Applied Energy 102 (2013) 204210

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Applied Energy
journal homepage: www.elsevier.com/locate/apenergy

Bioethanol production from mandarin (Citrus unshiu) peel waste using

popping pretreatment
In Seong Choi a, Jae-Hoon Kim b, Seung Gon Wi c, Kyoung Hyoun Kim d, Hyeun-Jong Bae a,c,d,
a

Department of Wood Science and Landscape Architecture, Chonnam National University, Gwangju 500-757, Republic of Korea
Faculty of Biotechnology, College of Applied Life Sciences, Cheju National University, Jeju 690-756, Republic of Korea
c
Bio-Energy Research Center, Chonnam National University, Gwangju 500-757, Republic of Korea
d
Department of Bioenergy Science and Technology, Chonnam National University, Gwangju 500-757, Republic of Korea
b

a r t i c l e

i n f o

Article history:
Received 23 December 2011
Received in revised form 28 March 2012
Accepted 31 March 2012
Available online 26 April 2012
Keywords:
Popping pretreatment
Ethanol productivity
Mandarin peel waste
Limonene

a b s t r a c t
In this study, we designed a biomass popping pretreatment, system using a red burner and a horizontal
cylinder rotating on an axis, to produce ethanol from mandarin (Citrus unshiu) peel (MP) waste. Popping
pretreatment was performed at 150 C for 10 min without chemical treatment. Popping pretreatment
reduced the size of particles to less than 1 mm and decreased the concentration of D-limonene, a yeast
fermentation inhibitor, from 0.21% to 0.01%. Enzymatic hydrolysis of pretreated MP was performed in
50 mM sodium acetate buffer (pH 4.8) at 45 C for 6 h, and the total saccharication rate was approximately 95.6%. The vacuum evaporation process increased the fermentable sugar concentration to 10%
(glucose 7.1% and fructose 2.9%). Subsequent fermentation at 30 C at pH 5.0 for 12 h in a laboratory bioreactor increased the ethanol yield to 90.6%, compared to 78% at 36 h from raw MP.
2012 Elsevier Ltd. All rights reserved.

1. Introduction
Bioethanol is a promising alternative to fossil fuels because it is
a renewable resource. It is a liquid fuel that can be adapted to
existing fuel supply systems and can replace fossil fuels in the
transportation sector [14]. However, starch-based bioethanol production is criticized because starch is generally derived from food.
Thus, the use of starch as a source of bioethanol can reduce food
supplies and increase food price. The use of starch- or sugar-rich
feedstock has directly increased food and bioethanol prices due
to the high price of raw materials, which account for 4075% of
the total expenses for ethanol production [5]. Therefore, alternative biomass sources are required for bioethanol production, such
as agricultural byproducts, forest residues, or energy crops (lignocellulosic biomass) [614].
Citrus peel waste is a suitable candidate for bioethanol production because it contains low lignin and high soluble sugar concentrations [15,16]. Citrus peel waste is rich in fermentable sugars
such as glucose, fructose and sucrose, along with insoluble polysaccharides such as cellulose and hemicellulose [16]. According
to the USDA [17], world citrus production was estimated at 49.9
million tons during the 2009/2010 growing season. After juice
Corresponding author at: Department of Bioenergy Science and Technology,
Chonnam National University, Gwangju 500-757, Republic of Korea. Tel.: +82 62
530 2097; fax: +82 62 530 0029.
E-mail address: baehj@chonnam.ac.kr (H.-J. Bae).
0306-2619/$ - see front matter 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.apenergy.2012.03.066

extraction, citrus peel waste (containing peel and saccharides)

