screened using a vibratory screen to a size normally used in industry (2. spectral evidence suggests that this sugar is present in our NMR samples at the level of 1–2 wt. Table 2 Repeatability of the NMR protocol for sugar analysis of wood pulp (Kiemle et al.9 3. 1966).5  2. mol%a Standard deviation Glucose Xylose Mannose Galactose Arabinose 90. At the end of the desired time.9 a Average of two separate mixtures and two separate NMR experiments.3 from Bruker BioSpin. we use the integrated intensity of either the C1-a or C1-b proton and then we use the published anomeric ratio (a/b) values (Robyt. Whistler and Rowell. 2. The normal heat-up time to heat wood chips to the desired temperature (typically 160 °C) in the M/K digester was about 25–30 min.8 3. Wood sample preparation In this study.7-liter M/K digester equipped with a centrifugal pump for liquor circulation and a PID temperature controller at a LSR of 6:1.2. we exploit a new methodology based on proton (1H) NMR spectroscopy for sugar quantification (Kiemle et al. All spectra were acquired at 10 °C with a Bruker AVANCE 600 spectrometer (600 MHz 1H frequency) equipped with a 5 mm triple resonance z-gradient probe. Table 2 highlights the excellent repeatability of this method. selected to be typical of a hardwood hydrolyzates.1. (2004) and Çöpür et al. and were screened and stored following the same procedures as that for the sugar maple chips.4. The screened chips were stored at room temperature in barrels in a single lot to avoid variation in the experiments. The milled wood samples obtained after the autohydrolysis treatment for each different time and temperature conditions were stored in a sealed zip-lock bag at room temperature (24 °C) for later testing of sugars and lignin as explained below. By using pre-heated dilution water.5 2. 2. Sample handling After autohydrolysis experiments. 2.6399 A. 2004).3 3. autohydrolysis experiments were conducted on two hardwood species: sugar maple (Acer saccharum) and aspen (Populus tremoloides) at various time and temperature conditions. 2004). 2004). 2004).7 8..8 4. wt.9 0. To reduce heat-up time. 2. Sjostrom and Alen. the solid residue obtained was weighed immediately and an aliquot was oven dried at 105 °C for 16 h for moisture determination. Teleman et al.0  0.6 5. dilution water was pre-heated in another digester before being charged to the chip containing M/K digester as shown in Fig.. 1966) and/or the comparative stability of its linkage to other sugars or oligosaccharides (Bertaud et al. Experimental procedures 2.4 10. / Bioresource Technology 100 (2009) 6398–6406 et al. For those sugars where the individual C1-a and C1-b proton resonances are not well resolved. To obtain quantitative data the 1H spectra were acquired with a relaxation delay set to 10 s. Autohydrolysis experiments with wood chips Wood chips were treated in a 4.3.5 24. 1. Sugar maple wood logs were obtained from ESF Forest Properties and were debarked and chipped in a Carthage chipper and were air-dried to a moisture content of 9– 12%.7 and 5. The rest of the samples were stored in plastic bottles in a cold room maintained at 4 °C for later analysis of sugars and sugar degradation products in the hydrolyzate.. this quantification algorithm provided values as shown in Table 1 below...

