enhancement of anaerobic biodegradability of flower stem wastes with vegetable wastes by...
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Journal of Environmental Sciences 20(2008) 297–303
Enhancement of anaerobic biodegradability of flower stem wastes withvegetable wastes by co-hydrolysis
ZHANG Bo, HE Pinjing∗, LU Fan, SHAO Liming
State Key Laboratory of Pollution Control and Resources Reuse, Key Laboratory of Yangtze River Water Environment, College of EnvironmentalScience and Engineering, Tongji University, Shanghai 200092, China. [email protected]
Received 21 May 2007; revised 10 September 2007; accepted 20 October 2007
AbstractThe vegetable wastes and flower stems were co-digested to evaluate the anaerobic hydrolysis performance of difficultly biodegradable
organic wastes by introducing readily biodegradable organic wastes. The experiments were carried out in batches. When the vegetable
wastes were mixed with the flower stems at the dry weight ratio of 1 to 13, the overall hydrolysis rate increased by 8%, 12%, and 2%
according to the carbon, nitrogen, and total solid (TS) conversion rate, respectively. While the dry weight ratio was designed as 1 to
3, there was a respective rise of 5%, 15%, and 4% in the conversion rate of carbon, nitrogen, and TS. The enhancement of anaerobic
hydrolysis from the mixed vegetable wastes and flower stems can be attributed to the formation of volatile fatty acids (VFA) and nutrient
supplement like nitrogen content. The maximum VFA concentration can achieve 1.7 g/L owing to the rapid acidification of vegetable
wastes, loosing the structure of lignocellulose materials. The statistic bivariate analysis revealed that the hydrolysis performance was
significantly related to the physical and biochemical compositions of the feeding substrate. Especially, the soluble carbon concentration
in the liquid was significantly positively correlated to the concentration of nitrogen and hemicellulose, and negatively correlated to the
concentration of carbon and lignocellulose in the feeding substrate, suggesting that the regulation and control of feedstock can have an
important influence on the anaerobic hydrolysis of organic wastes.
Key words: municipal solid wastes; hydrolysis; feedstock characteristics; VFA; bivariate analysis
Introduction
Municipal solid waste (MSW) management is nowadays
a critical concern in China. However, current practices
still count on the landfill of untreated, unsorted waste
as the main disposal route. High biodegradable organic
matter in the MSW leads to severe issues of public health
and environmental pollution, especially the emission of
leachate and landfill gas (He et al., 2003). Sustainable
waste management that favors waste recycling and re-
covery is considered to have the highest profits for the
environment. Anaerobic digestion of organic solid wastes
offers the advantage of both energy and fertilizer benefit
(Hartmann and Ahring, 2005). Therefore, anaerobic diges-
tion is envisaged to be an important addition to the list of
options available for managing MSW.
The anaerobic digestion of vegetable and food residues
has obtained very promising results owing to their high
biodegradability (Xu et al., 2002; Bouallagui et al.,2004; Zhang et al., 2005). However, for the difficultly
biodegradable organic wastes containing high content of
lignocellulose and cellulose, for example, plant stems, the
anaerobic digestion process will be limited and the biogas
yield will be consequently reduced owing to their complex
* Corresponding author. E-mail: [email protected].
structure (Converti et al., 1997). Generally, acid and alkali
conditions involving high temperature and high pressure
are applied to pretreat this kind of organic wastes (Vlysside
and Karlis, 2004), yet their decomposition products are
considered to be non-biodegradable and are not favorable
to the subsequent anaerobic digestion process; moreover,
chemical pretreatment is very expensive (Converti et al.,1997). Thereby, it is necessary to look for a milder and
cost-effective way to improve the hydrolysis of the organic
wastes containing lignocellulose and cellulose.
The formation of organic acids and ethanol from glucose
will play an important role in enhancing the hydrolysis
of lignocellulose (Yu et al., 2004). It was also proved
that mild acids can loosen the structure of lignocellulose
resulting in an improved overall rate of hydrolysis owing
to the increased accessibility of enzymes (Cassini et al.,2005). Hereinto, it can be deduced that the formation of
VFA from readily biodegradable organic wastes can help
the degradation of difficultly biodegradable organic wastes
containing lignocellulose. At present, the performance of
the anaerobic digestion process was improved by regulat-
ing the type of organic wastes fed considering the pH buffer
and nutrient adjustment (Callaghan et al., 2002; Kaparaju
et al., 2005). Nevertheless, there is still scarce information
about the contribution of readily biodegradable organic
298 ZHANG Bo et al. Vol. 20
wastes to the decomposition of difficultly biodegradable
organic wastes.
