ARTICLE IN PRESS

Renewable Energy 33 (2008) 726734

www.elsevier.com/locate/renene

Semi-continuous co-digestion of solid slaughterhouse waste, manure,

and fruit and vegetable waste
Rene Alvareza,b, Gunnar Lidenb,
a
IIDEPROQ, UMSA, Plaza del Obelisco 1175, La Paz, Bolivia
b
Department of Chemical Engineering, Lund University, P.O. Box 124, 221 00 Lund, Sweden
Received 24 May 2006; accepted 2 May 2007
Available online 15 June 2007

Abstract

The potential of semi-continuous mesophilic anaerobic digestion (AD) for the treatment of solid slaughterhouse waste, fruit-vegetable
wastes, and manure in a co-digestion process has been experimentally evaluated. A study was made at laboratory scale using four 2 L
reactors working semi-continuously at 35 1C. The effect of the organic loading rate (OLR) was initially examined (using equal proportion
of the three components on a volatile solids, VS, basis). Anaerobic co-digestion with OLRs in the range 0.31.3 kg VS m3 d1 resulted in
methane yields of 0.3 m3 kg1 VS added, with a methane content in the biogas of 5456%. However, at a further increased loading, the
biogas production decreased and there was a reduction in the methane yield indicating organic overload or insufcient buffering capacity
in the digester.
In the second part of the investigation, co-digestion was studied in a mixture experiment using 10 different feed compositions. The
digestion of mixed substrates was in all cases better than that of the pure substrates, with the exception of the mixture of equal amounts
of (VS/VS) solid cattleswine slaughterhouse waste (SCSSW) with fruit and vegetable waste (FVW). For all other mixtures, the steady-
state biogas production for the mixture was in the range 1.11.6 L d1, with a methane content of 5057% after 60 days of operation. The
methane yields were in the range 0.270.35 m3 kg1 VS added and VS reductions of more than 50% and up to 67% were obtained.
r 2007 Elsevier Ltd. All rights reserved.

Keywords: Anaerobic; Biogas; Co-digestion; Manure; Slaughterhouse waste; Vegetable waste

1. Introduction common problem in many developing countries, which

Untreated slaughterhouses waste entering into a munici-
pal sewage purication system may create severe problems,
due to the very high biological oxygen demand (BOD) and
chemical oxygen demand (COD) [1]. Obviously, this
problem is aggravated if the untreated waste directly
reaches the recipient, such as for example a river. In the
city of La Paz, Bolivia, an estimated volume of 137 m3 d1
of industrial wastewater (from about 60 cattle and 60 swine
slaughtered) is released daily into the Choqueyapu river.
This represents a loading of 160 kg BOD and 4.4 tons of
solid waste primarily composed of animal ruminal and
visceral content [2]. This environmental problem is not
unique for the city of La Paz. On the contrary, it is a
Corresponding author. Tel.: +46 46 222 0862.
E-mail address: Gunnar.Liden@chemeng.lth.se (G. Liden).
urgently requires a cost-effective solution.
Anaerobic digestion (AD) represents a potential possi-
bility to decrease the environmental burden, and at the
same time provide biogas for local energy needs. In
additionin the specic case of treatment of animal
wastethe remaining stabilized slurry after digestion may
be used as a fertilizer [3]. AD of slaughterhouse waste and
animal by-products has recently been considered as an
interesting alternative waste management option [4,5]. In
Sweden and Denmark, utilization of rumen, stomach and
intestinal content, blood waste fraction, and sludge from
slaughterhouse wastewater treatment in biogas plants is
rather common [4,69].
The solid substrates from slaughterhouses mainly consist
of untreated blood and the contents of rumen, stomachs
and intestines, as well as manure from the delivery hall
[4,6]. The digestion of mixed waste consisting of the blood

