ORBIT Symposium 1999
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LABORATORY TESTS DESCRIBING THE BIOLOGICAL
REACTIVITY OF PRETREATED RESIDUAL WASTES
Erwin Binner, Alexander Zach
Universität f. Bodenkultur Vienna, Dep. of Waste Management / IWGA,
Nußdorfer Lände 29-31, 1190 Wien
1 ABSTRACT
The Austrian Landfill Ordinance (1996) sets standards for the waste which is allowed to be landfilled. There are
limits for TOC, Ignition Loss, Calorific Value as well as for harmful substances, but no parameters describing the
biological reactivity of waste. The use of the Incubation Test as an experimental determination of gas generating
potential is very well suited for assessing biological reactivity because this anaerobe experimental method allows
the reproduction of most "natural conditions" in the laboratory. However, the incubation tests suffers from its
long duration, therefore other parameters had to be proven as well. Respiration activity, fermentation test,
analysis of substance groups (“Stoffgruppenanalysen” according to Van SOEST) and eluate parameters like
TOC, NH4-N, COD and BOD were discussed. Our research showed, that gas production in 90 days (GS 90) and
respiration activity in 7 days (AT 7) are the best suitable parameters. In Germany AT 4 and GB21 are favored
because of the shorter test duration. In September 1998 we started a research program in order to compare
incubation test (GS90) and fermentation test (GB21) by testing low reactive samples.
2 INTRODUCTION
In Austria as well as in Germany, several ideas have been put forward on how to describe the
activity of waste. Gas-generating potential, respiration activity and analysis of material groups
("Stoffgruppenanalysen" according to Van Soest, 1963) and eluate parameters like TOCeluate,
NH4-N, COD and BOD were discussed. The use of the Incubation Test (developed by Dep.
of Waste Management = ABF-BOKU) as an experimental determination of gas generating
potential is very well suited for assessing the reactivity of wastes because this experimental
method allows the reproduction of most "natural conditions" in the laboratory. Large sample
sizes (> 1 kg DS) and robust test conditions (short fluctuations of boundary conditions, e.g.
thermostat failure, only have a minimal impact on final results) ensure good reproducibility of
results. Due to the very long testing time, (an incubation test lasts approx. 3-8 months),
alternative methods were to be developed. That is why the Austrian Ministry of Environment
asked the Department of Waste Management of the Universität für Bodenkultur Wien
(University of Agricultural Sciences, Vienna) to undertake practical tests to describe the
reactivity of mechanically-biologically pretreated residual wastes from 4 different treatment
plants (results of these 8 samples of this "MBA 1 (=MBP, mechanical biological
pretreatment)-study" were presented in Sardinia '97; Binner et al 1997). In the meantime, 30
more samples have been analyzed (first results were presented in Lulea '98; Binner and Zach
1998a). Our research showed that gas production in 90 days (GS90) and respiration activity in
7 days (AT7) are the best suitable parameters for determining the biological reactivity of
waste. We suggest limit values for GS90 < 20 Nl/kg DS and a correlating AT7 of about
11 mg O2/g DS. This results in a decrease of the potential volume of landfill gas released
compared to untreated waste of more than 90%.
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Because of the shorter test duration, AT4 and GB21 are favored in Germany. In September
1998 we started a research program in order to compare incubation test (GS90) and
fermentation test (GB21) by testing low reactive samples.
3 MATERIALS AND METHODS
Most of the samples were taken out of the routine operation of the treatment plants. In order to
get very low reactive material, the samples of this study (MBA 2-study) were aerated 2 to 6
more weeks in laboratory test cells after sieving to < 20 mm.
The method for sampling mechanically-biologically pretreated waste was presented in detail
at Sardinia 1997 (Binner, et al., 1997). 10 single samples taken over the whole length of the
windrow were mixed and reduced to one mixed sample of about 50-100 kg. The mixed
samples were sieved with a drum-sieve  20 mm (wet grinding of samples is also possible).
