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Profile of hydrolases and biogas production

during two-stage mesophilic anaerobic
digestion of solid potato waste

Article in PROCESS BIOCHEMISTRY September 2005

DOI: 10.1016/j.procbio.2005.01.010

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 Process Biochemistry 40 (2005) 29452952
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Profile of hydrolases and biogas production during two-stage

mesophilic anaerobic digestion of solid potato waste
W. Parawira a,b, M. Murto a, J.S. Read c, B. Mattiasson a,*
a
Department of Biotechnology, Lund University, PO Box 124, SE-221 00 Lund, Sweden
b
Department of Biochemistry, University of Zimbabwe, PO Box MP167, Mt. Pleasant, Harare, Zimbabwe
c
Department of Applied Biology and Biochemistry, NUST, PO Box 939 Ascot, Bulawayo, Zimbabwe
Received 24 March 2004; accepted 12 January 2005

Abstract

Two two-stage systems, one consisting of a solid-bed reactor for hydrolysis/acidification connected to an upflow anaerobic sludge blanket
methanogenic reactor, and the other consisting of a solid-bed reactor connected to a methanogenic reactor packed with wheat straw biofilm
carriers, were investigated with regard to hydrolytic enzymes and methane production during mesophilic anaerobic digestion of solid potato
waste. Some of the enzymes used by microorganisms to degrade the potato were found to be amylase, carboxymethyl cellulase, filter paper
cellulase, xylanase, pectinase and protease. Both free and cell-bound enzyme activities were measured. The activity of the free enzyme was
higher than that of the cell-bound for all the enzymes. The amylase activity was highest, followed by carboxymethyl cellulase and filter paper
cellulase, while the other hydrolytic enzymes had low activities. The methane yield (0.39 m3/kg VSadded) and the cumulative methane
production was the same in both systems. The system with packed straw bed degraded the organic matter faster than the system with the
UASB methanogenic reactor.
# 2005 Elsevier Ltd. All rights reserved.

Keywords: Anaerobic digestion; Biogas; Methane yield; Hydrolytic enzymes; Amylase; Cellulase; Xylanase; Solid potato waste

1. Introduction
Anaerobic digestion of solid organic waste has gained
increased attention as a means of producing energy and
reducing problems associated with the disposal of organic
waste [1]. Anaerobic digestion is a multiple-stage process in
which hydrolysis is one of the main steps. During hydrolysis
the complex insoluble substrate macromolecules are hydro-
lysed into simpler and more soluble intermediates by bacteria.
A large number of microbial species, acting in concert, are
capable of utilising organic substrates such as carbohydrates,
proteins and lipids to produce volatile fatty acids (VFAs),
which can be converted to methane and carbon dioxide by
methanogenic microorganisms. Bacteria excrete enzymes
that hydrolyse the particulate substrate to small transportable
molecules, which can pass through the cell membrane. Once
* Corresponding author. Fax: +46 46 2224713.
E-mail address: bo.mattiasson@biotek.lu.se (B. Mattiasson).
inside the cell, these simple molecules are used to provide
energy and to synthesize cellular components. Polysacchar-
ides are converted to simple sugars; hydrolysis of cellulose by
the cellulase enzyme complex yields glucose; hemicellulose
degradation results in monosaccharides such as xylose,
glucose, galactose, pentoses, arabinose and mannose, while
starch is converted to glucose by amylase enzymes [24].
It is generally considered that two-stage anaerobic
digestion is more effective than the classical one-stage
process in the conversion of solid substrates to biogas [57].
In this process, an active hydrolytic and acid-forming
microbial population is maintained in a first acid-forming
reactor and a methane-forming microbial population is
maintained in a second reactor. The acidogens and
methanogens have different physical, biochemical and
environmental requirements. A successful two-stage anae-
robic digestion system based on physiological separation of
these two groups of microbes should theoretically result in a
more efficient anaerobic digestion system [8].

