4,4'-methylenebis(cyclohexylamine)

Long-term toxicity to aquatic invertebrates

Link to relevant study record(s)

Reference

Endpoint:

long-term toxicity to aquatic invertebrates

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 (OECD 211) 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, qualifying 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:

21 d

Dose descriptor:

LC50

Effect conc.:

5.48 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

(analytically verified)

Basis for effect:

mortality

Remarks:

(parental mortality)

Remarks on result:

other: 95% CL: 2.55 - 33.0 mg/L

Duration:

21 d

Dose descriptor:

EC50

Effect conc.:

> 7.2 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

(analytically verified)

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

7.2 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

(analytically verified)

Basis for effect:

reproduction

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

4 mg/L

Nominal / measured:

nominal

Conc. based on:

act. ingr.

Remarks:

(analytically verified)

Basis for effect:

reproduction

Details on results:

- Mortality of parent animals: control: 10%
- Mean living young per surviving adult after 21 d: control: 205
- Average time to first brood release or time to hatch: control: 8.1 d
- Effect concentrations exceeding solubility of substance in test medium: no
- The analytically measured concentrations of the test substance in the test water were within the range of ± 20% of the nominal concentration in all tested concentrations without any exception.

Results with reference substance (positive control):

- Reference substance: potassium dichromate
- 48-h EC50: 0.9 mg/L
- Results with reference substance valid? yes
(required: 0.6 - 2.1 mg/L according to OECD 202)

Table 1: Measured concentration of DMDC during a 21 -day exposure of Daphnia magna under semi-static conditions

Nominal conc [mg/L]	Measured conc [mg/L]						Time-weighted mean [mg/L]	Percent of nominal [%]
	0 day new	2 day old	6 day new	8 day old	13 day new	15 day old
Original test
Control	< 0.004	< 0.004	< 0.004	< 0.004	< 0.004	< 0.004	-	-
0.21	0.20	0.22	0.21	0.21	0.21	0.22	0.21	100.0
0.38	0.39	0.39	0.38	0.39	0.38	0.38	0.38	100.0
0.69	0.71	0.71	0.69	0.70	0.70	0.69	0.70	101.4
1.23	1.24	1.24	1.24	1.25	1.24	1.23	1.24	100.8
2.22	2.25	2.23	2.22	2.26	2.25	2.22	2.24	100.9
4.00	4.05	4.03	4.04	4.14	4.04	4.03	4.06	101.5
	0 day new	2 day old	7 day new	9 day old	14 day new	16 day old
Supplemental test
Control	< 0.004	< 0.004	< 0.004	< 0.004	< 0.004	< 0.004	-	-
7.20	8.36	8.08	7.28	7.84	7.28	7.82	7.77	107.9
13.0	14.4	14.0	- (a)	- (a)	- (a)	- (a)	13.0	109.2
23.3	24.6	24.0	- (a)	- (a)	- (a)	- (a)	23.3	104.3

new: freshly prepared test solutions

old: test solutions after 2 days exposure

(a): no measurement was made because all Daphnia magna were dead at this observation time

 

Table 2: Cumulative numbers of dead parental Daphnia

Nominal conc [mg/L]	Days
	0	1	2	3	4	5	6	7	8	9	10	11	12	13	14	15	16	17	18	19	20	21
Original test
Control	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	1	1
0.21	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	2	2	2	2
0.38	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
0.69	0	0	0	0	0	0	1	1	1	1	1	1	1	1	1	1	1	1	1	3	3	3
1.23	0	0	0	0	0	0	0	1	1	1	1	1	1	1	1	1	1	1	2	2	2	2
2.22	0	0	0	1	1	2	2	2	2	2	2	2	2	2	2	2	2	2	6	6	6	6
4.00	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	2	2	2	2
Supplemental test
Control	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0
7.20	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	0	1	1	2
13.0	0	0	8	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10
23.3	0	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10	10

 