For biological parameters (respiration activity, gas-production potential) and soluble
compounds (eluates DEV S4) the fraction < 20 mm is representative. The samples should be
analyzed as soon as possible. Stabilization is done by freezing. Stabilization by drying (even at
temperatures < 45°C) is not tolerable for biological parameters and soluble compounds.
3.1.1 Incubation Test (GS90)
The gas generation is determined by using incubation tests (Binner et al. 1998). A moist fresh
sample sieved to  20 mm with approximately 1 kg of dry matter is saturated to waterholding capacity and then incubated in glass reactors under anaerobic conditions at 40°C
(figure 1). Each material is tested 2 times; if both repetitions are parallel, one is stopped after
3 to 4 weeks. Gas generation is measured by a so-called "Eudiometer" and calculated to
normal-conditions (0°C, 1013 mbar). Even though gas generation is often not yet finished
after 240 days, it is still possible to make an adequately accurate assessment after 90 days.
The amount of gas generated within 90 days is between 65 and 95% of the volume produced
in 240 days (Binner et al. 1999). Short pretreated materials show quantities higher than 90%,
toxic impacts excepted. With longer pretreatment periods (GS90 <15-30 Nl/kg DS), the
amounts obtained are about 80% - 90%, very well pretreated material (GS90 <10 Nl/kg DS)
seems to produce 65% - 80% of the GS240. All the tests with very well pretreated material are
still running. Therefore most of the GS240 results are only estimated.
A test period shorter than 90 days leads to considerably lower findings. In 60-days a
production of about 50 – 90%, and for a 30-day period, results of about 20 - 75% can be
obtained.
The incubation test has some advantages over the fermentation test. The large sample amount
(> 1kg DS) and robust test conditions (short fluctuations of boundary conditions, e.g.
thermostat failure, only have a minimal impact on final results) ensure good reproducibility of
results. The high total gas amount in the incubation test cell (1 kg DS of a well pretreated
waste produces about 10 l gas) is to be collected more exactly than the low amount in the
fermentation test cell (50 g DS of the same waste only produce 0.5 l gas).
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Incubation Test Reactor - ABF-BOKU
tube
0 ml
6 mm
Glass tap
gas extrac tion
collecting
vessel
100 ml
150 m l
Eudiom etertube
600 m m
filled with 25% NaCl
1100 m l
water supply
tube
6 mm
100 mm
40 m m
teflontube
6 mm
200 mm
leachate
extraction
tube
6 mm
drainage bottom
150 m m
datei:skripten\grafik\versuch\bmu\inkuzy2e.cdr
Figure 1. Incubation Test Cell (ABF-BOKU)
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3.1.2 Fermentation Test (GB21, GB28)
Of course, the fermentation test also has advantages. Compared to incubation tests,
fermentation tests show significantly higher reactional behavior at the beginning of the test.
Toxic effects may influence the incubation test and lead to a longer lag-phase, fermentation
tests are, as a result of their dilution, not influenced. Fermentation tests are easier to handle; a
space-saving experimental arrangement allows the characterization of numerous samples in
one test series.
In Germany, a duration of 21 days is proposed for the fermentation test. Within 21 days, only
activity but no potential can be determined. First Tests in our laboratory showed that in 21
days the total gas amount is only about 10 to 60% of the amount produced in the incubation
test within 240 days (Binner and Zach, 1998a).
For the fermentation test we put 50 g DS + 1.000 ml H2O into a glass-reaction bottle. When
the test is inoculated (by adding leachate from the incubation test), the volume of added water
is reduced. Each material is tested twice. For the fermentation tests we don't use
eudiometertubes. The produced gas is collected in a glass bottle filled with 25% NaCl solution
(figure 2). The solution is pressed into a graduated cylinder by the produced gas, which is
measured daily and calculated to normal-conditions (0°C, 1013 mbar). The whole system is
situated in a climate-controlled room at 35°C. The test duration is 42 days minimum.
