United States
Environmental Protection
Agency
National Risk Management
Research Laboratory
Research Triangle Park NC 27711
Research and Development
EPA/6QQ/SR-97/071   September 1997
                                                        of
                                                    in
M. A. Barlaz, W. E. Eleazer, W. S. Odle, III, X. Qian, and Y-S. Wang
  The objective of this research was to
characterize the anaerobic biodegrad-
ability of the major biodegradable com-
ponents of  municipal  solid  waste
(MSW). Tests were conducted in  qua-
druplicate in  2-L  reactors operated to
obtain  maximum yields. Measured
methane (CH4) yields for grass, leaves,
branches, food waste, coated  paper,
old newsprint, old corrugated contain-
ers, and office paper were 144.4,  30.6,
62.6, 300.7, 84.4, 74.3, 152.3, and 217.3
mL CH4/dry g, respectively. While there
was a general trend of increasing CH4
yield with  increasing  cellulose  plus
hemicellulose  (carbohydrate) content,
many confounding factors  precluded
establishment of a quantitative relation-
ship. Similarly, the degree of lignifica-
tion of a particular component was not
a good predictor  of the extent of bio-
degradation.
  In parallel with the decomposition ex-
periments,  leachate from the  decom-
position of each refuse constituent was
analyzed for toxicity using a modified
anaerobic toxicity assay.  Leachate tox-
icity was not found in association with
the decomposition of any refuse com-
ponent other than food  waste. How-
ever, substantial toxicity was measured
in leachate from the food waste reac-
tors. This toxicity was consistent with
the behavior of the reactors but could
not be simulated  with high concentra-
tions of carboxylic acids and sodium.
The toxicity associated with food waste
leachate is  not likely to inhibit anaero-
bic decomposition in U.S. landfills due
to the relatively low concentration of
food waste in MSW.
  Most probable number (MPN) tests
were conducted to identify the compo-
nents  of refuse that  carry  refuse-de-
composing  microorganisms  into
landfills and to evaluate the significance
of two typical cover soils as  carriers of
refuse-decomposing microbes. Grass,
leaves, and branches were  the major
identifiable contributors  of  refuse-de-
composing microbes to landfills, while
the cover soils tested did not typically
contain populations with the activities
required for refuse methanogenesis.
  This Project Summary was developed
by EPA's Air Pollution Prevention  and
Control Division of the National Risk
Management Research Laboratory, Re-
search Triangle Park, NC, to announce
key findings of the research project that
is fully documented in a separate report
of the same  title (see Project Report
ordering information at back).

Introduction
  Approximately 62%  of  the  municipal
solid waste  (MSW) generated  in the U.S.
is disposed  of by burial in a sanitary land-
fill. The production of methane (CH4) from
sanitary landfills is well documented,  and
there are about  119 landfill gas  recovery
projects currently (January 1997) in op-
eration in the U.S. and Canada. While the
production of  CH4 from landfills is  well
established, there is large  uncertainty in-
volved  in estimating the  amount and  rate
of CH4 production. This uncertainty is in-
creasing as the  composition of the MSW
buried  changes due to increased recy-
cling.

-------
  Development of integrated solid waste
management programs, which include re-
cycling and  in some cases,  combustion,
have  led  to  a decrease in the use  of
landfills. However, there is a limit to the
types of waste that can be recycled, and
combustion has not  been the solid waste
management alternative of choice for many
communities. Thus, landfills will be a sig-
nificant part  of MSW management for the
foreseeable future.
  Both CH4  and carbon  dioxide CO2 are
greenhouse  gases that contribute to glo-
bal  climate change.  CH4 traps about 20
times more infrared energy than CO2 on a
volume  basis.  Consequently, although
landfill gas contains approximately  equal
proportions of CH4 and CO2,  CH4 is more
significant with respect to atmospheric cli-
mate change. Data on the amount of CH4
that can be expected from refuse already
buried, as well as CH4 that will result from
the  decomposition of refuse buried in the
future, are needed to better assess the
impact of landfills on global climate change.
  The overall objective  of this  research
was to develop information on the anaero-
bic  decomposition of refuse  that will im-
prove our ability to assess the impact  of
sanitary landfills on global CH4 accumula-
tion. Three sets of experiments were con-
ducted  to  meet  this  objective: (1)
measurement of the CH4 potential of the
major biodegradable components of MSW;
(2) assessment of whether leachate toxic-
ity, associated with whole refuse or some
individual constituent, inhibits the onset or
rate of CH4 production; and  (3) identifica-
tion of solid  waste constituents that carry
the anaerobic bacteria required for refuse
methanogenesis. The  results of each set
of experiments are summarized separately.

