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Low temperature anaerobic digestion of mixtures of llama,cow and sheep manure for improved methane production

Rene Alvareza,b, Gunnar Lidenb,*aIIDEPROQ, UMSA, Plaza del Obelisco 1175, La Paz, BoliviabDepartment of Chemical Engineering, Lund University, P.O. Box 124, 221 00 Lund, Sweden

a r t i c l e i n f o

Article history:

Received 16 June 2006

Accepted 29 August 2008

Published online 7 November 2008

Keywords:

Anaerobic digestion

Biogas

Biomethanation

Manure

Low temperature

* Corresponding author. Tel.: þ46 46 222 086E-mail address: gunnar.liden@chemeng.l

0961-9534/$ – see front matter ª 2008 Elsevidoi:10.1016/j.biombioe.2008.08.012

a b s t r a c t

Biogas production in anaerobic digestion in farm-scale units is typically performed under

mesophilic conditions when used for producing domestic fuel and stabilizing animal waste

for the use of digested manure as a fertilizer. Previous studies on the digestion of llama and

cow manure have shown the feasibility of producing biogas under altiplano conditions

(low pressure and low temperature) and of llama manure as a promising feedstock. The

present study concerns the utilization of various mixtures of feedstocks from the Bolivian

altiplano under low temperature conditions (18–25 �C). Laboratory scale experiments were

performed on the digestion of mixtures of llama, sheep and cow manure in a semi-

continuous process using ten 2-L stainless steel digesters to determine the effects of

organic loading rate (OLR) and the feed composition. The semi-continuous operation of

mixture of llama–cow–sheep manure proved to be a reliable system, which could be

operated with good stability. The results suggest that in a system digesting a mixture of

llama-cow-sheep manure at low temperature (18–25 �C) the maximum OLR value is

between 4 and 6 kg VS m3 d�1. The methane yields obtained in the mixture experiments

were in the range 0.07–0.14 m3 kg�1 VS added, with a methane concentration in the gas of

between 47 and 55%.

ª 2008 Elsevier Ltd. All rights reserved.

1. Introduction of raw material, and both the methane yield and the possible

Anaerobic digestion of manure, alone or in a mixture of

manure and others organic wastes, is widely used today.

A number of full-scale anaerobic mesophilic and thermophilic

digesters for biogas production have been developed in

Denmark and Sweden and have been in operation for the last

20 years [1,2]. Furthermore, about 3 million small-scale biogas

plants have been built in India, and about 7 million in China

[3,4]. These plants normally operate under mesophilic

conditions.

Temperature and the type of raw material are two of the

most important parameters in anaerobic digestion. The

anaerobic digestion is, of course, strongly affected by the type

2; fax: þ46 46 149156.th.se (G. Liden).er Ltd. All rights reserved

reduction of the solid content depends on the composition of

the waste material. Co-digestion, i.e. the simultaneous

digestion of a mixture of two or more substrates, is a tech-

nique, by which the bioconversion rate as well as the methane

yield can be increased. The process benefits of co-digestion

lies in effects such as an improved nutrient balance,

decreased effect of toxic compounds on the digestion process,

or improved rheological qualities of the substrate [1,5,6].

Anaerobic digestion is, technically speaking, an uncompli-

cated biological process for the treatment of different organic

wastes that can be used also at low temperatures for the

production of biogas, and thereby avoid the uncontrolled

release of methane directly into the atmosphere. According to

.

Table 1 – Characteristics of the fresh undiluted manures.

Type of analysis Cow Llama Sheep

Total solids

(% wet)

17.6(3.2) 52.1(3.6) 61.4(22.9)

Volatile solids

(% TS)

76.1(1.6) 64.4(8.3) 54.9(8.9)

Total Kjeldahl

nitrogen (% TS)

1.5(0.2) 1.7(0.2) 0.9(0.4)

Total organic

carbon (% TS)

28.6(3.1) 25.1(5.1) 18.1(1.1)

Phosphorus (% TS) 0.3(0.1) 0.4(0.1) 0.6(0.1)

Potassium (% TS) 0.7(0.1) 1.1(0.4) 1.8(0.9)

Parameter mean and standard deviation within each animal

species are comprised of 4 data points. Manures were sampled in

summer and winter.

