Technische Universität München

TECHNISCHE UNIVERSITÄT MÜNCHEN
Lehrstuhl für Pflanzenernährung
Nitrogen availability of biogas residues
Sara El-Sayed Fouda
Vollständiger Abdruck der von der Fakultät Wissenschaftszentrum Weihenstephan

für Ernährung, Landnutzung und Umwelt der Technischen Universität München zur
Erlangung des akademischen Grades eines
Doktors der Agrarwissenschaften
genehmigten Dissertation.
Vorsitzender: Univ.-Prof. Dr. J. Meyer

Prüfer der Dissertation:
1. Univ.-Prof. Dr. U. Schmidhalter
2. Univ.-Prof. Dr. K.–J. Hülsbergen
Die Dissertation wurde am 26.05.2011 bei der Technischen Universität München

eingereicht und durch die Fakultät Wissenschaftszentrum Weihenstephan für
Ernährung, Landnutzung und Umwelt am 07.09.2011 angenommen.
Dedicated to
The soul of my late Father (Prof. Dr. El-Sayed Fouda)

Acknowledgments
Among the many people and organization who have contributed to this work, I am
particularly indebted to the following ones:
First of all I like to thank my supervisor Prof. Dr. Urs Schmidhalter, for accepting me
as his Ph.D.student, giving me the opportunity to work on this project in his lab. I
enjoyed being a part of his group and greatly appreciate his confidence in me,
believing in me and calming me down with the right and constructive words. To him I
owe a great debt of gratitude for his patience and inspiration.
I respectfully express my sincere thanks to Dr. Sabine von Tucher for guiding the
progress of the experimental work, for the valuable discussions, suggestions in data
analysis and interpretation, thoroughly and consistently guiding the writing of the
thesis and for critical comments.
I especially thank Prof. Dr. Joachim Meyer for kindly accepting the position as
chairman of examination committee, and Prof. Dr. Kurt-Jürgen Hülsbergen to be the
co-examiner of this thesis.
I would be honoured to convey my heartfelt thanks and sincere gratitude to Prof. Dr.
Ali A. Abdel-Salam, Department of Soil Science, Faculty of Agriculture, Banha
University, Egypt, for his kind encouragement, permanent advice and the moral
support he always gave.
I am sincerely grateful to the member of the chair of Plant Nutrition, Center of Life
and Food Sciences, Weihenstephan, Technische Universität München, who
provided me with a friendly and supportive climate in which I could conduct my
research.
I take this opportunity to express my special and immense thanks to the Egyptian
Government represented by the General Mission Administration in Cairo and the
Cultural Office in Berlin, Germany for their financial support during my study in
Germany.
Really for achieving progress in science, it is not enough to have a place in a lab and
financial support, for me as a foreign student the environment was most important.
Therefore “Heartfelt Thanks” for the generous hospitality I found here in Germany.
I am also thankful to many friends that me and my family met and who contributed to
make Freising a pleasant and memorable part of our lives, although different
language and culture.
Finally but not least I wish to thank my husband (Abdallah), my children (Youssef
and Farah), my mother and my family at Egypt for their moral support, inspiration
and encouragement. I would praise their perseverance during the years which I
spent away from home striving to complete this work. Without their assistance, this
study would not have been realized.
Freising - Weihenstephan, May 2011
Sara Fouda
Table of contents I
Table of contents
1. Introduction .................................................................................................. 1
1.1 Biogas production as a renewable energy resource ......................................... 1

1.2 N availability of unseparated biogas residues .................................................. 2
1.2.1 Comparison of unseparated biogas residues with other organic fertilizers
......................................................................................................................... 2
1.2.2 Effect of organic fertilizers’ C:N ratio on N mineralization ....................... 3
1.2.3 Characteristics of the unseparated biogas residues compared with
animal slurry ............................................................................................ 4
1.2.4 Biogas residues from different substrates ............................................... 5
1.3 The role of soils in N availability from organic fertilizers ................................... 6
1.3.1 Factors affecting N mineralization in soils ............................................... 6
1.3.1.1 Temperature and water content ................................................... 6
1.3.1.2 Soil texture ................................................................................... 7
1.3.1.3 Soil organic matter content .......................................................... 8
1.3.2 Effect of organic fertilizers on N mineralization in soils ........................... 8
1.4 Separation of biogas residues and other organic fertilizers ............................ 10
1.4.1 NH3 losses from unseparated and separated organic fertilizers during
storage and after application to soil ....................................................... 10
1.4.2 Separation efficiency ............................................................................. 11
1.4.3 Characteristics of solid and liquid fractions of biogas residues and other
manures................................................................................................. 11
1.4.4 Nitrogen availability of separated solid and liquid biogas residues........ 12
1.5 Objectives of the present study ...................................................................... 14
2. Materials and methods .............................................................................. 15
2.1. Experimental conditions ................................................................................ 15

2.2 Fertilizers and soils used in the experiments .................................................. 16
2.2.1 First experiment .................................................................................... 16
2.2.2 Second experiment ............................................................................... 19
2.3. Experimental design ...................................................................................... 21
2.4 Analytical methods ......................................................................................... 26
2.4.1 Soil analysis .......................................................................................... 26
II Table of contents
2.4.2 Plant analysis ........................................................................................ 26

2.4.3 Analysis of biogas residues ................................................................... 26
2.5 Calculations and statistical of analysis ........................................................... 27
3. Results ........................................................................................................ 28
3.1 Chemical properties of unseparated biogas residues and cattle slurry .......... 28
3.2 Yield and N availability of seven different unseparated biogas residues and
cattle slurry as tested in two soils ................................................................... 29
3.2.1. Shoot dry matter yield .......................................................................... 29
3.2.3 Shoot N content .................................................................................... 31
3.2.4 Shoot N offtake ..................................................................................... 33
3.2.5 Apparent N utilization ............................................................................ 35
3.2.6. Additional apparent nitrogen utilization ................................................ 37
3.2.7 Soil nitrogen and carbon contents ......................................................... 39
3.2.8 Correlations between shoot N offtake and C and N in organic fertilizers
and soils ......................................................................................................... 40
3.3 Yield and N availability of ryegrass from three unseparated biogas residues as
tested in five soils ........................................................................................... 42
3.3.1 Nitrogen offtake of unfertilized soils ...................................................... 42
3.3.2 Shoot dry matter yield ........................................................................... 43
3.3.3 Shoot N content .................................................................................... 43
3.3.4 Shoot additional N offtake ..................................................................... 46
3.3.5 Additional apparent N utilization ............................................................ 49
3.3.6 Soil organic matter content ................................................................... 53
3.3.7 Net N mineralization rate from different soils......................................... 53
3.4 Chemical properties of solid and liquid biogas residues after separation ....... 55
3.5 Yield and N availability of ryegrass from liquid and solid biogas residues after
separation as tested in two soils .................................................................... 58
3.5.1 Shoot dry matter yield ........................................................................... 58
3.5.2 Shoot N content .................................................................................... 58
3.5.3 Shoot N offtake ..................................................................................... 63
3.5.4 Apparent N utilization ............................................................................ 66
3.5.5 Additional apparent nitrogen utilization ................................................. 69
3.5.6 N uptake (shoot, stubble and root) from selected treatments ................ 72
Table of contents III
3.5.6.1 N uptake from mineral treatments during five growth cycles ..... 72
3.5.6.2 N uptake in the last growth cycle ............................................... 72
3.5.7 Relationship between N offtake of the liquid and the solid biogas
residues and their Corg:Norg ratio ............................................................ 75
3.5.8 Relationship between N offtake of liquid and solid biogas residues and
the amounts of Norg and Corg applied in five growth cycles .................... 77
3.5.9 Soil total N and total C at the end of the experiment ............................. 78
3.5.10 Relationship between N offtake of the liquid and the solid biogas
residues and soil Nt at the end of the experiment .................................. 79
4. Discussion .................................................................................................. 81
4.1 N availability from different unseparated biogas residues .............................. 81

4.2 Effect of unseparated biogas residues’ C:N ratios on N mineralization .......... 82
4.3 N utilization from different unseparated biogas residues ................................ 84
4.4 Soil N accumulation after five repeated applications of different unseparated
biogas residues ............................................................................................. 84
4.5 N mineralization from different soils................................................................ 85
4.5.1 Effect of soil type and organic matter content on N release from
unfertilized and fertilized soils ................................................................ 85
4.5.2 Effect of organic fertilizers on N mineralization in different soils ........... 87
4.5.3. Soil N accumulation at the end of the experiment from different soils .. 88
4.5.4 Net mineralization of soil organic matter from different soils ................. 89
4.6 Separated biogas residues (liquid and solid) .......................................... 89
4.6.1 N availability from liquid and solid biogas residues ............................... 89
4.6.2 Accumulation of N in shoot, stubble and root at the final biomass harvest
....................................................................................................................... 93
4.6.3 Soil total N after ryegrass harvest at the end of the experiment ............ 93
5. Conclusions ............................................................................................... 95
6. Summary .................................................................................................... 96
7. Zusammenfassung .................................................................................... 99
8. References ............................................................................................... 103

IV List of Tables
List of Tables
Table 1 Monthly mean air temperature measured 2 m above soil surface during the
first experiment from November 2007 to October 2008 ............................. 15
Table 2 Monthly mean temperature measured in the greenhouse during the second
experiment from April 2010 to February 2011 ............................................ 16
Table 3 Characteristics of the soils used in the first experiment ............................... 18
Table 4 Characteristics of the soils used in the second experiment ......................... 21
Table 5 Chemical composition of the biogas residues obtained from different raw
materials and cattle slurry ......................................................................... 28
Table 6 Soil contents of total nitrogen and total carbon at the end of the experiment
.................................................................................................................... 39
Table 7 Correlations between shoot N offtake in the 1st, 2nd and 5th growth cycle and
Corg: Norg ratios of the organic fertilizers ..................................................... 40
Table 8 Correlation coefficients between soil N contents at the end of the experiment
and shoot N offtake (last unfertilized biomass harvest and fifth growth cycle)
................................................................................................................... 41
Table 9 F-statistics of two-way ANOVA of the effects of soils (1, 2, 3, 4 and 5) and
fertilizer sources (mineral, BGR1, BGR6 and BGR8) ................................... 46
Table 10 F-statistics of two-way ANOVA of the effects of soils (1, 2, 3, 4 and 5) and
the fertilizers sources (BGR1, BGR6 and BGR8)....................................... 50
Table 11 Soil Nt content at the end of the experiment .............................................. 53
Table 12 Chemical composition of liquid and solid biogas residues obtained from
different raw materials used for the first, second and third growth cycles .. 56
Table 13 Chemical composition of liquid and solid biogas residues obtained from
different raw materials used for the fourth and fifth growth cycles ............. 57
Table 14 Correlation coefficients between N offtake of five growth cycles and
Corg:Norg ratio of the solid and the liquid biogas residues ........................... 75
Table 15 Correlation coefficients between N offtake of five growth cycles and the
amounts of Corg and Norg applied as solid and liquid biogas residues in two
soils............................................................................................................ 77
Table 16 Soil contents of total nitrogen and total carbon at the end of the experiment
.................................................................................................................. 79
List of Figures V
List of Figures
Figure 1 Composition of substrates used for the fermentation process to obtain

biogas residues for the first experiment ................................................... 18
Figure 2 Composition of different substrates used for the fermentation process to
obtain biogas residues for the second experiment ................................... 21
Figure 3 Time schedule of fertilization and cutting during each growth cycle of
ryegrass in the first experiment ................................................................ 24
Figure 4 Time schedule of fertilization and cutting during each growth cycle of
ryegrass in the second experiment .......................................................... 25
Figure 5 Shoot dry matter yield of different biogas residues and cattle slurry collected
in the growth cycles 1, 2 and 5 in two different soils ................................ 30
Figure 6 Shoot N content of different biogas residues and cattle slurry collected in
growth cycles 1, 2 and 5 in two different soils .......................................... 32
Figure 7 Shoot N offtake of different biogas residues and cattle slurry collected in
growth cycles 1, 2 and 5 in two different soils .......................................... 34
Figure 8 Apparent N utilization of biogas residues and cattle slurry collected in growth
cycles 1, 2 and 5 in two different soils ..................................................... 36
Figure 9 Additional apparent N utilization of the organic fertilizers collected in growth
cycles 1, 2 and 5 in two different soils ..................................................... 38
Figure 10 Relationship between C org:Norg of unseparated biogas residue and cattle
slurry and the N offtake in the first growth cycle ...................................... 41
Figure 11 N offtake of unfertilized treatments in five different soils collected in the 1 st,
2nd and 5th growth cycles ......................................................................... 42
Figure 12 Shoot dry matter yield of different biogas residues collected in growth
cycles 1, 2 and 5 in five different soils ..................................................... 44
Figure 13 Shoot N content of different biogas residues collected in the growth cycles
1, 2 and 5 in five different soils ................................................................ 45
Figure 14 Shoot addtional N offtake of biogas residues collected in the growth cycles
1, 2 and 5 in five different soils ................................................................ 48
Figure 15 Additional apparent N utilization of biogas residues collected in the growth
cycles 1, 2 and 5 in five different soils ..................................................... 52
Figure 16 Net N mineralization rate of fertilizers collected in growth cycles 1, 2 and 5
in five different soils ................................................................................. 54
VI List of Figurs
Figure 17 Shoot dry matter yield of liquid and solid biogas residues collected in five
growth cycles in two different soils........................................................... 60
Figure 18 N content of liquid and solid biogas residues collected in five growth cycles
in two different soils. ................................................................................ 62
Figure 19 Shoot N offtake of liquid and solid biogas residues collected in five growth
cycles in two different soils ...................................................................... 64
Figure 20 Shoot N utilization of liquid and solid biogas residues collected in five
growth cycles in two different soils........................................................... 68
Figure 21 Additional apparent N utilization of liquid and solid biogas residues
collected in five growth cycles in two different soils ................................. 71
Figure 22 Partitioning of N uptake in shoots (all biomass harvests in each growth
cycle), stubbles and roots from the mineral treatment during five growth
cycles for two soils. .................................................................................. 73
cycle), stubbles and roots from the control, mineral fertilizer, liquid biogas
residue 1 and solid biogas residue 1 in the last growth cycle for two soils
................................................................................................................. 74
Figure 24 Relationship between N offtake in the five growth cycles and Corg:Norg ratio
of solid and liquid biogas residues ........................................................... 76
Figure 25 Relationship between ryegrass N offtake in the last cut and soil N t at the
end of the experiment after the applications of liquid and solid biogas
residues in two soils ................................................................................. 80
List of Abbreviations VII
List of Abbreviations
15 labelled nitrogen
N
AANU additional apparent nitrogen utilization
ANOVA analysis of variance
ANU apparent nitrogen utlization
BGR biogas residue
C carbon
C/N carbon/nitrogen ratio
CaCl2 calcium chloride
CAL calcium acetate lactate
CEC cation exchange capacity
Corg organic carbon
Ct total carbon
DF degree of freedom
DM dry matter
H2O water
ICP Inductively Coupled Plasma Emission Spectrometer
K potassium
KCl potassium chloride
LBR liquid biogas residue
LSD least significant difference
Mg magnesium
MS mass spectrometer
N nitrogen
NH4-N ammonium nitrogen
Nmin mineralized nitrogen

VIII List of Abbreviations
NO3-N nitrate nitrogen
Norg organic nitrogen
Nt total nitrogen
OM organic matter
P phosphorus
p≤0.01 1% probability level
p≤0.05 5% probability level
r2 coefficient of determination
SBR solid biogas residue

Introduction 1
1. Introduction
1.1 Biogas production as a renewable energy resource
Few countries are responsible for most of the world’s total greenhouse CO 2 gas
emission in 2009 (BMU, 2010). The United States emit 25% of the world total,
followed by China with 15%. Germany is ranged at the sixth position with 4% of the
world total CO2 gas emission. According to the Kyoto protocol Article 3, the countries
agreed to reduce anthropogenic carbon dioxide equivalent emissions of the
greenhouse gases by at least 5 per cent below the 1990 levels in the period 2008 to
2012. To reach this aim a number of measures are recommended in Article 2 such
as the enhancement of energy efficiency, protection and enhancement of sinks and
reservoirs of greenhouse gases, promotion of sustainable forest management and
agricultural practices, development and increased use of new and renewable forms
of energy. Therefore among other measures Germany’s current policy strives to
increase the contribution of renewable energy resources to substitute fossil energy
resources in order to decrease CO2 emissions but also to become less dependent on
imports of fossil fuel.
A substantial proportion of around 70% of the total renewable energy resources is
provided by biomass transformation (biogenic solid fuels, biogenic liquid and
gaseous fuels, biogenic portion of waste, biogas, sewage gas, land fill gas and bio
fuels). For electricity generation biogas is ranked fourth after wind energy,
hydropower, and biogenic solid fuels. It represents 11% of the renewable energy
resources in Germany in 2009. For heat generation the proportion for biogas in
renewable resources accounted for 8%, and the production of biogas provided 2.4%
of Germany’s total energy consumption (BMU, 2010).
With the aim to further increase energy formation from organic wastes and biomass
production biogas is financially supported by the German government. Generally in
Germany, most biogas digesters are fed with animal wastes as a basic substrate and
energy crops as a co-digestate at mesophilic temperatures (Weiland, 2000). Due to
the limited availability of organic wastes, the co-digestion of agricultural crops was
increased during the last years (Weiland, 2000), with crops such as maize forage,
sweet sorghum, ryegrass, clover and barley being the most frequently used. Analysis
of the biogas evaluation shows that maize and grass silage are the most used co-
substrates in biogas plants (Weiland, 2006). Eighty percent of all biogas plants used
2 Introduction
simultaneous fermentation of manure and maize silage and 50% of them used grass
silage for co-fermentation. More than 30 different energy–rich organic wastes from
agriculture as well as food- and agro-industry were used, and often treated
simultaneously with manure and energy crops (Weiland, 2006). In addition, the
digestion of energy crops as a single substrate in a mono-fermentation becomes also
more important.
1.2 N availability of unseparated biogas residues
1.2.1 Comparison of unseparated biogas residues with other organic fertilizers
Residues of the anaerobic digestion process are valuable sources of plant nutrients
(N, P, K, S, etc.) which have to be recycled in crop production, and particularly for
nitrogen the degradability is a decisive factor for nitrogen availability. In general,
organic fertilizers are derived from different raw materials and that is why they vary in
their nutrient composition. No clear recommendations exist for the application of
organic fertilizers and since they differ in their nutrient composition as well as the rate
of nutrient release, farmers tend to apply either too little or too much of them.
Considerable portions of total nutrients particularly N and P in the organic fertilizers
are organically bound. Chadwick et al. (2000) found that the plant availability of
manure N is influenced by manure composition, hence the N availability from organic
fertilizers is difficult to predict. Gutser et al. (2005) concluded that in the year of
application, plants would take up less than 50% of N applied as organic fertilizers
compared to 50-80% of N applied as mineral fertilizers. It is generally assumed that
in the year of organic fertilizer application, their contents in mineral nitrogen and in
organic substance are associated with the fractions of N which are available for crop
uptake (Olesen et al., 2004; Gutser et al., 2005).
Organic fertilizers contain mineral N in most cases ammoniacal N, which is a readily
plant-available form of N, ranging from very little to 90% of the total N. For example,
NH4-N proportion in total N is 80 to 90% in urine, 40 to 60% in animal slurry, 5 to 20%
in solid manure, and up to 15% in bio-composts or horn meal (Gutser et al., 2005).
Organic N may release ammoniacal N through mineralization. On the other hand,
organic C in the fertilizer may reduce the amount of plant-available N through
immobilization. After application of organic fertilizers the ammonium is available to
plant, but part of it is immobilized in soil during microbial decomposition of organic
Introduction 3
compounds in the cattle slurry (SØrensen and Jensen, 1995). The organic N must be
mineralized before it becomes available to the plant. However, the amount of plant
available N from slurry compared to NH4-N is depending on the soil type, way of
application and time after application.
The N released from the organic sources is difficult to estimate. Computer programs
that calculate manure application rates commonly presume that 100% of the NH 4+-N
and some additional percentage of up to 50% of the organic N becomes available for
plant uptake during the growing season immediately following the organic fertilizers
application (Thompson et al., 1997).
1.2.2 Effect of organic fertilizers’ C:N ratio on N mineralization
The C:N ratio of organic substance is used as a potential predictor for the N release.
Cabrera et al. (2005) summarized factors that control the net N mineralization of
organic residues and concluded from studies by Withmore (1996) that values of C:N
ratio of 20 to 40 in the organic material represent the break-even point between net N
mineralization and net N immobilization. In some cases, this break-even point was
found to be near C to N ratio of 15 (Gilmour, 1998). Another study (Seneviratne,
2000) found that plant residues with C:N ratio up to 27 lead to N mineralization and
ratios exceeding 27 cause N immobilization. Qian and Schoenau (2002) studied the
N release from different solid manures differing in C/N org ratio and indicate that net N
mineralization was related to the C:Norg ratio of the manures and that the highest N
uptake was observed from poultry and hog manures with the highest organic N
contents and the lowest C:Norg ratios of 7.6 and 6.6, respectively. Similar results are
reported by Gutser et al. (2005) who noticed that from organic fertilizers with C:N org
ratios below 6-7 a high N release can be excepted while organic fertilizers with a high
C:N ratio lead to immobilization. From these studies it can be concluded that the net
N mineralization can be enhanced by the application of organic sources with high
quality, i.e. low C:N ratio.
Organic residues with similar C:N ratio may mineralize different amounts of N due to
differences in their composition that are not explained by the C:N ratio. Several
studies have indentified the groups of the compounds present in organic residues
which are possibly involved in net mineralization or immobilization such as proteins,
lignin, soluble carbohydrates, hemicelluloses and cellulose. Wood et al. (2010)
4 Introduction
evaluated 87 different poultry wastes in an incubation experiment and found a