0960-1481/$ - see front matter r 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.renene.2007.05.001
 ARTICLE IN PRESS
R. Alvarez, G. Liden / Renewable Energy 33 (2008) 726734 727

and gut ll components in the proportion in which they are feed was increased, a deterioration in the digester
produced has shown possible. However, the process is performance was seen [18].
sensitive and prone to failure [5,10]. Most likely, this can be The composition of slaughterhouse waste varies con-
attributed to the accumulation of high levels of free siderably. In some regions there are centralized and
ammonia resulting from the degradation of the nitrogen- specialized slaughterhouse facilities, processing only one
rich protein components of blood. Potential inhibition of or two animal species, whereas in other regions small
methanogenic bacteria by ammonia plays a role in almost slaughterhouses handle many animal species. The technical
all media with high nitrogen content wastes, since feasibility of anaerobic co-treatment thus has to be
ammonia is the end product of AD of proteins. The evaluated carefully with respect to the available substrates
inhibitory concentration of ammonia varies depending on [4,8]. The goal of the present study was to investigate a
parameters such as origin of inoculum, substrate, pH, and semi-continuous and mesophilic wet digestion system
temperature [1115]. The concentration of uncharged processing a mixture of manure, solid slaughterhouse,
ammonia has been suggested to be the active component and FVWs. The substrates were evaluated separately and
causing ammonia inhibition [11,14]. Starting at a value of in mixtures in proportions of 0100% (VS/VS) of each
approximately 700 mg ammonia-N L1, the maximum substrate. Furthermore, the effect of the OLR on methane
specic activity of methanogenic bacteria has been found yield and solid volatile reduction was investigated.
to decrease with increasing ammonia concentrations
[13]. In unadapted methanogenic bacterial sludge, inhibi- 2. Material and methods
tion has been reported at even lower concentration of
ammonia (150 mg N L1) [16]. The possibility of adapting 2.1. Raw material
methanogenic bacteria to ammonia has been demonstrated
[1115]. The maximum tolerable ammonia concentration The FVW were obtained from the vegetable market, La
was 6.2 times higher than the initial toxicity threshold Paz, Bolivia. Each item of waste was weighed separately
level [13]. before mixing and the composition is specied in Table 1.
Co-digestion of slaughterhouse wastes with different The material was initially mixed and minced into smaller
co-substrates has been proposed as a solution to the pieces with an electric mincer before being further
problems mentioned above. The content of nutrients can
thereby be balanced, and the negative effect of toxic Table 1
compounds on the digestion process may be decreased Composition of the fruit and vegetable waste used
giving an increased gas yield from the biomass. Murto [7]
Waste fraction Part used as feedstock Percentage
co-digested slaughterhouse waste, pig manure, vegetable
(w/w)a
waste, and various kind of industrial waste. A highly
buffered system was obtained and the process worked well Achojcha (caigua) Whole rotten fruit 1.9
with gas yields of 0.81 m3 kg1 VS. Edstrom [6] studied Banana Whole rotten fruit 9.2
mixtures of animal byproducts, slaughterhouse waste (i.e., Carrot Leaves, roots 1.7
Cassava Peels, roots, whole plant 4.7
rumen, stomach and intestinal content), food waste, and Cucumber Whole rotten fruit 6.4
liquid manure at 37 1C, at laboratory and pilot scale. Stable Eggplant Whole fruit 0.8
processes at organic loading rates (OLR) exceeding Grapefruit Whole rotten fruit 8.6
2.5 kg VS m3 d1 and hydraulic retention times (HRT) Green bean Whole rotten fruit 0.9
less than 40 d could be obtained with total ammonia Lemon Whole rotten fruit 6.2
Lettuce Leaves 1.3
nitrogen concentrations in the range of 4 to 5 g L1. Gas Lime Whole fruit 3.1
yields obtained were 0.70.86 m3 kg1 VS. Locoto (Chile pepper) Whole rotten fruit 7.5
A particularly important aspect for the digester perfor- Onion Exterior peels, leaves 5.9
mance is the C/N ratio [17] and the buffer capacity [7]. Orange Whole rotten fruit 14.0
Blood and swine manure, for example, both have high Pea Skins 0.7
Peach Peels 2.1
nitrogen content, and should preferably be co-digested Pear mellon Whole rotten fruit 1.9
with waste that has a low nitrogen content. The nitrogen Peas Pods, leaves 0.2
and phosphorus content in fruit and vegetable wastes Pineapple Peels of fresh riped fruit 2.4
(FVW) is often low and for this reason it has been used in Potato Peels 0.8
co-digestions with wastes with higher N and P content Pumpkin Peels, seeds 1.8
Radish Leaves, whole plant 2.3
[7,1821]. Continuous co-digestion of cattle slurry with Sugar beet Leaves, whole plant 0.5
FVWs and chicken manure was tested by Callaghan et al. Sweet pepper Whole rotten fruit 5.2
[18]. When the proportion of FVW was increased from Tangerine Whole fruit 2.5
20% to 50%, the methane yield also increased from 0.23 to Tomato Whole rotten fruit 4.8
0.45 m3 kg1 VS added at a loading rate between 3 and Turnip Leaves, whole plant 1.5
Watermelon Whole rotten fruit 1.3
5 kg VS m3 d1 and a hydraulic residence time of 21 days.
However, when the proportion of chicken manure in the a
Wet weight basis.
 ARTICLE IN PRESS
728 R. Alvarez, G. Liden / Renewable Energy 33 (2008) 726734