In this study, the co-hydrolysis of vegetable wastes and
flower stems was studied to evaluate the interaction among
the different types of biochemical components of the di-
gested wastes. The direct association between the physical
and biochemical compositions of organic wastes fed and
the hydrolysis performance were statistically established.
1 Materials and methods
1.1 Compositions and characteristics of feeding veg-etable wastes and flower stems
Vegetable wastes, the readily biodegradable wastes, and
flower stems, the difficultly biodegradable wastes were
sampled from a vegetable and flower market, respectively.
The vegetable wastes included cabbage and celery, while
the flower wastes consisted of rose and lily. There were
four parallel operated hydrolysis reactors (Run-1, Run-
2, Run-3, and Run-4). At Run-1, the reactor was filled
with vegetable wastes, and at Run-2, it was loaded with
flower stems. At Run-3 and Run-4, the vegetable wastes
and flower stems were mixed as the dry weight ratio of
1:13 and 1:3, respectively. The physical and biochemical
compositions of the mixed wastes are indicated in Table 1.
The wastes were smashed in a blender to an average size
below 3 cm.
1.2 Experimental setup and operation
The scheme of anaerobic hydrolysis of the mixed wastes
is shown in Fig.1. Four parallel-operated hydrolysis re-
actors had the same dimensions with working volume of
1.4 L and were operated under the same conditions. The
process control was carried out in batches. The vegetable
and flower wastes were filled into the hydrolysis reactor
after mixing and the methanogenic effluent of 0.5 L was
sequentially pumped into it. After 2 d of recirculation, the
outlet from the hydrolysis reactor was discharged into the
storage container, and then the fresh methanogenic effluent
was fed into the hydrolysis reactor from the top over again
and the above-mentioned operation was repeated. The
methanogenic effluent was taken from an upflow anaerobic
filter. The total organic carbon (TOC) concentration in the
methanogenic effluent was lower than 200 mg/L and the
pH was 7.8–7.9. The experiment was carried out in the
temperature range of 33–37°C.
Table 1 Physical and biochemical compositions of the sampled organic
wastes
Reactor number Run-1 Run-2 Run-3 Run-4
TS (%) 6.7 89.4 48.1 23.2
VS (%) 83.5 76.4 76.9 78.0
C (%) 61.7 70.3 69.6 68.2
N (%) 5.0 2.6 2.8 3.2
Cellulose (%) 5.2 26.7 13.6 12.1
Lignocellulose (%) 10.9 14.5 14.2 13.7
Hemicellulose (%) 25.0 15.8 16.4 17.9
Total sugar (%) 10.4 10.7 10.6 10.6
TS: total solid; WS: waste solid.
Fig. 1 Scheme of anaerobic hydrolysis of vegetable wastes and flower
stems.
1.3 Analytical methods
1.3.1 Chemical analysispH, TS, volatile solid (VS), and Kjeldahl nitrogen (KN)
were analyzed according to standard methods (APHA,
1998). The total carbon (TC) and total nitrogen (TN) were
measured by a TC/TN analyzer (Multi N/C 3000, Ana-
lytik Jena, Germany). The elements of carbon, hydrogen,
nitrogen, and sulfur of solid materials were measured by
an elemental analyzer (CHNS-900, LECO, USA). The
culture media were centrifuged at 4000×g for 10 min.
After filtered by 0.45 µm polyester films, the supernatant
was measured for VFA (LC-20AD, Shimadzu, Japan).
The measurement of cellulose, hemicellulose, and lig-
nocellulose was based on the analysis of neutral detergent
fiber (NDF), acid detergent fiber (ADF), and ash contents
of the materials (Goering and van Soest, 1970). For the
determination of total sugar, the dry solid sample was first
pretreated by boiled HCl solution (6 mol/L), and then the 3,
5-dinitrosalycylic acid method was applied (Miller, 1959;
Nielsen, 2002).
1.3.2 Extracellular enzyme activity assayA 10-ml sample of the hydrolysis liquid was first
centrifuged at 3000×g for 10 min, and then the super-
natant was collected and used for the measurement of
carboxymethyl cellulose enzyme (CMCase) activity. The
samples were incubated with carboxymethyl cellulose in
sodium acetate buffer (pH 4.8, 0.2 mol/L), and pre-warmed
to the incubation temperature of 60°C (Nakamura and
Kitamura, 1988).