negative correlation between N mineralization and lignin content and a weak but
positive correlation with hemicellulose content. No correlation was observed between
N mineralization and cellulose content. Another study by Jensen et al. (2005) used
76 samples of plant materials to evaluate the C and N mineralization based on mg N
g-1 of added C and showed a negative correlation between cellulose C and
hemicellulose C content and N mineralization. Nourbakhsh. (2006) observed that
after 46 weeks of incubation a significant negative correlation between the lignin
content (r=-0.54*) or lignin/N (r=-0.68**) and N mineralization and a highly positive
correlation between the N content (r=0.96***) and N mineralization from different
plant materials existed.
1.2.3 Characteristics of the unseparated biogas residues compared with animal
slurry
Biogas residues are often described to resemble animal slurry with respect to their
composition, particularly in the value of NH4-N: total N (Nt) which is around 50% in
cattle slurry and between 60 to 70% in pig slurry. Raw undigested animal slurry in
comparison to residues from anaerobic digestion have lower pH value, NH4+-N
contents and the ratio of NH4-N:Nt. However C:N ratio, total C content and organic
dry matter content in anaerobic digestion residues are lower than that in undigested
cattle slurry (Gutser et al.,1987; Asmus et al., 1988; Möller et al., 2008). Gutser et al.
(1987) used 2-4 months stored cattle slurry and anaerobically digested cattle slurry
and found that cattle slurry had a pH value of 7.2, C:N ratio of 9.6, a dry matter
content of 8.5 % and an NH4/Nt ratio of 58 %, while the anaerobically digested cattle
slurry had a pH value of 7.7, C:N ratio of 5.9, a dry matter content of 7% and an
NH4/Nt ratio of 65%.
With regard to the N availability from biogas residues the knowledge is rather limited
and inconsistent. In some field experiments by Ross et al. (1989) there was no effect
of biogas residues on crop yield compared to the unfertilized control, only the plant N
content was found to be higher after the application of biogas residues. Studies by
Messner and Amberger (1987) based on equal amounts of applied NH4+-N showed a
slightly lower N removal from anaerobically fermented compared to untreated cattle
and pig slurry. Möller et al. (2008) reported that in spite of the higher NH4+-N content
Introduction 5
in liquid digested slurry, N uptake from this material resembled that from liquid
undigested slurry. In contrast, other studies found that N uptake from biogas slurry
exceeded that from unfermented slurry by about 10 to 20% (Gutser et al., 1987;
Asmus et al., 1988). De Boer (2008) compared a mineral fertilizer, undigested pig
slurry and pig slurry co-digested with different industrial wastes (such as yeast,
starch or dairy products) and found that the apparent N recovery of three cuts of
ryegrass from co-digested pig slurry was rather comparable to that from the mineral
fertilizer and higher than that from undigested raw slurry. With respect to plant
residues, application of solid and liquid residues from biogas digestion of field crop
residues gave higher N uptake within the crop rotation compared to a system without
the digestion of crop residues (Stinner et al., 2008). Recently, Gunnarsson et al.
(2010) concluded from a 180-day pot experiment based on equal amounts of mineral
N and with subsequent harvests of ryegrass that the N use efficiency and N recovery
were comparable between a biogas effluent and a mineral fertilizer. They further
concluded the organic N fraction to be rather recalcitrant because within this period
the estimated net mineralization of 12% of the organic N was assessed to be not
substantial.
1.2.4 Biogas residues from different substrates
Raw materials used as substrates for biogas production do not only consist of animal
excrements but are mostly mixtures with agricultural crops in widely varying
proportions or are 100% plant based. Digested crops may also cover a wide range of
chemical composition according to plant species and maturity (Kaiser, 2007). For
example in different maize samples the crude protein contents ranged from 7.1 to
9.1% (1.14 to 1.46% N) and neutral detergent fiber (NDF) contents, representing
cellulose, hemicellulose and lignin contents ranged from 38 to 62% in DM. For
ryegrass samples N contents between 1.6 to 5.0% and NDF contents from 38 to 60%
were observed. The effect of different agricultural crops used as raw materials for
anaerobic fermentation on the N availability of biogas residues need more attention.
6 Introduction
1.3 The role of soils in N availability from organic fertilizers
Arable soils are widely varying in physical, chemical and microbiological properties.
Depending on these factors the degradability of added organic fertilizers including
biogas residues may differ.
Ammonium as well as nitrate-N; i.e., mineral N present in organic fertilizers may be
subject to immobilization in building bodies of micro-organisms i.e., into the microbial
biomass. If such immobilized N is not released during the season, N availability for
plants will be reduced (Paul and Beauchamp, 1994). Immobilized N may remain in
the soil organic matter in forms which are recalcitrant to decomposition (Sørensen,
2004). The increase in the rate of N transformation does not necessarily affect the
net rate of N mineralization or N immobilization. Mineral N is continuously released
from the soil organic matter pool or organic fertilizers (N mineralization), predicting
the N mineralization from organic N is difficult because it is affected by several
factors. Because mineralization is a biological process, it only occurs when soil
conditions are suitable for biological activity.
1.3.1 Factors affecting N mineralization in soils
1.3.1.1 Temperature and water content
Temperature and soil water content play a very important role in the decomposition
of organic fertilizers and the uptake of nutrients by plants. Without application of
organic fertilizers, optimum decomposition of soil organic matter by soil
microorganisms occurs at a temperature of around 25 oC. Knoepp and Swank (2002)
examined the effect of temperature at levels of up to 25°C and of soil moisture
contents of 0.20 up to 0.45 g H2O g-1dry soil on soil N mineralization and found high
N mineralization rates at temperatures of 22 and 25°C. The lowest values for N
mineralization were obtained at a temperature of 5°C. With respect to soil water
contents an interaction between soil temperature and soil moisture was observed. An
increase in N mineralization occurred increasing soil water contents up to 0.45 g H 2O
g-1 soil when the temperature was 22 to 25°C. At low temperatures of up to 10°C,
and with soil moisture at about 0.30 to 0.45 g H2O g-1 either no or even a negative
effect on N mineralization was observed in soils. Zaman and Chang (2004)
measured net N mineralization from soils of an agroforestry system and reported that
after 30-day of incubation, soil net N mineralization occurred at 40°C only. At 25°C
Introduction 7
and 5°C there was no net N mineralization. N net mineralization decreased from
100% field capacity (FC) to 75% FC and from 75°C FC to 50°C FC.
1.3.1.2 Soil texture
Soil texture or type may affect the N release from soil organic pools in unfertilized
soils. Egelkraut et al. (2000) studied the effect of the N mineralizable in different soils,
and noticed that the highest N mineralization occurred in the soil with a medium clay
content (14 %) which was however also high in the content of organic N. From the
soils with comparable but lower organic N contents the lowest N mineralization rate
was found for the soil with the highest clay content, while soils high in sand content
could mineralize more nitrogen. Sørensen and Jensen (1995b) found that N
mineralization in unfertilized soils which differed in their texture was lower in the soil
amended with 75% sand than in a soil amended with 50% of sand which indicates
that the soil N accumulation decreased with increasing sand content. Clay soils have
generally more micropores than sandy soils. Hassink et al. (1993) found that the
relative increase in N mineralization after fine sieving was correlated to the
percentage of soil pore space occupied by pores with diameters <0.2 μm in clay soils
(r2 = 0.81). But the correlation with any other pore size class was poor. Fine sieving
enlarges pores and organic matter situated in small pores may become accessible to
microorganisms after sieving. Hassink et al. (1994) suggested that physical
protection plays a more important role in clay soils with relatively more micropores
than in sandy soils with fewer micropores thereby conditions in the soil should be
conducive to mineralization. Since a sandy soil has larger macro pores it is able to
release water (well drained) and allows air to occupy space. As a result of good
aeration in a sandy soil, microorganisms are able to breath. Therefore, the
breakdown of organic fertilizers to release nutrients (mineralization) is fast. In a wet
soil (clay soil) microorganisms are unable to get oxygen due to micro-pores which
are occupied by water under anaerobic conditions and, therefore, mineralization is
poor or slow. Bosstta and Ågren (1997) reviewed soil texture effects on organic
matter and concluded that soil N mineralization was positively correlated with clay
content. In a pot experiment Sørensen and Jensen (1995b) studied the effect of soil
texture on N uptake of ryegrass for 6 months, and found that the N uptake increased
by increasing clay content (loamy soil > sandy loam soil > sand soil). Thomsen et al.
(2001) showed that increasing soil clay content from about 10-40% leads to
8 Introduction
15
decreases in the net N mineralization of ryegrass. Moreover the addition of silt to
the soil did not affect net N mineralization.
1.3.1.3 Soil organic matter content
Basically soils varying in their soil texture also differ in their properties such as pH,
organic matter and Nt content (Hassink, 1993; Egelkraut et al., 2000; Thomsen et al.,
2001; Griffin et al., 2002). In a pot experiment, Legg and Stanford (1967) examined
the N uptake from 12 soils differing in their organic matter content and C/N ratio for 8
weeks and noticed that soils higher in total carbon content (1.9-3.9%) and total
nitrogen content (0.15-0.34%) showed a higher N uptake compared to soils with
lower Ct (0.53-1.58%) and Nt (0.041 - 0.083%) contents. Another study by Egelkraut
et al. (2000) investigated N mineralization in four unfertilized soils differing in their
texture and organic N content and found that N mineralization was higher in soils
which contained a higher Corg and Norg content (1.41 and 0.077%, respectively) while
it was lower from the other soils which had lower C org (0.26-0.44 %) and Norg (0.017-
0.031%) contents. Sørensen and Jensen (1995b) found that the soils which were
higher in a total N and total C content of 0.16 and 1.7%, respectively, resulted in a
higher N uptake by ryegrass than the other two soils which were lower in their total N
and total C content (0.11-0.14% N and 1.3-1.4% C). Li et al. (2003) studied the N
mineralization rate in an incubation experiment with soils differing in their Nt, Corg and
pH, and found that N mineralization increased with increasing soil N contents from
0.14 - 0.26% of Nt. Herlihy (1979) found that the N mineralization potential was
higher in a soil with a high N content compared to the soil that was lower in N
content. Reddy (1982) reported that organic soils very high in total N contents (2.1 -
3.7% in 0 – 30 cm soil depth) released high amounts of mineral N from 230- 870 kg
mineral N ha-1 y-1.
1.3.2 Effect of organic fertilizers on N mineralization in soils
Organic fertilizers may have a different behaviour when used in different soils. A
study by Egelkraut et al. (2000) tested organic fertilizers of cotton leaf residues,
cotton stem residues and compost in different soils in an incubation experiment for
179 days. They observed that the net N mineralization from cotton leaf residues
ranged from 25 to 39% of the applied N dependent on the soil they were applied to,
Introduction 9
but there was no clear relation to soil texture or soil organic matter content. However,
the differences in immobilization of applied cotton stem residues were ascribed to
differences in soil texture and it was concluded that the soils high in sand
immobilized less N compared with soils high in silt and clay. Sørensen and Jensen
(1995a) studied net N mineralization from fresh and anaerobically stored sheep
manure and observed a higher net N mineralization from both manures in the soil
which contained a mixture of 75% soil: 25% sand, compared with pure soil (100%
soil) and the soil containing a mixture of 50% soil: 50% sand.
A few reports deal with the interaction of incorporated organic fertilizers and soil OM.
For mineral fertilizers Hart et al. (1986) found a larger positive additional N interaction
15
for ( NH4)2SO4 in a soil with higher Corg content compared to a soil that was lower in
15
Corg. When ryegrass was grown in 21 different grassland soils fertilized with N la-
belled ammonium nitrate, it was found that yield without and with fertilizer was related
to the total OM of the soils (r = 0.68***, r = 0.69***), but was not significantly related
to their clay or sand content (Whitehead, 1984). Correlation coefficients between N
use efficiency and different soil properties of sand, silt and clay content; and organic
matter were not significant.
N mineralization can also be strongly affected by the way of fertilizer application into
the soil. In most cases, surface applied slurry has a weaker N fertilization effect than
does slurry incorporated into the soil (Sørensen and Amato 2002; Coelho et al.,
2006). NH3 volatilization reduces the recovery of ammoniacal N from surface-applied
slurry. Another possible factor, especially in dry conditions is the adsorption of
ammoniacal slurry N to the very top of the soil, where it is unavailable to the roots. In
a study by Mooleki et al. (2002) it was observed that injection of liquid swine manure
into the soil resulted in greater enhancement of available N, higher grain yield, and
better nitrogen use efficiency than broadcast application. Thus the most efficient way
to minimize N losses from applications of manure is incorporation into the soil.
Recently Meade et al. (2011) compared the N uptake efficiency (NUE) from inorganic
fertilizer and liquid pig manure, and found that it was higher from inorganic fertilizer in
both 2 years studied compared to the liquid pig manure, thereby the NUE was higher
from inorganic fertilizer and was between 58 to 73%, while for the liquid pig manure it
was between 33 to 36%. Most of the studies investigated the N availability from
organic fertilizers i.e., manure, poultry manures but the information about the N
availability from biogas residue application in different soils is still rare.
10 Introduction
1.4 Separation of biogas residues and other organic fertilizers
German livestock farms are producing a large quantity of manure from poultry, pig
and cattle husbandry (BML, 1995). Animal manure contains solid and liquid
components, the solid fraction being characterized by high concentrations of organic
matter and total N and P in comparison to the liquid fraction (MØller et al., 2007;
Bauer et al., 2009). Therefore the slurry separation is an efficient way to recover and
improve the nutrient efficiency. Separation could be done in untreated slurry and
biogas residues of fermented material (MØller et al., 2007). MØller et al. (2000) stated
that slurry separation into solid and liquid fractions can decrease the demand of
storage capacity and reduce transport costs. Hence, the liquid fraction can be
transported over low distances whereas the solid phase can be transported over long
distances to the arable land.
1.4.1 NH3 losses from unseparated and separated organic fertilizers during
storage and after application to soil
In most cases, solid and liquid fractions after separation are usually stored in
uncovered tanks which increase the susceptibility of ammonia loss. Oenema et al.
(2001) concluded that from unseparated organic fertilizers, total loss of gaseous N
from animal housing and slurry storage ranged from about 2 to 15% of total N in the
cattle slurry and 17 to 29% of total N in the pig slurry. It may be assumed that
separation and storage in uncovered tanks could still increase these losses. Large
losses of up to 50% of total initial N were reported in poultry manure (Oenema et al.,
2001). The liquid fraction is characterized by low dry matter content which leads to a
rapid infiltration into the soil thus reducing ammonia volatilization as reported by
Vandré et al. (1997) who tested NH3-N emission from separated and unseparated
slurry after different application methods, i.e., surface and injection application. They
reported that, surface fertilizer application (separated and unseparated slurry) in April
and May led to higher NH3-N losses compared to injection application. Their
unseparated slurry was characterized by a higher dry matter content of 71 g/kg but
after separation it dropped to 44 g/kg. Therefore, the higher NH 3-N loss was
observed from the unseparated slurry compared to the separated one even after
injection. They concluded that dry conditions increased NH3-N loss which led to
decreasing N uptake by the plant. Chantigny et al. (2007) studied ammonia
emissions from raw liquid swine manure (LSM), digested LSM and the liquid fraction
Introduction 11
after the separation of the raw swine manure after annual application for 3 years.
They found that the LSMs differed in their DM content (45.8, 25.9 and 16.5 kg DM/m 3
for raw, separated and digested, respectively). They concluded that the highest
ammonia emission was observed after the application of raw LSM (22 kg NH3-N/ha)
followed by the separated (17.8 kg NH3-N/ha) and the digested LSM (17.7 kg NH3-
N/ha). Bauer et al. (2009) concluded that the solid fraction after separation could be
used as a fertilizer before seeding of the plants whereas the liquid fraction could be
used as a rapidly available fertilizer in the vegetation period.
1.4.2 Separation efficiency
Bauer et al. (2009) reported that the efficiency of separation depends on the dry
matter content (DM) of the fermentation residue. Therefore the higher the dry matter
content is, the higher the proportion of solid phase after separation. They reported a
negative correlation between dry matter content in the fermentation residue and dry
matter content in the liquid phase after separation with r2 = 0.78. Also the relationship
between dry matter content in the unseparated slurry and the proportion of weight
transferred to the solid phase after the separation was studied by M Øller et al. (2002)
and found to be positively correlated with R2 of 0.76.
1.4.3 Characteristics of solid and liquid fractions of biogas residues and other
manures
Peters et al. (2011) stated that there are different techniques of animal manure and
slurry separation. The techniques included sedimentation, centrifugation, drainage
and pressurized filtration. They found that distribution of nutrients i.e., N, C and P
varied depending on the separation technique and particle size fraction. For all
techniques of separation from both pig and cattle slurry more than 50 up to 90% of C
was found in the solid fraction. Similar results were obtained for P. In contrast the
proportion of total N in the solid fraction was lower with about 15 up to 60 %. They
noticed that within the solid fraction organic nitrogen and phosphorus were mainly
associated with the smaller particles (25-250 µm), whereas organic C was mainly
associated within larger particles >250 µm. Additionally, they studied different kinds
of separation (mechanical and chemical separation), and found that the chemical
separation technique resulted in higher C:Norg and P proportions in the solid fraction
than in the liquid fraction. Whereas, the mechanical separation technique resulted in
12 Introduction
higher C, Norg and P proportions in the liquid fraction than the solid fraction. SØrensen
and Thomsen (2005) stated that separation could lead to a more efficient distribution
of nutrients. Therefore, after separation the Nt was higher in the solid fraction
compared to the unseparated manure but decreased slightly in the liquid fraction.
There was a decrease in NH4-N content in the solid fraction compared to the
unseparated manure, while in the liquid fraction the NH 4-N content did not change
compared to the unseparated manure. MØller et al. (2007) carried out a study to
compare the characteristics of manure before and after separation. They showed that
the solid fraction after the separation of pig manure, digested pig manure and dairy
cattle manure was markedly higher in total N and total P in comparison to
unseparated manure, while the contents of NH4-N and K remained rather unchanged.
On the other hand, the corresponding liquid fractions were characterized by very low
total P contents and lower total N contents compared the unseparated manures. The
NH4-N and K contents of the liquid fraction resembled those in the unseparated
manure. The most important differences between the liquid and solid fractions were
the higher contents of total N and P in the solid fraction compared to the liquid
fractions, whereas the NH4-N and K contents were similar in both fractions. MØller et
al. (2002) reported that total N in the solid fraction was 1.7 to 3.54 times higher than
in the untreated manure. MØller et al. (2007) explained the higher Nt content in the
solid fraction compared with the unseparated biogas fermentation residue by the fact
that the mechanical separation transforms more organic N to the solid phase,
contrarily, the dissolved NH4-N remains in the liquid phase. They concluded that solid
and liquid separation of digestate allows maintaining in the liquid fraction a higher
percentage of total N in form of ammonium, readily available to crops, and in the
solid fraction the slow release organic nitrogen will remain. Therefore, fertilizer value
is improved by separation because nutrient composition reflects crops demand.
1.4.4 Nitrogen availability of separated solid and liquid biogas residues
Chantigny et al. (2007) studied N uptake of timothy grass from inorganic fertilizer,
raw swine slurry, digested swine slurry and the liquid fraction of separated swine
slurry using two different soils, a loamy sand and sandy loam. They reported that
after 3 years of annual fertilizer application, the N uptake was higher from the
fertilized treatments compared to the control. Additionally, N uptake was higher from
Introduction 13
both digested and separated slurries than from the raw swine slurry. The order of N
uptake in both soils was, mineral > digested > separated > raw manure.
Chantigny et al. (2008) conducted a field experiment to study the yield of corn and N
availability from an inorganic fertilizer, from raw liquid swine manure (LSM) and its
separated liquid fraction and from digested manure (LSM) using two different soils (a
clay soil and a loam soil). They found that grain DM and grain N uptake were higher
from the different fertilizers compared to the control, while grain DM and grain N
uptake was similar for all LSMs and the mineral fertilizer. Laboski et al. (2010) tested
the potential available N (PAN) from different dairy manure, including raw,
anaerobically digested before and after liquid-solid separation, non-digested
separated liquids and solids, and composted bedded pack manures. Manure was
incubated for 112 d; generally, the digested separated liquid had greater PAN
(52.2%) of Nt than the raw liquid one (33.4%). However, the PAN for the digested
liquid before separation (43.5%) was identical to the raw or digested separated liquid.
On the other side, the digested separated solid manure had the lowest PAN with a
negative value of -14.6%, which indicated N immobilization from this manure. The
authors also studied the correlation between PAN and the composition of different
manures. They found that PAN from the manures was significantly correlated to the
organic fertilizers Nt and NH4+-N (r = 0.81* and 0.78*, respectively). Negative
correlations were found between PAN and the content of dry matter, acid detergent
fibre (ADF) and neutral detergent fibre (NDF) which accentuates that the higher
concentration of lignin, cellulose and hemicellulose in manure leads to both greater
microbial activity and demand of N therefore, reducing the potential available N from
manure. In addition a two-term regression model was developed to predict PAN from
parameters of manure composition. High correlations were obtained for the
parameters ADF:Nt + Ct:Norg (R²=0.913), NDF:Nt + Ct:Norg (R²=0.915) and NH4+-N:Nt +
Norg (R²=0.875). It was concluded that manure separation was effective on N
utilization with or without anaerobic digestion. While liquid and solid separation are
effective in reproducing a liquid fraction with lower dry matter and higher NH4-N
content as percent in Nt compared to the raw manure, which leads to increased the N
utilization from the liquid fraction. On the contrary, the separation of manure creates
opposite properties for the solid fraction compared to the liquid fraction and reduces
the N utilization of the solid fraction.
14 Introduction
1.5 Objectives of the present study
The objectives of the present study are (i) to describe biogas residues which differ in
substrates used for fermentation (plant residues or mixtures of plant residues and
animal wastes), (ii) to determine the effect of these different biogas residues on N
availability after one application as well as after repeated applications, (iii) to evaluate
the role of different soils on N utilization of these biogas residues, (iv) to characterize
the solid and liquid fractions obtained from physical separation of biogas residues
after co-fermentation of different substrates, and to determine the effect on N
availability.
To reach these objectives, as a test system two pot experiments were conducted
each lasting about one year using perennial ryegrass as a model plant in subsequent
harvests and with repeated fertilizer application. Several different biogas residues
unseparated and separated and differing in chemical composition were tested with
different soils to investigate their N availability.
Materials and methods 15
2. Materials and methods
2.1. Experimental conditions
Two pot experiments with perennial ryegrass were conducted at the research station
of the Institute of Plant Nutrition, Technische Universität München, Freising, Germany
(48.4°N, 11.7°E). Experiment 1 was designed with five soils from different sites under
greenhouse conditions from November 2007 to October 2008 (309 days). Experiment
2 was conducted from January 2010 to March 2011 (414 days). When natural day
length was shorter than 12 hours (between October and March) additional light was
applied (photon flux density 550 µmol photon m-2 s-1) to extend the light period to 12
hours and to improve photosynthetic irradiation. In the first experiment air temperature
was kept above 18º C/15°C (day/night). Monthly mean air temperature in Freising from
November 2007 to October 2008 is given in Tab. 1 (Bavarian Research Centre for
Agriculture). In summer the greenhouse walls were kept open for ventilation. Daily
mean temperature from April 2008 to October 2008 was 8.0°C (April 2008) to 17.6°C
(July 2008). For the second experiment the temperature was determined inside the
greenhouse during the eleven months of plant growth (Tab. 2).
Table 1 Monthly mean air temperature measured 2 m above soil surface during
the first experiment from November 2007 to October 2008
Temperature °C
Years Months
Average Min Max
November 1.6 -8.1 10.3