homogenized with a domestic electric blender (Hamilton
beach 908, Hamilton Beach Commercial, USA). Samples
were packed into polyethylene bags and stored at 10 1C in
a freezer until used.
A representative mixture of solid cattle and swine
slaughterhouse waste (SCSSW) were obtained from a local
municipal slaughterhouse (La Paz, Bolivia) in the propor-
tion in which they were produced. The minced and mixed
fractions of cattle rumen (57.1 wt%), stomach content and
gut ll of swine (9.4 wt%), blood cow (28.6 wt%), and
blood swine (4.9 wt%) was used as co-substrate in the tests.
Solid cattle and swine manure, SCSM (collected from
the same slaughterhouse) in the proportion cattle manure
71 wt% and swine manure 29 wt% were mixed, and the
samples were packed into polyethylene bags and stored in a
freezer until used. The characteristics of the cattle manure,
swine manure, cow rumen, swine paunch wastes, cattle
blood, swine blood, and the FVW, SCSM, SCSSW
mixtures are listed in Table 2. One thousand and eight
hundred grams of active slurry from a 10 L semi-
continuous mesophilic digester (35 1C) working with cattle
manure at 30 days of HRT, 1.8% w.w of TS content and
VS of 78 (% of TS) was used as initial inoculum in each
reactor.
a exible PVC tube to a separate water displacement glass
bottle lled with water acidied to pH 2. The positive
pressure in the bottles allowed the gas to be transferred to a
measuring gas cylinder. This was recorded at 24 h intervals.
2.3. Experimental procedure
During the semi-continuous experiments at laboratory
scale, seven OLR experiments and 10 co-digestion experi-
ments were performed at 35 1C in four digesters with 1.8 L
active volume and 2 L total volume, which were fed once a
day. The composition of the combination of mixtures used
as waste feed to reactors in the experiments are given in
Table 3.
2.3.1. OLR experiments
The effect of OLR on the digestion of mixtures of
manure, solid slaughterhouse waste, and fruitvegetable
wastes was tested in four mesophilic (35 1C) 2 L biogas
reactors (R1R4). Active slurry (1.8 L) from 10 L semi-
continuous mesophilic digester (35 1C), working with cow
manure at a HRT of 30 days with 1.8% w/w of TS content
and VS of 78 (% of TS), was used as initial inoculum in
each reactor (L1L4 experiments).

2.2. Experimental set-up

3

The digester experiments were carried out in four 4

8 2
identical, semi-continuously stirred stainless steel digesters,
each with a total volume of 2 L (see Fig. 1). The cylindrical
9
vessel was equipped with a anged top to which a ange 1
5
plate with stoppered ports was tted. This allowed gas
collection, and the mounting of a geared motor drive unit
6
for the reactor mixer. The contents of the reactor were
semi-continuously mixed as controlled by a timer, which
was activated for 15 min every hour. The digesters were 7
operated at 3571 1C by immersion in a water bath. Waste
Fig. 1. Digester set-up: (1) reactor, (2) transmission gear for stirrer, (3)
material was added and withdrawn through 12.7 mm ball timer, (4) biogas measurement system, (5) biogas reservoir ask, (6)
valves placed in the ank and the base of the reactor, thermostatic heater, (7) efuent valve, (8) inuent valve, and (9) water
respectively. Gas collection from the reactor was made via bath.