Table 3: Mean cumulative numbers of juveniles produced per adult

Nominal conc [mg/L]	Days
	0	1	2	3	4	5	6	7	8	9	10	11	12	13	14	15	16	17	18	19	20	21
Original test
Control	0.0	0.0	0.0	0.0	0.0	0.0	0.0	1.4	9.8	13.6	29.4	49.1	60.3	78.8	100.4	111.1	128.0	148.3	158.9	176.4	193.4	204.8
0.21	0.0	0.0	0.0	0.0	0.0	0.0	0.0	1.0	4.5	11.8	23.1	40.8	54.0	79.0	92.5	106.5	130.8	139.6	144.4	171.8	182.0	189.0
0.38	0.0	0.0	0.0	0.0	0.0	0.0	0.0	1.4	6.4	6.4	22.5	39.3	42.8	89.6	89.6	93.7	121.0	139.0	143.4	167.7	186.0	190.6
0.69	0.0	0.0	0.0	0.0	0.0	0.0	0.0	3.9	7.3	13.6	29.1	39.4	55.3	94.3	94.3	110.3	142.1	142.3	148.4	169.4	170.0	191.1
1.23	0.0	0.0	0.0	0.0	0.0	0.0	0.0	4.3	10.6	10.6	33.8	43.9	44.0	101.6	101.6	101.6	149.5	152.9	152.9	204.4	204.5	204.5
2.22	0.0	0.0	0.0	0.0	0.0	0.0	0.0	3.8	5.3	9.3	29.3	29.3	37.0	75.3	75.3	102.8	133.0	133.0	153.8	176.8	176.8	202.5
4.00	0.0	0.0	0.0	0.0	0.0	0.0	0.0	2.5	4.1	14.5	27.3	27.3	55.5	84.8	84.8	114.1	124.1	124.1	137.1	163.4	163.4	192.5
Supplemental test
Control	0.0	0.0	0.0	0.0	0.0	0.0	0.0	6.2	8.5	14.1	40.8	44.6	60.1	95.5	102.9	120.5	151.9	156.8	171.5	199.7	202.7	215.0
7.20	0.0	0.0	0.0	0.0	0.0	0.0	0.0	1.6	6.3	6.3	18.1	22.4	34.3	49.6	56.3	72.4	97.0	97.0	115.4	140.8	140.8	157.8
13.0	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)
23.3	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)	- (a)

(a): all parental daphnia were dead during 21 days of exposure period

 

BIOLOGICAL RESULTS

Reproduction and Mortality Summary after 21 days

Nominal concentration [mg/L]	Reproduction per surviving parent		Mortality
			Parent animals (%)
	Mean living young (S.D.)^a	% inhibition rate^b
0 (control)	205 (18.1)	-	10
0.21	189 (15.5)	8	20
0.38	191 (23.2)	7	0
0.69	191 (22.2)	7	30
1.23	205 (21.7)	0	20
2.22	203 (9.04)	1	60
4.00	193 (39.1)	6	20
Supplemental test
0 (control)	215 (21.5)	-	0
7.20	158 (54.8)	26.6*	20
13.0	-		100
23.3	-		100

^a:Standard deviation.

^b: Effect relative to control.

*: Indicates a significant difference (α = 0.05) from the control (by F&t-test procedure, two-sided test)

 

Other biological observations among surviving parent animals in each concentration after a 21 day exposure

Test conc. [mg/L, nominal]	Mean days to first brood
0 (control)	8.1
0.21	8.4
0.38	8.7
0.69	7.9
1.23	7.6
2.22	8.5
4.00	8.0
Supplemental test
0 (control)	7.6
7.20	8.0
13.0	-
23.3	-

*: p ≤ 0.05; **: p ≤ 0.01  (no significant effects were observed)

No additional adverse effects or abnormal behavior were observed in any of the test groups.

 

 

OTHER OBSERVATIONS

 pH-values(always measured in replicate 1)

Nominal concentration [mg/L]	Day of the test
	0^a	2^b	6^a	8^b	13^a	15^b
0 (control)	7.3	7.3	7.2	7.3	7.1	7.2
0.21	7.3	7.5	7.2	7.5	7.1	7.4
0.38	7.3	7.6	7.2	7.5	7.1	7.4
0.69	7.3	7.7	7.3	7.6	7.2	7.4
1.23	7.3	7.7	7.3	7.6	7.2	7.5
2.22	7.4	7.7	7.3	7.7	7.3	7.5
4.00	7.4	7.8	7.4	7.7	7.3	7.5
Day	0^a	2^b	7^a	9^b	14^a	16^b
Supplemental test
Control	7.2	7.1	7.4	7.4	7.4	7.4
7.20	7.4	7.2	7.5	7.6	7.8	7.6
13.0	7.7	7.3	-	-	-	-
23.3	8.4	7.4	-	-	-	-

^a: in the freshly prepared test solution

^b: before renewal in the old test solutions

 

Oxygen content [mg/L](always measured in replicate 1)