2007
December -0.1 -10.4 13.3
January 1.9 -5.7 12.6
February 2.6 -9.6 19.3
March 4.1 -8.1 19.3
April 8 -5.2 23
May 14.5 1.7 30.2

2008
June 17.3 4.1 31.8
July 17.6 7.8 31.3
August 17.3 5.7 32.2
September 11.9 -0.2 28.2
October 8.6 -1.8 20.9

16 Materials and methods
Table 2 Monthly mean temperature measured in the greenhouse during the

second experiment from April 2010 to February 2011
Temperature °C
Years Months
Average Min Max
April 25.7 20.5 30.8
May 20.4 11.9 40.6
June 24.7 21 30.7
July 21.3 13.9 35.3
2010 August 20.1 12.4 34.8
September 16.6 9.4 28.2
October 16.4 7.8 25.2
November 15.9 11.2 26.3
December 14.1 10.5 26.9
January 14.6 10.6 28.1
2011
February 15.2 10.6 28.9
2.2 Fertilizers and soils used in the experiments
2.2.1 First experiment
In the first experiment, seven different biogas residues were used coded (BGR1,
BGR2, BGR5, BGR6, BGR7, BGR8 and BGR11) and one undigested cattle slurry
coded CS. The biogas residues were collected from different biogas digesters. The
biogas residues including the information on substrate composition were kindly
provided by the Bavarian Research Centre for Agriculture from a research project. The
composition of the substrates digested in the biogas plants differed in their
composition (Fig. 1), some being exclusively plant based (BGR2, 6 and 8), others
consisted of a mixture of plant material and animal excrement (pig or cattle slurry,
poultry manure) (BGR 1, 5, 7 and 11). BGR7 was derived from cattle slurry (CS) used
in the experiment co-digested with 14% maize silage.
Figure 1 Composition of substrates used for the fermentation process to obtain

biogas residues for the first experiment
The organic fertilizers were each tested in five soils (Tab. 3). Topsoils were collected
from arable land near Freising from the locations Marktschwaben (soil 1), Dürnast (soil
2), Thalhausen (soil 3), Mintraching (soil 4) and Grünseiboldsdorf (soil 5). The soils
were air-dried and sieved at 10 mm. The physico-chemical characteristics of the soils
are given in Tab. 3.
Table 3 Characteristics of the soils used in the first experiment
Clay Silt Sand pH Nt Ct P1 K1 Nmin

Soil name
(--------------%----------------) CaCl2 g 100g-1 mg kg -1 mg pot-1
Soil 1 19 73 8 6 0.21 1.9 66 192 53
Soil 2 26 57 17 6.5 0.15 1.2 83 150 57
Soil 3 6 8 86 5.3 0.06 0.41 35 66 11

Soil 4 35 26 31 7.7 0.45 4.4 288 540 288
Soil 5 21 60 19 6.6 0.20 1.7 118 282 55
1
soluble in Ca-acetate/Ca-lactate
2.2.2 Second experiment
In the second experiment, biogas residues from 7 different biogas plants were
separated each into a liquid and a solid fraction to obtain 14 different biogas residues
(Fig. 2). In the 7 biogas digesters different substrates were used for fermentation. The
composition of the substrates is shown in Fig. 2. All substrates were exclusively plant
based except BGR1 and BGR7 consisting from a mixture between plant material and
animal excrement, where BGR1 contained 2% turkey manure and BGR7 contained
67% poultry manure. Generally the biogas residues contained different amounts of
maize silage ranging from 20% in BGR5 and 98% in BGR1.
Figure 2 Composition of different substrates used for the fermentation process

to obtain biogas residues for the second experiment
The biogas residues were studied in two different soils. Topsoils were collected from
two different sites Mühlfeld and Dürnast near Freising in southern Germany and
treated similarly like the soils in the first experiment. For the characteristics of the soils
see Tab. 4.
Table 4 Characteristics of the soils used in the second experiment
1
Clay Silt Sand pH Nt Ct P1 K
Soil
(--------------%--------------) CaCl2 g kg-1 mg kg
-1
Mühlfeld 13 21 66 5.7 0.09 0.80 66 192
Dürnast 57 26 17 6.5 0.13 1.09 83 150

1
extractable in Ca-acetate/Ca-lactate
2.3. Experimental design
Pots used in the experiments (24.5 cm length, 20.0 cm width and 24.0 cm height)
received 10 kg of air dried soil. Perennial ryegrass seeds (Lolium perenne. L. cv.
Belcampo used in the first experiment and Lolium perenne. L. cv. Ivanna used in the
second experiment) were sown at a rate of 2.0 g/pot in two rows. Soils were regularly
watered with distilled water to achieve 70% of the maximum water-holding capacity
(WHC) during the experiments.
Biogas residues and cattle slurry were applied to the pots in 3 rows each located
besides and between the plant rows. To avoid NH3 losses the organic fertilizers were
incorporated into the soil by slit application and immediately covered with soil. As
control treatments pots with mineral fertilization (NH 4NO3) and without fertilization were
prepared. All fertilizer applications with biogas residues or cattle slurry were based on
300 mg NH4+-N per pot and on 300 mg N as NH4NO3 for the mineral treatment.
Fertilizer application was done five times in both experiments, i.e. once at the
beginning of each growth cycle.
For the first experiment altogether nine cuts of ryegrass were taken (grass was cut 3
cm above the surface of the soil). Cuts No. 1, 2 and 3 represent the first growth cycle
(Σ of 1, 2 and 3 cuts), and the cuts 4 and 5 represent the second growth cycle (Σ of 4
and 5 cuts), and the cuts 8 and 9 represent the fifth growth cycle (Σ of 8 and 9 cuts).
Cut 1 was taken in mid-December 2007 and cut 9 was taken in mid-October 2008. For
equal experimental conditions one cut was removed prior to the first fertilization to take
up the mineral N present at the beginning of the experiment. Dates for fertilizer
application and cutting of ryegrass in the first experiment are given in Fig. 3. Because
of the optimal soil P (> 44 mg P kg -1) and K (> 120 mg K kg-1) contents (see Tab. 3)
only Mg (200 mg pot-1) and S (260 mg pot-1) were applied as basic fertilization. At the
end of the experiment shoots were tested for P, K, Mg and S concentrations and were
found to be in the optimal range.
The pots were placed in the greenhouse in a randomized block design (four replicates
per treatment). At the end of the experiment soil samples (4 pooled cores per pot from
top to bottom) were taken immediately after the last cut to be analyzed for total N, total
-1
carbon and mineral N. Mineral N was found to be less than 84 mg pot in
-1
Marktschwaben soil and 80 mg pot in Dürnast soil, and is therefore not considered
for further data interpretation.
For the second experiment also including five growth cycles (Fig. 4) the grass was
cut 10 times during the experiment, i.e. twice within each growth cycle (stubble height
3 cm from the soil surface). Cut 1 was taken in middle of March 2010 and the last cut
was taken in the first week of March 2011.
For an equal supply of K, P, Mg and S among the treatments and for balancing K, P,
Mg and S applied by the biogas residues control and mineral treatments were supplied
with K2HPO4 at rate of 500 mg K pot-1 corresponding to 200 mg P pot-1 and with
MgSO4 at rate of 50 mg Mg pot-1 and 67 mg S pot-1. Moreover liquid biogas residues
treatments were supplied with CaHPO4 at a rate of 200 mg P pot-1 and with MgSO4 at
a rate of 50 mg Mg pot-1 and 67 mg S pot-1.
The experiment included additional mineral treatments for the collection of roots and
stubbles after each growth cycle. The pots were placed in the greenhouse in a
randomized block design (three replicates per treatment). At the end of the experiment
a soil sample from each pot was taken immediately after the last cut. The soil was
mixed thoroughly, then sieved to 2 mm, ground and analyzed for Nt and Ct.
Fertilization
1st fert. 2nd fert. 3rd fert. 4th fert. 5th fert.
3.12.2007 6.2.2008 6.5.2008 13.6.2008 23.7.2008
Sowing
6.11.2007
Removed
cut
1st cut 2nd cut 3rd cut
17.12.2007 9.1.2008 6.2.2008
4th cut 8th cut 9th cut

Harvest 5th cut 6th cut 7th cut
First growth cycle (90 DAS) 4.3.2008 25.8.2008 15.10.2008
22.4.2008 5.6.2008 16.7.2008
(Σ of 1st, 2nd and 3rdcuts)
Fifth growth cycle (309 DAS)
Second growth cycle (166 DAS) th th
(Σ of 8 and 9 cuts)
(Σ of 4th and 5th cuts)
Figure 3 Time schedule of fertilization and cutting during each growth cycle of ryegrass in the first experiment
Fertilization st
1 fert. 2nd fert. 3rd fert. 4th fert. 5th fert.
11.02.2010 28.04.2010 15.7.2010 9.10.2010 21.12.2010
Sowing
13.01.2010
2 Removed
cut st nd
1 roots and 2 roots and
st nd stubbles stubbles
1 cut 2 cut
10.03.2010 sample sample 3nd roots and 4th roots and 5th roots and
27.04.2010 6.05.2010 14.07.2010 stubbles stubbles stubbles
sample sample sample
3th cut th
4 cut
th
5 cut
th
6 cut 8.10.2010 20.12.2010 8.3.2011
First growth cycle
Harvest (Σ of 1st and 2nd cuts)
2.06.2010 14.07.2010 17.8.2010 6.10.2010
Second growth cycle Third growth cycle

(Σ of 3th and 4th cuts) (Σ of 5th and 6th cuts)
7th cut 8th cut th

9 cut 10th cut
17.11.2010 20.12.2010 1.2.2011 7.3.2011
Fourth growth cycle Fifth growth cycle

th th th th
(Σ of 7 and 8 cuts) (Σ of 9 and 10 cuts)
Figure 4 Time schedule of fertilization and cutting during each growth cycle of ryegrass in the second experiment
2.4 Analytical methods
2.4.1 Soil analysis
Soil texture was determined by wet-sieving for the sand fraction and by the pipette
method for the silt and clay fractions Normenausschuss Wasserwesen (2002). The pH
value was measured in 0.01 mol L -1 CaCl2 solution (Hoffmann, 1997). Ct and Nt were
analyzed following the Dumas procedure (combustion method combined with an
isotopic ratio mass spectrometer (Anca SL 20-20 Crewe Europe Scientific UK)). Plant
available P and K in soil were measured after extracting with 0.1 mol L -1 Ca acetate +
0.1 mol L-1 Ca lactate + 0.3 mol L-1 acetic acid at pH 4.1 with soil-to-extracting ratio of
1:20, shaking for 2 h (Schüller, 1969). For the soil mineral N content NO3-N was
extracted with 0.0125 mol L-1 CaCl2 (1:2) and NH4-N with 1 mol L-1 KCl (1:5). Nitrate
was measured by HPLC according to Vilsmeier (1984) and ammonium according to
Mulvaney (1996).
2.4.2 Plant analysis
For each cut fresh and dry matter yield were determined. Root and stubble dry matter
yield were determined after each growth cycle. Dry matter yield was obtained after
drying for 24 hours at 105 °C. Total N was determined after grinding of dried material
by elemental analysis according to the Dumas method (combustion method combined
with an isotopic ratio mass spectrometer (Anca SL 20-20 Europe Scientific, Crewe,
UK)).
Plant analysis for P, K, Mg and S was conducted after wet digestion with HNO 3 and
H2O2 in closed vessels in a microwave oven. The nutrient concentrations were
measured with ICP (Liberty, Varian, Mulgrave, Australia).
2.4.3 Analysis of biogas residues
Total N and NH4+-N in the biogas residues were determined in fresh subsamples using
the Kjeldahl technique (Vapodest 12, Gerhardt, Germany) prior to and after the
digestion with H2SO4 conc. Norg content was calculated as the difference between N t
and NH4+-N. Dry matter was determined after drying at 105 °C. Total carbon was
determined in freeze dried subsamples according to Dumas combustion method
combined with an isotopic ratio mass spectrometer (Anca SL 20-20 Europe Scientific,
Crewe, UK). Carbonate content was measured following the Scheibler procedure
(Schlichting et al., 1995) according to DIN 18129. Corg was calculated as the difference
between Ct and CO3-C.
2.5 Calculations and statistical analysis
The apparent nitrogen utilization (ANU), which refers to the mineral N component of
the fertilizers (NH4+-N in biogas residues and slurry), was calculated as:
ANU = (PNF – PNO)/NF

Additional N offtake = (PNF – PNO)
where
PNF = the amount of N taken up by the plant shoots in the fertilized treatment,
PNO = the amount of N taken up by the plant shoots in the unfertilized treatment, and
NF = the amount of mineral N applied (i.e., as NH4-N in the organic fertilizers).
The additional apparent nitrogen utilization of organic fertilizers (AANU) was

calculated as:
AANU = (ANUorg – ANUmin) / ANUmin *100

where ANUorg and ANUmin = the apparent nitrogen utilization of the organic fertilizer
(biogas residues and cattle slurry) and mineral treatments, respectively.
Soil net mineralization rate was calculated as:

