\",
United States
Environmental Protection
Agency
Environmental Research
Laboratory
Athens GA 30601
Research and Development
EPA/600/S3-86/042 Nov. 1986
Project Summary
Anaerobic Transformation
RECEIVEl5rocesses: A Review of the
Microbiological Literature
: 0 l r 1986 a
This review evaluates and summa-
rizes available information about the
role of microorganisms in the anaerobic
transformation of xenobiotic com-
pounds in natural environments. The
review focuses on the anaerobic trans-
formation of xenobiotic compounds
and natural structural analogs. A dis-
cussion of the anaerobic degradation of
benzoate is used to introduce the
anaerobic microorganisms important
to the degradation of xenobiotics in the
environment. This is followed by a re-
view of the degradation of several com-
pound classes including hydroxy and
methoxy substituted and lignoaromatic
compounds, halogenated aromatic and
aliphatic compounds, aromatic and
aliphatic hydrocarbons, nitrogen con-
taining compounds, and ether linked
compounds. A comprehensive list of
compounds known to be degraded
under anaerobic conditions is pre-
sented in tabular form. A brief discus-
sion of environmental factors that may
affect anaerobic degradation processes
is presented along with a review of
methods important to measuring the
rate of disappearance of xenobiotics in
environmental samples.
Several common degradative path-
ways that appeared to be shared by all
bacteria and bacterial communities ca-
pable of anaerobic degradative activity
were identified. Ring reduction pre-
cedes ring cleavage in the degradation
of aromatic compounds. Substftuents
of aromatic compounds are generally
removed to produce benzoate or phe-
nol before ring reduction and ring cleav-
age occurs. In most substrates, includ-
ing sewage sludge, sediments, and
aquifer material, degradation results
from the activity of microbial consortia
having methanogenic and sulfidogenic
organisms acting as the final electron
acceptors of the community. The re-
moval of ring substituents, ring reduc-
tion and ring cleavage is generally per-
formed by fermentative members of
the consortia. Despite these advances
there is a lack of information on the
turnover rates of hazardous organic
chemicals in anaerobic environments.
Work in this area, however, may be fa-
cilitated by the increasing number of
aerobic studies directed toward predic-
tion of the persistence and concentra-
tion of toxic chemicals in natural envi-
ronments.
This Project Summary was devel-
oped by EPA's Environmental Research
Laboratory, Athens, GA, 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
Anaerobic and aerobic degradation of
hazardous organic chemicals are
markedly different processes. This dif-
ference is particularly evident in the
degradation of aromatic compounds.
Anaerobic degradation requires exten-
sive reduction of the ring nucleus before
ring cleavage. Benzoate, for example, is
converted to cyclohexanecarboxylate
by a six-electron reduction in the initial
steps of anaerobic degradation. Aerobic
processes, on the other hand, make di-
rect use of molecular oxygen to hydrox-
ylate the ring before ring cleavage, a
process that also requires interaction
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with molecular oxygen. Benzoate in this
case is initially oxidized under aerobic
conditions to catechol, which is subse-
quently cleaved by the incorporation of
molecular oxygen to produce cis,cis-
muconic acid.
Progress toward understanding the
biochemical details of the anaerobic
degradation of xenobiotics has been
slow because of the experimental diffi-
culties attending the use of communi-
ties of microorganisms. For the most
part, anaerobic degradation of xenobi-
otic compounds generally requires the
concerted effort of several metabolic
groups of bacteria. The initial group fer-
ments the parent compound to prod-
ucts such as saturated fatty acids, H2
and C02. An intermediate group, the
proton-reducing acetogenic bacteria,
converts the long chain fatty acids to
acetate, propionate, C02, and H2. The
terminal metabolic group includes the
many methanogenic species and possi-
bly sulfur-reducing species that catabo-
lize the acetate produced by the other
groups to C02 and CH4, and rapidly use
the H2 produced to reduce C02 to
methane.
Although the importance of anaero-
bic consortia is well known, a number of
anaerobic organisms have been shown
to grow on substituted aromatic com-
pounds as the sole source of carbon.
Such organisms include the purple non-
sulfur bacteria such as Rhodopseu-
domonas palustris and Rhodopseu-
domonas gelatinosa, nitrate reducing
bacteria such as Pseudomonas (PN1)
and Moraxella sp. {N.C.I.B. 11086) and
sulfate reducing organisms such as
Desulfococcus multivorans, Desul-
fonema magnum, and Desulfosarsina
varaibilis.
In recent years, a number of aromatic
and aliphatic compounds have been
shown to be degraded by anaerobic
microorganisms and consortia. For ex-
ample, the lignoaromatic compounds—
ferulic acid, vanillin, cinnamic acid, pro-
tocate-chuic acid, and catechol—can be
converted to methane by anaerobic
bacterial consortia. Recently it has been
demonstrated that anaerobic microor-
ganisms in lake sediments and sewage
sludge degrade halogen-substituted
benzoic acid by eliminating halide ion to
produce benzoate, which can then be
metabolized to methane. Anaerobic de-
halogenation may be more important
than the corresponding aerobic proc-
ess. Recent work has shown that ben-
zene and toluene can be degraded to
methane and carbon dioxide by a
ferulic-acid-adapted anaerobic enrich-
ment. These compounds were gener-
ally believed to be resistant to microbial
attack in the absence of oxygen.
