United States
Environmental Protection
Agency
Air and Energy Engineering
Research Laboratory
Research Triangle Park NC 27711
Research and Development
EPA/600/S7-85/050 Jan. 1986
4>EB\ Project Summary
Thermophilic Anaerobic
Biodegradation of Phenolics
Frank J. Castaldi and Barbara J. Hayes
This report presents the results of a
series of anaerobic microbial acclima-
tion and treatment performance tests
conducted with synthetic phenolic sub-
strates. The research is a feasibility
level assessment of substituting anaer-
obic biodegradation of phenolics for
solvent extraction. The tests demon-
strated the feasibility of biodegrading
phenol and p-cresol to methane under
thermophilic anaerobic conditions. The
experimental data indicate that anaero-
bic biodegradation of phenolics under
thermophilic conditions involves a dual
system of bioaccumulation and
biodegradation. Phenolics and un-
known metabolites were accumulated
in the anaerobic floes of sludge, and
these compounds were apparently
slowly degraded over time. Despite pe-
riodic upsets, treatment improved as
the phenolic loadings were incremen-
tally increased during the experiments.
This indicates that, although acclima-
tion of thermophilic anaerobic bacteria
to phenolics is difficult, it can be accom-
plished using normal microbial cultiva-
tion techniques. Moreover, degradation
intermediates identified during ther-
mophilic anaerobic treatment of pheno-
lics were similar to those found in efflu-
ents from anaerobic treatment systems
that operate at lower temperatures.
This Project Summary was devel-
oped by EPA's Air and Energy Engineer-
ing Research Laboratory, Research Tri-
angle 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
Fixed-bed coal gasification waste-
waters are generally high in biochemi-
cal oxygen demand. The largest fraction
of the carbonaceous biochemical oxy-
gen demand characteristic of these
wastewaters can be attributed to the
presence of phenolics. In addition to
phenols, the wastewaters contain ap-
preciable levels of cresols, xylenols, and
aromatic heterocyclic compounds with
nitrogen contained in the ring.
The chemistry of fixed-bed gasifier
wastewaters offers the potential for a
biological treatment alternative to sol-
vent extraction pretreatment of high
phenolic wastewaters. A conventional
wastewater treatment system for fixed-
bed coal gasification quench conden-
sates consists of tar/oil separation, sol-
vent extraction (phenolics removal),
steam stripping (ammonia and acid gas
removal), aerobic biodegradation, and
assorted tertiary treatments (specific to
discharge or reuse requirements). This
system is energy intensive and presents
both health and safety (worker expo-
sure to solvents) and hazardous materi-
als handling (phenolic by-product) con-
siderations.
This research is a feasibility level as-
sessment of substituting anaerobic
biodegradation of phenolics for solvent
extraction. This alternative treatment
system would consist of tar/oil separa-
tion, steam stripping, anaerobic bio-
degradation, aerobic biodegradation,
and assorted tertiary treatments. Anaer-
obic biodegradation would eliminate
many of the health and safety concerns
posed by solvent extraction treatment
while also producing methane as a by-
product. Methane may be generated in
sufficient quantity to meet the power re-
quirements of the entire alternative
wastewater treatment train.
The anaerobic treatment system
would provide simultaneous storage,
-------
equalization, and treatment of steam-
stripped quench condensates and other
process wastewaters. Since the process
would receive wastewaters at tempera-
tures in excess of 95°C, the anaerobic
biodegradations should occur in the
thermophilic temperature range (49 to
57°C) to minimize cooling requirements.
These temperatures are consistent with
equilibrium bioreactor temperatures es-
timated from heat balances on the
anaerobic process. Operation in the
thermophilic range would thus remove
much of the cooling requirement typical
of most mesophilic microbial treatment
processes. (Traditionally, hot waste-
waters are cooled by retention in large
aerated impoundment ponds. This
practice also permits the uncontrolled
release of volatile compounds to the at-
mosphere.) A thermophilic anaerobic
treatment process would restrict the re-
lease of volatile compounds Because
the emissions are controlled during
treatment. The major gaseous emis-
sions from the process are methane and
carbon dioxide. The thermophilic tem-
perature range should also optimize the
rate of microbial conversion.
Objectives
The purpose of this research was to
study the feasibility of biodegrading
phenolics to methane by thermophilic
anaerobic treatment. The study was a
feasibility level assessment which ex-
amined techniques for culturing ther-
mophilic bacteria, acclimating these mi-
croorganisms to phenolics, and
assessing toxicity thresholds for simple
phenol. The study was conducted with
bench-scale test reactors using syn-
thetic substrates composed of phenol,
p-cresol, organic acids, and basal salts.