1359-5113/$ see front matter # 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.procbio.2005.01.010
2946 W. Parawira et al. / Process Biochemistry 40 (2005) 29452952

Investigations have been made into the possibility of

using hydrolytic activity to monitor digestion in a number of
waste treatment processes, including landfilling, aerobic
digestion and composting [9,10]. This would thus provide a
reliable and direct measure of the functioning of such
processes, as an alternative to conventional methods, from
which the performance can only be inferred [10]. Conven-
tional process parameters such as concentration of volatile
fatty acids, gas composition and removal of chemical
oxygen demand determine the end products of metabolism
and are indirect measures of microbial activity [11]. Little
information is available on the hydrolytic enzymes present
in anaerobic digesters fed with solid potato waste. The initial
hydrolytic process when degrading solid potato waste is
important, as the waste is not a pre-digested material. Potato
is composed of approximately 80% water (w/w), 17%
carbohydrate, 2% protein, 0.1% fat and 0.9% vitamins,
inorganic minerals and trace metals, thus providing a good
substrate for microorganisms [12]. The present study
constitutes part of our efforts to understand the anaerobic
process with respect to the hydrolytic enzyme activities
acting on solid biomass during anaerobic digestion. Fig. 1. Two-stage experimental set-up for the anaerobic digestion of solid
In this study, we evaluated the performance of an upflow potato waste.
anaerobic sludge blanket (UASB) reactor and a reactor
packed with straw as a biofilm carrier in the methanogenic
stage of the two-stage anaerobic digestion of potato solids. reactor. This plant receives sewage sludge and industrial
The raw material studied here can be regarded as being effluents mainly from a potato-processing plant.
representative of many other kinds of starch-rich biomass Solid potato waste was cut into small pieces using a
suitable for conversion to biogas. kitchen blender and 1.0 kg was loaded into AR1 and AR2.
The potato waste had a TS content of 19% and a VS content
of 95% of the TS. To both reactors 0.8 l water was added,
2. Materials and methods together with 0.2 l of anaerobic sludge inocula (described
above). The reactors were sealed at the top with butyl rubber
2.1. Experimental set-up stoppers to maintain anaerobic conditions. A wire sieve
(3 mm gauge) was installed 5 cm above the bottom of each
Experiments were performed in two different two-stage reactor to support the solid waste substrate while allowing
systems, each consisting of an acidogenic reactor and a the liquid to pass through it. The leachate in AR1 and AR2
methanogenic reactor, as illustrated in Fig. 1. System I was recirculated at 9 ml/min and sprinkled over the packed-
included an acidogenic reactor (AR1) connected to a UASB bed of potato. Feeding of leachate to the methanogenic
methanogenic reactor (active volume 0.84 l). System II reactors was started after 24 h of recirculation of AR1 and
consisted of an acidogenic reactor (AR2) connected to a AR2. The organic loading rate (OLR) applied to the UASB
packed-bed methanogenic reactor (active volume 1.0 l) with reactor ranged from 1.4 to 3.0 g COD/l/day and from 1.0 to
pre-digested straw as a biofilm carrier. The pre-digestion 3.0 g COD/l/day for the packed-bed reactor with straw