1.4 Statistical analysis
Statistical analyses utilized SPSS 14.0 (SPSS, Inc.,
Chicago, USA). The dependence of hydrolysis perfor-
mance on a single physical and chemical factor was
analyzed by bivariate regression analysis.
2 Results
2.1 Soluble TC and TN in the liquid phase
Soluble TC and TN are the parameters that represent
the hydrolysis extent. The hydrolysis of organic matter at
four feeding compositions in terms of accumulated TC is
No. 3 Enhancement of anaerobic biodegradability of flower stem wastes with vegetable wastes by co-hydrolysis 299
shown in Fig.2a. The accumulated TC shows the highest
solubilization when vegetable wastes were fed at Run-1,
followed by the mixed vegetable wastes and flower stems
at Run-4 and Run-3, and the flower stems fed at Run-2 in
order. The maximum solubilization of carbon expressed as
TC can be 723, 296, 380, and 450 mg/g at Run-1, Run-2,
Run-3, and Run-4, respectively. The overall solubilization
of carbon at Run-3 and Run-4 was 13% more than the
combination of single wastes fed if taking no account of
the synergism among different types of wastes.
It can be seen from Fig.2b that adding the vegetable
wastes into the flower stems increased the soluble TN
concentration in the hydrolysis liquid. The TN solubi-
lization of about 691, 463, 518, and 639 mg/g at Run-1,
Run-2, Run-3, and Run-4 was obtained at the end of the
designed experimental period of 16 d, respectively. The
overall hydrolysis of nitrogen at Run-3 and Run-4 was
5% and 14% more according to the calculation from the
conversion rate of vegetable wastes and flower stems fed
at Run-1 and Run-2.
2.2 pH in the liquid phase
From Fig.3, it is seen that the addition of the vegetable
wastes caused a rapid drop of pH as compared to the
scenario of flower stems fed. The pH in the hydrolysis
reactors including the vegetable wastes decreased to the
minimum value (5.5–6) on day 6 of the experiment, while
the pH remained above 7 as for the flower stems fed at
Run-2.
2.3 VFA and reducing sugar
The VFA and reducing sugar in the hydrolysis products
were detected, whereas lactic acid could not be observed.
Fig.4 presents the variations of VFA and accumulated
VFA concentration in the hydrolysis liquid. The VFA
concentration had a peak value on days 6 and 8. The
VFA and accumulated VFA concentration at Run-1 were
3 times more than that at Run-2. The accumulated VFA
concentration was 189, 57, 71, and 62 mg/g, accounting
for about 27%, 19%, 19%, and 14% of TC in the hydrolysis
liquid for Run-1, Run-2, Run-3, and Run-4 at the end of the
experimental period, respectively.
As indicated in Fig.5, the reducing sugar concentration
at Run-1 was considerably lower than that at other reactors.
The variations of accumulated reducing sugar concen-
tration at Run-3 and Run-4 were similar. The maximal
concentration of 15 mg/g was obtained at Run-2, followed
by Run-3 and Run-4, and Run-1. The accumulated reduc-
ing sugar concentration accounted for about 0.4%, 5%,
3%, and 2% of TC in the hydrolysis liquid, respectively.
2.4 Conversion of TS, carbon, and nitrogen
It can be seen from Fig.6 that the conversion rates of
TS, carbon, and nitrogen at Run-1 were achieved 84%,
85%, and 94%, followed by Run-4, Run-3, and Run-2 in
sequence. The conversion rates of TS, carbon, and nitrogen
for the flower stems increased, when the vegetable wastes
were brought in. The total conversion rates of TS, carbon,
and nitrogen had a respective rise of about 2%, 8%, and
12% for Run-3, and about 4%, 5%, and 15% for Run-4
according to the calculation from the conversion rate of
vegetable wastes and flower stems at Run-1 and Run-2.
Figure 7 shows the degradation of hemicellulose,
cellulose, lignocellulose, and total sugar based on the
calculation from the mass balance. The conversion rate of
cellulose obviously increased after the vegetable wastes
were added into the flower stems. However, the hemicel-
Fig. 3 pH in the hydrolysis liquid.
Fig. 2 Accumulated TC (a) and TN (b) concentration in the hydrolysis liquid. Run-1: vegetable wastes; Run-2: flower stems; Run-3: vegetable
wastes:flower stems=1:13; Run-4: vegetable wastes:flower stems=1:3.
300 ZHANG Bo et al. Vol. 20
Fig. 4 VFA (a) and accumulated VFA (b) concentration in the hydrolysis liquid.
