United States
Environmental Protectior
Agency
Municipal Environmental Research
Laboratory ' " *
Cincinnati OH 45268
EPA-600/2-79-163
December 1979
Research and Development
vvEPA
Investigations of
Biodegradability and
Toxicity of Organic
Compounds
-------
RESEARCH REPORTING SERIES
Research reports of the Office of Research and Development, U.S. Environmental
Protection Agency, have been grouped into nine series. These nine broad cate-
gories were established to facilitate further development and application of en-
vironmental technology. Elimination of traditional grouping was consciously
planned to foster technology transfer and a maximum interface in related fields.
The nine series are:
1. Environmental Health Effects Research
2. Environmental Protection Technology
3. Ecological Research
4. Environmental Monitoring
5. Socioeconomic Environmental Studies
6. Scientific and Technical Assessment Reports (STAR)
7. Interagency Energy-Environment Research and Development
8. "Special" Reports
9. Miscellaneous Reports
This report has been assigned to the ENVIRONMENTAL PROTECTION TECH-
NOLOGY series. This series describes research performed to develop and dem-
onstrate instrumentation, equipment, and methodology to repair or prevent en-
vironmental degradation from point and non-point sources of pollution. This work
provides the new or improved technology required for the control and treatment
of pollution-sources to meet environmental quality standards.
This document is available to the public through the National Technical Informa-
tion Service, Springfield, Virginia 22161.
-------
EPA-600/2-79-163
December 1979
INVESTIGATIONS OF BIODEGRADABILITY AND TOXICITY OF ORGANIC COMPOUNDS
by
Jan R. Dojlido
Institute of Meteorology and Water Management..
Department of Water Chemistry and Biology
Warsaw, Poland
Grant No. PR-05-532-15
Project Officer
Robert L. Bunch
Wastewater Research Division
Municipal Environmental Research Laboratory
Cincinnati, Ohio 45268
MUNICIPAL ENVIRONMENTAL RESEARCH LABORATORY
OFFICE OF RESEARCH AND DEVELOPMENT
U.S. ENVIRONMENTAL PROTECTION AGENCY
CINCINNATI, OHIO 45268
&ms&^si Protection
Street
60804
-------
DISCLAIMER
This report has been reviewed by the Municipal Environmental Research
Laboratory, U.S. Environmental Protection Agency, and approved for publication,
Approval does not signify that the contents necessarily reflect the views and
policies of the U.S. Environmental Protection Agency, nor does mention of
trade names or commercial products constitute endorsement or recommendation
for use.
ii
-------
FOREWORD
The Environmental Protection Agency was created because of increasing
public and government concern about the dangers of pollution to the health
and welfare of the American people. Noxious air, foul water, and spoiled
land are tragic testimony to the deterioration of our natural environment.
The complexity of that environment and the interplay between its components
require a concentrated and integrated attack on the problem.
Research and development is that necessary first step in problem solu-
tion, and it involves defining the problem, measuring its impact, and
searching for solutions. The Municipal Environmental Research Laboratory
develops new and improved technology and systems for the prevention, treat-
ment, and management of wastewater and solid and hazardous waste pollutant
discharges from municipal and community sources, for the preservation and
treatment of public drinking water supplies, and to minimize the adverse
economic, social, health, and aesthetic effects of pollution. This publica-
tion is one of the products of that research; a most vital communications
link between the researcher and the user community.
Increasing numbers of products of commerce are reaching waste disposal
facilities, and little is known concerning their effects on treatment. Some
of these compounds are toxic, and others have profound physiological effects.
The future dictates that all compounds reaching our surface waters should
be biodegradable. Knowledge of what compounds that are not biodegradable
will aid us in logically planning what changes we should make to eliminate
them or develop treatment processes that will remove them.
This report presents the results of a testing program of several organic
compounds for toxicity and treatability.
Francis T. Mayo
Director, Municipal Environmental
Research Laboratory
111
-------
PREFACE
This project was conducted within the frame of the Marie SkJrodowska
Curie Fund, a bilateral monetary arrangement for cooperative scientific
programs between Poland and USA.
This final report is a concise summary of four interim reports. The
interim reports were not published but are on file in the libraries of
Institute of Meteorology and Water Management, 01-673, Warsaw ul. Podlesna 61,
Poland and U.S. Environmental Protection Agency, Environmental Research
Center, Cincinnati, Ohio 45268. The interim reports, date and subject matter
are:
Report #1, January, 1976 - MEK
Report #2, November, 1976 - MEK, DMF and DMA
Report #3, February, 1978, - PNP, OCP, TCP, and DCDEE
Report #4, June, 1979 - Five FWAs
The various sections of this report were prepared by the following
authors:
Principal Investigator - Jan R. Dojlido, Ph.D.
Biodegradability Testing - Andrzej Stojda
- Elzbieta Gantz
- Jan Kowalski
Analytical Methods - Halina Bierwagen
Toxicity Testing - J6zef Wojcik, Ph.D.
- Bozena Slomczyriska
- Anna Gwiazdowska
IV
-------
ABSTRACT
The biodegradability and toxicity of twelve organic compounds were
investigated. They were:
methylethyl ketone (MEK)
dimethyl amine (DMA)
dimethyl formamide (DMF)
p-nitrophenol (PNP)
o-chlorophenol (OCP)
trichlorophenol (TCP)
dichlorodiethyl ether (DCDEE)
five various fluorescent whitening agents, all of the same
stilbene-cyanuric type, but with different substitutes (FWA).
The investigations of the biodegradability were carried out by three
methods, namely: (1) respirometric measurements, (2) tests in the river
water, and (3) laboratory activated sludge units.
MEK, DMA, DMF, PNP, OCP, TCP were all biodegradable. DCDEE was found to
be a biologically inert substance under conditions of the tests. DCDEE is
a volatile substance; therefore, it may be partially blown off in the aeration
chamber. The FWA's are fairly resistant to biodegradation; however, they do
not affect the overall treatability of wastewater.
Long term respirometric measurements using a "Sapromat" that prints out
the amount of oxygen uptake every hour was very useful in supplying data for
kinetic parameters and inhibiting effects of various compounds. The acti-
vated sludge tests gave overall treatability information and time required
for acclimatization.
The toxicities of the test substances were determined by bioassays
using fish Lebistes reticulatus and Daphnia magna. The toxicity of MEK, DMF,
TCP are low while PNP, OCP, TCP were fairly high to fish. The FWA's varied
from low to high toxicity to fish depending on the molecular structure.
The work has been accomplished within the Polish American agreement
Project PL-480, Grant PR-05-532-15 by the Institute of Meteorology and Water
Management, Department of Water Chemistry and Biology at Warsaw, Poland, for
the U.S. Environmental Protection Agency.
The work was finished in 1979.
-------
CONTENTS
Foreword
Preface ................. ............ 1V
Abstract .............................
Figures .............................
vi ^
Tables .............................. .
Symbols ....... ...................... X1V
YV1
Acknowledgements ............. ............
\
1. Introduction ...... ................. *
2. Conclusions ........ ................ ^
3. Recommendations ...................... ~
4. Background Information .................. 11
5. Experimental Procedures .................. 17
Respirometric Measurements ............... •*•'
River Model ...................... 18
Treatability Tests . .................. 18
Toxicity Tests ..................... 20
Supplementary Experiments ............... 21
Analytical Procedures ................. 22
6. Methods of Calculations .................. 25
Respirometric Data ................... 25
Biodegradability in River Water ............ 31
Treatability Data ................... 31
Toxicity Tests ..................... 33
Supplementary Tests .................. 33
7. Results of Biodegradation ................. 35
Methylethyl Ketone, MEK ................ 35
Dimethyl Amine, DMA .................. 38
Dimethyl Formamide, DMF ................ 45
Para-nitrophenol, PNP ................. 50
Ortho-chlorophenol, OCP ................ 55
Trichlorophenol, TCP .................. 61
Dichlorodiethyl Ether, DCDEE .............. 66
Fluorescent Whitening Agents, FWAs ........... 71
8. The Results of Bioassays ................. 84
Methylethyl Ketone, MEK ................ 84
Dimethyl Amine, DMA .................. 85
Dimethyl Formamide, DMF ................ 86
Para-nitrophenol, PNP ................. 86
Ortho-chlorophenol, OCP ................ 87
Trichlorophenol, TCP .................. 89
Dichlorodiethyl Ether, DCDEE .............. 89
Fluorescent Whitening Agents, FWAs ........... 90
References ............................ 94
vii
-------
initial concentration of the tested substance c
o
15 The curves from Figure 14 after linearizing transformation . .
16 The final carbonaceous BOD L as function of
FIGURES
Number
1 Laboratory activated sludge unit 19
2 Typical shape of BOD curve of a solution containing
nutrient medium and a specific organic compound 25
3 The typical BOD curve transformed by equation
z = log dy/dt 27
4 The dependence of final carbonaceous BOD L on the
29
5 The typical curve of the decomposition of a specific
organic compound in river water 32
6 The final carbonaceous BOD Lc as a function of
initial MEK content c . . ,c
o o«>
36
7 Lag phase duration At-^ as function of CQ
8 Dependence of the ratio r on the initial MEK
concentration c ,,-
o 36
9 Decrease of BOD and MEK during the respirometric test 36
10 Linearized BOD curve from Figure 9 36
11 The decomposition of MEK in river water 37
12 MEK removal by activated sludge . . 38
13 The treatability coefficient K of wastewater
containing MEK LUU _0
« jo
14 BOD curve, DMA decrease and NH% increase during
the respirometric test . . ." 39
39
initial DMA content c0 . .c 40
Vlll
-------
Number
17 The influence of the seed acclimatization on the ^
DMA biodegradation process
18 The inhibition time of glucose decomposition Ati and of
DMA decomposition At2 as function of the DMA initial ^
content c
19 The dependence of the ratio r on the initial DMA ^ ^
concentration CQ
42
20 The decomposition of DMA in river water
43
21 DMA removal by activated sludge
22 The treatability coefficient KCQD of wastewater ^ ^
containing DMA
23 BOD curve, DMF decrease and NH3 increase during the ^
respirometric test
24 The curve from Figure 23 after linearizing transformation ... 45
25 The final carbonaceous BOD L and the final ammonia
content as a function of initial DMF content CQ
26 The influence of the seed acclimatization on the ^
DMF biodegradation process
27 The inhibition time of glucose decomposition Atj_ and
of DMF decomposition At2 as a function of the DMF
initial content CQ • '
28 The dependence of the ratio r on the initial DMF ^
concentration c
29 The decomposition of DMF in river water 48
30 DMF removal by activated sludge
31 The treatability coefficient KCQD of wastewater ^
containing DMF
32 BOD curve and PNP decrease during the respirometric test ... 51
33 The curves from Figure 32 after linearizing transformation . . 51
34 The final carbonaceous BOD LC as a function of
initial PNP content CQ 51
IX
-------
Number
35 The influence of the seed acclimatization on the
PNP biodegradation process ................. 52
36 The inhibition time of glucose decomposition Atj_ and
of PNP decomposition At2 as a function of the PNP
initial content c ..................... 52
37 The dependence of the ratio r on the initial PNP
concentration c
o .................
38 The decomposition of PNP in river water ............ 54
39 PNP removal by the activated sludge .............. 55
40 BOD curves of nutrient medium with various concentrations
of OCP ........................... 56
41 The final carbonaceous BOD L as a function of initial
o
OCP content c
42 The inhibition time of yeast extract decomposition Atj
as a function of the initial OCP content c ......... 57
43 The OCP content decrease during the respirometric test .... 57
44 The decomposition of OCP in river water ............ 58
45 Overall treatability and OCP removal by activated sludge ... 60
46 BOD curves of nutrient medium with various concentrations
of TCP ........................... 62
47 The final carbonaceous BOD L as a function of initial
TCP content c ..... ? ................. 62
48 The TCP content decrease at the initial concentration
12 mg/1 TCP ......................... 63
49 The influence of adaptation on BOD process in the
samples containing 10 mg/1 TCP ............... 63
50 The decomposition of TCP in river water ............ 65
51 Overall treatability and TCP removal by activated sludge ... 66
52 BOD curve of the sample containing DCDEE ........... 67
53 Final carbonaceous BOD L as a function of initial
DCDEE content c . . ? • AT
o ...................... o/
-------
Number
54
55
56
57
58
59
60
61
62
63
64
65
66
67
68
69
70
71
72
73
74
75
76
DCDEE changes in river water
Nitrites and ammonia changes in river water
The volatility of DCDEE from aqueous solution
DCDEE concentration in the effluent from activated
sludge unit
BOD curves of the samples containing nutrient medium
and 100 mg/1 of various FWAs ....
The curve of activated sludge endogenous respiration
in the presence of FWA-1
The curves of activated sludge endogenous respiration
in the presence of FWA-3
Photolytic FWA-1 decomposition in river and
distilled water
Photolytic FWA-2 decomposition in river and
distilled water . .
Photolytic FWA-3 decomposition in river and
distilled water
Photolytic FWA-4 decomposition in river and
distilled water
Photolytic FWA-5 decomposition in river and
distilled water
The mortality of fish at various MEK concentrations
The mortality of fish at various DMF concentrations
The mortality of fish at various PNP concentrations
The mortality of fish at various OCP concentrations
The mortality of fish at various TCP concentrations
The mortality of fish at various DCDEE concentrations
The mortality of fish at various FWA-1 concentrations
The mortality of fish at various FWA-2 concentrations
The mortality of fish at various FWA-3 concentrations
The mortality of fish at various FWA-4 concentrations
The mortality of fish at various FWA-5 concentrations
Page
68
69
70
70
72
72
73
76
76
76
77
77
84
87
88
88
89
90
91
91
92
93
93
XI
-------
TABLES
Number Page
1 The Respirometric Data 3
2 The Decomposition of Substances in River Water 4
3 Photolytic FWAs Decomposition in Water (Light
Intensity 1000 Lux) 5
4 Treatability of the Tested Substances by Activated Sludge ... 5
5 Lethal Concentrations of the Tested Substances to
Lebistes reticulatus and Daphnia magna 6
6 The Conditions of Respirometric Measurements of
DMA Biodegradability 38
7 Kinetic Parameters of DMA Biodegradation in River Water .... 42
8 The Course of Activated Sludge Acclimatization to DMA ..... 44
9 The Conditions of the Respirometric Measurements of
DMF Biodegradation 44
10 The Conditions of Respirometric Measurements of
PNP Biodegradability 45
11 Kinetic Parameters of PNP Biodegradation in River Water .... 54
12 The Conditions of the Tests Made by Means of Sapromat 56
13 Kinetic Parameters of OCP Biodegradation in River Water .... 59
14 Conditions of Respirometric Measurements of
TCP Biodegradability 61
15 BOD Process in the Samples Containing Various
TCP Concentrations 62
16 The Initial and Final DCDEE Content and NO and NO
Increase During the Tests in Sapromat (tRe First Series) ... 67
Xil
-------
Number Pa§e
17 The Final FWAs Concentrations in the Samples Containing Initial-
ly One of the Five FWAs (100 mg/1). The Tests Lasted 10 Days. 73
18 BOD and Final FWA Content in the Samples Containing
ca. 2 g/1 of the Activated Sludge and the Addition
of One of the FWAs . 74
19 The Velocity v of the FWAs Decomposition in River and
Distilled Water Exposed to Light 77
20 The Adsorption of FWAs on Activated Sludge 79
21 The Treatability of FWA-1 81
22 The Treatability of FWA-2 81
23 The Treatability of FWA-3 82
24 The Treatability of FWA-4 82
25 The Treatability of FWA-5 82
26 The Chlorophyll "a" Content in the Culture of Chlorella
After 7 Days of Exposure to the Presence of MEK 85
27 The Mortality of Lebistes reticulatus to DMA 85
28 Chlorophyll "a" Content in the Culture of Chlorella
After 7 Days of Exposure to DMA 86
29 The Chlorophyll "a" Content in the Culture of Chlorella
After 7 Days of Exposure to the Presence of DMF 87
Xlll
-------
SYMBOLS
a - Henry's coefficient of the equilibrium between volatile substance
content in the liquid and gaseous phase
a and b - Langmuir's isotherm coefficients
c - concentration of tested compound, mg/1
cg - - tested substance effluent concentration, mg/1
CQ - initial concentration or the inflow concentration, mg/1
kn ~ kinetic coefficient of the nutrient medium biodegradation, decimal
ks - kinetic coefficient of the tested compound biodegradation, decimal
day'1
Kad - the coefficient of the FWAs removal by the adsorption and removal
with excess sludge g~« l-h"^-
Kb - the coefficient of the FWAs removal due to the biodegradation
g-h-h-1
KCOD - overall treatability coefficient, being the COD decrease in the
activated sludge unit per 1 mg/1 COD in the effluent per 1 g/1 MLSS
and per 1 hour of retention time, g^l-h"1
Kph ~ the coefficient of the FWAs removal by photolytic decomposition,
g-.-4-h-1
Ks - the tested substance removal coefficient, being the decrease of the
tested substance contents in the activated sludge unit per 1 mg/1
in the effluent per 1 g/1 MLSS and per 1 hour of retention time
g-^l-h-1
L - the BOD, remaining at the time t, mg/1 02
LC - the final carbonaceous BOD, mg/1 02
Lu - the unit oxygen demand, caused by I mg of the given substance,
mg 02/mg substance
m - MLSS, dry mass, g/1
N - NH3 content at the time t, mg/1 NH3-N
N£ ' - final NH3 contents, mg/1 NH3-N
NOD - the oxygen demand caused by nitrification, mg 02/1
R - inhibition effect (the ratio of treatability coefficient in the
test unit K,™ and control unit K )
ks - The relative rate of the tested substance biodegradation (in the
r = — monomolecular reaction phase), dimensionless
K
n
tQ - lag phase of the tested substance biodegradation in river water
xiv
-------
t - time needed by activated sludge to restore normal treatment of
1 wastewaters background in the presence of the tested substance
t? - time needed by activated sludge to decompose the tested substance
Atj - inhibition time of nutrient medium BOD in the presence of the
tested substance, days
At2 - inhibition time of the tested substance BOD, days
v - the biodegradation rate in river water in the phase of constant
rate, mg/l-h
y - BOD at the time t, mg/1 02
t - the retention time, hours
Q - the sludge age, days
xv
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ACKNOWLEDGMENTS
We would like to thank Project Officer Dr. Robert L. Bunch from
Municipal Environmental Research Laboratory, Cincinnati, Ohio for his
great help during the course of this project, for his comments and
suggestions to the methodology of work and evaluation of data.
Our thanks to Dr. Joseph F. Malina from the University of Texas,
Austin and Dr. Robert D. Swisher for their help as consultants of
the project.
Technical contributions to these studies were made by technical
staff of the Department of Water Chemistry and Biology of the Institute
of Meteorology and Water Management. Their assistance is sincerely
appreciated.
xvi
-------
SECTION 1
INTRODUCTION
The development of elaborate industrial societies in the United States
and in Europe, has led to proliferation of a vast number of complex chem-
icals for industrial, agricultural and domestic use. Some portion of these
compounds eventually find their way into municipal and industrial wastewaters.
Unless specifically removed by waste treatment processes, they ultimately
appear in receiving waters and water supplies, thus no longer is it suffi-
cient to remove biochemical oxygen demand to protect the oxygen resource of
the receiving water but individual organic compounds become a concern.
