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Fuel
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Department of Food Storage and Technology, Wrocaw University of Environmental and Life Sciences, Chemonskiego 37/41, 51-630
Wrocaw, Poland
highlights
Waste wheat rye-bread ethanol fermentation using granular starch hydrolyzing enzyme. Three raw
material pretreatment methods studied for ethanol yield improvement.
Separate hydrolysis and fermentation process studied for comparison.
Fermentation of unpretreated waste bread yielded 354.36 g ethanol kg
Application of raw material pretreatment further improved ethanol yield.
raw material.
abstract
article info
Article history:
Received 7 March 2014
Received in revised form 20 May
2014 Accepted 27 May 2014
Available online 12 June 2014
Keywords:
Ethan
ol fuel
Wast
e
bread
Ethanol fermentation
Granular starch hydrolyzing enzyme
(GSHE)
The subject of this research project was assessment of direct starch to ethanol conversion
process course of waste wheat-rye bread using granular starch hydrolyzing enzyme (GSHE).
Several pretreatment meth-ods (enzymatic prehydrolysis, microwave irradiation, sonification) were
used to improve the course of fermentation and were compared with separate hydrolysis and
fermentation (SHF). Due to high water binding capacity of raw material fermentations were
1
conducted at a substrate loading of 150 g kg . Only during enzymatic pretreatment and the SHF
process the raw material was preliminary liquefied so its higher concentrations could be applied.
The dynamics of fermentation was similar in all studied variants. The fermentation of unpretreated
waste bread ended with 80.00% ethanol yield (354.36 g kg
material improved ethanol yield by ca. 38%.
Saccharomyces cerevisiae
1. Introduction
Ethanol is considered as one of the most promising
renewable fuel that can replace fossil fuels-based
transportation fuels. It is most commonly produced by
microbial (most often yeast) cata-lyzed fermentations using
plant biomass as a raw material. Starchy raw materials (i.e.
corn, wheat, sorghum) are still the most common feedstocks
for fuel ethanol production in temperate climate regions of
the world (Europe, North America, Central Asia). However its
use as fuel production resources may affect on the prices of
food prod-ucts manufactured from them [1]. The use of nonedible parts of the plant (straws, stalks), known as the
lignocellulose biomass, as the raw material in distillery is
nowadays considered as the most prom-ising opportunity for
ethanol production that does not affect the prices of
foodstuffs [2]. However, the conversion of lignocellusosic
biomass into fermentable sugars and, subsequently into
ethanol
http://dx.doi.org/10.1016/j.fuel.2014.05.08
1 0016-2361/ 2014 Elsevier Ltd. All rights
reserved.
1
1
251
2.85 g kg
of raw material dry matter. Raw material
moisture content was measured using WPS 50P weighing
1
252
specificity
(a-amylase,
b-glucanase,
pentosanase, cellulase, prote-ase). In
separate
hydrolysis
and
fermentation
experiment following enzymes were used (all
produced by Novozymes): Termamyl SC DS
(thermostable a-amylase from Bacillus
licheniformis with declared activity of 240 kilo
novo units per gram) and SAN Extra L
(glucoam-ylase from Aspergillus niger with
declared activity of 400 glucoam-ylase units
per gram) and, mentioned above, Neutrase
0.8 L. Commercial active dry yeast
Saccharomyces cerevisiae Ethanol Red
(Fermentis, France) was used as production
9
H2SO4,
1
L, 0.875 mL kg
of raw material dry matter)
were added and the hydrolysis was conducted
for 90 min. After this time the sam-ples were
cooled to 20 LC, the pH of the samples was
1
253
1
Shimadzu)
at
50
LC.
Obtained
chromatograms were integrated using
external standard method with CHROMAX 10
software (Pol-Lab, Poland).
On the basis of obtained results following
process parameters were calculated at 24 h
time intervals: ethanol production rate (r p [g L
1
120
(a)
90
60
30
0
0
20
150
(b)
II
III
IV
V
I
II
III
IV
V
100
50
0
60
80
20
40
60
150
80
Time [h]
Time [h]
(c)
I
II
III
IV
V
100
50
0
0
20
40
60
80
Time [h]
Fig. 1. Changes in physico-chemical properties of fermentation media from waste wheat-rye bread during direct starch
conversion to ethanol with GSHE (a) reducing sugars,
(b) dissolved solids, (c) non-dissolved solids (I unpretreated variant, II enzymatic pretreatment, III microwave
pretreatment, IV ultrasonic pretreatment, V SHF process). Results expressed as mean values standard deviation
(n = 3).
254
Table 1
Carbohydrate profiles during fermentation of media prepared from waste wheat-rye bread (I unpretreated variant, II enzymatic pretreatment, III microwave pretreatment, IV
ultrasonic pretreatment, V SHF process). Results expressed as mean values standard deviation (n = 3).
