International Biodeterioration & Biodegradation xxx (2010) 1e8

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International Biodeterioration & Biodegradation

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Effects of particle size on anaerobic digestion of food waste

Kouichi Izumi a, *, Yu-ki Okishio b, Norio Nagao a, Chiaki Niwa a, c, Shuichi Yamamoto a, Tatsuki Toda a
a
Department of Environmental Engineering for Symbiosis, Faculty of Engineering, Soka University, Tangi-cho, Hachioji, Tokyo 192-8577, Japan
b
School of Public and Environmental Affairs, Indiana University, 1315 East Tenth Street, Bloomington, IN 47405, USA
c
Institute of Technology, Shimizu Co., Etchujima, Koutou-ku, Tokyo 135-8530, Japan

a r t i c l e i n f o a b s t r a c t

Article history: The objective of this study was to investigate the effects of particle size reduction and solubilization on
Received 3 February 2010 biogas production from food waste (FW). To clarify the effects of volatile fatty acids (VFAs) in the
Received in revised form digestion process, the relationship between particle size and VFA accumulation was investigated in
21 June 2010
detail. For this purpose, substrates of various particle sizes were prepared by bead milling to support
Accepted 22 June 2010
Available online xxx
hydrolysis. Batch anaerobic digestion experiments were carried out using these pretreated substrates at
mesophilic temperature for a period of 16 days. The results of pretreatment showed that the mean
particle size (MPS) of substrates ground with a bead mill decreased from 0.843 to 0.391 mm, and
Keywords:
Anaerobic digestion
solubilization accounted for approximately 40% of the total chemical oxygen demand (total COD) for
Methane grinding pretreatment by bead milling. Anaerobic digestion batch experiments revealed that MPS
Particle size reduced by bead milling at 1000 rpm improved methane yield by 28% compared with disposer treat-
Pretreatment ment. Moreover, this may have increased microbial degradation during the VFA production process with
Food waste increasing total number of revolutions (operation time
revolutions per minute). However, excessive
Volatile fatty acids reduction of the particle size of the substrate resulted in VFA accumulation, decreased methane
production, and decreased solubilization in the anaerobic digestion process. These results suggest that
optimized reduction of the particle size of the substrate in conjunction with optimized microbial growth
could improve the methane yield in anaerobic digestion processes.
Ó 2010 Elsevier Ltd. All rights reserved.

1. Introduction
In Japan, an ever-increasing amount of food waste (FW) is gener-
ated, owing to population growth and rising living standards. At
present, the primary method for the disposal of FW in Japan
is incineration. Because FW has over 80% water content, extra fuel is
needed to treat FW by direct incineration (Sawayama et al., 1997;
Zhang et al., 2005). Anaerobic digestion has been widely studied as
an alternative method for the treatment of organic waste such as
sewage sludge and manure. Since FW has the advantage of high
organic content compared with traditional substrates, anaerobic
digestion is considered to be a feasible alternative method to decrease
treatment costs and recover renewable energy in the form of methane.
Anaerobic digestion is a method of FW treatment that can utilize
the biological processes of many classes of bacteria and generally
consists of four steps: hydrolysis, acidogenesis, acetogenesis, and
methanogenesis (Speece, 1996; Xu et al., 2002; Li, 2004; Metcalf
and Eddy Inc., 2004). In the case of methane fermentation of
* Corresponding author. Tel.: þ81 42 691 9455; fax: þ81 42 691 4086.
E-mail address: kizumi@soka.ac.jp (K. Izumi).
solid organic materials such as FW and waste activated sludge, the
methane yield is significantly affected by the mass transfer in each
biological step, as well as by food availability (Eastman and
Ferguson, 1981; Li and Noike, 1992). In particular, the hydrolysis
and acidogenesis stages are considered to be rate-limiting steps in
the process, since these two steps directly affect both the mass
transfer and the food availability in the process (Gomec et al.,
2002). To reduce the impact of these rate-limiting steps, pretreat-
ment of organic solids with heat (Kim et al., 2003), pressure
(Engelhart et al., 1999), ultrasonic irradiation (Shimizu et al., 1992),
or mechanical grinding (Nah et al., 2000) is required. These treat-
ments accelerate the solubilization (hydrolysis and acidogenesis) of
the substrate and reduce the particle size of the FW, subsequently
improving the anaerobic digestion.
Pretreatment to reduce particle size has two effects: first, if the
substrate has a high fiber content and low degradability, commi-
nution of the substrate increases gas production; second, it can lead
to more rapid digestion (Palmowski and Muller, 1999). Smaller
particles increase the surface area available to the microorganisms,
resulting in increased food availability to bacteria; thus, anaerobic
biodegradability increases (Mshandete et al., 2006). Sharma et al.
(1988) reported the effects of particle size on agricultural and