20 ml of 72% deuterated H2SO4 was added to 0. free acetyl groups.4 ml of D2O (NMR solvent) was added to the dispersion.55 (17) 2.24 <0.65 (52)a 15. 0. The difference in the monosaccharide concentration of the two aliquots provided Table 4 Composition (% dry basis) of raw materials (sugar maple and aspen). Repeatability of the NMR protocol In an effort to determine repeatability for the 1H NMR methods employed in this work. the carbohydrate composition of both solid wood samples and hydrolyzates were run in triplicate to determine a standard deviation with 95% CI. respectively.% since.8) 0.54 ± 0. To quantify the oligomers present in the hydrolyzate.0 (21) 2. was subjected to 1H NMR analysis. and HMF. 90° pulse angle.040 g of OD (oven dried) milled wood mass. b Reference values from Pettersen (1984).6400 A.05 ± 0. galactose.09 (0.5) 0.7) 0. 1. xylose.88 (0. After stirring. For NMR analysis. upon addition of 0. illustrate that the maximum variation for xylose and glucose in the extracted sugar maple wood was about 1. which was then autoclaved at 121 °C for 60 min.24 (2.3% and 0.7 s acquisition time and spectral widths of 16 ppm.1 ml of 72% deuterated sulfuric acid to 1 g of the sample.8% and 3.12 ± 0.42 ± 0. which was then autoclaved at 121 °C for 45 min to convert the oligomers into their monomers.42 ml of 96% H2SO4 was added.6.1) 0.% H2SO4 solution. T 222 om-88 and UM 250.3 (23) 3. 2. After this digestion.72 ± 0. Hydrolyzate analysis Monomeric sugars formed during the autohydrolysis of sugar maple and aspen wood chips (glucose. data included in Tables 4–6 suggest that the practical minimum detection limit for a specific sugar using NMR is around 0. respectively. respectively. which was followed by the addition of TMA (trimethylamine hydrochloride). due to . 2. TMA was used as an internal standard for both aliquots.83 (49)b 14.33%.16 (0.13 (2.25 (3.51 (15) 2. arabinose and rhamnose) as well as sugar degradation products (furfural and hydroxymethyl furfural (HMF)) were analyzed with 1 H NMR.63 ± 0.47 ± 0. Data provided in Tables 5 and 6.28 22. mannose.71 ± 0.17 (2.1) 1.48 (2.9 nd* * nd – not determined. The addition of extra acid at this point helped to shift the proton resonances of H2O to higher ppm out of the range of sugar proton signals. 0. Schematic diagram of the experimental apparatus for the experiments with wood chips.24 ± 0.22 0.79 ± 0.5. a measure of the concentration of the oligomers. another 5 g aliquot (Aliquot 2) was mixed with 96% H2SO4 to obtain a 4 wt.05 20. Composition Sugar maple Aspen Glucose Xylose Mannose Galactose Arabinose Rhamnose Acetate Furfural HMF Klason lignin Acid soluble lignin Extractives 45.25 wt. This sample was also filtered and.64 ± 0. at lower concentrations. the range of replicate results is on the order of the concentration itself.2 47. respectively. furfural. The standard deviation with 95% CI for sugar maple and aspen raw wood samples was calculated from quadruplicate and duplicate experiments. whereas in the hydrolyzate the maximum variation for xylose and glucose was about 2.0) 1.4%.19 3. an internal standard. respectively.08 ± 0. The composition of sugar maple and aspen along with standard deviations having a 95% confidence interval (CI) are reported in Table 4. Mittal et al.26 ± 0. Collectively. Klason lignin and acid soluble lignin were determined by standard TAPPI methods. A sample designated as Aliquot 1 of the hydrolyzate was filtered and used directly for the determination of monosaccharides. the dispersion was allowed to digest at 40 °C for 60 min in a water bath with stirring every 15 min. / Bioresource Technology 100 (2009) 6398–6406 P T P RV1 T RV2 P1 HC CW V HE P1: Pump P: Pressure gauge RV: Reaction vessel T: Temperature gauge V: Valve CW: Cooling water HE: Heat exchanger HC: Heating coil and liquor recirculation line Hydrolyzate Fig. a Includes glucose from glucomannans.67 ± 0. 5.3) 2. 2.64 ± 0.8 3.9) 0.17 3.78 ± 0. After cooling.53 ± 1.

