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Biological test systems for monitoring the operation of wastewater treatment plants
Chemosphere, Vol. 30, No. 2, pp. 327-338, 1995
Pergamon
0045-6535(94)00400-5
Copyright O 1995 Elsevier Science Ltd Printed in Great Britain. All rights reserved 0045-6535/95 $9.50+0.00
BIOLOGICAL TEST SYSTEMS FOR MONITORING THE OPERATION OF WASTEWATER TREATMENT PLANTS
Uwe J. Strotmann 1., Andreas Keinath 1,2 and Stefan H. Hfittenhain2 1 BASF AG, Ecology Laboratory, 67056 Ludwigshafen, Germany 2 Fachhochschule Darmstadt, Chemical Technology, 64289 Darmstadt, Germany
(Received in Germany 24 June 1994; accepted 12 August 1994)
Abstract Four different biological test systems were used for monitoring the biological activity of a laboratory-scale wastewater treatment plant which was operated with municipal wastewater. The heterotrophic respiration activity test (HRA), dehydrogenase activity test (DHA) and nitrification activity test were used for monitoring the biological activity of the activated sludge, and the luminescent bacteria test was used for screening the effluent of the treatment plant. In shock loading experiments with 2.3-dichlorophenol and 3.5-dichlorophenol it was shown that both the nitrification and heterotrophic respiration activities and the TOC degradation in the treatment plant were significantly reduced, whereas no clear effect on dehydrogenase activity was found. Screening of the effluent revealed a clear increase in luminescent bacteria inhibition during the shock loading experiments. Keywords Activated sludge, toxicity monitoring, respiration activity, dehydrogenase activity, nitrification activity, luminescent bacteria test, chlorinated phenols
(*): Corresponding author
327
328 1. Introduction
The toxicity for the activated sludge in a wastewater treatment plant is an important criterion for assessing the possibility of biological treatment ofa wastewater. Therefore, rapid and sensitive methods for the assessment of toxicity are of importance for the operators of wastewater treatment plants. Currently, measurement of the respiration activity of activated sludge is the most widely accepted parameter for monitoring activated sludge activity and is used in on-line monitoring systems and in toxicity screening tests ( Anderson et al., 1988; Dutka et al., 1983; Pagga and Gtinthner, 1981, Broecker and Zahn, 1977; Brown et al., 1981; King, 1984; King and Painter, 1986; Green et al., 1975; Thibault and Tracy, 1978; Summers and Slon, 1981). Besides this test system, the measurement of dehydrogenase activity and nitrification activity is becoming increasingly important (Strotmann et al., 1993; Kroiss et al., 1992). Furthermore, measurement of B-galactosidase activity, uptake of glucose and the estimation of the intracellular ATP level are biological test systems used for monitoring the biological activity of activated sludge. For the estimation of a possible inhibitory effect of the effluent of a treatment plant, the luminescent bacteria test has been frequently used in the last few years (Airbuckle and Alleman; 1992; Fentem and Balls; 1993). The main objectives of this investigation were (I) to compare different test systems for monitoring the activity of activated sludge in wastewater treatment plants, (II) to simulate shock loading situations to evaluate the reaction by the test systems used and (III) to obtain information about the sensitivity and reliability of the monitoring systems. 2. Materials and Methods
Chemicals The chemicals used were of analytical grade and obtained from Merck (Darmstadt, Germany), Riedel-de-H~ien (Seelze, Germany) and Fluka (Neu-Ulm, Germany). Wastewater treatment plants
A Husmann apparatus which was operated with municipal wastewater was used for the monitoring experiments. The characteristics of the Husmann apparatus have been described before (EEC, 1983). The system consisted of an aeration vessel with a volume of 4.6 1 and a separator with a volume of 2.5 1from which the sludge was recycled intermittently. The composition of the municipal wastewater is described in Table 1.
329
Table 1. Characteristics of the municipal wastewater
Parameter
Content
Unit
Total organic carbon (TOC)
80-300
mg 1-1
Chemical oxygen demand (COD)
180-1000
mg 1-1 mg 1-1
Biochemical oxygen demand (5 days) (BOD5) 150-700 Phosphate-P 0.2-40 Ammonium-N Nitrate-N
17-70 0.1-3
mg 1-1 mg 1-1 mg 1-1
The concentration of mixed liquid suspended solids (MLSS) in the treatment plant was between 2.5 and 3.0 g/l and the hydraulic residence time varied between 13 and 16 hours. The volumetric loading rate of the wastewater treatment plant was 0.9 to 1.4 kg COD m-3 d "1. The excess sludge production was 2.7 to 3.2 g MLSS -1 d-land the return sludge ratio I00 %. Shock loading experiments were performed by directly adding an appropriate amount of 2.3dichlorophenol or 3.5-dichlorophenoi to the storage vessel for the influent.
