Industrial Crops & Products 108 (2017) 598–603

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Industrial Crops & Products

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Evaluation of ethanol production from sugar and lignocellulosic part of MARK

energy cane
⁎
Sutticha Na-Ranong Thammasittironga,b, , Prasert Chatwachirawongc, Thada Chamduanga,
Anon Thammasittironga,b
a
Department of Microbiology, Faculty of Liberal Arts and Science, Kasetsart University, Kamphaeng Saen Campus, Nakhon Pathom 73140, Thailand
b
Microbial Biotechnology Unit, Faculty of Liberal Arts and Science, Kasetsart University, Kamphaeng Saen Campus, Nakhon Pathom 73140, Thailand
c
Department of Agronomy, Faculty of Agriculture at Kamphaeng Saen, Kasetsart University, Kamphaeng Saen Campus, Nakhon Pathom 73140, Thailand

A R T I C L E I N F O A B S T R A C T

Keywords: Energy cane is considered an ideal energy crop because it produces readily fermentable sugars and high yields of
Energy cane juice lignocellulosic biomass. Four improved clones of the energy cane series “TByEFC” (Tiphuyae and Banyang
Energy cane bagasse Energy and Forage-cane Clone), which was developed at the Kasetsart University Kamphaeng Saen campus in
Lignocellulosic biomass Thailand, were evaluated in terms of their potential ethanol yields from both sugar juice and bagasse. Juice from
Ethanol production
TByEFC05-1558 and TByEFC04-1208 displayed the highest total sugar production, ranging from
Energy crop
171.03–179.72 g/L, and reached the maximum ethanol production, ranging from 78.02–82.56 g/L, without any
nutrient supplementation at 37 °C using Saccharomyces cerevisiae ND48. All four energy cane clones showed a
maximum ethanol concentration from bagasse in the range of 9.11–10.68 g/L. The combination of agronomic
productivity and ethanol production yield indicated that the highest ethanol yield from juice was reached with
TByEFC04-1155 (2697.00 kg/ha), while the highest ethanol yield from bagasse was reached with TByEFC09-
0098 (1695.45 kg/ha). Among the tested clones, TByEFC04-1155 was shown to be useful for combined ethanol
production from both juice and bagasse, yielding 3923.48 kg/ha. This energy cane clone combines good agro-
nomic features for ethanol production from sugar substances and lignocellulosic biomass, making it an attractive
energy crop for the ethanol industry.

1. Introduction
With the depletion of limited fossil fuel stocks, countries worldwide
have been compelled to search for new energy sources to substitute for
petroleum following the “fossil fuel crisis” in the 1970s. In Thailand,
ethanol plays an important role in renewable energy for transport ac-
cording to government policy. Ethanol has been used as a substitute for
conventional gasoline by blending ethanol with gasoline (called ga-
sohol), i.e., E10, E20 and E80. E10 gasohol, made by blending 10%
ethanol and 90% octane 95 gasoline, was introduced into the market in
2004; E20 and E85 were introduced in 2008. Due to government pro-
motion strategies, the total ethanol consumption in Thailand has in-
creased from 1.2 million liters per day (M L/day) in 2010–3.2 M L/day
in 2014 (AEDP, 2015). The Thai renewable energy policy has set a
target to increase ethanol production to 11.3 M L/day by 2022. Cur-
rently, sugar cane molasses and cassava starch are the main raw ma-
terials for ethanol production in Thailand; however, these feedstocks
cannot sufficiently meet the policy production targets. In addition,
sugar cane molasses can be used as a raw material for several fer-
mentation industries, which can lead to an increase in the cost perfor-
mance of ethanol. Therefore, diverse feedstock sources are required for
ethanol production.
Currently, with the growing need for alternative sources of energy,
lignocellulosic biomass is a natural, abundant and renewable resource
that is a promising feedstock for achieving the Thai government’s en-
ergy policy goals. The government of the USA considers the use of
biomass for ethanol production and electricity generation as important
for reducing the use of fossil fuels, and the European Union has a plan
to supply 20% of its total energy needs with biomass energy by 2020
(Matsuoka et al., 2012). Thailand is a tropical, agriculture-based
country where a great effort has been applied to develop sugar cane as a
potential energy crop through sugar cane breeding programs. “Energy
cane” results from effectively selecting sugar cane that has a higher
fiber yield than sugar yield, making it more vigorous and rustic. These
characteristics bring a series of economic and environmental ad-
vantages (Carvalho-Netto et al., 2014; Kim and Day, 2011). Energy