0 1.32 0.67 1.23 0.13 0.91 ± 0.42 ± 0..50 ± 0. b-xylose.0 2.91 ± 0.50 ± 0. 2004). 2004.20 0. 2.15 0. Fig.05 0.40 ± 0.82 ± 0.63. NMR Quantification of sugars. 2003. Mittal et al.15 ± 0.7 4.09 6.6 2.6 Fig..82 1. Results and discussion 3.2 9.29 0.0 4.37 ± 0.53 0. The amount of HMF and free acetyl groups Acetate Furfural TMA HMF 0.01 ± 0..08 0.0 ppm Free 3.0 1. sugar degradation products and acetyl groups Table 6 Repeatability of 1H NMR sugar analysis for wood hydrolyzates (160 °C extraction.18 0. Lakshmi Bala et al.90 ± 1.33 0.07 <0.25 <0.65 59. HPLC with RI detector and High Performance Anion Exchange Chromatography with Pulsed Amperometric Detection (HPAEC-PAD) at different laboratories (Kiemle et al.11 0.77 1.23 2.07 ± 0.66 ± 0.11 8.46 0.68 ± 0.5 9.64 1.56 (Kabel et al.29 ± 0.35 0.50 ± 1.85 ± 0.08 0.54 ± 0.3 5.36 ± 0. g/ 100 g oven dried extracted wood).36 ± 0.25 0.17 <0.22 ± 0.33 ± 0.8 2.59 ± 3.5 h Sample 2 1h Sample 3 2h Xylose Mannose Glucose Galactose Arabinose Rhamnose Acetate Methanol Furfural HMF 12.3 0.85 ± 0.23 56.99 ± 0.46 ± 0. 2.2 4. respectively.26 0.05 ± 0.1 Rha 5.14 ± 0.32 0.3 1. and b-arabinose at 4.1.03 ± 0.42 ± 2.42 0.34 ± 0.23 0.5 Bound 149.93 ± 1.05 11.18 ± 0. 2004).9 8.29 1..05 8.9 4.34 51.2 ppm (Evtuguin et al.6 5.5 4 . 2003).31 0.11 ± 0.27 ± 0.38 0.61 ± 0.32 ± 0.31 ± 1.1 and 2.0 100.11 0.42 ± 0.17 2.5 7. and 4.4 5.51 ppm.16 0. Teleman et al. 2002. good agreement exists between this study and the compositional data published by Pettersen (1984) using an HPLC technique. respectively.2 0.11 0.97 ± 1. 2 contains the 1H NMR spectrum of a representative hydrolyzate sample (160 °C/1 h) obtained by hot water extraction of sugar maple wood chips (designated Aliquot 1).5 h Sample 2 1h Sample 3 2h Xylose Mannose Glucose Galactose Arabinose Rhamnose Acetate Methanol Furfural HMF 4.0 3.13 1.8 2...49 0. Results illustrated that NMR and HPAEC-PAD gave very similar compositional profiles while HPLC/RI severely underestimated the xylose fraction for all samples (Kiemle et al.2 8. 1H NMR spectra of treated sugar maple wood chips hydrolyzate: Aliquot 1 (see text). 3.26 ± 0.18 ± 0.22 ± 0..03 ± 0.5 3.77 ± 0.10 ± 0.23 0.0 8.09 Previously.3 1.23 ± 0.19 0.13 ± 0.2 3.8 0.25 0.16 ± 2. Sugars Extraction time Sample 1 0.18 1.5 4. This spectrum shows resonance peaks for free acetyl groups and acetyl groups bound to the xylan chain at 2. g/100 g oven dried extracted wood).49 ± 0.05 instrumental sensitivity and operator judgment in establishing signal integration limits.14 0.29 ± 0.11 ± 0.6401 A. / Bioresource Technology 100 (2009) 6398–6406 Table 5 Repeatability of 1H NMR sugar analysis of residual solid wood after hot water extraction (160 °C extraction. Sun et al.8 92. 4. Sugars Extraction time Sample 1 0.4 Gal Ara GLU Xyl Man 5.08 2.44 ± 0.5 0.35 ± 0. we have compared compositional results determined for three samples (willow wood. and distinct peaks for the anomeric protons of b-glucose.82 ± 0.10 0.31 ± 0.45 ± 0. Table 4 also shows that the repeatability of the NMR method is very good.19 0. paper birch wood and recycled paper) in a ‘‘round robin” test protocol employing NMR.18 3. Application of the NMR method to determine the carbohydrate and acetyl distribution of oven dried sugar maple and aspen woods provides the results given in Table 4. 2005).1 0.7 5.10 0.2 2. 2003. Notwithstanding differences in the actual samples analyzed.