Experiment 1: Measurement of
the CH4  Potential  of the Major
Biodegradable Components of
MSW
  The anaerobic biodegradability of the
major biodegradable components  of MSW
was characterized by measurement of their
CH4 yield and the biodegradation  of cellu-
lose and hemicellulose. The  components
that  were tested were  grass,  leaves,
branches,  food waste, and four types  of
paper—newsprint (ONP), old corrugated
containers (OCC), office paper (OFF), and
coated paper (CP).  These are the most
common types of paper in MSW and also
represent the range of biodegradability that
could  be  expected  from paper.  At one
extreme, newsprint contains all of the lig-
nin  of wood  pulp. At the other extreme,
office paper  has had almost all of the
lignin removed.  The  decomposition  of
mixed residential refuse was also charac-
terized.
  Tests were conducted in 2-L laboratory
reactors  in  quadruplicate.  Each  refuse
component was seeded with well-decom-
posed refuse to initiate refuse-decomposi-
tion. CH4 yield data have been corrected
for  the  background CH4 produced from
the  seed.  In the case  of food  waste, two
sets of reactors were  tested.  In the first
(F) series, there was insufficient seed, 30%
by volume, and the  reactors remained in-
hibited. A second set of food waste reac-
tors (SF) was then initiated with 70% seed
by volume, and these reactors produced
measurable CH4.
  The test conditions were designed  to
measure the maximum CH4  production
potential of each component. This included
shredding, incubation at  about 40°C, and
leachate recycle and neutralization. All re-
actors were  monitored until they were no
longer producing measurable CH4, except
for the old  corrugated container reactors
in which the CH4 yield increased by less
than 2% over the final 80 days of monitor-
ing. At the termination of the  monitoring
period,  reactors   were destructively
sampled for analysis of  the residual sol-
ids.
  The CH4 yield, solids composition, and
extent of cellulose and hemicellulose de-
composition for each MSW component and
mixed MSW are presented in Table 1. As
summarized in  Table  1, there was sub-
stantial variation in the  range of CH4  yields
(30.6 to  300.7 ml/dry g) and the extent of
decomposition (28  to 94%)  among the
components tested.  In previous research
with mixed  refuse, carbohydrates ac-
counted for 91% of the stoichiometric CH4
potential of MSW. Carbohydrates were the
major organic compounds analyzed  in the
waste components  tested  here, and the
relationship between carbohydrate concen-
tration and CH4 yield is presented in Fig-
ure  1. While the data in  Figure 1 show a
relationship, the relatively low correlation
coefficient (r2 = 0.49) and failure  of the
regression line to pass through zero, sug-
gest that factors in  addition to  carbohy-
drate concentration influence CH4 yield.
  Lignin decreases carbohydrate bioavail-
ability and  is expected  to confound the
relationship  presented in Figure 1. The
components with the lowest yields are the
two  sets of seed  reactors and  leaves.
These are also the  components with the
lowest  carbohydrate to  lignin  [(C+H)/Li)j
ratio. The  (C+H)/Li  ratio is a measure  of
the  degree of lignification. Values of 3 to 4
have been  reported for fresh refuse, and
lower values are associated with decom-
posed refuse. There is a general trend  of
more  extensive cellulose biodegradation
(MC decreasing) in the less lignified sub-
strates [(C+H)/Li increasing] e.g., food and
office paper (r2 = 0.28). However, the quan-
titative relationship is  weak because the
office  paper (C +  H)/Li is well above any
of the other components tested. The trend
towards increased cellulose  loss  with a
decreasing degree of lignification is most
definite among the four paper components.
  There is not  a linear relationship be-
tween (C+H)/Li and the extent of decom-
position  (r2  =  0.02).  However, it  is
interesting to note that grass,  which  is
highly lignified, underwent nearly complete
decomposition as  measured by either MC
or the extent of decomposition  (Table 1).
This suggests that the lignin concentra-
tion does not always reflect the degree to
which lignin inhibits cellulose bioavailabil-
ity. Apparently, the lignins in grass are not
as  restrictive to  microorganisms  as the
lignin  in  other  components   such  as
branches.  This result is consistent with a
report that stated, ".  . .the chemistry  of
grass  lignocellulose varies  considerably
from that of wood."
  The solids decomposition (MC and MH)
and CH4 yield data document the biode-
gradability of even the most lignified sub-
strates, leaves and branches, as well as
all other components of MSW tested. The
absence of  good linear  relationships  is
likely because a number of factors influ-
ence CH4 production and solids decompo-
sition.  The biodegradation  of  newsprint
measured  here is  in contrast to reports on
the  excavation of  readable newsprint that
had been  buried  in landfills decades  ear-
lier; however, these reported  data did not
represent  average values, but rather ob-
servations during an archaeological exca-
vation. The presence of readable newsprint
that had   not undergone biodegradation
may be due to  its isolation from other
factors required for biodegradation such
as bacteria,  moisture,  and nutrients.  The
biodegradability  of a newspaper buried in
a bag  that did  not break during waste
compaction would differ from the biode-
gradability of newsprint exposed to other
refuse components.
  Based on the CH4 yields presented  in
Table 1, a model was constructed to esti-
mate CH4  yields based on assumed com-
positions of  buried refuse. These  results
are  summarized  in Table 2. The actual
methane yield per wet kg of refuse buried
decreases by only 10% between the base
case (64.9 L CH4/wet kg) and  the case
with the most recycling  (58.6 L CH4/wet
kg).  However, the  appropriate  way  to
evaluate changes in  methane yield is  to
calculate the change in methane potential