Soil net N mineralization rate, % = N offtake of an unfertilized cut/ soil N t *100.
This rate was calculated for the last cut and the soil N t content after the experiment
and expressed per day. For comparison with data from the literature it was expressed
as % per year.
PASW version 17.0 (PASW Inc., Chicago, IL) and Microsoft Excel 2007 were used for
statistical analysis. The results of the pot experiments were subjected to one- or two-
way analyses of variance with the significance of the means tested with a Tukey,
HSD-test at P≤ 0.05. Correlation analyses were conducted to determine the
relationship between C org/Norg in biogas residues or soil N content after the experiment
and N offtake.
28 Results
3. Results
3.1 Chemical properties of unseparated biogas residues and cattle slurry
The biogas residues obtained from the biogas digesters after the fermentation of
different raw materials showed distinct differences in the chemical composition (Tab.
5). Compared to CS with 12.1%, the dry matter content of the biogas residues were
similar or lower ranging from 5.2 to 12.5%. The NH 4+-N contents were comparable
(BGR5 and 7) or markedly higher (0.49 to 0.51% in BGR11 and 1) than in CS with
0.22%. The total N contents ranged from 0.36 to 0.75%. Part of the total C (ranging
from 2.0 to 4.74%) was detected as CO3-C (0.20 to 0.48%). As a result from both
varying Corg and Norg contents the Corg/Norg ratio ranged from 8.6 (BGR1) to 13.4
(BGR7) in the biogas residues which is lower than in CS. Both the P and K contents
varied by the factor of about 3 in all organic fertilizers (data not shown).
Table 5 Chemical composition of the biogas residues obtained from different

raw materials and cattle slurry
NH4-
Nt Ct CO3-C Corg Norg
Dry N NH4-
Treatments pH Corg/Norg
matter N/Nt
% in FM
BGR 1 6.3 8.4 0.75 0.51 2.27 0.20 2.07 0.24 8.6 0.68
BGR 2 6.8 8.2 0.49 0.29 2.89 0.32 2.57 0.20 12.9 0.60
BGR 5 5.7 8.1 0.42 0.25 2.16 0.28 1.88 0.17 11.1 0.60
BGR 6 12.5 8.3 0.72 0.37 4.74 0.26 4.48 0.34 13.2 0.52
BGR 7 7.4 8.2 0.36 0.2 2.49 0.34 2.15 0.16 13.4 0.55
BGR 8 5.2 8.2 0.48 0.27 2.00 0.23 1.77 0.21 8.4 0.57
BGR 11 6.1 8.3 0.7 0.49 2.45 0.48 1.97 0.22 9.0 0.69
CS 12.1 7.9 0.44 0.22 4.41 0.14 4.27 0.23 18.6 0.49
% Norg % = N total % – NH4-N%; Corg % = C total % – CO3-C%
Results 29
3.2 Yield and N availability of seven different unseparated biogas residues and
cattle slurry as tested in two soils
3.2.1. Shoot dry matter yield
The biogas residues and cattle slurry application increased the shoot dry matter yield
of ryegrass over the five growth cycles in comparison with the control. In
Marktschwaben soil, the increase in dry matter yield in the first growth cycle ranged
from 16.1 g/pot (BGR2) to 18.4 g/pot (BGR8), while in Dürnast soil it ranged from 13.1
g/pot (CS) to 15.6 g/pot (BGR1). In the different treatments, including control, shoot
dry matter yield was high in the second and fifth growth cycle in comparison to the first
growth cycle; this is clear in both soils (Fig. 5).
In the first and the second growth cycle, the fertilized treatments were significantly
higher in the dry matter yield than the unfertilized treatment (control), but there was no
significant difference observed between the biogas residues and the mineral fertilizer
in both soils.
In the fifth growth cycle, the organic fertilizers treatments (CS, BGR5, 7 and BGR8) in
the Marktschwaben soil were significantly higher in the dry matter yield than the
mineral fertilizer. In the fifth growth cycle, the organic fertilizers treatments in Dürnast
soil were significantly higher in the dry matter yield than the mineral treatment, except
with BGR2 was significantly lower than the mineral treatment.
30 Results
Figure 5 Shoot dry matter yield of different biogas residues and cattle slurry
collected in the growth cycles 1, 2 and 5 in two different soils. Different
letters indicate statistically significant differences between the fertilizers (p 
0.05)
Results 31
3.2.3 Shoot N content
The whole biogas residues and cattle slurry had a large effect on the shoot nitrogen
content of the ryegrass, however it reached up to 4.5 and 4.0% in the first and fourth
cut (fertilized cuts), respectively in both soils (Fig. 6). Shoot N content was generally
higher in the fertilized cuts (first and fourth) than the unfertilized cuts (second, third and
fifth) and this is clear with the first and the second growth cycles in both soils (short
term fertilizers application). Shoot N content of the fertilized treatments in the
unfertilized cuts was decreased to half in the second and third cuts compared to the
first cut.
After five times of fertilizer application (long term of fertilizers application) this was not
the case with the eight cut after the fifth fertilizer application, where the grass N
content was lower in the eight cut compared to the ninth cut. Also after the fifth
fertilizer application (eighth cut), shoot N content was decreased to half in comparison
to the shoot N content in the first and second fertilizer application (first and fourth
cuts).
32 Results
Figure 6 Shoot N content of different biogas residues and cattle slurry collected
in the growth cycles 1, 2 and 5. in two different soils. Error bars
represent standard deviation
Results 33
3.2.4 Shoot N offtake
Shoot N offtake from all organic fertilizers, biogas residues and cattle slurry, was
generally comparable or higher than that from the mineral fertilizer in both soils (Fig.
7). However, there were significant differences in the N offtake between the different
organic fertilizers.
In the first growth cycle the N offtake from all biogas residues except BGR2 was
significantly higher compared to cattle slurry in the soil Marktschwaben. In soil Dürnast
only BGR1, 5, 8 and 11 yielded a higher N offtake compared to CS. Hence an
increase in the N offtake from cattle slurry co-fermented with maize (BGR7) compared
to undigested cattle slurry was found only in the soil Marktschwaben.
In the second growth cycle both the differences among the organic fertilizers and
between the organic fertilizers and the mineral treatment were higher in the soil
Marktschwaben than in the soil Dürnast. In the soil Marktschwaben all biogas residues
except BGR2 and BGR 7 resulted in a significantly higher N offtake to mineral N.
In the fifth growth cycle the N offtake from all organic fertilizers except BGR2
significantly exceeded that from mineral N in both soils. In this growth cycle a
significantly higher N offtake was obtained from the undigested cattle slurry compared
to BGR7.
34 Results
Figure 7 Shoot N offtake of different biogas residues and cattle slurry collected
in growth cycles 1, 2 and 5 in two different soils. Different letters indicate
statistically significant differences between the fertilizers (p  0.05)
Results 35
3.2.5 Apparent N utilization
The apparent N utilization (ANU) calculated on the basis of applied NH 4-N for the
organic fertilizers and on NH4NO3 for the mineral N ranged between about 80%
(mineral, CS and BGR2) up to about 100% (BGR1, 5 and 11 in soil Marktschwaben
and BGR 11 in soil Dürnast) in the first growth cycle (Fig. 8).
In the second and fifth growth cycle ANU differed between about 70% (mineral
treatment and BGR2) to 80 - 90% for most of the biogas residues or between about
50-60% (mineral treatment and BGR2) up to about 90% (i.e. BGR8) respectively. In
consequence with some exceptions ANU seemed to decrease from the first to the fifth
growth cycle.
36 Results
Figure 8 Apparent N utilization of biogas residues and cattle slurry collected in

growth cycles 1, 2 and 5 in the two different soils. Different letters
indicate statistically significant differences between the fertilizers of one
growth cycle (p  0.05)
Results 37
3.2.6. Additional apparent nitrogen utilization
Because for the mineral fertilizer ANU decreased across the growth cycles, the ANU
of the organic fertilizers was related to that of the mineral fertilizer in each respective
growth cycle. Such a relationship compensates for any differences in plant growth
caused by, for example, by different climatic conditions during the growing season and
enables the evaluation of the possible changes in the ANU of the organic fertilizers as
a consequence of repeated fertilizer application. The additional ANU, therefore, is a
sensitive measure for the additional net nitrogen release from the organic fraction of
the fertilizers that becomes available to the plant in addition to that from the applied
ammonium (Fig 9).
In the first growth cycle, up to 20% more N (e.g. for BGR11) was taken up by the rye
grass leaves from the organic fertilizers in both soils in addition to that from the
supplied mineral N. However, no additional N utilization was found for BGR2 and CS,
and for BGR2, 6, 7 and CS in the Marktschwaben and Dürnast soils, respectively.
AANU increased significantly by the repeated application of the organic fertilizers,
ranging from up to 20% in the first growth cycle (one application) to up to 70% in the
fifth growth cycle (five fertilizer applications). The greatest increase in AANU from the
first to the fifth growth cycles was observed for BGR8, CS and BGR6. By contrast,
both the final AANU in the fifth growth cycle as well as the increase compared to the
value in the first growth cycle was relatively low for BGR11, 7 and 2.
38 Results
Figure 9 Additional apparent N utilization of the organic fertilizers collected in

growth cycles 1, 2 and 5 in two different soils. Different lower case letters
indicate statistically significant differences between the fertilizers within one
growth cycle (p  0.05). Different capital letters indicate statistically
significant differences between the first, second and fifth growth cycle for one
organic fertilizer (p  0.05)
Results 39
3.2.7 Soil nitrogen and carbon contents
The five-fold repeated application of fertilizer across the duration of the experiment
changed soil Nt and Ct contents differently depending on the fertilizer applied (Tab. 6).
In both soils, repeated application of BGR 6 and 8 resulted in significantly higher Nt
and Ct soil accumulation compared to BGR 1 and 11; the value for the latter not being
significantly different from that for the mineral fertilizer application. High soil N t and Ct
content were also observed for CS, with values being comparable to those with BGR
6 and 8. BGR 2, 5 and 7 comprise an intermediate group characterized by moderate
Nt and Ct accumulation although the soil Ct content of BGR 7 was in the range of BGR
6 and 8. Generally, the soil levels of N t and Ct were lower in the Dürnast soil
compared to the Marktschwaben soil.
Differences in the final soil Nt contents were correlated with the different amounts of
organic N input from the organic fertilizers (r = 0.81* (p ≤ 0.05) and r = 0.93** (p ≤
0.01) in the Marktschwaben and Dürnast soils, respectively). No significant
relationship was observed between the organic C input and the final soil Ct content.
Table 6 Soil contents of total nitrogen and total carbon at the end of the
experiment
Marktschwaben soil Dürnast soil
Treatments 1 2 3 4
Nt% Ct% Nt% Ct%
a a a a
Control 0.19 1.84 0.13 1.17
ab ab ab a
Mineral 0.20 1.90 0.14 1.24
cd d e f
CS 0.21 2.13 0.18 1.63
bc bc bc abc
BGR1 0.20 1.97 0.15 1.31
cd cd cd bcd
BGR2 0.21 2.05 0.16 1.44
cd cd cd bcd
BGR5 0.21 2.05 0.16 1.42
d d de def
BGR6 0.22 2.13 0.17 1.48
cd d cd ef
BGR7 0.21 2.10 0.16 1.53
d d de ef
BGR8 0.22 2.13 0.17 1.54
bc bc bc ab
BGR11 0.20 1.96 0.15 1.30
Content before fertilizer application: 1Nt = 0.20, 2Ct = 1.90, 3Nt = 0.14, 4Ct = 1.19. Different
letters indicate statistically significant differences between the fertilizers for either N t or Ct for a
given soil (p  0.05)
40 Results
3.2.8 Correlations between shoot N offtake and C and N in organic fertilizers and
soils
With the aim to evaluate relations between the plant N offtake and properties of the
organic fertilizers correlations were calculated with the C org:Norg of the organic
fertilizers. The N offtake of the ryegrass showed a significant negative correlation to
the Corg:Norg of the organic fertilizers after the first growth cycle in both soils (Tab. 7).
However, for the second and fifth growth cycle i.e. after the second and the fifth
fertilizer application in none of the soils any significant correlation was found between
N offtake and Corg:Norg of the organic fertilizers. It seems that digestion of the cattle
slurry increased the N uptake (which is higher in C org: Norg) and was conducive to more
mineralization compared to BGR7 and consequently gave a higher N offtake (Fig. 10).
Table 7 Correlations between shoot N offtake in the 1 st, 2nd and 5th growth cycle
and Corg: Norg ratios of the organic fertilizers
Soil Growth cycle r
first -0.781*
Marktschwaben soil second -0.410 NS
fifth -0.105 NS
first -0.833**
Dürnast soil second -0.424 NS
fifth -0.071 NS
Significance levels: *(5%); ** (1%); NS (not significant)
Results 41
Figure 10 Relationship between Corg:Norg of unseparated biogas residues and

cattle slurry and the N offtake in the first growth cycle
After five repeated applications, however shoot N offtake from the fertilizers was
significantly correlated to the soil total N content at the end of the experiment (Tab. 8).
This was observed for the ryegrass N offtake of the whole fifth growth cycle but was
most prominent in the last, unfertilized cut where correlation coefficients of 0.83**
(Marktschwaben soil) and 0.80** (Dürnast soil) between the N offtake of ryegrass and
soil total N content were found.
Table 8 Correlation coefficients between soil N contents at the end of the

experiment and shoot N offtake (last unfertilized biomass harvest and
fifth growth cycle)
Shoot N offtake
N content in soil Shoot N offtake (fifth growth cycle)
(last cut)
Marktschwaben 0.72* 0.83**
Dürnast 0.70* 0.80**
Significance level: * (5%) and ** (1%)
42 Results
3.3 Yield and N availability of ryegrass from three unseparated biogas residues
as tested in five soils
3.3.1 Nitrogen offtake of unfertilized soils
The N offtake in plants from unfertilized soils can be used as indicator to describe the
N mineralization of organic nitrogen in soils. The N offtake of unfertilized soils ranged
between about 40 to 340 mg N/pot and differed between the soils (Fig. 11). In all
growth cycles the highest N offtake was observed in soil 4 followed by soil 5, 1 and 2.
Soil 3 showed the lowest N offtake. The N offtake from the different soils was an
indication of the soil total N content which was highest in soil 4 and lowest in soil 3 see
Tab. 3.
Figure 11 N offtake of unfertilized treatments in five different soils collected in

the 1st, 2nd and 5th growth cycles. Different letters indicate statistically
significant differences between the soils within one growth cycle (p ≤ 0.05)
Results 43
3.3.2 Shoot dry matter yield
Application of N either as mineral fertilizer or biogas residues, in the different soils

increased the ryegrass dry matter yield (Fig. 12). In the first growth cycle (after one
fertilizer application), dry matter yield of the fertilized treatments was significantly
higher compared to the unfertilized treatments. In the first growth cycle, there was no
significant difference in the dry matter yield between the biogas residues and the
mineral treatment in soil 2 and soil 3, but in soil 1, 4 and 5 only BGR8 was significantly
higher compared to the inorganic treatment.
In the second and the fifth growth cycle, ryegrass dry matter yield of the different
biogas residue treatments were significantly greater compared with the inorganic
treatment for soil 1 and 2. However in soil 3 the yield from either BGR6 or BGR8 was
significantly higher than from the mineral fertilizer. For soil 5, only BGR8 resulted in a
significantly higher yield than from the inorganic fertilizer treatment.
Treatments which received fertilizers showed considerably higher N content compared

to the unfertilized control, particularly after the first applications of fertilizers. The shoot
N concentration was higher in both cuts after the fertilization (first and fourth cuts)
compared to the unfertilized cuts (second, third and fifth cut) but this is not true for the
eighth cut (Fig. 13).
After the fifth application of fertilizers, the eighth cut had a lower N content than the
ninth cut. In addition, for the eighth cut the N content dropped to half after the fifth
fertilizer application compared to the first and fourth cuts (fertilized cuts). Different soils
had similar effects on N content in all growth cycles.
44 Results
Figure 12 Shoot dry matter content of different biogas residues collected in

growth cycles 1, 2 and 5 in five different soils. Different letters indicate
statistically significant differences between the fertilizers (p  0.05)
Results 45
Figure 13 Shoot N content of different biogas residues collected in the growth

cycles 1, 2 and 5 in five different soils. Error bars represent standard
deviation
46 Results
3.3.4 Shoot addtional N offtake
Statistical analysis of the effects of different soils and mineral or organic fertilizers
applied (Tab. 9) showed that for all growth cycles there were significant effects for both
the soil-factor and the fertilizer-factor on the N offtake of ryegrass but there were no
significant interactions between soil and fertilizer.
The effect of the soil-factor, averaged over all fertilizers was inconsistent and varied
from one growth cycle to another; i.e. the lowest addtional N offtake in the first growth
cycle was observed in soil 2 whereas in the second growth cycle this was true for soil
4; and in the fifth growth cycle it was true for soil 1 (Fig. 14).
In each growth cycle the additional N offtake from the biogas residues, averaged over
all soils, was significantly higher than that from the mineral fertilizer and this effect
seemed to increase with time (see also Tab. 9). While in the first and second growth
cycle no differences appeared between the biogas residues, in the fifth growth cycle
the additional N offtake increased significantly from biogas residue 1 to 6 and 8.
Table 9 F-statistics of two-way ANOVA of the effects of soils (1, 2, 3, 4 and 5)

and fertilizer sources (mineral, BGR1, BGR6 and BGR8)
ANOVA 1st growth cycle 2nd growth cycle 5th growth cycle
main
effects+ DF F Significance DF F Significance DF F significance
value value value
interactions
Soil 4 4.98 0.002* 4 8.31 0.000*** 4 2.58 0.046*
Fertilizer 3 9.28 0.000*** 3 30.45 0.000*** 3 104.0 0.000***
Soil × 12 1.48 0.157 NS 12 1.05 0.420 NS 12 1.548 .132 NS

Fertilizer
Significant level: * P ≤ 0.05. * P ≤ 0.001. * P ≤ 0.0001
Results 47
Main
factor AB A AB B B
soil
Main
factor A B B B
fertilizers
Main
BC AB AB A C
factor soil
Main
factor A B B B
fertilizers
48 Results
Main
factor A AB AB B AB
soil
Main
factor A B C D
fertilizers
Figure 14 Shoot addtional N offtake of biogas residues collected in the growth

cycles 1, 2 and 5 in five different soils. Shoot N offtake in unfertilized
treatments was subtracted for each soil respectively. Different lower
case letters indicate statistically significant differences between the
fertilizers within one soil (p ≥ 0.05)
Results 49
3.3.5 Additional apparent N utilization
In order to evaluate the biogas residues for the amount of N that was plant available
additionally to the applied ammonium the additional N offtake was calculated as the
difference in N offtake between the organic fertilizers and the mineral fertilizer for each
soil related to the N offtake of the mineral fertilizer. This procedure also allows
comparing different soils and growth cycles irrespective of different levels of N offtake
achieved from the mineral fertilizer.
In the first growth cycle the additional apparent N utilization (AANU) for all soils and
fertilizers accounted for an average value of 12%. In the second growth cycle the
AANU increased to 24 % and further to 46% in the fifth growth cycle (Fig. 15).
Statistical analysis of the effects of the soils and fertilizers in each growth cycle
showed that the effect of soil on the AANU was significant (Tab. 10) whereas the effect
of the fertilizers was significant only in the fifth growth cycle. No significant interaction
was found between the two factors.
The effect of soil, averaged over all fertilizers in the first and second growth cycle
showed that the highest additional apparent N utilization occurred in soil 4 and the
lowest AANU was in soil 2 (Fig. 15). In the fifth growth cycle however, the AANU in soil
3 was significantly lower compared to the other soils. Averaged over all soils there was
no effect of the fertilizers on additional N offtake in the first and second growth cycle.
However in the fifth growth cycle the AANU was significantly higher and with BGR 1 it
ranged from 7% in soil 3 to 39% in soil 4, with BGR 6 it ranged from 24% in soil 3 to
59% in soil 4 and with BGR8 it ranged from 52% in soil 3 to 72% from soil 1. These
differences between the fertilizers reflect the different amount of organic N input as a
consequence of the fertilizer application based on equal amounts of ammonium-N.
With one exception i.e., BGR 1 in soil 3 AANU of all fertilizers in all soils increased
from the first to the fifth growth cycle.
50 Results
Table 10 F-statistics of two-way ANOVA of the effects of soils (1, 2, 3, 4 and 5)

and fertilizer sources (BGR1, BGR6 and BGR8)
ANOVA st nd th
1 growth cycle 2 growth cycle 5 growth cycle
main DF F significance DF F Significance DF F value Significance

effects+ value value
interactions
Soils 4 3.14 0.023* 4 3.37 0.017* 4 9.69 0.000***
Fertilizer
2 2.31 0.111NS 2 2.06 0.139 NS 2 39.48 0.000***
Soil ×
8 1.69 0.127NS 8 0.98 0.464 NS 8 1.16 0.341 NS
Fertilizer
Significant level: * P ≤ 0.05. * P ≤ 0.001. * P ≤ 0.0001
Results 51
Main factor
AB A AB B AB
soil
Main factor
a a a
fertilizers
Main factor
AB A AB B AB
soil
Main factor
a a a
fertilizers
52 Results
Main factor B B A B B
soil
Main factor a b c
fertilizers
Figure 15 Additional apparent N utilization of biogas residues collected in the

growth cycle 1, 2 and 5 in five soils different. ANOVA results present
the effect of fertilizers, soils and their interaction on the apparent N
utilization. Different lower case letters indicate statistically significant
differences between the fertilizers within one growth cycle (p  0.05).
Different capital letters indicate statistically significant differences between
the first, second and fifth growth cycle for one organic fertilizer (p  0.05)
Results 53
3.3.6 Soil organic matter content
After five repeated fertilizer applications soil total N contents of the fertilized treatments
increased as compared to the unfertilized control (Tab. 11). While the application of
NH4NO3 as a mineral fertilizer had only a small effect, the application of the biogas
residues significantly increased soil total nitrogen content.
Soil total N accumulation was higher after the application of the biogas residues with
BGR6 and BGR8 compared with BGR1. This is due to the different amounts of organic
N applied which were 2.6 or 2.9 times higher, respectively for BGR 6 and BGR8
compared with BGR1.
Table 11 Soil Nt content (g kg-1) at the end of the experiment
Treatments soil 1 soil 2 soil 3 soil 4 soil 5
a a a a a
Control 1.90 1.33 0.52 4.29 1.81
ab ab ab a a
Mineral 1.95 1.40 0.53 4.33 1.86
bc b b ab b
BGR1 2.05 1.50 0.68 4.46 1.95
c c c b c
BGR6 2.18 1.68 0.79 4.62 2.06
c c c ab c
BGR8 2.18 1.70 0.78 4.41 2.08
Different letters indicate statistically significant differences between the fertilizers (p 0.05)
3.3.7 Net N mineralization rate from different soils
After five repeated fertilizer applications the rate of net N mineralization from soil total
nitrogen (extrapolated to one year) was calculated for the ninth cut which is the
unfertilized cut this was done to assess differences in the N release affected by soils
or fertilizers. N mineralization rate in unfertilized soils ranged from 2 to 4 % of soil total
N (Fig. 16). Compared with the unfertilized soils mineralization rate after application of
biogas residues was significantly greater in all soils except in soil 4. The N
mineralization rate of soil N after biogas effluent application ranged between 3 to 7 %
of the soil total N with the highest mineralization rate in the sandy soil 3 followed by the
silty soils 5, 1 and 2 and the lowest rate occurred in the organic clayey soil 4. No
significant differences in N mineralization rate appeared between the different biogas
residues.
54 Results
Figure 16 Net N mineralization rate of fertilizers collected in the growth cycles 1,