⁎
Corresponding author at: Department of Microbiology, Faculty of Liberal Arts and Science, Kasetsart University, Kamphaeng Saen Campus, Nakhon Pathom 73140, Thailand.
E-mail addresses: sutticha.n@ku.ac.th (S.N.-R. Thammasittirong), tiphuyae@live.com (P. Chatwachirawong), kaatoa@gmail.com (T. Chamduang),
faasant@ku.ac.th (A. Thammasittirong).

http://dx.doi.org/10.1016/j.indcrop.2017.07.023
Received 10 November 2016; Received in revised form 30 June 2017; Accepted 10 July 2017
Available online 21 July 2017
0926-6690/ © 2017 Elsevier B.V. All rights reserved.
S.N.-R. Thammasittirong et al.
cane is considered an ideal energy crop because it produces readily
fermentable sugar juice and a high yield of lignocellulosic biomass.
After the juice extraction process, the bagasse can be used either as a
feedstock for cellulosic ethanol production or as a raw material for
thermal energy in electricity plants. Sugar cane breeders in the De-
partment of Agronomy, Faculty of Agriculture at Kamphaeng Saen,
Kasetsart University, Kamphaeng Saen campus, Thailand, have at-
tempted to breed the energy cane series “TByEFC” (Tiphuyae and Ba-
nyang Energy and Forage-cane Clone) by crossing sugar cane (Sac-
charum spp.) with wild cane (S. spontaneum). Compared with sugar
cane, the energy cane series TByEFC contains a higher fiber content, has
better tolerance to insects and diseases and requires less fertilizer and
less water input. In addition, the crop cycle of the energy cane series
TByEFC is 8–10 months, shorter than that of sugar cane, which requires
a harvesting period of 12 months (Chatwachirawong et al., 2009).
Much of the research to date has focused on the lignocellulosic part
of energy cane as a bioethanol feedstock. Pretreatment methods and
optimizations have been reported for maximum enzymatic hydrolysis
and ethanol yield from energy cane bagasse (Aita et al., 2011; Qiu et al.,
2012, 2013; Oladi and Aita, 2017). However, for economic ethanol
production, the juice, fiber and cane yields per cultivated area are
important parameters that are associated with ethanol production. In
this current study, four improved TByEFC energy cane clones (with
higher biomass, juice and total sugar contents) and one commercial
sugar cane variety, LK92-11 (reference material), were evaluated for
their efficiency in ethanol production from both the juice and bagasse.
This study is the first to report an evaluation of ethanol production from
the juice and bagasse of energy cane at the laboratory scale in combi-
nation with their agronomic productivity. Additionally, the chemical
compositions of the energy cane juice and bagasse are also reported.
2. Materials and methods
2.1. Raw materials and preparation of samples
The experiment was conducted from October 2014 to March 2016
on silt loam soil texture at the Field Crops Building, Kasetsart
University, Kamphaeng Saen campus, Thailand. A randomized com-
plete block design with 4 replications was used. Four energy cane
clones (TByEFC04-1155, TByEFC04-1208, TByEFC05-1558, and
TByEFC09-0098) and a commercial sugar cane variety (LK92-11, used
as a reference material) were harvested 10 months after planting. Each
plot consisted of 5 adjacent 1.50-m-wide rows (8 m long) with a 1.50-m
alley between each plot. The plants in the 3 middle rows were manually
cut at the ground level and topped at their natural break point for yield
determination. At the same time, 10 sampling cane stalks were crushed
twice in a small local roller press immediately after being cut. The juice
was filtered through cheesecloth and kept at −20 °C for further studies.
The cane yield was calculated from the millable stalk weight of the 3
middle rows. The juice volume was used to calculate the juice yield.
The sugar yield was calculated from juice volume and total sugar
content (sucrose, glucose and fructose), whereas the fiber yield was
calculated from the cane yield and fiber content.
2.2. Physical and chemical composition determination of the energy cane
juices
The energy cane juices were measured for total soluble solids (TSS,
°Brix) using a refractometer (N.O.W., Japan). Sugars in the juices were
quantified using a High Performance Liquid Chromatography (HPLC)