Fig. 5 Reducing sugar (a) and accumulated reducing sugar (b) concentration in the hydrolysis liquid.
lulose content in the residual wastes was higher than in the
feeding wastes, as a result of lignocellulose degradation.
2.5 Variations of extracellular enzyme activity
The CMCase activity at Run-2 and Run-3 was higher
than that at Run-4 and Run-1 (Fig.8). The CMCase activity
in the four reactors differed with different substrates fed. It
is tightly related to the substrate compositions (Parawira etal., 2005). Even if the hydrolysis rate of vegetable wastes
was higher than flower stems, the CMCase activity in the
hydrolysis reactor containing vegetable wastes showed the
minimum value.
2.6 Association of hydrolysis with physical and bio-chemical compositions
Table 2 shows that bivariate correlation analysis be-
tween the hydrolysis performance and the physical and
biochemical compositions of feeding wastes. The soluble
TC, TN, VFA, and the reducing sugar concentration in
the hydrolysis liquid were related to the carbon, nitrogen,
TS content, hemicellulose, cellulose, and lignocellulose
as well in the feeding substrate. The TC concentration
was significantly positively correlated to nitrogen and
hemicellulose, and negatively correlated to carbon and
lignocellulose. The TN concentration was negatively cor-
related to the TS content. The VFA concentration was
positively correlated to the nitrogen content, and nega-
tively correlated to lignocellulose and hemicellulose. The
reducing sugar concentration was positively correlated to
lignocellulose and negatively correlated to hemicellulose.
No. 3 Enhancement of anaerobic biodegradability of flower stem wastes with vegetable wastes by co-hydrolysis 301
Fig. 6 Conversion rate of TS, C, and N.
3 Discussion
The anaerobic hydrolysis of the mixed wastes realized
mass and volume reduction, but the hydrolysis yield was
affected by the constituents of wastes feedstock (Chanakya
et al., 1999; Nguyen et al., 2007). The hydrolysis of
vegetable wastes was two times higher than flower stems
according to the solubilization of carbon. Adding the
vegetable wastes improved the hydrolysis of the flower
stems. For the hydrolysis of the difficultly biodegradable
flower stems, about 30% of carbon contained in the wastes
was converted in the leachate form; however, when the
vegetable wastes were mixed with these, more than 50%
Fig. 7 Conversion rate of hemicellulose, cellulose, lignocellulose, and
total sugar.
of carbon was converted. The introduction of vegetable
wastes into the flower stems obviously enhanced the
overall solubilization of carbon and nitrogen, and the
degradation of TS, which can be attributed to the weak
acid environment caused by the rapid acidogenesis of the
vegetable wastes and the supplement of nutrients (Cassini
et al., 2005).
The cellulose and hemicellulose are difficultly
biodegradable mainly owing to their intimate combination
with lignin (Ren et al., 1996). The initial reaction in
the process of organic acids pretreatment involves a
mild acid-catalyzed hydrolysis of the glycosidic bonds
of hemicellulose and the α-ether linkage in lignin, in
Table 2 Bivariate correlation analysis between the hydrolysis characteristics and chemical compositions of feeding wastes
Hydrolysis characteristics Carbon Carbon conver- Cellulose Hemicellulose Lignocellulose Nitrogen Reducing
(%VS) sion rate (%) (%VS) (%VS) (%VS) (%VS) sugar (mg/g)
Carbon (%VS) 1 –0.484 0.870 –0.916* 0.941 –0.911 0.865
Carbon conversion rate (%) –0.484 1 –0.316 0.789 –0.748 0.800 –0.801
Cellulose (%VS) 0.870 –0.316 1 –0.720 –0.756 –0.723 0.816
Hemicellulose (%VS) –0.916 0.789 –0.720 1 –0.998** 1.000** –0.951*
Lignocellulose (%VS) 0.941 –0.748 –0.756 –0.998** 1 0.996** 0948*
Nitrogen (%VS) –0.911 0.800 –0.723 0.816 0.996** 1 –0.958*
Reducing sugar (mg/g) 0.865 –0.801 0.816 –0.951* 0948* –0.958* 1
TC (mg/g) –0.907* 0.796 –0.788 0.986** –0.985** 0.990** –0.989**
TN (mg/g) –0.144 0.894* –0.093 0.490 –0.435 0.509 –0.587