accounts for approximately 44% of the wet fruit mass [18].
Mandarin peel (MP) waste is also considered an important source
for bioethanol production because the soluble sugar and carbohydrate contents are similar to citrus peel waste [15]. In Korea, the
average production of fresh mandarin fruits and mandarin waste
per year are approximately 700,000 and 70,000 tons, respectively.
Discarded mandarin fresh fruit and waste will be likely increased
over time due to the increase in imported citrus fruit from the
USA. Approximately 30% of mandarin waste is currently used in
medicinal herbs and the remaining 70% is disposed into the ocean.
However, ocean disposal causes environmental problems and is
forbidden by the London Convention and London Protocol of
2013 in Korea.
Citrus peel waste contains 0.81.6% D-limonene, which is an
inhibitor of yeast fermentation [19]. When using of citrus peel
waste for bioethanol production, acidic steam explosion pretreatment is required to lower the limonene content to below 0.05%
because D-limonene inhibit microbial growth [1921]. Steam
explosion pretreatment has been used to produce bioethanol from
MP [15].
Pretreatment is required to improve the accessibility of substrates to cellulose-degrading enzymes [22]. We have designed a
biomass pretreatment system with a red burner and horizontal
cylinder, which rotates on an axis, called popping pretreatment
(there is a pop out as biomass is produced from the pretreatment). Previously, we applied popping pretreatment to rapeseed

I.S. Choi et al. / Applied Energy 102 (2013) 204210

straw and obtained a high glucose yield after enzymatic hydrolysis

[23].
In this study, we investigated bioethanol production using MP
after popping pretreatment and established an efcient bioethanol
production process base. We investigated the effect of popping
pretreatment on MP composition, limonene concentration, and
particle size. We also studied the efciency of enzyme cocktails
on hydrolysis and ethanol yields after individually optimizing the
hydrolysis and fermentation of the enzymatic hydrolysate of pretreated MP.
2. Materials and methods
2.1. Mandarin peel wastes and popping pretreatment
Fresh mandarin (Citrus unshiu) peel wastes (MP), with a dry
matter content of 17.8% (wet basis), was obtained from Jeju Island.
Raw MP was ground in a homogenizer, frozen ( 20 C), lyophilized
( 50 C), and stored at 20 C until further use. The pretreatment
experiment after defreeze, 1 kg of fresh MP matter without any
earlier cut or chemical pretreatment was performed by modied
popping equipment. The main structure of the reactor, with an inner horizontal cylinder of 300 mm from left to right, had a total
volume of 3 L (Supplementary Fig. S1). The heating and pressure
mechanism was adapted using an external heating plate on the
surface of the vertical cylinder and internal steam pressure was increased proportionately with followed by increasing internal temperature without any external steam injection. The conditions of
the popping machine were temperature 150 C, reaction time
10 min and pressure 15 kgf/cm. During the popping pretreatment,
the right side cap lock was tightly sealed on the cylinder and operating conditions were maintained by an automatic controller with
a pressure gauge and temperature probe in the cylinder. After the
end of the reaction, the side cap lock was quickly loosened to reduce the internal pressure of the reactor to atmosphere pressure.
Both internal steam and MP were erupted to pressure relief tank
to cause the disruption of MP.
2.2. Chemical analysis
MP (raw and popping-pretreated) was analyzed for holocellulose, Klason lignin, ash, and organic solvent extractives by TAPPI
Standard Method [24]. Each powdered sample was obtained after
being sieved through a 251422 lm of sieve. For organic solvent
extraction, the Soxhlet apparatus was used with a 2:1 mixture of
benzene:ethanol. The extract-free samples (2.5 g) were treated
with 1 g of sodium chlorite and 0.2 mL of glacial acetic acid
(18 M), and then incubated at 70 C for 3 h with homogenizer
treatment every hour. The treated samples were analyzed by gas
chromatography (GC). Klason lignin was determined as an unhydrolyzed residue remaining after sulfuric acid hydrolysis for extract-free samples. The samples were treated with 2.5 mL of 72%
H2SO4 for 45 min at room temperature and diluted with water to
4% H2SO4. After hydrolysis for 1 h at 121 C, insoluble residue
was quantitatively ltered off (Pyrex, 1G4 glass lter) with
500 mL of hot water. The Klason lignin residue was dried at
105 C for 12 h.
2.3. Scanning electron microscopy (SEM)
For analyzing cell wall structure differences between raw MP
(ground form) and the popping-pretreated MP, samples were prepared using 2% glutaraldehyde (v/v) and 2% paraformaldehyde
(v/v) in 0.05 M cacodylate buffer (pH 7.2). After rinsing in cacodylate buffer, samples were dehydrated in a graded ethanol series and