Table 2
Characterization of slaughterhouse wastes and fruit and vegetable waste

Analysis Cow Swine Cow Swine paunch Cow Swine Mixtures

manure manure rumen wastes blood blood
FVWa SCSMb SCSSWc

Total solids (% w.w) 19.8 30.8 14.9 31.7 19.8 22.3 12.7 20.5 20.0
Volatile solids (% of TS) 75.0 57.7 89.4 82.7 75.0 95.6 93.4 83.9 94.0
Volatile solids (% w.w) 14.9 17.6 13.3 26.2 14.9 21.3 11.9 17.2 18.8
Total nitrogen (% of TS) 1.6 2.5 2.2 1.9 15.0 8.3 1.1 2.2 5.5
Total organic carbon (% of TS) 22.3 ** ** ** ** ** 10.5 17.5 18.8
Total phosphorous (% of TS) 0.4 2.2 0.7 0.5 0.1 0.1 0.2 1.6 0.3
Total potasium (% of TS) 0.7 1.1 0.9 0.9 0.3 1.1 1.6 1.3 0.7
pH 7.1 9.2 6.1 5.9 7.4 7.2 4.9 8.2 7.1
a
FVW, fruit and vegetable wastes.
b
SCSM, solid cattle and swine manure.
c
SCSSW, solid cattle and swine slaughterhouse wastes, it includes rumen, stomach content, and blood from slaughtered animals.
 ARTICLE IN PRESS
R. Alvarez, G. Liden / Renewable Energy 33 (2008) 726734 729

Table 3
Average waste mixtures wet weight composition, total solid, volatile solid, organic carbon, nitrogen and phosphorous content used

Trial SCSSWa SCSMb FVWc Dilution Total Volatile Total Total organic Total Total
(% of WM) (% of WM) (% of WM) (% of WM) solids solids nitrogen carbon phosporous potassium
(% of WM) (% of TS) (% of TS) (% of TS) (% of TS) (% of TS)

L1 2 2 3 93 1.1 84.7 3.2 16.2 0.7 1.2

L2 8 7 11 74 4.5 87.5 3.1 15.7 0.7 1.2
L3 15 12 20 54 6.9 89.1 3.5 18.0 0.8 1.3
L4 8 7 11 74 4.3 88.8 3.2 16.5 0.7 1.2
L5 1 1 1 97 0.6 77.6 3.0 15.5 0.7 1.1
L6 8 7 11 74 4.4 89.7 3.2 16.2 0.7 1.2
L7 8 7 11 74 3.9 88.4 3.6 18.3 0.8 1.4
M1 0 20 0 80 4.3 83.8 2.1 16.9 1.5 1.3
M2 L2 8 7 11 74 4.5 87.5 3.1 15.7 0.7 1.2
M3 25 0 0 75 4.0 92.4 7.0 23.9 0.4 0.9
M4 11 12 0 78 4.3 89.1 3.9 18.8 1.0 1.1
M5 4 13 6 77 3.8 86.7 3.0 18.8 1.3 1.4
M6 17 3 6 74 3.8 91.7 5.5 21.8 0.6 1.2
M7 11 0 17 73 4.4 88.2 3.2 14.1 0.2 1.1
M8 4 4 23 70 4.2 91.7 2.1 13.9 0.5 1.5
M9 0 12 17 72 4.3 88.3 1.8 14.9 1.0 1.5
M10 0 0 34 66 4.0 93.2 1.2 11.1 0.2 1.7
a
SCSSW solid cattle and swine slaughterhouse wastes composed of 57.1% rumen, 33.5% blood, and 9.4% pigs paunch wastes.
b
SCSM solid cattle and swine manure, 71% cattle manure, and 29% swine manure.
c
FVW fruit and vegetable wastes.