system (Water 600E, Waters Inc., USA) with a refractive index detector
and a sugar pak I column at 85 °C. The mobile phase was deionized
water at 0.5 mL/min (Senatham et al., 2016). Free amino nitrogen
(FAN) was determined using the ninhydrin official method with glycine
as the standard (AOAC, 1980). The total phenolic content in the juices
was determined with the Folin-Ciocalteu reagent using gallic acid as the
599
Industrial Crops & Products 108 (2017) 598–603
standard (Vinson et al., 2001).
2.3. Fermentation of the energy cane juice
Saccharomyces cerevisiae ND48 cells (with high ethanol production
from sucrose, unpublished data) pre-cultivated overnight in YPD broth
(2% glucose, 1% yeast extract and 2% peptone) were inoculated into
energy cane juice medium without nutrient supplementation and pH
adjusted to 5.0 with 0.1 N NaOH. The initial cell concentration was
adjusted to a cell density of 5 × 105 cells/mL. Fermentation was per-
formed on a rotary shaker at 100 rpm and 37 °C for 72 h. Samples were
withdrawn every 12 h to determine the cell density at 600 nm (OD600)
using UV–vis spectrophotometer (GENESYS™ 10S UV–vis, Thermo
Scientific, USA). The concentrations of ethanol and residual sugar were
monitored using HPLC.
2.4. Chemical composition determination of energy cane bagasse
The energy cane bagasse chemical composition was determined
using a slight modification of the standard procedure developed by the
National Renewable Energy Laboratory (NREL), as detailed in Sluiter
et al. (2008). In brief, 2–10 g of a milled, dried bagasse sample was
extracted with 95% ethanol for 12 h using a Soxhlet apparatus. A
sample of 0.3 g of dried extractive-free material (moisture below 10%)
was treated with 3 mL of 72% H2SO4 at 30 °C for 60 min. The acid was
diluted to 4% H2SO4 by adding 84 mL of deionized water, and the
mixture was then autoclaved at 121 °C for 60 min. After cooling to
room temperature, the mixture was filtered, and the solid fraction was
rinsed with warm deionized water until a neutral pH was recorded,
followed by oven drying at 105 °C to a constant weight. The material
was then heated in a furnace set at 575 °C for 24 h. The weight dif-
ference before and after incineration was considered to be the acid
insoluble lignin in the bagasse. The soluble lignin-in-liquid fraction was
determined from absorbance measurements at a wavelength of 280 nm
using UV–vis spectrophotometer. The sugar concentrations were ana-
lyzed using HPLC with a sugar pak I column and refractive index de-
tector as described above. Furfural and 5-hydroxymethyl furfural
(HMF) were eluted with 20% acetonitrile in deionized water (80%) at a
flow rate of 1.1 mL/min. Acetic acid was eluted with 1% acetonitrile in
0.05 M KH2PO4 (99%) at a flow rate of 1.0 mL/min (Senatham et al.,
2016).
The concentrations of glucose, cellobiose and HMF were used for
determination of the cellulose content, while the concentrations of
xylose, arabinose, glucuronic acid, acetic acid and furfural were used
for determination of the hemicellulose content. The cellulose content
and hemicellulose content were calculated using the formulas used by
the NREL (Sluiter et al., 2008).
2.5. Preparation of energy cane bagasse hemicellulosic hydrolysate using
dilute-acid hydrolysis
The milled bagasse was dried at 60 °C for 24 h. The dried bagasse in
a solid-liquid ratio of 1:10 was soaked in 1% H2SO4 for 30 min at
ambient temperature and then autoclaved at 121 °C for 30 min. The
hydrolysate was filtered and neutralized with CaO to pH 5.5. The
precipitate that formed was removed using centrifugation and filtration
(Senatham et al., 2016). Detoxification of energy cane bagasse hemi-
cellulosic hydrolysate was performed with slightly modified method
described by Branco et al. (2011). The neutralized hemicellulosic hy-
drolysate was treated with 2% (w/v) activated charcoal with stirring at
150 rpm for 1 h at 30 °C. The solids were then removed using vacuum
filtration.
2.6. Fermentation of the energy cane bagasse hemicellulosic hydrolysate
Scheffersomyces shehatae TTC79 (Senatham et al., 2016) cells pre-
S.N.-R. Thammasittirong et al. Industrial Crops & Products 108 (2017) 598–603