step was achieved by incubating the packed straw bed with carriers (COD is chemical oxygen demand). The overflow
water recirculation for more than 3 months at 37 8C, to from the methanogenic reactors was recycled back to the
ensure that all the easily degradable compounds had been hydrolysis/acidification reactors to replenish the acidogenic
digested. Both AR1 and AR2 reactors were identical and had reactors and to provide buffering capacity to prevent
a working capacity of 2.0 l. Both the acidogenic and excessive acidification. The experiments were run for a
methanogenic reactors were operated at 37 8C. period of up to 50 days at which time biogas production in
The inoculum (0.2 l) used in the UASB reactor was the methanogenic reactors was insignificant, and the potato
granular sludge from a full-scale UASB reactor processing waste had been completely degraded in the acid reactors.
papermill wastewater (the Netherlands). Anaerobic sludge
with a total solids (TS) content of 1.7% and volatile solids 2.2. Analytical methods
(VS) content of 59% of TS from the Ellinge Municipal
Wastewater Treatment Plant in Eslo v, southern Sweden, was Samples from the hydrolysis/acidification reactors were
used as inoculum in the straw packed-bed methanogenic analysed to determine enzyme activities during the study
 W. Parawira et al. / Process Biochemistry 40 (2005) 29452952
Table 1
pH and temperature optima for hydrolytic enzymes in anaerobic digestion
of potato solids
Enzyme pH Temperature (8C)
Amylase 5.0, 9.0 50
Carboxymethyl cellulase 5.0 60
Xylanase 6.0 50
Pectinase 6.0 50
Protease 7.0, 9.0 50
Filter paper cellulase 6.0 50
period. Several parameters including pH, alkalinity, con-
centration of VFAs, soluble COD, and gas composition,
were measured in samples taken from both the acidogenic
and the methanogenic reactors.
Since the optimum conditions for assaying activity were
unknown it was necessary to determine the pH and
temperature optima for each enzyme assay. Studies on
temperature optimisation for each type of enzyme analysis
were performed in sodium acetate buffer (pH 6.0, 0.05 M)
by incubating the reaction mixtures for 1 h at 2090 8C.
Studies on pH activity optimum for each type of enzyme
analysis were carried out by incubating the reaction mixtures
at the optimal temperature in buffers with pH ranging from 4
to 10. The activity analysis of the enzymes was performed at
optimal pH and temperature conditions (Table 1).
Enzyme activities were determined by centrifuging a
10 ml liquid sample at 3000  g for 10 min. The supernatant
was collected and used for free enzyme activity assays. The
pellet was washed twice by re-suspension in 10 ml
potassium dihydrogen phosphate buffer (pH 7.0, 0.1 M),
and centrifuged again at 3000  g for 10 min. The pellet was
re-suspended in 2 ml 0.05 M sodium acetate buffer at pH 6.0
(assay buffer) for analysis of cell-bound enzyme activity
according to the method of Fay et al. [13]. Both the
supernatant fraction and cell fraction were kept on ice until
enzyme activity determination. The chemicals used in
enzyme assays were all obtained from Sigma (St. Louis,
MO, USA). In all the enzyme analyses, the total reducing
sugars produced were measured with the dinitrosalicylate
reagent method at 540 nm using an Ultrospec 1000 UV/
visible spectrophotometer (Biochron Ltd., Cambridge, UK).
Glucose was used as the standard unless otherwise stated
[14].
Amylase activity was measured using the method of
Giraud et al. [15]. Appropriately diluted culture samples