Table 2 – Fraction of the different manures (wt%) used inthe organic loading rate experiments.

Experiment Llamamanure (%)

Cowmanure (%)

Sheepmanure (%)

Water (%)

1 and 6 1.0 2.8 1.5 94.6

2 and7 6.3 16.9 9.3 67.6

3 and 8 8.3 22.5 12.4 56.8

4 and 9 6.3 16.9 9.3 67.6

5 and 10 8.3 22.5 12.4 56.8

b i o m a s s a n d b i o e n e r g y 3 3 ( 2 0 0 9 ) 5 2 7 – 5 3 3528

previous studies, reasonable methane yields from anaerobic

digestion can be expected at low temperatures (14–23 �C) if the

digester organic loading rate (OLR) is reduced appropriately by

extending the hydraulic retention time [7]. From a simple

Arrhenius type expression one may expect an exponential

decay of productivity with temperature. However, the

methane yield at a certain residence time has been deter-

mined to typically decrease linearly as temperature is

decreased over the range of 10–23 �C [8–10]. Biogas production

rate has been reported to vary from 0.09 m3 kg�1 VS in the

winter to 0.33 m3 kg�1 VS in the summer for treating swine

waste in a covered lagoon digester. The methane concentra-

tion in the biogas was 70–75% at a loading rate of

0.75 kg VS m�3 day�1 [11]. The biogas production from swine

wastes started at temperatures as low as between 3 and 9 �C,

and methane became the primary biogas component at

approximately 10 �C [8]. Unfortunately, biomethanation at low

temperatures has not been the much studied [12], although

recent efforts have made [13]. While the anaerobic digestion of

manure and other agriculture wastes appears to be an inter-

esting technological alternative for solving the energy prob-

lems of villages in developing countries, the technology and

feedstocks studied are not always applicable in extreme

environmental conditions, where it would be of particular

usefulness. The Bolivian altiplano has an average elevation of

nearly 4 km, and encompasses an area of over 600 000 km2. It

is swept by strong, cold winds, and has an arid, chilly climate,

with sharp differences in daily temperature. Average highs

during the day range from 15 �C to 20 �C and the average lows

from �15 �C to 3 �C with an atmospheric pressure around

460–500 mmHg. The most important group of domestic

animals are llamas, cows, and sheep. The population of

llamas on the altiplano is nearly 3 million, with nearly 70%

located in the Bolivian part. The current study investigates the

potential for using the manures from these animals as

feedstocks for anaerobic digestion on the altiplano. Llama,

cow and sheep manure were used as raw material in a semi-

continuous biomethanation process at low temperature

(18–25 �C). Important process parameters such as the OLR

effect and feed composition were studied and the methane

yield and volatile solids reduction were determined.

2. Materials and methods

2.1. Feedstock

Llama, sheep and dairy cattle manure were collected from

farms in the Bolivian altiplano situated at 19�S Latitude and

68� W longitude at an altitude of 3800 m above mean sea level.

Samples were collected in two seasons: winter (July) and

summer (February). Llama and sheep manure separately were

minced and pulverized with a semi-industrial cutter (CUT-3,

Metvisa, Brazil), and afterward they were packed into poly-

ethylene bags and stored at �10 �C in a freezer until used. The

characteristics of manure are listed in Table 1.

2.2. Apparatus

The experiments were carried out using ten stainless steel

digesters, each with a total volume of 2 L. The cylindrical

vessels were equipped with a flanged top to which a flange

plate with stoppered port was fitted; this allowed gas collec-

tion. Each digester was equipped with a port for feeding and

effluent drawn through 12.7 mm ball valve placed on the flank

of the reactor close the base. Two immersion thermostats in

each bath controlled the water bath temperature.