From the compositional data for sugar maple wood provided in Table 4. Based on the 96% mass balance closure on total sugars it appears that the high resolution NMR technique is an accurate method for the quantification of sugars in pretreated solids and hydrolyzates obtained during autohydrolysis process.4 p Fig.0 2. 3. Objective 2 for this work was to assess the utility of 1H NMR spectroscopy in the quantification of sugars resulting from the solubilization of hemicelluloses while evaluating the kinetics of the autohydrolysis of hardwoods.0 96.7 GluA 4. As a result. an additional 1–2% of mass could potentially be accounted for by GlucA. The compositions of the recovered solid and liquid streams show that about 96% of the total sugars and Klason lignin (Total2) are recovered whereas on the overall basis (Total3) about 91% of the material is recovered.4 3.7 4. 1H NMR spectra of treated sugar maple wood chips hydrolyzate: Aliquot 2 (see text).5. 3.1 0. rhamnose) which were not quantified in Aliquot 1 were quantified from 1H NMR spectra obtained for Aliquot 2 as shown in Fig. 7.3 5. 3. Since a fraction of the hydrolyzate obtained after autohydrolysis could not be identified as sugars or Klason lignin.6 Glu 4. The difference between the amount of xylose in Aliquots 1 and 2 provided the amount of xylooligomers present in the hydrolyzate.1 5.0 Xyl Ara 4.6 4. sugars or Klason lignin) was lost during autohydrolysis.5 4. The increase in the amount of furfural between Aliquots 1 and 2 provided the amount of xylose (pentose) degraded to furfural and this was added to the total xylose concentration.8 p 294. furfural and HMF). / Bioresource Technology 100 (2009) 6398–6406 Acetate Furfural TMA HMF 3. However. During the .8 4. acid soluble lignin.5 Man Xyl GluA 2.4 2.5 7.0 5. As mentioned previously. NMR spectra do show weak signals associated with GlucA but these are not accurately quantified.8 15. Mittal et al. Since the xylooligomers conversion step to xylose was carried in the acidic conditions (4% by weight).0 Rha 8.0 Man 5. individual mass balances were performed on the total sugars (Total1).6 2.3 Gal Ara GLU 5. and extractives. galactose.2 100.2 ppm 58.e.5 ppm.45 and 2.7 7. in the hydrolyzate was quantified by integrating the peaks obtained at 9.6 9. total sugars and Klason lignin (Total2) and overall material (Total3).1 6. during this step some amount of xylose undergoes a dehydration reaction to yield furfural Root et al.2. Mass balance To estimate the amount of material recovered and the material lost in both the treated wood and the hydrolyzate sample obtained after autohydrolysis.. The results of the material balance for a selected experiment (autohydrolysis of sugar maple wood chips at 160 °C for 1 h) is given in Fig.9 3.8 1.0 ppm. it can be seen that the total extractives and acid soluble lignin in sugar maple wood adds up to 6.9 and 7. To identify which fraction of the raw material (i. Kinetics of wood autohydrolysis As mentioned earlier. the overall mass balance was always lower than the mass balance for total sugars or total sugars plus Klason lignin. A second aliquot of the same hydrolyzate was further treated with acid to reduce its oligomers to monomer sugars (designated as Aliquot 2). It is plausible that the majority of the 9% mass loss observed on the overall basis (Total3) was due to the formation of unidentified compounds originating from sugar degradation products (i.0 2.0 2.3.0 12. 4 and Table 7. (1959).5 8.2 5.5 14.7 0. 3. these xylooligomers and acetyl groups attached to the xylooligomers and other sugar monomers (mannose. Furfural was quantified by averaging the integrated intensity of the three peaks obtained at 9.e.6402 A. a mass balance was performed on a 100-g oven-dry basis (ODB) of the raw material.6 9.5% of the total wood mass.9 5. respectively.

%XlnRM is the amount of xylan in the raw material ex- pressed as g xylose/100 g ODW and %Xln is the amount of xylan in the treated material expressed as g xylose/100 g oven dried extracted wood (ODEW).7 g Mannose 0. 1 h Hydrolyzate (510.3 92.5 g Arabinose 0.7 g Galactose 0.2 85.8 g solids odb 44. mannose and acetate in the residual xylan in the extracted wood are shown in Fig.0 g Acid soluble lignin 3. / Bioresource Technology 100 (2009) 6398–6406 Sugar maple (100 g odb) 45. the percentage of xylan remaining in the extracted wood chips (PRXln) can be calculated using an equation proposed by Garrote (Garrote et al.2 45. lignin Total1 Total2 Total3 a Untreated wood (g) Extracted wood (g) Hydrolyzate (g) Total (g) %a 100.3 96.8 82.0 66.3 g Hemicellulose 10.5 g Water 0.61 0.2 0. Yields and compositions in the autohydrolysis of sugar maple wood chips at 160 °C and 1 h.5 g Mannose 0.6 61.72 0.3 14.8 0.0 7.4 g Hemicellulose 4.2 g Extractives 600 g Water (6:1) Autohydrolysis o 160 C.2 96.7 44. The time courses for the xylan remaining (percentage of the initial).8 g Acetate 1.0 g Xylose 2.6 g Arabinose 0.8 7.00 0.7 g Glucose 16.34 2.77 0.6 84.6 0.6 90.5 g Galactose 0.54 0.7 g) 76.6403 A. The values of %Xln for three different autohydrolysis experiments are given in Table 5. PRXln ¼ %Xln  SY RM %Xln ð1Þ where SY is the solid yield of the treated solid (g/100 g oven dried wood (ODW)).4 1.9 g Furfural and HMF 21.05 g Furfural and HMF 6.7 g Acid soluble lignin 1.2 g Rhamnose 1. Mittal et al.9 48.2 g Xylose 0.3 g Furfural and HMF 22.52 0.6 g Klason lignin 1.0 15.2 g Xylose 1.7 0.8 10.8 90.1 100.3 g Rhamnose 2.6 0.8 g Glucose 7.4 71.3 95.8 g Rhamnose 3.9 g Water + 85.27 21.0 Percentage of the mass obtained in the treated wood and hydrolyzate as a total mass of untreated wood.01 0.50 0. Basis.1 85.4 2.7 g) 496.20 1. 