cultivated overnight in YPX broth (2% xylose, 1% yeast extract and 2%
peptone) were inoculated into the hydrolysate medium supplemented
with 5 g/L KH2PO4, 2 g/L (NH4)2SO4, 0.2 g/L MgSO4·7H2O, 1 g/L
peptone and 5 g/L yeast extract, pH 5.5. The initial cell concentration
was adjusted to a cell density of 1 × 107 cells/mL. Ethanol production
was performed at 30 °C and 100 rpm for 72 h. Samples were withdrawn
every 12 h to determine the cell density at 600 nm (OD600) using
UV–vis spectrophotometer. The concentrations of ethanol and residual
sugar were monitored using HPLC.
2.7. Statistical analysis
All recorded data were subjected to analysis of variance, using the
SPSS 16.0 statistical package (SPSS Inc., Chicago, USA). Least sig-
nificant difference (LSD) test at 5% level of significance (p < 0.05) was
used to compare means.
this nutrient content in energy cane juice.
The other parameter used in the energy cane juice characterization
was the total phenolic content. The phenolic compounds have been
found to have negative effects on microbial growth and fermentation
(Heipieper et al., 1994; Orozco et al., 2012). The ability of phenolic
acids to form complexes with proteins and inhibit enzyme activity is
well known (Salunkhe et al., 1982). Mullins and Lee (1991) reported
that a level of 3.00 g/L of phenolic compounds is the critical con-
centration that allows the precipitation of more than 70% of the pro-
teins in the fermentation media. The total phenolic acid concentrations
of all of the energy cane juices were low, with 0.56–0.87 g of gallic acid
equivalent (GAE)/L, similar to the amounts observed for sugar cane
juice in this study as well as those reported by Payet et al. (2006)
(0.83 g GAE/L) and Manohar et al. (2014) (0.45–0.90 g GAE/L).
3.2. Chemical composition of energy cane bagasse