2 and 5 in five different soils. Different letters indicate statistically
significant differences between the fertilizers (p  0.05)
Results 55
3.4 Chemical properties of solid and liquid biogas residues after separation
After separation of the biogas residues dry matter content was highly enriched in the
solid fraction compared to the liquid fraction (Tab. 12). Thus the liquid biogas residues
(LBRs) and the solid biogas residues (SBRs) dry matter content in the SBRs ranged
from 22.4 to 30.3% in fresh matter, the dry matter content in the LBRs ranged from 5.4
to 10.2% in FM. For most of the residues the pH value of the SBRs (pH range 7.7 to
8.8) was comparable or markedly higher than that of the liquid fraction which had pH
values from 7.7 to 8.2. The NH4-N content ranged from 0.27% to 0.47% in the LBRs
and from 0.20% to 0.62% in the SBRs. In five of the seven residues the NH4-N content
tended to be higher in the LBRs, in the other two residues 3 and 7, the NH4-N content
of the SBRs was higher than that of the LBRs. Regarding total N there was no
consistent difference between LBRs and SBRs. For example, in residues 1, 3 and 7
total N in the SBRs exceeded that of the LBRs whereas in residue 2 and 4 total N was
higher in the LBRs. For all liquid and solid biogas residues total N was between 0.51
and 1.08% of the fresh matter. The content in organic nitrogen ranged from 0.04 to
0.46% in fresh matter and was higher in the SBRs except for residues 2 and 4 where
the organic N content was lower in the SBRs compared to the LBRs. The SBRs were
characterized by higher organic carbon contents and higher C:N ratios compared to
the liquid fraction. The Corg:Norg ratio ranged from 6.9 to 9.8 for the LBRs and from 16.8
to 30.3 for the SBRs. The extraordinary higher Corg:Norg ratio of LBR7 (41.5) derived
from its exceptionally low organic N content. Both P and K contents varied between
the SBRs and the LBRs. Phosphorus content in the LBRs was between 0.06-0.22%
and in the SBRs was between 0.13-1.20% being higher in the SBRs than in the LBRs.
Potassium content in LBRs was between 0.45- 0.91% and in the SBRS was between
0.36- 0.78 (data not shown), with rather higher contents in the LBRs in all cases. The
biogas residues were reanalyzed for total N and NH4-N prior to the fourth fertilizer
application (Tab. 13). Most of the biogas residues changed only slightly in NH4-N or
total N contents. However, based on this analysis SBR6 showed very low NH4-N
contents, most probably because of high NH3 losses during storage. Therefore, a new
sample of SBR6 from the same biogas digester was collected to replace the former.
56 Results
Table 12 Chemical composition of liquid and solid biogas residues obtained from different raw materials (% in FM)
used for the first, second and third growth cycles
Biogas
pH DM Nt Norg NH4-N Ct Corg Ct/Norg Ct/Nt Corg/Norg NH4-N/Nt
residues
LBR 1 7.7 6.32 0.56 0.26 0.30 2.5 2.3 9.4 4.4 8.6 0.53
LBR 2 7.7 9.11 0.76 0.42 0.34 3.5 3.2 8.3 4.6 7.6 0.44
LBR 3 8.1 5.39 0.54 0.23 0.31 2.0 1.6 8.6 3.7 6.9 0.57
LBR 4 8 7.63 0.63 0.35 0.28 3.0 2.7 8.6 4.8 7.8 0.44
LBR 5 7.8 10.15 0.76 0.35 0.41 3.7 3.5 10.6 4.9 9.8 0.53
LBR 6 7.8 6.14 0.53 0.27 0.27 2.3 2.2 8.9 4.4 8.2 0.50
LBR 7 8.2 7.57 0.51 0.04 0.47 2.3 1.8 53.6 4.5 41.5 0.92
SBR 1 7.7 26.63 0.58 0.35 0.23 10.3 10.2 29.6 17.9 29.4 0.40
SBR 2 8.8 23.55 0.63 0.36 0.27 9.8 9.7 27.1 15.6 26.7 0.43
SBR 3 8.8 23.64 0.76 0.40 0.36 9.5 9.5 24.1 12.6 23.9 0.48
SBR 4 8.4 24.65 0.58 0.34 0.24 10.4 10.3 30.6 17.8 30.3 0.42
SBR 5 8.8 30.3 0.75 0.42 0.33 11.6 11.4 27.9 15.6 27.4 0.44
SBR 6 8.8 22.43 0.53 0.33 0.20 9.4 9.3 28.8 17.8 28.5 0.38
SBR 7 7.8 29.85 1.08 0.46 0.62 8.3 7.7 18.2 7.7 16.8 0.57
Results 57
Table 13 Chemical composition of liquid and solid biogas residues obtained from different raw materials used for the
fourth and fifth growth cycles
Biogas residues* Nt Norg NH4-N Ct/Norg Ct/Nt Corg/Norg NH4-N/Nt

LBR 1 0.57 0.25 0.31 9.8 4.4 9.0 0.55
LBR 2 0.75 0.41 0.34 8.5 4.6 7.8 0.45
LBR 3 0.54 0.22 0.32 9.0 3.7 7.2 0.59
LBR 4 0.63 0.35 0.29 8.7 4.8 7.9 0.45
LBR 5 0.76 0.34 0.42 11.1 4.9 10.2 0.55
LBR 6 0.53 0.26 0.27 8.9 4.4 8.2 0.50
LBR 7 0.56 0.21 0.35 10.8 4.1 8.3 0.62
SBR 1 0.56 0.37 0.19 27.7 18.4 27.5 0.34
SBR 2 0.64 0.33 0.32 30.3 15.3 29.9 0.50
SBR 3 0.77 0.37 0.40 25.7 12.4 25.4 0.52
SBR 4 0.57 0.32 0.25 32.4 18.0 32.1 0.44
SBR 5 0.74 0.37 0.37 31.3 15.7 30.7 0.50
SBR 6 0.78 0.48 0.30 21.8 13.3 21.8 0.39

SBR 7 0.79 0.46 0.33 18.3 10.6 16.9 0.42
*New analysis was done before the fourth and fifth fertilization and a new source of SBR6 was applied
58 Results
3.5 Yield and N availability of ryegrass from liquid and solid biogas residues after
separation as tested in two soils
3.5.1 Shoot dry matter yield
Ryegrass dry matter yield increased after the application of the different biogas residues
(Fig. 17). In the first growth cycle, yield of the mineral fertilizer treatment and LBRs were
rather similar. Yield of the SBRs treatments were significantly lower than those of the
mineral fertilizer treatment in the Mühlfeld soil only. However yield of the SBRs
treatments were comparable to those of the mineral fertilizer in the Dürnast soil. In the
second growth cycle, the ryegrass yield was comparable between the mineral treatment
and the LBRs treatments in both soils except for LBR2, 4 and 7 which were significantly
higher than that of the mineral treatment in the Mühlfeld soil. Yield of the SBRs was
similar to the yield of the mineral treatment. Only SBR1 was significantly lower than that
of the mineral treatment. In the Dürnast soil, yield was similar in both the mineral and
LBR treatments, but was significantly lower for the SBR treatments than for the mineral
treatment. In the third growth cycle, yield from LBR 4, 5, 6 and 7 was significantly higher
than the mineral one in the Mühlfeld soil. In the Dürnast soil all LBRs resulted in higher
yield than given by the mineral treatment. Yield from the SBRs was comparable to of
the mineral treatment in both soils. In the fourth growth cycle, the ryegrass yield of the
LBR treatments was comparable or higher than that of the mineral treatment. However
yield of the SBRs was comparable to that of the mineral treatment. After five repeated
fertilizer application the highest yield was obtained from the LBR treatments in both
soils, while yield of the SBR treatments was similar to that of the mineral treatment.
The application of LBRs, SBRs and mineral fertilizers increased the shoot N content
(Fig. 18). It is obvious that the fertilized cut (cuts 1, 3, 5 and 7) contained higher N
content than the unfertilized ones (cuts 2, 4, 6 and 8). After the last fertilizer application,
the N content was similar from cut 8 and 9. However the highest shoot N content
occurred in cuts 5 and 7 with N contents of 3.7% and 3.0%, respectively, in the Mühlfeld
soil. and with 2.9% and 2.3%, respectively, in the Dürnast soil. The lowest N content
occurred in the cut 3 being 2% in the Mühlfeld soil and 1.8 in the Dürnast soil.
Results 59
60 Results
Figure 17 Shoot dry matter yield of liquid and solid biogas residues collected in
five growth cycles in two different soils. Different letters indicate
statistically significant differences between fertilizers (p 0.05)
Results 61
62 Results
Figure 18 N content of liquid and solid biogas residues collected in five growth
cycles in two different soils. Error bars represent standard deviation
Results 63
3.5.3 Shoot N offtake
Over all the five growth cycles the ryegrass N offtake in the two soils was generally
higher after application of liquid biogas residues (LBRs) compared to solid biogas
residues (SBRs) (Fig. 19). In the first growth cycle after one fertilizer application and in
both soils, shoot N offtake of LBRs was similar to that from the mineral treatment,
except LBR2 in Mühlfeld soil of the shoot N offtake which was significantly higher than
the mineral one. The N offtake for the SBRs was similar or lower compared with the
mineral treatment. In the second growth cycle the N offtake from the LBRs was
comparable to that of the mineral treatment in the Mühlfeld soil, but comparable or lower
than the mineral treatment in the Dürnast soil. The SBRs treatments released less N
compared with the mineral treatment in the Dürnast soil. However N release from SBRs
in Mühlfeld soil was comparable to the mineral treatment, only SBR1 released less N
compared to the mineral treatment in this soil. In the third growth cycle the N offtake
from all LBRs was significantly higher than from the mineral treatment in both soils. The
N offtake from SBRs was comparable or higher than that of the mineral treatment in
both soils. The N offtake of the SBR2 was higher than that of the mineral treatment in
the Mühlfeld soil; however N offtake from SBR2, 3 and 5 were higher than from the
mineral treatment in the Dürnast soil. In the fourth growth cycle the N offtake values of
all LBRs and SBRs treatments were similar to the N offtake of the mineral treatment.
Only N offtake from LBR7 was higher than from the mineral treatment in both soils. After
five fertilizer applications (fifth growth cycle), the N offtake for all LBRs treatments was
higher than for the mineral treatment in both soils, only LBR5 was comparable to the
mineral treatment in the Dürnast soil. Most of the SBRs provided N offtake similarly to
that of the mineral treatment in both soils but the N offtake of SBR1 was markedly lower
than that of the mineral treatment.
Generally, the ryegrass shoot N offtake showed that liquid biogas residues provide
available N at least similar to that of the mineral fertilizer in all growth cycles and in both
soils. N offtake of the various SBRs was in most cases not only significantly lower than
that of LBRs, but also sometimes lower compared with that of the mineral fertilizer
particularly after the first two fertilizer applications. In the course of the five fertilizer
applications it became more obvious that N availability from the SBRs was lower than
that from the LBRs.
64 Results
Results 65
Figure 19 Shoot N offtake of liquid and solid biogas residues collected in five
growth cycles in two different soils. Different letters indicate
statistically significant differences between fertilizers (p 0.05)
66 Results
3.5.4 Apparent N utilization
In the five growth cycles the ANU of the mineral fertilizer ranged between 40% and 80%
in Mühlfeld soil and between 60% and more than 80% in Dürnast soil (Fig. 20). In the
first growth cycle, the ANU in Mühlfeld soil from the liquid biogas residues was between
80-85%, in contrast to only 45-68% from the solid biogas residues. However in Dürnast
soil, the ANU from LBRs ranged from 85-100%, but from SBRs it ranged only from 55 to
80%. From both soils it is clear that LBR2 had the highest ANU i.e., an increase of 10%
compared to the mineral treatment. By contrast, SBR1 had the lowest ANU which
shows a reduction of 43% compared to the ANU of the mineral fertilizer.
In the second and the third growth cycle, the ANU of the liquid biogas residues ranged
between 65 – 85% in both soils, and the ANU from the solid biogas residues was
between 40 up to 60%.
In the fourth growth cycle, the highest ANU was observed from LBR7 (up to 100%) in
both soils. On the other hand, the lowest ANUs were observed from SBR5 and 6 in
Mühlfeld soil, while in Dürnast soil the lowest ANU with only 40% was found for SBR1.
In the fifth growth cycle, LBR7 reached the highest ANU with up to 110% in both soils.
For the SBRs the ANU from SBR2, 3 and 4 was similar but the lowest ANU of 18-20%
was observed for SBR1. It is interesting to notice that the ANU from SBRs, except SBR
1 was comparable to that of the mineral treatment in this growth cycle.
Generally from the first to the fifth application the ANU of the liquid biogas residues
seemed to increase when compared to the mineral fertilizer. Similarly however on a
distinctly lower level, the ANU of the solid biogas residues seemed to increase in the
course of five repeated fertilizer applications with most SBRs reaching the ANU level of
the mineral fertilizer in the fifth growth cycle.
Results 67
68 Results
Figure 20 Shoot N utilization of liquid and solid biogas residues collected in five
growth cycles in two different soils. Different letters indicate statistically
significant differences between the fertilizers (p 0.05)
Results 69
3.5.5 Additional apparent nitrogen utilization
The additional apparent nitrogen utilization (AANU) for all LBRs was increasing
significantly in the course of the five fertilizer applications (Fig. 21). This was most
obvious in the Mühlfeld soil, while in the Dürnast soil the increase in AANU was more
heterogenous for the different LBRs treatments. The AANU for the LBRs in the first
growth cycle ranged between -3 to 13% in the Mühlfeld soil, in the fifth growth cycle it
reached 70 to 130%. In this cycle the highest AANU occurred with the LBR 2 and 7.
Lowest AANU occurred in the LBRs 1, 3 and 5.
In the Dürnast soil the highest AANU after five fertilizer applications was also found with
LBR7, and the AANU of all other LBRs was significantly lower.
Generally, the increase in the AANU was lower in the Dürnast soil than in the Mühlfeld
soil. In contrast to the LBRs the pattern of the AANU for the SBRs is characterized by
mostly negative values indicating an immobilization of the mineral nitrogen in the SBRs
treatments. The AANU for the SBRs also increased from the first to the fifth growth
cycle in most SBRs in the Mühlfeld soil up to positive value of about 20%. After five
repeated fertilizers applications low and negative values for AANU were observed in
SBR1.
In the Dürnast soil no positive values for the AANU could be found. Even a tendency to
an increasing AANU by repeated fertilizer applications was rather missing, with the
exception of SBR4. In both soils SBR1 showed the lowest and continuously negative
values of AANU indicating severe immobilization of the mineral fertilizer nitrogen even
after repeated applications.
70 Results
AANU from liquid biogas residues

Mühlfeld soil
200
AANU first growth cycle AANU second growth cycle AANU third growth cycle C
180 AANU fourth growth cycle AANU fifth growth cycle c
160
B
C C
140 b
bc ab
120 C C B C
ab BC abc
100 a B BC ab abc
AANU%
B BC
ab ab ab
80 AB ab
AB A a A
A AB
60 ABB a a a AB a
a Aa
40 Ba Aa A A B
a A
a b A Aa
20 a ab a
ab ab
0
-20 A AA A
a aa a
-40
LBR1 LBR2 LBR3 LBR4 LBR5 LBR6 LBR7
AANU from liquid biogas residues

200
Dürnast soil
180
160
140
120 B
b
AANU%
100 B
b
80 A
ab B C
B C B
A ab a
60 ab ab A ab B
B A A a
AB AB
BC a A
40 a b AB b a AB AB a A
AB AB a a b A A
a A ab A abc
a A a a a
20 a ab A A A a A A
A ab
c c a abc bc
ab
0
-20
LBR1 LBR2 LBR3 LBR4 LBR5 LBR6 LBR7
Results 71
AANU from solid biogas residues

Mühlfeld soil
120
AANU first growth cycle AANU second growth cycle AANU third growth cycle
100
AANU fourth cycle AANU fifth growth cycle
80
B B C
A B a b B a
60 b
aA aB A
40 C A b b a A BC
AANU%
a b AB A ab b
20 B b b A
ab b
0
-20 AB
A A AB
c b AA b AA A AB A c AB
-40 A
c ab c ab A bc ab ab ab
-60 AB ab A
A a AB b a
-80 a a
SBR1 SBR2 SBR3 SBR4 SBR5 SBR6 SBR7
AANU from solid biogas residues

