4,4'-methylenebis(cyclohexylamine)

Toxicity to microorganisms

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

activated sludge respiration inhibition testing

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Justification for type of information:

GLP guideline study (ISO 8192) of the structural analogue DM-PACM (CAS 6864-37-5). PACM and DM-PACM are considered read-across analogues based on structural similarity and similar physico-chemical and toxicological properties. The common structural features of the two substances are: a common functional primary amine group; the amines are bound to a cyclic aliphatic organic substituent; there are no elements other than carbon, hydrogen and nitrogen; identical structures except for a methyl group on each cyclohexane, ortho to the amine on DM-PACM similar molecular weights, both under 500 daltons, qualifiyng as “low molecular weight” compounds. For a detailed read across justification see also the attached justification document.

Reason / purpose for cross-reference:

read-across source

Duration:

30 min

Dose descriptor:

other: EC20

Effect conc.:

160 mg/L

Nominal / measured:

nominal

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Duration:

30 min

Dose descriptor:

other: Highest test concentration below a respiration inhibition of 20%

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Results on activated sludge respiration over 30 min

Parameters	Nominal concentration [mg/L]
	10	50	100	500	1000
DOC concentration of the test item [mg/L]	7	37	74	570	740
Respiration speed [mg/L*h]	25	23	21	18	18
Respiration inhibition [%] compared to blank value*	0	8	16	28	28

*significant effect > 20% inhibition compared to blank value

Validity criteria fulfilled:

yes

Conclusions:

Read across to target substance DMDC: EC20 of 160 mg/l was determined

Executive summary:

The analogue chemical DMDC shows comparable physico-chemical behavior, ecotoxicity and toxicity to that of the target chemical PACM. Available data on aquatic toxicity indicate that toxicity is generally slightly lower for PACM than for its analogue DM-PACM. Therefore, read across to ecotoxicity data from DMDC is justified(for detailled justification see field "justification for type of information")and can be used in a worst-case approach.

A short-term respiration inhibition study according to ISO 8192 with domestic activated sludge was conducted. After 30 minutes an EC20 of 160 mg/l was determined.

Reference 2

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

supporting study

Study period:

1987-08-20

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study without detailed documentation

Qualifier:

according to guideline

Guideline:

other: German Industrial Standard DIN 38412, Part 27 (draft): ROBRA- Test; Test on inhibition of oxygen consumption

GLP compliance:

no

Analytical monitoring:

no

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Stock solution (10000 mg/L, pH 11.8): stirred for 20 h.
- Evidence of undissolved material (e.g. precipitate, surface film, etc): no

Test organisms (species):

Pseudomonas putida

Details on inoculum:

- Laboratory culture: yes
- Initial biomass concentration: extinction (E: 436 nm/1 cm; dilution of 1:100) = 0.320
- Age of bacterial suspension: 22 h

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

30 min

Test temperature:

25 °C

pH:

7.2 (lowest test conc.) to 11.8 (highest test conc. and stock solution)

Nominal and measured concentrations:

nominal: 0 (control), 19.5, 39, 78, 156, 312, 625, 1250, 2500, 5000, and 10000 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Material, size, headspace, fill volume: glass vessels with flattened bottom and silicon-foam caps, 10 mL fill volume
- No. of organisms per vessel: 1 mL of bacterial pre-culture (10 TE/F)
- No. of vessels per concentration (replicates): 2
- No. of vessels per control (replicates): 2
- TEST MEDIUM: AK-Medium according to DIN 38412, Part 8 (draft)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Fluorometric determination of growth as extinction at 436 nm after 17 h


TEST CONCENTRATIONS
- Spacing factor for test concentrations: 2

Duration:

30 min

Dose descriptor:

EC10

Effect conc.:

80 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other: based on not pH-adjusted test solutions

Duration:

30 min

Dose descriptor:

EC50

Effect conc.:

ca. 156 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Remarks on result:

other: not reported, but taken from presented data.

Respiration

Test substance conc. (mg/L)	respiration rate: replicate 1 (mg/mL*min)	respiration rate: replicate 2 (mg/mL*min)	respiration rate: mean value (mg/mL*min)	% of Control	pH
C	2.3	2.10	2.200	100
19.5	>C	>C	>100	>100	7.2
39	>C	>C	>100	>100	7.2
78	2.15	2.25	2.200	100	7.2
156	1.15	1.15	1.150	52.3	7.9
312	0	0	0	0	10.1
625	0	0	0	0	10.5
1250	0	0	0	0	10.7
2500	0	0	0	0	11.1
5000	0	0	0	0	11.3
10000	0	0	0	0	11.8

C = Blank

Conclusions:

Based on the nominal concentrations of the test item, EC10 in the Pseudomonas cell multiplication inhibition test (ROBRA) was 80 mg/L and the EC50 was 156 mg/L.

Executive summary:

The toxicity of the test item was examined according to German Industrial Standard DIN 38412, Part 27 (draft): ROBRA- Test; Test on inhibition of oxygen consumption

Psaudomonas putida was exposed with ten nominal test concentrations (Blank, 19.5, 39, 78, 156, 312, 625, 1250, 2500, 5000, 10000 mg/L) of the test item and the reduction of oxygen consumption was determined.