-------
Table 1.  CH4 Yield and Initial and Final Solids Composition Data Summary3
Reactor
Series
Seed
sd
Seed-2d
sd
Grass
sd
Raleigh
grass
sd
Leaves
sd
Branch
sd
Yield
mL CH4/
dryg
25.5
5.7
5.8
0.6
144.4
15.5
127.6

21.8
30.6e
8.6
62.6e
13.3
Cellulose
23.4

18.3

26.5

25.6


15.3

35.4

Hemi-
cellulose
4.7

3.7

10.2

14.8


10.5

18.4

Lignin
22.5

22.1

28.4

21.6


43.8

32.6

MCb
0.18
0.02
0.34
0.01
0.19
0.01



0.43
0.05
0.52
0.05
MHb
0.36
0.03
0.69
0.11
0.42
0.06



0.68
0.10
0.59
0.02
Extent of
Decomposition0
21.8

6.3

94.3

75.5


28.3

27.8

Food
                                46.1
                                                  6.2
                                                                    8Q
                                                                    .6
Second
Food
sd
ONP
sd
OCC
sd
OFF
sd
CP
sd
MSW
sd
300.7e
10.6
74.33
6.802
152.3
6.7
217.3
14.96
84.4
8.1
92.0e
4.1
55.4
48.5
57.3
87.4
42.3
28.8
7.2
9
9.9
8.4
9.4
9
11.4
23.9
20.8
2.3
15
23.1
0.24
0.02
0.73
0.05
0.36
0.01
0.02
0
0.54
0.01
0.25
0.03
0.58
0.04
0.46
0.06
0.38
0.01
0.09
0.01
0.58
0.06
0.22
0.05
84.1
31.1
54.4
54.6
39.2
58.4
" Data represent the average for each reactor set. Standard deviations (sd) are presented below the average where data are the average of all reactors in a set.
b The ratio of the cellulose (MC) or hemicellulose (MH) recovered from a reactor divided by the mass added initially.
c The measured CH4 yield divided by the yield calculated by assuming conversion of 100% of the cellulose and hemicellulose (and protein in the case of food waste) to CH4 and
 CO2.
d Seed used for second set of food waste reactors.
8 Yield data for the leaf reactors exclude L2, data for the branch reactors exclude B4,and data for the second food and MSW reactors were corrected for leakage.
based on the yield multiplied by the mass
landfilled. Using this calculation, the  po-
tential reduction in methane production is
25.5% and  38% for the  recycling  cases
based on national averages and local re-
cycling  rates,  respectively. Thus,  these
data  suggest that recycling  can have a
substantial impact on the volume of meth-
ane  available for  recovery over the  de-
composition period.
  Where CH4  is released  to  the  atmo-
sphere,  recycling   clearly  reduces  the
amount  of  CH4  released from  landfills.
However, at landfills where there is an
active program to  compare the relative
benefits of recycling  and energy recovery.
Given the CH4 potential data for individual
constituents measured here, this analysis
could be done on  a component-specific
basis because the results may be differ-
ent for two  different types  of paper  or
between yard waste and paper.
  The calculated composite CH4 yields in
Table 2 range from 58.6 - 64.9 L CH4/wet
kg of MSW. These values are low relative
to landfill gas  models that generally as-
sume a yield of 62.3 to 112.2 L/kg. This is
surprising in that the CH4 yields measured
here were measured under optimal condi-
tions  and should  be  considerably  higher
than values assumed for field conditions.
There are  two potential explanations for
this discrepancy. The first explanation  is
that the assumed waste composition is in
error. The  data presented in an EPA re-
port  represent an  estimate of MSW gen-
eration  and exclude a number of wastes
that are buried in landfills.  Some of these
other wastes have high CH4 yields (waste-
water treatment plant sludge and agricul-
tural and food  preparation wastes),  while
others have little or no CH4 potential (wa-
ter treatment plant sludge and construc-
tion  and  demolition  debris).  A  second
explanation for the  discrepancy  in  yield
calculations pertains  to  the assumptions
used by the landfill gas models. The range
of values  used, 62.3 to  112.2 L/kg,  is
based  on field measurements and an esti-
mate  of the mass of waste buried in a
landfill. While  this  mass  is  accurately
known in  newer landfills where all waste
received is weighed, this mass  represents