were incubated in pre-warmed soluble starch in sodium
acetate buffer (pH 6.0, 0.05 M) at 50 8C. Enzyme activity
was expressed as mmol glucose released/min. To measure
carboxymethyl cellulase (CMCase), the samples were
incubated with carboxymethyl cellulose in sodium acetate
buffer (pH 6.0, 0.05 M), pre-warmed to the incubation
temperature, 60 8C [16,17]. To measure filter paper activity
(FPase) the samples were incubated with Whatman No. 1
filter paper strips in sodium acetate buffer (pH 6.0, 0.05 M),
pre-warmed to the incubation temperature, 50 8C. In each
2947
case one unit of enzyme activity was defined as the amount
of enzyme that releases 1 mmol of glucose equivalent/min
[18]. Xylanase (endo-1,4-(-D-xylan xylanohydrolase) activ-
ity was assayed using oat spelt xylan as substrate [19]. The
amount of reducing sugars released was estimated using
xylose as a standard. One unit of enzyme activity was
defined as the amount of enzyme that releases 1 mmol of
xylose equivalent/min. Pectinase (polygalacturonase) activ-
ity was measured by incubating samples with pre-warmed
pectin (from citrus fruits) in sodium acetate buffer (pH 6.0,
0.05 M) at 50 8C. Reducing sugars were estimated using
monogalacturonic acid as standard [14]. One unit of enzyme
activity was defined as the amount of enzyme that releases
1 mmol of monogalacturonic acid equivalent/min [18].
Protease activity was measured by incubating the sample
with pre-warmed azocasein in TrisHCl buffer (pH 7.4,
0.1 M) at 50 8C for 1 h [20]. The reaction was stopped by
adding 2 ml 10% (w/v) trichloroacetic acid. The mixture
was then centrifuged at 3000  g for 10 min and the
absorbance (OD) of the supernatant was read at 380 nm. The
activity of the protease was expressed in arbitrary units,
where one unit was equivalent to an OD380 change of
0.1 nm/min compared with the absorbance of the enzyme
blank [21].
The biogas composition was measured using Varian 3350
gas chromatography (Walnut Creek, CA, USA) fitted with a
Haysep Q 80/100 mesh column, a molecular sieve column
and a thermal conductivity detector was used. Helium was
used as the carrier gas at a flow rate of 12 ml/min. The
column temperature was 70 8C and the injector and detector
temperatures were 110 and 150 8C, respectively. The
compounds detected were methane, carbon dioxide, nitro-
gen and oxygen. The biogas produced in the reactors was
collected in gas-tight bags. The volume of the biogas
produced was measured using a wet-type precision gas
meter (Schlumberger, Karlsruhe, Germany). The TS, VS and
COD were determined according to standard methods [22].
The concentrations of VFAs were determined using HPLC
according to Bjo rnsson et al. [23].
3. Results
The optimal pH and temperature values for the enzymes
are given in Table 1. Amylase showed optimal activity at pH
5.0 and 9.0, but the enzyme was assayed only at 5.0.
CMCase showed optimal activity at pH 5.0, while xylanase,
pectinase and FPase activities were optimal at pH 6.0.
Protease activity was optimal at pH 7.0, although there were
small peaks at pH 4.0 and 9.0. The activity was assayed at
pH 7.0. Multiple pH peaks are probably due to hetero-
geneous mixtures consisting of several inherent enzymes.
The studies on the effect of temperature indicated optimal
enzyme activities at 50 8C for amylase, xylanase, protease,
pectinase and FPase, and at 60 8C for CMCase. The assays
were thus performed at these two temperatures.
2948 W. Parawira et al. / Process Biochemistry 40 (2005) 29452952