Along with the benefits of chemicals there are risks. Unfortunately,
exposure to some chemicals, sometimes in very small amounts, can lead to
tragic and irreversible biological effects including cancer (carcinogenesis),
transmissible genetic damage (mutagenesis), and birth defects (teratogenesis).
The negative effects of toxic organic substances are greater when the sub-
stances are resistant to biodegradability. New organic compounds introduced
into the water environment should be biodegradable, easily removed in treat-
ment and their residual easily degraded in water bodies. Knowledge of the
toxicity and biodegradability of organic compounds will aid in designing
wastewater treatment processes.
This report describes the testing of twelve compounds both for bio-
degradability and toxicity. The compounds tested were: methylethyl ketone
(MEK), dimethyl amine (DMA), dimethyl foramide (DMF), p-nitrophenol (PNP),
o-chlorophenol (OCP), trichlorophenol (TCP), 2,2'-dichlorodiethyl ether
(DCDEE) and five flourescent whitening agents (FWA) used as components of
household detergents.
-------
SECTION 2
CONCLUSIONS
Biodegradation and toxicity of twelve organic compounds were studied.
The biodegradation tests performed were respirometric measurements, river
model, and activated sludge model. Additionally, for some compounds supple-
mentary tests were made for evaluation of their volatility, photolysis and
adsorption on activated sludge. The toxicity was measured with use of fish
Lebistes reticulatus and crustacean Daphnia magna.
The obtained data permitted the calculation of several parameters char-
acterizing the properties of tested compounds. From the respirometric data,
the unit oxygen uptake LU caused by 1 mg of a given substance, was calculated.
The processing of BOD curves parameters gave three kinetic characteristics:
the time A t of the inhibition of the standard medium decomposition (yeast
extract or glucose), the time A t of the inhibition of the tested substance
decomposition, and the ratio r = Rs/kn of the Streeter-Phelps coefficients
corresponding with the tested substance decomposition ks and standard
medium decomposition kR. This parameter indicates, whether the tested
substance biodegradation is faster r > 1 or slower r < 1 than the bio-
degradation of the standard medium.
The river model tests indicate the lag phase duration to, which is the
time of acclimatization of river water biocenosis. The same data permitted
the calculation of the rate of the tested substance decomposition v, mg/l-h,
during the phase of the constant rate of the decomposition.
The tests with the activated sludge supplied data on the acclimatization
time ti, after which the activated sludge normally decomposes the wastewaters
background, and ^2, after which also the tested substance is decomposed.
The treatability of wastewaters containing the tested substance was char-
acterized by means of K coefficients, one referring to the tested substance
removal KS and the second one referring to the overall treatment KCQD meas-
ured as COD decrease. Both coefficients are the decrease of tested substance
or of the COD, calculated per 1 mg/1 in the effluent, and per 1 g/1 MLSS
and per 1 hour of retention time. The inhibiting effect of the tested
substance on the overall treatment of wastewaters was characterized by the
ratio R of KCQD values observed in the test unit and in the control unit.
The results of the tests are summarized in Tables 1-5.
-------
TABLE 1. THE RESPIROMETRIC DATA
Substance
MEK
DMA
DMF
PNP
OCP
TCP
DCDEE
FWAsx
Concentration Measured unit
test range oxygen demand, Lu
mg/1 mg C>2/mg subst.
50 - 800 2.12
40 - 900 2.13
80 - 440 1.40
10 - 100 1.30
5 - 1200 0
10 - 540 0.9
0.7 - 500 0
100 0
Kinetic parameters at
concentration CQ
co
mg/1
50
800
300
450
80
440
50
100
10
200
1200
12
90
0.7
500
100
At!
days
0.5
1.0
0
0
0
0
0
4
0
0.5
> 11
0
0
0
0
0
At 2
days
0.5
1.0
5
> 13
3
4.5
0.8
> 15
2
> 11
> 11
5
> 18
7
> 16
the
r
1
0.2
0.75
0
0.8
0.8
1.5
0
0.3
0
0
n.dxx
0
0
0
x The tests with all FWAs gave the same results.
not possible on the basis of respirometric data
demand, despite the partial FWAs decomposition.
The determinations of At2 and r were
because of the lack of the oxygen
xx not determined.
-------
TABLE 2. THE DECOMPOSITION OF SUBSTANCES IN THE RIVER WATER
Substance
MEK
DMA
DMF
PNP
OCP
TCP
DCDEE
FWAsx
Concentration
mg/1
20
22
28
5
20
2
20
5
15
3
7
10
Kinetic parameters,
biocenosis not acclimatized
to
days
0.8
1.3
2
0
1.4
13
22
6.5
5
> 18
> 18
> 30
V
mg/l-h
0.6
0.4
0.55
0.1
0.35
0.02
0.10
0.13
0.27
0
0
0
Kinetic parameters, biocenosis
previously acclimatized
t0
days
0
0.8
0
0
0
0
0
0
2
_
-
V
mg/l-h
1.2
0.55
0.8
~ 0.3
~ 0.5
0.03
0.10
0.20
0.38
-
In the absence of the light.
-------
TABLE 3. PHOTOLYTIC FWAs DECOMPOSITION IN WATER
(LIGHT INTENSITY 1000 LUX)
Substance
Concentration
mg/1
Rate of photolysis
in river water,
mg/l-h
Rate of photolysis
in distilled water,
mg/l'h
FWA-1
FWA-2
FWA-3
FWA-4
FWA-5
10
50
10
50
10
50
10
50
10
50
5
6
9
13
1.5
5
2
4
5
7
7
11
14
13
5
5
8
9
11
12
TABLE 4. TREATABILITY OF THE TESTED SUBSTANCE BY ACTIVATED SLUDGE
Substance
mg/1
days days g'-'-l'h-l
MEK
DMA
DMF
PNP
OCP
TCP
DCDEE
FWA-1
FWA-2
FWA-3
FWA-4
FWA-5
200
400
20
135
70
5
200
5
200
5
25
5
10
10
40
6
200
5
80
4
44
4
40
0
0
0
14
38
0
0
0
-
0
-
0
0
0
0
0
0
0
0
0
0
0
0
8
9
6
12
38
~ 15
3*
6
-
0
-
_
-
0
0
_
-
0
0
-
-
_
-
high
high
high
high
high
high
0.7
0.2
0
high
0
0
0
2.1
2.5
0
0
0.03
0.01
0
0
0
0
1
1
1
~ 0.7
~ 1
~ 1
1
1
0
1
0
1
1
1
~ 1
~ 1
0.7
~ 1
0.3
~ 1
1
1
~ 1
xthe value is the additional time needed by the sludge previously acclimatized
to the presence of 40 mg/1 PNP.
5
-------
TABLE 5. LETHAL CONCENTRATIONS OF THE TESTED SUBSTANCES TO
Lebistes reticulatus AND Daphnia magna
LC50- 96 h to LC50- 24 h to
Substance Lebistes reticulatus Daphnia magna
MEK
DMA
DMF
PNP
OCP
TCP
DCDEE
FWA-1
FWA-2
FWA-3
mg/1
5700XX
60XX
1300XX
19
12
4.5
190
16
1800
110
mg/1
n.tx
ti
!!
tl
1!
1!
1!
22
3900
surpasses
solubility
FWA-4 3000 9400
FWA-5 6400 5600
x not tested
xx LC5o- 24 h
The obtained data lead to the following conclusions:
MEK is easily biodegradable. The respirometric data show that MEK
practically does not inhibit the microbiological activity and is biodegraded
at its initial concentration up to 800 mg/1. At the high concentrations,
the biodegradation rate is lower. The river model tests and treatability
tests confirm that MEK is easily biodegraded and does not affect the overall
treatability. The toxicity of MEK is low, its LC50 for fish Lebistes is
near 6 g/1. This is much more than the value, which could be expected in the
wastewaters.
DMA, is easily biodegradable at the concentrations not surpassing
300 mg/1. The biodegradation rate is rather high, and the time needed for
microbiological acclimatization is short. In the treatment by activated
sludge, DMA is easily removed from wastewaters and has little effect on the
-------
overall treatability of wastewaters. The toxicity of DMA is high. A con-
centration of 60 mg/1 of DMA causes 50 percent mortality of fish.
DMF is easily biodegraded. Its unit oxygen demand and unit amount of
NH3 involved in the biodegradation are not very high. The DMF biodegradation
in the rivers and its removal by means of activated sludge needs a period
for microorganisms acclimatization. The rate of DMF removal by acclimatized
sludge is high, and no harmful affect of DMF on the overall treatability
occurred. The toxicity of DMF is low. The value of LC50- h f°r fisn is
above 1 g/1.
PNP is biodegradable at concentrations up to 50 mg/1. At the higher
concentrations, 100 mg/1 and above, PNP is toxic and biodegradation does not
occur. In the river water at concentrations up to 20 mg/1, PNP is bio-
degraded at a rather high rate. The activated sludge removes PNP from
wastewaters with high efficiency. Also the overall efficiency of wastewater
treatments is not affected by the PNP presence. PNP is highly toxic to
higher animals; its LCso- 96 h is ca. 20 mg/1.
OCP is biodegradable only at low concentrations up to 10 mg/1. At
concentrations up to 200 mg/1 OCP is not biodegraded, but it does not inhibit
significantly the other biochemical processes. OCP concentration of 1200
mg/1 causes the poisoning of the microorganisms. OCP biodegradation in the
river water needs a long period of acclimatization, lasting 2 to 3 weeks.
Even after this period, the biodegradation rate is low. The activated
sludge removes OCP with low efficiency. The overall treatability is not
affected by 50 mg/1 OCP, but 100 mg/1 OCP causes the poisoning of the
activated sludge. The toxicity of OCP to fish is high, LC50- 96 h being
equal to 12 mg/1.
TCP is biodegradable at concentrations up to 35 mg/1, but needs a
certaln~period for microorganisms acclimatization. The TCP biodegradation
in river water proceeds at a rather high rate. The activated sludge
removes TCP from wastewaters containing up to 18 mg/1. This TCP content
does not affect the overall efficiency of the wastewater treatment, but
25 mg/1 causes poisoning of the sludge, and both TCP and COD removal
drop down to zero. TCP is very toxic to higher animals. Its LCsg- 96 h
to fish amounts to ca. 5 mg/1.
DCDEE is a biologically inert substance. No DCDEE biodegradation
was observed, even at a low concentration of 0.7 mg/1. DCDEE does not
affect the microbiological activity, even at concentrations as high as
500 mg/1. The only effect observed was the inhibition of the nitrification
partially occurring at 3 mg/1 DCDEE. The river model and results obtained
with use of the activated sludge model confirm these conclusions. DCDEE is
a volatile substance and may be partially blown off the wastewater in the
aeration chamber. The LC50- 96 h for fish is 190 mg/1.
FWAs do not cause any oxygen demand but they are partially decomposed
at thT^nditions of the respirometric tests. All FWAs are quite resistant
in the river water in the absence of light. Illumination of the FWAs
solutions causes their decomposition at a high rate. Both in the
-------
respirometric and the river water tests, the FWAs decomposition consist most
probably in transfiguration of the molecules or in their breaking into large
fragments.
In the treatment by the activated sludge, FWAs were resistant, except
FWA-1 which was biodegraded. FWA-3 was partially removed, but at a very
low rate. FWAs do no affect the overall treatability of wastewater until
their contents surpass 80 mg/1. The results of the bioassays show that
FWA-1 is the most toxic, the toxicity of FWA-3 is moderate and the toxic-
ities of other FWAs are low.
-------
SECTION 3
RECOMMENDATIONS
The biodegradation study of organic compounds in the respirometer, in
the river water model and in the activated sludge model gives useful infor-
mation on the condition of organic compounds removal in the treatment plant
and on their fate in river water. Those data supplemented by the toxicity
data can be useful in elaborating the criteria for the safe concentration
of organics in the discharged wastewater and the receiving water.
Based on the obtained results of the study, the general recommendations
formulated are:
MEK is a substance of weak toxicity and is easily biodegraded, there-
fore Its" harmfulness to the environment is limited. However, the BOD of MEK
solutions is high and may cause oxygen deficiency in the rivers polluted with
MEK or extensive BOD5 load in the wastewater. MEK does not contain hetero-
atoms in the molecules, therefore the biodegradation of the high MEK contents
will require the addition of mineral nutrients.
DMA is moderately toxic and easily biodegraded. The high BOD and the
ammoniT~development in the DMA biodegradation process may cause, in .rivers
polluted with DMA, an oxygen deficiency and an increase in ammonia.
DMF is weakly toxic and easily biodegraded by acclimatized microorganisms,
The treatment of wastewater, containing DMF, by means of the activated sludge
needs the acclimatization of the sludge to the presence of DMF.
PNP is a highly toxic substance. PNP biodegradation occurs in river
water~atf low concentrations. The wastewater containing PNP can be treated
by activated sludge after sludge acclimatization.
OCP is a highly toxic substance. OCP is biodegraded at low concentra-
tionsTrf the river water and in the activated sludge process, but at a low
rate. Long retention times might be needed.
TCP is the most toxic of all substances tested. The TCP biodegradation
proceeds" at low concentrations. It is suggested that severe limitation be
placed on TCP discharge into sewage systems and into rivers.
DCDEE is moderately toxic, but is a biologically resistant substance.
Its presence in the wastewater does not affect treatability, but DCDEE is not
removed except by the partial blowing off with the air in the aeration basin.
Special treatment may be needed.
-------
FWAs are weakly toxic except highly toxic FWA-1 and moderately toxic
FWA-3. In the treatment process FWAs are resistant, except FWA-1 which is
removed at a high rate and FWA-3 which is removed slowly. In the natural
environment, FWAs are easily decomposed due to photolysis but this process
has not led to FWAs complete decomposition. Rather, photolysis causes FWAs
molecules to break into large fragments.
Not enough compounds were examined to form any conclusions as to any
relationship between chemical structure and biodegradability. It is recom-
mended that such be done. It will not be possible to test all the organic
compounds now found in wastewater and all the new ones that will be developed
in the future. Some chemical, physical or structural property or a combin-
ation of several parameters must be found that would lead to the prediction
of the behavior of organic compounds in wastewater treatment plants and
surface waters.
10
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SECTION 4
BACKGROUND INFORMATION
Methylethyl ketone CH3.CO.C2H5 is a liquid boiling at 81 °C; one liter of
water dissolves 292 g of MEK at 20 °C. MEK is used in industry as a solvent.
MEK is found in some industrial wastes (e.g., in the waste from production
of synthetic leather Corfam, paints and lacquers).
MEK in concentrations up to 50 mg/1 does not influence the biochemical
processes (67).
According to Turnbull (61), the TLm for fish Lepomis macrochirus is
5640 mg/1 and a noticeable influence on these fish was found at 3380 mg/1.
TLm for fish Gambusia affinis was found to be 5600 mg/1 (64). Bringmann and
Meinck (9) determined the toxic concentration of MEK for bacteria Pseudo-
monas as 2.5 g/1, for algae Scenedesmus as 12.5 g/1, and for protozoa
Colpoda as 5 g/1. The experiments performed by Wjodek (70) have shown that
LC50- 48 h for Asellus aquaticus is 2850 mg/1. WXodek proposes that the
permissible, safe concentration is 28.5 mg/1.
The following methods of MEK determination are known:
- Gas chromatographic method (after distillation with butyl alcohol). The
limit of detection is equal to 0.004 mg/1 (12).
- Spectrophotometric method based on the reaction of MEK with vanillaldehyde.
The developed color is measured at 600 nm (62).
- Redox titration with the iodine in alkali solution (59).
- Amperometric titration with hydroxylamine hydrochloride in a waterless
solution of pyridine and isopropyl alcohol (7).
Dimethyl amine CH5.NH.C% is a gas condensing at 7 °C. DMA is easily
dissolved in water, forming an alkali solution. DMA is found in some
industrial waste (e.g., from the production of synthetic leather Corfam). It
is used in the synthesis of many substances.
The toxic influence of DMA on higher organized life is more marked than
on lower organisms. Toxic concentration for fish Leuciscus cephalus is 30 -
50 mg/1 (19) whereas for algae Scenedesmus and for protozoa Colpoda it is
250 mg/1. A toxic influence was found for bacteria Pseudomonas at the
concentration 1 g/1 (38) . Corti (13) observed that DMA in concentration
of 205 mg/1 influences fish Salmo irideus after 33 - 40 sec and death
11
-------
occurred after 10 - 13 min. Water Quality Criteria (66) states 0.7 mg/1 DMA
changes the taste of the fish meat of trout. Dzhanashvili (15) proposed the
value of permissible concentration of DMA as 0.1 mg/1.
DMA is determined usually by GC method (17), (30), (41). Kubelka (30)
set the detection limit of DMA in water as 0.01 mg/1. Other methods of DMA
determination in water are as follows:
- Spectrophotometric method with carbon disulfide and ammonia solution of
cupric acetate (10), (52).
- Fluorometric method by the exciting with light of 365 nm and by the
measurement of the emitted fluorescent light at 535 nm (650 . The samples
are prepared by use of the thin layer chromatography.
- Polarographic method (1).
Dimethyl formamide (CH3)2N.CHO is a liquid boiling at 150 °C. DMF is
soluble in water in any ratio. DMF is used in chemical industry as solvent.
It occurs in wastewater from the production of synthetic leather Corfam.
According to Zamyslova and Smirnova (74) DMF in concentration 10 - 500
mg/1 does not change the BOD of water and wastes. High amounts of DMF in
water causes an increase of ammonia. Permissible concentration of DMF in
receiving water in the Soviet Union is 10 mg/1.
The bioassays of DMF with Asellus aquaticus (73) indicated that LCcn-
48 h is equal to 9628 mg/1.
DMF contents in water is usually determined by the gas chromatographic
method (30), (34).
The Spectrophotometric method is based on the reaction of DMF with
picric acid forming a yellow colored complex (11).
p-Nitrophenol is a solid compound, melting at 114 °C and boiling at
279 °C, sublimating at a temperature below b.p. Depending on pH, PNP may
exist in two tautomeric forms: the benzene or quinone form:
pH
2
high pH
0 = N - OH
benzene form quinone form
PNP is used for wood impregnation. It is used also as raw material in
production of pesticides and azo dyes.
Biodegradation of PNP was studied by Fitter (47), (48). Comparative
investigation of various nitro phenols showed that biodegradation decreases
12
-------
with the increase of nitro groups on the molecule. The highest rate of
biodegradation was shown by the nitrophenols with the nitro group in the
para position. Fitter (48) measured the rate of PNP biodegradation with
use of COD determination and found it equal to 17.5 mg COD/g-h.
The following methods of PNP determination are known:
- Spectrophotometric method based on reduction of nitro groups to amine
groups and ,by diazotization, the dye is developed and measured (44).
- Spectrophotometric method based on development of yellow colored sodium
pseudosalt of nitrophenol in presence of sodium hydroxide (4), (45), (56).
- Polarographic measurements in the presence of methanol and sodium hydroxide
made at Ej/2 =0.98 - 1.05 V (73).
- Gas chromatographic method (8), (26), (55).
- Thin layer chromatographic method (40), (42), (58).
- Phosphorescence method (37).