only an estimate at older facilities and
errors  of 20 to 30% would  not appear to
be  unreasonable. Thus,  the  values  as-
sumed in practise may be inaccurate.

-------
      350 -i
                                        Second  Jk
                                        food    ^
                                                                  r2 = 0.493
                                                                       100
                                 Cellulose + Hemicellulose (%)
Figure 1. CH4 yield vs. carbohydrate concentration.
Table 2.  Calculated CH4 Yield Based on Measured Yields and Assumed MSW Composition
Case
Base Case-No Recycling
Recycling at National Average
Recycling-Typical Local Program
Yield
(L CH4/
wet kg)
64.9
59.9
58.6
Methane
Reduction3
(%)
na
25.5
38.0
Recycle
Rate
(%)
na
19.4
30.9
1 Calculated from the CH4 yield multiplied by the mass buried after recycling relative to the CH4 yield and mass buried
 in the base case.
Experiment 2: Measurement of
the Anaerobic Toxicity of
Leachate Associated with the
Decomposition of Individual
Refuse Components
  The anaerobic toxicity of leachate asso-
ciated  with the  decomposition  of  each
refuse component tested above was mea-
sured in  parallel with the decomposition
experiments. Leachate was sampled three
times from each  reactor. For food waste,
four samples  were collected from the F
reactors,  but  no  samples were collected
from the  SF reactors.  Six  samples  were
collected  from the MSW reactors. The ini-
tial  strategy was to sample each reactor
twice during the acid phase and  twice
during  the  decelerated  CH4  production
phase.  However, except for the first set of
food reactors  (F),  the  acid  phase was
very brief. As  a result,  only one  sample
was collected from most reactor sets dur-
ing  the acid phase.
  Leachate toxicity was  evaluated using a
modified  anaerobic toxicity assay (ATA).
The ATA included  anaerobic medium,
ground refuse as a carbon source, and an
inoculum. The inoculum was  a methano-
genic consortium enriched from decom-
posed  refuse  with  ground refuse  as a
carbon source. CH4 production from the
ground refuse  was  measured  in triplicate
in the presence and absence  of leachate.
Leachate was tested at final  concentra-
tions in the ATA of 25 and   80%  of its
original strength. Two sets of controls were
also inoculated. Controls containing inocu-
lum and medium but no  refuse were used
to  measure background CH4  production
from the inoculum. Controls containing in-
oculum, medium, and ground refuse were
used to compare CH4 production  in the
presence and absence of leachate.
  Leachate toxicity was  not measured  in
association with the decomposition  of any
refuse component other than food waste.
However,  leachate  associated with the
food waste reactors containing 30% seed
and  70%  food waste  (F) exhibited sub-
stantial toxicity, and this toxicity was gen-
erally consistent with the behavior of the
reactors.
  The toxicity of the food waste leachate
could not be  simulated with  synthetic
leachate containing high concentrations  of
carboxylic acids and sodium. ATAs with
20,  5,  15, and 12 g/L of acetate,  propi-
onate, butyrate, and  sodium, respectively,
suggested that high concentrations of bu-
tyric acid and sodium  inhibited  the onset
of CH4 production, but that refuse  micro-
organisms could  acclimate to these con-
centrations within 5 to 10 days  under the
conditions of the ATA.  The corresponding
concentrations of undissociated acetic, pro-
pionic,  and butyric acids were 113, 27,
and 96.8 mg/L, respectively. Comparison
of carboxylic  acid  concentration  data
from the S and SF  reactors series indi-
cated that the refuse  ecosystem was
able to tolerate  and recover from 142,
35,  24,  and 305 mg/L of undissociated
acetic,  propionic, i-butyric,  and  butyric
acids,  respectively.  These concentra-
tions of undissociated, carboxylic acids
are higher than concentrations reported
to be inhibitory in previous research with
anaerobic digesters.