Batches of feedstocks from SCSM, SCSSW, and FVW in
a proportion of 33.3% (VS/VS) were prepared for the
experiments L1L7 (cf. Table 3). The HRT was set at 30
days in experiments L1L3 (R1R3), 10 days in L4 (R4).
Subsequently, the HRT was changed to 50, 30, and 70 days
in experiments L6 (R1), L5 (R2), and L7 (R4), respectively.
The range of OLR studied in the experiments was
0.143.80 kg VS m3 d1. Thoroughly mixed substrate was
fed to the reactor once a day. The stirrer was activated
for 15 min every hour at 30 rpm. The atmospheric pressure
was 495 mmHg (the mean atmospheric pressure in La Paz,
Bolivia). Biogas was collected and measured by displace-
ment of water once a day at zero gauge pressure and
ambient temperature. The volumes were then recalculated
to normal temperature and pressure conditions (0 1C,
760 mmHg). The pH value and solid content of the slurry
was analyzed every 10 days.
2.3.2. Co-digestion experiments
The co-digestion of organic wastes depends on the
relative proportion of the components, the amount of the
mixture, and other physical process variables such as
temperature and pressure. In the current design, the
proportion of SCSSW, SCSM, and FVW were varied
while maintaining the %VS wet weight at 4%. In a mixture
experiment, the fractions of the different components
always sum to 1 (100% total of volatile solids). Therefore,
the factor space for a three-component mixture experiment
is a two-dimensional plane represented by an equilateral
triangle the vertices of which are the three pure compounds
A: SCSM x1 1, B: SCSSW x2 1, and C: FVW
x3 1. These points have the coordinates (1,0,0), (0,1,0),
and (0,0,1) in a 3D space. The points along the edges of the
triangle represent mixtures of only two components,
whereas points in the interior (x1, x2, x3) represent mixture
of all three components. A number of batches of feedstocks
(denoted M1 to M10) were prepared with the composition
as specied in Table 3.
The four 2 L reactors (denoted R1 to R4) had a working
volume of 1.8 L and operated at a temperature of 35 1C.
They operated for periods of 60 days, with a HRT of 30
days following the OLR experiments. The sequence of
experiments in each reactor was: R1: M5, M9; R2: M1,
M4, M7; R3: M3, M8, and R4: M6, M10.
Sampling procedures were identical to previously
described for OLR experiments.
2.4. Analytical methods
Methane and carbon dioxide concentration in the biogas
were measured with a gas chromatograph (Shimadzu
Model GC14B, Japan) equipped with a thermal conduc-
tivity detector (TCD) and Carboxen-1010 plot Capillary
column 0.53 mm ID (Supelco, USA), helium served as the
carrier gas. The injector, detector, and oven temperatures
were 150, 200, and 120 1C, respectively. VFA concentra-
tions were determined by gas chromatography. A Shimad-
zu GC14B Gas Chromatograph with a ame ionization
detector (FID) and a Nukol, 15 m  0.53 ID, 0.5 mm
(Supelco, USA) column was used. The carrier gas was
He. An initial oven temperature of 110 1C was maintained
for 1 min; then, it was increased to 158 1C at 8 1C min1,
and maintained at that temperature for 1 min. Injector
and detector temperatures were 250 1C. Samples were
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730 R. Alvarez, G. Liden / Renewable Energy 33 (2008) 726734

centrifuged (3000g) for 3 min and the resulting supernatant
ltrated through a Millipore lter (0.42 mm). This sample
was used for VFA measurements.
The total solids (TS), volatile solids (VS), pH, total
nitrogen and total phosphorous, in the feed and samples of
the substrates were all analyzed using standard methods as
described by Clesceri et al. [22]. The TS was determined
after a repeated heating (105 1C for 1 h), cooling, desiccat-
ing, and weighing procedure until the weight change was
less than 4%. VS were determined by ignition of the residue
produced in TS analysis to constant weight in a mufe
furnace at a temperature of 550 1C. Total Kjeldahl nitrogen
(TKN) was measured by semi-micro-Kjeldahl method as
described in Standard methods [22]. Potassium and
phosphorus were measured by spectrophotometry (method
3500-K and 4500-P, respectively). Total organic carbon
(TOC) was determined by high-temperature combustion
method (method 5310 B).
3. Results
3.1. Effect of loading rate
The methane yield increased from 0.14 to 0.34 m3 kg1 VS
added, when the OLR increased from 0.14 to 0.49 kg
VS m3 d1 and at the same time the VS reduction
increased from 9% to 60% (cf. Table 4 and Fig 2). There
was a stable performance of the AD in experiments with
OLR from 0.49 to 1.31 giving almost the same methane
yields (0.310.34 m3 kg1 VS added) and methane content
into the biogas (5456%). The pH in the efuent was
stabilized at values between 7.4 and 8.1. At higher loading,
the methane yield decreased (from 0.34 to 0.12 m3 kg1 VS
added at 3.8 kg VS m3 d1) and also the VS reduction
decreased (cf. Fig 2). The drop in pH in the efuent to 6.7
and the decreased methane content (down to 44%)
indicated hydraulic overload (wash-out of the microorgan-
isms), organic overload, or insufcient buffering capacity
in the digester giving a reduction in the methanogenic
activity.
3.2. Co-digestion of manure, slaughterhouse, fruit and
vegetable wastes
Based on the experiments above, an OLR of
1.11.3 kg VS m3 d1 was chosen for the mixture co-
digestion study. A stable steady state with a biogas
productivity well above 500 ml day1 was obtained in
seven of the 10 experiments (M1, M2, M4, M5, M6, M8,
and M9), as shown by steady proles of biogas productiv-
ity, biogas composition, and reactor pH (Fig. 3) as well as
by negligible levels of VFA (o0.2 g L1 data not shown).
Co-digested mixtures between all three components gave a
better process performance in terms of methane yield and
productivity compared to digestion of only one, or a
mixture of two substrates (cf. Table 5). The highest
methane yield was 0.35 m3 kg1 VS added obtained for
experiment M8 (i.e. 17% SCSSW, 17% SCSM, and 67%
FVW). The biogas productivity and the reduction of VS
was also the highest in this experiment.
Mixtures of two substrates gave a relatively good
performance, with steady-state biogas productivities values
and methane yields similar to the case with three
substrates, with the exception of the mixture 50%
SCSSW50% FVW (M7).
In three experiments (M3only SCSSW; M7equal
proportions of FVW and SCSSW, M10only FVW) it
was not possible to reach a steady state with a substantial
methane production. The biogas productivity gradually
declined as did the methane concentration (Fig. 3, right).
In experiment M10 (only FWV used as substrate), there
was a drop in biogas production during the rst 10 days
of operation, coinciding with a drop in methane concen-
tration and a drop in pH (Fig. 3). The digester reached the
steady-state operation at a pH value of 4.4 and with a
biogas production of 0.3 L d1 with almost negligible
methane content (2%). There was a sharp increase of
total VFAs from day 1 to day 10 from 2.5 to 8.1 g L1,
followed by a slow increase until day 25, whereafter
no further increase was observed (Fig 4a). The principal
VFA was acetic acid, which accounted for 5070% of the
total VFA.