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forest residues used as feedstock for biogas generation through Table 1

anaerobic digestion in batch digesters at 37  C. Out of five particle Composition of food waste used in this study; the wet weight percentages were
based on Izawa et al. (2001).
sizes (0.088, 0.40, 1.0, 6.0, and 30.0 mm), the maximum quantity of
biogas was produced from 0.088-mm to 0.40-mm particles. Kim Large Percentage Content in each category
et al. (2000) reported the effects of particle size on anaerobic classification (% wet weight) (% wet weight)

thermophilic digestion in FW treatment. The maximum substrate Rice 4.0 Rice

Noodle 2.5 Buckwheat (50.0), spagetti (50.0)
utilization rate coefficient doubled with a decrease in the average
Bread 1.7 Bread (100.0)
particle size from 2.14 to 1.02 mm, indicating that particle size is Tea leaves 8.0 Coffee (50.0), oolong (50.0)
one of the most important factors in anaerobic FW digestion. Vegetables 53.6 Carrot (10.5), spinach (10.1), tomato (8.5),
Smaller particle size increases the surface area available to the radish (7.5), cabbage (6.0), onion (5.5),
microorganisms. On the other hand, it is possible that it accelerates pumpkin (5.0), cucumber (4.5),
komatsuna (4.0), Chinese cabbage (2.2),
the hydrolysis and acidogenesis steps as well as the production of green pepper (2.2), broccoli (2.1),
soluble organic materials such as volatile fatty acids (VFAs), others (31.9)
resulting in excessively high organic loading in the anaerobic Fruits 24.8 Citrus (40.5), apple (26.0), banana (8.0),
digestion reactor. The early stages of the anaerobic solubilization strawberry (4.7), processed fruits (2.5),
others (18.3)
process, especially the hydrolysis and acidogenesis steps, are
Fish 2.7 Saury (100.0)
significantly affected by physicochemical conditions such as Meat 2.2 Bacon (35.5), chicken (26.5), beef (17.4),
temperature and pH rather than by the effects of biological factors mince (17.0), duck (3.6)
(Cha et al., 1992; Komemoto et al., 2009). In such an overloading Egg shell 0.5 Egg shell (100.0)
operation, the imbalance in production and consumption of VFAs
leads to accumulation of VFAs and decreased pH, resulting in
inhibition of the biogas production rate (Griffin et al., 1998; Kim revolutions per minute (II-300, III-1000, IV-1000, V-4000, VI-
et al., 2003; Hori et al., 2006; Ward et al., 2008). 20000, and VII-40000; Table 3).
Ahring et al. (1995) reported that the accumulation of acetate Mesophilic anaerobic sewage sludge was collected from the
occurred faster than that of any other VFA after organic overloading Hokubu Sludge Treatment Center, Yokohama, Japan, in two 20-l tanks.
and that butyrate was a good indicator of organic overloading in The chemical characteristics of this seed sludge are shown in Table 2.
a continuous stirred tank reactor. Ahring et al. (1995) also showed
that VFAs at concentrations of 100e200 mM reduced the overall 2.2. Anaerobic digestion
methane production rate in batch experiments. The inhibition of
methane production rate by organic overloading leads to problems Experiments were carried out using various pretreated
of delayed digestion and inefficient treatment in batch operation substrates (I-DP, II-300, III-1000, IV-1000, V-4000, VI-20000,
(Neves et al., 2004). In the case of continuous operation, the and VII-40000) in mesophilic anaerobic digestion. Eight batch
reduction of the chemical oxygen demand (COD) removal efficiency experiments including a control experiment were conducted in
has been observed as a result of organic overloading (Kim et al., duplicate in 2-l glass reactors (working volume: 1200 ml) at a mean
2002). Therefore, the effects of particle size reduction as mesophilic temperature of 37  1  C. Materials were added to the
a method of pretreatment on the anaerobic digestion process reactor in the following sequence: (1) seeded sludge, (2) substrate
should be evaluated in order to avoid organic overloading. (10 g-COD l1 total reactor volume) and (3) sufficient Milli-QÒ
A bead mill is a grinding device used to reduce the particle size water for a total liquid volume of 1200 ml. The reactors were sealed
of fine particles and microalgae in pretreatment. This device is with silicone stoppers, and a 1-l aluminum gas pack (GL Sciences,
expected to provide smaller particles and higher solubilization AAK-2) was attached for biogas collection. The batch reactors were
compared with traditional pretreatment methods and to enable kept under constant agitation of 80 rpm using a shaker (Taitec,
higher methane production rates. The objective of this study was to NR-150, Japan) for a period of 16 days. The 10 sampling points taken
investigate the effects of particle size on FW soluble COD and for the 16-day experiment were as follows: 0 h, 12 h, D1, D1.5, D2,
methane production; specifically, particle size was modified by D2.5, D3, D5, D10, and D16. Depending on the biogas production
bead milling (BM). Moreover, to clarify the effects of VFAs on the rate, the collection interval varied from once every few hours at the
digestion process, the relationship between particle size and VFA start of the experiment to once every several days. Means of
accumulation was investigated in detail. duplicates are shown in each table and figure.

2. Materials and methods

2.3. Experimental parameters and analytical methods
2.1. Preparation of FW and seed sludge
Total solids (TS), total volatile solids (TVS), pH, total organic
carbon (TOC), total organic nitrogen (TON), total chemical oxygen
The FW used in this study was based on the standard waste
prepared by Izawa et al. (2001), which is based on the composition
Table 2
of actual food waste in Japan (Table 1). The chemical characteristics
Characteristics of food wastes before grinding and seed sludge.
of the collected FW are shown in Table 2. The FW was ground
to reduce the particle size by combining two methods: grinding Parameter Food wastes Seed sludge
with a household disposer (Anaheim, KDF55JK, USA: DP) and with Total solids: TS (% wet weight) 16.5 2.9
a bead mill (Aimex, RMB, Japan). First, 1 l of FW and 1 l of Milli-QÒ Total volatile solids: TVS (% wet weight) 15.5 1.9
TVS/TS (e) 0.94 0.66
water were mixed and ground with a DP (I-DP), as shown in Table 3. pH (e) 6.7 7.7
Next, 150 ml of I-DP was ground by BM in the presence of 150 ml of Salinity (g l1) 0.35 0.39
Milli-QÒ water and 300 ml of glass beads (Aimex, 0.71e1.00 mm). Total COD (g-CODcr l1) 250.5 19.1
Six types of pretreated substrates were prepared by changing the Soluble COD (g-CODcr l1) e 0.8
Total organic carbon (% dry weight) 42.3 32.4
total number of revolutions of the BM between 300 and 40,000
Total organic nitrogen (% dry weight) 3.2 5.6
revolutions by varying the operation time and number of

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Table 3
Operation conditions and characteristics of experiment in this study.