The biogas was collected in the collecting glass-bottles,

acting as gas reservoirs. The overpressure in the bottles

allowed the gas to be transferred to a measuring gas cylinder.

This was recorded at 24-h intervals. All reactors were mixed

by shaking the reactors by hand once a day for about 2 min

before to feed.

2.3. Experimental procedure

The study basically consisted of two parts. First, the effect of

organic loading rate was studied at 18 �C and 25 �C. Secondly,

a set of mixture experiments were made at the previously

determined most suitable OLR to find the optimal feedstock

composition. Both experiments were preceded by preparation

of respective batches of feedstocks.

2.3.1. Preparation of feedstocksBatches of feedstocks were prepared according to Tables 2

and 3. Each batch was diluted with tap water to obtain the

desired solid content was homogenized with a domestic

electric blender (Hamilton Beach 908, Hamilton Beach

Commercial, USA), fractionated (volume defined by the

hydraulic residence time, HRT), and packed into polyethylene

bags and stored in a freezer until used. The samples for each

Table 3 – Fraction of the different manures, given both aswt% and % of the total volatile solids used in mixturedesign experiments. The hydraulic residence time was50 days in the experiments.

Exp. Llamamanure

Cowmanure

Sheepmanure

Water

(%Mass)

(% VS) (%Mass)

(%VS) (%Mass)

(VS) (%Mass)

D1 17.2 100.0 0.0 0.0 0.0 0.0 82.8

D2 8.6 50.0 0.0 0.0 6.3 50.0 85.1

D3 5.7 33.3 13.4 33.3 4.2 33.3 76.6

D4 0.0 0.0 40.3 100.0 0.0 0.0 59.7

D5 0.0 0.0 0.0 0.0 12.6 100.0 87.4

D6 0.0 0.0 20.2 50.0 6.3 50.0 73.5

D7 8.6 50.0 20.2 50.0 0.0 0.0 71.2

D8 11.5 67.0 6.9 16.5 2.1 16.5 79.5

D9 2.9 16.5 6.9 16.5 8.5 67.0 81.8

D10 2.9 16.5 27.0 67.0 2.1 16.5 67.9

b i o m a s s a n d b i o e n e r g y 3 3 ( 2 0 0 9 ) 5 2 7 – 5 3 3 529

day were withdrawn from the freezer and allowed to thaw

overnight.

2.3.2. OLR experimentsDigestion of a mixture of llama–cow–sheep manure at five

different OLR (0.5, 3, 4, 6 and 8 kg VS m�3 d�1) was tested at

18 �C and 25 �C. 10 reactors with a working volume of 1.8 L

each were used in this study. The initial load of 1800 g in each

reactor was prepared with: 52 g llama manure, 121 g cow

manure, 38 g sheep manure, 689 g water, and 900 g of active

slurry. The inoculum for the reactors with TS content of 7.3%

and VS of 72 (% of TS) were taken from a bioreactor working

with cow manure at 25 �C and HRT of 50 days. After closing

each vessel with flange plates the reactors were flushed for

2 min with an anaerobic gas containing 98% CO2 to ensure

anaerobic conditions in the head space of the digesters. The

temperatures in reactors’ number 1–5 were controlled at 18 �C

and the temperature in reactors 6–10 was controlled at 25 �C.

In the start-up period of 30 days the reactors were first run as

batch processses for 10 days followed by a semi-continuous

process with a HRT of 50 days and an OLR of 1.2 kg VS m�3 d�1.

The daily feed to the reactors were a mixture of llama–cow–

sheep manure (33.3% each VS basis). On day 30, the daily feed

Table 4 – Composition of the used mixtures.