Respiration activity The total respiration activity (TRA) was determined by measuring the oxygen consumption by an activated sludge sample from the aeration vessel with an oxygen electrode (WTW EO 90). Prior to measurement the activated sludge sample was saturated with oxygen by sparging with air. The respiration activity is defined as mg ofO 2 per liter consumed per hour and the specific respiration activity is the respiration activity per g of MLSS. The heterotrophic respiration activity (HRA) is the total respiration activity minus the respiration activity caused by nitrification processes.
Nitrification activity The nitrification activity was determined as the oxygen consumption caused by nitrification processes. For this purpose an activated sludge sample from the aeration basin was incubated together with allylthiourea (ATU; 25 mg 1-1 final concentration) for 30 minutes to inhibit nitrification processes. The respiration activity measured after this treatment was due to heterotrophic respiration. The difference between total respiration activity (TRA) measured in an untreated sample and heterotrophic respiration (HRA) measured in an ATU treated sample
330 was the respiration caused by nitrification processes. The specific nitrification activity was the oxygen consumption by nitrification per g of MLSS. Dehydrogenase activity
The dehydrogenase activity of activated sludge was determined by measuring the reduction of resazurin to resorufin. The detailed test procedure has been described earlier (Strotmann et al., 1993). One unit of deh~,drogenase activity is defined as the activity catalyzing the reduction of 1 ~tmoi of resazurin per minute. The specific dehydrogenase activity is the dehydrogenase activity measured per g of MLSS. Luminescent bacteria inhibition
The inhibition ofPhotobacteriumphosphoreum luminescence was measured according to DIN 38412 part 341 (1993). For the tests the equipment from Dr. Lange (Diasseldorf, Germany) with conserved bacteria was used. The incubation period of the luminescent bacteria with the different water samples was 30 minutes. The inhibition is expressed in terms ofG L values indicating the highest dilution factor not causing a 20 % inhibition of the luminescent bacteria. 3. Results Normal operation of the wastewater treatment plant
The normal operation parameters of the treatment plant are summarized in Table 2. The extent of TOC elimination and ammonium elimination were in the region of 75-90 % and 70-100 %, respectively. During the whole operation period these parameters proved to be quite constant with only minor changes. The specific total respiration activity was between 17 and 32 mg 0 2 g MLSS -1 h-1, the specific heterotrophic respiration activity was between 10 and 20 nag 0 2 g MLSS -1 h"1 and the specific respiration activity due to nitrification processes varied between 7 and 12 mg 0 2 g MLSS -1 h -1 . Therefore the oxygen consumption by nitrification of ammonium reached as much as 40 % of the total oxygen consumption. The dehydrogenase activity of the activated sludge was stable in the range from 1.3 to 2.5 U g MLSS -1. The influent GLValues widely varied between 6 and 64, whereas the effluent GLvalues were stable at GL=2 indicating that the activated sludge was operating well and the effluent toxicity was low.
331 Table 2. Operation parameters of the wutewater treaunent plant
Parameter
Content
Unit
TOC elimination Ammonium elimination Dehydrogenase activity Heterotrophic respiration Nitrification activity In_fluent G L Effluent G L
75-90 70-100 1.3-2.5 10-20 7-12 6-64 2
% % U g MLSS" 1 mg 0 2 g MLSS" 1 h" 1 mg 0 2 g MLSS "1 h"1
Shock loading with 3.S-dichlorophenol A shock loading of 20 mg/i 3.5-dichlomphenol over a period ofg days caused a decrease in ammonium degradation by 85 to 95 %, resulting in a sharp increase in ammonium in the effluent of the treatment plant. The TOC elimination was reduced by 25 to 45 %. On the other hand, the dehydrogenase activity was not significantly affected and remained stable in the range between 1.5 and 2.1 U g MLSS "1. The heterotmphic respiration was reduced by 30 to 45 % to a rate of 8 to 10 mg 0 2 g MLSS "1 h"1 (Fig. 1). The shock loading was also detected by an increase in the G L values of the effluent (Fig. 2). Ten days after the end of shock loading the treatment plant regained its normal nitrification activity, indicating the reversible character of the inhibition process during the shock loading. Shock loading with 2.3-dicblorophenol An initial shock loading with 20 mg/i 2.3-dichlorophenol over a period of 20 days immediately caused an inhibition of the nitrification activity by 90 to 100 %. The TOC degradation was reduced by 20 to 40 %. A stepwise increase in the 2.3-dichiorophenol concentration (40 mg/l, 60 rag/I, 100 mg/I) resulted in a clear increase in heterotmphic respiration inhibition, whereas dehydrogenase activity was not affected (Fig. 3). The effluent toxicity as measured by the luminescent bacteria inhibition test showed an increased inhibition response with increasing influent concentrations of 2.3-dichlorophenol (Fig. 4). The time for reestablishment of the sensitive nitrification process after terminating the shock loading was about 12 days which also indicates the reversible type of inhibition caused by 2.3-
332
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Time (days) Fig. 1. Monitoring of the biological activity of activated sludge before and after shock loading with 3.5-dichlorophenol. (A) Ammonium degradation (e), dehydrogenase activity (O) and (B) TOC degradation (11), heterotrophic respiration activity (17) were followed. The 3.5-dichlorophenol concentration is indicated by a dotted line. Details are given in the materials and methods section
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T i m e (days) Fig. 2. Effluent monitoring with the luminescent bacteria test before and after shock loading with 3.5-dichlorophenol. The GLvalues in the effluent (o) and the 3.5-dichlorophenol concentration (dotted line) are indicated.