Exp. No. Grinding methods Pretreatment MPSa (mm) Solubilityb (%) SD

Total revolutions (rpm
min) Revolution (rpm) Time (min)

I-DP Disposer e e e 0.888 28.1 0.6
II-300 þ Beads mill 300 300 1 0.843 39.0 3.3
III-1000 þ Beads mill 1000 500 2 0.718 37.3 2.6
IV-1000 þ Beads mill 1000 2000 0.5 0.715 42.1 3.5
V-4000 þ Beads mill 4000 2000 2 0.508 41.4 1.0
VI-20000 þ Beads mill 20,000 2000 10 0.391 39.7 0.7
VII-40000 þ Beads mill 40,000 2000 20 0.393 40.3 1.0
a
Mean particle size.
b
Solubility was calculated as soluble COD/total COD after filtration through 0.45-mm membrane.

demand (total COD), soluble chemical oxygen demand (soluble
COD), long-chain fatty acids (LCFAs), volatile fatty acids (VFAs),
and particle size distribution of the pretreated substrate and
digestion samples were measured. The TS, TVS, TOC, TON, LCFA,
pH, and total COD of pretreated substrates and digestion samples
were measured before filtration with a combusted 0.45-mm glass
filter (Advantec, GC-50). We measured TS and TVS according to
sewage analysis methods of the Japan Sewage Works Association
(1997). The pH of all batch reactors was measured using a pH
meter (Horiba, B-212). By catalytic oxidation, TOC and TON were
measured on an elemental analysis instrument (Fisons, EA1108).
At the start of the experiment, LCFAs (oleate and stearate) in the
loaded substrate were analyzed using the off-line tetramethy-
lammonium hydroxide (TMAH)eGCeMS method. The GCeMS
analyses were carried out on an Agilent 6890N-GC/5973MSD
system equipped with a DB-5MS capillary column (Agilent,
30 m
0.25 mm; film thickness: 0.25 mm). Chromatographic
separation was achieved by using the following temperature
program: 60  C (1 min isothermal), raised at 6  C min1 to 310  C
(20 min isothermal). Helium was the carrier gas at 1.90 ml min1;
the injector temperature was 300  C in spilt injection mode for
1 min. Mass spectra were obtained in El mode (70 eV), scanning in
the range of m/z 50e750. Compound identification was based on
comparison of mass spectra with the NIST02 and Wiley7n library
databases, published spectra, and purchased compounds used as
standards. The values of total COD, soluble COD, and VFA
concentration of the resultant filtrate were quantified in accor-
dance with standard methods of the American Public Health
3. Results and discussion
3.1. Effects of particle size and solubilization
on grinding pretreatment
The mean particle size (MPS) of I-DP was 0.888 mm, which was
the largest of all the pretreated substrates (Table 3 and Fig. 1A). The
MPS of substrates pretreated by BM decreased exponentially from
0.843 to 0.391 mm as the total number of revolutions increased to
20,000, and did not decrease any further at 40,000 total revolu-
tions. The particle size reduction by BM pretreatment of 20,000
total revolutions was 50% greater than that of DP pretreatment.
Comparing pretreatments III-1000 and IV-1000 (Table 3), the size
reduction was enhanced for IV-1000, which had a higher number of
revolutions per minute in comparison with III-1000, which had
a longer operation time. Izawa et al. (2001) have reported that the
1.0 A
0.8
I-DP
Beads mill
0.6
0.4
Mean particle size (mm)

0.2
Association (APHA, 1998). The VFAs (acetic acid, propionic acid, n-
butyric acid, i-butyric acid, n-valeric acid, i-valeric acid) were
measured on a gas chromatograph (Shimadzu, GC-9A, Japan) 0
0 10000 20000 30000 40000
equipped with a packed column (Shincarbon A) and flame ioni-
zation detector. The column temperature was maintained at Total revolution (min rpm)
140  C. The temperature at the injector and detector was main-
tained at 200  C. Helium was used at the carrier gas at a flow rate
1.0
B
of 50 ml min1. The particle size of the substrate was measured
using a laser particle size analyzer (Beckman Coulter, LS320, USA) 0.8
after filtration through a 2.0-mm mesh (Nonaka Rikaki, Testing
Sieve: JIS Z 8801). Particle sizes over 2.0 mm were measured at 0.6
105  C for 4 h. The mean particle size (MPS) was calculated
according to the method of Izawa et al. (2001). The volume of
0.4
biogas was quantified by the water displacement method. Carbon
dioxide and methane were monitored on a gas chromatograph
(Shimadzu, GC-2014AT, Japan) equipped with a packed column 0.2
(Shincarbon ST) and thermal conductivity detector. The injector
and detector temperatures were maintained at 120  C and 260  C, 0
respectively. The column temperature was gradually increased 0 5 10 15 20 25
from 40  C to 250  C. Helium was used as the carrier gas at a flow
Operation time at 2000 rpm (min)
rate of 40 ml min1. The volumes of accumulated methane
and carbon dioxide were corrected for standard conditions of Fig. 1. Effect of grinding process on mean particle size (MPS). (A) Total revolutions
temperature (0  C) and pressure (1 atm). denote grinding condition by bead milling. (B) At 2000 revolutions.