Exp. Composition

Llama manure(% w.w)

Cow manure(% w.w)

Sheep manure(% w.w)

D1 15.5 0.0 0.0

D2 7.7 0.0 5.7

D3 5.2 12.1 3.8

D4 0.0 36.3 0.0

D5 0.0 0.0 11.4

D6 0.0 18.2 5.7

D7 7.7 18.2 0.0

D8 10.4 6.2 1.9

D9 2.6 6.2 7.6

D10 2.6 24.3 1.9

according Table 2 was started. The reactors were maintained

at each OLR for a minimum period of three HRT. Biogas was

collected and measured by displacement of water once a day

at zero gauge pressure and ambient temperature. The

volumes were then recalculated to normal conditions (0 �C,

760 mmHg). The pH value and solid content of the slurry was

analyzed every 10 days.

2.3.3. Mixture experimentsTen anaerobic digestion experiments in semi-continuous

process with different proportions of llama–cow–sheep

manure were made in parallel at 25 �C. All experiments were

started as batch processes at 25 �C, which ran for 20 days,

before daily feeding was commenced. The initial content of all

ten reactors (1800 g) were prepared according to Table 4. The

inoculum for the reactors was taken from a bench scale

anaerobic digester (ST of 1.89%, VS of 68.6%) working with cow

manure at a HRT of 50 days and a temperature of 25 �C. On day

20, the daily feed according Table 3 was started with a HRT of

50 days and an OLR of 1.2 kg VS m3 d�1. The reactors were

maintained for a period of two HRT. A volume of slurry (36 ml)

was withdrawn from each reactor daily and was replaced with

the same volume of fresh feedstock via the slurry sampling

tube. The establishment of a steady state in the reactors was

assessed by measuring the daily gas production and the

methane content of the gas, as well as the solids content and

pH. Measurements taken for a period of five days were aver-

aged to obtain a final value for the productivity and methane

concentrations at steady state. Sampling procedures were

identical to previously described for the OLR experiments.

2.4. Analytical methods

Methane and carbon dioxide concentrations in the biogas

were determined with a gas chromatograph (Shimadzu Model

GC14B, Japan) equipped with a thermal conductivity detector

(TCD) and Carboxen-1010 plot Capillary column

30 m� 0.53 mm ID (Supelco, USA). The injector, detector and

oven temperatures were 150 �C, 200 �C and 120 �C, respec-

tively. Helium served as the carrier gas.

Total solids (TS), volatile solids (VS), pH, total organic

carbon (TOC), total Kjeldahl nitrogen (TKN), potassium and

Total Volatile pH

Dilution(% w.w)

Inoculum(% w.w)

Solids(% w.w)

Solids(% of TS)

74.5 10.0 8.1 74.3 7.5

76.6 10.0 7.5 73.7 7.6

69.0 10.0 7.8 72.6 7.6

53.7 10.0 7.8 75.3 7.1

78.6 10.0 7.2 71.7 7.7

66.2 10.0 7.8 72.3 7.6

64.1 10.0 7.4 75.7 7.6

71.5 10.0 8.2 74.7 7.8

73.6 10.0 8.5 73.0 7.8

61.1 10.0 7.8 74.6 7.5

Table 5 – Results from anaerobic digestion at different organic loading rates of mixture of llama-cow-sheep manure at 18and 25 8C.

Trial Bioreactors conditions Feedstocks Fluid into reactor Biogas

T (�C) HRT(Days)

OLR(kg VS m�3 d�1)

pH VS(%w,w)

(%)

pH VS reduction(%)

Dailybiogas

(ml)

CH4

conc(%)

Methane yield(m3 kg�1 VS

added)

CH4 prod.Rates (m3 CH4 m�3

reactor)Initial Final

1 18 20 0.5 8.0 1.0 7.2 7.5(0.1) 26.5 57 (11) 42 (2) 0.028 (0.013) 0.013 (0.006)