Table 4
OLR, HRT, VS content, and steady-state values of the measured parameters

Experiment OLR HRT VSa pH Reduction of Methane Biogas productionb Methane yieldb
name (kg VS m3 d1) (days) (% w.w.) VS (%) contentb (%) (mL day1) (m3 kg1 VS added)

L5 0.14 30 0.4 7.4 9.3 5972.1 60712 0.1470.02

L1 0.31 30 0.9 7.4 8.7 5271.7 264731 0.2570.03
L7 0.49 70 3.4 7.7 60.4 5570.1 545725 0.3470.01
L6 0.78 50 3.9 8.1 67.6 5471.6 817721 0.3170.01
L2 1.31 30 3.9 7.4 52.4 5672.4 1359720 0.3270.01
L3 2.03 30 6.1 7.7 51.8 5570.1 1736763 0.2670.01
L4 3.80 10 3.8 6.7 47.1 4473.0 1810769 0.1270.01
a
VS % wet weight in the feed.
b
95% condence limits from ve consecutive measurements.
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0.6 1.0

Methane yield (l CH4/g VS added)

Methane production (l/day)

0.8

0.4
0.6

0.4
0.2

0.2
CH4yield methane vol

0.0 0.0

10 70

Reduction of VS (%)
50

6
pH

30
4

10
2
pH %VS remov

0 -10
0.1 0.3 0.5 0.8 1.3 2.0 3.8
OLR

Fig. 2. Organic loading rate (kg VS m3 d1) effect in anaerobic co-digestion of manure, slaughterhouse waste, and fruit and vegetable waste.

2500 2500
Biogas production
Biogas production

2000 2000
(ml/d)
(ml/d)

1500 1500
1000 1000
500 500
0 0
0 10 20 30 40 50 60 0 10 20 30 40 50 60

10 10
9 9
8 8
7 7
pH
pH

6 6
5 5
4 4
3 3
0 10 20 30 40 50 60 0 10 20 30 40 50 60
Composition (%CH4)

Composition (%CH4)

90
80 80
70 60
60 40
50
40 20
30 0
0 10 20 30 40 50 60 0 10 20 30 40 50 60
Time (d) Time (d)

Fig. 3. Daily biogas production (mL d1), pH, and biogas composition in experiments M1 (m), M2 (x), M4 (n), M5 (), M8 (+), and M9 (), (left); M3
(~), M7 (&), and M10 (*) (right).
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732 R. Alvarez, G. Liden / Renewable Energy 33 (2008) 726734

Table 5
Contribution percentage to VS and steady-state values for the measured and calculated parameters from semi-continuous experiments