1999a).6 68.6 96.9 101. g Xylose Mannose Glucose Galactose Arabinose Rhamnose Acetate Furfural HMF K.4 g Klason lignin 4.82 3.2 1.2 0.0 g Galactose 0.4 45.50 0.5 2.56 22.5 g Glucose 26.0 g Arabinose 0.0 95.9 58.58 0.5 g Other (by difference) Extracted Wood (162.62 0. 4.02 0.8 7.7 0. Table 7 Mass balance closure during the autohydrolysis of sugar maple wood chips at 160 °C and 1 h.6 g Acetate 0.03 0.8 100.0 82. and the mass fractions of xylan.7 99.0 14. course of hydrothermal treatment on xylan.0 g Hemicellulose 15.5 g Other (by difference) Fig.4 g Mannose 2.0 g Acetate 0.54 3. 5 where it can be seen that the amount of residual xylan decreases with time as the hydrolysis rate increases .25 21..5 93.1 0.0 2.2 4.2 91.

2 0. with the temperature. The concentration of total xylose in the hydrolyzate increases with reaction time and. and acetate in the extracted wood for all the autohydrolysis temperatures remain constant with hydrolysis time. It is interesting to note that at the maximum recovered yield of sugars in the hydrolyzate.2 0. The time courses for total xylose (xylose monomer and xylooligomers). whereas it took only about 2 h to achieve the same conversion at 175 °C. 2009) and can be attributed to the postulate that xylan is composed of two fractions. and the mass fractions of total xylose. Tunc and van Heiningen. follows a slower rate. 160 °C (s). Nabarlatz et al. It can be observed from Fig. mannans and acetate are similar and. 2008). At 152 °C. about 6–9% of initial cellulose removal is observed with a maximum cel- . it is possible that mannose and acetate are formed at the same rate as xylose. and 160 °C () for aspen. The compositional analysis of wood hydrolyzates obtained for three different autohydrolysis experiments are given in Table 6. 175 °C (D) for sugar maple.6 0.0 80 0. 1999a. after a further reaction period due to the increased reaction severity. after achieving a removal of about 75–80%.0 0 1 2 3 0 1 2 3 0 1 2 3 0. mass fractions of total xylose. 167 °C (h). 1984. / Bioresource Technology 100 (2009) 6398–6406 100 1. their proportion in the hydrolyzate remains constant. During autohydrolysis. and therefore. a hydrolysable fraction which is susceptible to the hot water extraction with the other being an un-reactive fraction which does not show any affinity towards hydrolysis under normal operational conditions.. the percentage of cellulose remaining in the extracted wood chips was also calculated (Garrote et al. 5.0 0. The plausible causative mechanism for the biphasic behavior of hemicellulose solubilzation may be due to the presence of chemically and structurally different portions of hemicelluloses that can be differentiated on the basis of the degree of association between hemicelluloses and lignin (Tunc and van Heiningen.. their proportion in the extracted wood remains constant over time. a higher yield of xylose monomers is obtained due to the further hydrolysis of xylooligomers. 2004.1 0. and acetate in the extracted wood for all the autohydrolysis temperatures remain constant with hydrolysis time suggesting that the hydrolysis rates for xylan. Conner. Mittal et al. after reaching a maximum of about 65% of the initial xy- lan at 160 °C after 2 h. the fraction of residual xylan reached about 20% of the initial xylan in about 6 h. 7 that during the initial one hour of reaction time that PRGlc remains near 100% and.4 0. To study the impact of hydrothermal treatment on cellulose... Further. it starts decreasing due the degradation of xylose into furfural. ð2Þ where %GlcRM is the amount of cellulose in the raw material expressed as g glucose/100 g ODW and %Glc is the amount of cellulose in the treated material expressed as g glucose/100 g ODEW. mannose and acetate in the autohydrolysis hydrolyzate are shown in Fig. which is consistent with values reported in the literature (Garrote et al.6404 A.1 40 0.. 1999a). This phenomenon of an initially fast followed by a slow solubilization of xylan has been also observed in earlier studies (Carrasco and Roy. The hydrothermal treatment of sugar maple resulted in little degradation of cellulose. 2008. With a prolonged reaction time. mannan (mannose). 1992. PRGlc ¼ %Glc  SY %Glc RM . about 60–70% of the sugars remains in the oligomeric form (data not shown).3 20 Acetate Xylan remaining in extracted wood (% of initial) Xylose 0. Mittal et al. xylan solubilization initially follows a fast rate and.8 0. Mittal et al. 2002).3 Mannose 60 0. Yields and composition of the hemicelluloses remaining in the extracted wood chips at autohydrolysis temperatures of 152 °C (e). but it results in a lower overall yield of the desired reaction products due to the formation of degraded compounds like furfural via the dehydration reaction of xylose monomers (Jacobsen and Wyman. mannose. It is interesting to note from Fig. 6.2 0. 2009). therefore. 5 that the mass fractions of residual xylan (expressed as xylose).0 0 0 1 2 3 Time (h) Time (h) Fig. As mentioned earlier.