Fig. 2. Production, with time, of free and cell-bound amylase (a), carboxymethyl cellulase (b), filter paper activity (c), xylanase (d), pectinase (e) and protease
(f) during two-stage anaerobic digestion of solid potato waste. (^) free enzyme activity in AR1; (^) cell-bound activity in AR1; (~) free enzyme activity in
AR2; (~) cell-bound activity in AR2.

The activity of both free and cell-bound enzyme was
measured for a range of hydrolytic enzymes. In general, the
activity of free enzymes was higher than that of cell-bound
enzymes for all the enzymes assayed: amylase, CMCase,
FPase, xylanase, protease and pectinase (Fig. 2). The free
protease activity was higher than that of the cell-bound, during
the first 20 days after which the cell-bound activity was higher
than the free activity until the end of the hydrolysis period
(Fig. 2f). It was furthermore seen that the maximal activities
of the different enzymes occurred at different times during the
study period. Starch-hydrolysing and pectin-hydrolysing
enzymes exhibited their maximum activities early during
the study period, while the maximum activities of the
cellulases, xylanase and protease appeared later during
incubation. A comparison of the enzyme activities in AR1 and
AR2 shows some similarities in the levels and times of peak
activity. The total amylase activity ranged from 2.4 to 4.4 IU/
ml in AR1 and from 2.2 to 3.7 IU/ml in AR2. The total
CMCase activity varied between 0.3 and 0.6 IU/ml in both
hydrolysis/acidification reactors. The total FPase activity
varied between 0.3 and 0.5 IU/ml in both AR1 and AR2. The
total xylanase activity ranged between 0.2 and 0.6 IU/ml in
AR1 and between 0.2 and 0.4 IU/ml in AR2. Pectinase and
protease activity levels were low in both acidogenic reactors.
The maximum total pectinase activity reached was 0.004 IU/
ml in both reactors. The maximum total protease activity was
0.01 IU/ml in both AR1 and AR2. All the enzymes showed a
decline in activity at the end of the digestion period.
 W. Parawira et al. / Process Biochemistry 40 (2005) 29452952
Fig. 3. Methane content and cumulative methane production by the 2 two-
stage systems. (^) %CH4, UASB; (^) %CH4, Straw bed; (~) %CH4,
AR1; (~) %CH4, AR2; (*) CH4 production, UASB; (*) CH4 production,
Straw bed.
The methane content and cumulative methane production
during the two-stage digestion of solid potato waste are
shown in Fig. 3. The methane content varied between 67 and
77% in the UASB reactor and between 69 and 82% in the
packed-bed reactor. The cumulative methane production
was 70 l in both methanogenic reactors. There was little
methane production in the acidogenic reactors until day 30,
when the methane content gradually increased to 53% in
AR2, at which time the soluble COD and VFA concentra-
tions were very low (data not shown). The methane content
increased from day 38 in AR1 and peaked at 56% during the
final days of operation, at which time the COD and VFA
concentrations were very low. The methane yield was
(0.39 m3/kg VSadded) in both the methanogenic reactors and
is compared with results in literature in Table 2. The System
II with the straw packed-bed performed better in degrading
the potato solid waste and completed the digestion in 38
days compared with 50 days for system I with the UASB
reactor. Organic matter digestion was considered complete
when the methane production was insignificant.
The pH remained between 4.0 and 5.0 for about 30 days
in AR1 and for 24 days in AR2, and thereafter increased to
around 7.0 in both reactors (Fig. 4). The pH in the methane
reactors remained at around 7.8 throughout the operation of
the systems. The total VFAs (TVFAs) increased rapidly,
reaching concentrations of 23 g/l in AR1 in 10 days and
20 g/l in AR2 in 8 days. The maximum amount of TVFAs
achieved was 24 and 22 g/l in AR1 and AR2, respectively.
Acetic, propionic and butyric acids were the major VFAs
Table 2
Comparisons of our results with methane yield data found in the literature
Feed System
Solid potato waste Two-stage lab-scale
Potato waste CSTR 20 l continuous
Food waste Two-stage pilot-scale
Sugar beet pulp Two-stage pilot-scale
Fruit and vegetable mixture Two-stage lab-scale
2949
Fig. 4. Profiles of pH in the acidogenic reactors and methanogenic reactors,
together with total VFAs in the acidogenic reactors. (~) pH, AR1; (~) pH,
AR2; (^) pH, UASB; (^) pH, Straw bed; (*) TVFAs, AR1; (*) TVFAs,
AR2.
produced, with low amounts of valeric acids (data not
shown). The TVFAs then decreased towards the end of the
study to 1 g/l.
Effective biodegradation in the methanogenic reactors
was evidenced in terms of COD removal efficiencies, which
ranged from 80 to 97% during the period of the study
(Fig. 5). The soluble COD increased to a maximum of 41 g/l
in AR1 and 37 g/l in AR2 after 8 days, and decreased to
around 4 g/l at the end of operation (Fig. 5). Soluble COD is
a parameter that represents the degree of hydrolysis and
solubilisation achieved by the acidogenic bacteria.
4. Discussion