2 18 30 2.0 7.4 5.9 7.4 7.4 (0.3) 13.1 336 (12) 58 (1) 0.055 (0.003) 0.097 (0.006)

3 18 20 4.0 7.4 8.0 7.4 7.3 (0.1) 14.1 420 (23) 56 (1) 0.032 (0.001) 0.116 (0.005)

4 18 10 6.2 7.5 6.2 7.4 7.2 (0.3) 12.5 296 (13) 44 (2) 0.012 (0.001) 0.067 (0.005)

5 18 10 8.1 7.2 8.1 7.3 7.0 (0.1) 8.3 349 (33) 46 (2) 0.011 (0.001) 0.080 (0.006)

6 25 20 0.5 8.0 1.0 7.5 7.6 (0.2) 32.4 120 (12) 39 (1) 0.047 (0.010) 0.022 (0.004)

7 25 30 2.0 7.4 5.9 7.5 7.4 (0.3) 16.3 530 (15) 52 (1) 0.078 (0.003) 0.139 (0.006)

8 25 20 4.0 7.4 8.0 7.3 7.3 (0.1) 20.7 832 (35) 54 (1) 0.061 (0.002) 0.220 (0.006)

9 25 10 6.2 7.5 6.2 7.4 6.8 (0.2) 15.3 456 (11) 40 (2) 0.016 (0.001) 0.091 (0.004)

10 25 10 8.1 7.2 8.1 7.2 7.0 (0.1) 8.8 861 (37) 45 (1) 0.027 (0.001) 0.194 (0.008)

Standard deviation from 5 consecutive days in parenthesis.

b i o m a s s a n d b i o e n e r g y 3 3 ( 2 0 0 9 ) 5 2 7 – 5 3 3530

phosphorus were determined according to standard methods

[14]. The total solids (TS) content was determined after

a repeated heating (105 �C for 1 h), cooling, desiccating, 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 muffle furnace at a tempera-

ture of 550 �C. TOC was determined by high-temperature

combustion method (Method 5310 B). TKN was measured by

semi-micro-Kjeldahl method (Method 4500-Norg C), potassium

and phosphorus were measured by spectrophotometry

(Method 3500-K and 4500-P, respectively).

0

400

800

1200

1600

0 20 40 60 80 100Time (d)

Bio

gas p

ro

du

ctivity (m

l d

-1)

Fig. 1 – Daily biogas production measured in OLR

experiments corrected at normal conditions (0 8C,

760 mm Hg), Experiments: D1 (D), D2 (,), D3 (6), D4 (B),

D5 (>), D6 (3), D7 (-), D8 (:), D9 (C), and D10 (A).

3. Results and discussion

3.1. Organic loading rate experiments

The effect of varying the organic loading rates on the biogas

productivity and the degree of volatile solid reduction in

anaerobic digestion was studied in ten 2 L reactors at two

different temperatures. A mixture of equal proportion of

llama–cow–sheep manure (volatile solid basis) was used in

these experiments. The experiments were carried out at 18 �C

(exp. 1–5) and 25 �C (exp. 6–10) with organic loading rates

between 0.5 and 8.1 kg VS m�3 d�1 and lasted until steady state

had been reached (2–8 HRT). The bioreactor conditions

(temperature, HRT and OLR) for each trial are given in Table 5,

and the daily biogas production measured at local conditions

and corrected at normal conditions (0 �C, 760 mmHg) is shown

in Fig. 1.

Experiments with an OLR lower than 4 kg VS m�3 d�1

proceeded without indications of instability. However, the

digesters with higher OLR values were more sensitive.

Fluctuating behaviour caused difficulties in stabilizing the

experiments with low HRT and a high VS content in the feed

(after 8 HRTs in experiments 4, 5, 9, 10). Experiment 4 showed

pH oscillations between 6.9 and 7.4 and experiment 9 had

a decreasing pH profile (7.4–6.8), indicating possible organic

overload. Similar behaviour has been reported previously

[15–17]. This behaviour could be attributed to wash out of the

microorganisms or some kind of organic overload. At low OLR

the pH showed stable behaviour, indicating sufficient

buffering capacity.

