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Toxicity to microorganisms

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Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
GLP compliance:
yes
Specific details on test material used for the study:
Nipacide BIT
Batch number: A88
Analytical monitoring:
no
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on test conditions:
Type: aquatic

The effect of Nipacide BIT on the respiration rate of activated sludge was assessed by the methods detailed in OECD Test Guideline 209, 'Activated Sludge, Respiration Inhibition test' adopted 4 April 1984.

Samples of activated sludge (suspended solids 1.6 g/l), fed with synthetic sewage, were exposed to suspensions of the test material for three hours. Their rates of oxygen consumption were determined using an oxygen electrode and compared with those of controls, containing activated sludge and synthetic sewage alone, which were established at the beginning and end of each culture series. Nominal test concentrations of 1, 10, 32, 56 and 100 mg/l were investigated. The reference inhibitor 3,5-dichiorophenol (3,5-DCP) was employed in each test, at 3.2, 10 and 32 mg/l, as a positive control.
Key result
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
13 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Key result
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
12.8 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
inhibition of total respiration
Details on results:
The control and positive control results show that the tests were valid and that the samples of activated sludge employed were sensitive to inhibition. Concentration-related inhibition of respiration rates was obtained at and above test concentrations of 10 mg/l. The highest tested concentration of Nipacide BIT (100 mg/l) caused 90% inhibition. The three-hour 50% effect concentration (EC50) for Nipacide BIT was calculated by the Moving Average method from the results of the final test to be 13.0 mg/l (95% confidence limits 10.3 and 16.0 mg/l). The EC20 and EC80 of Nipacide BIT were estimated to be 3.3 and 44.0 mg/l respectively.
Conclusions:
Results and discussion

The specific respiration rates of the control cultures established at the end of the two test series (31.9 and 30.4 mg O2/g/h) were 101% and 93% of the rates of those established at the start (31.5 and 32.6 mg O2/g/h).
The three-hour 50% effect concentration (EC50) for 3,5-DCP in the final test was calculated to be 9.9 mg/L (95% confidence limits of 7.7 and 12.6 mg/L).
These results show that the tests were valid and that the samples of activated sludge employed were sensitive to inhibition.
Concentration-related inhibition of respiration rates was seen at and above test concentrations of 10 mg/L. The highest tested concentration of BIT (100 mg/L) caused 90% inhibition.
The EC50 for BIT was calculated from the results of the final test to be 13.0 mg/L (95% confidence limits 10.3 and 16.0 mg/L). EC20 and EC80 for BIT were estimated to be 3.3 and 44.0 mg/L, respectively.


EC20 EC20 = 3.3 mg/L BIT

EC50 EC50 = 13.0 mg/L BIT (with confidence limits of 10.3 and 16.0 mg/L)

EC80 EC80 = 44.0 mg/L BIT
The study was conducted with BIT which has a BIT purity of 98.8%. The EC20, EC50 and EC80 data are therefore reported as BIT.

Conclusion
The procedures described in OECD Guideline 209 have been followed and the acceptance criteria of the guideline have been met. The three hour 50% effect concentration (EC50) for the reference inhibitor (3,5 Dichlorophenol) was within the acceptable range of 5 to 30 mg/l and the percentage difference between the respiration rates of the control at the start and end of the test also met the acceptance criteria of ≤ 15%. The highest tested concentration of BIT (nominal concentration of 100 mg/L BIT) resulted in 90% inhibition.
The study was conducted with Nipacide BIT which has a BIT purity of 98.8%. The data reported in this study summary are therefore reported as BIT.

Executive summary:

A study was conducted to determine the toxicity of the substance to activated sludge microorganisms according to OECD Guideline 209. The test system was activated sludge of the municipal treatment plant treating mainly domestic sewage. The study was carried out under static conditions at nominal test substance concentrations of 0, 1, 10, 32, 56 and 100 mg/L for a duration of 3 h. The respiration rates of the control, reference and test substance replicates were measured after a contact time of three hours, and the inhibitory effects of the test and reference substance (3,5 -dichlorophenol) were determined in comparison to the control respiration rates. The validity criteria of the test guideline were fulfilled. Under the study conditions, the nominal 3 h EC50 of the substance was determined to be 13 mg/L (equivalent to 12.8 mg a.i./L) (Jenkins, 1994).