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particle size reduction of FW increased with increasing total
number of revolutions. Furthermore, increasing the number of
revolutions per minute was more effective at reducing particle size
than increasing the operation time, even at the same total number
of revolutions. At the maximum number of revolutions (2000 rpm)
used in this study, MPS decreased exponentially with increasing
operation time up to 10 min, after which there was no further
decrease in MPS (Fig. 1B). These results indicate that particle size
decreased as the number of revolutions per minute increased, and
that long operation times over 10 min were not effective at
reducing MPS. Thus, to reduce the size of FW, the optimal condi-
tions for pretreatment by BM were around 2000 rpm and 10 min.
The COD solubility of substrates pretreated by I-DP was 28.1%
(Table 3). The average COD solubility of substrates pretreated by BM
was 39.8% regardless of the total number of revolutions, and this
value was 42% higher than I-DP. The solubility of FW treated by
different grinding methods, as measured by Izawa et al. (2001), was
about 30%, regardless of the total number of revolutions and
operation time; this solubility was lower than that for the BM
pretreatment in this study. This result is possibly attributable to the
different grinding devices, which have differing efficacy of size
reduction. Grinding the substrate with rotary crushing hammers,
DP treatment produces particle sizes in the millimeter range
(Takezaki et al., 2001). On the other hand, BM is used for dispersing
agglomerated particles and producing small particles in the sub-
micrometer range (Inkyo et al., 2006). As a result, the solubility of
II-300 was found to be 1.39 times higher than that of I-DP in spite of
their similar MPS. These results clearly show that BM pretreatment
effectively promoted solubilization even at a low total number of
revolutions. On the other hand, a high total number of revolutions
were effective at reducing particle size. Therefore, grinding
pretreatment by BM would be an effective method for solubiliza-
tion and particle size reduction of FW.
3.2. Effect of particle size on pH and biogas production
The time course of pH variation in mesophilic anaerobic diges-
tion of FW is shown in Fig. 2. The initial pH value in the conditions
of I-DP, II-300, III-1000, IV-1000, V-4000, VI-20000, and VII-40000
were 7.5, 7.3, 7.3, 7.2, 7.2, 7.2, and 7.2, respectively. In all conditions,
sharp drops in pH to around 6.9 were observed on the first day. The
pH values in all experiments gradually increased and reached
approximately 7.5 by the end of the experimental period. The pH of
the reactor running with only seed sludge (control) was around 7.8
throughout the experimental period. The pH value of VII-40000 in
8.0
7.8
7.6
7.4
pH
7.2
Control IV-1000
7.0 I-DP V-4000
II-300 VI-20000
6.8
III-1000 VII-40000
6.6
0 2 4 6 8 10 12 14 16
Digestion time (day)
Fig. 2. Time course of pH in mesophilic anaerobic digestion of food waste.
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the early phase was lower than that of the other conditions. The
lower pH value of VII-40000 suggests the possibility that excessive
particle size reduction of the substrate accelerated hydrolysis and
acidogenesis in the early stage of anaerobic digestion, resulting in
accumulation of VFAs.
Cumulative biogas production increased with digestion time
(Fig. 3). Considerable increases in biogas production were observed
for the initial 6 days in all experimental conditions. Cumulative
biogas production differed among I-DP, II-300, III-1000, IV-1000,
V-4000, VI-20000, and VII-40000, and reached, respectively, 375,
439, 503, 455, 470, 455, and 404 ml g-total COD1 at the end of
experiment. In the case of the seed sludge (control), biogas was not
detected throughout the 16 days (data not shown). The cumulative
biogas production of III-1000 reached the highest value of
503 ml g-total COD1, which was 34% higher than that of I-DP. The
average concentrations of methane gas were similar among I-DP,
II-300, III-1000, IV-1000, V-4000, VI-20000, and VII-40000, and
were 66.9, 62.1, 64.0, 65.8, 65.5, 65.7, and 62.9%, respectively.
The methane production rate increased with decreasing MPS to
peak at around 330 ml g-total COD1, and then decreased with
more intense pretreatment beyond the maximum value (Fig. 4).
Methane production rate increased 28% when the MPS of FW was
decreased from 0.888 to 0.718 mm by BM pretreatment. Mshandete
et al. (2006) reported that the methane production rate increased
by 22% when the fibers were cut to a size of 2 mm compared with
untreated fibers (100 mm). Smaller MPS makes a larger surface area
available to the microorganisms, resulting in an increased methane
production rate. The results obtained in this study possibly indicate
that particle size reduction increased substrate utilization for
methane fermentation, thereby enhancing the biogas production
rate. These results also suggest that BM pretreatment was effective
for methane production rate even at short operation times. It is
thought that the grinding pretreatment by BM was effective for
improving methane production efficiency to a certain extent,
beyond which methanogenesis was inhibited. In BM pretreatment,
the MPS decreased with increasing operation time and total
number of revolutions. However, the solubility values were similar
at about 40%, regardless of the difference in the number of revo-
lutions per minute and operation time. In anaerobic digestion
experiments using substrates pretreated by BM, high methane
production rates were observed for the smaller MPS substrates.
This suggests that the availability of the particulate substrate as
food for the microorganisms increased with increasing surface area
of the substrate in the digestion period. However, a low methane
production rate for VII-40000 was observed in spite of its
600
Cumulative biogas production
500
(mL g-total COD-1)
400
300
I-DP V-4000
200 II-300 VI-20000
100 III-1000 VII-40000
IV-1000
0
0 2 4 6 8 10 12 14 16 18
18
Digestion time (day)
Fig. 3. Time course of cumulative biogas production in mesophilic anaerobic digestion
of food waste.
 K. Izumi et al. / International Biodeterioration & Biodegradation xxx (2010) 1e8 5

study, the theoretical maximum concentrations of NHþ 4 and H2S

Cumulative methane production

340
were 210 mg N l1 and 11 mg H2S l1, respectively. These low values
320 suggest that process inhibition by NHþ4 and H2S did not occur during
(mL g-total COD-1)