Steady state values of the measured variables and the

operational conditions of the reactors are given in Table 5. The

methane yield increased from 0.02 to 0.08 m3 kg�1 VS added at

25 �C and from 0.01 to 0.06 m3 kg�1 VS added at 18 �C as the

OLR decreased from 6.2 to 2 kg VS m�3 d�1 (Fig. 2). The

methane production rates tended to increase from 0.01 to

0.12 m3 CH4 m�3 at 18 �C and from 0.02 to 0.22 m3 CH4 m�3 at

25 �C as the loading rates increased until the fermentors were

overloaded (at OLR> 4 kg VS m�3 d�1). The lower values of VS

reductions, the low methane content in the biogas produced,

together with the low methane productivity at higher OLR

(>4 kg VS m�3 d�1 in our system) are probably signals of

hydraulic overload (wash out) or organic overload. This is

supported by previous finding that such stress of the biolog-

ical population may occur as a result of short residence time

leading to bacterial wash out or some kind of inhibition

[10,17].

The observed results suggest that the maximum OLR value

lies between 4 and 6 kg VS m�3 d�1 (Fig. 2a and b) for a system

digesting mixture of llama, cow and sheep manure at

0

10

20

30

40

0 1 2 3 4 5 6 7 8 9OLR (kg VS m

-3d

-1)

Vo

latile S

olid

red

uctio

n (%

)

b

a

0.00

0.02

0.04

0.06

0.08

0.10M

eth

an

e yield

(m

3 k

g-1 V

S ad

ded

)

Fig. 2 – Methane yield (a), and volatile solid reduction (b)

versus organic loading rate using mixture of llama–cow–

sheep manure at 18 8C (A) and 25 8C (>).

0

200

400

600

800

0

200

400

600

800

0

20

40

60

80

a

Bio

gas p

ro

du

ctivity (m

l d

-1)

Bio

gas p

ro

du

ctivity (m

l d

-1)

b

0 20 40 60 80 100 120

Meth

an

e in

b

io

gas (%

)c

b i o m a s s a n d b i o e n e r g y 3 3 ( 2 0 0 9 ) 5 2 7 – 5 3 3 531

temperatures between 18 and 25 �C. Working at higher

temperatures, Hashimoto [18] reported maximum OLRs of

7 kg VS m�3 d�1 at 35 �C and 20 kg VS m�3 d�1 at 55 �C for cattle

waste fermentation. In addition to the temperature effect, also

the difference between waste materials used, especially the

differences in biodegradability, are necessary to consider

[19,20].

Time (d)

Fig. 3 – Biogas productivity (a–b) and methane

concentration (c) from semi-continuous digestion at 25 8C

in mixture experiments of llama-cow-sheep manure from

experiments: D1 (-), D3 (6), D6 (:), and D8 (>).

3.2. Digestion of llama–cow–sheep manure: feedingcomposition effect

The effect of the feeding composition was studied in another

set of experiments with ten anaerobic digesters. The digesters

were fed with mixtures of llama, cow and sheep manure in

different proportions according to Table 3, and were operated

semi-continuously over a period of 100 days at 25 �C after

a start-up period of 20 days. The daily biogas production and

the methane concentration in biogas are shown in Fig. 3.

The digestion of mixture of manures was shown to be quite

stable with respect to daily biogas productivity and methane

concentration after the initial adaptation period (Fig. 3).