In this study, amylase, CMCase, FPase, xylanase,
protease and pectinase enzyme activity was demonstrated
during the hydrolysis of solid potato waste in anaerobic
digestion. It is useful to determine enzyme activities over
time since hydrolytic enzyme activity levels can be used to
monitor waste treatment processes [10,24]. Assuming that
enzymatic breakdown is the dominant mechanism in
hydrolysis, then determination of enzymatic activity is
one of the direct ways to study the hydrolysis process.
Higher free enzyme activities than cell-bound enzyme
activities were expected since the efficiency in degrading
Temperature (8C) CH4 yield (m3 CH4/kg VSadded) Reference
37 0.39 This study
3337 0.43 Stewart et al. [29]
3538 0.44 Lee et al. [34]
35 0.36 Weiland [35]
35 0.51 Virtua et al. [36]
2950 W. Parawira et al. / Process Biochemistry 40 (2005) 29452952
Fig. 5. Temporal variation of organic matter solubilisation in terms of
soluble COD and percentage COD reduction efficiency in the methanogenic
reactors. (^) COD, AR1; (^) COD, AR2; (~) COD reduction, UASB;
(~) COD reduction, straw-bed.
particulate substrates are far higher for an excreted enzyme
than for a cell-bound one [9]. Depolymerisation is generally
carried out by extracellular hydrolases. While depoly-
merases, those enzymes acting on larger polymers, are
located at or outside the cell wall, oligomerases, which act
on depolymerase products may be located in the periplasmic
space, or even in the cytoplasm [25]. Kosugi et al. [26]
reported similar results for xylanase activity of Clostridium
cellulovorans, an anaerobic, mesophilic, cellulolytic bacter-
ium. Free amylase enzyme activity was also reported to be
greater than cell-bound enzyme activity during resistant-
starch hydrolysis under anaerobic conditions, by Sharp and
Macfarlane [27]. Vetter et al. [28] concluded that released
extracellular enzymes provide individual bacteria with a
powerful feeding mechanism, particularly in wastewater
conditions. Furthermore, the integrated action of the
excreted and the cell-bound enzymes makes it possible
for the cells to take up the soluble reaction products [9].
The high levels of amylase activity were understandable
since starch is the predominant polymer in potato and its
breakdown product was probably the preferred carbon
source. In this respect, the results of this work support the
finding of Ugwuanyi et al. [10] that the activity needed for
the degradation of the principal polymer in a waste stream
appears rapidly and at high levels during the digestion of the
waste. The high levels of amylase activity were probably
also due to the choice of inoculum. The wastewater
treatment plant from were the sludge was taken routinely
converts starch. Thus, there was probably a degree of
adaptation of the anaerobic consortium to degrade the
substrate. It is, therefore, important to choose adapted
inoculum to reduce the lag phase in hydrolysis. Low levels
of proteases were detected in this study and this is in
agreement with the observations Sarada and Joseph [24]
made during anaerobic digestion of tomato processing
waste. The inoculum used in our study contained a mixed
culture of microorganisms. The advantage of using the
mixed culture enabled a high rate of hydrolysis of the
substrate since it was degraded by symbiotic associations
(each microbial species having its own niche for growth and
substrate degradation). Comparison of the enzyme activities
reported here was not possible because, to the best of the
authors knowledge, no data for enzyme production during
anaerobic digestion of solid potato waste have yet been
published.
The rise and fall in the levels of the enzyme activities
observed may be due to various factors, such as microbial
enrichment, production and stability of enzymes, avail-
ability of substrate, pH and temperature of the liquor [24].
According to Ugwuanyi et al. [10] the fluctuation in the
amylase activity was probably a response to fluctuations in
the availability of reducing sugars. Variations in the
concentration of these sugars could lead to repression and
de-repression of amylase production in response to
microbial demand for carbon sources. It is possible that
different groups of microorganisms were supported in the
reactors at different times, and certain enzymes were stable
and active at different times. Potato waste consists of 86%
soluble cell contents (sugars, carbohydrate, protein, lipids
and pectin) and cell wall components composed of 11%
hemicellulose, 1% cellulose and 1% lignin [29]. The
expected major components of the plant cell wall are
cellulose, lignocellulose, hemicelluloses (e.g., xylan, ara-
ban, galatan and mannan) and pectin. Of the hemicelluloses
the dominant compound is xylan. Enzymatic depolymerisa-
tion of these structural polysaccharides of the plant cell wall
is an important process in the degradation of plant material
in nature. The increase in xylanase and pectinase activities
towards the end of the study period could possibly be related
to the degradation of cell wall material, as the potato solids