Variant
I
II
III
IV
V
Glucose (g L )
Maltotriose (g L )
Dextrins (DP4+) (g L )
24 h
48 h
72 h
24 h
48 h
72 h
40.78 3.99
42.99 3.54
52.86 3.95
49.33 0.66
40.10 3.47
2.58 2.03
0.16 0.07
2.42 0.15
4.75 0.90
0.06 0.04
0.09 0.05
0.00 0.00
0.00 0.00
0.00 0.00
0.00 0.00
0.24 0.07
0.47 0.06
0.77 0.16
0.37 0.04
0.89 0.18
0.37 0.07
0.35 0.07
0.58 0.01
0.84 0.33
0.30 0.02
0.36 0.04
0.26 0.03
0.39 0.02
0.11 0.09
0.23 0.01
24 h
8.46
11.23
13.90
9.95
12.57
0.72
0.48
0.66
0.17
0.71
48 h
72 h
7.40 0.98
8.28 0.26
9.75 0.39
8.46 0.26
7.93 0.13
7.71 0.56
8.02 0.38
8.43 0.11
6.90 0.61
7.86 0.16
60
(Fig. 1a). Its concentration ranged ca. 53, 10 and 0.2 g L after 24, 48
and 72 h of fermentation respec-tively. In enzymatically pretreated
samples sugars concentration after the first day increased by ca. 12 g
1
L
in comparison to ini-tial state. After subsequent days of
fermentation the reducing sugars content in this variant of experiment
was below
(a)
(b)
Glycerol [g L-1]
45
Ethanol [g L-1]
30
I
II
III
15
4
I
II
III
IV
IV
0
0
20
40
60
80
20
40
Time [h]
60
80
Time [h]
1,5
Lactic acid [g L -1
(c)
I
II
III
IV
V
0,5
0
0
20
40
60
80
Time [h]
Fig. 2. Formation of ethanol (a), glycerol (b) and lactic acid (c) during direct starch to ethanol conversion process of waste wheat rye-bread media with GSHE (I unpretreated
variant, II enzymatic pretreatment, III microwave pretreatment, IV ultrasonic pretreatment, V SHF process). Results expressed as mean values standard deviation (n = 3).
W. Pietrzak, J. Kawa-Rygielska / Fuel 134 (2014) 250256
255
0.47 0.11c
0.52 0.06bc
0.73 0.03ab
0.92 0.01a
0.55 0.02bc
0.21
0.13 0.04a
0.10 0.07a
0.06 0.03a
0.01 0.14a
0.05 0.03a
0.26
48 h
1.61 0.11a
1.81 0.01a
1.62 0.01a
1.36 0.02b
1.81 0.03a
0.19
20.11b
4.47a
5.73a
19.70a
2.42a
(g L1 h 1 )
24 h
72 h
0.5 g L . In the SHF variant of experiment yeast did consume half of the
available sugars in the fermentation feed after 24 h of the process and almost
all available sugars after subsequent days. HPLC analysis of carbohydrates in
media during the fermentation also shown that almost all available glucose
was utilized by yeast by the second day of the process (Table 1). The content
15.12b
7.25a
5.78a
2.45a
6.00a
22.25
waste bread sam-ples, while the lowest amount of alcohol (ca. 32.6 g L ) was
determined in media with subsequently microwave irradiated waste bread.
The production of ethanol by yeast in studied fer-mentation tests lasted by the
second day of the process and did not significantly change by the third day.
The lowest concentra-tion of ethyl alcohol was found in unpretreated samples
1
difference.
48 h
24 h
practical ethanolyield, r
(g kg 1 of raw material)
72 h
354.36
389.41
384.60
366.78
385.98
38.8 g L ) was found after the same time in unpretreated and sonificated
72 h
48 h
24 h
24 h
48 h
72 h
(g kg
p
(%)
I
58.33 3.93a 75.25 3.41b
II
65.52 0.54a 84.24 1.64a
III
58.53 0.31a 84.78 1.31a
IV
49.11 0.63b 82.42 0.55a
V
65.53 1.05a 85.50 1.35a
LSD
6.77
6.94
5.36
of sugars)
408.79 23.20b
449.21 5.15a
443.66 6.61a
423.11 22.72a
445.26 2.79a
Variant Y
Table 2Ethanol yield and production rate at specified time intervals of fermentation of waste wheat-rye bread with GSHE (Ymicrowavepretreatment,IVultrasonicpretreatment,VSHFprocess).
6.06.5 g L ). The lac-tic acid content in fermentation media was similar in all
256
4. Conclusions
Present study proved that waste wheatrye bread is a high eth-anol yielding material
in direct conversion to ethanol process using
1
References
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