300
280
260
240
I-DP II-300 III-1000 IV-1000
220
V-4000 VI-20000 VII-40000
0 total VFA concentration of 5600 mg l1 was observed in the
0 0.2 0.4 0.6 0.8
Mean particle size (mm)
Fig. 4. Relationship between methane production and mean particle size of pretreated
substrate for each pretreatment condition.
maximum degree of pretreatment, which is possibly the result of
organic overloading due to excessive size reduction.
3.3. Evaluation of inhibitors in anaerobic digestion
Although MPS was smallest for the VII-40000 substrate, cumu-
lative methane production was the lowest at 254 ml g-total COD1
(Fig. 4). Generally, methanogenesis is inhibited by lower pH values,
which result from accumulation of VFAs (Boone and Xum, 1987;
Griffin et al., 1998), accumulation of NHþ
4 (Gallert and Winter,
1997), accumulation of LCFAs such as oleate and stearate
(Angelidaki and Ahring, 1992; Masse et al., 2002), production of H2S
from protein materials (Koster et al., 1986), and accumulation of
persistent substrates (Moen et al., 1997). To evaluate the effects of
inhibition factors such as NHþ
4 , LCFAs, H2S, and VFAs, the concen-
tration of each inhibition factor was measured and calculated. These
concentrations were compared with the corresponding inhibition
levels reported in the literature (Table 4). The theoretical maximum
concentrations of NHþ
4 and H2S were calculated under the
assumption that all nitrogen and sulfur initially present in the
loaded substrate were converted in the batch reactor to NHþ
4 and
H2S, respectively. The oleate and stearate in the FW substrate were
directly measured, and each VFA in the digested sludge was also
measured. It is unlikely that process inhibition was caused by the
accumulation of persistent organic materials (Moen et al., 1997),
because the substrates were loaded only once at the start of the
batch experiment. Hansen et al. (1998) have reported that process
inhibition occurs at the NHþ 4 concentration of 6000 mg l
1
in mes-
ophilic anaerobic digestion of swine manure. In the literature, the
levels of H2S required for inhibition of methane production vary; the
reported IC50 values are 90 mg H2S l1 at pH 7.8e8.0 and
250 mg H2S l1 at pH 6.4e7.2 (Koster et al., 1986). In the present
the batch experiments. The oleate and stearate levels required for
inhibition of methane production have been reported to be
100e200 mg l1 and 500 mg l1, respectively (Angelidaki and
Ahring, 1992). The maximum concentrations of oleate and stea-
rate were 39 mg l1 and 10 mg l1 in the present experiment. These
values were less than 40% of the inhibition levels for both LCFAs.
These low LCFA concentrations suggest that no experimental
conditions were substantially inhibited by LCFAs. In contrast, a high
1.0 1.2 VII-40000 condition. This high concentration was equivalent to 96%
of the VFA inhibition level reported by Siegert and Banks (2005).
Therefore, the high VFA concentration possibly inhibits the diges-
tion process in the VII-4000 condition.
The relationship between the rate of pH change per day during
the early phase of the experiment (from 1 to 3 days) and cumulative
methane production is shown in Fig. 5. This relationship was similar
to the relationship between MPS and cumulative methane produc-
tion. The methane production increased with increasing rate of pH
change to peak at around 0.2 per day, and then decreased thereafter.
The rate of pH change in VII-40000 was the lowest among all
conditions as the increase in pH was most gradual in this condition.
This result suggests that a low recovery rate from decreased pH for
the substrate with the smallest MPS reduced microbial activity,
causing a decrease in methane production. The pH of VII-40000 was
low compared to that of other pretreatment conditions.
In this study, smaller MPS resulted in an increased methane
production rate due to the increased surface area available to the
microorganisms. On the other hand, excessive size reduction of the
substrate caused low pH, resulting in a decreased methane
production rate. Neves et al. (2004) reported that the waste/seed
sludge ratio is an important parameter in batch high-solid anaer-
obic digestion processes, and the use of a large seed sludge amount
in a batch process allows for successful digestion without pH
adjustment. In this study, it was thought that optimization of the
waste/seed sludge ratio could further improve methane produc-
tion, even for substrates with excessively small MPS. For larger MPS
values (>0.7 mm) where the rate of pH change is high, the
microorganisms are not limited by low pH. However, the surface
area available to the microorganisms is smaller, leading to their
starvation. Therefore, the anaerobic biodegradability is not expec-
ted to be enhanced, even if the waste/seed sludge ratio is increased.
3.4. Effect of particle size on the concentration
and characteristics of VFAs
As mentioned above, the decline of pH is caused by the accu-
mulation of VFAs. In the present study, VFA concentrations increased

Table 4
The concentration of inhibitory substances in this study and each inhibition level in the previous studies.