4. Discussion
The data obtained show that during the normal operation of the wastewater treatment plant the various parameters measured were rather stable. Both TOC removal and ammonium oxidation by nitrification reached 90 to 100 %. The inhibitory effect of the effluent on luminescent bacteria was low with an GL value of 2. During shock loading with 2.3-dichlorophenol and 3.5-dichlorophenol a strong inhibition of the nitrification process was observed. This inhibitory effect was also confirmed by the reduction of TOC degradation. An effect on the dehydrogenase activity could not be observed, whereas the heterotrophic respiration was clearly inhibited by 2.3-dichlorophenol and 3.5-dichlorophenol. The inhibitory effect on luminescent bacteria was significant indicating that the luminescent bacteria test was an appropriate test system for screening effluent toxicity. The results obtained are comparable to literature data summarized in Table 3. The highly toxic effect of dichlorophenols on nitrifying bacteria is in good agreement with data already reported. Also the inhibitory effect on luminescent bacteria is in good agreement with other data from this test system. Concerning heterotrophic respiration activity and dehydrogenase activity
334
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Time (days) Fig. 3. Monitoring of the biological activity of activated sludge before and after shock loading with 2.3-dichlorophenol. (A) Ammoniumdegradation (o), dehydrogenase activity (O) and (B) TOC degradation (Ill), heterotrophic respiration activity ([]) were followed. The 2.3-dichlorophenol concentration is indicated by a dotted line. Details are given in the materials and methods section.
335 there are discrepancies in the literature data. For dichlorophenols it was recently shown that the toxicity in the dehydrogenase test strictly depends on the pH of the test system, the toxicity increasing with decreasing pH (Strotmann et al., 1993).Therefore, it must be remembered that the conditions under which the toxicity of a chemical compound for activated sludge is determined can influence the results found (Klecka et al., 1985). Furthermore, there may also be differences in data obtained in laboratory tests and experiments to simulate real shock loading situations. The response of wastewater treatment plants to inhibitory compounds may also vary due to different organic loading, biomass concentration, hydraulic residence time and a possible biological adaption. Therefore correlations may not be valid for different systems. Nevertheless, it can be stated that the different test procedures provide reliable data and can be regarded as useful screening tools for identifying problems.
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T i m e (days) Fig. 4. Effluent monitoring with the luminescent bacteria test before and after shock loading with 2.3-dichlorophenol. The GLValues in the effluent (e) and the 2.3-dichlorophenol concentration (dotted line) are indicated.
336 Table 3. Bacterial toxicity data for 2.3-diehlorophenol and 3.5-dichlorophenol
Compound
Test system
EC50 (mg/i)
References
2.3-DCP
Nitrification inhibition Luminescent bacteria inhibition
0.09 4
Strotmann and Egls~ier, 1994 Strotmann and Egls~ier, 1994
Respiration inhibition
120
Strotmarm et al., 1994
Growth inhibition
40-60
Strotmann et al., 1994
Nitrification inhibition
0.5
Luminescent bacteria inhibition Luminescent bacteria inhibition Luminescent bacteria inhibition Respiration inhibition Respiration inhibition Respiration inhibition Respiration inhibition Respiration inhibition Growth inhibition
5 3.9 3 10 25-60 11-12 11 20-30 5- 10
Strotmann and Egls~er, 1994 Strotmann and Egls~er, 1994 Einabarawy et al., 1988 Dutka et al., 1983 Strotmann et al., 1994 Broecker and Zahn, 1977
Pagga, 1981 Strotmann et al., 1994
DHA DHA (pH 7.0)
80 >100
Dutka et al., 1983 Strotmann et al., 1993
DHA (pH 6.5)
50
Strotmann et al., 1993
DHA (pH 6.0)
32
Strotmann et al., 1993
3.5-DCP
Klecka et al., 1985 EInabarawy et al., 1988
Acknowledgements The authors thank C.Schmitt for skilful technical assistance and U. Pagga for helpful discussions. We also thank D. Charles for correcting the manuscript. 5. References Airbuckle, W.B. and Alleman, J.E (1992). Effluent toxicity testing using nitrifiers and Microtox®. Water Environ.Res. 64: 263-267.