Total sugar (%VS) –0.520 –0.493 –0.592 0.135 –0.200 0.120 –0.099
TS (%) 0.455 –0.865 0.531 0.677 0.647 –0.696 0.836
TS conversion rate (%) –0.579 0.993** –0.386 0.854 –0.818 –0.863 –0.848
VS (%TS) 0.165 0.777 0.316 0.244 –0.179 0.258 –0.256
VFA (mg/g) –0.761 0.883* –0.519 0.942* –0.927* 0.943* –0.870
Hydrolysis characteristics TC TN Total TS TS conversion VS VFA
(mg/g) (mg/g) sugar (%VS) (%) rate (%) (%TS) (mg/g)
Carbon (%VS) –0.907* –0.144 –0.520 0.455 –0.579 0.165 –0.761
Carbon conversion rate (%) 0.796 0.894* –0.493 –0.865 0.993** 0.777 0.883*
Cellulose (%VS) –0.788 –0.093 –0.592 0.531 –0.386 0.316 –0.519
Hemicellulose (%VS) 0.986** 0.490 0.135 0.677 0.854 0.244 0.942*
Lignocellulose (%VS) –0.985** –0.435 –0.200 0.647 –0.818 –0.179 –0.927*
Nitrogen (%VS) 0.990** 0.509 0.120 –0.696 –0.863 0.258 0.943*
Reducing sugar (mg/g) –0.989** –0.587 –0.099 0.836 –0.848 –0.256 –0.870
TC (mg/g) 1 0.536 –0.133 –0.763 0.854 0.239 0.912*
TN (mg/g) 0.536 1 –0.703 –0.883* 0.849 0.870 0.580
Total sugar (%VS) –0.133 –0.703 1 0.346 –0.394 –0.928* –0.124
TS (%) –0.763 –0.883* 0.346 1 –0.848 –0.586 –0.667
TS conversion rate (%) 0.854 0.849 –0.394 –0.848 1 0.706 0.920*
VS (%TS) 0.239 0.870 –0.928* –0.586 0.706 1 0.475
VFA (mg/g) 0.912* 0.580 –0.124 –0.667 –0.667 0.475 1
* corresponds to 0.01 confidence level; ** corresponds to 0.05 confidence level.
302 ZHANG Bo et al. Vol. 20
Fig. 8 CMCase activity in the hydrolysis liquid.
which the organic acids, formed by cleavage of the
labile ester groups, catalyze the hemicellulose hydrolysis.
The fraction is achieved by an enlargement of the inner
surface (Schmidt and Thomsen, 1998). The VFA can be
continuously produced owing to the addition of vegetable
wastes, and the maximal VFA concentration can reach
1.7 g/L. The pH in the hydrolysis liquid decreased from
7.8–7.9 to 5.6. VFA functioned as a conditioning to the
structure of lignocellulose and cellulose, making the
extracellulose enzyme easily accessible to the surface
of the material (Yu et al., 2004). Successful mixing of
different wastes can result in better digestion performance
by improving the content of the nutrients and can
even reduce the negative effect of toxic compounds on
the digestion process (Murto et al., 2004). From the
point of physical and biochemical compositions, some
compositions, such as moisture and nitrogen source
had an obvious influence on the hydrolysis of organic
wastes (Misi and Forster, 2001; Sponza and Agdag, 2004;
Sanphoti et al., 2006). It was found from the bivariate
analysis in this study that the increase of nitrogen and
hemicellulose content was favorable to the solubilization
of carbon expressed as TC. The increase of nitrogen could
meet the need of microbial nutrients for the hydrolysis
of organic wastes rich in carbon (Altaf et al., 2007;
Ohkouchi and Inoue, 2007). The single hemicellulose
matter was considered as easily biodegradable, while
the lignocellulose matter was difficultly biodegradable.
The addition of vegetable wastes increased the nitrogen
and hemicellulose content, resulting in the improved
hydrolysis rate.
4 Conclusions
The introduction of vegetable wastes into the flower
stems can increase the overall hydrolysis rate of the mixed
organic wastes, caused by the VFA formation and nutrient
supplement from the hydrolysis of vegetable wastes. The
soluble carbon concentration in the liquid was significantly
positively correlated to the concentration of nitrogen and
hemicellulose, and negatively correlated to the concentra-
tion of carbon and lignocellulose in the feeding substrate.
Acknowledgements
This work was supported by the National Key Technol-
ogy Research and Development Program of China (No.
2006BAC02A03), the Key Project of Chinese Ministry of
Education (No. 107122), the 2006 Shanghai-Rhone Alpes
Region (France) Scientific Research Cooperation Fund
(No. 06SR07105), and the China Postdoctoral Science
Foundation (No. 20060390653).
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