Inhibitors Inhibitor concentration Inhibition level Inhibitor concentration/Inhibitor Reference

in this study a (mg l1) b (mg l1) level a/b (%)
NH4eNþa 210 6000 3.5 Hansen et al. (1998)
H2Sb 11 90e250 4.2e12 Koster et al. (1986)

LCFA
Oleate 39 100e200 20e39 Angelidaki and Ahring (1992)
Stearate 10 500 2.0 Angelidaki and Ahring (1992)

Total VFAb 4200e5600 6000 70e93 Siegert and Banks (2005)

Propionic acid 930e1300 3000 31e43 Boone and Xum (1987)
a
Calculated maximum values.
b
Total VFA was calculated as sum of each VFA.

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6 K. Izumi et al. / International Biodeterioration & Biodegradation xxx (2010) 1e8

340 from the start of the experiments to day 2 (Fig. 6), and the maximum
Cumulative methane production

total VFA concentration for I-DP, II-300, III-1000, IV-1000, V-4000,

320 VI-20000, and VII-40000 was 4330, 4210, 4270, 4750, 4500, 5030,
(mL g-total COD-1)

and 5600 mg l1, respectively. Acetic acid, butyric acid, and propionic
300
acid accounted for most of the total VFA concentration. Generally,
280 macromolecular organic compounds such as carbohydrates and
lipids are decomposed and transformed to VFAs and alcohol via
260 biochemical reactions such as hydrolysis and beta-oxidation (Speece,
1996). The microorganisms that produce methane gas consume only
240 acetic acid and hydrogen. Other VFAs such as butyric acid are
I-DP II-300 III-1000 IV-1000 transformed into acetic acid, and then acetic acid is converted to
220
V-4000 VI-20000 VII-40000 methane and carbon dioxide by methanogenic bacteria (Mata-
0 Alvarez, 2003). In this study, all VFA concentrations reached
0 0.1 0.2 0.3 0.4 a maximum value 1e3 days from the start of the experiment and
Rate of pH change at early phase then sharply decreased within 6 days, with the exception of pro-
pionic acid (Fig. 7). Propionic acid, which is known to degrade slowly,
Fig. 5. Relationship between methane production and rate of pH change in the initial was also consumed within 16 days. The maximum values of acetic
phase for each pretreatment condition. acid and propionic acid generated at the initial stages of the

6000 6000
I-DP V-4000
5000 5000

4000 4000

3000 3000

2000 2000

1000 1000

0 0
6000 6000
II-300 VI-20000
5000 5000

4000 4000

3000 3000
VFAs concentration(mg L-1)

2000 2000

1000 1000

0 0
6000 6000
III-1000 VII-40000
5000 5000
4000 4000
3000 3000
2000 2000

1000 1000

0 0
6000 0 2 4 6 8 10 12 14 16 18
IV-1000
5000

4000

3000

2000

1000

0
0 2 4 6 8 10 12 14 16 18
Digestion time (day)

Acetic acid Propionic acid Butyric acid Total-VFAs

Fig. 6. Variation of VFA production as a function of digestion time for different grinding processes.

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 K. Izumi et al. / International Biodeterioration & Biodegradation xxx (2010) 1e8 7

3000 3000
Acetic acid Butyric acid
2500 2500

2000 2000

1500 1500

VFAs concentration (mg L-1)

1000 1000
500 500
0 0

3000 6000
Propionic acid Total-VFAs
2500 5000

2000 4000

1500 3000

1000 2000

500 1000

0 0
0 2 4 6 8 10 12 14 16 18 0 2 4 6 8 10 12 14 16 18

Digestion time (day)

I-DP II-300 III-1000 IV-1000

V-4000 VI-20000 VII-40000

Fig. 7. Variation of VFA production as a function of digestion time for production of different VFAs.

experiments increased with decreasing MPS (Fig. 8). Maximum These VFAs are converted by methanogenic bacteria to methane
butyric acid production decreased with decreasing MPS and and carbon dioxide (Thiele and Zeikus, 1988). The formation and
comprised the main component in the maximum total VFAs with consumption of some VFAs, such as propionic and butyric acid,
values of 48.4, 63.4, 60.8 55.7, 40.2, 37.0, and 31.7% in the I-DP, II-300, demonstrate the conversion to acetic acid by acetogenic bacteria;
III-1000, IV-1000, V-4000, VI-20000, and VII-40000 treatments, acetic acid is considered to be the major precursor of methane.
respectively (Figs. 7 and 8). These results indicate that the difference Shimizu et al. (1992) reported that the VFA concentration of wasted
in MPS values affected the production and composition of VFAs. activated sludge pretreated by ultrasonic pretreatment was higher
The maximum cumulative concentration of VFAs varied with than that of the non-pretreated control. Comparing the solubility of
MPS. Acetic acid and propionic acid increased while butyric acid high-molecular-weight biological polymers with the VFA produc-
decreased with decreasing MPS. The amount of seed sludge in all tion rate of pretreated wasted activated sludge, the VFA production
conditions was fixed in this study. Therefore, the difference in rate was higher (Shimizu et al., 1992). These observations indicate
production of VFAs may have been caused by the increased surface that the conversion of VFAs from pretreated wasted activated
area of the substrate and solubilization of intracellular macromol- sludge to methane was the rate-limiting step. In the present study,
ecules due to mechanical breakage by grinding. excessive reduction of the MPS caused VFA accumulation, resulting
in decreased methane production. On the other hand, smaller MPS
improved acetic acid accumulation and led to increased methane
production. Smaller MPS values also increased the solubility in the
3000 anaerobic digestion process.
Maximum VFA concentration (mg L-1)

2500
4. Conclusions

2000 The following conclusions can be drawn from this study

regarding the effects of particle size and solubilization with
1500 pretreatment by BM in anaerobic digestion processes.