337 Anderson, K., Koopman, B. and Britton, G. (1988). Evaluation of INT-dehydrogenase assay for heavy metal inhibition of activated sludge. Water Res. 22: 349-353. Broeeker, B. and Zahn, R. (1977). The performance of activated sludge plants compared with the results of various bacterial toxicity tests - a study with 3.5-dichlorophenol. Water Res. 11: 165-172. Brown, D., Hitz, H.R. and Schaefer, L. (1981). The assessment of the possible inhibitory effect of dyestuffs on aerobic wastewater bacteria - experience with a screening test. Chemosphere 10: 245-261. DIN 38412 Part 341 (1993). German standards for the determination of water, wastewater and sludge; bioassays; determination of the inhibitory effect ofwastewater on the light emission of
Photobacterium phosphoreum. Dutka, B.J., Nyhol, N. and Petersen, J. (1983). Comparison of several microbiological toxicity screening tests. Water Res. 17: 1363-1368. EEC Directive 79831 Annex V (1983). Part C: Methods for determination of ecotoxicity; 5.2. Degradation- biotic degradation; C.5. Activated sludge simulation test. Elnabarawy, M.T., Robideau, R.R. and Beach, S.A. (1988). Comparison of three rapid toxicity test procedures: Microtox®, Polytox® and activated sludge respiration inhibition. Toxicity Assessment 3: 361-370. Fentem, J. and Balls, M. (1993 ).Replacement of fish in ecotoxicology testing: use of bacteria, other lower organisms and fish cells in vitro. In: Richardson, M (ed.) Ecotoxicology monitoting, pp. 71-81, VCH Publishers, Weinheim. Green, BB., Willets, D.G. and Bennett, M. (1975). Applications of toxicity testing to sewage treatment processes. Water Pollut. Control 74:40-54. King, E.F. (1984). A comparative study of methods for assessing the toxicity to bacteria of single chemicals and mixtures. In: Liu, D. and Dutka, B.J. (eds.) Toxicity screening procedures using bacterial systems, pp. 175-194, Marcel Dekker, New York.
338 King, E.F. and Painter, H.A. (1986). Inhibition of respiration of activatexl sludge" variability and reproducibility of results. Toxicity Assessment 1: 27-39. Klecka, G.M., Landi, L.P. and Bodner, KM. (1985). Evaluation of the OECD activated sludge respiration inhibition test. Chemosphere 14:1239-1251. Kroiss, H., Schweighofer, P., Frey, W. and Matsche, N. (1992). Nitrification inhibition. A source identification method for combined municipal and/or industrial wastewater treatment plants. Wat.Sci.Tech. 26:1135-1146. Pagga, U. (1981). The short-term respiration test - a simple method to check the respiratory activity of activated sludge. Vom Wasser 57: 263-276. Pagga, U. and Gtinthner, W. (1981). The BASF Toximeter- a helpful instrument to control and monitor biological wastewater treatment plants. Wat.Sci.Tech. 13: 233-276. Strotmann U. J. and Eglsaer, H. (1994). The toxicity of substituted phenols in the nitrification inhibition test and the luminescent bacteria test. Ecotoxic.Environm.Saf., in press. Strotmann, U.J., Egls~ier, H. and Pagga, U. (1994). Development and evaluation of a growth inhibition test with sewage bacteria for assessing bacterial toxicity of chemical compounds. Chemosphere 28: 755-766. Strotmann, U. J., Butz, B. and Bias, W.R. (1993) The dehydrogenase assay with resazurin: practical performance as a monitoring system and pH-dependent toxicity of phenolic compounds. Ecotoxic.Environm.Saf. 79-89. Summers, S.M. and Slon, R.A. (1981). Real-time monitoring of biomass respiration in an activated sludge system. In: Proceedings of the 36th Industrial Waste Conference, pp. 701-710. Ann Arbor Science, MI. Thibault, G.T. and Tracy, K.D. (1978). Controlling and monitoring activated sludge units. Chem. Eng. 85: 155-160.