Experiment* Feed Reactor Biogas

SCSSW SCSM FVW pH Reduction of Methane Biogas production Methane yield

(%) (%) (%) VS (%) content (%) (ml day1) m3 kg1 VS added

M3 100 0 0 7.7 34.5 4573 297722 0.0670.01

M1 0 100 0 7.4 38.8 5672 804712 0.2170.01
M10 0 0 100 4.4 19.2 270.2 316737 0.00274E-4
M4 50 50 0 7.7 51.7 5772 1063750 0.2670.02
M9 0 50 50 7.4 56.6 5073 1467735 0.3270.02
M7 50 0 50 5.3 53.8 2574 401773 0.0470.01
M2 33 33 33 7.4 52.4 5672 1359720 0.3270.01
M6 67 17 17 7.7 67.3 5171 1112740 0.2770.01
M5 17 67 17 7.8 54.2 5372 12117105 0.3270.03
M8 17 17 67 7.7 67.4 5172 1602732 0.3570.02

*HRT 30 days, T 35 1C.

concentration decreased (from 60% to 45%), indicating

12
ammonia inhibition related to the degradation of the
10 protein rich feed. The VFA concentration in the reactor
8 increased from 1.1 to 4.7 g L1 with 65% of acetate, 19%
VFA (g/l)

6 propionate, and 16% of butyrate at day 60.

4
2 4. Discussion
0
0 10 20 30 40 The loading rate is obviously a critical process parameter
Time (d) in anaerobic treatment. In the case of a CSTR system, a
decreased hydraulic residence time may result in biomass
10 washout, which inevitably leads to process failure [23,24].
8 Overloading may, however, not necessarily be the result of
VFA (g/l)