205

then freeze-dried. The samples were then coated with gold (20 nm)
in a sputter coater prior to being observed and photographed with
a scanning electron microscope (S2400, Hitachi, Japan).
2.4. D-limonene extraction
Extraction and quantication of D-limonene were performed
according to the method described previously [25]. Matters (raw
and popping-pretreated) were homogenized in 10 mL of hexane
and known amount of camphor as an internal standard. After treatment for 3 h, the residue was discarded and 5 mL of supernatant
was transferred to a tube. 0.2 mL of 2 N KOH in methanol was
added and the samples were mixed for 1 min. After the addition
of 1 mL of distilled water, the samples were vigorously mixed
and then centrifuged for 5 min at 3000 rpm. The hexane phase
was analyzed by gas chromatography (CP-9100, Chrompack) using
a CP-Sil 5 CB fused silica capillary column (25 m  0.32 mm i.d.,
1.2 lm lm thickness, Chrompack) operated with He, injector temperature 280 C, ame ionization detector (FID) at 280 C.
2.5. Enzymatic hydrolysis
Enzymatic hydrolysis was performed on matter with individual or mixed commercial cellulase (Celluclast 1.5L), pectinase
(Pectinex SP-L), xylanase (X2629 endo-1,4-b-D-xylanase), and bglucosidase (Novozym 188) at 45 C for 24 h as follow previous
studies [8,19]. Cellulase, pectinase and b-glucosidase were purchased from Novozyme A/S (Bagsvaerd, Denmark) and xylanase
was provided by Sigma (St. Louis, MO, USA). Cellulase and pectinase activities were determined as shown previously [19] and
xylanase activity was measured according to Bae et al. [27]. The
activities of cellulase, pectinase and xylanase were 0.122 lter paper unit (FPU)/mg protein, 240 international units (IU)/mg protein
and 2.65U/mg xylanase, respectively. The activity of b-glucosidase
was 2.6 IU/mg solid as reported by the supplier.
The raw and popping pretreated matters were diluted in 15 mL
test tubes to 1% (w/v) dry matter with 50 mM sodium acetate buffer (pH 4.8). The optimal enzyme cocktails for measuring the sugar
composition of MP was determined with different enzyme cocktails on raw or pretreated MP as compare with non enzyme treated
(control). Based on a small scale study, the enzymatic hydrolysis
system was scaled up to a 5 L bioreactor (1% dry matter).
2.6. Sugars analysis
After complete hydrolysis, all samples in test tube with cap
were boiled at 105 C for 5 min to inactivate enzymes and then
centrifuged for 20 min at 13,000 rpm and 26,000g (Avanti JE,
Beckman, USA) before analyzing the soluble sugars. The sample
supernatants were analyzed for soluble sugar content using high
performance liquid chromatography (HPLC) with a refractive index
detector (2414, Waters, USA). The mixtures were treated with
50 ll of enzyme and loaded into the REZEX RPM (Phenomenex,
USA) column (300  7.8 mm) at 85 C and eluted with deionized
water at a ow rate of 0.6 mL per min.
The insoluble solids were washed ve times in distilled water
and pelleted. Pellets were lyophilized before analyzing the sugars.
Each pelleted samples were treated using a modied alditol acetate method and the neutral sugar contents were measured by
gas chromatography (GC) [28]. Gas chromatography analysis conditions were as follows, gas chromatography (GC-2010, Shimadzu,
Japan) using a DB-225 capillary column (30 m  0.25 mm i.d.,
0.25 lm lm thickness, J&W) operated with He, injector temperature of 220 C, ame ionization detector (FID) at 250 C, and
oven temperature programming: 100 C for 1.5 min, 5 C/min to
220 C.

