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Article in Energy Sources Part A Recovery Utilization and Environmental Effects · April 2015
DOI: 10.1080/15567036.2011.601794
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To cite this article: S. Ceylan & S. Ünal (2015) The Saccharification of Hazelnut Husks to Produce
Bioethanol, Energy Sources, Part A: Recovery, Utilization, and Environmental Effects, 37:9, 972-979,
DOI: 10.1080/15567036.2011.601794
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Energy Sources, Part A: Recovery, Utilization, and Environmental Effects, 37:972–979, 2015
Copyright © Taylor & Francis Group, LLC
ISSN: 1556-7036 print/1556-7230 online
DOI: 10.1080/15567036.2011.601794
1
Bioengineering Department, Engineering Faculty, Marmara University, Kadıköy-Istanbul, Turkey
Biofuels have been produced industrially by fermentation of sugars derived from wheat, corn, sugar
beets, sugar cane, and molasses. But because of economic and environmental issues an efficient
substrate must be used for bioethanol production. Hazelnut husk, a renewable and low-cost natural
recourse, is not used industrially. The objective of this study was to examine the saccharification of
hazelnut husk for the production of fermentable sugars. Hazelnut husks were treated with sulphuric acid
for varying periods (30–90 min), with varying acid concentrations (1–5%, w/w) and sample concentra-
tions (1/5, 1/10, 1/15, 1/20 solid/liquid ratio). The maximum sugar yield of 18.63 ± 0.51% was obtained
with 90 min incubation time, 4% acid concentration, and 1/20 sample concentration. The sugar
composition of the acid hydrolyzed hazelnut husks was analyzed using thin layer chromatography.
Arabinose, glucose, mannose, and xylose were found to be present. Fermentability of the hyrolysate
was also tested.
Keywords: acid hydrolysis, agricultural residues, bioethanol, biomass, fermentation, hazelnut husk
1. INTRODUCTION
An increase in energy consumption due to rapid industrialization and exponential growth in the
population has caused a decline in fossil fuel reserves. Therefore, there is a great interest in
exploring alternative energy sources. Biofuels are attracting growing interest around the world,
and in order to reduce both greenhouse gas emissions and dependence on petroleum-based fuels,
some governments have been announcing commitments to bio-fuel programs. The European
Commission has indicated that biomass will play an important role in the future (Balat and Balat,
2009). Bioethanol production from lignocellulosic materials, such as agricultural residues, has
gained interest because of its strategic and economic benefits and attractive characteristics of
lignocellulosic biomass. A variety of biomass feedstocks can be used for bioethanol production,
e.g., mill wastes, urban wastes, agricultural residues, and forest residues (Balat and Balat, 2009;
Mohanty et al., 2009). Among these resources, agricultural residues dominate in terms of tonnage
and afford a renewable and low cost raw material for bioethanol production (Balat and Balat,
2009; Kadam and McMillan, 2003). Current ethanol production processes generally use crops,
such as sugar beet, sugar cane, and corn. Usage of a cheaper and more abundant substrate could
render bioethanol more competitive in comparison to fossil fuels, but processing and usage of the
Address correspondence to Dr. S. Ceylan, Ondokuz Mayıs University, Chemical Engineering Department, 55139,
Samsun, Turkey. E-mail: selimceylan@marmara.edu.tr
972
SACCHARIFICATION OF HAZELNUT HUSKS FOR BIOETHANOL 973
cheaper substrate is complex, and differs in many aspects from the crop-based ethanol production
(Sanchez et al., 2003). It is also very important to ensure a high degree of utilization of all the
carbohydrate components in the feedstock and to attain a high yield of both glucose and
hemicellulosic sugar (Söderström et al., 2003). However, because of pre-treatment of biomass
feedstock and optimization of the hydrolysis process, there is a lack of cost-competitive bioetha-
nol production process.