Dürnast soil
120
100
80
60
B
40 A a
AANU%
d A B
20 AB B A a A BC A A AB
a ab a ab
a ab a ab a
0
-20 A A A
A cd cd AB B A
cd d C ABC A
-40 b A A bcd cd A
A bcd bc c A
ab A bc A
-60 B bc A ab A AB abc A
AB a a bc bc a b
a ab
-80
SBR1 SBR2 SBR3 SBR4 SBR5 SBR6 SBR7
Figure 21 Additional apparent N utilization of liquid and solid biogas residues

collected in five growth cycles in two different soils. Different letters
indicate statistically significant differences between the fertilizers (p 0.05)
72 Results
3.5.6 N uptake (shoot, stubble and root) from selected treatments
3.5.6.1 N uptake from mineral treatments during five growth cycles
The N uptake from the mineral treatment generally was higher in shoots followed by
stubbles and the roots which showed the lowest N uptake in both soils (Fig. 22). In both
soils and from the different growth cycles, shoot N uptake ranged from 43 to 60 % of the
total N uptake, stubble N uptake ranged from 14 to 37% and root N uptake ranged from
12 to 27% of the total N uptake. Therefore in general, in both soils shoots took up about
55% from the total N uptake while in the stubbles 24% and in the roots about 21% of the
total N uptake was found. Although the absolute N uptake in mg/pot decreased from the
first to the fifth growth cycle, the proportion of shoots and stubbles did not significantly
change with time.
3.5.6.2 N uptake in the last growth cycle
The fertilized treatments had a greater plant N uptake (shoots, roots, stubbles) than the
unfertilized treatment (Fig. 23). At the end of the experiment the highest N uptake was
found in the shoots of LBR1 (61% for the Dürnast and 64% for the Mühlfeld soil of the
total N uptake), followed by the mineral treatment (59% for the Dürnast and 57% for the
Mühlfeld soil) and finally by SBR1 (29% for the Dürnast and 36% for the Mühlfeld soil)
which indicates a higher N immobilization from the solid compared to the liquid biogas
residues (Fig. 23). The highest amount of N was accumulated in the roots of SBR1
(45% of all N uptake for the Mühlfeld soil and 52% of total N uptake for the Dürnast
soil). In general, the amount of N in roots at the final growth cycle tended to be higher in
the LBR1 and SBR1 than in the mineral fertilizer treatment. With the application of
SBR1 the absolute N uptake of the roots as well as the proportion of root N in total N
uptake exceeded that of the shoots.
Results 73

cycle), stubbles and roots from the mineral treatment during five
growth cycles for two soils. Numbers above the columns represent the
relative distribution of N in shoots, stubbles and roots in each growth
cycle (sum of N in shoots + stubbles + roots = 100%)
74 Results
Figure 23 Partitioning of N uptake in shoots (all cuts in each growth cycle),

stubbles and roots from the control, mineral fertilizer, liquid biogas
residue 1 and solid biogas residue 1 in the last growth cycle for two
soils. Numbers above the columns represent the relative distribution
of N in shoots, stubbles and roots in each growth cycle (sum of N in
shoots + stubbles + roots = 100%)
Results 75
3.5.7 Relationship between N offtake of the liquid and the solid biogas residues
and their Corg:Norg ratio
The C:N ratio can be a good indicator for the N mineralization from organic fertilizers.
Tab. 14 shows the correlation between the N offtake and the Corg:Norg ratio for all liquid
and solid biogas residues. In both soils the negative correlation between the Corg:Norg
ratio and the N offtake of the ryegrass ranging from -0.76 to 0.93 was highly significant
(p≤ 0.01) and valid from the first to the fifth growth cycle. However analyzing single data
points makes clear that when the data set is subdivided into liquid (Corg:Norg 6.9 to 9.8)
and solid (Corg:Norg 16.8 to 30.3) biogas residues this correlation is lower for LBRs or
almost missing for SBRs (Fig. 24).
Table 14 Correlation coefficients between N offtake of the five growth cycles and
Corg:Norg ratio of the solid and the liquid biogas residues
Corg:Norg
N uptake
Mühlfeld soil Dürnast soil
st
1 growth cycle -0.883** -0.850**
2nd growth cycle -0.843** -0.847**
3th growth cycle -0.759** -0.756**
4th growth cycle -0.886** -0.931**
5th growth cycle -0.851** -0.810**
Significance level: * (5%) and ** (1%)
76 Results
a b
c d
Figure 24 Relationship between N offtake in the five growth cycles and Corg:Norg
ratio of solid and liquid biogas residues. The letters a,b,c,d,e represent
the growth cycles from the first to the fifth growth cycle
Results 77
3.5.8 Relationship between N offtake of liquid and solid biogas residues and the
amounts of Norg and Corg applied in five growth cycles
As the fertilizer application was based on equal amounts of NH 4-N, different amounts of
organic N and also organic C were applied (Tab. 15). When comparing LBRs and SBRs
the organic carbon application from SBRs exceeded that from LBRs by far whereas the
application of organic nitrogen was not much differing between LBRs and SBRs. With
some exceptions e.g., a significant positive correlation coefficient of 0.969** in the
second growth cycle in Mühlfeld soil, there was no significant correlation between the
amount of Norg or Corg applied and the N offtake of the ryegrass in the five growth cycles
although correlation coefficients were mostly negative for SBRs and positive for LBRs.
Table 15 Correlation coefficients between N offtake of five growth cycles and the
amounts of Corg and Norg applied as solid and liquid biogas residues in
two soils
Growth cycle Liquid Solid Liquid Solid
Norg Corg Norg Corg Norg Corg Norg Corg

Applied
amounts/ one
0.22-0.37 1.6-3.5 0.22-0.48 7.7-11.1 0.22-0.37 1.6-3.5 0.22-0.48 7.7-11.1
application
(g/pot)
First growth
0.628 NS 0.145 NS -0.516 NS -0.647 NS 0.509 NS 0.132 NS -0.183 NS -0.229 NS
cycle
Second growth
0.969** 0.407 NS -0.097 NS -0.318 NS 0.774 NS 0.338 NS -0.211 NS -0.259 NS
cycle
Third growth
0.579 NS 0.735 NS 0.310 NS 0.659 NS 0.421 NS 0.183 NS -0.270 NS -0.267 NS
cycle
Fourth growth
0.542 NS -0.603 NS -0.627 NS -0.574 NS 0.165 NS -0.603 NS -0.696 NS -0.574 NS
cycle
Fifth growth
0.346* 0.379 NS -0.166 NS -0.174 NS 0.884* 0.105 NS -0.229 NS 0.030 NS
cycle
Significance levels: *(5%); ** (1%); NS (not significant)
78 Results
3.5.9 Soil total N and total C at the end of the experiment
At the end of the experiment i.e., after the last cut and after five repeated applications of
different LBRs and SBRs, soil total nitrogen and total carbon contents were determined
(Tab. 16). The N contents in the soils changed compared to the start of the experiment
in all organic fertilizer treatments. Compared to the mineral fertilizer soil N contents after
the application of LBRs did not change significantly in both soils (except LBR2 in
Mühlfeld soil). In contrast the application of most of the SBRs significantly increased soil
N content compared with the mineral fertilizer treatment (except SBR 3 in both soils and
SBR2 in the Dürnast soil). Similar results were obtained for the soil C t contents. The
application of LBRs did not significantly change soil Ct contents compared to the
mineral treatment in both soils. By contrast soil C t contents in both soils were
significantly increased after five repeated applications of SBRs (except SBR3 in the
Mühlfeld soil).
Soils C:N ratio significantly increased by the application of SBRs compared to the
application of the mineral treatment in both soils except for SBR7, whereas the soil C:N
ratio remained largely unchanged after the applications of LBRs. In addition comparing
LBRs to the corresponding SBRs the soil C:N ratio was found to be higher with SBRs
except for SBR3 and SBR7 in the Mühlfeld soil and for SBR 2 and SBR7 in the Dürnast
soil.
Results 79
Table 16 Soil contents of total nitrogen and total carbon at the end of the
experiment
Treatments 1 2 3 4
Nt% Ct% C:N ratio Nt% Ct% C:N ratio
a a ab a a
Control 0.087 0.80 9.14 0.134 1.205 8.99a
ab ab a ab ab ab
Mineral 0.093 0.828 8.90 0.136 1.253 9.22
bcde abcd ab bcd abcde ab
LBR1 0.106 0.991 9.34 0.153 1.414 9.25
LBR2 0.109cdef 1.022bcde 9.37ab 0.152abc 1.422bcde 9.38abc
LBR3 0.100abc 0.947abc 9.85ab 0.151abc 1.399abcde 9.25ab
LBR4 0.106bcde 0.995abcd 9.41ab 0.156bcd 1.428bcdef 9.15ab
LBR5 0.103abc 0.956abc 9.28ab 0.153bcd 1.452bcdef 9.50bcde
LBR6 0.105bcde 0.992abcd 9.43ab 0.151abc 1.424bcde 9.43abcd
abc abc ab abc abc ab
LBR7 0.101 0.919 9.08 0.147 1.336 9.09
f g e cd fg f
SBR1 0.125 1.399 11.21 0.160 1.641 10.27
SBR2 0.115cdef 1.215efg 10.56de 0.155bcd 1.526cdef 9.85cdef
SBR3 0.103bcd 1.015bcde 9.86bcd 0.154bcd 1.524cdef 9.92def
SBR4 0.114cdef 1.182defg 10.36cde 0.162cd 1.609defg 9.96ef
SBR5 0.119ef 1.267fg 10.62de 0.167d 1.676g 10.02f
SBR6 0.117def 1.256efg 10.75e 0.159cd 1.626efg 10.20f
cdef cdef abc cd cdefg abcd
SBR7 0.110 1.052 9.56 0.157 1.485 9.43
Before the experiment 1Nt=0.09, 2Ct =0.80, 3Nt= 0.13 and 4Ct =1.09
Different letters indicate statistically significant differences between the fertilizers (p 0.05)
3.5.10 Relationship between N offtake of the liquid and the solid biogas residues
and soil Nt at the end of the experiment
Correlation analysis between N offtake in the last cut and soil N t at the end of the
experiment was done to study the long term effect of five repeated applications of LBRs
and SBRs (Fig. 25). A weak positive correlation between N offtake of the last cut and
soil Nt at the end of the experiment was found only in the liquid biogas residues in
Mühlfeld soil. If LBR7 that is characterized by a very low Norg is excluded from the
correlation analysis the correlation coefficient increases from 0.396 NS to 0.857*. A
weak negative correlation between N offtake of the last cut and soil Nt content at the
end of the experiment was found for the solid biogas residues in both soils. In contrast,
a significant correlation between the N offtake of the last cut and soil Nt at the end of the
experiment was observed in both soils when both liquid and solid biogas residues were
included (r =-0.665** in the Mühlfeld soil and r =-0.623* in the Dürnast soil).
80 Results

LBR7
Figure 25 Relationship between ryegrass N offtake in the last cut and soil Nt at
the end of the experiment after the applications of liquid and solid
biogas residues in two soils
Discussion 81
4. Discussion
4.1 N availability from different unseparated biogas residues
The present study investigates the “short term” effects of a single fertilizer application
of different biogas residues on the N offtake of ryegrass, as well as the “long term”
effects after five repeated applications (one application at the start of each of five
growth cycles being based on equal amounts of ammonium).
After the application of organic fertilizers the ammonium will become available to
plants, but part of it is immobilized in soil during microbial decomposition of organic
compounds in the cattle slurry (SØrensen and Jensen 1995a). In contrast after the
first growth cycle, the shoot N offtake of ryegrass from biogas residues and the
undigested CS was comparable to or higher than that from the inorganic N fertilizer
of ammonium nitrate (Fig. 7). Minimizing or avoiding ammonia volatilization by
immediate incorporation of the organic fertilizers and nitrate leaching by using closed
pots most likely accounts for this observation given that Matsunaka et al. (2006)
showed that more N had to be applied as anaerobically digested CS compared to
mineral fertilizer to achieve an equal N uptake because of considerable N losses
through nitrate leaching and ammonia volatilization from organic fertilizers. However,
our results are congruent with those of Gunnarsson et al. (2010), who found that N
uptake of ryegrass foliage and stubbles from the application of a biogas residue in a
pot experiment was equal to that from an inorganic fertilizer following one application
(i.e., after one growth cycle). Similar results were also obtained by de Boer (2008),
who compared the apparent N utilization from calcium ammonium nitrate with that
from pig slurry co-digested with different co-products. The higher N release from
organic fertilizers was particularly shown in the 2nd and 5th growth cycles (Fig. 9).
This indicates that the superiority of organic sources over the mineral source was
more marked with time. It must be pointed out that N application of the organic
sources as well as the mineral source was done on the basis of their content of
NH4+-N. Since the organic sources have additional N in the form of organic N as
compared to the mineral source, organic sources may provide more N to plants upon
its mineralization. Kirchmann and Lundvall (1993) reported that animal slurries
provide efficient slow release from N sources. Accumulation of N in the soil was
reported by Flowers and Arnold (1983) as a result of using organic N for fertilization.
82 Discussion
The apparent N utilization (ANU) from all fertilizers decreased with time (Fig. 8), and
the highest utilization occurred in the first growth cycle and the lowest was in the fifth
growth cycle. An increasing proportion of non-harvested stubbles may at least partly
account for this. In the first growth cycle of the present study, there was no
significant difference between digested cattle slurry with 14% of maize silage and
undigested cattle slurry (Fig. 7). Rubaek et al. (2001) found that digested slurry
resulted in a higher N uptake than given by undigested slurry, and attributed this to
the higher ammonia volatilization from the latter source.
The present study showed higher N offtake in the fifth growth cycle from CS than
from BGRs (BGR 2,7 and 8 in Marktschwaben soil and BGR7 and 11 in Dürnast
soil). This may be due to the higher C:N ratio of CS. Gutser et al. (2005) reported
that the residual effect of N increased with a decreasing short term N release from
organic fertilizers. Further soil microorganisms may have increased in the slurry
treatment immobilizing N in their bodies. Such biomass N would need some years to
mineralize again for crops (SØrensen and Amato 2002). These results agree with
results by Martyniuk et al. (2002) who concluded that the long term fertilizer
application for 17 years increased soil fertility more than mineral fertilizers. These
results are similar to those found by Rubaek et al (2001) who observed more
microbial immobilization of N in raw slurry than in digested slurry. On the other hand,
results obtained by Loria et al., (2007) showed no difference after a 3-year
experiment between raw and digested slurry.
4.2 Effect of unseparated biogas residues’ C:N ratios on N mineralization

The C:N ratio can be a reliable predictor for the N mineralization to distinguish
nitrogen-rich from nitrogen-poor materials. Thus the C:N ratio can be a useful tool for
the N availability. The N offtake of ryegrass after the first growth cycle was
significantly related to the composition of the biogas residues or the CS (Fig 10). In
both soils tested, a significant negative correlation was found between the C org:Norg
ratio of the organic fertilizers ranging from 8.6 to 18.6 and the N offtake of ryegrass.
Similar negative correlations between N uptake and the C org:Norg or C:N ratio of
organic fertilizers were also reported in other studies (Gutser et al. 1987; Chadwick
et al. 2000; SØrensen et al. 2003; de Boer 2008). Chadwick et al. (2000) found a
highly significant negative correlation between C:N org of organic manures and N
mineralization during the first 30 days of an incubation experiment, but such
Discussion 83
correlation was not significant after 45 days of incubation. Such a relation was not
found with the second and the fifth growth cycles in the present study (Tab. 7). Van
Kessel and Reeves (2002) studied correlations between N mineralization and
manures composition after a 56-day incubation experiment and obtained a highly
significant correlation between organic N mineralization and C:N t ratios of manures.
When co-fermented slurry is compared to undigested raw slurry, it was often found
that the C/N ratio decreased through the process of anaerobic digestion (Gutser et
al. 1987; Asmus et al. 1988; de Boer 2008; Möller et al. 2008), thereby enabling
increased plant N uptake. Similarly, in the present experiment, CS co-fermented
with 14% maize silage was characterized by a lower C org/Norg ratio compared to
undigested CS. However, the Corg/Norg ratio for the former is still relatively high
compared with the other tested biogas residues. This fact may be the reason why no
significant increase in either the N offtake or in the apparent N utilization was
observed in the first and second growth cycle between undigested and co-fermented
CS.
Although the biogas residues differed considerably in their ammonium N, N org and
Corg contents, no consistent relationship existed between the composition of the
biogas residues and the different fermentation substrates used for the digestion (i.e.,
whether the residues were either purely plant based or derived from co-fermentation
with animal excrements). Residues obtained from co-fermentation (BGR 1, 7 and 11)
contained both high and low C org/Norg ratios or NH4-N contents, with the chemical
composition of the 100% plant based residues being similar to those from co-
fermentation. It remains unclear, however, whether the N availability to ryegrass from
the biogas residues is related to the composition of the fermentation substrates used
for digestion. Although the lowest N offtake in both soils was obtained from the
application of a purely plant based residue (BGR2) and the highest N offtake from
one derived from co-fermentation (BGR11), the N offtake from another purely plant
based residue (BGR8) was not significantly different from the co-fermentation
residues BGR1 and 11.
84 Discussion
4.3 N utilization from different unseparated biogas residues

The apparent N utilization (ANU) after the first growth cycle ranged from about 80%
for the mineral fertilizer to about 100% for some biogas residues (e.g., BGR11). This
additional ANU of up to 20% compared to NH4NO3 indicates a significant
mineralization of organic N even after a single fertilizer application (Fig 8). No
statistically significant net N immobilization was found for either biogas residues or
for CS. This latter result seems to contrast with observations by Sorensen and
Jensen (1995a) and Flowers and Arnold (1983), who both report a net N
immobilization of 24 to 40% compared to ammonium sulfate when cattle or pig slurry
were mixed into the soil. However, such high net N immobilization can be explained
by the comparatively high C/Norg ratio of the CS used in their experiment (about 30
compared to 18.6 here).
The additional apparent N utilization (AANU) from some of the organic fertilizers as
compared to the mineral fertilizer increased significantly after repeated fertilizer
application (Fig 9). Such increasing N releases after regular application of organic
fertilizers with high proportions of organic N has been reviewed by Gutser et al.
(2005). Organic N from organic fertilizers acts mainly via the soil N pool because of
the slow release characteristics of the organically bound N. Residual effects
expressed as an increase in plant N uptake from N org after long-term application of
animal slurry are well described (Martynuik et al. 2002; Schröder et al. 2005; Angers
et al. 2010) and are often associated with an observed increase in soil N t.
4.4 Soil N accumulation after five repeated applications of different

unseparated biogas residues
Mineral N content in the soil at the end of the experiment reflects the balance
between the processes of addition, transformation and loss in the soil. Removal of N
from soil occurs mainly by N uptake by the plants. The N remaining in the soil which
received a mineral N source would be expected to be less than that which received
an organic source. Most of the N remains in the soil at the end of the experiment
adding to the long term pool of soil organic N (Morvan et al. 1997).
In the present study, both an accumulation of soil N t and an increased additional
ANU were observed after five fertilizer applications over the course of nearly one
year. This result may be due to the intensive soil-fertilization-crop system used as a
model here, which seems to accelerate such processes.
Discussion 85
Moreover, because all fertilizer applications in this experiment were based on an

equal amount of ammonium, the amount of N org applied differed considerably among
the organic fertilizers resulting in different N accumulations in the soil at the end of
the experiment (Tab. 6). Consequently, N offtake by ryegrass in the fifth growth cycle
and particularly that of the last biomass sampling, which received no direct fertilizer
application, were closely related to the soil N content. In contrast to the effect
observed after the first growth cycle, no correlation between N offtake and the
Corg/Norg ratio of the organic fertilizers was found after repeated fertilizer application,
possibly indicating that agricultural soils that are regularly supplied with organic
fertilizers might react less to different qualities of organic fertilizers over the long
term. Dick (1992) noted a positive relationship between soil organic matter content
and soil microbial biomass, and concluded that any practice that increases organic
matter in soil increases the biological activity in the soil. Organic and inorganic input
of N, when combined with appropriate management, increases the amount of
residue returning to the soil, and thus increases the soil biological activity.
Concerning the C content, the highest was that within the cattle slurry treatment. The
importance of N-mineralization in determining N uptake from manure is dependent
on the amount of organic N initially present as well as the potential for mineralization
(Douglas and Magdoff 1991). Many animal manures, particularly poultry manure and
slurry, contain considerable proportion of their N in soluble inorganic forms
(Beauchamp 1983) with as much as up to 80% of the total N. However in some
cases a considerable proportion may be lost by volatilization from such sources
(Beauchamp 1986).
4.5 N mineralization from different soils
4.5.1 Effect of soil type and organic matter content on N release from
unfertilized and fertilized soils
The ammonium present in organic fertilizers may be immobilized into the microbial
biomass. If the immobilized N is not released for plants during the season its
availability will be decreased (Paul and Beauchamp 1994). Immobilized N may
remain in the soil organic matter in forms recalcitrant to decomposition (SØrensen
2004). However the increase in the rate of N transformation does not necessarily
affect the net rate of N mineralization or immobilization. Mineral N is continuously
released and mineralized from the soil organic matter pool or organic fertilizers but
86 Discussion
predicting the N mineralization from organic N is difficult because it is affected by

several factors such as soil temperature, soil water content, soil texture and soil
organic matter content. Because mineralization is a biological process, it only occurs
when soil conditions are suitable for biological activity. Soil type can affect the
dynamic processes of soil organic matter build up as well as the dynamics of N
mineralization. Moreover, N transformation depends on soil type and organic matter
content. Therefore the N offtake in soil supplied with organic fertilization as compared
with that in soils not supplied gives a relative indicator to interpret the decomposition
of the organic fraction in soil. Hence in the present study the N offtake by the control
soils (without fertilizer application) was greatest in soil 4 (organic soil). Soils 1, 2 and
5 (silt loam soils) mineralized intermediate amounts of N and soil 3 (sandy soil)
mineralized the least N (Fig. 14). The N offtake of plants from unfertilized soils can be
used as an indicator to describe the N mineralization of organic nitrogen in soils. The
N offtake in unfertilized soils which did not receive fertilizer ranged from 40 to 340 mg
N/pot and varied among the soils (Fig. 11). In all growth cycles the highest N offtake
was observed in the organic soil followed by silt loam soils. The sandy soil showed
the lowest N offtake when unfertilized. The N offtake from different soils was
associated with the soil total N content which was highest in the organic soil and
lowest in the sandy soil. These results agree with those reported by Egelkraut et al.
(2000) who found that in the untreated soils, the sandy loam and loamy sand soils
high in clay content showed a high N uptake as compared with the sandy soil.
Accordingly, the highest N offtake would be in the organic soil which contained 0.45 g
Nt/kg soil and the lowest N offtake was in the sandy soil which contained 0.15 g Nt/kg
soil. Thomsen et al. (2001) showed that increasing the soil clay content from 10% to
40% leads to a decrease in the net 15N mineralization of the ryegrass.
The additional apparent nitrogen utilization (AANU) can predict the increasing net N
mineralization of the BGRs (Fig. 15). Results show no effect obtained from different
soil types on the N mineralization from the biogas residues. These results disagree
with results obtained by Amato and Ladd (1992); Hassink (1992); Gordillo and
Carbrere (1997) who studied the N mineralization in different soils and found that N
mineralization was related to sand content. They explained these results due to soil
aeration being suitable for the microbial activates.
SØrensen and Jensen (1995b) found that N uptake by grass in unfertilized soils was
increased by increasing the soil clay content. They attributed this to the fine soil
Discussion 87
particles causing reduction of decomposition of organic residues through a protection