Pergamon
0045-6535(94)00400-5
Copyright O 1995 Elsevier Science Ltd Printed in Great Britain. All rights reserved 0045-6535/95 $9.50+0.00
BIOLOGICAL TEST SYSTEMS FOR MONITORING THE OPERATION OF WASTEWATER TREATMENT PLANTS
Uwe J. Strotmann 1., Andreas Keinath 1,2 and Stefan H. Hfittenhain2 1 BASF AG, Ecology Laboratory, 67056 Ludwigshafen, Germany 2 Fachhochschule Darmstadt, Chemical Technology, 64289 Darmstadt, Germany
(Received in Germany 24 June 1994; accepted 12 August 1994)
Abstract Four different biological test systems were used for monitoring the biological activity of a laboratory-scale wastewater treatment plant which was operated with municipal wastewater. The heterotrophic respiration activity test (HRA), dehydrogenase activity test (DHA) and nitrification activity test were used for monitoring the biological activity of the activated sludge, and the luminescent bacteria test was used for screening the effluent of the treatment plant. In shock loading experiments with 2.3-dichlorophenol and 3.5-dichlorophenol it was shown that both the nitrification and heterotrophic respiration activities and the TOC degradation in the treatment plant were significantly reduced, whereas no clear effect on dehydrogenase activity was found. Screening of the effluent revealed a clear increase in luminescent bacteria inhibition during the shock loading experiments. Keywords Activated sludge, toxicity monitoring, respiration activity, dehydrogenase activity, nitrification activity, luminescent bacteria test, chlorinated phenols
(*): Corresponding author
327
328 1. Introduction
The toxicity for the activated sludge in a wastewater treatment plant is an important criterion for assessing the possibility of biological treatment ofa wastewater. Therefore, rapid and sensitive methods for the assessment of toxicity are of importance for the operators of wastewater treatment plants. Currently, measurement of the respiration activity of activated sludge is the most widely accepted parameter for monitoring activated sludge activity and is used in on-line monitoring systems and in toxicity screening tests ( Anderson et al., 1988; Dutka et al., 1983; Pagga and Gtinthner, 1981, Broecker and Zahn, 1977; Brown et al., 1981; King, 1984; King and Painter, 1986; Green et al., 1975; Thibault and Tracy, 1978; Summers and Slon, 1981). Besides this test system, the measurement of dehydrogenase activity and nitrification activity is becoming increasingly important (Strotmann et al., 1993; Kroiss et al., 1992). Furthermore, measurement of B-galactosidase activity, uptake of glucose and the estimation of the intracellular ATP level are biological test systems used for monitoring the biological activity of activated sludge. For the estimation of a possible inhibitory effect of the effluent of a treatment plant, the luminescent bacteria test has been frequently used in the last few years (Airbuckle and Alleman; 1992; Fentem and Balls; 1993). The main objectives of this investigation were (I) to compare different test systems for monitoring the activity of activated sludge in wastewater treatment plants, (II) to simulate shock loading situations to evaluate the reaction by the test systems used and (III) to obtain information about the sensitivity and reliability of the monitoring systems. 2. Materials and Methods
Chemicals The chemicals used were of analytical grade and obtained from Merck (Darmstadt, Germany), Riedel-de-H~ien (Seelze, Germany) and Fluka (Neu-Ulm, Germany). Wastewater treatment plants
A Husmann apparatus which was operated with municipal wastewater was used for the monitoring experiments. The characteristics of the Husmann apparatus have been described before (EEC, 1983). The system consisted of an aeration vessel with a volume of 4.6 1 and a separator with a volume of 2.5 1from which the sludge was recycled intermittently. The composition of the municipal wastewater is described in Table 1.
329
Table 1. Characteristics of the municipal wastewater
Parameter
Content
Unit
Total organic carbon (TOC)
80-300
mg 1-1
Chemical oxygen demand (COD)
180-1000
mg 1-1 mg 1-1
Biochemical oxygen demand (5 days) (BOD5) 150-700 Phosphate-P 0.2-40 Ammonium-N Nitrate-N
17-70 0.1-3
mg 1-1 mg 1-1 mg 1-1
The concentration of mixed liquid suspended solids (MLSS) in the treatment plant was between 2.5 and 3.0 g/l and the hydraulic residence time varied between 13 and 16 hours. The volumetric loading rate of the wastewater treatment plant was 0.9 to 1.4 kg COD m-3 d "1. The excess sludge production was 2.7 to 3.2 g MLSS -1 d-land the return sludge ratio I00 %. Shock loading experiments were performed by directly adding an appropriate amount of 2.3dichlorophenol or 3.5-dichlorophenoi to the storage vessel for the influent.
Respiration activity The total respiration activity (TRA) was determined by measuring the oxygen consumption by an activated sludge sample from the aeration vessel with an oxygen electrode (WTW EO 90). Prior to measurement the activated sludge sample was saturated with oxygen by sparging with air. The respiration activity is defined as mg ofO 2 per liter consumed per hour and the specific respiration activity is the respiration activity per g of MLSS. The heterotrophic respiration activity (HRA) is the total respiration activity minus the respiration activity caused by nitrification processes.
