United States
Environmental Protection
Agency
Municipal Environmental Research ^
Laboratory "
Cincinnati OH 45268
Research and Development
EPA-600/S2-82-017 Sept. 1982
Project Summary
Nonvolatile Organics in
Disinfected Wastewater
Effluents: Chemical
Characterization and
Mutagenicity
R. L Jolley, R. B. Cumming, N. E. Lee, L R. Lewis, J. E. Thompson, W. W. Pitt,
M. S. Denton, and S. J. Hartman
The overall objectives of this research
program were to examine the effects
of three disinfectants on the chemical
composition and mutagenic activity of
wastewater effluents. The disinfec-
tants studied were chlorine, ozone,
and ultraviolet light.
Nine different treatment plant efflu-
ents were examined before and after
disinfection (Table 1); fractions of con-
centrates were isolated to determine the
identity of mutagenic components.
The investigation used the following
methodology: grab sample collection,
concentration by lyophilization, high-
pressure liquid chromatography (both
analytical and preparative scales), and
mass spectrometry.
Mutagenic activity was determined
largely by the Ames Bacterial Test
Method using two strains of organisms
(TA 1535 and TA 1538). On one
occasion, another test (in vivo un-
scheduled DNA synthesis in mice),
was used to determine whether the
sample constituents or metabolites
reached the germ cells and damaged
DNA in these cells.
No consistent pattern was noted in
this study. Interplant comparison
revealed considerable variability.
Moreover, samples collected at the
same plant at different periods varied
significantly in chemical composition
and in mutagenic activity.
This Project Summary was devel-
oped by EPA's Municipal Environ-
mental Research Laboratory, Cincin-
nati, OH. 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
The severity of waterborne disease
epidemics has been greatly reduced
because of disinfection treatment of
wastewater and potable waters, mainly
with chlorine. Recently, however,
harmful chloro-organics have been
established as byproducts of this
disinfection process. Ozonation and
ultraviolet light irradiation are two
disinfectants that are being considered
as viable alternatives. But first an
evaluation is needed of possible harmful
byproducts, like those produced with
chlorination.
Within the last decade, rapid progress
has been made in the analysis of volatile
organic constituents of water and
wastewater. The determination and
characterization of nonvolatile organics,
however, have not advanced as rapidly,
even though the greatest portion of
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Table 1. Sources of Effluent Samples
Name of
Treatment Plant
Oak Ridge West Wastewater
Treatment Plant
Oak Ridge East Wastewater
Treatment Plant
Lyons Bend Fourth Creek
Wastewater Treatment Plant
Moccasin Bend Wastewater
Treatment Plant
Marlborough Easterly
Advanced Waste Treatment
Plant
Upper Thompson Sanitation
District
Taft Center, USEPA Pilot
Wastewater Treatment Plant
Meander Creek Wastewater
Treatment Plant
Northwest Bergen County
Wastewater Treatment Plant
Location
Oak Ridge, TN
Oak Ridge, TN
Knoxville. TN
Chattanooga, TN
Marlborough, MA
Estes Park, CO
Cincinnati, OH
Mineral Ridge, OH
Waldwick. NJ
Type of
Treatment
Primary
Secondary13
Secondary*
Secondary*
Secondary0
Secondary0
Secondary*
Secondary*
Secondary*
Disinfection
Method
Chlorine
Ozone"
Ultraviolet
Chlorine
Ozone*
Ultraviolet*
Chlorine
Chlorine
Ozone
Chlorine
Ozone
Ozone
Ozone
Chlorine
Ultraviolet"
Type of
Wastewater
Primarily
residential
Primarily
residential
Primarily
residential
Residential
Industrial
Primarily
residential
Residential,
relatively clean
Residential and
industrial
Primarily
residential
Primarily
residential
"Conducted in laboratory tests.
"Activated sludge.
soluble organics in wastewater and
natural water fall into that category- An
understanding of the possible environ-
mental and health-related effects of
these pollutants based on definitive
data is essential.
Scope
This research program was developed
into four major tasks: (1) laboratory-
scale disinfection of wastewater effluent
to assess the chemical and mutagenic
effects of each disinfectant on the
effluent; (2) analysis of concentrated
samples from eight operational waste-
water treatment plants and one pilot
plant, representing all three disinfec-
tants and varying levels of domestic and
industrial wastewater sources; (3)
characterization and identification of
constituents separated from the effluent
concentrates of operating plants, and (4)
screening of concentrated effluents for
mutagenic activity. In the later phases of
the research program, mutagenic activity
was used as the criterion for character-
ization of the separated chromatographic
components.
Results and Conclusion
Disinfection of secondary effluent by
chlorination and ozonation was found to
destroy some nonvolatile organics and
to produce others, as illustrated in
Figure 1 showing high-pressure liquid
chromatograms of effluent before and
after treatment with three disinfectants.
This pattern was found to prevail in all
samples.
A few more mutagenic fractions were
found in chlorinated and ozonated
samples than in the corresponding
nondisinfected control effluent of most
plants sampled. But inconsistent data
prevent the conclusion that disinfection
caused this effect. Many mutagenic
fractions were isolated from a nonmu-
tagenic concentrate and vice versa.
Chlorination of primary effluent
produces far more mutagenic compo-
nents that does chlorination of secondary
effluent. This pattern was not observed
with ozonation. Ultraviolet irradiation
had only a slight effect on the chemical
composition of secondary effluent, but it
destroyed the mutagenic activity of
several fractions.
Samples collected from the same
plant on different dates showed varying
chemical compositions and mutagenic
components.