The methane concentrations of the biogas were in the range

44–60% and the daily biogas productivity was in the range of

290–570 ml d�1. The change of the mixture composition in the

feed did in no case cause a pH change by more than a half unit,

and the pH value was in the range of 7.3–7.8 in all experiments

(Table 6). The anaerobic digestion of the three different

unmixed manures (llama, cow and sheep) showed clear

differences. The highest methane yield (0.12 m3 kg�1 VS

added) was obtained with sheep manure (exp. D5). In this

experiment a VS reduction of 19% and biogas productivity of

about 500 ml d�1 was obtained. The least satisfying perfor-

mance of the three unmixed manure was that of the llama

manure (exp. D1), giving a methane yield of 0.09 m3 kg�1 VS

added and a 15% reduction of VS. The biogas productivity was

about 300 ml d�1 and the methane concentration in the gas

phase 53%. The results for the cow manure (exp. D4) were in

between the other two (methane yield 0.1 m3 kg�1 VS added,

13% VS reduction, 374 ml d�1 biogas productivity and 55%

methane concentration).

Previous anaerobic digestion studies on cow manure at

25 �C report methane yields of 0.16 m3 CH4 kg�1 VS added

(29.5 L biogas kg�1) [21] and 0.18 m3 CH4 kg�1 VS added [22].

The methane yield obtained in the present study for cow

manure from Bolivian altiplano is lower, which is likely

caused by the differences in manure composition which

affects the degradation process. The theoretical maximum

methane yield (Bu) can be calculated from the amount of VS

consumed (m3 CH4 kg�1 VS con) from Bushwell’s formula [23].

This value represents the methane yield if all organic

components are converted to methane only taken into

account the limits set by the degree of reduction of the

substrates. Based on the composition of cow manure –

approximately 65% carbohydrates, 15% protein, 10% lipids and

10% lignin – the Bu value should be close to 0.5 m3 CH4 kg�1 VS

con [24,25]. Actual experimentally obtainable methane yield –

the so-called the ultimate methane yield (Bo, methane yield as

HRT approaches infinity) – will always be lower than the

Table 6 – Results from anaerobic digestion experiments at different mixture composition at a hydraulic residence time of 50days.

Trial Composition of feedstocks Feed Slurry into bioreactor Biogas

Llama(% VS)

Cow(% VS)

Sheep(% VS)

pH % SVw,w

pHInitial

pHFinal

VS reduction(%)

Biogas vol.(ml d�1)

CH4 conc%

Methane yield(m3 kg�1 VS added)

1 100 0 0 7.90 6.45 7.53 7.49 (0.37) 14.88 382 (15) 53.0 (1.6) 0.09

2 50 0 50 7.45 5.85 7.57 7.42 (0.15) 19.66 567 (10) 50.6 (1.8) 0.14

3 33 33 33 7.45 6.06 7.64 7.57 (0.22) 15.02 485 (12) 53.1(1.2) 0.12

4 0 100 0 7.04 6.23 7.07 7.38 (0.26) 12.52 374 (11) 54.8 (1.8) 0.10

5 0 0 100 7.72 5.63 7.66 7.41 (0.13) 19.54 501 (18) 49.9 (1.0) 0.12

6 0 50 50 7.58 6.02 7.56 7.43 (0.17) 15.78 446 (22) 51.5 (2.5) 0.10

7 50 50 0 8.07 6.06 7.59 7.41 (0.29) 13.70 398 (19) 46.5 (2.2) 0.09

8 67 17 17 8.23 6.35 7.80 7.37 (0.31) 16.38 364 (16) 54.3 (2.1) 0.09

9 17 17 67 8.03 6.21 7.83 7.73 (0.03) 18.36 540 (20) 47.5 (2.5) 0.12

10 17 67 17 7.72 6.21 7.53 7.37 (0.34) 12.08 296 (17) 52.7 (2.6) 0.07

Standard deviation from 5 consecutive days in parenthesis.