Both continuous-feed and batch treat-
ment tests were conducted and treat-
ment performance monitored. Where
possible, degradation intermediates
were identified. These data provide the
basis for assessing the feasibility of
thermophilic anaerobic biodegradation
of phenolics.
Results and Conclusions
The report presents the results of a
series of anaerobic microbial acclima-
tion and treatment performance tests
conducted with synthetic phenolic sub-
strates. These tests were designed to
demonstrate the feasibility of biode-
grading phenolics under thermophilic
anaerobic conditions. Two types of ex-
periments were conducted:
• microbial seed acclimation tests
using mixed cultures of anaerobic
microorganisms grown under ther-
mophilic conditions, and
• treatment performance tests using
acclimated microorganisms sub-
jected to both continuous and
batch biodegradation conditions.
The experimental results are discussed
relative to the feasibility of achieving
thermophilic anaerobic biodegradation
of phenolics characteristic of most
fixed-bed coal gasification wastewaters.
Microbial Seed Acclimation
Testing
Mesophilic anaerobes from a munici-
pal sewage sludge digester were used
to seed two test bioreactors. These
sludges were fed a mixture of phenol
and organic acids and were subjected to
mixed liquor temperatures between 52
and 55°C. These two bioreactors were
operated throughout the acclimation
period at different substrate loadings.
Although the bioreactors were loaded
similarly, they performed differently be-
cause of differences in the nature of the
microbial cultures developed during
seed acclimation.
The microorganisms showed an in-
herent ability for thermophilic anaero-
bic biodegradation of phenol early in
the acclimation cycle. After three hy-
draulic residence times, a flocculent mi-
crobial mass developed that settled well
and produced an effluent that was low
in phenol. Overall chemical oxygen de-
mand (COD) reduction lagged phenol
removal: optimum COD removal was
not obtained until after about six hy-
draulic residence times. Methane was
detected in the off gas from the bioreac-
tor whenever these gases were sam-
pled during the acclimation period.
Although the bioreactors consistently
reduced the phenol concentration in the
wastewater, the apparent level of
methane produced did not keep pace
with the rate of phenol removal during
the early stages of the acclimation.
However, this was not unexpected be-
cause methane-forming bacteria are
usually slow to develop in most anaero-
bic treatment processes. The bioreac-
tors also maintained relatively low con-
centrations of volatile cell mass
throughout the test, and the apparent
rate of growth of new biosludge was
negligible.
Performance Testing
Performance testing consisted of two
continuous-feed biodegradation experi-
ments with similar populations of accli-
mated thermophilic anaerobic bacteria.
Each experiment was designed to track
the change in phenol, p-cresol, acetic
acid, and propionic acid in a test reactor.
Treatment performance data for one
test are presented in Table 1. This reac-
tor was fed phenol at 1000 mg/L,
p-cresol at 50 mg/L, and volatile organic
acid as equal quantities of acetic and
propionic acids (i.e., 200 to 400 mg/L).
The bioreactor was operated with an hy-
draulic residence time of 20 days and a
mean cell residence time of about
60 days. The system maintained a mass
loading of 0.04 mg phenol/mg total
volatile suspended solids (TVSS)/day
throughout the test run.
Test data indicate relatively uniform
treatment performance with the re-
moval of cresol and acetic acid. How-
ever, variable effluent qualities for phe-
nol and propionic acid may be the result
of several system specific factors. The
mass loading of phenol to the bioreac-
tor could change the equilibrium con-
centration within the mixed liquor by no
more than 50 mg/L for each day of oper-
ation. Therefore, an apparent rise in the
mixed liquor phenol concentration of
between 78 and 230 mg/L over a single
day of bioreactor operation must be the
result of factors other than the normal
changes in biochemical reaction rate
typical of microbial processes. One
such condition is the accumulation of
phenol within the floes of sludge before
actual biodegradation occurs. This ac-
cumulation probably results from sorp-
tion of phenol to the activated sludges
because of the lipophilic properties of
the floes. Therefore, the biofloc may
carry much higher concentrations of
phenol than actually are measured in
the liquid of the reactor. From time to
time, desorption may occur, liberating
phenol and unknown metabolites to the
reactor liquid. These compounds would
be quickly taken up by the biomass and
biodegraded in the normal course of
process operation. However, point-in-
time measurements will show effluent
quality variability as reported by the test
data. The accumulation and eventual
biodegradation of degradable pollu-
tants on floes is a common occurrence
in activated sludge treatment. Whether
this phenomenon presents problems
specific to thermophilic anaerobic treat-
ment of phenolics cannot be assessed
from the experimental data.