- Atomic absorption spectrophotometry (AAS) method based on the creation of
a complex compound of PNP with cobalt and determination of cobalt by AAS
method (39).
o-Chlorophenol is a liquid melting at 9 °C and boiling at 175 °C. The
solubility in water at 20 °C is equal to 28 g/1. OCP has
very strong antiseptic properties. It is used in produc-
tion of antiseptic agents and pesticides. Additionally
OCP can be developed in water during the chlorination
process.
Ingols (22), (23), (24) performed an extensive study on the degradation
of chlorophenols by means of activated sludge. He found that the persistence
of chlorophenols increases with the increase of the number of chlorine atoms
on the molecule and depends on the isomeric position of those atoms in the
aromatic ring. It was found that OCP in concentration 100 mg/1 was fully
degraded within 3 days. Fitter (47), (48) found the rate of OCP biodegrada-
tion by activated sludge, determined on the basis of COD measurement, was
equal to 25 mg COD per hour and per 1 g of activated sludge.
Lammering and Burbank (31) found that the introduction of chlorine into
phenol ring increases the toxicity. The toxicity of chlorophenols increases
with the increase of the number of chlorine atoms in the phenol ring (13), (63),
(66). Ingols (22), (23) found that TLm at 23 °C for fish is equal to 58 mg/1.
OCP content can be determined by the following methods:
- Spectrophotometric method based on the formation of the indophenol dyes
in the reaction of phenols with 4-aminoantipyrine (21), (53), (56). It is
the method used most commonly for determination of phenols.
13
-------
- Spectrophotometric method based on reaction of chlorophenols with diazo-
tizated p_-nitroamine and magnesium ions (72) , (32) .
- Titrimetric method based on bromination or chlorination of aromatic ring
and iodometric titration of the excess oxidation agents (3) , (29) , (49) .
- Gas chromatographic method (2), (27), (46).
Tr i ch 1 oropheno 1 is a solid compound, melting at 67 °C and boiling at
244 °C and soluble in water in the amount of 0.8 g/1 at 20 °C. The use of
TCP and occurrence in wastes and water is similar to OCP.
Biodegradation of TCP was studied by Ingols (23) . In the tests lasting
two days TCP was totally degraded at a concentration of 50 mg/1, but at a
concentration of 400 mg/1 TCP, the biodegradation was completely inhibited.
According to Ingols (23) the TLm at 25 °C for fish is equal to 3.2 mg/1.
TCP can be determined by the following methods:
- Spectrophotometric method based on the formation of indophenol dyes in
reaction of phenols with 4-aminoantipyrine (2) , (53) , (69) .
- Thin layer chromatographic method (68), (75).
- Gas chromatography (20), (43), (51), (50).
2,2'-dichlorodiethyl ether a^Cl.O^O.Ct^.O^Cl is a colorless liquid
boiling at 178 DC. The solubility in water is 1 g/1 at 20 °C. DCDEE is used
as an insecticide, as cleaning agents and as a solvent. It is a byproduct
in the production of the synthetic fibers, glycoles, antifreezing agents and
some pesticides (36) . Meager data exist on toxicity of DCDEE (60) .
DCDEE is determined in water mostly by gas chromatography (14),
and gas chromatography-mass spectrometry method (6).
(28)
Fluorescent whitening agents are used for the improvement of the color
of the textiles, paper, plastics and other materials. They are added to
detergents used in households. FWAs are solid compounds, soluble in water
in the range 0.5 - 100 g/1 depending on the kind of FWA. In the frame of
the present work five commercial FWAs were studied. The samples contained
8 - 30% of the sodium chloride.
The structural formulas of five tested FWAs are given below:
H
FWA-l
H2f ^H2
H2C CH2
14
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I
Ul
I
-p-
* f-
2-rt
X
u>
o
o
\
O/ Wo''
x
ro
O
M ,
1
2-X
I
^^V
IV)
-------
Jensen (25) found that FWA can accumulate in fish. He analysed various
species of fish taken from rivers in Sweden and found measurable quantitites
of FWA in the muscle of fish. When fish were returned to water not con-
taining any whitener, they lost the FWAs very quickly (57).
The study of toxicity of FWAs for trout showed that LCcn- 96 h ranged
from 108 - 1780 mg/1 (76).
The most common method for determination of the FWA in water is
fluorimetric method (16), (18), (33). The mercury lamp is used for
excitation with the wavelength of 370 nm, and the fluorescence is meas-
ured at 440 - 480 nm. Thin layer chromatography method is used also for
determination of FWA.
There are some advantages to the combination of the two above mentioned
methods, i.e. the separation of the FWA by use of thin layer chromatography
and the identification by means of fluorimetric measurement (18). The
combined technique of TLC with radiometric measurement and with IR-spectro-
photometry is also known (54).
16
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SECTION 5
EXPERIMENTAL PROCEDURES
RESPIROMETRIC MEASUREMENTS
Apparatus
The BOD tests were carried out by means of "Sapromat" respirometer.
The principle of measurement is the following: The sample of solution,
in which the BOD is to be measured, is put into the 0.5 1 flask in the
amount of 332 ml. The flask is then hermetically sealed. The oxygen uptake
in the solution causes a reduction of the pressure in the air space in the
flask which switches on the electrolytic oxygen generator.
The automatic counter records the amount of the produced oxygen, which
is equal to the amount of oxygen taken up by the test solution. This is
printed every hour on a paper tape.
In the air space of the flask, a basket is filled with soda lime to
absorb C02 produced in the biochemical process.
One apparatus is equipped with 6 or 12 independently operating flasks
and printers. All these flasks are kept in the same room at 20 ±1° C. Mag-
netic stirrers, individual to each flask, allow the equilibrium between
oxygen contents in the liquid and gaseous phase to be reached in a short
time.
Materials
The dilution water was prepared with tap water, previously aerated by
air during 24 h, and with addition of the following mineral salts: 8.5 mg/1
KH2P04, 21.75 mg/1 K2HP04, 33.4 mg/1 Na2HP04 - 7H20, 1.7 mg/1 NH4C1, 22.5
mg/1 MgS04 • 7H20, 27.5 mg/1 CaCl2, 0.25 mg/1 FeCl3 • 6H20.
The nutrient medium was prepared by dilution of 200 mg yeast extract
or glucose in 1 liter of the dilution water. The tested solutions were
prepared by adding the determined volumes of the tested substances stock
solutions into the dilution water.
As inoculum the following seeds were used:
- the municipal wastewater, obtained from the Warsaw sewerage and
aerated for 24 hours before use
- the supernatant sample from the laboratory activated sludge
installation
17
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- the activated sludge from the same installation.
In the tests of activated sludge respiration in the presence of FWAs,
the test solutions were prepared with settled activated sludge, to which the
FWAs solutions in the dilution water were added.
Measurements
In every case, the BOD was automatically recorded every hour during the
test lasting usually 1 to 3 weeks. Additionally, chemical analyses were
carried out at the beginning and at the end of the tests on the concen-
tration of the tested substance and, in some cases, the NHv, NO? and NO?
content. In some tests the concentration of the tested substance was
determined every day. A flask from six flasks containing the same solu-
tion was withdrawn daily from the apparatus and its contents analyzed.
RIVER MODEL
Apparatus
The experiments were carried out in 10-liter bottles, filled with the
river water and the known amounts of tested substances. The water in the
bottles was stirred mechanically at a rate causing the aeration conditions
similar to the aeration in the rivers. The water in the bottles was
exposed to the light of the fluorescent tubes (at the intensity of approx-
imately 1000 lux). The tests were performed in all day illumination or in
regulated illumination (12 h with light and 12 h in darkness).
Materials
The river water was obtained from the Vistula river upstream of
Warsaw, and next it was seasoned during at least 2 weeks in the 100 liter
polyethylene barrel. During this seasoning the water was aerated by
bubbling the air. The test solutions were prepared by adding the tested
substances stock solutions into this river water.
Measurements
From the river models, the samples were withdrawn periodically and
analysed. The tested substance concentration and in the certain tests
also NHg, N02 and NO^ content were determined. The frequence of sampling
and analyzing depended on the speed of biodegradation of any tested sub-
stance, usually it was made every day.
TREATABILITY TESTS
Apparatus
The test of the treatability of wastewater containing tested sub-
stances was carried out by means of a laboratory activated sludge unit,
presented schematically in Fig. 1.
18
-------
Figure 1. Laboratory activated sludge unit.
This unit consisted of a storage tank 1 (30 liter volume), containing
the wastewaters with tested substance, the peristaltic pump 2 feeding the
aeration chamber 3 with wastewater. The aeration chamber was separated
by the wall from the settler 4. The volume of aeration chamber was equal to
1 liter, the volume of settler 0.5 liter. The mixed liquor in the chamber
was aerated by means of the air pump 5.
The parameters of operation were:
- retention time: tr= 1 to 6 hours.
- sludge age:
- MLSS:
0=5 days.
m = 3 to 4 g/1 (dry mass).
Materials
The activated sludge was obtained from the municipal treatment plant
and acclimatized during 2 weeks in the storage unit, similar to the unit
presented above, but larger (volume of aeration chamber equal 8 liter).
The wastewaters were simulated by preparing a solution of 0.6 g/1 meat
extract, 0.12 g/1 NH4C1, 0.1 g/1 MgS04 • 7H20, 0.033 g/1 NaHC03 in the tap
water. BODr of these wastewaters was equal to ca. 360 mg/1 02; COD was ca.
420 mg/1 02. These wastewaters were used to feed the activated sludge
units. Usually 4 units were in operation simultaneously. The test units
were fed with the wastewaters, with addition of the tested substance at
known concentration. The control unit, operating simultaneously was fed
with wastewaters without any addition of test substances.
Measurements
The following analyses were carried out:
- the determinations of tested substance concentration in the waste-
waters flowing into the test units and in the effluent,
19
-------
- the determination of COD in the inflow and in the effluent,
- the MLSS (dry mass).
Additionally, the technological parameters were controlled. The
retention time was controlled by means of the effluent volume measurements
and the DO concentration was maintained at 2 mg/1 or higher.
TOXICITY TESTS
Two kinds of bioassays were carried out. The first was the deter-
mination of the lethal concentrations of tested substances, i.e. the
concentrations causing the death of 50 percent organisms used in the test
during 24 or 96 hours.
LC5Q- 24 h or LC50- 96 h.
The following organisms were used in the bioassays: the fish was
Lebistes reticulatus and the crustacean was Daphnia magna.
In the second type of bioassay, the influence of tested substances
on the changes of the chlorophyll content in the cells of algae Chlorella
was analyzed.
Tests with Lebistes reticulatus
The tests were carried out following the procedure described in
Standard Methods (53).
In every test, 10 test organisms were put into the 400 ml solution
of tested substance in the open vessel. The organisms dead after 24, 48,
72 and 96 hours were counted and removed to avoid the pollution of test
solutions. In this kind of test, the concentration of tested substance
may decrease slightly during the test.
Tests with Daphnia magna
In every test 10 organisms were put into the 200 ml solution of
tested substance in an open vessel. The organisms dead after 24 hours
were counted.
Tests with algae
The procedure of tests was based on a method recommended by EPA (5).
The tests with use of algae Chlorella consisted in the measurement of
the chlorophyll "a" content changes after 7 days of exposure to the
presence of the tested substance. The tests were carried out in the
300 ml conical flask, containing tested substances in 100 ml nutrient
solution, inoculated with 20 ml supernatant of algae culture. The
culture of algae was kept at 18 °C ±1 °C and illuminated with the
fluorescent tubes.
20
-------
Materials
All test solutions were prepared with use of activated carbon treated
tap water, pH equal 7.6 to 8, the calcium content 80 mg/1 and magnesium
content 15 mg/1. Before use, the water was aerated.
The tested substances were always added to the test solutions, using
the same stock solutions as in the biodegradability tests. The concen-
trations of tested substance in the test solutions represented always a
geometric series, e.g. 1.0, 1.8, 3.2, 5.6, and 10 mg/1.
Test organisms
The fish Lebistes reticulatus were selected before the tests, so that
all individuals were the same age (just before they revealed the sexual
dimorphism).
The population of the crustacean Daphnia magna used in the tests was
prepared in the following way. The matured females of Daphnia were put into
the aquarium in the amount of 20 to 30 individuals. After 1 to 2 days, the
new generation was hatched. After the next 5 days, these young organisms
were used for the tests.
The algae Chlorella sp. wild strain were raised with use of nutrient
solution pre'pared following the EPA standard (5). The test solutions con-
tained the same nutrient.
SUPPLEMENTARY EXPERIMENTS
The proper interpretation of the results of experiments needs some
information on the physico-chemical properties of tested substances. If
this information was not available in the literature, additional experi-
ments were carried out to obtain the needed data.
So, the following experiments were made:
- the tests of volatility of MEK, DCDEE and OCP from the dilute
aqueous solutions
- tests of photolytic decomposition of FWAs
- tests of adsorption of FWAs on the floes of activated sludge.
The tests of volatility were made by means of dry air bubbling through
the dilute solutions of tested substances and measurement of the loss of
the sample weight and the decrease of tested substance concentration in the
sample. Henry's coefficient of equilibrium between the concentration of
tested substance in the liquid and the gaseous phase was calculated.
The tests of photolysis consisted of the measurements of FWAs con-
centration changes in the solutions of FWAs in the distilled water,
21
-------
exposed to the light of fluorescent tube at the known intensity. As pH has
the influence on this process, the solutions contained buffers, so that pho-
tolysis was tested in the pH range 4 - 9. -
The tests of FWAs adsorption were made by means of measurements of FWAs
concentration decrease in the supernatant after the contact with activated
sludge lasting 1 hour. At the same time, the activated sludge dry mass was
determined which permitted the Langmuir isotherm coefficients to be
calculated.
ANALYTICAL PROCEDURES
COD was determined by use of dichromate method with addition of HgS04
and AgS04. The samples were kept at the boiling temperature under the reflux
condenser during 2 hours and after cooling they were titrated with the
solution of Mohr's salt in the presence of 1,10-phenanthroline-ferrous
sulfate complex.
The Kjeldahl nitrogen was determined by means of digestion with cone.
H2S04 without the former distillation of NH3. After digestion, the ammonia
was distilled off and determined by means of Nessler method. The Kjeldahl
nitrogen content was calculated by substraction of NH3 content in the raw
sample from the result of the determination.
Nitrites determinations were made by spectrophotometric method with
sulfanilamide and N-(l-naphtyl)-ethylenediamine.
Nitrates were reduced to nitrites by means of amalgamated cadmium and
determined as N02 by the method described above.
Mixed liquor suspended solids MLSS (dry mass) were determined by
filtering a known amount of mixed liquor through Whatman's filter paper and
drying at 105 °C until constant weight.
Special procedures
The tested substances were determined by means of procedures developed
in the frame of the present work. Three methods were used: GC, spectro-
photometric and fluorometric methods. All GC determinations were made by use
of the chromatograph Pye Unicam 104, equipped with flame ionization ""
detector. Spectrophotometric determinations were carried out by means
of Pye Unicam spectrophotometer SP 600 equipped with cells of 1 cm light
path. Fluorometric determinations were made by use of the fluorometer
Aminco.
Methylethyl ketone, MEK
Gas chromatographic method was used. The column, 2.1 m long and 4 mm
diameter, was filled with Porapak Q (polystyrene), coated with polyethylene
glycol 1000 as stationary phase. The carrier gas (nitrogen) flow rate was
30 ml/min. The temperature of column was 83 °C. 'The aqueous samples of
MEK were injected in the amount of 5 yl each. The retention time was
22
-------
9 min 45 sec. The area of the peaks was proportional to the MEK concentration
within the range 0.3 to 200 mg/1.
Dimethyl amine, DMA
The GC method was used. The column 2.7 m in length and 4 mm in diameter
was filled with Chromosorb T (PTFE), 30 - 60 mesh, with 20 percent addition of
Triton X - 100 and 2 percent addition of KOH. Carrier gas (nitrogen) flow
rate was 60 ml/min. The column temperature was 96 °C, the retention time
1 min 45 sec. The samples of DMA were treated with KOH solution to keep
pH ca. 12 and were injected into the apparatus as aqueous solution in the
volume of 5 yl. The detection limit was 1 mg/1 DMA.
Dimethyl formamide, DMF
DMF was determined by means of hydrolysis with use cf cone. HC1 (1+1)
and heating the sample on the boiling water bath under reflux condenser for
1 hour. After cooling, KOH was added to reach pH ca. 12 and next the DMA
involved was determined by GC method.
Para-nitrophenol, PNP
The spectrophotometric method was applied. The quinone form of PNP
forms with NaOH the yellow pseudo-salt intensively colored. The absorbance
of the light at the wavelength 405 nm is proportional to the PNP content
within the range 0.2 to 10 mg/1.
Ortho-chlorophenol, OCP
The spectrophotometric method was applied. The samples were treated
with the 4-aminoantipyrine and potassium ferricyanide. The involved
indophenol dye was measured spectrophotometricly at light wavelength 510 nm.
Direct determination was possible within a range of OCP concentration from
0.5 to 5 mg/1. The substances contained in the yeast extract, mineral
nutrients and in the river water did not interfere with the determinations.
Trichlorophenol, TCP
TCP was determined similarly as OCP with 4-aminoantipyrine method. The
absorbance of the formed dye was measured at the wavelength 510 nm. Direct
determination of TCP in the tested solutions is possible within a range
of concentrations 0.5 to 5 mg/1.
Dichlorodiethyl ether, DCDEE
DCDEE was determined by means of GC method. The column 2.1 m long and
4 mm diameter was filled with Chromosorb T 40 - 60 mesh, coated with
Triton X - 100 as stationary phase, added in the amount of 5 percent per
weight. The flow of the nitrogen as the carrier gas was 30 ml/min. At a
column temperature of 118 °C the retention time was 6 min 10 sec. The aqueous
solutions of DCDEE were injected in a volume of 10 yl. The detection limit
amounted to 0.2 mg/1 DCDEE.
23
-------
Fluorescent Whitening Agents, FWAs
The fluorometric method was applied. In this method, a UV beam passes
through the cell with the tested solution and the light emitted by the
solution at the direction rectangular to the UV beam is measured at the
wavelength 450 nm.
Turbid samples were previously cleared by means of centrifuging at
12000 rpm. As the measured signal depends on pH, phosphate buffer was used
to keep pH at the constant level of 7.2. The apparatus was standardized
with quinine sulfate solution. The calibration curves were prepared
individually for each FWA. The apparatus gave linear response within the
FWA concentration range 1 to 10 mg/1. The solutions with higher FWAs content
were adequately diluted before measurement.
24
-------
SECTION 6
METHODS OF CALCULATIONS
RESPIRCMETRIC DATA
Long Term BOD Curve Examination
If the sample, which BOD is measured, contains the nutrient medium
(yeast extract or glucose and mineral salts) with addition of the specific
organic compound, which biodegradability is tested, the resulting BOD curve
has usually the shape schematically presented on the Fig. 2.
BOD, mg/l 02
Figure 2. Typical shape of BOD curve of a solution containing nutrient
medium and a specific organic compound.
25
-------
The process of biochemical oxygen uptake can be divided into the
following subsequent phases:
- Phase A, being lag phase, during which oxygen uptake is very small or
none. This phase corresponds with the period of microorganisms accli-
matization to the conditions of test.
- Phase B, being growth phase, during which the microorganisms are
multiplying, so that oxygen uptake is accelerated. The curve is then
concave.