3. Results and discussion
3.1. Chemical composition of the energy cane juice
The chemical composition of the energy cane juice from the four
energy cane clones, TByEFC04-1155, TByEFC04-1208, TByEFC05-1558
and TByEFC09-0098, and the commercial sugar cane variety, LK92-11,
used as a reference material in this study, were characterized, and the
results are shown in Table 1. In this study, the TSS and total sugars
detected in the energy cane juice samples were between 16.2–18.4°Brix
and 153.48–179.72 g/L, respectively, while the corresponding values
for the sugar cane juice samples were 19.0°Brix and 187.40 g/L, re-
spectively. The energy cane clones TByEFC04-1208 and TByEFC05-
1558 had the highest total sugar contents (171.03–179.92 g/L), fol-
lowed by TByEFC04-1155 (157.48 g/L) and TByEFC09-0098
(153.48 g/L). The energy cane variety L79-1001(L) was reported to
contain TSS between 10.0–12.0°Brix and total sugars of 98.00 g/L (Kim
and Day, 2011). Energy cane varieties Ho-02-113 and HoCP72-114
were found to contain TSS of 9.0 and 10.1°Brix, respectively (Hale,
2010; Gravois et al., 2010). Rao et al. (2007) reported that energy cane
juice varieties WI81456, WI79458 and WI79460 contained TSS values
of 12.2, 14.0 and 14.2°Brix, respectively. While it is difficult to directly
compare the juice chemical compositions from different studies, it is
still a useful indicator of the competitiveness of the energy cane clones
TByEFC04-1208 and TByEFC05-1558, which could improve the yield of
high fermentable sugars used in the ethanol and sugar industries.
The initial concentration of soluble nitrogenous compounds mea-
sured as free amino nitrogen (FAN) was also characterized in the energy
cane juices. This nitrogen source plays significant roles in yeast growth
and fermentation efficiency (Thomas and Ingledew, 1990). In this
study, energy cane juice from TByEFC05-1558 showed the highest FAN
level (184.74 mg/L), while TByEFC04-1155 showed the lowest FAN
level (74.90 mg/L). Energy cane juice from TByEFC04-1208 and
TByEFC09-0098 and sugar cane juice from LK92-11 showed FAN values
in the range 103.20–136.92 mg/L. Liliana and Silvana (2014) reported
that the FAN level of sugar cane juice variety CC8592 was 45 mg/L.
Unfortunately, no other previous reports were available that monitored
The chemical composition of untreated energy cane bagasse sam-
ples is summarized in Table 2. The contents of cellulose, hemicellulose,
lignin and other components (e.g., ash and extractive) were in the range
of 40.00–45.72%, 25.39–30.05%, 12.04–17.51% and 11.59–15.78%,
respectively. These values were similar to those found for sugar cane
bagasse and energy cane bagasse reported in the literature, as shown in
Table 2.
3.3. Agronomic properties of energy cane
The average agronomic properties of the four energy cane clones are
shown in Table 3. All of the energy cane clones differed significantly in
their juice, sugar, fiber and cane yields. Among the energy cane clones,
TByEFC04-1155 had the highest juice yield (37,200 L/ha) and sugar
yield (5.86 t/ha), which were similar to the values of LK92-11 of
36,600 L/ha and 6.86 t/ha, respectively.
The highest fiber yield was from energy cane clone TByEFC09-0098
(16.20 t/ha), which is comparable to the average yield of sugar cane
(HoCp 96-540, L99,266, L99-233) and energy cane (L79-1002) planted
in three planting dates (15.90–18.90 t/ha) (Viator and Richard, 2012).
The lowest fiber yields was from TByEFC04-1208 (5.82 t/ha). For cane
yield, the energy cane clones TByEFC04-1155 and TByEFC05-1558
showed the highest cane yields (85.56–85.88 t/ha), followed by
TByEFC09-0098 (57.15 t/ha). Salassi et al. (2014) reported similar cane
yields from five energy cane varieties, ranging from 57.20–99.00 t/ha.
Energy cane Ho02-113 also showed a similar cane yield (89.00 t/ha)
(USDA-ARS, 2010).
The commercial sugar cane variety, LK92-11, is one of the most
widely cultivated varieties in Thailand, and it is used for sugar manu-
facture due to its good agronomic characteristics including high cane
yield, high tiller number and high sugar content (Thongpaiyai et al.,
2012; Tippayawat et al., 2012). The agronomic properties of
TByEFC04-1155 showed high juice and sugar yields similar to LK92-11
but also contained a higher fiber content. These results suggest that
TByEFC04-1155 combines good agronomic features for ethanol pro-
duction from its sugar and lignocellulosic components.

Table 1
Chemical composition of the energy cane juices and sugar cane juice.

Clone °Brix Free amino nitrogen (mg/L) Total phenolics (g GAE/L) Sucrose (g/L) Glucose (g/L) Fructose (g/L) Total sugars (g/L)

c e c e a a
TByEFC04-1155 16.3 ± 0.7 74.90 ± 2.16 0.56 ± 0.01 137.69 ± 1.31 11.59 ± 2.40 8.20 ± 1.53 157.48 ± 0.83c
TByEFC04-1208 18.4 ± 0.6a 120.24 ± 3.29c 0.87 ± 0.01a 165.77 ± 3.13b 9.59 ± 1.77ab 4.39 ± 0.27c 179.72 ± 3.60b
TByEFC05-1558 17.2 ± 0.4b 184.74 ± 1.90a 0.75 ± 0.03b 159.18 ± 3.73c 7.70 ± 0.95b 4.16 ± 0.24c 171.03 ± 4.36b
TByEFC09-0098 16.2 ± 0.7c 103.20 ± 1.63d 0.75 ± 0.04b 142.92 ± 3.50d 6.68 ± 1.52b 3.89 ± 0.27c 153.48 ± 3.93d
LK92-11 19.0 ± 0.8a 136.92 ± 1.25b 0.83 ± 0.02a 171.34 ± 1.12a 9.97 ± 1.84ab 6.08 ± 0.24b 187.40 ± 2.03a

Different lowercase superscript letters in a column indicate significant differences between the clones (p < 0.05).