0 0 0 1 2 3 Time (h) Time (h) Fig. and 175 °C (D). Yields and composition of the hydrolyzate obtained at autohydrolysis temperatures of 152 °C (e). a plantation energy crop (Serapiglia et al. NMR Biomed.8 0. and 160 °C () for aspen. Wood Fiber Sci.0 0. Conner.. 105 Cellulose. Sundberg. 319–324..A. 160 °C (s). U.3 20 0. C. 69–75. Technol. 4. Roy. Khetrapal. Holmbom. Nagana Gowda. Although not discussed in this paper.6405 A. A. Misra. that result from genetic optimization of this biomass resource. this NMR procedure would be useful to determine the effect of other common pretreatments as well in the production of sugars and ethanol from biomass. Results have illustrated that this new technique has good repeatability and compares relatively well to established procedures for wood analysis for sugar maple and aspen wood. lignin which is solubilized during the autohydrolysis could also be quantified (approximately) from the intensity of phenyl proton resonances using NMR.2 0. Overall. 7. Bhandari.. Polym. C. 1984. 167 °C (h).H. Xylan removal by water prehydrolysis. 48. 167 °C (h). 26 (3). 1992. Kinetic modeling of hardwood prehydrolysis.0 0 1 2 3 0 1 2 3 0 1 2 3 0. and to determine the impact of electron beam pretreatment of lignocellulosic feedstocks on enzyme digestibility.. 189–207. Bertaud.. Kinetic studies on the autohydrolysis of hardwoods allowed selective solubilization of hemicelluloses and resulted in 75–80% removal of the initial xylan and little (5–7%) degradation of cellulose. % of initial 102 99 96 93 90 0 1 2 3 Time (h) 4 5 6 Fig. 2009).. lulose removal of about 7% observed at 160 °C after a reaction period of 4 h. F.0 40 Xylose 60 Xylooligomers composition (mass fraction) Mannose Total xylose (% of initial) 80 0.. Cellulose removed from sugar maple wood chips at autohydrolysis temperatures of 152 °C (e).3 0..6 0. A.4 0. 16 (2). F.2 Acetate 0. / Bioresource Technology 100 (2009) 6398–6406 100 1. 160 °C (s). it can be concluded that under the autohydrolysis conditions studied in this work up to 75–80% of the initial xylan could be solubilized with the degradation of only about 6–9% of initial cellulose.). Mittal et al. 268–277. 175 °C (D) for sugar maple. 1H NMR spectroscopic method for diagnosis of malabsorption syndrome: a pilot study. 6. M. .1 0. 2002. Part I. Lakshmi. The majority of the solubilized xylan (about 70%) in the hydrolyzates was in the oligomeric form. B. G. A. Conclusions The objectives of this study were to introduce a new NMRbased technique to quantify the carbohydrates present in solid wood and wood hydrolyzates and to use this technique to study the kinetics of wood autohydrolysis.. In principle. Carrasco. Ghoshal.1 0.. 2004..C.. References Bala.2 0. Evaluation of acid methanolysis for analysis of wood hemicelluloses and pectins.L. Kinetic study of dilute-acid prehydrolysis of xylancontaining biomass. Wood Sci. Carbohyd. We also exploit this method routinely to determine compositional changes for willow (Salix sp. 17.

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