- Phase C, during which the organic components of the nutrient medium
are in increasing scarcity in comparison with amount of bacteria, so
that the process is restrained and the BOD curve is convex.
- Phase D, being second growth phase, during which the population of
bacteria, able to decompose the tested substance is growing, so that
oxygen uptake for tested substance decomposition is accelerated and
the BOD curve is concave.
- Phase E, analogous to phase C, during which the tested substance con-
tents is in the increasing scarcity in comparison with the amount of
bacteria able to decompose this substance. The BOD curve is then
convex.
- Phase F, during which the BOD accelerates once more because of the
oxygen uptake for nitrification of ammonia present in the tested
solution.
- Phase G, during which the BOD curve is approximately straight line
with little slope. This phase corresponds with the period, during
which the nutrients and the tested substance are almost completely
removed and BOD is caused mostly by endogenous respiration of micro-
organisms.
In almost all experiments made in the frame of the present work, the
following equation can be used to describe the BOD curve sections:
- lag phase (phase A), lasting to the moment t :
y a 0, 0 < t < t0,
where y is BOD at the moment t
- growth phase B, lasting from the moment t to t-, :
y = cn • 10n-0} t0 < t
where cn and bn are constants, the last being the exponential growth coeffi-
cient corresponding with the decomposition of the nutrient medium components.
26
-------
- monomolecular reaction phase C, lasting from the moment
described by the classic Streeter-Phelps equation:
to
where y, is the BOD at the moment t,, LI is the residual BOD at the moment tl
and k^ is the kinetic coefficient of the decomposition of the nutrient medium
components.
- the phases D and E are analogous to the phases B and C, but correspond
with the tested substance biodegradation:
y =
y =
3,
L[l -
where C and b are the constants, L3 is the residual BOD at the moment t3,
k is tile kinetic coefficient of tested substance biodegradation.
s
Nitrification phase F proceeds usually in some irregular way and was not
described with any equation. In the phase G (endogenous respiration), the
oxygen uptake rate is almost constant, equal Re:
y =
Re(t-t5)
The expression z = log dy/dt transforms any of equations to the linear
function of time t. Thus, the graph of the dependence of z_ on t has the
shape schematically presented on the Fig. 3.
log Re
time, days
Figure 3. The typical BOD curve transformed by equation z = log dy/dt,
27
-------
The BOD record made every hour enables one to calculate the BOD incre-
ments Ay, approximately equal to dy/dt, the results of BOD measurements can
be marked on the graph in the coordinate system: log dy/dt versus t. The
slope of any straight line section drawn through the marked points indicates
one of the coefficients bn, kn, bs or kg. Also the values of t^, t2 and t3
can be easily read out from this graph. The final carbonaceous BOD of the
tested solution can be calculated on the base of BOD recorded at the moment
t3, equal y^ and the maximum value of log dy/dt at the moment t?, equal zmax.
According to the above written equations, this maximum value z is equal:
zmax = log 2.303 L3 kg, thus: L3 = .^ fc - 10Zraa*
The final carbonaceous BOD L amounts to the sum of BOD measured up to
moment t, and BOD residual at the moment t?, so
L = y + U, and L = y_ + _ 1 .l
c 3 3 c 3 2.303 ks
The value /„ is recorded and the values k and z are read out from the
graph, so the value of L can be calculatid. max
In certain cases, when the nitrification phase F was superposed on the
phase E, the recorded BOD data were corrected by substraction of theoretical
oxygen demand for nitrification NOD from the recorded BOD data:
NOD = 3.2 • A NO + 4.3 A NO , where
A N02 and A N03 are the analytically determined increase of nitrites and
nitrates contents (expressed as mg/1 N) .
If the final carbonaceous BOD L was determined in the series of samples,
each containing different initial concentration c of tested substance, we
can make the graph presenting the dependence of L on c (Fig. 4).
The slope of the straight line drawn through the marked points indicates
the unit oxygen demand of tested substance L , i.e., the amount of oxygen
needed by the solution containing 1 mg/1 of the tested substance. L is
expressed in mg 02/mg substance. Apart from the mean value of L , the
confidence limits 95 percent were always calculated following the Bartlett's
equation. The intercept of the straight line indicates the oxygen demand
caused by organic components of nutrient medium and by the inoculum used in
the test.
It should be stated that in special cases, the shape of BOD curve differs
from the above given pattern. For example, if the tested substance is as
easily biodegradable as organic components of the nutrient medium are, the
phases D and E are superposed on the phase B and C, i.e., the decomposition
of the tested substance proceeds simultaneously with the decomposition of
nutrient medium components. If the tested substance is not biodegradable,
the phases D and E do not occur at all.
28
-------
Figure 4. The dependence of final carbonaceous BOD L on the initial
concentration of the tested substance c .
The Tested Substance Decomposition
In some tests, the tested substance contents were analyzed every day in
subsequent samples, initially identical. (BOD was also recorded.)
On the graph z = log dy/dt versus the time the supplementary values of
log c were added (where c is the tested substance concentration at the
moment t). It allows to confirm that BOD process during the phase D and E
is corresponding with the decrease of the tested substance contents. The
slope of the straight line log c = f(t) is also the kinetic coefficient of
tested substance decomposition kg.
In the cases when ammonia was one of the final products of tested sub-
stance decomposition, the values of log N£ were marked versus the time
(where N.p means final NH_ contents and N s"eans NH^ contents at the moment t) .
The slope of this line also indicates the kinetic coefficient kg.
The Choice of Parameters Characterizing the Biodegradability
Practice shows that many of the above described parameters are not use-
ful to characterize the biodegradability of tested substances. The reason is
that their values depend on the properties of the inoculum used in the tests,
and the control of these properties is very difficult. Even if the source
of the seed is always the same (e.g., the wastewaters sampled from the same
municipal sewage system), the quantity and kind of sewage microorganisms
29
-------
highly differs in different samples. As the results of experiment should
describe the properties of tested substance and not the random conditions of
the test performing, it was necessary to choose those parameters which are
reproducible.
It was found, that the best reproducibility can be obtained when the
relative parameters were chosen, namely, the differences or the ratios of the
parameters corresponding with the tested substance and the standard nutrient
medium biodegradation.
Thus, the influence of a given substance on the microbiological activity
can be determined by the calculation of the inhibition period of the nutrient
medium biodegradation. If t^(0) and t^(co) denote the moments of maximum
rate of nutrient medium decomposition, respectively in the blank sample not
containing the tested substance and in the sample containing initially the
tested substance at the concentration co, the inhibition time Atj is equal to:
Ati = Vco> - h^
The inhibiting influence of various concentrations of the tested sub-
stance on the yeast extract or glucose decomposition can be described by the
graph At-, = f(c_). The examination of this graph allows one to conclude
whether a given substance inhibits the microbiological activity and if so,
at what concentration it does.
The biodegradability of the tested substance can be determined by com-
parison with the biodegradability of nutrient medium components. The first
method is to calculate the inhibition time of tested substance decomposition
At2- It is equal to:
where t.r(co) is the moment of maximum rate of the tested substance decompo-
sition in the sample containing the initial concentration c , and t-^(O) is,
as formerly, the moment of maximum decomposition rate of the yeast extract
or glucose in the blank sample.
The data obtained in experiments with various initial concentrations CQ,
allow to prepare the graph At2 = f(c ). Examination of this graph leads to
the conclusion how much the given substance is more difficult to biodegrade
than the standard substance.
Another method is to calculate the ratio r of the values k corresponding
respectively with the tested substance decomposition and the decomposition of
the nutrient medium components:
k (c )
r = s °
kn(0) ,
where ks(c ) is the value of k coefficient read out from BOD graph,
corresponding with the phase E of the process in the sample containing
initial concentration co. The kn is the k value read out from the BOD
curve, phase C, concerning the blank sample (co = 0).
30
-------
As formerly, the dependence of r value on c value can be presented
graphically. If r = 1, we can conclude that the tested substance is decom-
posed at the same rate as organic components of the nutrient medium (after
certain period of microorganisms acclimatization). If r < 1, the biodegrada-
tion of tested substance is slower, and if r > 1, it is faster than the bio-
degradation of the nutrient medium components.
BIODEGRADABILITY IN THE RIVER WATER
The only information gathered during the experiments with the river model
was the results of determination of the tested substance concentration during
the tests. In several cases this information was supplemented by the data of
NH?, NC>2 and NO, contents during the tests.
It was found that the process of specific organic compound decomposition
in the river water can be divided into three phases. In the first phase, the
decomposition is slow or does not occur. This lag phase corresponds with the
period of acclimatization of microorganisms to the presence of the given sub-
stance. In the second phase, the decomposition of the tested substance pro-
ceeds at the approximately constant rate:
v = -dc/dt = const.
In the third phase, when the concentration c of the tested substance
drops down below certain critical value, the decomposition proceeds at a
decreasing rate and the concentration c decreases asymptotically to zero.
The process in this phase usually could not be exactly observed, because the
concentrations were close to the detection limit.
The above given pattern of process is illustrated by the typical graph
of a tested substance decrease in the river water as shown in Fig. 5.
As the parameters characterizing the biodegradation of tested substances
in the river water, the following ones were chosen:
- the time of lag phase t
- the decomposition rate v during the constant rate phase.
The values of the above given parameters can depend on the kind of micro-
organisms living in the river water used in the tests. However, it can be
assumed, that they are adequate to comparatively study the fate of all tested
substances in the river water.
TREATABILITY DATA
In the treatability tests, the inflowing wastewaters and the effluent
from the activated sludge unit were analyzed. The determined parameters were:
COD and tested substance content. Additionally, the MLSS (dry mass), the
retention time and D.O. content were controlled.
31
-------
c, mg/l
v=const
log phase
constant
-velocity_£hqse\velocity
the phase of
1 decreasing
time, days
Figure 5. The typical curve of the decomposition of a specific organic
compound in river water.
The simple method of treatment efficiency characterization is the cal-
culation of coefficient K, defined as below:
- c.
K =
mt
r-
where:
- c and c denote the concentration of the pollutants in the inflowing
wastewaters and the effluent, respectively, mg/l
- m denotes MLSS in the aeration chamber, g/1
- t.r is the retention time, hours
The treatment efficiency coefficient K determines the treatability of a
given substance by means of activated sludge, and its value is approximately
independent from the technological parameters of treatment process (MLSS and
retention time).
In the present work two values of K coefficient were calculated: the
value K , corresponding with the tested substance removal, and the value
characterizing the overall treatability of wastewaters. In this last case,
the COD values of the inflow and of the effluent have to be substituted into
the above equation for c and c .
32
-------
Usually, when c was increased, the values KCQD and Kg decreased, but
after certain time t°, KCOD reached a stabilized level. Similarly, the value
KS reached the constant level after the time t2. The values t-^ and t2 deter-
mine the time of sludge acclimatization which restores the normal activity of
microorganisms at the time t-^ and makes the microorganisms able to decompose
the tested substance after the moment t2.
As characteristics of the treatment process, the following parameters
were presented:
- times of acclimatization t-^ and t2
- overall treatability coefficient KQQQ
- the specific substance treatability coefficient Kg.
All these characteristics are the functions of tested substance content
in the inflow c .
TOXICITY TESTS
The 50 percent lethal concentrations LC50-96 h or 24 h were calculated
by use of the "probit" method. This method of calculation is based on the
assumption that the mortality of the test organisms is a Gaussian function of
the logarithm of the toxicant concentration. So, the observed mortality were
marked on the probability paper versus the logarithm of tested substance con-
centration and the best fitting straight line was drawn. LC5Q value was read
out as the abscissa, corresponding with the value of ordinate equal to 50 per
cent. To present the results of mortality determinations, the empirical
points were marked in the coordinate system: mortality (%) versus the tested
substance concentration, and next the best fitting log normal distribution
curve was drawn.
SUPPLEMENTARY TESTS
The Tests of Volatility
According to the Henry's law, the concentration of a given substance in
gaseous phase c is proportional to the concentration of this substance in
the liquid phase C]_: eg = a • Ci
If a volume dVa of the air is buobled through a volume Vw of solution,
the decrease of volatile substance concentration in the liquid is:
dcx = - cgdVa/Vw, or: dc-^^ = -a dVa/Vw.
At the same time, the bubbling of the air causes the evaporation of dVw, the
volume dVw of water, equal to:
0.018 is the molecular mass of water kg/mole
24 is the molar volume of water vapor at the test temperature 22
and at the normal pressure 760 mm Hg
33
-------
- 20 is the partial water vapor pressure at the test temperature mm Hg
- 760 is the atmospheric pressure, mm Hg
The rearranging of these equations leads to:
dc, _ dvw cj V
—L = 50.7 x 103 x a x -y- or log — = 50.7 x a x W6 log ^f- ,
cl w o o
where c0 and VQ are the volatile substance concentration and the liquid volume
at the beginning of the tests.
The results of the determinations of c-^ and the weighing of the solution
giving V^ values are marked on the graph log c^ versus log V'. The slope of
the obtained straight line equals the factor 50.7 x 103 x a,Wso the value a
can be calculated. ~~
The blowing off rate of the volatile, but not biodegradable substance,
from the aeration chamber can be calculated with use of mass balance equation:
co ' Vw = ce ' Vw + Va • cg , where
- V and V are the volume rate of inflowing wastewaters and the air
1/h.
- CQ, ce and c are the volatile substance concentration, respectively
in the inflow, in the effluent and in the air flowing off the chamber.
As c = a-ce, the volatile substance concentration in the effluent is
equal:
c
o
34
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SECTION 7
RESULTS OF BIODEGRADATION
METHYLETHYL KETONE, MEK
Respirometric Measurements
Three series of experiments were made. In the first series, BOD was
measured by means of Sapromat. The samples contained the nutrient medium
(yeast extract) with various MEK additions within the range 0 - 800 mg/1.
The test solutions were inoculated with municipal wastewater. In the second
series, both BOD and MEK content changes were observed in the samples with
initial MEK concentration equal 200 mg/1. After the second test was
completed, the third series was carried out with the same conditions except
that the solution remaining from the second test was used as inoculum (test
with the microorganisms acclimatized to the presence of MEK).
In the first series, in which different initial concentrations of MEK
c0 were applied, the phase of nutrient medium decomposition was superposed on
the phase of MEK decomposition.
are presented:
- final carbonaceous BOD (L(
In Figs. 6, 7 and 8 the following relations
as function of initial MEK content c0)
- the time of process inhibition Ati as a function of CQ
- the ratio r of the coefficient of MEK decomposition k to the
coefficient of nutrient medium components k .
2000
1500
1000
500
Figure 6. The final carbonaceous BOD L as function of initial MEK content cc
c ^*
35
-------
1.5
1.0
0.5-
A todays
200
400
660
800 .
c0. mg/IMEK
r = ks/kn
200
400
600 80C
c0, mg/l MEK
Figure 7. Lag phase duration At-,
as function of c0.
Figure 8. Dependence of the ratio r
on initial MEK concentra-
tion cQ.
400
300
200-
100
0
BOD. mg/l 03
— MEK x-
2nd test )/
ja^/\
c. mg/l MEK
200
3rd test
1 23450
100
1234
time, days
Figure 9. Decrease of BOD and MEK during Figure 10. Linearized BOD curve
the respirometric test. from Figure 9.
The results of the second and third series are presented graphically in
Figs. 9 and 10.
Discussion
It may be assumed that the basic reaction of MEK biodegradation follows
the equation:
CH3.CO.C2H5 + ii 02 »• 4 C02 + 4 H20
The stoichiometric value of oxygen uptake in the above given process of
MEK oxidation amounts to 2.44 mg 02/mg MEK. The experimental value (Fig. 6)
was determined as equal to 2.12 ±0.05 mg 02/mg MEK.
Comparing of experimental and stoichiometric values leads to the con-
clusion that ca. 87% of MEK is mineralized and the resting 13% is assimilated
by microorganisms to produce the biomass.
36
-------
Determination of inhibition time of biochemical processes in the pres-
ence of MEK leads to the conclusion that MEK at a concentration of 100 mg/1
and above causes the inhibition of the microbiological processes during 1 day
in comparison with the process in the blank sample.
The coefficient ks of monomolecular reaction of MEK decomposition
decreased at the initial MEK concentrations of 200 mg/1 and above. The
reason may be that at higher MEK concentrations the mineral nutrients may not
be adequate, thus slowing the process.
The acclimatization testing showed that the previous adaptation of the
seed used to inoculate the sample significantly shortens the lag phase.
Biodegradation in the river water
The results of MEK biodegradation in the river water are presented on
Fig. 11. The right side of the graph presents the MEK content changes
after the repeated addition of MEK to the water in which the first portion
of MEK was decomposed (i.e. to the water containing acclimatized micro-
organisms) .
c. mg/l MEK
t0=0
T
time, days
Figure 11. The decomposition of MEK in river water.
It is visible that MEK is easily biodegraded by the microorganisms
living in the river water, especially when they are previously acclimatized
to MEK.
Treatability tests
During the experiments two activated sludge test units and one control
unit were operating in parallel The first was fed with wastewater containing
200 mg/1 MEK; the second one 400 mg/1 MEK. The results of MEK content in the
effluent and the results of calculations of the overall treatability coeffi-
cient K are presented in Figs. 12 and 13.
It is visible that MEK is easily removed from the wastewater after
acclimatization of the activated sludge, lasting 8 to 9 days. The waste-
37
-------
c«, mg/l MEK
0 2 3 5 6 78
OA
0.3
0.2'
KCOD'9~'''h~
width ot contidenee strip
\Z c.=200mgVlMEK
c.=400mq/lMEK
C0=0i control chamber
Figure 12. MEK removal by activated
sludge.
0123456789 10
time, days
Figure 13. The treatability coeffi-
cient KQQP of wastewater
containing MEK.
water treatment (COD removal) is practically independent from MEK content
in the inflow. The differences between KCOD values in the control unit ai
in the test units were statistically not significant.
DIMETHYL AMINE, DMA
Respirometric measurements
Seven series of measurements were made. In the first three series
different initial DMA concentrations were applied, whereas in the remaining
series, the samples contained initially the same DMA concentrations.
TABLE 6. THE CONDITIONS OF RESPIROMETRIC MEASUREMENTS
OF DMA BIODEGRADABILITY
Number of
No. samples in
the series
I
II
III
IVa
IVb
V
VI
VII
5
6
8
4
4
9
6
5
Test
duration, Nutrient
days medium Seed
18
12
13
5
5
11
7
8
yeast extr. sewage
yeast extr. sludge
glucose sludge
" sludge
" adapted
sludge
" sludge
" sludge
" sludge
Range of
concentration,
mg/l
0 - 135
0 - 270
0 - 900
180
180
170
185
180
38
-------
The exemplary curves of BOD, DMA decrease and NH3 increase are shown in
Fig. 14 (the data resulting from series V). On Fig. 15 the same data are
presented in the linearizing coordinates system.
BOD, mq/l 02
c, mg/l N-NH3
c, mg/l DMA
BOD of nutrient medium
Figure 14. BOD curve, DMA decrease and NH3 increase during the respiro-
metric test.
8 9 10
time, days
Figure 15. The curves from Figure 14 after linearizing transformation.