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I.S. Choi et al. / Applied Energy 102 (2013) 204210

2.7. Vacuum evaporation

3. Results and discussion

For the increase of fermentation efciency, low concentrated

fermentable sugar solutions from raw MP or enzyme hydrolysis
MP were vacuum evaporated to produce 10% (w/v) fermentable
sugars (Supplementary Fig. S2). The vacuum evaporation process
was undertaken with vacuum evaporation equipment, which consisted of a vertical stainless reactor (50 L) and an electronic heater
on the bottom of the reactor. The vacuum device, thermometer and
condenser were connected to the reactor and a cold trap cooled by
water was placed between a vacuum pump and a storage tank
connected to the condenser. The vapor, which was evaporated at
3740 C under pressure 0.10.09 Mpa, was collected in the storage tank. As a consequence of evaporation, a low sugar concentration solution was separated into a high level sugar solution in the
reactor.

3.1. Effect of popping pretreatment on mandarin peel composition

2.8. Yeast fermentation

The yeast strain Saccharomyces cerevisiae KCTC 7906 was obtained from the Korean Collection for Type Culture (KCTC, Korea)
and activated in 4 ml of Yeast Peptone Dextrose media (YPD),
which consisted of glucose 20 g/L, yeast extract 10 g/L, peptone
10 g/L, MgSO4
7H2O 0.4 g/L, (NH4)2
SO4 0.2 g/L and KH2PO4 3.3 g/
L. The yeast inoculum was placed in a 500 mL Erlenmeyer ask
containing 100 mL of autoclaved YPD media for 24 h at 30 C.
The 100 mL of inoculated yeast was added to the bioreactor with
YPD media except glucose. The fermentation process after enzyme
hydrolysis of pretreated MP (glucose 7.1%, fructose 2.9% by HPLC)
was performed by a 3 L bioreactor with 1 L working volume
(FMT-05/C-B, Fermentec, Korea). The bioreactor had an auto-control system for maintenance at 30 C and was adjusted to pH 5.0
by 2 M NaOH. Samples were withdrawn aseptically from the bioreactor periodically from 0 h to 36 h for analysis of total sugar and
produced ethanol.

Popping pretreatment uses whole MP without chemical treatments or mechanical cuts. This procedure simplies the MP pretreatment compared to other methods such as steam explosion.
Generally, steam explosion requires diluted chemicals (sulfuric
acid or sodium carbonate) or sample cutting to produce ethanol
from citrus species [15,18].
We evaluated the observed structural and chemical changes
through the popping pretreatment on MP. The D-limonene concentration was also analyzed before and after popping pretreatment
because terpenoid is a fermentation inhibitor. To enhance ethanol
production, MP required a pretreatment process that effectively
disrupts cell walls, increasing access of polysaccharides to the enzymes, as reported previously [29].
SEM images indicated that popping treatment reduced MP
particle size, and consequently increased the substrate surface
area (Supplementary Fig. S3A and B). The increased cell wall surface area improved accessibility of cell wall-degrading enzymes,
and the reduced particle size was economically benecial.
Mechanical pretreatments are generally applied during bioethanol production. A particle size reduction to 251 lm improves
the hydrolysis yield by 525% [30,31]. However, due to the high
energy requirement, mechanical pretreatments are not economically feasible [32]. According to Carroll and Somerville [7], a
hardwood particle size of 1.6 mm required 130 kWh/kg, whereas
grass straws required 7.5 kWh/kg. In comparison, an approximately 10  10 cm MP particle was reduced to 1 mm after popping pretreatment.
The MP cell wall carbohydrate concentration was affected by
popping pretreatment (Table 1). GC analysis of the MP cell wall
carbohydrate showed glucose and xylose as the primary components with concentrations of 24.8% and 3.1%, respectively. There

Table 1
Chemical composition of raw, popping-pretreated MP and various lignocellulosic feedstocks. Organic solvent extractives, lignin, and ash were analyzed by TAPPI standard method
and monosaccharide was analyzed by using GC. The values for raw and popping pretreated mandarin peel from analysis of this study.
(% of dry matter)