Conclusions
The following conclusions result from
the study of thermophilic anaerobic
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Table 1
Test
Day
1
4
5
6
7
8
W
11
12
13
14
15
16
17
18
19
20
22
25
27
Thermophilic Performance Test
Influent
Phenol
(mg/L)
_b
—
1030
-
-
7070
-
570
—
-
-
-
-
-
7000
-
7000
-
7020
-
Cresol
(mg/U
50
-
31
-
-
38
-
48
-
-
_
-
-
_
47
-
48
14
31
<5
Acetic
Add
(mg/L)
300
370
-
-
-
350
-
230
320
310
770
97
-
-
250
-
200
<5
-
-
Propionic
Acid
(mg/L)
280
310
-
-
-
440
-
230
490
240
760
775
-
-
300
-
590
<5
95
-
COD
(mg/L)
3040
3100
2970
3120
-
-
-
2440
-
3140
-
2890
-
-
2570
-
2960
2680
2810
Phenol
(mg/L)
<5
-
<5
79
<5
-
230
<5
55
70
6.2
<5
<5
75
790
720
<5
76
<5
733
Cresol
(mg/L)
<5
-
<5
<5
<5
-
<5
<5
<5
<5
<5
<5
<5
<5
-
<5
<5
<5
<5
<5
Effluent3
Acetic
Acid
(mg/L)
<5
<5
<5
<5
<5
<5
<5
<5
<5
<5
-
<5
<5
36
-
84
6
<5
<5
-
Propionic
Acid
(mg/U
97
-
96
-
94
-
703
770
86
770
-
88
70S
95
-
76
90
700
80
-
COD
(mg/U
170
-
237
234
724
_
570
790
775
750
726
734
753
282
-
350
270
204
702
224
"Reactor total volatile suspended solids (TVSS) equals 1400 mg/L during test.
bBlank (-) means parameter not analyzed.
degradation of synthetic phenolic sub-
strates:
• Experimental results indicate that
phenol and p-cresol can be metabo-
lized to methane under ther-
mophilic anaerobic conditions. This
microbial treatment process was
sustained for a period of 6 months
with continued improvement in
treatment performance. A phenolic
toxicity threshold for the microbial
process was not evident when the
degradations were performed with
acclimated microorganisms. Rela-
tively short acclimation periods
were required to achieve consistent
treatment performance (i.e., greater
than 90 percent removal of phenol)
at low substrate concentrations.
However, extended periods of accli-
mation (i.e., greater than six hy-
draulic residence times) were re-
quired to achieve acceptable
treatment at higher substrate (phe-
nolic) loadings. Generally, accept-
able treatment occurred coincident
with the development of a floccu-
lated biomass. However, the pres-
ence of dispersed microorganisms
signalled poor treatability. These
observations indicate that stable
treatment performance could be
maintained with conventional mi-
crobial acclimation techniques.
• The experimental data indicate that
thermophilic anaerobic biodegrada-
tion of phenolics involves a dual
system of bioaccumulation and
biodegradation. Phenolics and un-
known metabolites are accumu-
lated in the anaerobic floes, and
these compounds are released to
the liquid at various times during
treatment. Process control would
specify a condition that optimizes
the degradation of toxics in the liq-
uid phase while maintaining an
equivalent level of treatment for the
sludges. Operating under this con-
dition would minimize the produc-
tion of new cell mass.
• Although the test bioreactors expe-
rienced upset conditions in the early
stages of the acclimation study,
phenolic wastes were continuously
fed to the systems throughout the
upset periods. Despite these peri-
odic upsets, system performance
improved as the waste loadings
were incrementally increased dur-
ing the experiments. This indicates
that, although acclimation of ther-
mophilic anaerobic bacteria to phe-
nolics is difficult, it can be accom-
plished using normal microbial
cultivation techniques.
• The experimental observations indi-
cate that proportional increases in
reactor biomass relative to pollutant
loading will not guarantee that the
individual phenolic compounds and
intermediates will be uniformly de-
graded. This would imply that sys-
tem kinetics may not fit a conven-
tional microbial substrate utilization
model.
The experimental data indicate that
degradation intermediates identi-
fied during thermophilic anaerobic
treatment of phenolics were similar
to those found in effluents from
anaerobic treatment systems that
operate at lower temperatures.
-------
F. J. Castaldi andB. J. Hayes are with Radian Corp., Austin, TX 78766.
William J. Rhodes is the EPA Project Officer (see below).
The complete report, entitled "Thermophilic Anaerobic Biodegradation of
Phenolics," (Order No. PB 86-122 603/AS; Cost: $11.95. 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 and Energy Engineering 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
EPA/600/S7-85/050
0000339 PS
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