39
-------
final carbonaceous BOD,
Lc, mg/l02
100
200 300
initial DMA concentration,
C0, mg/t DMA
2.5'
2.0
15
1.0
sample with the
adapted seed
time, days
Figure 16. The final carbonaceous BOD Figure 17,
Lc as function of initial
DMA content c^.
Figure 18. The inhibition time of
glucose decomposition
and of DMA decomposition
At2 as function of the DMA
initial content c~.
1.0
06
0.4
02
=ks/kn
X
Figure 19.
The influence of the seed
acclimatization on the DMA
biodegradation process.
/correlation coeticient 046 «
(not significant)
X
100
200
300
The dependence of the
ratio r on the initial
DMA concentration c.
The dependence of final carbonaceous BOD L on the initial DMA concen-
tration c0 is exemplified in Fig. 16 (data obtained in the II series). The
mean value of LC calculated from data obtained in all 7 series was equal to
1.56 +0.05 mg 02/mg DMA. The ammonia produced in the biodegradation of DMA
Nu calculated in a similar way was NU = 0.21 +_ 0.006 mg NH3-N/mg DMA. Fig. 17
presents the results of biodegradation tests in which the samples were seeded
with non-acclimatized and acclimatized inoculum to DMA.
The data obtained in all 7 series were marked on the graphs in a similar
way, and the moments of maximum BOD t-L and t2 were read out. Next, the
time of inhibition of the nutrient medium decomposition Atj was calculated
and presented in Fig. 18 as the function of initial DMA contents c . In the
same Fig. 18 the inhibition time of DMA decomposition At2 is presented also.
40
-------
Additional analyses of final DMA contents have shown that at initial concen-
trations up to 270 mg/1 DMA was completely decomposed; whereas, DMA decomposi-
tion at initial concentrations of 450 and 900 mg/1 proceeded more slowly.
The velocity of DMA decomposition in comparison with the velocity of
yeast extract or glucose decomposition is characterized by the values of r,
defined as r = ks/k , where ks is the coefficient of DMA decomposition, kn is
the coefficient of ^composition of glucose or yeast extract. The dependence
of r on initial DMA concentration co is shown in Fig. 19.
Discussion
According to the equation of DMA oxidation: (CH3)2NH + 302 «- 2C02 +
2HoO + NHx the unit oxygen uptake should amount to 2.13 mg 02/mg DMA, and
the unit ammonia involved should be Nu = 0.31 mg N/mg DMA. Comparison of
this last value with experimentally determined Nu = 0.21 leads to the con-
clusion that approximately 2/3 of DMA content is mineralized according to
the above equation and 1/3 is assimilated by the microorganisms.
The experimentally determined value of unit oxygen demand Lu is equal to
1.56 in comparison with the stoichiometric value of 2.13 mg 02/mg DMA. There-
fore, the most probable summary reaction equation is as follows:
2/3 mg DMA 1.42 mg 02_ 0<25 mg NH3 + 1.3 mg C02 + 0.53 mg H20
<"+ energy
0.14 mg 02_
1/3 me DMA T~-—I"** biomass + byproducts
1/0 m& U1U^ + nutrients r
The ratio of assimilated nitrogen coming from DMA to the final BOD is
equal to 1 mg NH3-N:15 mg 02 which is near to the value of N:BOD ratio
optimal in the treatment process.
The results of BOD determination show that DMA at concentrations up to
270 mg/1 do not inhibit the microbiological activity which is proved by the
horizontal course of graph At].. = f(c0) (Fig. 18). The DMA decomposition
proceeds with some delay which increases as the initial DM content increases,
Additional analyses of final DMA content show that at co = 450 and co = 900
the DMA decomposition was strongly inhibited during all 13 days of testing.
The ratio of DMA decomposition coefficient to the coefficient corre-
sponding to the nutrient medium decomposition is ca. 0.75 and it depends
just a little on the DMA contents within the range of co 0 - 270 mg/1 (the
dependence is statistically not significant). Therefore, it can be stated
that DMA biodegradation after a certain time of microorganisms acclimatiza-
tion proceeds with a velocity not much lower than the velocity of nutrient
medium decomposition.
The acclimatization of the inoculum to the DMA shortens the inhibition
periods ca. 0.5 day in comparison with the biodegradation process without
acclimatization. Beyond this, the process is very similar in both cases.
41
-------
c0, mg/l DMA
'=0.56mg/l-h
0 1
235
time, days
Figure 20. The decomposition of DMA in river water.
TABLE 7. KINETIC PARAMETERS OF DMA BIODEGRADATION IN RIVER WATER
Lag phase to,
days
Decomposition
velocity (v),
mg/l-h
Without acelima-
tization
After acclimati-
zation
1.3
0.8
~ 0.4
~ 0.55
Biodegradation in the river water
During the test, the DMA content was measured. When DMA content had
decreased down to zero, the next dose of DMA was added once more to the same
solution (adaptation test). The results are presented in Fig. 20. The
kinetic parameters read out from the graph are gathered in Table 7. The
data show that DMA can be biodegraded in the river water during several days
The process is advanced and accelerated by previous acclimatization of micro-
organisms to the DMA.
Treatability tests
Three series of experiments were made. In the first series the acti-
vated sludge unit was fed with wastewater containing 20 mg/l DMA. In the
second series two activated sludge units were operating in parallel (DMA
content in the inflow was 45 and 90 mg/l during the first week and changed
to 90 and 135 mg/l, respectively). In the third series one unit was fed with
wastewater with 90 mg/l DMA; the second one with 135 mg/l DMA
42
-------
ig/L DMA t
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17
Figure 21. DMA removal by activated sludge.
control chamber
c.=135 ma/1 DMA
0 12 3 5 6 7 8 9 10 11 12 13 14 15 16 17
Figure 22. The treatability coefficient KCOD of wastewater containing DMA.
The results of DMA removal determinations and the calculations of the
overall treatability coefficient KCOD are presented in Figs. 21 and 22 (third
series).
On the basis of these graphs, as well as of the data obtained in the
remaining series, the following conclusions can be made.
- The overall treatability, characterized by the value of K^QQ, did not
depend on the DMA content (within the period of K^Qp value stabilization after
acclimatization of the sludge). The differences of these KCQD values
corresponding with the control unit and two test units were statistically
not significant.
- The DMA treatability after several days of sludge acclimatization was
high and no DMA could be detected in the effluent at DMA content in the inflow
c0 up to 135 mg/l and with a retention time of 4 hours. However at the very
43
-------
short retention time (1 hour) and at c0 equal to 90 and 135 mg/1, the effluent
contained measurable quantities of DMA which enabled one to calculate K™UA
value equal to 2.0 g^-l-h"1. UMA
- The acclimatization of the activated sludge is characterized by the
values t^ of the time necessary to stabilize the COD removal rate after any
increase of DMA content in the inflow and by the value t2 required for DMA
removal rate stabilization. These values are gathered in Table 8.
TABLE 8. THE COURSE OF ACTIVATED SLUDGE ACCLIMATIZATION TO DMA
Series
I
I la
after co increase
up to ...
lib
after co increase
up to ...
Ilia
Illb
DMA in inflow
c0, mg/1
20
45
90
90
135
90
135
Time t1}
days
-
5
3
5
4
15
14
Time t2,
days
6
5
0
6
0
10
12
TABLE 9. THE CONDITIONS OF THE RESPIROMETRIC MEASUREMENTS
OF DMF BIODEGRADATION
Number of
No. samples in
the series
I
II
III
IV
V
VI
VII
4
4
6
6
6
6
6
Test
duration
days
10
9
7
9
8
9
6
Range of
, Nutrient concentration,
medium Seed mg/1
yeast extr. sludge
glucose sludge
" sludge
" sludge
" adapted
sludge
" sludge
" adapted
sludge
0 - 440
0 - 440
145
145
145
290
290
44
-------
c, mg/l NH-,
BOD mg/l 02
150
6 7
time, days
; logc
DMF;
log-^-
9N,-N
01
k=Q6
2345678
time, days
Figure 23. BOD curve, DMF decrease
and NHg increase during
the respirometric test.
Figure 24. The curve from Figure
23 after linearizing
transformation.
DIMETHYL FORMAMIDE, DMF
Respirometric measurements
Seven series of tests were carried out. In two series, BOD of the
samples with various DMF content was measured. In the other series, just one
DMF concentration was used but the measurements consisted of BOD, DMF, NH3,
N02 and N03 determinations. The conditions of the tests are given in Table 9.
The exemplary curves of BOD, DMF decrease and NH3 increase are given in
Fig. 23 (the data coming from the third series). The same data after the
linearizing transformation are presented in Fig. 24.
On the basis of the data obtained in all the series, the final carbona-
ceous BOD LC was calculated. The exemplary graph presenting Lc as a function
of initial DMF concentration co is given in Fig. 25 where the final ammonia
content is also marked.
The mean value of unit oxygen demand related to the decomposition of
1 mg DMF is equal to: LU = 1.40 ±0.02 mg 02/mg DMF. The amount of
ammonia produced in the process of the decomposition of 1 mg DMF is equal
to: Nu = 0.087 ±0.004 mg NH3-N/mg DMF.
The influence of the previous acclimatization of the seed micro-
organisms on the DMF decomposition kinetics is visible from Fig. 26 where
the linearized BOD curves are marked: curve I corresponds with the sample
seeded without acclimated inoculum and curve II corresponds with the sample
45
-------
1000
100
200 300
400 500
c0, mg/l DMF
LC and the final ammonia
content as a function of
initial DMF content cn.
20
1.0
somple with not adopted seed
/sample with the adapted seed
789
time, days
Figure 25. The final carbonaceous BOD Figure 26.
The influence of the
seed acclimatization on
the DMF biodegradation
process.
5 doys
X X \^
8 « * ^-^
X
1.0-
X
{*
0_- 0 \ 0
r = ks/kn
X
X
300 40°c, mg/D°SF 0 100 200 300 400 500
C0. mg/l DMF
Figure 27. The inhibition time of
glucose decomposition Atj_
and of DMF decomposition
At2 as a function of the DMF
Figure 28. The dependence of the
ratio r on the initial
DMF concentration co.
initial content co.
seeded with solution remaining from the former sample.
The: DMF decomposition kinetics are characterized by the curves presented
in Figs. 27 and 28. In Fig. 27 two curves are presented, both curves being
calculated on the basis of data coming from all 7 series, and representing
the following:
46
-------
- the dependence of inhibition time At-, of nutrient medium decomposition
on the initial DMF concentration co.
- the dependence of inhibition time At~ of DMF decomposition on co.
In Fig. 28 the ratio r of the coefficient ks of DMF decomposition to the
coefficient kn of nutrient medium decomposition in the blank sample is
plotted versus c .
Discussion
Assuming that the basic reaction of biochemical oxidation of DMF
follows the summarizing equation:
H.CO.N(CH3>2 + 7/2 02 . - ». 3 C02 + 2 H20 + NH3
we can calculate the stoichiometric oxygen uptake as equal to 1.53 mg 02/
mg DMF, and the unit ammonia production equal to 0.19 mg NH^-N/mg DMF.
The measured unit oxygen uptake was equal to 1.40 mg 02/mg DMF which
corresponds to 90 percent of the stoichiometric value. The unit ammonia
production was measured as equal ca. 0.09 mg NH3-N/mg DMF, i. e. a little
less than 50 percent of stoichiometric value. This last figure shows that
the ratio of nitrogen assimilated by microorganisms to the carbonaceous BOD
is equal to ca. 1:14 (mg N:mg 62) which is near to the optimal ratio in the
activated sludge treatment.
The determinations of the kinetic parameters lead to the conclusion
that DMF at concentrations up to 440 mg/1 do not affect the nutrient medium
biodegradation which is proved by the horizontal course of straight line
= 0 (Fig. 27).
The inhibition time of DMF decomposition At2 changed irregularly. Up
to DMF concentration 300 mg/1 this inhibition time was equal to ca. 3 days
and at the concentration of 440 mg/1 this time increased up to 4.5 days.
At all concentrations tested, the DMF decomposition was always complete,
i.e. no final DMF content was measurable.
The previous acclimatization of the seed microorganisms to the DMF
presence shortens the inhibition time of nutrient medium biodegradation
(t^ is average 3.5 days shorter) as well as DMF biodegradation (tj is
average 3 days shorter) .
The monomolecular reaction rate coefficient ks of DMF decomposition
amounts to 80 percent of kn value corresponding with yeast extract or
glucose biodegradation. Therefore, it can be stated that except for a
period of delay needed for microorganisms acclimatization, the biodegra-
dation of DMF processes is not much slower than the decomposition of above
mentioned standard substances.
47
-------
c,
10-
01 2 t 3 A 5 frt) 1 23
time, days
Figure 29. The decomposition of DMF in river water.
Biodegradability in the river water
Three series of measurements were made. In the first and second series,
the DMF was added to the fresh river water. The initial DMF content was
equal to ca. 30 mg/1. The third series consisted of the repeated addition
of DMF to the water in which DMF was completely decomposed during the second
series, i.e. to water with acclimatized microorganisms. The results of the
second and third series are presented in Fig. 29.
The tests without acclimatized microorganisms gave the following results
(mean of the values from first and second tests):
- lag phase t = 2 days
- the degradation rate v = 0.5 mg/l:H
The test with previously acclimatized microorganisms showed the fol-
lowing results:
48
-------
- lag phase to ~= 0
- the degradation rate v = 0.8 mg/l-h
These values prove that DMF is easily biodegraded in the river water,
but it needs a certain period of microorganisms acclimatization.
The treatability tests
One test lasting 42 days was made in which wastewater containing 70 mg/1
DMF was treated. The results of DMF determinations in the effluent and the
values of overall treatability coefficient KCQD are given graphically in
Figs. 30 and 31.
•=4h ., _tr=4h
DMF concentration in the inflow c.=73
• 2 £ 6 8 10 12 14 16 18 20 22 24 26 28 30 32 34 36
Figure 30. DMF removal by activated sludge.
"40 "42 44
time, days
0.5
KCOD- 9
''~
n/
0.3
02
0.1
0
T
K
1\
J\
\C0=73 mq/l DMF J_\
f* T
\
< /
^\\ /width ot confidence
^\/ strip
0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30 32 34 36 38 40 42 44
time, days
Figure 31. The treatability coefficient
of wastewater containing DMF.
49
-------
TABLE 10. THE CONDITIONS OF RESPIROMETRIC MEASUREMENTS OF PNP
BIODEGRADABILITY
Number of
No. samples in
the series
I
II
III
IV
V
VI
VII
5
5
5
6
6
3
3
Test Range of
duration, Nutrient concentration,
days medium Seed mg/1
10 glucose wastewater
15 " "
5 " "
5 " "
4 ii ii
3.5 yeast extr. not adapted
wastewater
3.5 " adapted
wastewater
0 - 100
0 - 100
0-80
30
30
70
70
It is visible that 35 days was necessary to acclimatize the activated sludge
to DMF. After this period DMF was removed almost completely from the
wastewater.
During the acclimatization period, the overall treatability coefficient
KCOD was much l°wer than the value corresponding with the control unit, but
after this period the KCOD value increased and any influence of DMF on the
wastewater treatment was not observable.
PARA-NITROPHENOL, PNP
Respirometric Measurements
Seven series of tests were carried out. The conditions of the
tests are given in Table 10.
The exemplary curves of BOD and the decrease of the PNP content are
given in Fig. 32. The same data, after linearizing transformation are
plotted in Fig. 33.
The unit oxygen demand Lu can be read out from Fig. 34 in which the
data coming from the second series are plotted.
The mean value, obtained on the basis of all data was equal to: Lu =
1.04 ±0.12 mg 02/mg PNP.
The amount of developed mineral nitrogen (mainly as N03) was equal to
ca. 50 percent of the nitrogen bound in PNP.
50
-------
c, mg/f PNP
BODf mg/102
130
50
Figure 32.
BOD curve and
PNP content dur-
ing the respiro-
metric test.
time, days
1.8-
1.4'
1.0-
Q6
0.2-
0
log
c A, log c
k=1.2 d'
2345
time, days
Figure 33. The curves from Figure 32
after linearizing trans-
formation.
Lu=1.03 mgO^/mg PNP
10 20 30 40
initial PNP concentration. c0,
mg/l PNP
Figure 34. The final carbonaceous
BOD Lc as a function of
initial PNP content c0.
The influence of the seed acclimatization on PNP biodegradation is seen
in Fig. 35 in which two curves are drawn, corresponding to the samples inoc-
ulated with and without acclimatized seed.
The mean values of the inhibition time of the nutrient medium decompos-
ition in the presence of PNP Atj are graphically presented in Fig. 36 as a
function of initial PNP content co. On the same graph, the mean values of
the inhibition time of PNP decomposition At2 are plotted also.
The mean values of the ratio r of the coefficient ks to the coefficient
kn (corresponding with PNP and nutrient medium biodegradation) are plotted
versus the initial PNP content co (Fig. 37).
51
-------
150
Figure 35. The influence of the seed
acclimatization on the
biodegradation process.
time, days
r=ks/kn
5.0
4.0
3JO-
2.0-
1JO
0
, Atj ; days
so
Figure 36. The inhibition time of
glucose decomposition Atj
and of PNP decomposition
At 2 as a function of the
PNP initial content c.
Figure 37. The dependence of the
ratio r on the initial
PNP concentration co.
Discussion
It was found from the experiments that around one half of the nitrogen
bound in PNP is converted to nitrates. However, the calculations of unit
oxygen demand were based not on the total BOD but on the final carbonaceous
BOD which is a more reproducible parameter.
Therefore, the biodegradation of PNP could be expressed by the equation:
52
-------
HO.C6H4.N02 + 5 02 to. 6 C02 + H20 + NH3
According to this equation, the stoichiometric value of the final carbonaceous
BOD equals 1.15 mg 02/mg PNP. The experimentally determined value (corrected
by substracting nitrogen oxygen uptake) was equal to 1.04 ±0.12 mg 02/mg PNP,
i.e. nearly 90 percent of the stoichiometric value. It should be noted that
the total BOD of PNP is somewhat higher. If one half of the nitrogen bound
in PNP is converted to N03 and the rest is assimilated, the experimental unit
oxygen demand would be ca. 1.3 mg 02/mg PNP.
The determination of kinetic parameters allows one to conclude that PNP
concentrations up to 80 mg/1 inhibits glucose biodegradation just a little.
However, at the initial PNP amount co = 100 mg/1, the inhibition of glucose
biodegradation was distinctly marked (4 days of inhibition period). This
proves that at this concentration PNP is toxic for the microorganisms.
At a low PNP content (up to 50 mg/1), the PNP biodegradation proceeds
almost simultaneously with the nutrient medium decomposition. At PNP content
within the range from 50 to 80 mg/1, the PNP biodegradation was inhibited
during 2 days, and at 100 mg/1 PNP was almost resistant.
Once the microorganisms are acclimatized to the PNP presence the PNP
decomposition proceeds fast. After the inhibition period, which is necessary
to microorganisms acclimatization, the ratio r of kinetic coefficients
corresponding with PNP and glucose decomposition ranged from 1.0 to 2.5, i.e.
PNP was decomposed faster than glucose was.
The previous adaptation of the seed shortens the inhibition time about
0.5 day.
Biodegradation in the river water
Four series of tests were made, each with use of three installations,
operating in parallel, everyone with another PNP addition. In every series,
PNP was repeatedly added at the moments in which the former PNP dose had
been completely decomposed (acclimatization testing).