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S.N.-R. Thammasittirong et al. Industrial Crops & Products 108 (2017) 598–603

Table 2
Chemical composition on a % dry basis for untreated bagasse from the energy canes and sugar cane.

Clone/Variety Cellulose Hemicellulose Lignin Other References

TByEFC04-1155 42.75 ± 2.00bc 30.05 ± 2.90a 15.60 ± 0.5b 11.59 ± 1.28b In this study
TByEFC04-1208 45.72 ± 0.30a 25.39 ± 1.39a 14.19 ± 0.54b 14.69 ± 1.00ab In this study
TByEFC05-1558 44.86 ± 0.34b 27.31 ± 1.01a 12.04 ± 0.12c 15.78 ± 1.45a In this study
TByEFC09-0098 40.00 ± 1.20c 27.84 ± 1.09a 17.51 ± 0.19a 14.63 ± 0.45a In this study
LK92-11 40.04 ± 1.78c 31.30 ± 3.26a 14.92 ± 0.59b 13.72 ± 1.12ab In this study
RB92579a 39.07 31.02 12.89 0.88 (ash value) Pereira et al. (2015)
NRa 45.00 25.80 19.10 10.10 Canilha et al. (2011)
NRa 41.60 25.10 20.30 NR Kim and Day (2011)
L79-1001Lb 43.30 23.80 21.70 NR Kim and Day (2011)
NRb,c 44.20 21.10 23.50 NR Ogata (2013)

Different lowercase superscript letters in a column indicate significant differences between the clones (p < 0.05).
Mean ± standard deviation (n = 3).
NR: Not reported.
a
Sugar cane.
b
Energy cane.
c
Mean value of 207 energy cane clones.

3.4. Ethanol production from energy cane
Energy cane, including juice and bagasse, were assessed for ethanol
production, and the results are shown in Fig. 1 and Table 4. Nutrient
supplementation has been shown to be required for increased ethanol
production under very high gravity (VHG) conditions (Nuanpeng et al.,
2011). Thus, due to the TSS (16.2–18.4°Brix) and total sugar
(153.48–179.72 g/L) values of the energy cane juices (Table 1), ethanol
production in this study was evaluated without any nutrient supple-
mentation in the energy cane juice.
The time profiles of the total sugar, ethanol and FAN concentrations
and cell growth are shown in Fig. 1. The total sugar contents of all of
the energy cane juices were almost completely consumed within 36 h of
fermentation (Fig. 1A). The sugars seemed to be efficiently utilized and
fermented to ethanol by S. cerevisiae ND48. The maximum ethanol
production from all of the juices was obtained at 36 h of fermentation
(Fig. 1B). As expected, the ethanol yield was correlated with the initial
concentration of sugars in each of the energy cane juice clones. Juice
from TByEFC04-1208, which contained the highest total fermentable
sugars, produced the maximum ethanol concentration and theoretical
yield at 37 °C using S. cerevisiae ND48 with values of 82.56 g/L, and
89.29%, respectively (Table 5).
The initial FAN content in all of the four energy cane juices was in
the range of 74.90–184.74 mg/L (Table 1), but at the beginning of
fermentation, the contents were approximately 84.63–196.61 g/L
(Fig. 1C). The increased FAN contents in the energy cane juice culture
media is related to the use of YPD broth for yeast propagation. The YPD
medium contained 10 g/L of yeast extract and 20 g/L of peptone, which
can increase the initial concentration of nitrogenous compounds.
Barredo-Moguel et al. (2001) and Davila-Gomez et al. (2011) also re-
ported that sorghum juice inoculated with yeast cultured in malt-yeast
extract media displayed increased FAN levels. In the current study, FAN
was available in the medium until the end of the fermentation. Residual
FAN was detected in the range of 9.47–25.60 mg/L at 60 h of fermen-
tation. These results revealed that energy cane juices contain sufficient
FAN for yeast growth and fermentation, and consequently, nitrogen
supplementation may be not necessary to achieve improved fermenta-
tion performance using S. cerevisiae ND48. This present finding in-
dicates economic advantages for industrial ethanol production using
these energy cane juices as feedstocks.
To evaluate energy cane for lignocellulosic ethanol production, all
of the bagasse samples of the energy cane clones were hydrolyzed using
the dilute-acid method. The sugars and inhibitors released during acid
hydrolysis depend on the type of biomass and the operating conditions,
temperatures, acid concentrations and reaction times used (Almeida
et al., 2007; Canilha et al., 2011). The conditions for sugar extraction
were 1% H2SO4 with a solid to liquid ratio of 1:10 (w/v) at 121 °C for
30 min, followed by treatment with activated charcoal, which resulted
in levels of xylose (11.41–14.52 g/L), glucose (6.22–9.71 g/L) and
arabinose (5.98–9.58 g/L) in the hydrolysate. For stress inhibitors, the
bagasse hydrolysate medium contained 5.34–5.50 g/L acetic acid,
0.16–0.18 g/L furfural and 1.12–1.15 g/L HMF (data not shown). The
maximum ethanol concentrations, ethanol yields, theoretical yields and
ethanol productivities in the ranges of 9.11–10.68 g/L, 0.44–0.46 g/g,
87.37–90.00% and 0.25–0.30 g/L/h, respectively, were observed for all
of the four energy cane bagasse hemicellulosic hydrolysates and in the
sugar cane bagasse hemicellulosic hydrolysate fermented using Sch.
shehatae TTC79 at 30 °C.
For economic ethanol production, the juice, fiber and cane yields
per cultivated area are important parameters that are associated with
ethanol production. In this study, significant differences in ethanol
yield per area were noted among the energy cane clones. Sugar cane
LK92-11 showed the highest ethanol production from juice
(3087.94 kg/ha) followed by TByEFC04-1155 (2697.00 kg/ha) and
TByEFC05-1558 (2148.67 kg/ha). For ethanol production from ba-
gasse, TByEFC09-0098 had the highest ethanol production per area
(1695.45 kg/ha) followed by TByEFC04-1155 (1226.48 kg/ha).
The combination of agronomic productivity and ethanol production
yield from juice and bagasse indicated that the highest overall ethanol
yield was achieved with TByEFC04-1155 (3923.48 kg/ha), followed by