Fermentation Test Reactor
gaertst1.cdr
parafilm
Glass
reaction
bottle
[1l]
graduated
zylinder
25%
NaCl
sample
[50g DS]
Figure 2. Fermentation Test Cell (ABF-BOKU)
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3.1.3 Respiration Activity Test (AT4, AT7)
The fraction < 20 mm is wetted to an optimized water content ("Faustprobe") of about 4050%. 30 - 40 g of this wet sample are put into the reaction bottle. Hardly known samples are
tested in triplicates. Well known material is tested only 2 times. If one repetition differs
significantly, stabilized material (frozen) must be tested again.
The respiration activity is analyzed by Sapromat E. The reaction bottle is connected with an
oxygen generator and a pressure control gauge. The whole system is situated in a water bath
(20 °C). The CO2 produced by aerobe microorganisms is absorbed by the sodium hydroxide.
The negative pressure starts the oxygen generator which produces O2 until normal pressure
conditions are reached. O2 production is recorded by a computer (figure 3).
Recorder
Control Device
Pressure
Oxygen
Control Cage
Reaction
Generator
Bottle
Figure 3. Respiration Activity Test (Sapromat E)
Respiration activity, in contrast to the incubation test, does not assess the potential of a
sample, but rather its activity, meaning its momentary oxygen consumption per unit of time.
Normally in this test oxygen consumption remains constant over a period of 7 to 10 days
following a short lag-phase. Toxic impacts can cause longer lag-phases and therefore lead to a
misinterpretation of the AT4 value. In almost every case, this toxic impact no longer produces
false results after 7 days, (longest lag-phase: 6 days). After 7 days such long lag phases can be
recognized. The determination of AT10 does not provide enough additional information to
justify its application. A test period of 7 days is sufficient and, therefore, recommended for
respiration activity determination (Binner and Zach, 1998).
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4 RESULTS AND DISCUSSION
4.1 Relationship between Gas Generation (GS90) and Respiration Activity (AT7)
The research showed very good correlation between respiration activity and gas generation
(figure 4). The dotted correlation line includes 42 samples (r = 0.907). Gas generation of the
"toxic" samples is corrected by eliminating the lag-phase. Samples with oxygen starvation
during the biological pretreatment show lower results of respiration activity than expected.
That indicates the importance of doing anaerobic tests and aerobic tests in combination.
Focusing samples with low reactivity (GS90 < 50 Nl/kg DS) indicates that the correlation line
rises slightly (dark line in Figure 4, r = 0.891). The respiration activity (AT7) correlating to the
GS90 = 20 Nl/kg DS in that case is about 11.3 mg O2/g DS.
Relationship between Gas Generation (GS90) and
Respiration Activity (AT7)
toxic effects (corrected)
Respiration Activity (AT7) [mg O2/g DS]
100
MBA 1-study
75
MBA-routine output
oxygen starvation
42 samples: r = 0.907
MBA-routine output
50
MBA 2 - routine output
25 samples (GS90< 50): r = 0.891
MBA 2 - routine output
+ treatment in laboratory
25
MBA 2 - routine anaerob
11
MBA 2 - routine anaerob
+ treatment in laboratory
0
0
20
40
60
80
100
120
140
160
180
200
220
240
Gas Generation (GS90) [Nl / kg DS]
Figure 4. Relationship between gas generation GS90 (incubation test) and respiration activity
AT7 (Sapromat) of mechanically-biologically pretreated residual waste. The dark correlation
line is calculated with 25 low reactive samples (GS90 < 50 Nl/kg DS). The dotted line shows
the correlation of 42 samples; toxic effects were corrected.
4.2 Comparison of Incubation Test (GS90) and Fermentation Test (GB21)
As said before, the test duration of 90 days is too long for routine analysis of pretreated waste.
In the incubation test of well treated waste, inoculation has no important effect; neither to the
duration of the lag-phase not to the gas generation rate. Because of the shorter test duration
(21 days) we compared the results of the incubation test to those of the fermentation test as
suggested in Germany. For inoculation we used leachate of well running incubation tests.