Table 2 lists the 103 components
identified in concentrates of the controls
and disinfected effluents. Seventy-
seven were in the mutagenic fractions,
and 48 were in the nonmutagenic
fractions, with 22 of these compounds
being common to both sets. None of the
identified compounds are known to be
mutagenic.
The full report was submitted in
fulfillment of Interagency Agreement
EPA-IAG-D7-01027, DOE 40-593-76,
by Oak Ridge National Laboratory under
the sponsorship of the U.S. Environ-
mental Protection Agency.
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Control
UV irradiated
Chlorinated
Ozonated
Concentrated
Untreated - 2350X
21,200 uW sec/cm2 ---- 2350X
0.5-mg/L residual ------ 2220X
8-mg Os/ L --------- -2220X
0 1 23 4 5 678 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32
Time, hr
Figure 1. Chromatograms of ultraviolet- absorbing constituents in secondary effluents disinfected with chlorine,
ozone, and ultraviolet light.
Table 2. Summary of Identified Compounds in High-Pressure Liquid Chromatography Fractions of Effluent Concentrates
Alcohols
4-Butoxy-1 -butanola
Cyclobutanol*
2,4-Dimethyl-3-pentanola
4-Methyl-2-propyl-1 -pentanol*
Aliphatic Acids
Diglycolic acid*1"
2,2-Dimethyl-3-hydroxybutyric acid*
3,2-Dimethyl-3 -hydroxybutyric acida
Glutamic acid"
3-Hydroxybutyric acid*
2-Hydroxyisobutyric acid*1*
3-Hydroxyisovaleric acid*
tsovaleric acid"
Ketoglutaric acida
Lactic acid3"*
Malonic acid*
2-Methyl-2-hydroxybutyric acid"
Muconic acid*
Oxalic acida'*
Propanoic acida
Amines
Canavanine*
N,N'-diethyl-N,N'-dimethyl-p-phenylenediaminea
Amino Acids
Alanine"'*
Aminoisobutyric acid*
Aspartic acid*
Cystine*
Glycine*
Histidine*
Homocystine*
Phenylalanine"
Serine"
Tyrosine*
Aromatic Acids
Benzole acid*'*
4-Chlorobenzoic acid"*
Dichlorobenzoic *
0-hydroxyphenylacetic acid*
Phthalic acid"
Phenylpuruvic acid"
2-Pyridinecarboxylic acid*
Carbohydrates
Erythrose*'*
Erythrono- 1.4-lactone*
Galactose*
Glucose*
Gluconic acid*
1 -Methylmannofuranoside*
2-Methylxylopyranoside*
Rhamnose*
Rib/no- 1,4-lactone*
Threono-1,4-lactone*"*
Xylono- 1,4-lactone*
Xylose*'*
Epoxide
2-Methyl-3-propyl oxiranea
Fatty Acids
Linoleic acida
Myristic acid*
Oleic acid*
Palmitic acid*'*
Stearic ac«/a'b
Tetradecanoic acid*
Hydrocarbons
Decane*
2,4-Dimethylhexanea
Dodecane*
Ethylindan*
4-Ethyltetradecane*
Methyleicosane*
5-Methyl-5-ethyldecane*
2-Methylhexadecane*
2-Methyloctanea
2-Methyl-1,3,6-trioxocane*
Nordodecane*
Pentacosane*
Pentadecane*
2,2,3,3- Tetramethylhexane*'*
Phenols
Phenol*'*
Phthalates
Di-n-Butylphthalate*
Diethylphthalate*
Diisobutylphthalate*'*
Polyols
Cyclohexane- 1.4-diof*
Dihydroxyacetone*
2,5-Dimethyl-3,4-hexanediol*
Erythritol*1*
« U.S.QOVERNMENTPBINTINQOFFICE: 19B2-559-017/0827
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Table 2. (continued)
Ethylene glycol^"
G/uc/fo/"*
Glycerol**
Inositol*
T,3-Propanediota*
Propylene gtyco/*
Xytitol*
Threitol^
Purines and Pyrimidines
Adenosine*
Caffeine*
6-Dimethyladenosine"
5-Hydroxyuridine"
Thymine*
Miscellaneous
2-Aminoethanola
Decylhydroxyl amine"
2,4-Dimethyl-2,4-disiffipentanea
3,7-Dioxa-2,8--disilanortane-2-one-2,2,8,8-tetramethy/a
L -ct-G/ycerophosphate"
Phenytacetamide"
Phenylpyruvic ox/me*
1,2,3- Trimethoxypentane"
3.6,9- Trioxa-2-silaundecane-2-3-dimethyla
Urea"
Identified in mutagenic high-pressure liquid chromatography fraction.
^Identified in a nonmutagenic high-pressure liquid chromatography fraction.
R. L Jolley, R. B. Gumming, N. E. Lee. L. R. Lewis, J. E. Thompson, W. W. Pitt, M.
S. Denion, and S. J. Hartman are with Oak Ridge National Laboratory, Oak
Ridge, TN3783O.
Charles I. Mashni is the EPA Project Officer (see below).
The complete report, entitled "Nonvolatile Organics in Disinfected Wastewater
Effluents: Chemical Characterization and Mutagenicity." (Order No. PB 82-
254 053; Cost: $18.00, 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
Municipal Environmental Research Laboratory
U.S. Environmental Protection Agency
Cincinnati, OH 46268
United States
Environmental Protection
Agency
Center for Environmental Research
Information
Cincinnati OH 45268
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