were observed to completely disintegrate during this time.
Cellulases and xylanases encompass a collection of enzymes
whose primary function is to hydrolyse b-1,4-glycosidic
linkages in the major plant structural polysaccharides,
cellulose and xylan [30]. The extracellular protease activity
was higher than that of the cell-bound, during the first 20
days of operation, after which the cell-bound activities
became higher than the extracellular activities until the end
of the hydrolysis period (Fig. 2f). The increase in protease
activity observed on days 2838 may have been due to
microflora that had developed and degraded proteinaceous
material, some of which was possibly cell debris.
These results suggest that the acidogenic microorganisms
produced several enzymes to degrade the organic material.
A study by Palmisano et al. [31] reported such a wide range
of hydrolytic enzymes in landfilled refuse, in which polymer
degradation occurs under anaerobic conditions The com-
bined activity of the enzymes resulted in complete
hydrolysis of the solid potato waste due to complementary
interactions. Ghose [18] and Bisaria and Ghose [32] reported
synergism between exo- and endo-glucanases since the
degradation of crystalline cellulose is a complex process
requiring the participation of many enzymes. A considerable
 W. Parawira et al. / Process Biochemistry 40 (2005) 29452952
degree of synergy was also reported to exist between
xylanases in the degradation of xylan, which is more
heterogeneous in structure than cellulose [32].
The anaerobic digestion of the same load of solid potato
waste was completed in 37 days in system II compared with
50 days in system I. A possible reason for the faster
degradation in system II may be the improved retention of a
high content of slow-growing methanogenic microorgan-
isms in the process. Andersson and Bjo rnsson [33] reported
better performance in terms of methane yield of the packed-
bed reactor with straw as carrier compared with other
carriers during digestion of crop residues. The advantage of
using straw is that it is a common agricultural by-product
and requires limited resources in terms of handling and cost.
The methane yields were similar in the two methanogenic
reactors. Our results are comparable to those reported in the
literature, although the differences in the waste and
characteristics of the substrate(s) are believed to be the
major reason for the difference in methane yield (see
Table 2). Andersson and Bjo rnsson [33] reported that the
degradation of the straw itself could contribute to biogas
production. However, in this study, pre-digested straw was
used to reduce the contribution of degradation products from
straw. A disadvantage when using straw as the carrier in a
packed-bed is the risk of clogging as the straw becomes
degraded.
When the hydrolysis/acidification reactors were coupled
to the methanogenic reactors, biogas production with high
methane content commenced immediately. The organic load
decreased with decreasing amount of organic matter in the
leachate in the acidogenic reactors as it was converted into
biogas. Although there were large fluctuations in the overall
organic loading rate of the methanogenic reactors, both
systems remained stable, and this is one of the advantages of
two-stage systems treating easily degradable substrates [34].
Effective biodegradation in the methanogenic reactors was
also evidenced by the high COD removal efficiencies
measured.
5. Conclusions
When performing two-stage anaerobic digestion it is
important to employ an efficient first hydrolytic stage since
waste solubilisation can be rate limiting in waste treatment.
The hydrolysis can be improved by inoculating the waste
with microorganisms producing hydrolytic enzymes specific
to the waste in question. The levels of enzyme activity can be
useful monitoring parameters during anaerobic digestion of
solid waste as they are easy and cheap to measure, and may
be waste-type specific. Both extracellular and cell-bound
enzymes produced by microorganisms led to hydrolysis of
solid potato waste. Among the enzyme activities, amylase
was especially high, indicating the importance of substrate
concentration in enzyme regulation. The two-stage system
with the packed-bed methanogenic reactor with straw as a
2951
biofilm carrier degraded the potato waste in a shorter time.
The performance in terms of methane yield was the same in
the straw packed-bed reactor and the UASB reactor. The
experiments demonstrated that separation of acidogenesis
and methanogenesis was achieved, as indicated by the
differences in pH and methane content in the acidogenic and
methanogenic reactors. The high methane yield obtained in
this study favours the application of the two-stage process to
digestion of solid potato waste.
Acknowledgement
Funding for this research was provided by the Swedish
Agency for Research Cooperation (SAREC) and the
Swedish National Energy Administration (STEM).
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