Nitrification activity The nitrification activity was determined as the oxygen consumption caused by nitrification processes. For this purpose an activated sludge sample from the aeration basin was incubated together with allylthiourea (ATU; 25 mg 1-1 final concentration) for 30 minutes to inhibit nitrification processes. The respiration activity measured after this treatment was due to heterotrophic respiration. The difference between total respiration activity (TRA) measured in an untreated sample and heterotrophic respiration (HRA) measured in an ATU treated sample
330 was the respiration caused by nitrification processes. The specific nitrification activity was the oxygen consumption by nitrification per g of MLSS. Dehydrogenase activity
The dehydrogenase activity of activated sludge was determined by measuring the reduction of resazurin to resorufin. The detailed test procedure has been described earlier (Strotmann et al., 1993). One unit of deh~,drogenase activity is defined as the activity catalyzing the reduction of 1 ~tmoi of resazurin per minute. The specific dehydrogenase activity is the dehydrogenase activity measured per g of MLSS. Luminescent bacteria inhibition
The inhibition ofPhotobacteriumphosphoreum luminescence was measured according to DIN 38412 part 341 (1993). For the tests the equipment from Dr. Lange (Diasseldorf, Germany) with conserved bacteria was used. The incubation period of the luminescent bacteria with the different water samples was 30 minutes. The inhibition is expressed in terms ofG L values indicating the highest dilution factor not causing a 20 % inhibition of the luminescent bacteria. 3. Results Normal operation of the wastewater treatment plant
The normal operation parameters of the treatment plant are summarized in Table 2. The extent of TOC elimination and ammonium elimination were in the region of 75-90 % and 70-100 %, respectively. During the whole operation period these parameters proved to be quite constant with only minor changes. The specific total respiration activity was between 17 and 32 mg 0 2 g MLSS -1 h-1, the specific heterotrophic respiration activity was between 10 and 20 nag 0 2 g MLSS -1 h"1 and the specific respiration activity due to nitrification processes varied between 7 and 12 mg 0 2 g MLSS -1 h -1 . Therefore the oxygen consumption by nitrification of ammonium reached as much as 40 % of the total oxygen consumption. The dehydrogenase activity of the activated sludge was stable in the range from 1.3 to 2.5 U g MLSS -1. The influent GLValues widely varied between 6 and 64, whereas the effluent GLvalues were stable at GL=2 indicating that the activated sludge was operating well and the effluent toxicity was low.
331 Table 2. Operation parameters of the wutewater treaunent plant
Parameter
Content
Unit
TOC elimination Ammonium elimination Dehydrogenase activity Heterotrophic respiration Nitrification activity In_fluent G L Effluent G L
75-90 70-100 1.3-2.5 10-20 7-12 6-64 2
% % U g MLSS" 1 mg 0 2 g MLSS" 1 h" 1 mg 0 2 g MLSS "1 h"1
Shock loading with 3.S-dichlorophenol A shock loading of 20 mg/i 3.5-dichlomphenol over a period ofg days caused a decrease in ammonium degradation by 85 to 95 %, resulting in a sharp increase in ammonium in the effluent of the treatment plant. The TOC elimination was reduced by 25 to 45 %. On the other hand, the dehydrogenase activity was not significantly affected and remained stable in the range between 1.5 and 2.1 U g MLSS "1. The heterotmphic respiration was reduced by 30 to 45 % to a rate of 8 to 10 mg 0 2 g MLSS "1 h"1 (Fig. 1). The shock loading was also detected by an increase in the G L values of the effluent (Fig. 2). Ten days after the end of shock loading the treatment plant regained its normal nitrification activity, indicating the reversible character of the inhibition process during the shock loading. Shock loading with 2.3-dicblorophenol An initial shock loading with 20 mg/i 2.3-dichlorophenol over a period of 20 days immediately caused an inhibition of the nitrification activity by 90 to 100 %. The TOC degradation was reduced by 20 to 40 %. A stepwise increase in the 2.3-dichiorophenol concentration (40 mg/l, 60 rag/I, 100 mg/I) resulted in a clear increase in heterotmphic respiration inhibition, whereas dehydrogenase activity was not affected (Fig. 3). The effluent toxicity as measured by the luminescent bacteria inhibition test showed an increased inhibition response with increasing influent concentrations of 2.3-dichlorophenol (Fig. 4). The time for reestablishment of the sensitive nitrification process after terminating the shock loading was about 12 days which also indicates the reversible type of inhibition caused by 2.3-
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0
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20
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30
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50
60
0
70
80
Time (days) Fig. 1. Monitoring of the biological activity of activated sludge before and after shock loading with 3.5-dichlorophenol. (A) Ammonium degradation (e), dehydrogenase activity (O) and (B) TOC degradation (11), heterotrophic respiration activity (17) were followed. The 3.5-dichlorophenol concentration is indicated by a dotted line. Details are given in the materials and methods section
333 50
~ , . , l , , , , i , , ~ , l , , , , l ~ , , ~ l , , , , i , , , , i , , ~ ,
20
40
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~
211
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T i m e (days) Fig. 2. Effluent monitoring with the luminescent bacteria test before and after shock loading with 3.5-dichlorophenol. The GLvalues in the effluent (o) and the 3.5-dichlorophenol concentration (dotted line) are indicated.