Experiment 3: Identification of
Solid  Waste Constituents that
Carry the Anaerobic Bacteria
Required for Refuse
Methanogenesis
  The objective of part of this study was
to identify the components of refuse that
carry refuse-decomposing microorganisms
into landfills. A  second objective was  to
evaluate  the  significance of two typical
cover soils as carriers of refuse-decom-
posing  microbes. Refuse  buried in a sani-
tary landfill is typically covered with  15 cm
of soil  daily. Recently, geotextile sheets
and foams have been proposed as alter-
natives to soil to minimize the volume  of
soil in a landfill. While soil may  contribute
refuse-decomposing  microorganisms  to
landfills, the proposed  alternatives almost
certainly do not.
  The total anaerobic  population and the
subpopulations of cellulolytic, hemicellu-
lolytic,  hydrogen-producing  acetogenic
(based on butyrate  catabolism) bacteria

-------
and acetate- and hydrogen (H2)/CO2-uti-
lizing  methanogenic bacteria were enu-
merated  by the most probable number
(MPN) technique on several waste com-
ponents:  grass,  branches, leaves, food
waste, whole  refuse, and landfill  cover
soil.  For  each component, the objective
was  to enumerate  microbial  populations
on a representative  sample in the form in
which  it  would typically  enter  a landfill.
Although paper represents  37.6%  of
refuse, it was  not  tested  because  it  is
likely  populated  with  bacteria  originating
in wet components of refuse.
  Microbial enumerations were performed
by MPN  tests. Thus, it was necessary  to
form a liquid inoculum from solid samples.
The technique used here was similar to a
technique developed previously to process
smaller samples. In  the laboratory,  refuse
samples  were placed in a 113-L  plastic
garbage  can which  had been wiped  with
ethanol and purged with sterile argon.  A
measured volume of filter sterilized anaero-
bic phosphate buffer  (23.7 mM, pH  7.2)
was  then added to a sample to form a
slurry. The  sample was then stirred by
hand  (covered  with  arm  length gloves).
Next,  four samples were removed using a
1-L beaker, and the  liquid was poured into
a sterile, 4-L,  argon-purged,  Erlenmeyer
flask.  The liquid in this flask served  as the
inoculum for MPN enumerations. Inocula
were  serially diluted in phosphate buffer
(23.7  mM, pH 7.2). For soil, 250 to 300 g
of each sample  was added directly to a
nitrogen-purged flask, 2.5 L of sodium py-
rophosphate (0.1%, pH 7) was added, and
the slurry was shaken for 2 minutes. The
slurry was then allowed to settle for 1
minute after  which  a liquid  sample was
removed for use as  an inoculum.
  Microbial  populations on  each  waste
component and whole refuse are reported
in  Table  3.  Total  anaerobic  and
hemicellulolytic  populations were present
on all components tested, while the pres-
ence  of cellulolytic,  acetogenic, and
methanogenic bacteria was more  limited.
Thus, identification  of the waste compo-
nents that  are the  major  contributors of
cellulolytic,  acetogenic, and methanogenic
bacteria is evaluated here. Yard  waste
(grass, leaves, and  branches) most con-
sistently carried  the microorganisms  re-
quired  for  refuse methanogenesis.
Surprisingly, food waste did not carry  ei-
ther cellulolytic or methanogenic bacteria,
and one of two food waste samples con-
tained only one acetogen per gram. Popu-
lations of  cellulolytic,  acetogenic, and
methanogenic  bacteria were  generally
lower in the mixed  refuse samples com-
pared to the grass,  leaves, and  branch
samples.
Table 3. Anaerobic Microbial Populations on Refuse Components (Most Probable Number—Iog10
        cells/dry g)a