Extractives

Rhamnose

Arabinose

Xylose

Mannose

Galactose

Glucose

Lignin

Ash

Raw mandarin peel

Popping pretreated mandarin peel
Citrus peela
Sugarcane bagasseb
Wheat strawb
Switchgrass (alamo)b

38.3
44.3
nd
3.8
12.9
17.0

3.8
0.5
0.9
nd
nd
nd

11.2
1.7
8.4
2.1
2.4
2.8

3.1
5.8
3.7
22.1
19.2
20.4

2.0
1.7
3.0
0.4
0.3
0.3

4.2
1.9
6.4
0.5
0.8
0.9

24.8
32.0
25.8
39.0
32.6
31.0

8.6
9.2
nd
23.1
16.9
17.6

4.2
3.1
nd
3.7
10.2
5.8

nd = no data.
a
From Hendriks and Zeeman [32] for citrus peels from French citrus industrial factory.
b
From Carroll and Somerville [7].

Table 2
Concentration of saccharide as soluble sugars by enzymatic hydrolysis on raw and popping pretreatment material. Each enzymes loading follow as; pectinase 5 mg/g dry matter,
xylanase 5 mg/g dry matter and b-glucosidase 2 mg/g dry matter. Abbreviations used monosaccharides (Suc; sucrose, Glu; glucose, Gal; galactose, Ara; arabinose, Fruc; fructose)
and enzymes (Pec; pectinase, Xyn; xylanase, b-G; b-glucosidase.).
Content (g/10 g)

Enzyme cocktails

Suc

Glu

Gal

Ara

Fruc

Total

Raw

Cont.
Pec
Xyn
Pec + Xyn
Pec + Xyn + b-G

0.94
0.00
0.46
0.00
0.00

1.73
3.10
3.99
4.71
4.60

0.04
0.11
0.03
0.12
0.00

0.00
0.07
0.07
0.10
0.09

1.52
1.90
1.77
2.21
2.16

4.23
5.18
6.31
7.14
6.84

Popping pretreatment

Cont.
Pec
Xyn
Pec + Xyn
Pec + Xyn + b-G

0.04
0.00
0.09
0.00
0.00

1.75
3.17
4.05
4.42
4.51

0.02
0.18
0.03
0.15
0.12

0.10
0.24
0.19
0.26
0.28

1.47
1.45
1.55
1.33
1.84

3.39
5.04
5.91
6.15
6.76

Mean values of three determinations.

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Raw and pretreated MP was mostly hydrolyzed at pH 4.8, while

the optimal pH for commercial pectinase was pH 3.8. Raw and pretreated MP naturally contains soluble sugars such as sucrose, glucose, and fructose. These sugars affect the analysis of soluble
sugars after enzymatic hydrolysis. HPLC results showed that various enzyme cocktails produced different yields of soluble sugars
from MP with or without popping pretreatment (Table 2). For
enzyme hydrolysis of raw MP, xylanase (5 mg/g dry matter) and
pectinase (5 mg/g dry matter) enzyme cocktails were most effective, producing 7.14 g sugar/10 g dry matter after enzymatic hydro-

may be some loss of pectin, which is a highly complex polymer

consisting of galacturonic acid bound by many different monosaccharides, such as arabinose and rhamnose [33].
3.2. Effect of popping pretreatment on monosaccharide yield after
enzymatic hydrolysis
Cellulase, pectinase, and b-glucosidase are required for hydrolysis of orange or MP waste [19,20,26,34,35]. In our study, we included xylanase with the above enzymes.

Table 3
Carbohydrate concentration of solid residues from MP (raw and popping pretreatment) after enzymatic hydrolysis by GC. Abbreviations used, monosaccharides (Rham;
rhamnose, Ara; arabinose, Xyl; xylose, Man; mannose, Gal; galactose, Glu; glucose) and enzymes (Pec; pectinase, Xyn; xylanase, b-G; b-glucosidase.).
Content (g/kg)