Nominal concentration [mg/L]	Day of the test
	0^a	2^b	6^a	8^b	13^a	15^b
0 (control)	8.9	8.6	9.0	8.5	8.3	8.2
0.21	8.7	8.7	8.6	8.4	8.7	8.2
0.38	8.6	8.6	9.0	8.7	8.5	8.2
0.69	8.4	8.6	8.7	8.7	8.3	8.3
1.23	8.6	8.8	9.0	8.4	8.4	8.4
2.22	8.9	8.6	9.0	8.7	8.5	8.4
4.00	8.2	8.9	8.8	8.5	8.3	8.2
Day	0^a	2^b	7^a	9^b	14^a	16^b
Supplemental test
Control	7.7	7.2	8.1	8.0	9.0	8.1
7.20	8.3	7.9	8.8	7.9	9.0	8.3
13.0	8.1	8.0	-	-	-	-
23.3	8.1	7.7	-	-	-	-

^a: in the freshly prepared test solution

^b: before renewal in the old test solutions

 

Total hardness [mmol/L](always measured in replicate 1)

Nominal concentration [mg/L]	Day of the test
	0^a	2^b	6^a	8^b	13^a	15^b
0 (control)	2.50	2.45	2.54	2.53	2.44	2.50
0.21	2.49	2.49	2.57	2.55	2.52	2.53
0.38	2.49	2.49	2.57	2.55	2.51	2.53
0.69	2.49	2.50	2.57	2.54	2.51	2.53
1.23	2.49	2.49	2.56	2.53	2.51	2.51
2.22	2.48	2.48	2.55	2.53	2.50	2.51
4.00	2.46	2.46	2.53	2.51	2.48	2.49
Day	0^a	2^b	7^a	9^b	14^a	16^b
Supplemental test
Control	2.50	2.50	2.54	2.52	2.52	2.50
7.20	2.54	2.55	2.55	2.56	2.53	2.54
13.0	2.54	2.55	-	-	-	-
23.3	2.52	2.54	-	-	-	-

^a: in the freshly prepared test solution

^b: before renewal in the old test solutions

 

Instantaneous temperature [°C]

Nominal concentration [mg/L]	Day of the test
	0^a	2^b	6^a	8^b	13^a	15^b
0 (control)	20.1	19.4	20.8	20.8	20.4	20.1
0.21	20.2	19.4	20.6	20.6	20.3	20.0
0.38	20.4	19.4	20.6	20.6	20.2	19.9
0.69	20.5	19.5	20.5	20.5	20.2	20.0
1.23	20.5	19.6	20.6	20.6	20.2	20.0
2.22	20.4	19.7	20.6	20.6	20.3	20.0
4.00	20.4	19.9	20.6	20.6	20.3	20.0
Day	0^a	2^b	7^a	9^b	14^a	16^b
Supplemental test
Control	20.2	20.1	20.4	20.4	19.8	20.2
7.20	19.9	20.0	20.2	20.3	20.0	19.8
13.0	19.8	20.0	-	-	-	-
23.3	20.0	20.0	-	-	-	-

^a: in the freshly prepared test solution

^b: before renewal in the old test solutions

 

Validity criteria fulfilled:

yes

Conclusions:

Read across to target substance DMDC: The NOEC of 4.0 mg/L indicates that the substance does not present a long-term hazard according to the criteria of Regulation (EC) No 1272/2008) for non-rapidly degradable substances.

Executive summary:

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") and can be used in a worst-case approach.

 

In order to determine chronic effects of 2,2'-dimethyl-4,4'-methylenebis(cyclohexylamine) (CAS 6864 -37 -5) on aquatic invertebrates, a GLP-study according to OECD 211 (21 d-reproduction test) was performed with Daphnia magna. The test concentrations were analytically verified. The mean measured concentrations for the whole exposure period were within ±20% of the nominal concentration. The NOEC was determined to be 4.0 mg/L (reproduction; nominal) (NITE Japan, 2003; NMMP/E00/3090). All validity criteria were fulfilled in this study. Based on the NOEC of 4.0 mg/L chronic effects on aquatic invertebrates are not expected.

Description of key information

The NOEC for aquatic invertebrates was based on read across to the 21 d-reproduction inhibition from the key study on Daphnia magna (NITE Japan, 2003; NMMP/E00/3090).

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates

Dose descriptor:

NOEC

Effect concentration:

4 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.

This value represents one of the long-term aquatic toxicity values for each of three trophic levels (fish, aquatic invertebrates and aquatic algae and plants). The NOEC of 4.0 mg/L was determined for Daphnia the most sensitive species from the three short-term aquatic toxicity studies. Moreover, based on available long-term data (ACR approach for fish and 72hrs ECr10 for green algae) this value represents also the lowest value for long term effects and was used as the point of departure for the derivation of the PNECaquatic (freshwater).