6 the residence time, but may result from organic overload

4
2 capacity. In the present work (cf. Fig 2), there was a linear
0 relationship between methane yield and loading rate at low
0 10 20 30 40 50
Time (d)
Fig. 4. Time course over VFAs (Acetic acid x, Propionic acid &, Butyric
acid W, Total VFA K) from the co-digestion of (a) experiment M7, i.e.
mixture of FVW and SCSSW and (b) experiment M10, i.e. FVW alone.
In experiment M7 (equal proportion of FVW and
SCSSW) it took about 30 days until the methanogenic
inhibition was evident by decreases in pH, biogas produc-
tion, and methane concentration (Fig 3). The total VFA
concentration remained at about 1 g L1 for 20 days.
Subsequently, there was an almost linear increase up to a
nal value of 8.3 g L1 at day 60 (Fig. 4b). Also in this case,
acetic acid was the predominant acid accounting for more
than 60% of the VFA.
A different kind of malperformance was seen in
experiment M3 (feed consisting of SCSSW only). In this
case, the pH value increased from 7.5 to 8, and at the same
time there was a gradual reduction in biogas production
from 1500 down to 300 mL d1. Also the methane
giving inhibition of the microorganisms due to accumula-
tion of VFAsin combination with a low buffering
60 loading rates. At intermediate loading rates, the methane
yield was almost constant up to a certain loading at which
it starts to decrease. This breakpoint indicates the
beginning of biological stress and beyond this point, the
methane production rate decreases sharply. Edstrom [6]
reported that it was difcult to operate a stable digestion
process at an OLR above 1 kg of VS m3 d1 with mixtures
of animal byproducts, stomach content, sludge, and food
waste. The slurry in the reactor reached high ammonia
nitrogen level (6.5 g L1), and to improve the digestion
process, the substrate mixture was diluted. After a gradual
increase in the OLR, the digester could then in fact be
operated at 3 kg VS m3 d1 with a concentration of
total ammonia nitrogen at 4.5 g L1. The improvement
was attributed to adaptation of the bacterial consortia to
the ammonia-rich environment. Murto [7] working in
co-digestion of sewage sludge and potato processing
industrial waste observed that the maximum OLR
(3.94.2 kg VS m3 d1) before organic overload decreased
as the proportion of carbohydrates-rich sludge increased
in the substrate. The process imbalance was due to the
 ARTICLE IN PRESS
R. Alvarez, G. Liden / Renewable Energy 33 (2008) 726734
low-buffered system by the composition of the incoming
substrate, the accumulation of VFAs resulted in decreasing
pH and nally led to digester failure.
The limit in OLR of 1.3 kg VS m3 d1 obtained in the
present work for mixtures of equal proportion in volatile
solids from SCSM, SCSSW, and FVW is lower than the
values obtained by Murto, indicating that these mixtures
are somewhat more inhibitory at mesophilic conditions.
However, it is to be kept in mind that the values in the
present study were obtained without any pretreatment of
the material.
Methane yields are to be expected to vary depending on
feedstock. The methane yields (0.270.35 m3 kg1 VS
added) and the VS reduction (5267%) obtained here are
in the range reported for co-digestion of similar waste
materials [57]. However, the methane content in the
biogas (5157%) was somewhat lower than what has
previously been reported [6,7,25,26]. A co-digestion process
using two of the substratesi.e. SCSM and FVW, and
SCSM and SCSSWworks well, whereas a mixture of
FVW and SCSSW is inhibited. Phrased differently, the
manurewhich is characterized by a low TS concentra-
tion, a high fraction of bers, many nutrients and a high
buffering capacityacts as an excellent carrier co-
substrate in the AD of concentrated slaughterhouse wastes
or FVW. The methane yields from FVW co-digested with
manure previously reported vary considerably. Callaghan
et al. [18] have reported methane yield between 0.23 and
0.45 m3 kg1 VS added and a 3050% VS reduction using a
HRT of 21 days, with an OLR in the range 35 kg
VS m3 d1. Kaparaju and Rintala [20] working with
potato tuber and its industrial by-products in co-digestion
with pig manure at loading rate of 23 kg VS m3 d1 at
different feed ratios of pig manure to potato waste reported
methane yields between 0.21 and 0.33 m3 kg1 VS added.
The methane yield in the co-digestion of FVW with SCSM
(cattle and pig manure mixture) in the present study
(0.3 m3 kg1 VS added) is comparable with these earlier
results.
The methane yield (0.26 m3 kg1 VS added) and VS
reduction (51.7%) obtained for the other binary substrate
mixture, i.e. SCSSW and SCSM (M4), was comparable to
reported data which combined treatment of manure
together with organic matter from slaughterhouses and
food-processing industries. Miladenovska et al. [27]
reported that lab scale CSTR reactors at 37 1C co-digesting
manure and lipids exhibited a signicantly higher specic
methane yield (0.38 m3 CH4 kg1 VS) and removal of VS
(51%) than the reactor treating manure (0.22 m3 CH4
kg1 VS, 37% VS reduction).
The inhibition shown in experiments M7, M3, and M10,
were characterized by a gradual reduction in biogas
production and methane composition, increased levels of
total VFA and reduction (M7 and M10) or increase (M3)
of pH, is likely an effect of methanogenic inhibition. The
sharp pH reduction in the digestion of only FWV (M10)
followed by accumulation of VFA and a concomitant
733
decreased methane production agree with inhibition of the
methanogenic bacterial species which are more inhibited
than the acidogenic bacteria [2022,28,29]. The resistance
to a pH change in the digester liquid depends on the
buffering capacity and the additional buffering as a result
of the addition of new ions; they also contribute to the
alkalinity. In experiment M7 (i.e. the co-digestion of
mixture of 50% SCSSW and FVW), inhibition occurred
after 30 days. The proteins present in SCSSW were
degraded forming ammonium bicarbonate, which results
in additional buffering of the digester liquid [30].
In contrast, in the digestion of only SCSSW (M3), there
was an increase in pH (from 7.5 to 8), and at the same time
a reduction in biogas production by a factor of ve. Also,
the methane concentration in the biogas decreased. In this
case, ammonia is likely the cause of methanogenic
inhibition. The nitrogen content of SCSSW is as high as
7% of TS (Table 3). The reactor was initially inoculated
from a low ammonia concentration. The nitrogen con-
centration, therefore, slowly increased at the on-set of
operation (from 100 to 190 mg N L1) and after about 30
days of operation there was a strong decrease in biogas
productivity and a notable decrease in methane content in
the biogas, in agreement with results by Banks and Wang
[5,10].
5. Conclusion
This study shows that a combined treatment of different
Bolivian waste types like manure (cattle and swine), solid
slaughterhouse wastes (rumen, paunch content, and blood
from cattle and swine), and FVW in a mesophilic co-
digestion process gives the possibility of treating waste,
which cannot be successfully treated separately. A semi-
continuous co-digestion process using these substrates
can be expected to result in a reduction of the volatile
solid contents of between 50% and 65% and give a
methane yield of about 0.3 m3 kg1VS added at OLRs up
to 1.3 kg VS m3 d1.
Acknowledgements
This work was nancially supported by SIDA, the
Swedish International Development Cooperation Agency.
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