Enzyme cocktails

Rham

Ara

Xyl

Man

Gal

Glu

Total

Raw

Cont.
Pec
Xyn
Pec + Xyn
Pec + Xyn + b-G

3.8
0.5
3.0
0.3
0.8

11.2
0.5
1.1
0.1
0.2

3.1
1.0
0.9
0.7
0.8

2.0
0.3
0.2
0.1
0.4

4.2
0.3
0.3
0.1
0.2

24.8
4.3
4.4
1.2
4.3

49.1
6.9
9.9
2.5
6.7

Popping pretreatment

Cont.
Pec
Xyn
Pec + Xyn
Pec + Xyn + b-G

0.5
0.1
0.1
0.1
0.0

1.7
0.3
0.4
0.2
0.2

5.8
0.8
0.6
0.6
0.8

1.7
0.2
0.2
0.1
0.1

1.9
0.2
0.1
0.1
0.0

32.0
3.2
1.5
1.3
0.8

43.6
4.8
2.9
2.4
1.9

Mean values of three determinations.

Fig. 1. Time course changes in glucose conversion rate (A) and carbohydrate concentration (B) during enzymatic hydrolysis.

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I.S. Choi et al. / Applied Energy 102 (2013) 204210

lysis. For the popping-pretreated MP, total sugars obtained after

xylanase (5 mg/g dry matter), pectinase (5 mg/g dry matter) and
b-glucosidase (2 mg/g dry matter) mixed enzyme cocktail hydrolysis were 6.76 g sugar/10 g dry matter.
After enzymatic hydrolysis of raw and popping-pretreated MP,
the raw MP contained as high as 0.38 g sugar/10 g dry matter,

Fig. 2. Limonene concentration of MP and orange peel (raw and popping pretreated). MP Raw; raw mandarin peel, MP Popping; popping-pretreated mandarin
peel, OP Raw; raw orange peel, OP Popping; popping-pretreated orange peel.
Popping pretreatment applied to orange peel decreased D-limonene concentration
from 1.70% to 0.02%.

which may be because sucrose in MP was converted to glucose

and fructose. This was due to the commercial pectinase (Pectinex
SP-L) and b-glucosidase (Novozym 188), produced by Aspergillus
aculeatus and A. niger, respectively (Supplementary Fig. S4). These
ascomycetes produce several hydrolysis enzymes, such as pectinase, cellulase and fructosyltransferase, which hydrolyze glucose
and fructose produced from sucrose [36,37].
After enzymatic hydrolysis, insoluble solid sugars from raw and
pretreated MP were analyzed to identify suitable enzyme cocktails
by GC. Total insoluble solid content was low (2.5% w/w) in pectinase and xylanase cocktails of raw MP, while the pectinase, xylanase, and b-glucosidase cocktail was most efcient for pretreated
MP (Table 3). Based on the enzyme cocktails, time course changes
of popping pretreated MP during enzymatic hydrolysis are shown
in Fig. 1. Cell wall carbohydrates accounted for 43.6% of pretreated
MP mass prior to hydrolysis and reached a maximum glucose conversion rate of 97.5% after 6 h of enzymatic action. Glucose and
fructose are the primary saccharides from MP and can be fermented by S. cerevisiae. Eliasson et al. [38] applied S. cerevisiae
for fermentation of a glucosexylose mixture. However, in MP, xylose was very low concentrations. In this study, we used glucose
and fructose for fermentation.
3.3. Effect of popping pretreatment on ethanol yield
Terpenoid is the essential oil D-limonene, which possesses antimicrobial activity [21,35]. Thus, it is not surprising that terpenoid
inhibits S. cerevisiae fermentation. This inhibitory mechanism

Fig. 3. Overall mass balance of MP waste.