Turkey is a major hazelnut producer. The residues of hazelnut are not used industrially. In the
industrial processing of hazelnut crops either for oil or nuts, large amounts of husks are generated.
Because of their large storage volumes, they present a disposal problem. Being basically of
lignocellulosic material, hazelnut husks can be utilized as a renewable and low-cost raw material
for the production of fermentable sugars. This article is a report of the study conducted on
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saccharification of hazelnut husk as a feedstock for bioethanol fermentation. The effects of acid
hydrolysis on bioconversion of hazelnut husk to sugar at laboratory-scale were investigated. As far
as it is known, no study about the saccharification of hazelnut husk and fermentability of hazelnut
husk hydrolysate has yet been reported.
Hazelnut husks (HH), an agricultural residue especially abundant biomass in Turkey, used in
this work were supplied by the growers in northern Turkey at the end of the 2004 harvest.
Chemicals used in this work were supplied by Merck AG (Darmstadt, Germany) and by Sigma
Chem. Ltd. (USA). S. cerevisiae FY23 strain used for ethanol production was kindly supplied
by the Chemical Engineering Department, Bosphourus University, Istanbul, Turkey. HH was
ground and samples were washed twice with tap water and once with distilled water, and
cloth filtered after each washing. The solids were then dried at 70°C overnight to constant
weight.
intervals and analyzed for growth of S. cerevisiae FY23 strain and ethanol production. Growth of
microorganism was measured at 600 nm by a spectrometer (Perkin Elmer, USA) using medium as
blank. The experiments for growth of S. cerevisiae FY23 strain and ethanol production were
performed in duplicate. Mean values are used in the figures shown.
nose, fructose, glucose, maltose, mannose, rafinose, rhammnose, and xylose. Solutions of sugars
listed above (0.02 g/ml) were obtained by dissolving 0.100 g of sugar in 5 ml 0.02% (w %) sodium
azide. Each solution was diluted to 1/20 by taking 200 μl from the sugar solution and completing it
with 0.02% (w %) sodium azide to 4 ml in a test tube. A TLC layer with 20 cm length marked with
a pencil and 1 μl from each standard sugar solution and hydrolyzate was injected on marked places
using a Hamilton syringe. After the addition of all samples, the layer was dried. The dried layer
was immersed in 85:15 (v/v) acetonitrile-water solution. At the end of the run, the layer was taken
from the solution and dried. The dried layer was placed in the solution again. This procedure was
repeated three times. At the end of the third run, the layer was dried again and dipped in 0.05 g/ml
H2SO4–0.005 g/ml α-naphthol solution once. A wet layer was placed on a hot plate at 110°C for 10
min to visualize the sugar bands. Ethanol concentration in the fermentation media was determined
by using the Ethanol UV-Test Kit of Roche (R-Biopharm, Roche) as described by the manufac-
turer’s instructions.
FIGURE 1 Variation of sugar yield from acid hydrolysis of HH for (a) 30 min, (b) 60 min, and (c) 90 min
incubation period (▲: 1/5; □: 1/10; ●: 1/15; Δ: 1/20 sample concentration).
976 S. CEYLAN AND S. ÜNAL
20
1% acid conc.
18
2% acid conc.
Sugar yield (w%)
16 3% acid conc.
4% acid conc.
14
5% acid conc.
12
10
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8
0 30 60 90
Hydrolysis period (min)
FIGURE 2 Variation of sugar yield from acid hydrolysis of HH with different hydrolysis period at 1/20 sample
concentration.
for 1/20 sample concentration and 4% acid concentration. This yield was also the maximum
sugar yield obtained from the acid hydrolysis of HH. The reducing sugar concentration in the
hydrolyzate at this point was 7.0 g/L. The sugar yield increased with decreasing sample concentra-
tion. For 1/20 sample concentration the sugar yield was overall higher than the other sample
concentrations for 30-, 60-, and 90-min hydrolysis periods.