 Trophic   Total      Hemicellu-                           Methanogen  Methanogen
 Group  Anaerobes	lolytic	Cellulolytic    Acetogen	Acetate	H2/CO2
Grass
(April 92)
Grass
(July 92)
Branches
9.8

9.8

6.5
7.9

9.5

4.2
1.4

t>

2.5
0.7

1.8

1.3
1.6

t>

1.1
1.8

1.3

0.8
Leaves
(Nov. 91)
Leaves
(Nov. 92)
Food
(Mar. 92)
Food
(Aug. 92)
Refuse
(July 92)
Refuse
(Sept. 92)
5.8

6.9

>8.0C

9.4

9.3

8.4

4.1

4.4

5.3

6.2

6.6

6.3

1.0 <0.4 1.0

1.8 4.4 3.0

<-0.4 <-0.1 <-0.1

<-0.4 0 <0

0.4 0.2 3.6

<-0.2 <-0.1 <-0.1

0.7

3.8

<-0.1

<0

5.0

0.8

  Grass, leaves, and branches were the
major identifiable contributors of refuse-
decomposing microbes to landfills. About
9% of the refuse stream is characterized
as "miscellaneous" and contains many dif-
ferent items. In  addition to diapers and
pet wastes,  there  may  be other compo-
nents in the miscellaneous fraction that
carry refuse-decomposing microbes. How-
ever, their presence is small, and they are
likely to be poorly distributed. The  impor-
tance of yard waste should be considered
as solid waste management programs are
implemented. Where there is  interest  in
CH4 recovery from landfills, banning yard
waste from landfills may be self-defeating.
Unless,  of course,  the landfill is receiving
substantial volumes of other wastes known
to carry refuse-decomposing  microbes.
The cover soils tested did  not typically
contain  populations with the activities  re-
quired for refuse methanogenesis. Thus,
efforts to develop  lower volume alterna-
tives to cover soil  will  not adversely im-
pact the  input  of refuse-decomposing
microbes to  landfills.
  Financial support for this research was
provided by the Climate Change Research
Program of  the U.S. Environmental Pro-
tection  Agency, Waste  Management  In-
corporated,  the  National   Science
Foundation,  and S.  C.  Johnson Wax  &
Son. This support is  gratefully acknowl-
edged. We are also grateful for the assis-
tance of Kathi McBlief in  editing, typing,
and figure preparation.
" Data reported as less than a number indicate that no positive tubes were detected. The number reported assumes
 one positive tube in the first dilution.
b MPN results code was anomalous and not reported.
0 All tubes were positive at the highest dilution tested.

-------
  M. A. Barlaz, W. E. Eleazer, W. S. Odle, III, X. Qian, and Y-S. Wang are with North
   Carolina State University, Raleigh, NC 27695-7908.
  Susan A. Thorneloe is the EPA Project Officer (see below).
  The complete report, entitled "Biodegradative Analysis of Municipal Solid Waste in
   Laboratory-Scale  Landfills," (Order No. PB97-189674; Cost: $35.00, subject to
   change) will be available only from
          National Technical Information Service
          5285 Port Royal Road
          Springfield, VA 22161
          Telephone: 703-487-4650
  The EPA Project Officer can be contacted at
          Air Pollution Prevention and Control Division
          National Risk Management Research Laboratory
          U.S. Environmental Protection Agency
          Research Triangle Park, NC 27711
United States
Environmental Protection Agency
Center for Environmental Research Information
Cincinnati, OH 45268
Official Business
Penalty for Private Use
$300
      BULK RATE
POSTAGE & FEES PAID
         EPA
   PERMIT No. G-35
EPA/6QO/SR-97/071

-------