4. Discussion
The data obtained show that during the normal operation of the wastewater treatment plant the various parameters measured were rather stable. Both TOC removal and ammonium oxidation by nitrification reached 90 to 100 %. The inhibitory effect of the effluent on luminescent bacteria was low with an GL value of 2. During shock loading with 2.3-dichlorophenol and 3.5-dichlorophenol a strong inhibition of the nitrification process was observed. This inhibitory effect was also confirmed by the reduction of TOC degradation. An effect on the dehydrogenase activity could not be observed, whereas the heterotrophic respiration was clearly inhibited by 2.3-dichlorophenol and 3.5-dichlorophenol. The inhibitory effect on luminescent bacteria was significant indicating that the luminescent bacteria test was an appropriate test system for screening effluent toxicity. The results obtained are comparable to literature data summarized in Table 3. The highly toxic effect of dichlorophenols on nitrifying bacteria is in good agreement with data already reported. Also the inhibitory effect on luminescent bacteria is in good agreement with other data from this test system. Concerning heterotrophic respiration activity and dehydrogenase activity
334
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,,, , , ~ I I ~ , , ; I, I , , ....
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Time (days) 100
,,,l
....
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Time (days) Fig. 3. Monitoring of the biological activity of activated sludge before and after shock loading with 2.3-dichlorophenol. (A) Ammoniumdegradation (o), dehydrogenase activity (O) and (B) TOC degradation (Ill), heterotrophic respiration activity ([]) were followed. The 2.3-dichlorophenol concentration is indicated by a dotted line. Details are given in the materials and methods section.
335 there are discrepancies in the literature data. For dichlorophenols it was recently shown that the toxicity in the dehydrogenase test strictly depends on the pH of the test system, the toxicity increasing with decreasing pH (Strotmann et al., 1993).Therefore, it must be remembered that the conditions under which the toxicity of a chemical compound for activated sludge is determined can influence the results found (Klecka et al., 1985). Furthermore, there may also be differences in data obtained in laboratory tests and experiments to simulate real shock loading situations. The response of wastewater treatment plants to inhibitory compounds may also vary due to different organic loading, biomass concentration, hydraulic residence time and a possible biological adaption. Therefore correlations may not be valid for different systems. Nevertheless, it can be stated that the different test procedures provide reliable data and can be regarded as useful screening tools for identifying problems.
80
,,,~l,,,,i,,,,i,,,,i,,,,i,,,,i,,,,i,,~,l,,,,i,,,,-
140 70 120
~
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0=
80
~
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=
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~
20
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60 50 ¢d
40
E
30 20 10 0 0
0
10
20
30
40
50
60
70
80
90
I00
T i m e (days) Fig. 4. Effluent monitoring with the luminescent bacteria test before and after shock loading with 2.3-dichlorophenol. The GLValues in the effluent (e) and the 2.3-dichlorophenol concentration (dotted line) are indicated.
336 Table 3. Bacterial toxicity data for 2.3-diehlorophenol and 3.5-dichlorophenol
Compound
Test system
EC50 (mg/i)
References
2.3-DCP
Nitrification inhibition Luminescent bacteria inhibition
0.09 4
Strotmann and Egls~ier, 1994 Strotmann and Egls~ier, 1994
Respiration inhibition
120
Strotmarm et al., 1994
Growth inhibition
40-60
Strotmann et al., 1994
Nitrification inhibition
0.5
Luminescent bacteria inhibition Luminescent bacteria inhibition Luminescent bacteria inhibition Respiration inhibition Respiration inhibition Respiration inhibition Respiration inhibition Respiration inhibition Growth inhibition
5 3.9 3 10 25-60 11-12 11 20-30 5- 10
Strotmann and Egls~er, 1994 Strotmann and Egls~er, 1994 Einabarawy et al., 1988 Dutka et al., 1983 Strotmann et al., 1994 Broecker and Zahn, 1977
Pagga, 1981 Strotmann et al., 1994
DHA DHA (pH 7.0)
80 >100
Dutka et al., 1983 Strotmann et al., 1993
DHA (pH 6.5)
50
Strotmann et al., 1993
DHA (pH 6.0)
32
Strotmann et al., 1993
3.5-DCP
Klecka et al., 1985 EInabarawy et al., 1988
Acknowledgements The authors thank C.Schmitt for skilful technical assistance and U. Pagga for helpful discussions. We also thank D. Charles for correcting the manuscript. 5. References Airbuckle, W.B. and Alleman, J.E (1992). Effluent toxicity testing using nitrifiers and Microtox®. Water Environ.Res. 64: 263-267.