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Incubation Test and Fermentation Test
5 Weeks Aerobe Pretreated Waste
Gas Generation [Nl / kg DS]
100
Inkubation Test, not inoculated
GS21 = 27,7 Nl/kg DS
Fermentation Test, not inoculated
GB21 = 22,5 Nl/kg DS
75
GS 90 = 63 Nl gas / kg DS
Fermentation Test, inoculated
GB21 = 32,4 Nl/kg DS
50
25
0
0
30
60
90
120
Test Duration [days]
Figure 5. Gas Generation of 5 weeks pretreated waste in incubation test and fermentation test
Incubation Test and Fermentation Test
27 Weeks Aerobe Pretreated Waste
25
Inkubation Test, not inoculated
GS21 = 5,0 Nl/kg DS
Gas Generation [Nl / kg DS]
20
Fermentation Test, not inoculated
GB21 = 0,1 Nl/kg DS
Fermentation Test, inoculated
GB21 = 0,9 Nl/kg DS
15
10
GS 90 = 11,8 Nl / kg DS
5
0
0
30
60
90
120
Test Duration [days]
Figure 6. Gas Generation of 27 weeks pretreated waste in incubation test and fermentation test
In the fermentation test inoculation makes the process faster (figure 5), but the inoculated
fermentation test (duration 21 days) only produces 2% to 60% of the GS240 detected in the
incubation test (table 1). For well pretreated waste, the gas production within 21 days in the
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incubation test is equal or higher (15% to 60%) than in the fermentation test (figure 6). Only
for high reactive material (which is not the usual object of assessment) does the fermentation
test start faster than the incubation test. After a test duration of about 21-28 days both tests
produce similar gas amounts.
Table 1. Gas generation in the incubation tests (GS21, GS90, GS240) compared to fermentation
tests (GB21). Amounts in italics are estimated.
Sample
1)
Duration GB21
GS21
GS90
GS240
GB21
of Preinocul.
not
not
not
treatment
inocul. inocul. inocul.
[Nl/kgDS] [Nl/kgDS] [Nl/kgDS] [Nl/kgDS] [% 240]
[weeks]
53
8,9
91
165
32
SAB 3-01) 0
SAB 3-8
8
2,1
5,4
16
25
8
OP 7-5
5
73
24,0
114
120
61
ZS 3-10
10
6,4
16,1
23,5
27
24
ZS 3-13
13
2,9
7,9
14,6
18
16
ZS 3-16
16
1,7
4,5
10,7
14,5
12
Incubation tests are still running
MBA 2 – study:
OP8-27
27
0,9
5,0
11,8
15,6
6
OP8-31
31
0,2
0,1
0,5
1,3
15
LZ1-16
16
0,4
4,1
8,2
12,1
3
LZ1-20
20
0,0
0,5
2,4
3,8
0
MV4-5
5
32,4
27,7
63
69
47
MV4-9
9
0,8
3,5
10,9
13,7
6
MV4-11
11
0,0
1,3
6,5
10,2
0
ZS5-14
14
34,1
44,8
95
100
34
ZS5-16
16
11,8
17,7
29,5
35
34
ZS5-19
19
1,0
2,0
8,3
12,5
8
ZS6-16
16
0,3
5,5
11,7
16,1
2
LÜN 1-16 16
0,0
0,8
2,2
n.e.
LÜN 1-18 18
0,0
0,5
1,1
n.e.
BA 1-11
11
20,3
34,3
58
64
32
BA 1-13
13
2,4
10,7
20,8
n.e.
BA 1-15
15
1,8
5,7
n.e.
n.e.
toxic effects (low pH-values)
n.e. = not estimable
GS21
GS90
[% 240]
5
22
20
60
44
31
[% 240]
22
64
95
87
81
74
32
8
34
13
40
26
13
45
51
16
34
54
-
76
38
68
63
91
80
64
95
86
66
73
91
-
Figure 7 shows that there is a very high correlation (mathematical function 2nd order),
r = 0.975, but for many low reactive samples no gas production was shown in the fermentation
test whereas in the incubation test 1 - 5 Nl/kg DS were obtained (table 1). A possible reason
could be the small sample size (50 g DS) for the fermentation test, which is not enough for the
gas collection system. Another reason could be the aquatic milieu which might limit the
availability of hardly degradable organic compounds for the anaerobic microorganisms. To
gain more information concerning this, a low reactive material will be tested under aquatic
conditions in the incubation test reactor (270 g DS + 1,800 ml H2O + 450 ml inoculum). In
May 1999 a "Ringtest" about fermentation test and respiration activity was started. To
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complete the assessment the material will also be tested by incubation test. The results will be
presented at the symposium.