Figure 38 presents exemplary results of PNP biodegradation in the river
water (third series).
On the basis of the data coming from all four series, the mean values
of inhibition time to and the decomposition velocity v were calculated and
are shown in Table 11.
The data in the table show that PNP is easily biodegraded in the river
water, especially after the acclimatization of river water biocenosis to the
PNP. It was observed that at low concentrations, e.g., 5 mg/1, the PNP bio-
degradation proceeds more slowly than at the higher concentrations.
53
-------
20
1234501 23450
Figure 38. The decomposition of PNP in river water.
1 234567
time, days
TABLE 11. KINETIC PARAMETERS OF PNP BIODEGRADATION IN RIVER WATER
After PNP
addition
Initial PNP
content c ,
mg/1 PNP
5
10
15
20
to the
river
V
mg/l-h
0.1
0.18
0.35
0.35
fresh
water
t
days
0
0.6
1.3
1.4
After repeated
PNP addition
v t
mg/l-h days
0.16 0
0.43 0
0.48 0
0.45 0
After subsequent
PNP addition
v t
mg/l'h days
0.28 0
0.53 0
0.50 0
0.48 0
Treatability Tests
The tests were carried out with use of two activated sludge units
operated in parallel. The results of the determinations of PNP removal
are presented in Fig. 39. On this graph CQ denotes PNP content in the
inflow and c denotes PNP content in the effluent.
C
The curves in the graph show that non-adapted activated sludge removes
PNP with low efficiency. After ca. 2 weeks of acclimatization, the PNP
removal efficiency becomes high and the effluent contains just traces of PNP.
54
-------
c, mg/i PNP ist
Cor5 mg/l PNP
V^ ^ --—__-.
unit
c»=:10 mg/l PNPi C0=15 mg/ PNP
•> •>/. 9fi OR
^
c, m/l PNP
2nd unit
13
10
5
*--«^^^
C.-5 """"V Co = 20 mg/l PNP c«= 30 mg/l PNP c,=40 mg/l PNP
3 2
u
yie
s
v — ./"NN. *.
\
4 6 8 10 12 14 16 18 20 22 24 26 28 30 32 34 36 38 40 42 44
time, days
Figure 39. PNP removal by the activated sludge.
The overall treatability coefficient KCOD did not depend significantly
on the PNP presence in the wastewater. KCQD value in the tests units was
almost the same as in the control unit. Additional tests were made in which
the activated sludge, acclimatized during the former tests, was fed with
wastewater containing 200 mg/l PNP. The results showed that after additional
three days of acclimatization to such PNP conent, PNP was removed at the
efficiency of 90 percent, Ks = 0.7 g'Al-h-1.
ORTHO-CHLOROPHENOL, OCP
Respirometric Measurements
Four series of tests were carried out. The first two series consisted
in the BOD measurements in the samples containing nutrient medium with
various OCP additions ranging 0 - 200 mg/l in the first series and 0 -
1200 mg/l in the second. After these tests were completed, the residual
OCP content was determined and was found in all samples as equal to ca.
50 percent of the initial value. It could be caused by biodegradation or
by the evaporation of OCP from the solution and by the absorption of OCP
vapors by the soda-lime absorbers with which the flasks of Sapromat are
equipped. The aim of the third series was to determine this process of
evaporation. The samples with the initial OCP content ranging 125 - 1240
mg/l were incubated in Sapromat at the same conditions as formerly, but
every day the flasks in Sapromat were open and small samples withdrawn for
analyses. The soda-lime mixture was analysed also, but the opening of
flasks made the BOD record not feasible. The results of this series
showed that all the OCP lost by the samples was found in the soda-lime
absorbers, and therefore, no biodegradation of OCP occurred.
The fourth series aimed to determine whether OCP at small concentrations
of 5 and 10 mg/l is biodegraded. The method was similar to the procedure
55
-------
used in the third series: the flasks were open every day and OCP content in
the samples and in the soda-lime was determined.
The conditions of the tests performed are described in Table 12 The
results of the first and second series enable one to calculate the final
carbonaceous BOD Lc of the samples with various OCP content. Exemplary BOD
curves are given in Fig. 40 (data coming from the second series). The
dependence of Lc on the initial OCP content c0 resulting from these BOD
toTS 1AnnreS!"ted ^o^g- I1' °n the last graPh> the P°ints corresponding
to c? - 600 and c0 = 1200 mg/1 represent 11 days BOD because it was not
possible to determine the final BOD in these cases.
TABLE 12. THE CONDITIONS OF THE TESTS MADE BY MEANS OF SAPROMAT
No.
I
II
III
IV
Number of Test R of
samples in duration, Nutrient concentration,
the series days medium Seed ma/1
6 14
6 11
5 19
11 11
* BOD. mg/l02
yeast extr. wastewater 0 - 200
0 - 1200
" 125 - 1240
0 - 10
Omq/l OCP, Lr=94
q/\ OCP. Lc=85
Q/t OCP. LC=7A
0
5 6 7 8 9 10 12 13
time, days
Figure 40. BOD curves of nutrient medium with various concentrations of OCP.
56
-------
BOD, mg/i 02
P-0.085mg02/mgPNP
500
1000 ,
C0, mg/l OCR
Figure 41.
The final carbonaceous
BOD Lc as a function of
initial OCP content co-
A »!, days
500
1000
c0< mq/l OCP
Figure 42,
The inhibition time of
yeast extract decomposi-
tion At^ as a function of
the initial OCP content CQ.
109 c
days
blank sample; kn=0.15 d
k=Cl055 d"1
1
9 10 11
Figure 43. The OCP content decrease
during the respirometric
test.
time, days
The value of the unit oxygen demand Lu read out from Fig. 41 amounts to
-0.085 mg 02/mg OCP. The mean Lu value, calculated on the basis of the
first and the second series, amounted to -0.08 mg 02/mg OCP which means that
OCP does not cause any oxygen demand, but it inhibits the process of oxygen
uptake for nutrient medium decomposition. Such inhibiting effect is indi-
cated not only by the decrease of final BOD of nutrient medium by the
presence of OCP, but also by the delay of nutrient medium biodegradation.
The values of this lag duration versus initial OCP content are plotted in
Fig. 42.
The tests of OCP biodegradation at low concentrations (5 and 10 mg/l)
gave the results presented in Fig. 43. As the BOD was not recorded (except
57
-------
of blank sample BOD), the curves of OCP content decrease are the only results
presented. The OCP concentration found in the soda-lime at the end of the
tests are measurable, but it was so small that it could be neglected. The
slope of the straight lines on Fig. 43 indicates the value of kinetic
coefficient ks to equal -D-OSd'1 which can be compared with the value k
determined for BOD process in the blank sample, equal to 0.15 d"1. Therefore
the ratio r = ks:kn is equal to ca. 0.3 in both cases (co = 5 and co = 10 mg/1)
The graph corresponding to 10 mg/1 (Fig. 43) shows"also a lag phase, lasting
ca. 2 days which indicates that the OCP biodegradation needs a certain
period for acclimatization.
Discussion
The obtained results show that OCP is only slightly biodegraded in
respirometric tests. At its low concentrations, up to 10 mg/1 OCP, the
decomposition of this substance was observed but the decomposition rate
was low. At an initial OCP content of 10 mg/1, two days for microorganisms
acclimatization was needed. At higher initial concentrations, OCP is
not biodegraded.
However, even at concentrations up to 350 mg/1, OCP just slightly
affects the microbiological activity which is expressed by the small
decrease of final BOD of nutrient medium and a certain delay of the process
beginning. At an initial OCP content of 600 mg/1, the inhibition of the
BOD process was distinctly marked, and at 1200 mg/1 the BOD process was
completely suspended.
Biodegradation in river water
The tests were carried out with the use of four parallel operated in-
stallations. The initial OCP content in each of the installations was,
respectively: 2, 5, 10 and 20 mg/1 OCP. When the OCP content in any of
the installations had dropped down to zero, the next OCP dose was added to
raise the OCP content to the former initial value. The results of OCP
content determinations during these tests are given in Fig. 44.
I c, mg/1 OCP
c, mg/1 OCP
10
c. mg/l OCP
time, days
0 -0 20
c, mg/l OCP0
30 40
time, days
time, days
30 40
time, days
Figure 44. The decomposition of OCP in river water.
58
-------
TABLE 13. KINETIC PARAMETERS OF OCP BIODEGRADATION IN RIVER WATER
Initial OCP
content co
mg/1 OCP
2X
5
10
20
After OCP
addition to
the fresh
river water
t V
days mg/l'h
13 0.02
15 0.07
17 0.17
22 0.10
After
OCP
t
days
0
0
0
0
repeated
addition
V
mg/l-h
0.02
0.10
0.09
0.10
After
OCP
t
days
0
0.5
0
-
subsequent
addition
V
mg/l-h
0.03
0.13
0.09
-
After
OCP
t
days
-
0
0.5
-
subsequent
addition
V
mg/l-h
-
0.09
0.13
-
x At 2 mg/1 OCP the decomposition velocity v was not constant. The data in
the table are the maximum v values.
The values of parameters read out from Fig. 44 are shown in Table 13.
This data led to the conclusion that OCP is biodegraded in the river water
but that a long period of acclimatization is needed. This period lasted
from 2 to 3 weeks, depending on the initial OCP content. Most probably
this microbiological adaptation is a sociological, not physiological one,
i.e. that adaptation consists in a change of the species domination in
the river water in the presence of OCP. The population of these species
which are able to decompose OCP are growing in number during the acclima-
tization period. Once the biocenosis is adapted to the OCP presence, the
decomposition of OCP after the repeated OCP addition begins without any
delay. However, even the adapted biocenosis degradates OCP at a low rate,
equal approximately to 0.1 mg/l'h. At a low initial content (2 mg/1 OCP),
the velocity of the decomposition is not constant and it is lower than
at the higher concentrations.
Treatability tests
The tests were carried out^with use of three parallel operated instal-
lations. The results of OCP determinations in the effluent and the
determinations of overall treatability coefficient KCOD are presented
in Fig. 45.
The data given in Fig. 45 show that the activated sludge acclima-
tization needed 6 to 12 days, depending on the OCP content in the inflow,
ranging from 5 to 10 mg/1. In the test in which the non-adapted activated
sludge was fed with wastewater containing 20 mg/1 OCP, the acclimatization
was not reached.
The overall treatability coefficient K^QQ was the same at 5 mg/1 OCP
as in the control unit. The adapted activated sludge treated the wastewater
59
-------
0.3
Q2
0.1
0
0.3
0.2
0.1
0.3
.i
0.1
o-
-< — 1
KCOD* 9 *'*"
_____
KCOD in the control unit 20%
KrOD in tne test unit\ \ width ot tne confidence strip /
/
i TsOVo removal
'v r/^-^fev^c
"""---j^ H - Jj80% rempyal unJS ^^
0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30 32 34 36 38 40
time, days
KCOD- g'-'-h"1
j 18%
KPQQ in the control unit i remowal
i T L jf^
\ *\ \l /
KQQD m the test unit\ \width of tire confidence strip V /
15% remowal, / \
25% ! /I \
1 remowal {/ \
Co I W—2S^ M H J ^\ ^P
^_ _j ),/ "T x w
M MH^ O^V ~^7fl°/ f
32468 10 12 14 16 18 20 2'2 24 26 28 30 3-2 34 36 38 40
time, days
u- -1 i k-1
KCOD • 9 '''n
KQQQ in the control unit 1 f
\ I /
• i
\ !/
width of the confidence strip \ 7\
j \
^2. , _j \
_\ J2°_)l LSEfliRL r***~ 39ct^"^W<
X " " T *
c. m^/IOCP
50
40
30
•20
•in
c, mo/I OCR
100
50
c. ma/l OCR
200
150
inn
50
n
0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30 32 34 36 3"8 40
time, days
Figure 45. Overall treatability and OCP removal by activated sludge.
60
-------
with the same value of Kcnn at OCP concentrations up to 40 mg/1 (gradually
increased).
Similarly, the feeding of the activated sludge with wastewater with
10 mg/1 OCP, gradually increased to 40 mg/1, did not affect the overall
treatability. In the test in which the non-adapted activated sludge was
fed with wastewater containing 20 mg/1 OCP, the value of K^QQ slightly
decreased. However, it remained at the level of ca. 75 percent of the
value corresponding with the control unit even when OCP content was in-
creased gradually up to 80 mg/1. OCP at concentrations of 100 mg/1 and
above poisoned the activated sludge and KPQD coefficient decreased down
to a very low value.
The OCP removal from wastewater containing up to 10 mg/1 OCP proceeds
with quite good efficiency Ks = 0.2 g-i-l-h"1. With the increase of OCP
content in the inflow, this efficiency decreases and does not depend on
the time of the activated sludge acclimatization.
TRICHLOROPHENOL, TCP
Respirometric Measurements
Four series of tests were made. In the first three series, BOD was
recorded and the final TCP content was determined. The third series con-
sisted in comparative determinations of the process in the samples seeded
with the inoculum alternatively not adapted or previously adapted to the
TCP presence. The fourth series consisted in the determination of the TCP
content during the test (BOD was not recorded). As TCP is a somewhat
volatile substance and because of its acidic properties it can be absorbed
by the soda-lime mixture, the determinations of TCP content in the absorbers
were also made similarly as in the tests with OCP. The results showed that
TCP is much less volatile than OCP. The soda-lime mixture contained measur-
able quantities of TCP after the tests were completed but these quantities
were small and could be neglected.
The conditions of the test performance are given in Table 14.
TABLE 14. CONDITIONS OF RESPIROMETRIC MEASUREMENTS OF TCP BIODEGRADABILITY
Number ofTestRange of
No. samples in duration, Nutrient concentration,
the series days medium Seed mg/1
I
II
Ilia
Illb
IV
8
6
3
3
6
18
13
7
7
7
yeast extr. wastewater
ii n n
" " not adapted
" " adapted
" " wastewater
0 - 540
0-20
10
10
12
61
-------
200-
BOD, mg/l 02
Lc, mg/l02
170
160
1 234 56 7 8 9 10 11 12 13 14 15 16 17 18
time, days
Figure 46. BOD curves of nutrient
medium with various con-
centrations of TCP.
Figure 47. The final carbonaceous
BOD Lc as a function of
initial TCP content co.
TABLE 15. BOD PROCESS IN THE SAMPLE CONTAINING VARIOUS TCP CONCENTRATIONS
Initial TCP
content CQ
mg/1
0
10
20
35
90
210
540
Final TCP
content
mg/1
0
0
0
0
80
190
500
Final carbo-
naceous BOD
mg/1 02
140
145
160
170
140
145
145
NOD
mg/1 02
35
32
30
25
20
10
10
k
of1
0.09
0.09
0.08
0.09
0.09
0.085
0.08
The exemplary BOD curves resulting from the first test are presented in
Fig. 46. The logarithmic transformation of those data follows straight lines
lying very close to one another. The kinetic parameters read out from the
logarithmic curves are presented in more readable tabular form in Table 15.
62
-------
log c
£56
time, days
BOD, mg/f 02
inoculum adapted
Lc--165 k=0,2
5 6
time, days
Figure 48. The TCP content decrease
at the initial concentra-
tion 12 mg/1 TCP.
Figure 49. The influence of adapta-
tion on BOD process in
the samples containing
10 mg/1 TCP.
The dependence of the final carbonaceous BOD of the samples containing
nutrient medium and TCP on the initial TCP content CQ is shown in Fig. 47.
The points on this graph correspond only with those samples in which TCP
was fully decomposed at the end of the tests. This was confirmed by the final
TCP content analyses.
The value of unit oxygen demand L read out from this graph is equal to
L = 0.91 mg 0 /mg TCP. The similar determination of LU in the second series
gave the result L = 0.85 mg 02 TCP. These values are not precise because
they are based onurather low BOD values of samples with small TCP addition
that causes the relative large error.
The test of TCP decomposition in Sapromat apparatus gave the result
presented in Fig. 48.
The influence of the seed acclimatization on the BOD process in the
presence of 10 mg/1 TCP is shown in Fig. 49. The curves are for the seed
alternatively adapted and not adapted.
Discussion
If it can be assumed that the TCP biodegradation follows the equation:
C1-.C,H..OH + 5.5 0_
3 o 2. 2.
6 CO + 3 HC1
the stoichiometric value of the unit oxygen uptake would amount to 0.89 mg
0?/mg TCP. The mean value resulting from the tests is equal to 0.88 mg 0 /mg
TCP which is near to the stoichiometric value. It should be stated that
this value is adequate to calculate the final BOD of the samples containing
initially 35 mg/1 TCP or less. At the higher initial concentration TCP is
not biodegraded and does not cause any oxygen demand.
63
-------
The determination of the kinetic parameters shows that TCP does not
inhibit the process of nutrient medium biodegradation. Even at high TCP
content of 540 mg/1 no delay of nutrient medium decomposition was observed.
Similarly the value of kn coefficient and the final BOD of nutrient medium
were not decreased by the presence of TCP.
The TCP decomposition proceeded only at the initial TCP concentration
up to 35 mg/1. Any delay of the TCP decomposition could not be read out
from the BOD curves. However, the curve of TCP content decrease (Fig. 48)
indicates that TCP decomposition at its initial contents 12 mg/1 needed ca. 5
days for microorganisms acclimatization. After that the TCP decomposition
proceeded at a high rate (k =0.9 d"-*-) .
The previous acclimatization of the seed to the TCP presence accelerates
the BOD process. This is indicated by some increase of the coefficient k
value (Fig. 49).
Biodegradation Tests in River Water
Two series of the tests were made. In the first series two river models
were operating in parallel with the initial TCP content 8 and 15 mg/1,
respectively. The second series was carried out with use of three river
models with initial TCP content 5, 8 and 10 mg/1. In any case, when TCP
was fully decomposed, the TCP dose was repeated (the tests of microorganisms
acclimatization to the TCP presence).
The results of both series are presented in Fig. 50. These graphs show,
that the TCP biodegradation in the river water needs a certain period of
biocenosis acclimatization (ca. 6 days) during which TCP is decomposed at
a low rate. Once the biocenosis is acclimatized, the next dose of TCP is
biodegraded without any delay (or after short delay) and at a rate of about
0.35 mg/l-h. At low initial TCP content (5 mg/1), this rate is somewhat
lower.
Treatability Tests
One test lasting 38 days was carried out. The activated sludge was fed
with wastewater with gradually raising the TCP content from 5 mg/1 at the
test beginning to 25 mg/1 at the end of the test.
The results of the determinations of the TCP in the effluent c and of
the overall treatability coefficient K are presented graphicallyein Fig. 51,
On the basis of these results, the following conclusions can be made. The
activated sludge did not need any acclimatization period for its normal
activity until the TCP in the inflow surpassed 18 mg/1. The overall treat-
ability coefficient K was stable during the tests and equal the value of
KCOD corresPondin8 to ™e control unit. However, when the TCP content in the
inflow equaled 25 mg/1 it poisoned the activated sludge, and K dropped to
zero. UJU
The TCP removal from wastewater needs some period of adaptation t7
depending on the TCP content in the inflow: at 5 mg/1 TCP t, = 0; at
64
-------
c. mg/1 TCP
log phase 6.5 days
0.20
v=0.20
0 42 3 I 5 6 1 8: 9
c, mg/l TCP
11 12 13 ft". 1? 1"6
time, days
v=0.33
lag phase 6.5 days
7 5 9 10 11 12 13 U 15 1
0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16
time, days
c. mg/l TCP
11 12 13 U 15 ,
time.