Table 3
Agronomic characteristics of the energy canes and sugar cane.

Clone Juice yield (L/ha) Sugar yield (t/ha) Fiber yield (t/ha) Cane yield (t/ha)

a a c
TByEFC04-1155 37,200.00 ± 786.39 5.86 ± 0.34 12.58 ± 1.40 85.56 ± 7.12b
TByEFC04-1208 11,017.50 ± 254.71d 1.98 ± 0.09c 5.82 ± 1.10e 28.82 ± 5.49d
TByEFC05-1558 27,540.00 ± 546.94b 4.71 ± 0.19b 15.28 ± 0.30b 85.88 ± 1.71b
TByEFC09-0098 14,605.00 ± 594.57c 2.24 ± 0.02c 16.20 ± 0.70a 57.15 ± 4.41c
LK92-11 36,600.00 ± 760.00a 6.86 ± 0.63a 8.72 ± 0.24d 97.63 ± 2.69a

Different lowercase superscript letters in a column indicate significant differences between the clones (p < 0.05).

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Table 4
Ethanol production from the bagasse of energy canes and sugar cane using Sch. shehatae TTC79 at 30 °C.

Clone Ethanola (g/L) Ethanol yieldb (gp/gs) Theoretical yieldc(%) Productivity (g/L/h)

TByEFC04-1155 9.42 ± 0.88ab 0.44 ± 0.01a 87.37 ± 2.29a 0.26 ± 0.03ab

TByEFC04-1208 9.11 ± 0.11b 0.45 ± 0.00a 89.14 ± 0.82a 0.25 ± 0.00b
TByEFC05-1558 10.37 ± 0.13a 0.45 ± 0.01a 88.43 ± 2.26a 0.29 ± 0.00a
TByEFC09-0098 10.68 ± 0.50a 0.46 ± 0.01a 89.97 ± 1.58a 0.30 ± 0.01a
LK92-11 10.34 ± 0.20a 0.46 ± 0.02a 90.00 ± 3.45a 0.29 ± 0.00a