Fermentation Test (GB21) [mg/kg DS]
Relationship between Incubation Test (GS90) and
Fermentation Test (GS21)
75
y = 0,0039x + 0,1492x
r = 0,975
50
25
0
0
20
40
60
80
100
120
Gas Generation (GS90) [Nl / kg DS]
Figure 7. Relationship between gas generation GS90 (incubation test without inoculation)
and GB21 (fermentation test with inoculation) of
mechanically-biologically pretreated residual waste
5 CONCLUSIONS
Alternative parameters describing the reactivity of mechanically-biologically pretreated waste
were tested by analyzing 42 samples taken from 9 different MBP-facilities. 25 of these
samples showed very low reactivity (GS90 = 0.5 to 40 Nl/kg DS).
 The amount of gas generated in the incubation test in 90 days (GS90) is about 75% to 90%
of the gas generating potential of well pretreated residual waste (GS90 = 15 – 30 Nl/kg DS).
 A limit value for gas generation GS90 < 20 Nl/kg DS indicates a reduction of reactivity by
> 90% compared to untreated residual waste. Corresponding to the incubation test GS90
limit value of 20 Nl/kg DS, the fermentation test GB21 limit value would be about
5 Nl/kg DS.
 There is a significant correlation between gas generation (GS90) and respiration activity
(AT7). The correlation coefficient (42 samples) is r = 0.907, for low reactive materials
(GS90 < 50 Nl/kg DS) r = 0.891, respectively. The AT7 correlating to GS90 = 20 Nl/kg DS
is about 11 mg O2/g DS in both cases.
 A fermentation test with inoculation (duration 21 days) as suggested in Germany, produces
only 2% to 60% of the GS240. For well pretreated waste the gas production in the
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incubation test is equal or higher (15% to 60%). Only for higher reactive material does the
fermentation test show higher gas production than the incubation test in 21 days.
 The fermentation test, however, starts a little bit faster than the incubation test. After a test
duration of about 21-28 days the incubation test shows equal or even higher gas amounts.
 There is a high correlation between incubation test and fermentation test (r = 0.975). The
incubation test seems to be more exact for low reactive materials. In the fermentation test
of very low reactive material (GS90 < 5 Nl/kg DS) very often no gas can be detected,
whereas the incubation test shows gas production.
LIST OF REFERENCES
Binner E., Lechner P., Widerin M., Zach A. (1997). Laboratory Test Methods Characterizing
the Biological Reactivity of Wastes. Proc. of the 6th International Landfill Symposium,
Sardinia. Italy. R. Cossu (Ed.), vol. 1, pp.485-494 .
Binner E., Zach A., Widerin M., Lechner P. (1998). Auswahl und Anwendbarkeit von
Parametern zur Charakterisierung der Endprodukte aus mechanisch-biologischen
Restmüllbehandlungsverfahren. Schriftenreihe des BMUJF, Vienna, Austria. vol.
9/1998 (a short version is published in Waste Reports 7/98. Lechner P. (Ed). ABFBOKU, Vienna A.)
Binner E. and Zach A. (1998a). Biological Reactivity of Residual Wastes in dependence on
the duration of Pretreatment. Presented at the 3rd Swedish Landfill Symposium, Lulea,
Sweden.
Binner E., Zach A., Lechner P. (in press). Test Methods Describing the Biological Reactivity
of Pretreated Residual Wastes. Proc. of the 7th International Landfill Symposium,
Sardinia. Italy. R. Cossu (Ed.).
Van SOEST J.P. (1963). Use of Detergents in the Analysis of Fibrous Feeds. II. A rapid
Method for the Determination of Fiber and Lignin. J.Assoc.Off.Anal.Chem., 46 (5), p.
829-835.
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