Figure 2 shows the change in sugar yield with hydrolysis period. As the sample concentration
decreased, a better extraction was obtained. Roberto et al. (2003) studied the effect of sample
concentration on sugar yield and they observed similar behavior. For the 1/20 sample concentra-
tion, as the hydrolysis period increases the sugar yield increased for all acid concentrations.
However, the rate of increase in sugar yield with time is higher between 60 to 90 min than that
observed up to 60 min. The best yield 18.63 ± 0.51% was obtained at 4% acid concentrations for
the 90-min hydrolysis period. Sun and Cheng. (2005) observed the increase in sugar yield with
hydrolysis period in their study on both Bermuda grass and rye straw. They obtained the best sugar
yields after the 90-min acid hydrolysis period, in which Liao et al. (2004) also observed the same
behavior on the acid hydrolysis of dairy manure. The decrease in sugar yield in the present study
between 4–5% acid concentration (Figures 1a, 1b, 1c) can be explained by the degradation of
reducing sugars (Choteborska et al. 2004). For example, xylose in the hydrolyzate can be further
degraded to produce furfural. Liao et al. (2004) and Iranmahboob et al. (2002) explained the
decrease in sugar yield with the increase in acid concentration by degradation of hemicellulose and
cellulose. The acid hydrolyzates obtained from HH were in a red/brown color. This is considered to
be due to the dehydrating and oxidizing actions of sulphuric acid on the different hydrolytic
products. Formation of furfural from xylose has been suggested as another reason (Agu et al.,
1997). Liao et al. (2004) also observed the red/brown color of the solutions and suggested that this
color may be due to reaction between reducing sugars and protein.
4
(a) Control medium
1
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0
0 20 40 60 80
Time (h)
12
(b) Control medium
10
Ethanol concentration (g/L)
0
0 20 40 60
Time (h)
FIGURE 3 Growth curve of S. cerevisiae FY23 (a) and time course of ethanol production (b) in the acid
hydrolyzate and control medium.
reported that the soluble carbohydrate part of lignocellulosic materials of agricultural residues is
composed of mostly glucose and xylose. Minor constituents are arabinose, galactose, and rham-
nose. Liao et al. (2004) and Saha et al. (2005) also found arabinose, galactose, xylose, and glucose
in their acid hydrolyzates of cellulosic materials. Choteborska et al. (2004) reported that xylose,
arabinose, galactose, and glucose were found in acid hydrolysate of wheat bran.
increased rapidly after 12 h. Maximum ethanol production was obtained after 28 h in acid
hydrolyzate medium. Ethanol production was 7.65 g/L in acid hydrolyzate medium and 9.20 g/L
in control medium after 28 h. Ethanol concentrations were lower than the control mediums. The
poor fermentability is considered to be primarily due to the presence of toxic components, which
inhibited the growth and fermentation activity of the yeast (Nigam, 2001). Lag phase before the
exponential phase is due to inhibitors also (Palmarola-Adrados et al., 2004). The most important
inhibitors are furfural and 5-hydroxymethyl-2-furaldehyd (HMF). Pentose degradation results in
increased yield of furfural. HMF is the degradation product of hexoses under more severe
pretreatment conditions. These substances were found to be toxic for yeasts; their presence in
the slurry causes insufficient conversion of sugars to ethanol, or even total inhibition of fermenta-
tion (Taherzadeh et al., 2000; Choteborska et al., 2004).
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4. CONCLUSION
The improvement in bioconversion of wastes into valuable chemicals and fuels will provide
reduced pollution problems and valorization of wastes. Lignocellulosic wastes can be thought as
a major resource for bioethanol production. In this study, hydrolyzates of lignocelluloic waste
biomass hazelnut husk used for bioethanol production. Results showed that his type of waste has a
potential as a resource for bioethanol production.
ACKNOWLEDGMENT
The authors wish to thank Prof. Dr. Dilek Kazan for reviewing the manuscript.
FUNDING
This research was funded by Marmara University through the project BSE-071/131102.
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