Based on the nominal concentrations of the test item, EC10 in the Pseudomonas cell multiplication inhibition test (ROBRA) was 80 mg/L and the EC50 was 156 mg/L.

Reference 3

Endpoint:

activated sludge respiration inhibition testing

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

comparable to guideline study with acceptable restrictions

Remarks:

e.g. activated sludge from industrial source used instead of municipal wwtp

Qualifier:

according to guideline

Guideline:

ISO 8192 (Water quality - Test for inhibition of oxygen consumption by activated sludge for carbonaceous and ammonium oxidation)

Version / remarks:

part determination of respiration inhibition of activated sludge

Deviations:

not specified

GLP compliance:

no

Analytical monitoring:

no

Vehicle:

no

Test organisms (species):

activated sludge, industrial

Details on inoculum:

- Laboratory culture: Activated sludge from the industrial wastewater treatment plant of BASF, production site Ludwigshafen, Germany

Test type:

static

Water media type:

freshwater

Remarks:

inocolum: activated sludge from industrial plant

Limit test:

no

Total exposure duration:

180 min

Test temperature:

ambient

Nominal and measured concentrations:

nominal: 0 (control), 100, 500, 1000 mg/L
Stock solution 3000 mg/L

Details on test conditions:

syntheticsewage consisting of peptone, meat extract,urea, sodium chloride, 7 mg; calcium chloride, magnesium sulphate deionised water

Reference substance (positive control):

no

Duration:

30 min

Dose descriptor:

other: EC20

Effect conc.:

> 1 000 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Duration:

180 min

Dose descriptor:

EC50

Effect conc.:

ca. 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

inhibition of total respiration

Remarks:

respiration rate

Details on results:

No inhibition after 30 min-exposure even at the highest concentration (1000 mg/L)

Duration of incubation: 30 min

Test concentration	Oxygen concentration (mg/L)			Respiration rate	Respiration change
(mg/L; nominal)	start	end	difference	[mg O2/L/h]	[%]
0 (blank)	4.61	2.65	1.96	29.4	0.130
100	4.95	3.32	1.64	24.5	-16.5
500	5.33	3.60	1.73	26.0	-11.5
1000	4.53	2.82	1.71	25.6	-12.8

 

Duration of incubation: 180 min

Test concentration	Oxygen concentration (mg/L)			Respiration rate	Respiration change
(mg/L; nominal)	start	end	difference	[mg O2/L/h]	[%]
0 (blank)	5.39	3.80	1.59	23.8	0.040
100	6.40	5.46	0.943	14.1	-40.6
500	6.75	6.30	0.454	6.81	-71.4
1000	6.76	5.76	1.00	15.0	-36.9

 

Conclusions:

Based on the nominal concentrations of the test item, 30 min EC20 in the Test for inhibition of oxygen consumption by activated sludge according to ISO 8192 was >1000 mg/L and the 180 min EC50 was approximately 100 mg/L.

Executive summary:

The toxicity of the test item was examined by determination of respiration inhibition of activated sludge according to ISO 8192.

Activated sludge was exposed with 3 nominal test concentrations (Blank, 100, 500, 1000 mg/L) of the test item and the reduction of oxygen consumption was determined.

Based on the nominal concentrations of the test item, EC20 was 80 mg/L and the EC50 was >1000 mg/L and the 180 min EC50 was approximately 100  mg/L.

Description of key information

- Read across from a short-term respiration inhibition study according to ISO 8192 with the structural analogue DMDC and domestic activated sludge was used as key study. After 30 minutes an EC20 of 160 mg/l was determined.

The read across result is supported by two experimental studies with PACM:

- Pseudomonas putida 30 -min respiration rate EC10 was 80 mg/L.

- Respiration inhibition study with activated sludge from industrial WWTP, the 30 -min EC20 was > 1000 mg/L.

Additional information

Read across to structural analogue substance:

The analogue chemical DMDC shows comparable physico-chemical behaviour, ecotoxicity and toxicity to that of the target chemical PACM. Available data on aquatic toxicity indicate that toxicity is generally slightly lower for PACM than for its analogue DMDC. Therefore, read across to ecotoxicity data from DMDC is justified (for detailed justification see field "justification for type of information" in the study endpoint record for the target chemical PACM) and can be used in a worst-case approach.

 

 

A short-term respiration inhibition study according to ISO 8192 with DMDC and domestic activated sludge was conducted. After 30 minutes an EC20 of 160 mg/l was determined.

 

The read across result from DMDC revealing low toxicity towards microorganisms is supported by two experimental studies with PACM:

In a study with Pseudomonas putida a 30 -min EC10 was determined to be 80 mg/L based on the respiration rate (BASF AG, 1987; report no. 01/87/0792).

In a short-term respiration inhibition study with activated sludge from a WWTP treating industrial wastewater, the 30 -min EC20 was determined to be > 1000 mg/L (BASF AG, 1986; report no. 01.114/86).