1000 1. Grinding pretreatment by BM resulted in 30% higher solubili-

zation than pretreatment with a DP, even at 300 revolutions,
500 and yielded a particle size that was 57% smaller at 20,000
Acetic acid Butyric acid Propionic acid revolutions.
2. Smaller MPS improved the methane yield by 28% at 1000
0
0 0.2 0.4 0.6 0.8 1.0 1.2 revolutions compared with I-DP, and enhanced VFA production
with an increasing total number of revolutions.
Mean particle size (mm)
3. Excessive size reduction of the substrate caused VFA accumu-
Fig. 8. Relationship between maximum VFA concentration and mean particle size in lation, resulting in decreased methane production and
mesophilic anaerobic digestion of food waste. decreased solubility in the anaerobic digestion process.

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8 K. Izumi et al. / International Biodeterioration & Biodegradation xxx (2010) 1e8
4. Optimizing the size reduction of the substrate as well as the
microbial growth could potentially improve the methane yield
of anaerobic digestion processes.
These results suggest that size reduction by BM pretreatment
effectively promoted solubilization and increased methane yield in
the anaerobic digestion process. However, the energy required for
grinding the FW was not measured, and should be evaluated from
the point of view of energy balance. In this experiment, we inves-
tigated the effects of size reduction as pretreatment in the batch
anaerobic digestion process. The conclusions reached in this study
apply to small-scale batch operations, and further investigation is
necessary to relate these findings to the operation of full-scale
continuous digestion processes. Furthermore, we plan to investi-
gate the effects of size reduction using persistent organic substrates
such as seaweed and phytoplankton in a future study. The effects of
the seed sludge concentration and the composition of the bacterial
community are also of interest.
Acknowledgements
This work was supported by the University-Industry Joint
Research Project for Private Universities and matching funds from
the Ministry of Education, Culture, Sports, Science and Technology,
Japan, MEXT, 2004e2008. We are grateful to the Hokubu Sludge
Treatment Center, Yokohama, Japan, for preparation of the seed
sludge.
References
Ahring, B.K., Sandberg, M., Angelidaki, I., 1995. Volatile fatty acids as indicators of
process imbalance in anaerobic digesters. Applied Microbiology and Biotech-
nology 43, 559e565.
American Public Health Association, 1998. Standard methods for the examination of
water and wastewater, 20th ed. American Public Health Association, American
Water Works Association, and Water Environment Federation, Washington DC,
USA.
Angelidaki, I., Ahring, B.K., 1992. Effects of free long-chain fatty acids on thermophilic
anaerobic digestion. Applied Microbiology and Biotechnology 37, 808e812.
Boone, D.R., Xum, L., 1987. Effect of pH, temperature and nutrients on propionate
degradation by a methanogenic enrichment culture. Applied and Environ-
mental Microbiology 53, 1589e1592.
Cha, G.C., Li, Y.-Y., Noike, T., 1992. Effect of temperature and hydraulic retention time
on the characteristics of anaerobic acidogenesis in the low temperature range.
Process of Environment and Sanitary Engineering Research 28, 29e37.
Eastman, J.A., Ferguson, J.F., 1981. Solubilization of particulate organic carbon during
the acid phase of anaerobic digestion. Journal Water Pollution Control Feder-
ation 53, 352e355.
Engelhart, M., Kruger, M., Kopp, J., Dicht, N., 1999. Effect of disintegration on
anaerobic degradation of sewage excess sludge in downflow stationary fixed
film digesters. In: Mata-Alvarez, J., Tilche, A., Cecchi, F. (Eds.), Proceedings of the
second international symposium on anaerobic digestion of solid wastes, Bar-
celona, pp. 153e160.
Gallert, C., Winter, J., 1997. Mesophilic and thermophilic anaerobic digestion of
source-sorted organic wastes: effect of ammonia on glucose degradation and
methane production. Applied Microbiology and Biotechnology 48, 405e410.
Gomec, C.Y., Kim, M., Ahn, Y., Speece, R.E., 2002. The role of pH in mesophilic
anaerobic sludge solubilization. Journal of Environmental Science and Health
10, 1871e1878.
Griffin, M.E., McMahon, K.D., Mackie, R.I., Raskin, L., 1998. Methanogenic population
dynamics during start-up of anaerobic digesters treating municipal solid waste
and biosolids. Biotechnology and Bioengineering 57, 342e355.
Hansen, K.H., Angelidaki, I., Ahring, B.K., 1998. Anaerobic digestion of swine
manure: inhibition by ammonia. Water Research 32, 5e12.
Hori, T., Haruta, S., Ueno, Y., Ishii, M., Igarashi, Y., 2006. Dynamic transition of
a methanogenic population in response to the concentration of volatile fatty
Please cite this article in press as: Izumi, K., et al., Effects of particle size on anaerobic digestion of food waste, International Biodeterioration &
Biodegradation (2010), doi:10.1016/j.ibiod.2010.06.013
acids in a thermophilic anaerobic digester. Applied and Environmental Micro-
biology 72, 1623e1630.