337 Anderson, K., Koopman, B. and Britton, G. (1988). Evaluation of INT-dehydrogenase assay for heavy metal inhibition of activated sludge. Water Res. 22: 349-353. Broeeker, B. and Zahn, R. (1977). The performance of activated sludge plants compared with the results of various bacterial toxicity tests - a study with 3.5-dichlorophenol. Water Res. 11: 165-172. Brown, D., Hitz, H.R. and Schaefer, L. (1981). The assessment of the possible inhibitory effect of dyestuffs on aerobic wastewater bacteria - experience with a screening test. Chemosphere 10: 245-261. DIN 38412 Part 341 (1993). German standards for the determination of water, wastewater and sludge; bioassays; determination of the inhibitory effect ofwastewater on the light emission of
Photobacterium phosphoreum. Dutka, B.J., Nyhol, N. and Petersen, J. (1983). Comparison of several microbiological toxicity screening tests. Water Res. 17: 1363-1368. EEC Directive 79831 Annex V (1983). Part C: Methods for determination of ecotoxicity; 5.2. Degradation- biotic degradation; C.5. Activated sludge simulation test. Elnabarawy, M.T., Robideau, R.R. and Beach, S.A. (1988). Comparison of three rapid toxicity test procedures: Microtox®, Polytox® and activated sludge respiration inhibition. Toxicity Assessment 3: 361-370. Fentem, J. and Balls, M. (1993 ).Replacement of fish in ecotoxicology testing: use of bacteria, other lower organisms and fish cells in vitro. In: Richardson, M (ed.) Ecotoxicology monitoting, pp. 71-81, VCH Publishers, Weinheim. Green, BB., Willets, D.G. and Bennett, M. (1975). Applications of toxicity testing to sewage treatment processes. Water Pollut. Control 74:40-54. King, E.F. (1984). A comparative study of methods for assessing the toxicity to bacteria of single chemicals and mixtures. In: Liu, D. and Dutka, B.J. (eds.) Toxicity screening procedures using bacterial systems, pp. 175-194, Marcel Dekker, New York.
338 King, E.F. and Painter, H.A. (1986). Inhibition of respiration of activatexl sludge" variability and reproducibility of results. Toxicity Assessment 1: 27-39. Klecka, G.M., Landi, L.P. and Bodner, KM. (1985). Evaluation of the OECD activated sludge respiration inhibition test. Chemosphere 14:1239-1251. Kroiss, H., Schweighofer, P., Frey, W. and Matsche, N. (1992). Nitrification inhibition. A source identification method for combined municipal and/or industrial wastewater treatment plants. Wat.Sci.Tech. 26:1135-1146. Pagga, U. (1981). The short-term respiration test - a simple method to check the respiratory activity of activated sludge. Vom Wasser 57: 263-276. Pagga, U. and Gtinthner, W. (1981). The BASF Toximeter- a helpful instrument to control and monitor biological wastewater treatment plants. Wat.Sci.Tech. 13: 233-276. Strotmann U. J. and Eglsaer, H. (1994). The toxicity of substituted phenols in the nitrification inhibition test and the luminescent bacteria test. Ecotoxic.Environm.Saf., in press. Strotmann, U.J., Egls~ier, H. and Pagga, U. (1994). Development and evaluation of a growth inhibition test with sewage bacteria for assessing bacterial toxicity of chemical compounds. Chemosphere 28: 755-766. Strotmann, U. J., Butz, B. and Bias, W.R. (1993) The dehydrogenase assay with resazurin: practical performance as a monitoring system and pH-dependent toxicity of phenolic compounds. Ecotoxic.Environm.Saf. 79-89. Summers, S.M. and Slon, R.A. (1981). Real-time monitoring of biomass respiration in an activated sludge system. In: Proceedings of the 36th Industrial Waste Conference, pp. 701-710. Ann Arbor Science, MI. Thibault, G.T. and Tracy, K.D. (1978). Controlling and monitoring activated sludge units. Chem. Eng. 85: 155-160.