Endpoint:
activated sludge respiration inhibition testing
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Qualifier:
according to
Guideline:
OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test
GLP compliance:
yes
Specific details on test material used for the study:
Proxel Press Paste
Batch Sample Reference Bx 1445 M. Wird 33/9/3
The Test Substance employed was pre-dried technical grade active substance
Purity 93.8%
Analytical monitoring:
no
Test organisms (species):
activated sludge of a predominantly domestic sewage
Details on test conditions:
Type: aquatic
Key result
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
24 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Key result
Duration:
3 h
Dose descriptor:
EC50
Effect conc.:
22.5 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
inhibition of total respiration
Key result
Duration:
3 h
Dose descriptor:
NOEC
Effect conc.:
11 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
inhibition of total respiration
Key result
Duration:
3 h
Dose descriptor:
NOEC
Effect conc.:
10.3 mg/L
Nominal / measured:
nominal
Conc. based on:
act. ingr.
Basis for effect:
inhibition of total respiration
Conclusions:

Results and discussion
The median effect concentration (EC50) defined as the concentration resulting in a 50% reduction in respiration rate of activated sludge within the period of the test was estimated from a graphical plot of percentage inhibition against nominal test concentrations. The three hour EC50 was estimated to be 24.0 mg/l Proxel press paste which is equivalent to 23 mg/l BIT. The EC10 was not reported since 10% inhibition was within the expected experimental variability of the test and was not considered to be an effect of the Proxel press paste. The no observed effect concentration (NOEC) is therefore equivalent to the EC10 in this test. The three hour NOEC was estimated from the concentration/response curve to be 11.0 mg/l Proxel press paste which is equivalent to 10 mg/l BIT
The reference substance, 3,5-dichlorophenol, caused substantial inhibition of the respiration rate of the activated sludge. The 3 hour EC50 value was estimated from a plot of percentage inhibition against nominal concentration of 3,5-dichlorophenol to be I8 mg/l. This is within the expected normal range of 5-30 mg/l, indicating the sludge was responding normally and confirming the viability of the sludge organisms.
The pH measured at the end of the study was in the range 8.0 to 8.1 in all the flasks.

EC20 Not stated.

EC50 Three Hour EC50 - based on nominal concentrations and estimated from the concentration/response curve

Calculated EC50 = 24.0 mg/l Proxel Press paste (dry)

EC80 Not stated.

Conclusion
The procedures described in OECD Guideline 209 have been followed and the acceptance criteria of the guideline have been met. The three hour 50% effect concentration (EC50) for the reference inhibitor (3,5 Dichlorophenol) was within the acceptable range of 5 to 30 mg/l and the percentage difference between the respiration rates of the control at the start and end of the test also met the acceptance criteria of ≤ 15%. The highest tested concentration of Proxel press paste (nominal concentration of 110 mg/1) caused > 90% inhibition.
Executive summary:

A study was conducted to determine the toxicity of the substance to activated sludge microorganisms according to OECD Guideline 209. The test system was activated sludge of the municipal treatment plant treating mainly domestic sewage. The study was carried out under static conditions at nominal test substance concentrations of 0, 1.1, 3.4, 11, 34 and 110 mg/L for a duration of 3 h. The respiration rates of the control, reference and test substance replicates were measured after a contact time of three hours, and the inhibitory effects of the test and reference substance (3,5 -dichlorophenol) were determined in comparison to the control respiration rates. The validity criteria of the test guideline were fulfilled. Under the study conditions, the nominal 3 h NOEC and EC50 of the substance was determined to be 11 mg/L (equivalent to 10.3 mg a.i./L) and 24 mg/L (equivalent to 22.5 mg a.i./L) respectively (Daniel, 2004).

Description of key information

Key value for chemical safety assessment

EC10 or NOEC for microorganisms:
10.3 mg/L

Additional information

A study was conducted to determine the toxicity of the substance to activated sludge microorganisms according to OECD Guideline 209. The test system was activated sludge of the municipal treatment plant treating mainly domestic sewage. The study was carried out under static conditions at nominal test substance concentrations of 0, 1.1, 3.4, 11, 34 and 110 mg/L for a duration of 3 h. The respiration rates of the control, reference and test substance replicates were measured after a contact time of three hours, and the inhibitory effects of the test and reference substance (3,5 -dichlorophenol) were determined in comparison to the control respiration rates. The validity criteria of the test guideline were fulfilled. Under the study conditions, the nominal 3 h NOEC and EC50 of the substance was determined to be 11 mg/L (equivalent to 10.3 mg a.i./L) and 24 mg/L (equivalent to 22.5 mg a.i./L) (Daniel, 2004).

A study was conducted to determine the toxicity of the substance to activated sludge microorganisms according to OECD Guideline 209. The test system was activated sludge of the municipal treatment plant treating mainly domestic sewage. The study was carried out under static conditions at nominal test substance concentrations of 0, 1, 10, 32, 56 and 100 mg/L for a duration of 3 h. The respiration rates of the control, reference and test substance replicates were measured after a contact time of three hours, and the inhibitory effects of the test and reference substance (3,5 -dichlorophenol) were determined in comparison to the control respiration rates. The validity criteria of the test guideline were fulfilled. Under the study conditions, the nominal 3 h EC50 of the substance was determined to be 13 mg/L (equivalent to 12.8 mg a.i./L) (Jenkins, 1994).