-------
0.3
0.2-
0.1
KCOD. 9 •'•"'
KQQQ in the control unit
L\__—
width of the confidence strip
in the test unit
_t
A A
c, mg/l TCP
24 T
20
10
0 2 4 6 8 10 12 14 16 18 20 22 24 26 28 30 32 34 36 38 40
time, days
Figure 51. Overall treatability and TCP removal by activated sludge.
10 mg/l TCP t, = 2; at 18 mg/l TCP t = 5 days. The adapted activated sludge
removed TCP from wastewater with hign efficiency; the effluent contained just
traces of TCP. An increase of TCP contents in the inflow up to 35 mg/l
caused a decrease of TCP removal efficiency down to near zero.
DICHLORODIETHYL ETHER, DCDEE
Respirometric Measurements
Two series of the tests were carried out. Yeast extract (200 mg/l) was
used as nutrient medium and the samples were seeded with the wastewater
previously aerated. The DCDEE contents ranged from 0 to 500 mg/l in the
first series and from 0 to 15 mg/l in the second one.
Exemplary BOD curve from the first series is presented in the Fig. 52.
All BOD curves obtained in this series were posed very close one to another.
Therefore, to make the graph more readable Fig. 52 shows only the BOD curve
of the sample containing 100 mg/l DCDEE, compared with the BOD curve of
blank sample and the carbonaceous BOD of the blank sample, corrected on
the basis of N0? and NO, content determinations.
The values of final carbonaceous BOD were determined for every BOD curve
and they are plotted versus the initial DCDEE content (Fig. 53). The slope
of the straight line in Fig. 53 does not differ significantly from zero.
It means that the unit oxygen uptake caused by DCDEE is equal to zero or
too small to be measurable.
66
-------
uo
01234567
10 11 12 13 14 15 16
time, days
200
300
400
500 .
C0, mg/l DCDEE
Figure 52. BOD curve of the sample Figure 53.
containing DCDEE.
Final carbonaceous BOD
L as a function of
initial DCDEE content c
The values of k coefficient of nutrient medium biodegradation ranged
irregularly within tRe narrow limits 0.15 - 0.17 d-1 in the first series and
0.2 to 0.25 d"1 in the second series. The lag phase of nutrient medium bio-
degradation was not observed in any test.
The final DCDEE content and the N02 and NO increase during the tests
are given in Table 16. In the second series no change of DCDEE content
during the test was observed either.
TABLE 16. THE INITIAL AND FINAL DCDEE CONTENT AND N02 AND NO
INCREASE DURING THE TESTS IN SAPROMAT (THE FIRST SERIES)
Initial
CQ, mg/l
0
14
41
107
205
512
DCDEE
Final
c£J mg/l
0
15
40
109
203
501
N02 + NO
mg/l N
9.5
1.2
0.5
0.3
0.2
0.6
67
-------
c, mg/l DCDEE
01 2 3 4 5 6 7
c, mg/l DCDEE
9 10 11 12 13 U 15 16 17 18
time, days
0 1 23 A 567
9 10 11 12 13 U 15 16 17 18
time, days
Figure 54. DCDEE changes in river water.
Discussion
The unit oxygen uptake caused by DCDEE was measured as equal to zero
(L = 0). No lag phase of nutrient medium decomposition was observed. The
coefficient k of nutrient medium decomposition was also not affected by the
DCDEE. The determination of the final DCDEE content showed that DCDEE is not
degraded during the tests. Summarizing, it can be concluded on the basis of
these results that DCDEE is an inert substance which is not biodegraded but
does not affect in anyway the biodegradation of the other organics (yeast
extract components).
The only effect of DCDEE on the biochemical processes was the inhibition
of nitrification. At a DCDEE concentration of 14 mg/l and higher, the
nitrification was almost completely inhibited; whereas, in the blank sample
almost all the ammonia was converted to NO,, and N0_.
Biodegradation in River Water
As
Two series of tests were made using two units operating in- parallel.
DCDEE is a volatile substance, the river models were sealed to avoid any
DCDEE loses due to evaporation. In both series the DCDEE content was analyzed
during the tests. The results are presented in Fig. 54.
In the second series the NH and NO changes were also observed during
the test. The aim was to determine the influen.ce of DCDEE on the nitrification
of ammonia in river water. The results are presented in Fig. 55.
68
-------
3 mg/l DCDEE
6 mo/1 DCDEE
0 2 4 6 8 10 12 14 16 18
time, days
c, mg/l N-NH3
blank sample
3mg/i DCDEE
6 m^/l DCDEE
=***
0 2 • 4 6 8 10 12 14 16 18
time, days
Figure 55. Nitrites and ammonia changes in river water.
Discussion
The DCDEE content curves show that this substance is resistant in the
river water. A small decrease of DCDEE was observed at the beginning of
tests. Beyond this, no further DCDEE decrease was observed.
The observation of the nitrification process leads to the conclusion
that DCDEE at concentrations up to 6 mg/l does not inhibit the nitrification
but causes a delay of this process (lasting 1.5 to 2 days).
Treatability Tests
One series of test was carried out by use of two parallel operating
activated sludge units fed with the wastewater containing 5 and 10 mg/l
DCDEE, respectively. As DCDEE is a volatile substance, the supplementary
determination of its Henry coefficient a_ was made. The air was bubbled
through the DCDEE solution in distilled~~water. DCDEE content and the weight
of the solution were measured during the test. The results were plotted in
the logarithmic coordinates system (Fig. 56). The slope of the straight
line was read out, and according to the equation given in Section 6, the
value of coefficient a_ was calculated equal to 0.37 x 10~3. Assuming that
the evaporation of DCDEE is the only cause of its content decrease in the
activated sludge chamber, the DCDEE content in the effluent would be equal
to: :
c =
e
1+a • —
where v is the rate of air blowing, equal to 300 1/h and V is the rate of
wastewater inflow, equal to 0.17 1/h. As the value of a amounts to 0.37 x
10~3, c should be equal to 0.6 • c .
e o
69
-------
2.00-
1.99-
1.98
1.97
1.96
1.95
log c
slope 18.74
2.397 2398 2399 2.400
log ot the weight of solution
Figure 56. The volatility of DCDEE from aqueous solution.
6
5i
i
3
2
1
c, my'l DCDEE
UJ Ol
, iu£ —
\ o i ^
\ / ^ f\ rt
\ ,°l°
\ y
V-A
v^ . i: ^
v*"1 • — i|r
ce i
i
^Bh .!. ^Sh
1
0 i 2
9
8
7
6
5
t.
3
2
1
n
3 i 5 6 7 8 9 10 11 12 13 U 15 16 17 18 19 2'0 21
time, days
c, mo/1 DCDEE
2)
LU °
LLl 01
Q c
0 >
DO
n
V^
V
/\ A
^^ ^^ / ^^r ^V
./ HI, / ^ ^^
v<- v \
time, days
Figure 57. DCDEE concentration in the effluent from activated sludge unit.
70
-------
The results of the determinations of the DCDEE content in the effluent
c are presented in Fig. 57 on which the theoretical DCDEE content decrease
by the blowing out is also marked.
The overall treatability coefficients K were calculated on the basis
of COD determinations. The obtained values snow just irregular scattering.
The mean values and their confidence limits are as below:
control unit KCQD = 0.20 ±0.06
at DCDEE content in the inflow of 5 mg/1, KCQD = 0.20 + 0.07
at DCDEE content in the inflow of 10 mg/1, KCOD = 0.16 ± 0.09
The scattering of the results causes rather wide confidence limits.
Because of it, any decrease of K value in the presence of DCDEE cannot
i (jUJJ
be stated.
It can be concluded that DCDEE content in wastewater does not affect
significantly the overall treatability of wastewater (i.e. the COD decrease).
Thus, DCDEE is an inert substance in the treatment process. Its removal
is caused just by the blowing off with the air in the aeration chamber.
FLUORESCENT WHITENING AGENTS, FWAs
Respirometric Measurements
Two kinds of the tests were carried out. The tests of the first kind
(BOD tests) consisted in the recording of the BOD of the samples, each
containing nutrient medium and one of the five various FWAs (100 mg/1).
For the inoculum, the activated sludge supernatant was used. After com-
pleting the tests, lasting 12 days, the final FWAs content was determined.
Next the tests were repeated, similarly as before, except that the solu-
tions remaining from the previous tests were used as inoculum (acclimatization
tests).
The tests of the second kind consisted in the measurement of the
activated sludge endogenous respiration in the presence of various FWAs.
The procedure was as follows: the settled activated sludge was put into
the flasks of Sapromat and the FWAs solutions in the dilution water were added.
The activated sludge respiration was recorded during 10 days and compared
with the respiration of the blank sample (the same activated sludge without
any FWA addition). Every FWA was tested in three identical samples to reach
the higher reliability of the results. After the tests were finished, the
FWAs content were determined.
The results of the BOD test for FWA-1 is presented in Fig. 58. The BOD
curves for the other FWAs are very similar to that shown in Fig. 58, e.g.,
the blank sample and the sample with 100 mg/1 FWA have identical oxygen
uptake. Figs. 59 and 60 present exemplary curves of the activated sludge
endogenous respiration in the presence of FWA-1 and FWA-3.
71
-------
inoculum: supernatant of the
activated sludge
inoculum: the liquid
ot test I
123456789 10 11 12 1 2 3 I 5 £>
time, days
Figure 58. BOD curves of the samples containing nutrient medium and 100 mg/1
of FWA-1.
1200
1000
800
600
400
200
Figure 59. The curve of activated sludge endogenous respiration in the
presence of FWA-1.
72
-------
1500-
1000
500
BOD, mg/f 02
8 100
time, days
6 8 10 0
time, days
4 6
time, days
Figure 60. The curves of activated sludge endogenous respiration in the
presence of FWA-3.
TABLE 17. THE FINAL FWAs CONCENTRATIONS IN THE SAMPLES
CONTAINING INITIALLY ONE OF THE FIVE FWAs
(100 mg/1). (THE TESTS LASTED 10 DAYS.)
Final FWA FWA-1 FWA-2 FWA-3 FWA-4 FWA-5
concentration
mg/1 5
21
91
69
87
The results of final FWAs content determination are summarized in Table
17 (BOD tests) and in Table 18 (activated sludge respiration tests). The
last table contains also the data of final BOD (mean values and the limits
at 90 percent confidence).
73
-------
TABLE 18. BOD AND FINAL FWA CONTENT IN THE SAMPLES CONTAINING
CA. 2 g/1 OF ACTIVATED SLUDGE AND ONE OF THE FWAs.
Initial
FWA concentration
mg/1
FWA-1
FWA- 2
FWA- 3
FWA-4
FWA-5
0
100
0
100
180
300
0
20
0
100
0
200
0
20
100
0
20
100
Final BODio
concentration
mg/1 mg/1 0?
0
5
0
10
15
30
0
4
0
70
0
180
0
15
70
0
10
100
1125
1170
1500
1520
1610
1780
1450
1400
1130
1080
900
680
920
925
1060
1530
1550
1440
± 130
+ 90
± 140
± 110
± 100
± 180
± 120
± 100
± 80
± 30
± 150
± 110
± 190
± 180
± 120
± 80
Discussion
The BOD curves of the samples containing nutrient medium and FWAs
additions show that the FWAs do not affect the BOD process. It means
that the FWAs are not subject to very much decomposition, which would
cause the oxygen uptake, and that they do not inhibit the process of.
nutrient medium biodegradation. Acclimatization of microorganisms to the
FWAs presence does not change the BOD processes.
The similar conclusion can be made on the basis of the activated
sludge respiration data. The oxygen uptake by activated sludge endogenous
respiration in the FWAs presence was almost the same as in the blank sample
74
-------
(the differences were statistically not significant). One exception was the
FWA-3 at 200 mg/1 which inhibited the respiration of activated sludge.
The data of final FWAs contents show that FWA-1 and FWA-2 are not resistant
at the conditions of the tests (i.e. in the samples aerated and kept in
darkness). The final FWA-4 content was not much lower than its initial
content. The FWA-3 and FWA-5 were almost resistant in the solutions with
their high initial contents. But in the case of FWA-3 at the low initial
content equal to 20 mg/1, it decreased to a low value. It may be caused by
the FWA-3 adsorption on the activated sludge floes because this FWA is easily
adsorbed as it was confirmed in the experiments described further.
It should be underlined that the above described partial FWAs decomposi-
tion was based on fluorimetric analyses. The FWAs concentrations were deter-
mined based on the fluorescent properties of FWAs molecules. Such results do
not mean that the FWAs are really degraded. The FWAs molecules could lose
their fluorescent properties as a result of a simple transfiguration (e.g.,
cis-trans) or as a result of their breaking into large fragments (e.g., by
the oxidation and breaking of the double bond of styrene structure). As no
measurable oxygen uptake was observed in the samples in which FWAs content
decreased, it can be concluded that no large amount of degradation occurred.
The Decomposition in River Water
Two kinds of tests were made. The tests of the first kind consisted in
the long term observation of the FWAs content changes in river water kept
in darkness. In parallel, FWAs in distilled water, kept at the same condi-
tions, were also observed.
In the tests of the second kind, the river water with FWAs was exposed
to the light of fluorescent tubes at an intensity of 1000 lux. Also in these
tests, FWAs solutions in the distilled water, exposed to the light, were used
as the blank samples. In the last case, the total organic carbon (TOC) was
also measured. Additionally, the river water models and blank samples were
kept in the darkness to compare the FWAs decomposition in the presence and
in the absence of the light.
As the rate of the FWAs photolytic decomposition depends on pH, phosphate
buffer solution was always added to the blank samples (FWAs solutions in the
distilled water) to keep pH at the same level as pH of river water used in
the tests (8 - 8.5) .
The results of 30 days testing of the FWAs fate in the river water in the
absence of the light showed that FWAs at these conditions are almost resistant.
The FWAs contents decreased just a little, 10 to 20 percent of the initial
value (10 mg/1). Even this small decrease probably was not related to the
biodegradability because in the blank samples, which were almost microbiolog-
ically sterile, some FWAs decrease was observed.
The decomposition of FWAs exposed to light proceeded at a high rate so
that FWAs content had to be measured every half hour. In the samples
75
-------
c,,mg/ FWA-1
= 11 mg/l-h
10
0 1
234567
time, days
c, mg/l FWA-2
v=13 mg/l-h
distilled water
0 1
5 6 7
time, days
Figure 61. Photolytic FWA-1 decom-
position in river and
distilled water.
Figure 62. Photolytic FWA-2 decom-
position in river and
distilled water.
50
30-
20-
c, mg/l FWA-3
v = 5 mg/l-h
v = 5 mg/l-h
river water
distilled water
v=1.5 mg/l-h
0 1 23456.7
time, days
Figure 63.
Photolytic FWA-3
decomposition in
river and dis-
tilled water.
76
-------
c, mg/ FWA-4
distilled water
river water
v=3.6 mg/l-h
50Jc.mg/FWA-5
5 6 7
time, days
river water
v=7 mg/l-h
distilled water
0
time, days
Figure 64. Photolytic FWA-4 decom-
position in river and
distilled water.
Figure 65. Photolytic FWA-5 decom-
position in river and
distilled water.
TABLE 19. VELOCITY v OF THE FWAs DECOMPOSITION IN RIVER AND DISTILLED
WATER EXPOSED TO LIGHT
Velocity of FWAs decomposition,
FWA
In river water at
initial FWA content
FWA-1
FWA- 2
FWA- 3
FWA-4
FWA-5
c = 10
o
5
9
1.5
2
5
c = 50
o
6
13
5
4
7
mg/l'h.
In distilled water at
initial FWA content
c = 10
o
7
.14
5
8
11
c = 50
o
11
13
5
9
12
77
-------
simultaneously kept in darkness no FWAs change was observed. The curves of
FWAs decomposition are presented in Figs. 61, 62, 63, 64 and 65.
The curves show the decrease of FWA down to the residual value c related
most probably to the fluorescence of the FWA decomposition product.
During the initial period of FWAs photolysis, the decomposition velocity
is usually constant. The values of decomposition velocity v, being equal to
the slope of the initial section of lines, are read out from the graphs and
are shown in Table 19.
The additional TOG determinations in the samples of FWAs in distilled
water show that the FWAs decomposition is not related to any TOC decrease.
Final TOC values were almost the same as the initial values, whereas the FWAs
content dropped down to the residual values c^.
Discussion
The long term tests of the FWAs fate in the river water in the absence
of light have shown that all FWAs tested are resistant. No FWAs biodegrada-
tion was observed during 30 days. The observed small FWAs decrease, equal
to 10 to 20 percent of the initial FWAs concentrations, is probably due to
physical processes.
The tests of FWAs decomposition in the presence of the light have shown
that all FWAs tested are subject to the photolytic decomposition. According
to the tests with FWAs solutions in distilled water, the most easily decom-
posed FWAs are FWA-2 and FWA-5, whereas FWA-3 is the most difficult to decom-
pose with a rate less than one half of the FWA-2 or FWA-5 decomposition rate.
The decomposition rate is initially almost not dependent on the FWAs
concentration and next it is decreasing asymptotically to zero. The FWAs
solutions after a long time of exposition to the light (1 days or more)
retain a residual fluorescence, corresponding with 5 to 10 percent of their
initial value.
The decomposition of FWAs in river water proceeded similarly as in the
distilled water. At the initial FWAs concentration of 50 mg/1, the
decomposition rate in river water was equal to or less than in the distilled
water. At the concentration of 10 mg/1, the FWAs were decomposed in river
water at significantly lower rates. The fact that the decomposition of FWAs
is slower in the river water than in the distilled water can be explained
as a result of the light extinction in the river water because of colored
impurities. This absorption of the light decreases the amount of light needed
for photochemical reaction of FWAs decomposition.
The fact that the solutions keep the residual fluorescence and that the
TOC is not changed during the tests, leads to the conclusion that chemical
changes of FWAs molecules were very little. It could be a cis-trans trans-
figuration or division to large fragments.
78
-------
TABLE 20. THE ADSORPTION OF FWAs ON ACTIVATED SLUDGE
c c m a b
o e
"^*r^ ~ ~~
10 4.86
FWA-2 5 3.52
teSt X 10 7.96
3.1 5.6
test 2 5 3.45 _
/ • 3
10 8.26
FWA-3 5 1.5 , ,
«Stl 10 5.1 '6
1.0 1.0
test 2 5 0.9
/ • O
10 3.75
FWA-4 ^ ^7?
hWA 4 b • '/Z 4.8 1.3 9.2
10 8.03
FWA-5 5 5
7.3
10 10
Treatability Tests
As the adsorption of FWAs on the floes of activated sludge can interfere
with the FWAs biodegradation determinations, supplementary tests of FWAs
adsorption were carried out. The procedure was as follows: into the sample
of the settled activated sludge of a known density (dry mass) the FWAs stock
solution was added and after 1 hour of FWA contact with the activated sludge
the FWA content in the supernatant was determined. Next, the coefficients
of the Langmuire's isotherm of adsorption were calculated, following the
equation:
c - c c ,
o e _ e , where:
m a«c +b
e
CQ is the initial FWA content in the supernatant, mg/1
cg is the FWA content in the supernatant at the state of equilibrium
with the adsorbed FWA, mg/1
m is the dry mass of the activated sludge, g/1
a_ and b_ are Langumire's coefficients.