I.S. Choi et al. / Applied Energy 102 (2013) 204210

may be related to disruption of yeast cellular membranes and subsequent H+ and K+ transport in glycolysis. D-limonene concentration decreased after popping pretreatment. Raw MP contains
0.21% D-limonene, but its concentration was reduced to 0.01% after
popping pretreatment (Fig. 2). Interestingly, the D-limonene
concentration decreased in orange peel (from 1.70% to 0.02%), possibly below the fermentable concentration [18,19].
Separate hydrolysis and fermentation (SHF), semi-simultaneous
saccharication and fermentation (SSSF), and simultaneous saccharication and fermentation (SSF) are generally considered to
be the primary methods for bioethanol production [39]. We modied SHF by including vacuum evaporation (SHEF).
A 10% (glucose 7.1%, fructose 2.9%) sugar solution was obtained
using vacuum evaporation from low concentrate hydrolysates.
After vacuum evaporation, fermentation was performed in a fermentor containing 1 L of fermentation medium inoculated with
10% high density cell inoculum. Ethanol yields were based on the
total fermentable sugars of both raw and pretreated MP. For raw
MP, fermentation peaked at 36 h when ethanol concentration
and yield efciency were 39.8 g/L and 78%, respectively. The ethanol yield of raw MP was lower than pretreated MP because the
concentration of D-limonene was higher than after pretreatment.
In contrast to raw MP, the fermentation of pretreated MP hydrolysate was almost complete after 12 h with 46.2 g/L ethanol concentration and 90.6% ethanol yield. More interestingly, for ethanol
productivity, pretreated MP fermentation produced 3.85 g/L h,
while raw MP fermentation produced 1.11 g/L h.
According to a previous study on the fermentation of orange
peel or MP wastes, 2448 h fermentation is required to achieve
over 90% ethanol yield [15,19,34,35,40]. Our process achieved a
greater ethanol yield with a much shorter fermentation time,
which is important for industrial applications because it reduces
the overall production cost. The high ethanol productivity and
yield efciency in our study could be due to several factors, including suitable hydrolysate concentrations, optimized pH, temperature, and low concentrations of D-limonene.
3.4. Overall mass balance
We used an overall mass balance diagram to describe the popping-pretreated MP and conversion of carbohydrates to ethanol
by enzymatic hydrolysis (Fig. 3). Compared to the raw MP values,
popping pretreatment increased the solid glucose concentration
by 22.5%, while D-limonene concentration decreased from 0.21%
to 0.01%. During enzymatic hydrolysis of pretreated MP, 311.7 g
of glucose were hydrolyzed from an initial 320 g of solid glucose,
while 8.3 g of residual glucose remained as unhydrolyzed cellulose.
The mass balance for glucose after enzymatic hydrolysis was 97.5%.
Hydrolysate was concentrated to 10%, and 93.6 L of water was
recovered by vacuum evaporation. Fermentation using S. cerevisiae
effectively converted monomeric glucose and fructose with 90.6%
fermentation yield and 373 mL of ethanol. These results suggest
that popping pretreatment combined with vacuum evaporation is
an acceptable industrial process for ethanol production from MP.
4. Conclusion
Mandarin (C. unshiu) peel waste is an attractive source of biomass for bioethanol production and has several advantages over
lignocellulosic biomass rich in hemicellulose and lignin (Table.
1). Popping pretreatment ruptures MP cell walls, producing particles with diameters less than 1 mm and also reduces D-limonene
concentrations to less than 0.01%. The pretreated MP does not yield
higher sugar concentrations in the hydrolysate after enzymatic
treatment but signicantly reduces D-limonene content. By com-

209

bining popping pretreatment, enzymatic hydrolysis, and vacuum

evaporation, we obtained an ethanol concentration of 46.2 g/L
and ethanol productivity of 3.85 g/L h, compared to 39.8 g/L and
1.11 g/L h for raw MP. This study on the popping pretreatment of
MP demonstrates the potential benets of the popping methods
for ethanol production. Further investigations regarding ethanol
fermentation are part of an ongoing economical analysis.
Acknowledgements
This work was supported by the New & Renewable Energy of
the Korea Institute of Energy Technology Evaluation and Planning
(2010T100100573) grant funded by the Korea government Ministry of Knowledge Economy and by Priority Research Centers Program (2011-0018393), and WCU (World Class University) project
(R31-2009-000-20025-0) through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science
and Technology to H.-J. Bae. I.S. Choi is grateful for the BK21 program provided by the Ministry of Education.
Appendix A. Supplementary material
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.apenergy.2012.
03.066.
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