The relative importance of each
mechanism to the degradation of a
xenobiotic released to a particular envi-
ronmental niche will depend on the
chemical structure of the xenobiotic and
the physical/chemical characteristics of
the site. Microorganisms active in the
degradation of the xenobiotic will be
limited to those organisms that have or
can produce enzymes and possibly
transport systems that can recognize
and act on the xenobiotic. The closer in
structure a xenobiotic compound is to
the structure of a common natural sub-
strate, the more likely that it will be
transformed or degraded by a large va-
riety of microorganisms. These organ-
isms are only active if they are able to
grow and compete under the physical/
chemical constraints of the particular
environment. For example, in estuarine
sediments the microbial populations
are generally dominated by sulfate re-
ducing microorganisms. Alternatively,
in fresh water sediments where the con-
centration of oxidized sulfur species is
low, methanogenic microorganisms
dominate. If the dominating organisms
are capable, the xenobiotic has a good
chance of being rapidly degraded. Un-
fortunately, if minor populations, such
as methanogenic microorganisms in an
estuarine environment, are the active
members of the community, the chemi-
cal would most likely be degraded at a
slower rate.
Objectives
The purpose of this review is to evalu-
ate and summarize available informa-
tion concerning the role of microorgan-
isms in the anaerobic transformation of
xenobiotic compounds in natural envi-
ronments. Unfortunately, information
that relates directly to anaerobic trans-
formation of xenobiotics in the environ-
ment is limited. A large body of infor-
mation, however, does exist concerning
the anaerobic transformation (or degra-
dation) of these synthetic chemicals and
related natural compounds by pure or-
ganisms, mixed cultures, and adapted
and unadapted sewage sludge and sed-
iment systems. In addition to a compila-
tion of all the available data, this report
represents an attempt to relate the data
from tangential areas to the transforma-
tion of xenobiotics in natural environ-
ments. To focus this review, only infor-
mation concerning the anaerobic
transformation of xenobiotic com-
pounds and natural structural analogs
was included.
Conclusions
The anaerobic degradation of poten-
tially hazardous chemicals released to
the environment has received increased
attention over the last 10 to 20 years.
This irtcr'eased interest'teas led to signif-
icant advances in identifying the princi-
pal microorganisms involved in the
anaerobic degradation process, unrav-
elling the complex degradative path-
ways used by these organisms, and
demonstrating the" potential for anaero-
bic degradation of a large number of
compounds. As these studies have de-
veloped, a number of key processes
have been recognized. Several common
degradative pathways appear to be
shared by all bacteria and bacterial
communities capable of anaerobic
degradative activity. Ring reduction pre-
cedes ring cleavage in the degradation
of aromatic compounds. Substituents
of aromatic compounds are generally
removed to produce benzoate or phenol
before ring reduction and ring cleavage
occurs. In most substrates, including
sewage sludge, sediments, and aquifer
material, degradation results from the
activity of microbial consortia having
methanogenic and sulfidogenic organ-
isms acting as the final electron accep-
tors of the community. The removal of
ring substituents, ring reduction and
ring cleavage is generally performed by
fermentative members of the consortia.
Despite these advances there is a lack
of information on the turnover rates of
hazardous organic chemicals in anaero-
bic environments. The lack of attention
apparently results from the difficulties
in working with bacteria and bacterial
communities under strict anaerobic
conditions, the long adaptation periods
before degradation is observed for
many compounds, and the statistical
problems inherent in working with
undisturbed sediments.
Work in this area may, however, be
facilitated by the increasing number of
aerobic studies directed toward predic-
tion of the persistence and concentra-
tion of toxic chemicals in natural envi-
ronments. A number of kinetic models
have been proposed to characterize
degradation in natural environments
and in pure cultures of microorganisms.
Many of these models are based on the
assumption that substrate disappear-
ance can be modeled with information
only on substrate concentration and
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population density together with
parameters of classical Monod kinetics.
Recently, models were described for the
kinetics of biodegradation of organic
compounds of bacteria growing on an
alternative carbon source. Statistical
means have been proposed for evaluat-
ing the fits of these models to data ob-
tained from laboratory studies.
The EPA author J. E. Rogers (see below) is with the Environmental Research
Laboratory, Athens. GA 30613.
The complete report, entitled "Anaerobic Transformation Processes: A Review of
the Microbiological Literature," (Order No. PB 86-230 042/AS; Cost: $11.95,
subject to change) will be available only from:
National Technical Information Service
5285 Port Royal Road
Springfield, VA22161
Telephone: 703-487-4650
J. £. Rogers can be contacted at:
Environmental Research Laboratory
U.S. Environmental Protection Agency
Athens, GA 30613.
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United States
Environmental Protection
Agency
Center for Environmental Research
Information
Cincinnati OH 45268
Official Business
Penalty for Private Use $300
EPA/600/S3-86/042
0000329
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