Different lowercase superscript letters in a column indicate significant differences between the clones (p < 0.05).
a
The maximum ethanol concentrations produced by the yeast strain obtained at 36 h.
b
Ethanol yield (gp/gs) is calculated as the ethanol accumulation divided by the glucose and xylose consumed.
c
The theoretical yield of ethanol is 0.511 gp/gs from glucose and 0.511 gp/gs from xylose; the theoretical yield is calculated as the ethanol yield multiplied by 100 and divided by
0.511.

sugar cane (3570.15 kg/ha), TByEFC05-1558 (2988.64 kg/ha), 4. Conclusion

TByEFC09-0098 (2688.59 kg/ha) and TByEFC04-1208 (1179.84 kg/
ha). Even though the sugar cane (LK92-11) obtained the highest cane
yield and ethanol yield from juice, this sugar cane had a low fiber yield,
which led to lower total ethanol production per area compared with the
energy cane TByEFC04-1155. Our results are in agreement with a
previous study by Kim and Day (2011), who reported that the theore-
tical ethanol yield of energy cane is superior to that of sugar cane. It is
possible to achieve ethanol production of 12,938.00 kg/ha and
3609.00 kg/ha from energy cane and sugar cane, respectively, under
conditions where all of the cellulose and hemicellulose are converted
into monomeric sugars, all of the fermentable sugars from the juice and
lignocellulose are also completely fermented to ethanol and there is no
loss of sugar substrates or ethanol product throughout the entire pro-
cess. To our knowledge, this is the first report to evaluate ethanol
production from the juice and bagasse of energy cane at the laboratory
scale in combination with their agronomic productivity.
The juice and bagasse from four TByEFC energy cane clones were
successfully fermented to ethanol. The combined high ethanol yield
from the juice and bagasse per cultivated area of TByEFC04-1155 was
the best overall ethanol yield (3923.48 kg/ha). An energy cane clone
that maximizes both the biomass and sugar yields holds considerable
promise in the expansion of future opportunities for both the ethanol
and sugar industries.
Acknowledgement
This research was financially supported by the Kasetsart University
Research and Development Institute (KURDI), Kasetsart University,
Bangkok, Thailand (grant number 141.57).

Fig. 1. Total sugar (a), ethanol (b), FAN (c)

and cell growth (d) profiles during the fer-
mentation of energy cane juices and sugar
cane juice using S. cerevisiae ND48.

602
S.N.-R. Thammasittirong et al. Industrial Crops & Products 108 (2017) 598–603

Table 5
Ethanol production at the laboratory scale and per area from the juice and bagasse of energy canes and sugar cane.

Clone Ethanol production at the laboratory scale Ethanol production per area

Juice Bagasse Juice Bagasse Total

g/La %b g/La %b kg/ha kg/ha kg/ha

b a ab a b b
TByEFC04-1155 72.50 ± 2.09 89.25 ± 0.49 9.42 ± 0.88 87.37 ± 2.29 2697.00 ± 77.00 1226.48 ± 64.29 3923.48 ± 165.10a
TByEFC04-1208 82.56 ± 2.27a 89.29 ± 0.81a 9.11 ± 0.11b 88.21 ± 0.82a 909.61 ± 40.12d 270.23 ± 9.68e 1179.84 ± 95.87c
TByEFC05-1558 78.02 ± 2.95b 86.10 ± 0.97b 10.37 ± 0.13a 88.43 ± 2.26a 2148.67 ± 81.40c 839.97 ± 21.16c 2988.64 ± 196.02b
TByEFC09-0098 68.00 ± 1.15c 86.10 ± 1.69b 10.68 ± 0.50a 89.97 ± 1.58a 993.14 ± 42.00d 1695.45 ± 50.00a 2688.59 ± 59.63d
LK92-11 84.37 ± 0.59a 87.36 ± 0.59b 10.54 ± 0.20a 90.00 ± 3.45a 3087.94 ± 219.40a 482.21 ± 17.58d 3570.15 ± 405.51ab

Different lowercase superscript letters in a column indicate significant differences between the clones (p < 0.05).
a
The maximum ethanol concentrations produced by the yeast strain obtained at 36 h.
b
The theoretical yield of ethanol is 0.511 gp/gsfrom glucose and 0.511 gp/gs from xylose;the theoretical yield is calculated as the ethanol yield multiplied by 100 and divided by 0.511.

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