The results are summarized in Table 20.
79
-------
The tests of the FWAs treatability were carried out with the use of three
parallel operated activated sludge units as follows:
- test unit, fed with wastewater containing the FWA addition
- control unit I, fed with wastewater without any FWA addition
- control unit II, fed with distilled water with FWA addition.
The measured parameters were:
- FWAs content in the inflow c and in the effluent c
o e
- COD in the inflow and in the effluent
- activated sludge dry mass m in the test unit and control unit I.
On the basis of the obtained data, the following treatability coeffici-
ents were calculated. The general FWAs removal coefficient K was determined
by use of the equation
c - c
K=
c mt
e r
where tris the retention time. It can be assumed that the general coefficient
K is the sum of the coefficient K, of FWA removal by degradation, the coef-
ficient Kph of FWA removal by photolysis and the coefficient Kad °f FWA removal
by adsorption on the sludge. The values Kp^ were determined by use of the
equation
v - ACH • 1
ph c mt
where Ac is the FWA decrease in the test unit II; c , m and t^. are corres-
ponding with the test unit. The values of K were calculated on the basis
of formerly determined Langumire's coefficients:
ad 9 a-c +b
e
where 9 is the sludge age equal to 120 hours, a_ and b are Langumire's
coefficients (Table 20) , c@ is FWA content in the effluent from the
test unit. The values of the coefficient of FWAs removal by the biodegrada
tion are equal toK, =K-K,-K,.
80
-------
The overall treatability coefficient K
equation
was calculated by use of the
K
COD
COD - COD
o e_
COD,,
mt
In cases when the tests were made at high FWAs concentrations in the inflow,
the Kmn values were corrected by substracting the value of COD corresponding
with FWA content both from the measured COD of the inflow and of the efflu-
ent. This corrected value is denoted as K' .
L/UJJ
The results of subsequent FWAs tests are presented in Tables 21, 22, 23,
24 and 25. In the case of FWA-1, the photolysis significantly decreased FWA
content in the effluent; the general coefficient of FWA removal was approxi-
mately twice the value of K . In the case of FWA-2, the photolysis caused
a small decrease of FWA content. Practically, it was the only cause of FWA
removal .
TABLE 21. THE TREATABILITY OF -FWA-1
Test
duration
days
20
27
FWA-1 in FWA-1 in
the inflow the effluent
CQ mg/1 cg mg/1
9.9 1.6
40.0 8.0
TABLE 22. THE
FWA-1
Overall treatability
biodegradation coefficients
coefficient
g~i LIT1
0.08
0.10
TREATABILITY OF
V
b test unit
K'
COD
g-1 i-h-1
0.17
0.17
FWA-2
control
unit K
COD
-1 . . -1
g . 1-h
0.17
0.17
Test FWA-2 in FWA-2 in
duration the inflow the effluent
FWA-2
biodegradation
coefficient K,
Overall treatability
coefficients
test unit control
KCOD Unit KCOD
days
10
18
24
34
40
CQ mg/1
6
10
45
100
200
cg mg/1
4.9
10.1
44.3
~ 100
~ 200
g-1. Lh-1
~ 0
~ 0
- o
~ 0
- o
-1 - , -1
g • 1-h
0.18
0.19
0.18
0.14
0.13
-1 . , -1
g • 1-h
0.17
0.18
0.19
0.17
0.18
81
-------
TABLE 23. THE TREATABILITY OF FWA-3
Test
duration
days
5
11
18
24
32
FWA-3 in
the inflow
CQ mg/1
5
10
16
24
80
FWA-3 in
the effluent
ce mg/1
3.0
6.2
8.1
18.0
64.5
TABLE 24. THE
Test
duration
days
11
16
FWA-4 in
the inflow
c mg/1
4
44
FWA-4 in
the effluent
cg mg/1
4.3
44.4
TABLE 25. THE
Test
duration
days
11
16
FWA-5 in
the inflow
C0 mg/1
4
40
FWA-5 in
the effluent
ce mg/1
4.2
39.5
FWA-3
biodegradation
coefficient K,
b
-1 . , -1
g ' 1-h
0.029
0.021
0.042
0.012
0.008
Overall treatability
coefficients
test unit control
KCOD Unlt KCOD
-1 , , -1 -1 . , -1
g • l.h g • 1-h
0.18 0.17
0.15 0.19
0.14 0.19
0.06 0.18
0.05 0.18
TREATABILITY OF FWA-4
FWA-4
biodegradation
coefficient K,
b
g~* 1-h"1
~ 0
~ 0
Overall treatability
coefficients
test unit control
KCOD Unit KCOD
g"1 1-h"1 g-1 1-h"1
0.16 0.16
0.15 0.15
TREATABILITY OF FWA-5
FWA-5
biodegradation
coefficient K,
-1 . , -1
g • 1-h
~ 0
~ 0
Overall treatability
coefficients
test unit control
KCOD Unit KCOD
-1 . , -1 -1. , -1
g • 1-h g • 1-h
0.16 0.16
0.16 0.15
82
-------
Discussion
The obtained results show that FWA-2, FWA-4 and FWA-5 are not biodegraded
in the activated sludge process. The highest removal rate was observed in the
case of FWA-1 but even in this case the treatability coefficient K is rather
low being equal to ca. 0.1 g-*1 1-h'1. FWA-3 is less biodegradable than FWA-1;
its Kb value is equal to ca. 0.03 g-^l-h'1 and it decreases down to 0.01
when FWA-3 content in the inflow increases above 20 mg/1.
The overall treatability of wastewater is not affected significantly by
any of FWAs in the inflow, except FWA-3 which decreases the KCOD value when
its content in the inflow surpasses ca. 20 mg/1. Certain decrease of K^
was observed also in the case of FWA-2 at its high content in the inflow
(100 mg/1 and above).
83
-------
SECTION 8
THE RESULTS OF BIOASSAYS
METHYLETHYL KETONE, MEK
Behavior. The test organisms (fish, Lebistes reticulatus) reacted
with darkening when they were exposed to MEK at the concentration of 2000 mg/1.
At 2600 mg/1 MEK, the disturbing of the balance was observed. At 3400 mg/1
MEK, the fish were laying flat on the vessel bottom. At MEK concentrations
near LC5Q, i.e. 5000 mg/1, the fish were paralysed.
The mortality of fish at various MEK concentrations is shown in Fig. 66.
LC50~24 h Was found as equal to 5700 mg/1 MEK.
The results of a supplementary test with algae Chlorella sp. are given
in Table 26.
100
8 . 10
c, g/l MEK
Figure 66. The mortality of fish at various MEK concentrations,
84
-------
TABLE 26. CHLOROPHYLL "a" CONTENT IN THE CULTURE OF CHLORELLA AFTER
7 DAYS OF EXPOSURE TO THE PRESENCE OF MEK
Test MEK content, Chlorophyll "a" content, mg/1
no* mg>/1 blank culture test culture
1 806 435 487
2 806 640 660
The differences in the chlorophyll content in the test samples and
blank samples are within the range of analytical error. Therefore, any
impact of MEK on the assimilation activity of Chlorella cannot be stated.
DIMETHYL AMINE, DMA
Behavior. After 20 rain of exposure to the DMA at 64 mg/1, the fish
reacted with fluttering movements. At 73 mg/1 DMA the fish turn pale and
their balance was disturbed (partial paralysis).
The mortalities of fish at various concentrations of DMA are presented
in Table 27. From the results of the test (Table 27) the approximate value
of LC,-n- 24 h was calculated as equal to 55 mg/1.
O \J
TABLE 27. THE MORTALITY OF Lebistes retlculatus
N. Time
Concen- N.
tration
mg/1
Blank
41.6
54.6
72.8
97.5
130.0
10
\
mm
0
20
20
20
30
20
30
min
0
20
20
20
40
60
1
h
0
20
20
40
60
100
3
h
0
20
20
60
100
100
24
h
0
20
30
100
100
100
85
-------
TABLE 28. CHLOROPHYLL "a" CONTENT IN THE CULTURE OF CHLORELLA AFTER
7 DAYS OF EXPOSURE TO DMA
Test DMA content, Chlorophyll "a" content, mg/1
no.
1
2
3
4
5
6
mg/i
20
20
40
40
60
60
blank culture
34
40
34
40
57
59
test culture
44
44
48
41
54
56
The test with algae Chlorella sp. indicates that DMA does not affect the
assimilation activity of these algae. It is visible from the data presented
in Table 28.
DIMETHYL FORMAMIDE, DMF
Behavior. The fish reacted with their balance being disturbed within
the first hour of exposure to 650 mg/1 DMF. After 24 hours, fish turned
dark. At 750 mg/1 DMF, the lateral or ventral bend of the fish body was
observed. The 870 mg/1 DMF caused fluttering movements of the fish.
The mortality data are presented in Fig. 67. The value of LC - 24 h
was found to be 1300 mg/1. 50
Supplementary test with algae Chlorella did not indicate any affect of
DMF on the assimilation activity of Chlorella. The results of this test
are given in Table 29.
PARA-NITROPHENOL, PNP
Behavior. At the lower PNP concentration the reaction of fish was not
significant. At 18 mg/1 PNP the animals moved slowly at the botiom and
after 30 min they turned somewhat pale and the first symptoms of partial
paralysis were observed. At the concentration of 32 mg/1, after 20 minutes
the fish turned on their side (they looked almost not alive except for the
weak action of heart and sporadic movements of the fin). After 2 or 3 hours
all organisms were dead and dropped to the bottom. The mortality of fish
at various PNP concentrations is marked on Fig. 68. The LC _- 96 h was
found as equal to 19 mg/1.
86
-------
100
60
o
S
E
20
1000
1500
c, m
DM
Figure 67. The mortality of fish at various DMF concentrations.
TABLE 29. THE CHLOROPHYLL "a" CONTENT IN THE CULTURE OF CHLORELLA AFTER
7 DAYS OF EXPOSURE TO DMF
Test
no.
1
2
3
4
DMF content,
mg/1
945
945
1890
1890
Chlorophyll "a"
blank culture
57
40
34
57
content, mg/1
test culture
46
46
43
50
ORTHO-CHLOROPHENOL, OCP
Behavior. Within the first 10 minutes in all solutions tested, the
movements of fish were rapid and violent. At lower concentrations the fish
were acclimated to OCP presence after one hour and their behavior became the
same as in the control flask. At the concentration of 15 mg/1 and higher
most of the fish were paralysed after one hour. The others got together at
the bottom. At the highest concentration and after one or two hours, the
87
-------
100
80-
^60
o
o
E
40-
20
12 U 16 18 20 22 24 26 28 30 32
c, mg/l PNP
Figure 68. The mortality of fish at various PNP concentrations.
100
80
£
£60
~B
o
E^O
20
10
12
U
16 18
c, mgj/l OCR
Figure 69. The mortality of fish at various OCP concentrations.
fish were almost completely paralysed. Several of them were alive in this
state during one day.
Mortality of the fish at various OCP concentrations is plotted in Fig.
69. The calculated LC - 96 h was 12 mg/1.
88
-------
100
80-
£
^60
"5
-*-
t_
£40
20
7 , 8
c, mg/l TCP
Figure 70. The mortality of fish at various TCP concentrations.
TRICHLOROPHENOL, TCP
Behavior. Four or five minutes after the fish were placed in the test
flasks the solutions in the flasks became turbid. After the next 5 or 10
minutes the animals began to move rapidly; later on their movements became
slower and the fish kept themselves near the surface. After 30 to 40 minutes
from the beginning of the experiment, a fine precipitate, causing turbidity,
settled so that the solution became clear. This precipitate, agglomerated
with fish slime, was visible on the mouths, gill covers and fins of the fish.
At the concentrations above 4.9 mg/l most of the fish were paralysed.
Mortality of fish in the presence of TCP is shown in Fig. 70.
interpolated LC „- 96 h value was 4.5 mg/l.
The
DICHLORODIETHYL ETHER, DCDEE
Behavior. After 20 to 30 minutes the animals turned slightly dark and
their colors became more grayish. At lower concentrations, the fish got
together at the surface, moving slowly. At higher concentrations the
symptoms of paralysis were visible after 1.5 hours: the fish kept abnormal
positions. The dead animals floated at the surface.
Mortality is presented in Fig. 71.
equals 190 mg/l.
The
calculated LC - 96 h value
89
-------
100
80
tf-
>;,
o
e
20-
100
200 300
c, mg/l DCDEE
Figure 71. The mortality of fish at various DCDEE concentrations.
FLUORESCENT WHITENING AGENTS, FWAs
FWA-1
Behavior. During the first period of the test the fish reacted to the
FWA-1 presence with rapid movements. After several minutes the fish were
posed on the bottom of the vessel rapidly moving with their chest fins. In
the test media with the highest FWA-1 content (21 - 37 mg/l), the fish were
floating, turned with their backs down, and almost did not react when
touched. At FWA-1 content of 15.5 mg/l the fish were partially paralysed.
The mortality of fish is presented in Fig. 72. LC - 96 h was equal
to 16 mg/l. The determination of LC „- 24 h to Daphnia magna gave the
result of 23 mg/l. bu
FWA-2
Behavior. At the lower FWA-2 content (below 1 g/1) the fish were
animated during the first minutes of the test, and then they moved more
slowly. After 24 hours of the test, the behavior and the look of the
fish were normal. At the highest tested concentration (above 3 g/1),
paralysis and death of fish were observed. Extravasations were visible
on various parts of the fish body.
The mortality of fish is presented in Fig. 73.
Lebistes reticulatus was equal to 1760 mg/l.
for Daphnia magna was found to be 3900 mg/l.
LC - 96 h for
The LC5Q- 24 h of FWA-2
90
-------
100
80
40
20-
10 20 30 , 40
c, mg/l FWA-1
Figure 72. The mortality o£ fish at various FWA-1 concentrations.
100
0 1 2 3,4
c, g/l FW*-2
Figure 73. The mortality of fish at various FWA-2 concentrations.
FWA-5
Behavior. During the initial three days of the test, in the solutions
with ca. 100 mg/1 FWA-3, the fish were sluggish but reacted when touched.
During the fourth day, the same symptom was visible in the fish exposed
91
-------
80 . 100 120
c, mg/l FWA-3
Figure 74. The mortality of fish at various FWA-3 concentrations.
to 89 mg/l FWA-3. At the FWA-3 concentration of 120 mg/l, little extra-
vasations on the fish body were visible.
The mortality of fish is presented in Fig. 74. The LC5Q- 96 h of
Lebistes reticulatus was equal to 110 mg/l. The LC50- 24 h of Daphnia
magna surpassed the solubility of FWA-3 in water. At the saturated FWA-3
solution, the mortality of Daphnia magna amounted to 20 percent..
FWA-4
Behavior. At FWA-4 content of 4 to 10 g/1, the fish were paralysed
and the extravasations were visible near the gills of the fish. The backs
of the dead fish were bent near the caudal fins.
The mortality of the fish is presented in Fig. 75. The LC50- 96 h
of Lebistes reticulatus was 3000 mg/l. The LC50- 24 h to Daphnia magna
equaled 9400 mg/l.
FWA-5
Behavior. At the lower FWA-5 concentration (4.5 g/1) the fish were
sluggish but reacted when touched. After several hours their behavior
was quite normal. At the higher FWA-5 concentration (6-7 g/1), the
fish were paralysed and extravasations were visible. The skin of the dead
fish was chapped.
92
-------
100
Figure 75. The mortality of fish at various FWA-4 concentrations
5 6 , 7
c. mg/l FWA-5
Figure 76. The mortality of fish at various FWA-5 concentrations.
The mortality of fish is presented in Fig. 76. The LC „- 96 h of
Lebistes reticulatus was 6400 mg/l. The LC - 24 h of Daphnia magna was
5600 mg/l. bu
93
-------
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99
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TECHNICAL REPORT DATA
(Please read Instructions on the reverse before completing)
REPORT NO.
EPA-600/2-79-163
3. RECIPIENT'S ACCESSION NO.
4. TITLE AND SUBTITLE
INVESTIGATIONS OF BIODEGRADABILITY AND TOXICITY OF
ORGANIC COMPOUNDS
5. REPORT DATE
December 1979
(Issuing Date)
6. PERFORMING ORGANIZATION CODE
. AUTHOR(S)
8. PERFORMING ORGANIZATION REPORT NO.
Jan R. Dojlido
9. PERFORMING ORGANIZATION NAME AND ADDRESS
Institute of Meteorology and Water Management
Department of Water Chemistry and Biology
01-673 Warsaw ul. Podlesna 61
Poland
10. PROGRAM ELEMENT NO.
PL-480
11. CONTRACT/GRANT NO.
PR-05-532-15
12. SPONSORING AGENCY NAME AND ADDRESS
Municipal Environmental Research Laboratory--Cin.,OH
Office of Research and Development
U.S. Environmental Protection Agency
Cincinnati, Ohio 45268
13. TYPE OF REPORT AND PERIOD COVERED
Final 1975 - 1979
14. SPONSORING AGENCY CODE
EPA/600/14
15. SUPPLEMENTARY NOTES
Project Officer: Robert L. Bunch (513)684-7655
16. ABSTRACT
The development of elaborate industrial societies has led to proliferation of a
vast number of complex chemicals for industrial, agricultural and domestic use. Some
portion of these compounds eventually find their way into municipal and industrial
wastewater. Unless specifically removed by waste treatment processes, they ultimate-
ly appear in receiving waters and water supplies, thus no longer is it sufficient to
remove biochemical oxygen demand to protect the oxygen resource of the receiving
water but individual organic compounds become a concern.
Knowledge of the toxicity and biodegradability of organic compounds will aid in
designing wastewater treatment processes and be useful in elaborating the criteria foi
safe concentrations of organics in wastewaters discharged to surface waters7
This report describes the testing of twelve compounds both for biodegradability
and toxicity. The compounds tested were: methylethyl ketone, dimethyl amine, di-
methyl foramide, p-nitrophenol, o-chlorophenol, trichlorophenol, 2,2'dichlorodiethyl
ether and five flourescent whitening agents used as components of household detergents
The biodegradation tests performed were respirometric measurements, river model and
activated sludge model. Additionally, for some compounds supplementary tests were
made for evaluation of their volatility, photolysis and adsorption on activated sludge
The toxicity was measured with use of fish Lebistes reticulatus § crustacean Daphnia
mnanx
17.
KEY WORDS AND DOCUMENT ANALYSIS
DESCRIPTORS
b.lDENTIFIERS/OPEN ENDED TERMS
COS AT I Field/Group
Water pollution
Biodeterioration
Organic wastes
Toxicology
Sewage treatment
Industrial waste treatment
Biodegradability
Activated sludge
17C
13B
18. DISTRIBUTION STATEMENT
Release to Public
19. SECURITY CLASS (ThisReport)
Unclassified
21. NO. OF PAGES
116
20. SECURITY CLASS (Thispage)
Unclassified
22. PRICE
EPA Form 2220-1 (9-73)
100
I U.S. GOVERNMENT PRINTING OFFICE: 1980 -657-146/5557
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