Cuprate(6-), [2-[[[[3-[[4-chloro-6-[[4-[[4-chloro-6-[(3-sulfophenyl)amino]-1,3,5-triazin-2-yl]amino]-2,5-disulfophenyl]amino]-1,3,5-triazin-2-yl]amino]-2-hydroxy-5-sulfophenyl]azo]phenylmethyl]azo]-5-sulfobenzoato(8-)]-, pentasodium hydrogen, (SP-4-3)-

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

activated sludge respiration inhibition testing

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

other:

Justification for type of information:

Chapter R.7b: Endpoint specific guidance
Version 4.0, June 2017

Guidance on Information Requirements and Chemical Safety Assessment (ECHA)

Information from biodegradation and simulation tests
Absence of microbial toxicity can often be inferred from biodegradation studies in the laboratory. The information content of ready biodegradability tests (available as of 1 t/y) can under certain conditions also be used to derive a NOEC. This can be used to avoid new testing. The assumption that the substance under investigation is not inhibitory to the microorganisms when dosed in the test system is implicit in ready biodegradability testing (i.e., EC C.4 A-F, OECD 301A-F (OECD, 1992) and OECD 310 (2006)). If a compound degrades well in a ready biodegradability test, or does not inhibit the degradation of a positive control at a certain concentration, this concentration can be used as a NOEC value. Any Ready Biodegradability Test relying on continuous monitoring, e.g. the MITI I test (EC C.4F; OECD 301C) or the Manometric Respirometry test (EC C.4D; OECD 301F) is considered more reliable for observing the effects of a chemical on the inoculum. A partial or transient toxic effect often results in a delayed mineralisation of the test substance and/or the positive control.

Reference 2

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

1974

Reliability:

4 (not assignable)

Rationale for reliability incl. deficiencies:

documentation insufficient for assessment

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 209 (Activated Sludge, Respiration Inhibition Test

GLP compliance:

not specified

Analytical monitoring:

no

Vehicle:

no

Test organisms (species):

activated sludge

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

3 h

Nominal and measured concentrations:

10 mg/L

Reference substance (positive control):

not specified

Duration:

3 h

Dose descriptor:

IC10

Effect conc.:

> 10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Validity criteria fulfilled:

not specified

Conclusions:

The substance has a 3-h IC10 of >10 mg/L. No negative effects the biodegradation activity of activated sludge in pre-adapted wastewater treatment plants are anticipated when applied at low concentrations.

Reference 3

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

weight of evidence

Study period:

2016-11-15 to 2016-12-15

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Qualifier:

according to guideline

Guideline:

other: OECD 301F (Ready Biodegradability: Manometric Respirometry Test)

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

no

Vehicle:

no

Test organisms (species):

activated sludge of a predominantly domestic sewage

Details on inoculum:

- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Activated sludge from the municipal sewage treatment plant, 31137 Hildesheim, Germany
- Receipt: 2016-11-10
- Preparation of inoculum for exposure: The activated sludge was washed twice with chlorine free tap water. After the second washing the settled sludge was resuspended in mineral salts medium and was maintained in an aerobic condition by aeration for 2 hours. Thereafter the sludge was homogenized with a blender. After sedimentation the supernatant was decanted and maintained in an aerobic condition by aeration with CO2-free air for 6 days. 10 mL/L were used to initiate inoculation.
- Initial cell/biomass concentration: approx. 1.66 x 107 CFU/L in the final test solution
- Water filtered: no

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

28 d

Test temperature:

20.5 - 21.5

pH:

7.7 - 7.9

Nominal and measured concentrations:

45 mg/L

Reference substance (positive control):

no

Duration:

28 d

Dose descriptor:

EC50

Effect conc.:

> 75 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Details on results:

Results of the Toxicity Control
In the toxicity control the biodegradation achieved 56% after 14 days. The biodegradation came to a maximum of 58% on day 19. The validity criterion that the test item should not inhibit the biodegradation of the reference item was fulfilled.