mechanism. Bossta and Ågren (1997) reviewed that N mineralization from the soil
was related to clay content. Hassink et al. (1994); SØrensen and Jensen (1995);
Griffin et al. (2002) reported that the decomposition rate of organic residues in the
soil is lower in fine-textured soils than in coarse-textured soils.
The N release in our present study from the five unfertilized soils is obviously related
to its total N and C, in all growth cycles. These results agree with those obtained by
Vellinga and André (1999) who studied soil nitrogen supply from peat, clay and sand
soils and observed a higher soil nitrogen supply from the peat soil than from the clay
or the sand soil. They attributed the high soil nitrogen supply from peat soil to its high
organic matter compared with the clay soil or the sand soil. On the other hand
Whitehead (1984) did not observe such relationship between the N mineralization in
soil and its organic matter content. The present study shows that the additional N
offtake from the fertilizers differed between soils in each growth cycle but that it was
not related to soil organic matter content or soil texture. This result disagrees with the
results of Legg and Stanford (1967) who examined the N uptake from 12 soils
differing in their organic matter content and C/N ratio for 8 weeks and reported that
soils which were higher in total carbon content (1.9-3.9%) and total N (0.15-0.34%)
showed a higher N uptake compared to soils with lower total C (0.53-1.58%) and
total N (0.041-0.083%) contents.
4.5.2 Effect of organic fertilizers on N mineralization in different soils
For all growth cycles, there were significant effects of both soils and fertilizers on the
N offtake of ryegrass but there were no significant interactions between soils and
fertilizers (Table 9). This result agrees with the result obtained by Chantigny et al.
(2008) who found a significant effect of different organic fertilizers on N uptake in the
plant grown in loam and clay soils.
The effect of the soil, averaged over all fertilizers was inconsistent and changed from
one growth cycle to the other. The lowest additional N offtake in the first growth cycle
was observed in soil 2 (silt loam) whereas in the second growth cycle this was true
for soil 4 (organic soil) and in the fifth growth cycle it occurred in soil 1 (silt loam) (Fig.
14).
In each growth cycle, the average additional N offtake from the biogas residues
fertilizers, was significantly higher than from the mineral fertilizer and seemed to
88 Discussion
increase with time. In the first and second growth cycles no differences appeared
between the biogas sources. In the fifth growth cycle the N offtake was significantly
greater from biogas residue BGR1 to BGR6 and BGR8. The N mineralization from
the three BGRs (BGR1, 6 and 8) differed. BGR 6 and 8 gave higher shoot additional
N offtake particularly in the fifth growth cycle. This reflects the higher organic N
applied with BGR6 and BGR8 (22 and 25 g Norg/pot) compared to BGR1 (6 g
Norg/pot). Based on an equal amount of N applied N mineralization will depend on the
N content of the material. For example, Egelkraut et al. (2000) reported a higher net
mineralization rate from cotton leaves of 28.3 g Norg/kg compared to cotton stems of
15.4 g Norg/kg. SØrensen and Jensen (1995b) found that N mineralization was higher
from an-aerobically stored sheep manure of 33.6 g Norg/kg than from fresh manure of
31.7 g Norg/kg. Whitemore and Groot (1997) found that the N availability was strongly
dependent upon the amount of N added in the crop residues since spinach with C:N
ratio of 6 released more N compared with other plant residues having C:N ratios of
13 to 15 and concluded that sandy soils mineralize N earlier than the silt loam soils.
From the present study it is concluded that the soil and fertilizers represented
important factors for N mineralization but no interaction was obtained between soil
and fertilizers.
4.5.3. Soil N accumulation at the end of the experiment from different soils
After five repeated fertilizer applications soil total N contents of fertilized treatments
were higher as compared with the unfertilized treatment (Tab. 11). While application
of NH4NO3 as a mineral fertilizer had little effect, application of biogas residues had
greater effect. Soil total N accumulation was higher after the application of BGR 6
and 8 compared with BGR 1. This is associated with the difference in amounts of
organic N applied which were 2.6 or 2.9 times higher for BGR 6 and 8, respectively,
compared with BGR 1. At the end of the experiment, the soils varied in N
accumulation. The N accumulation in the soil seemed to be associated more to the
silt content which was high in soil 1 (of 73% silt) followed by soil 5 (of 21% of silt) and
was lowest in soil 3 (of 8% silt). This result agrees with the finding by Hassink et al.
(1994). Higher N accumulation was obtained in soils treated with organic fertilizers
than in those treated with mineral fertilizers. SØrensen and Jensen (1995b) attributed
the higher N mineralization in manure-treated soils compared to the mineral fertilizer
treated ones to the mineralizable organic N in manure. SØrensen and Amato (2008)
Discussion 89
found that the N accumulation of barley crops from the injection of pig slurry in the
first year was comparable from a loamy sand soil having 1.4 g total N/kg soil and a
sandy loam soil having 1.6 g total N/kg. They attributed that to the lower clay fixation
of N in the loamy sand soil due to its lower clay content. They concluded that soils of
high clay content have a higher capacity for physical protection. Herlihy (1979) found
that N accumulation in the soil depends on soil texture; higher N accumulation was
obtained in a loam soil and lower accumulation occurred in a loamy sand soil.
4.5.4 Net mineralization of soil organic matter from different soils
Results show that the highest net N mineralization rate/year occurred in the soil 3
(sandy soil) and the lowest was in soil 4 (clay soil). Net N mineralization of soil
organic matter was found to be more rapid in sandy soils than in clayey soils
(SØrensen 1981; Hassink et al. 1993). Giardina et al. (2001) found in a short term
litter decomposition study lasting 2 years that soil N transformation was little related
to soil clay content. Soil aeration would enhance microbial and microfauna
populations present in this soil as well as cause a physical protection of the soil
organic matter (Hassink et al.1993; Hassink et al.1994). Barker and Pilbeam (2007)
reported that the turnover of humus N could be about 1 to 3% of the soil N depending
on the type of the soil, climate and other factors.
4.6 Separated biogas residues (liquid and solid)

4.6.1 N availability from liquid and solid biogas residues
Liquid biogas residues applied to the ryegrass increased the shoot N offtake from the
first until the fifth growth cycle compared to that of the solid biogas residues (Fig. 18).
This result is in agreement with findings reported by Manfredini et al. (2010) who
found more plant available nitrogen from the liquid digested pig manure compared to
that from the solid digested one from the 10th until the 75th day of the incubation
period.
The liquid biogas residues which were used in the present study were characterized
by a low C:N ratio (6.9 to 9.8). Persson and Kirchmann (1994) concluded that the
decomposing process in arable soils are favoured by a C:N ratio of about 10 and by
suitable aeration, sufficient nutrient status and suitable pH. Gutser et al. (2005) noted
that a high N release can be expected from organic fertilizers with a C:Norg ratio
below 6 to 7 while organic fertilizers with a high C:N ratio lead to immobilization. Qian
90 Discussion
and Schoenau (2002) studied the N release from different solid manures differing in
their C/Norg ratio and found that the net N mineralization was associated with a low
C:Norg leading to a high N uptake from the animal manures with high organic N
contents and the low C:N ratios of about 7.6 to 6.6. In the present study the N offtake
was comparable between the liquid biogas residues and mineral fertilizer in the first
growth cycle. These results are similar to those found by Chantigny et al. (2008) who
reported that the N offtake by plants after one fertilizer application from separated
liquid swine manure was similar to that from mineral fertilizer, while the N offtake by
plants from solid biogas residues was less compared to the mineral treatment.
However (using labelled N) SØrensen and Thomsen (2005) showed a higher
recovery of N from the mineral (55%) than from separated liquid pig slurry (51%)
after one fertilizer application, while the recovery of N from solid separated pig slurry
was 26.5%. After 3 years recovery of N dropped to 3.5% for separated solid pig slurry
followed by 2.3% for liquid separated pig slurry and 1.7% for the mineral fertilizer.
In the present study the long term effect was studied after four and five applications.
After five applications the N offtake from the liquid biogas residues exceeded that
from the mineral fertilizer by a factor of about 2, indicating that about all the organic N
applied in addition to the fertilizers’ ammonium might have become plant available
(approx. the NH4-N/Nt ratio is 50% see Tab. 12 and 13). These results agree with the
results reported by Chantigny et al. (2008) who found that after annual applications
for three years based on total N, the N export in grains was similar from the liquid
swine manures as compared to the mineral fertilizer. They attributed this to the
intermediate incorporation of the digested and liquid swine manure into the soil
resulting in a decreased NH3 volatilization. Chantigny et al. (2007) studied the N
uptake of imothy grass from inorganic fertilizer, raw swine slurry, digested swine
slurry and the liquid fraction of separated swine slurry using two different soils, a
loam and sandy loam. They reported that after 3 years of annual fertilizer application,
the N uptake was higher from the fertilized treatments and that N uptake was higher
from the digested as well as from the separated slurry than from the raw swine slurry.
Moreover, N uptake in the loam was higher compared to the sandy loam, a fact that
is in agreement with the results obtained here as well.
To achieve comparable N offtake from mineral and solid biogas residues five
repeated applications were needed.
Discussion 91
In the five growth cycles of the present study the apparent N utilization (ANU) of the
mineral fertilizer ranged from 40% to 80% in the Mühlfeld soil and between 60% to
more than 80% in the Dürnast soil (Figure 20). In the first growth cycle, the ANU in
the Mühlfeld soil from the liquid biogas residues was between 80 and 85% but it
increased up to 100% in the fifth growth cycle from both soils. Meade et al (2011)
found that based on total N the nitrogen uptake efficiency (NUE) of an inorganic N
fertilizer was significantly higher (67%) compared with that of the liquid pig manure
(33%) after two fertilizer applications during two years. Sørensen and Amato (2002)
attributed the lower pig manure NUE to the immobilization of NH 4-N during microbial
decomposition of the organic matter in the manure after application to the soil. They
also reported that a higher proportion of applied N remained in the soil system.
SØrensen and Jensen (1996) found that the N recovery from a sandy loam soil was
62% from urine and 78% from urea, but was between 51 and 53% from urine and
urea fertilizers in a sandy soil after 6 months from grass sowing, with high N
mineralization in the sandy soil.
Additional apparent nitrogen utilization (AANU) for all liquid biogas residues (LBRs)
increased significantly along the course of the five fertilizer applications (Fig. 21). The
increase was most obvious in the Mühlfeld soil, while in the Dürnast soil the increase
was lower. The AANU from the LBRs in the first growth cycle ranged between -3 to
13% in the Mühlfeld soil and reached 70 up to 130% in the fifth growth cycle. Higher
N uptake occurred in the Mühlfeld soil than in the Dürnast soil, and this reflects the
difference between the properties of the two soils. This denotes a difference in
cycling and turnover of organic nitrogen through the process of immobilization–
mineralization, soil aeration and the suitable water status enhance the microbial
activity (Hassink et al., 1993). The AANU from the solid biogas residues was up to -
42% in both soils. These results agree with results by Laboski et al. (2010) who
found that plant available nitrogen after applying digested separated liquid cattle
manure was 52.2%, but from the solid digested separated manure it was -14.6%.
They stated that the negative values of available nitrogen from separated solid
manures indicate high N immobilization. In the present study the N availability
increased over time especially from liquid as well as solid biogas residues. The
current results are also in good agreement with those by Manfredini et al (2010) who
found that after 80 days of incubation available N was higher from digested liquid pig
manure (60%) than that from digested solid pig manure (-22%). Results are also in
92 Discussion
agreement with those by Pereira et al. (2010) who found that mineralization
/immobilization efficiency from the liquid fraction and the solid fraction after the
separation was -9.7 and -35.6 mg N/kg, respectively, after 93 days of incubation in a
silt loam soil.
High correlations between N offtake and C:N ratio were observed in both soils from
the first to the fifth growth cycle. In the first growth cycle in the Mühlfeld soil a highly
significant negative correlation (r = -0.883**) occurred, a similarly significant one (r =
-0.850**) was found in the Dürnast soil (Tab. 14). Therefore N mineralization from
both liquid and solid residues was related to the C:N ratio with the lowest one of 6.9
for a liquid residue and the highest one of 30.5 for a solid residue. Solid biogas
residues had higher C:N ratio and lower N mineralization, while liquid had lower C:N
ratio and higher N mineralization (Fig. 24). The C:N ratio may not be an adequate
parameter for mineralizable N. In the third and the fourth growth cycles for the liquid
biogas residues there was a significant correlation with r = - 0.862* in the third growth
cycle and with r = - 0.910* in the fourth growth cycle. This result conflicts with the
result found by Qian and Schoenau (2002) who studied the relationship between N
uptake and available N supply over time from 11 solid manures of different C:N ratios
(6 to 21). They found that the relationship increased over time (67days) in a loam and
a sandy loam soils. For the group of the solid biogas residues alone, the Corg:Norg
ratios of 16.8 up to 30.3 were not sufficient to explain the N mineralization from the
solid biogas residues used in the present experiment. The Corg:Norg ratio of the liquid
biogas residues of 6.9 to 9.8 is also not adequate to explaine the N mineralization
from the liquid biogas residues. Thus the Corg:Norg ratio differs between the liquid and
the solid biogas residues, but not among the liquid biogas residues or among the
solid biogas residues (Fig. 24). Organic residues with similar Corg:Norg ratio may
mineralize different amounts of N due to the differences in their composition that are
not explained by the Corg:Norg ratio. Wood et al. (2010) evaluated 87 different poultry
wastes in an incubation experiment and found a significant negative correlation
between N mineralization and lignin content and a positive correlation between N
mineralization and hemicellulose content. Nourbakhsh (2006) observed that after 46
weeks of incubation using different plant materials there was a highly significant
positive correlation between the N content and N mineralization.
Cabrera et al. (2005) summarizing factors that control the net N mineralization of
organic residues concluded from studies by Withmore (1996) that a range in C:N
Discussion 93
ratios of 20 to 40 in the organic material represents the break-even point between net
N mineralization and net N immobilization. In some cases, this break-even point was
found to be near a C:N ratio of 15 (Gilmour, 1998). It was concluded by Laboski et al.
(2010) that manure separation was effective for N utilization with or without anaerobic
digestion.
4.6.2 Accumulation of N in shoot, stubble and root at the biomass harvest
The fertilizer treatments increased the N uptake by roots. Gunnarsson et al. (2010)
mentioned that N fertilization led to decreased organic matter as well as decreased
root respiration. The amount of N in roots at the final cut tended to be higher in the
solid biogas residues than the liquid one and the mineral fertilizer (Fig. 23). These
results are in agreement with the results by Gunnarsson et al. (2010) who found that
N recovery in ryegrass roots after 172 days tended to be higher with biogas effluents
than with inorganic sources. They noted the direct impact on C and N status.
SØrensen and Thomsen (2005) found that N recovery by the barley stubbles + roots
after 3 years application was higher from the separated solid pig slurry (0.5%)
followed by the separated liquid one (0.3%) and the lowest was obtained from the
mineral fertilizer (0.2%).
4.6.3 Soil total N after ryegrass harvest at the end of the experiment
Soil total N after the last harvest of the ryegrass was significantly greater in fertilized
treatments of different types of biogas residues (solid and liquid fraction) applied
during one year compared to the unfertilized treatment. A high accumulation of N in
the soil after the harvest from the solid biogas residues compared to the liquid biogas
residues was observed (Tab. 16). Gunnarsson et al. (2010) reported an increase in
soil mineral nitrogen after 172 days after the application of biogas effluent compared
to the mineral treatment. They concluded that based on an equal amount of N
applied the soil delivery capacity increased when the N supply source was changed
from inorganic fertilizer to biogas residues. SØrensen and Thomsen et al. (2005)
found a high N accumulation in the soil 0-20 cm top layer after three years of
applying separated solid pig slurry compared to the separated liquid pig slurry and
mineral fertilizer. The present results also agree with the results reported by
Chantingny et al. (2007) who found higher N accumulation after 3 years applying
94 Discussion
separated liquid swine manure compared to mineral fertilizers, and more N

accumulated in a loam soil than a sandy loam soil. Higher N accumulation in the soil
at the end of the experiment occurred in the Dürnast soil (silt loam) than in the
Mühlfeld (sandy soil). This result also agrees with those found by Qian and
Schoenau (2001) who reported higher available N supply in a clay loam soil than in a
sandy loam soil, and that N mineralization in coarse textured soils was more rapid
than in the fine textured soil (Hassink et al.1994).
Bertora et al. (2008) reported a higher N accumulation in the soil after 85 days of
incubation was obtained from urea and the liquid fraction of pig slurry than from the
solid fraction of pig slurry (138 mg NO3-N/kg soil). They concluded that the amount of
soil nitrate was strongly affected by the type of the slurry.
Conclusions 95
5. Conclusions
The unseparated biogas residues derived from different biogas plants fed with
different substrates for fermentation varied extremely in their chemical composition. A
similar high variability in the chemical composition was observed for liquid as well as
solid biogas residues derived from different digesters after separation. Further
research will be needed to elucidate the possible relationship between the
composition of biogas residues and the composition of substrates as well as the
efficiency of the fermentation process.
Based on the same amount of NH4+-N application, the unseparated biogas residues
and the separated liquid biogas residues provided N at least corresponding to their
NH4-N contents particularly after one time fertilizer application. However, the
separated solid biogas residues provided similar or less N than it would be expected
from their NH4-N contents. After one fertilizer application the Corg:Norg ratio of
unseparated and separated biogas residues, including both the liquid and solid
fractions, was a decisive factor for the short term N availability to ryegrass. Whereas
the Corg:Norg ratio revealed to an unsatisfactory parameter for the short term N
availability from liquid biogas residues or solid biogas residues alone. After repeated
applications, a positive correlation between soil Nt content and plant N offtake from
unseparated biogas residues indicated that the accumulation of Norg from the organic
fertilizers leads an increased release of N from the soil N pool. In contrast, the
increased soil Nt after the repeated application of solid biogas residues did not result
in an increased soil N mineralization whereas the soil C:N ratio was increased
instead. The increased additional apparent N utilization (AANU) from the first growth
cycle to the fifth growth cycle found for all unseparated biogas residues and all
separated liquid fractions, indicated a considerable N mineralization from the organic
N of these fertilizers. However, the negative AANU from the solid biogas residues
demonstrates their potential for N immobilization. Consequently, it can be suggested
that the continuous application of biogas residues leads to an increase in soil organic
matter and soil total N which can possibly be mineralized by time. Such increased N
mineralization will increase soil fertility and plant available N but also the risk of N
losses by denitrification and nitrate leaching. In addition, for an efficient use of biogas
residues application techniques, such as effective incorporation by injection, needs to
be improved in order to avoid N losses by NH3 volatilization.
96 Summary
6. Summary
The residues of the anaerobic digestion process contain valuable nutrients (N, P, K,
S, etc.) which need to be recycled in crop production in order to fulfil the aims of a
sustainable and environmentally efficient biogas production. However, biogas
residues are obtained from different substrates used for digestion, being either
exclusively plant based or derived from co-fermentation with animal excrements, a
fact which may lead to differences in their composition and possibly also in nutrient
availability. In addition physical separation of the biogas residues into a solid and a
liquid fraction may change their composition which may affect the nutrient efficiency
from both of them.
The objectives of this study were to

 characterize biogas residues either unseparated or separated into liquid and solid
phases from the fermentation of different raw materials with respect to their N and
C content,
 investigate short and long term effects of the application of these biogas residues
on the N availability and N utilization by ryegrass
 evaluate the role of different soils on the N utilization of these biogas residues.
To reach these aims, as a test system two pot experiments each lasting for about
one year were conducted with perennial ryegrass as a model plant in being regularly
harvested. Short and long term effects on N offtake were investigated after single
and repeated fertilizer application. In the first experiment, seven biogas residues
obtained from the digestion of different raw materials and one undigested cattle slurry
were incorporated into the soil based on an equal amount of NH 4-N (300 mg NH4-
N/pot*fertilization). In order to investigate the effect of soils on ryegrass N utilization,
a number of soils with different soil texture (two silt loam soils, one clay loam soil,
one sandy soil and one organic soil) were used. In the second experiment, biogas
residues from seven different biogas plants separated each into a liquid and a solid
fraction were used. The different biogas residues (liquid and solid fractions) were
applied to two soils (one sandy and one silt loam soil) based on an equal amount of
NH4-N (300 mg NH4-N/pot*fertilization). In both experiments, the grass was grown in
five growth cycles each starting with a fertilizer application. Within each growth cycle
the grass was cut two up to three times.
Summary 97
The following results were obtained:

(1) Based on an equal amount of NH4-N applied, the N offtake of ryegrass from the
unseparated biogas residues and cattle slurry was comparable or higher than
that from NH4NO3. However, already after one fertilizer application there were
significant differences in the N offtake of up to about 20% between the biogas
residues. In the 1st growth cycle the N offtake of ryegrass was significantly
related to the Corg/Norg ratio of the organic fertilizers. The additional crop N offtake
from the organic fertilizers as compared to NH 4NO3 increased from the 1st to the
5th growth cycle. At the end of the experiment, after five subsequent fertilizer
applications the N offtake of ryegrass was positively correlated to the soil organic
matter content which increased in response to the different amounts of organic N
applied.
(2) The influence of soil type was investigated with five soils and three different
biogas residues. The N offtake from the unfertilized five soils in all growth cycles
was affected strongly by the soil Nt content. Consequently after five repeated
applications, the highest N offtake was found in soil 4 with the highest organic
matter content (4.4 g Nt/kg soil), but the lowest was observed with soil 3 with the
lowest organic matter content (0.6 g Nt/kg soil). The additional nitrogen offtake
from the fertilizers differed between soils in each growth cycle but with no
consistent relationship to soil organic matter content or soil texture being
observed. Moreover there was no interaction between soils and fertilizers. Soil
net N mineralization rate of the accumulated organic matter at the end of the
experiment did not differ between biogas residues.
(3) Based on the same amount of NH4+-N, after one fertilizer application shoot N
offtake from the LBRs was comparable to the inorganic fertilizer in both soils. In
contrast shoot N offtake from SBRs was comparable or lower than that from the
inorganic fertilizer in both soils. Including LBRs and SBRs there was a significant
negative correlation between the Corg:Norg ratio of the biogas residues and the N
offtake of ryegrass after one as well as after the following four successive
fertilizer applications. However, within the respective groups of LBRs or SBRs the
Corg:Norg ratio was not a reliable parameter to predict ryegrass N offtake.
The additional apparent N utilization (AANU) describing the N release from the
organic N fraction of the organic fertilizers increased with time from the first to the
fifth growth cycle for all biogas residues.
98 Summary
However, while the positive values for the AANU of the LBRs indicated a
significant mineralization of fertilizer organic N, the AANU of the SBRs were
mostly negative, suggesting a considerable N immobilization of the applied
ammonium.
After five repeated fertilizer applications, soil Nt and Ct contents increased after
the application of SBRs but not of LBRs. However, this increase in soil organic
matter content did not result in any increase in plant available N.
It is concluded that unseparated biogas residues from both 100% plant based
substrates and from co-fermentation with animal excrements provide N at least
corresponding to their ammonium content and that after a first fertilizer application
the Corg:Norg ratio of the biogas residues was a crucial factor for the N availability.
After repeated i.e., long term application the organic N accumulated in the soil leads
to an increased release of N. Similarly, the application of the liquid fraction of
separated biogas residues will provide N at least corresponding to their ammonium
content with an increased release of N form the organic fraction after repeated
application. In contrast, depending on the soil and the type of biogas residue part of
the ammonium applied from the solid fraction after separation may be unavailable to
plants. Moreover, in the case of solid biogas residues N accumulated in soil seems to
be rather recalcitrant to mineralization.
Zusammenfassung 99
7. Zusammenfassung
Rückstände aus der Anaerobvergärung enthalten wertvolle Mineralstoffe (z.B. N, P,

K, S etc.), deren Rückführung in die landwirtschaftliche Produktion erforderlich ist,
um das Ziel einer nachhaltigen und umweltfreundlichen Biogasproduktion zu
erreichen. Biogasgärreste entstehen jedoch aus Vergärung einer Vielzahl
unterschiedlicher Substrate, die ausschließlich aus pflanzlichen Rohstoffen wie auch
aus Mischungen von pflanzlichen Rohstoffen mit tierischen Ausscheidungen
stammen können. Es ist zu vermuten, dass dies zu Unterschieden sowohl in der
Zusammensetzung der Gärreste als auch in deren Nährstoffwirkung führt.
Zusätzliche Veränderungen in der Zusammensetzung können sich durch die
mechanische Separierung in eine flüssige und eine feste Fraktion (Dünnseparat und
Feststoff) ergeben, was ebenfalls die Nährstoffwirkung beeinflussen könnte.
Ziel der vorliegenden Arbeit ist es daher
 Rückstände aus der Vergärung von unterschiedlichen Substraten hinsichtlich
ihrer N und C Gehalte zu charakterisieren, wobei nicht separierte Gärreste sowie
Dünnseparate und Feststoffe nach der Separierung berücksichtigt wurden,
 die kurz- und langfristige Wirkung des Einsatzes dieser Gärreste auf die N
Verfügbarkeit und die N Ausnutzung durch die Pflanze zu untersuchen und
 die Bedeutung verschiedener Böden für die N Ausnutzung aus diesen Gärresten
zu bewerten
Als Modellsystem hierfür wurden zwei Gefäßversuche von jeweils etwa einem Jahr
Dauer durchgeführt, in denen Weidelgras regelmäßig gedüngt und geschnitten
wurde. Dieses System erlaubte eine Untersuchung der kurzfristigen Wirkung nach
einmaliger Düngung wie auch der langfristigen Wirkung nach wiederholter Düngung.
Im ersten Experiment wurden sieben Gärreste aus der Vergärung unterschiedlicher
Substrate und eine unvergorene Rindergülle eingesetzt. Gedüngt wurde auf der
Basis des enthaltenen Ammoniums (300 mg NH 4-N pro 12-l-Gefäß und Düngung) in
Form einer Bandapplikation und sofortiger Einarbeitung. Der Einfluss des Bodens auf
die N Verwertung wurde mit fünf Böden unterschiedlicher Textur bzw.
unterschiedlichen Gehalts an organischer Substanz (2 schluffige Lehme, 1 toniger
Lehm, 1 Sandboden, 1 organischer Boden) untersucht. Im zweiten Experiment
wurden Gärreste aus sieben verschiedenen Biogasanlagen eingesetzt, die jeweils in
Dünnseparat und Feststoff separiert worden waren. Diese 14 Gärreste wurden
100 Zusammenfassung
jeweils in zwei Böden (1 sandiger Boden und 1 schluffiger Lehm) ebenfalls auf der
Basis von 300 mg NH4-N pro 12-l-Gefäß und Düngung nach sofortiger Einarbeitung
untersucht. Als Vergleich diente jeweils eine Mineraldüngung mit 300 mg N als
NH4NO3 und eine Kontrolle ohne N Düngung. In beiden Experimenten wurde das
Weidelgras in fünf Wachstumszyklen mit jeweils ein bis drei Aufwüchsen kultiviert,
wobei die Düngung zu Beginn jedes dieser Wachstumszyklen erfolgte.
Folgende Ergebnisse wurden erzielt:
(1) Auf der Basis des gleichen Angebots an Ammonium- bzw. Mineral-N war die N
Aufnahme von Weidelgras aus den nicht separierten Gärresten und der
Rindergülle vergleichbar oder höher als die N Aufnahme nach Düngung mit
NH4NO3. Bereits nach der ersten Düngung bestanden jedoch auch Unterschiede
in der N Aufnahme von bis zu 20% zwischen den Biogasgärresten. Die N
Aufnahme nach dem ersten Wachstumszyklus war signifikant negativ mit dem
Corg:Norg Verhältnis der organischen Dünger korreliert. Die über die N Aufnahme
aus NH4NO3 hinausgehende zusätzliche N Aufnahme aus den organischen
Düngern stieg vom ersten zum fünften Wachstumszyklus an. Am Ende des
Versuchs nach fünf aufeinander folgenden Düngungen zeigte sich eine
signifikante positive Korrelation zwischen der N Aufnahme des letzten
Aufwuchses und dem im Boden akkumulierten organischen Stickstoff, der
wiederum in enger Beziehung zur unterschiedlichen Menge an appliziertem
organischem N stand.
(2) Der Einfluss der Bodenart wurde mit fünf Böden und mit drei unterschiedlichen
Biogasgärresten untersucht. Die N Aufnahme aller Wachstumszyklen aus den
ungedüngten Böden war durch deren unterschiedliche Gehalte an Gesamt-N
geprägt. Demzufolge war die N Aufnahme in dem organischen Boden mit 4,4 g
Nt/kg Boden am höchsten, während der Sandboden mit einem Gehalt von 0,6 g
Nt/kg Boden die geringste N Aufnahme zeigte. Die N Aufnahme aus den drei
Düngern unterschied sich zwischen den Böden in jedem Wachstumszyklus,
zeigte aber keine konsistente Beziehung zur Bodenart oder dessen Gehalt an
organischer Substanz. Außerdem bestand keine signifikante Wechselwirkung
zwischen Düngern und Böden. Die Nettomineralisationsrate des über die Dünger
im Boden akkumulierten organischen Stickstoffs zeigte keine Unterschiede
zwischen den Gärresten.
Zusammenfassung 101
(3) Auf der Basis des gleichen Angebots an Ammonium- bzw. Mineral-N war die N
Aufnahme nach der ersten Düngung mit Dünnseparaten in beiden Böden
genauso hoch wie nach der Zufuhr von Mineraldünger. Im Unterschied dazu war
die N Aufnahme nach der Düngung mit Feststoffen eher niedriger als mit
Mineraldünger. Unter Einbeziehung sowohl der Dünnseparate wie auch der
Feststoffe bestand eine signifikant negative Beziehung zwischen der N
Aufnahme von Weidelgras und dem C org:Norg Verhältnis in den Gärresten. Dies
galt für die erste wie auch für alle vier folgenden Düngungen. Innerhalb der
jeweiligen Gruppe der Dünnseparate bzw. der Feststoffe bestand jedoch keine
Beziehung zwischen N Aufnahme und Corg:Norg Verhältnis im Gärrest.
Die zusätzliche scheinbare N Ausnutzung (additional apparent N utilization;
AANU), die die N Freisetzung aus der organischen Fraktion des Gärrestes
beschreibt, zeigte von der ersten bis zur fünften Düngung einen deutlichen
Anstieg. Während diese zusätzliche scheinbare N Ausnutzung durchwegs für die
Dünnseparate erhöht war, zeigten die negativen Werte für die Feststoffe eine
mehr oder weniger deutliche Immobilisierung des gedüngten Ammoniums nach
der Anwendung dieser Fraktion.
Nach fünf aufeinander folgenden Düngungen mit Feststoffen konnte eine
Erhöhung der Nt und Ct Gehalte im Boden beobachtet werden, während nach der
Düngung mit Dünnseparaten die Gehalte an N t und Ct im Boden auf dem Niveau
der Mineraldüngung blieben. Der Anstieg der Boden Nt Gehalte nach
Feststoffdüngung führte jedoch nicht zu einer Zunahme an pflanzenverfügbarem
N.
Aus den vorliegenden Untersuchungen kann geschlussfolgert werden, dass nicht
separierte Biogasgärreste unabhängig davon, ob sie ausschließlich aus pflanzlichen
Substraten oder aus der Co-Fermentation mit Wirtschaftsdüngern stammen,
mindestens eine dem Ammoniumgehalt entsprechende Menge an
pflanzenverfügbarem N bereitstellen. Dabei steht die N Freisetzung nach der ersten
Anwendung mit dem C org:Norg Verhältnis der Gärreste in Beziehung. Nach
wiederholter, d.h. Langzeitanwendung führt der durch die erhöhte Zufuhr an
organischem Stickstoff gestiegene Gehalt an organischem N im Boden zu einer
Erhöhung der N Freisetzung. Ähnliches zeigt sich nach der Anwendung von
Dünnseparaten, deren N Verfügbarkeit nach einmaliger Düngung ihrem Gehalt an
Ammonium-N entspricht und die mit zunehmender Häufigkeit der Anwendung
102 Zusammenfassung
steigende Mengen an N aus dem organischen Anteil freisetzen. Im Gegensatz dazu

ist, abhängig vom Boden und der Art des Gärrestes, nach der Düngung von
Feststoffen ein mehr oder weniger großer Teil des gedüngten Ammoniums nicht
mehr pflanzenverfügbar d.h. wird immobilisiert. Zudem scheint der durch die
Feststoffe im Boden akkumulierte organische N weniger gut mineralisierbar zu sein.
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Curriculum Vitae
Curriculum Vitae
Personal data:
Name: Sara El-Sayed El-Sayed Fouda

Nationality: Egyptian
Date of birth: 09.01.1981
Place of birth: Zagazig, Sharkea, Egypt
Material status: Married with two children
Work address: Department of Soil Science, Faculty of Agriculture,
Zagazig University, Zagazig, Egypt
Home Address: Elfateh Eleslamy Street 11, Mubarak Area, Zagazig, Egypt
E- Mail: [email protected]
Education:
12/2007 - present Ph.D. Student at Department of plant sciences (Chair of

Plant Nutrition), Center of Life and Food Sciences,
Weihenstepan, Technische Universität München, Freising,
Germany.
2001 - 2007 M. Sc. degree in Soil Science from Faculty of Agriculture,
Zagazig University, Egypt
1998 - 2001 B. Sc. degree in Agriculture Science from Faculty of
Agriculture, Zagazig University with final degree Excellent
with honour
1995 - 1997 Eldamam Secondary School, Saudi Arabia
1992 - 1994 Eldamam Preparatory School, Saudi Arabia
1987 - 1991 Notre Dame Des Apoters School, Zagazig, Egypt
Professional Work:
12/2007 - present Assistant Researcher at Department of plant sciences

(Chair of Plant Nutrition), Center of Life and Food
Sciences, Weihenstepan, Technische Universität
München, Freising, Germany
Curriculum Vitae
2001 - 2007 Demonstrator at Soil Science Department, Faculty of

Agriculture, Zagazig University, Egypt
Lebenslauf
Lebenslauf
Persönliche Daten:
Name: Sara El-Sayed El-Sayed Fouda
Nationalität: Ägyptisch
Geburtsdatum: 09.01.1981
Geburtsort: Zagazig, Sharkea, Ägypten
Familienstand: Verheiratet, Zwei Kinder
Dienstanschrift: Lehrstuhl für Bodenkunde, Fakultät der Landwirtschaft,
Universität, Zagazig, Ägypten
Home Address: Elfateh Eleslamy Str. 11, Zagazig, Ägypten
E- Mail: [email protected]
Ausbildung:
12/2007 - present Doktorandin am Lehrstuhl für Pflanzenernährung,
Wissenschaftszentrum Weihenstephan für Ernährung,
Landnutzung und Umwelt, Technische Universität
München, Freising, Deutchland
2001 - 2007 Erlangung des Mastergrades (M.Sc.) in Bodenkunde,
Fakultät für Landwirtschaft, Universität Zagazig, Ägypten
1998 - 2001 Studium der Agrarwissenschaften an der
Landwirtschaftlichen, Universität Zagazig, Ägypten (mit
Auszeichnung bestanden)
1995 - 1997 Gymnasium in Eldamam, Saudia Arabian
1992 - 1994 Mittlere Reife in Eldamam, Saudia Arabian
1987 - 1991 Notre Dame Des Apoters Schule, Zagazig, Ägypten
Berufstätigkeit:
12/2007 - present Wissenschaftliche Mitarbeiterin am Lehrstuhl für
Planzenernährung, Wissenschaftszentrum Weihenstephan
für Ernährung, Landnutzung und Umwelt, Technische
Universität München, Freising, Deutchland
2001 - 2007 Demonstrator am Department für Bodenkunde, Fakultät
der Landwirtschaft, Universität, Zagazig, Ägypten

Technische Universität München

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Technische Universität München

Uploaded by

TECHNISCHE UNIVERSITÄT MÜNCHEN

Lehrstuhl für Pflanzenernährung

Nitrogen availability of biogas residues

Sara El-Sayed Fouda

Vollständiger Abdruck der von der Fakultät Wissenschaftszentrum Weihenstephan

Doktors der Agrarwissenschaften

Vorsitzender: Univ.-Prof. Dr. J. Meyer

Die Dissertation wurde am 26.05.2011 bei der Technischen Universität München

The soul of my late Father (Prof. Dr. El-Sayed Fouda)

Freising - Weihenstephan, May 2011

1.1 Biogas production as a renewable energy resource ......................................... 1

2. Materials and methods .............................................................................. 15

2.1. Experimental conditions ................................................................................ 15

2.4.2 Plant analysis ........................................................................................ 26

4.1 N availability from different unseparated biogas residues .............................. 81

8. References ............................................................................................... 103

Figure 1 Composition of substrates used for the fermentation process to obtain

ANOVA analysis of variance

ANU apparent nitrogen utlization

BGR biogas residue

C/N carbon/nitrogen ratio

CaCl2 calcium chloride

CAL calcium acetate lactate

CEC cation exchange capacity

Corg organic carbon

ICP Inductively Coupled Plasma Emission Spectrometer

KCl potassium chloride

LBR liquid biogas residue

LSD least significant difference

NH4-N ammonium nitrogen

Nmin mineralized nitrogen

NO3-N nitrate nitrogen

Norg organic nitrogen

p≤0.01 1% probability level

p≤0.05 5% probability level

SBR solid biogas residue

1.1 Biogas production as a renewable energy resource

1.2 N availability of unseparated biogas residues

1.2.1 Comparison of unseparated biogas residues with other organic fertilizers

1.2.2 Effect of organic fertilizers’ C:N ratio on N mineralization

evaluated 87 different poultry wastes in an incubation experiment and found a

1.2.3 Characteristics of the unseparated biogas residues compared with animal

1.2.4 Biogas residues from different substrates

1.3 The role of soils in N availability from organic fertilizers

1.3.1 Factors affecting N mineralization in soils

1.3.1.1 Temperature and water content

1.3.1.2 Soil texture

1.3.1.3 Soil organic matter content

1.3.2 Effect of organic fertilizers on N mineralization in soils

1.4 Separation of biogas residues and other organic fertilizers

1.4.2 Separation efficiency

1.4.4 Nitrogen availability of separated solid and liquid biogas residues

1.5 Objectives of the present study

2. Materials and methods

2.1. Experimental conditions

November 1.6 -8.1 10.3

January 1.9 -5.7 12.6

February 2.6 -9.6 19.3

March 4.1 -8.1 19.3

May 14.5 1.7 30.2

July 17.6 7.8 31.3

August 17.3 5.7 32.2

September 11.9 -0.2 28.2

October 8.6 -1.8 20.9

Table 2 Monthly mean temperature measured in the greenhouse during the

2.2 Fertilizers and soils used in the experiments

2.2.1 First experiment

Figure 1 Composition of substrates used for the fermentation process to obtain

Table 3 Characteristics of the soils used in the first experiment

Clay Silt Sand pH Nt Ct P1 K1 Nmin

Soil 2 26 57 17 6.5 0.15 1.2 83 150 57

Soil 3 6 8 86 5.3 0.06 0.41 35 66 11

2.2.2 Second experiment

Figure 2 Composition of different substrates used for the fermentation process

Table 4 Characteristics of the soils used in the second experiment

Mühlfeld 13 21 66 5.7 0.09 0.80 66 192

Fertilizer 3 9.28 0.000* 3 30.45 0.000* 3 104.0 0.000***