b i o m a s s a n d b i o e n e r g y 3 3 ( 2 0 0 9 ) 5 2 7 – 5 3 3532

theoretical yield (Bu). The reason is that a fraction of the

organic material is used to synthesize bacterial biomass, and

recalcitrant organic matter contained in manure (as lignin)

will only be degraded to a limited degree [26]. Typical values of

the ratio Bo/Bu are between 0.2 and 0.5. The amount and type

of bedding material, their degradation processes during pre-

storage, and the species-type, breed and growth stage of the

animals determine the quality of the manure [27]. IPCC

(International Plant Protection Convention) [28] estimated the

Bo of dairy cattle in developed countries to be

0.24 m3 CH4 kg�1 VS added, whereas lower values were

reported by Hill [25] (0.13 m3 CH4 kg�1 VS added) and Moller et

al. [24] (0.15 m3 CH4 kg�1 VS added).

The harsh climatic conditions on the altiplano and the

frost-tolerant forages give a rather different animal diet on

the altiplano compared to low-land developed countries.

The low methane yields obtained in the anaerobic digestion

from manures of the altiplano suggest the presence of high

content of low-soluble and recalcitrant compounds, and the

cow manure from the altiplano also contain a low

percentage of protein (w10%) and a high content of lignin

(w25%) [29].

An improved anaerobic digestion was observed as a result

of mixture of the manures, i.e. a so-called co-digestion effect

was found. Llama–sheep mixture digestion increased the

methane yield from llama with 56% (see Table 6, exp. 1 and 2)

and llama–cow–sheep manure increased the methane yield by

35% (see Table 6, exp. 3). However, the binary mixture llama–

cow manure digestion increased the performance only

marginally. The methane yield from cow manure was also

possible to increase by 18% in cow–llama–sheep manure

(Table 6, exp. 4 and 3). Only a modest synergetic effect from

co-digestion was obtained with sheep manure. Co-digestion

with llama and cow manure increased the methane yield by

13% (cf. Table 6). Previous studies have shown that a mix of

different manures may results in better digestion perfor-

mance through improving the C/N ratio [30], associated

increase in buffering capacity [6], and decreased effect of toxic

compounds on the digestion process [1]. Digestion of llama–

cow–sheep resulted in slightly improved methane yield. The

results indicated that some aspects of llama, cow, sheep

manures benefited the mixture digestion, e.g. the relatively

high nitrogen content from llama manure reduces cow

nitrogen deficiency (Table 1, [29]). Additionally, ammonia

inhibition in a pure llama manure digestion may be avoided by

dilution.

Apart from the advantages in terms of process perfor-

mance demonstrated shown above, the co-digestion of

several manures has additional process economical advan-

tages. Co-digestion on a farm-scale can considerably decrease

the investment cost per unit of methane produced.

4. Conclusions

The results of the present study support that llama manure is

a possible feedstock for anaerobic digestion on the Bolivian

altiplano [29], and the productivity is improved by mixing with

cow and/or sheep manure. Anaerobic digestion of the

mixtures of llama, cow and sheep manure proceeded without

indications of failure at organic loading rates lower than

4 kg VS m�3 d�1 in a semi-continuous process. In conventional

non-stirred digestion at 25 �C with loading rate between 1.2

and 1.3 kg VS m�3 d�1, the methane yields varied from 0.07 to

0.14 m3 kg�1 VS added with a reduction of VS between 12 and

20%. An improved anaerobic digestion was observed as

a result of the mixture of the three manures, with an

increased methane yield in comparison to pure llama manure

with more than 50%. All llama–cow–sheep manure digestion

experiments showed that the system is reliable, with enough

buffer capacity to be used on the altiplano without risk of

instabilities as a result of mixture of these manures. Aspects

that deserve further studies are the special environmental

conditions on the altiplano, with for instance very large

temperature fluctuations between night and day.

Acknowledgements

This work was financially supported by the Swedish Agency

for Research Cooperation (SAREC).

b i o m a s s a n d b i o e n e r g y 3 3 ( 2 0 0 9 ) 5 2 7 – 5 3 3 533

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