Biodegradation [%] of the Test Item Reactive Blue 160 in Comparison to the Functional Control and Toxicity Control

		Biodegradation [%]
Date	[d]	Functional Control	Test Item		Toxicity Control Test Item + Reference Item
		R1	P1	P2	T1
2016-11-17	1	2	0	0	4
2016-11-18	2	51	0	0	33
2016-11-19	3	66	0	0	40
2016-11-20	4	74	0	0	46
2016-11-21	5	78	1	0	49
2016-11-22	6	80	1	0	50
2016-11-23	7	81	3	2	51
2016-11-24	8	82	2	0	52
2016-11-25	9	83	1	0	52
2016-11-26	10	85	2	0	53
2016-11-27	11	87	1	0	54
2016-11-28	12	88	0	0	55
2016-11-29	13	90	1	0	56
2016-11-30	14	91	2	0	56
2016-12-01	15	91	0	0	56
2016-12-02	16	93	2	0	57
2016-12-03	17	93	1	0	57
2016-12-04	18	93	0	0	57
2016-12-05	19	95	1	0	58
2016-12-06	20	95	1	0	57
2016-12-07	21	96	1	0	58
2016-12-08	22	97	2	0	57
2016-12-09	23	97	1	0	57
2016-12-10	24	97	1	0	57
2016-12-11	25	97	2	0	57
2016-12-12	26	97	1	0	57
2016-12-13	27	98	3	0	57
2016-12-14	28	98	5	0	57

Validity criteria fulfilled:

yes

Conclusions:

The substance was tested for total respiration inhibition as toxicity control in an OECD 301F ready biodegradability study. The functional control with 45 mg/L sodium benzoate reached a maximum oxygen depletion of 98% on day 27. The mean oxygen depletion with 75 mg/L Reactive Blue 160 was 3% (not readily biodegradable). The toxicity control with 45 mg/L sodium benzoate and 75 mg/L Reactive Blue 160 reached a maximum oxygen depletion of 58% on day 21. The substance was not considered inhibitory to activated sludge according to the validity criteria of OECD 301F. Since the biodegradation of sodium benzoate was not inhibited by >50% at a test item concentration of 75 mg/L, the EC50 for total respiration inhibition is > 75 mg/L.

Description of key information

The substance was tested for total respiration inhibition as toxicity control in an OECD 301F ready biodegradability study. The functional control with 45 mg/L sodium benzoate reached a maximum oxygen depletion of 98% on day 27. The mean oxygen depletion with 75 mg/L Reactive Blue 160 was 3% (not readily biodegradable). The toxicity control with 45 mg/L sodium benzoate and 75 mg/L Reactive Blue 160 reached a maximum oxygen depletion of 58% on day 21. The substance was not considered inhibitory to activated sludge according to the validity criteria of OECD 301F. Since the biodegradation of sodium benzoate was not inhibited by >50% at a test item concentration of 75 mg/L, the EC50 for total respiration inhibition is > 75 mg/L.

A second study (1974) stated a 3-h IC10 >10 mg/L to activated sludge (respiration inhibition). However, due to lack of a sufficient documentation, the study was not used for an assessment.

Key value for chemical safety assessment

EC50 for microorganisms:

75 mg/L

Additional information

Chapter R.7b: Endpoint specific guidance

Version 4.0, June 2017

Guidance on Information Requirements and Chemical Safety Assessment (ECHA)

Information from biodegradation and simulation tests

Absence of microbial toxicity can often be inferred from biodegradation studies in the laboratory. The information content of ready biodegradability tests (available as of 1 t/y) can under certain conditions also be used to derive a NOEC. This can be used to avoid new testing. The assumption that the substance under investigation is not inhibitory to the microorganisms when dosed in the test system is implicit in ready biodegradability testing (i.e., EC C.4 A-F, OECD 301A-F (OECD, 1992) and OECD 310 (2006)). If a compound degrades well in a ready biodegradability test, or does not inhibit the degradation of a positive control at a certain concentration, this concentration can be used as a NOEC value. Any Ready Biodegradability Test relying on continuous monitoring, e.g. the MITI I test (EC C.4F; OECD 301C) or the Manometric Respirometry test (EC C.4D; OECD 301F) is considered more reliable for observing the effects of a chemical on the inoculum. A partial or transient toxic effect often results in a delayed mineralisation of the test substance and/or the positive control.

The substance did not significantly inhibit the biodegradation activity of activated sludge in a toxicity control of an OECD 301F study. Since the biodegradation of sodium benzoate was 58% maximum with Reactive Blue 160 and 98% maximum without the test item, the applied concentration of 75 mg/L was not considered a NOEC but a "worst-case" EC50.