Inkyo, M., Tahara, T., Iwaki, T., Iskandar, F., Hogan Jr., C.J., Okuyama, K., 2006. Exper-
imental investigation of nanoparticle dispersion by beads milling with centrif-
ugal bead separation. Journal of Colloid and Interface Science 304, 535e540.
Izawa, S., Somiya, I., Fujii, S., Park, C.W., Kishimoto, N., 2001. Solubilization and
break down in size of particulate raw garbage with stirring grinder. Environ-
mental and Sanitary Engineering Research 15, 54e58.
Japan Sewage Works Association, 1997. Examination of water quality. In:
Kashiwaya, K. (Ed.), Sewage examination methods, Vol. 1. Japan Sewage Works
Association, Tokyo, Japan.
Kim, I.S., Kim, D.H., Hyun, S.-H., 2000. Effect of particle size and sodium ion
concentration on anaerobic thermophilic food waste digestion. Water Science
and Technology 41, 67e73.
Kim, M., Ahn, Y.-h., Speece, R.E., 2002. Comparative process stability and efficiency of
anaerobic digestion; mesophilic vs. thermophilic. Water Research 36, 4369e4385.
Kim, J., Park, C., Kim, T., Lee, M., Kim, S., Kim, S.-W., Lee, J., 2003. Effect of various
pretreatments for enhanced anaerobic digestion with waste activated sludge.
Journal of Bioscience and Bioengineering 95, 271e275.
Komemoto, K., Lim, Y.G., Nagao, N., Onoue, Y., Niwa, C., Toda, T., 2009. Effect of
temperature on VFA’s and biogas production in anaerobic solubilization of food
waste. Waste Management 29, 2950e2955.
Koster, I.W., Rinzema, A., De Vegt, A.L., Lettinga, G., 1986. Sulfide inhibition of the
methanogenic activity of granular sludge at various pH levels. Water Research
20, 1561e1567.
Li, Y.Y., 2004. Recent development of biomethanation technology. Journal of Japan
Society on Water Environment 27, 622e626.
Li, Y.Y., Noike, T., 1992. Upgrading of anaerobic digestion of waste activated sludge
by thermal pretreatment. Water Science and Technology 26, 857e866.
Masse, L., Masse, D.I., Kennedy, K.J., Chou, S.P., 2002. Neutral fat hydrolysis and long-
chain fatty acid oxidation during anaerobic digestion of slaughterhouse
wastewater. Biotechnology and Bioengineering 79, 43e52.
Mata-Alvarez, J., 2003. Biomethanization of the organic fraction of a municipal solid
wastes. IWA Publishing, London.
Metcalf and Eddy Inc., 2004. Wastewater engineering: treatment and reuse.
McGraw Hill, New York.
Moen, E., Horn, S., Ostgaard, K., 1997. Biological degradation of Ascophyllum nodo-
sum. Journal of Applied Phycology 9, 347e357.
Mshandete, A., Bjornsson, L., Kivaisi, A.K., Rubindamayugi, M.S.T., Mattiasson, B.,
2006. Effect of particle size on biogas yield from sisal fiber waste. Renewable
Energy 31, 2385e2392.
Nah, I.W., Kang, Y.W., Hwang, K.-Y., Song, W.-K., 2000. Mechanical pretreatment of waste
activated sludge for anaerobic digestion process. Water Research 34, 2362e2368.
Neves, L., Oliveria, R., Alves, M.M., 2004. Influence of inoculum activity on the bio-
methanization of a kitchen waste under different waste/inoculum rations.
Process Biochemistry 39, 2019e2024.
Palmowski, L., Muller, J., 1999. Influence of the size reduction of organic waste on
their anaerobic digestion. In: Mata-Alvarez, J., Tilche, A., Cecchi, F. (Eds.),
Proceedings of the second international symposium on anaerobic digestion of
solid wastes, Barcelona, pp. 137e144.
Sawayama, S., Inoue, S., Minowa, T., Tsukahara, K., Ogi, T., 1997. Thermochemical
liquidization and anaerobic treatment of kitchen garbage. Journal of Fermen-
tation and Bioengineering 83, 451e455.
Sharma, S.K., Mishra, I.M., Sharma, M.P., Saini, J.S., 1988. Effect of particle size on
biogas generation from biomass residues. Biomass 17, 251e263.
Shimizu, T., Kudo, K., Nasu, Y., 1992. Ultrasonic treatment of excess sludge as a pretreat-
ment stage for anaerobic digestion. Journal of Water and Waste 34, 221e226.
Siegert, I., Banks, C., 2005. The effect of volatile fatty acid additions on the anaerobic
digestion of cellulose and glucose in batch reactors. Process Biochemistry 40,
3412e3418.
Speece, R.E., 1996. Anaerobic biotechnology for industrial wastewaters. Archae
Press, Tennessee, USA.
Takezaki, Y., Uryu, K., Tuji, T., Shimizu, Y., Sankai, T., 2001. Study on estimation of
biological dissolved rate for the crushed garbage using a disposer. Japanese
Journal of Water Treatment Biology 37, 173e181.
Thiele, J.H., Zeikus, J.G., 1988. Control of interspecies electron flow during anaerobic
digestion: the role of formation versus hydrogen transfer during syntrophic
methanogenesis in flocs. Applied and Environmental Microbiology 54, 20e29.
Ward, A.J., Hobbs, P.J., Holliman, P.J., Jones, D.L., 2008. Optimisation of the anaerobic
digestion of agricultural resources. Bioresource Technology 99, 7928e7940.
Xu, H.L., Wang, J.Y., Zhang, H., Tay, J.H., 2002. A comparative study of anaerobic
digestion of food waste in a single pass, a leachate recycle and coupled solid/
liquid reactors. Water Science and Technology 46, 319e324.
Zhang, B., Zhang, L.L., Zhang, S.C., Shi, H.Z., Cai, W.M., 2005. The influence of pH on
hydrolysis and acidogenesis of kitchen wastes in two-phase anaerobic diges-
tion. Environmental Technology 26, 329e339.