N-[3-(trimethoxysilyl)propylcyclohexylamine]

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

26 - 30 Apr 2004

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

guideline study with acceptable restrictions

Remarks:

analysis of test media at the end of the test was without algae

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Version / remarks:

1992

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

1984

GLP compliance:

yes (incl. QA statement)

Remarks:

Hessisches Ministerium für Umwelt, Landwirtschaft und Forsten, Wiesbaden, Germany

Analytical monitoring:

yes

Remarks:

TOC

Details on sampling:

- Concentrations: Samples were taken from all concentrations, the control and the blank controls at test start and at 24, 48 and 72 h.
- Sampling method: Duplicate samples were taken at the start of the test from the freshly prepared test media (without algae) of all test concentrations and from the control. For the determination of the stability of the test item under the test conditions, sufficient volumes of the freshly prepared test media of all test concentrations and the control were incubated under the same conditions as in the actual test (but without algae), and were sampled in duplicates at the end of the test. From the control samples only one of the duplicates was analyzed.
- Sample storage conditions before analysis: Most of the samples were analyzed directly. Samples which were not measured directly were stored in the refrigerator at 4 ±4 °C.

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: Since the test item hydrolyses in water quickly, the hydrolysis product was mainly tested. Thus, the highest concentration of 100 mg/L was prepared by dosing 99.5 mg test item into 995 mL test water and stirred for 24 h at room temperature. Then adequate volumes of the stock solution were diluted with the test water to prepare the remaining concentrations.

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: green algae
- Source (laboratory, culture collection): Originally from "Sammlung von Algenkulturen, Pflanzenphysiologisches institut der Universität Göttingen, Germany".
- Age of inoculum (at test initiation): Cells were taken from an exponentially growing pre-culture, which was set up 4 d prior to the test start at the same conditions as in the test.
- Method of cultivation: Algae were cultivated in the laboratory under standardized conditions according to the test guidelines.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

0.24 mmol/L (=24 mg/L) as CaCO3

Test temperature:

23 °C

pH:

8.0 - 8.2 at test start
8.3 - 10.5 at test end

Nominal and measured concentrations:

Nominal: control, 0.05, 0.19, 0.67, 2.33, 8.16, 28.57, and 100 mg test item/L
86% and 87% of the nominal test item concentrations were found in the analyzed test media at the start and at the end of the test respectively (TOC).

Details on test conditions:

TEST SYSTEM
- Test vessel: 50 mL Erlenmeyer flasks
- Type: Erlenmeyer flasks were covered with glass dishes
- Fill volume: 50 mL fill volume
- Initial cells density: 10 000 algal cells/mL test medium
- Control end cells density: 76 x 10^4 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6
- Other: One replicate per test concentration was prepared without algae and was used as a blank for the spectrophotometrical measurements.

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: according to the guideline
- Intervals of water quality measurement: The pH of the test media was measured in all concentrations and the control at the start and at the end of the test. The temperature was measured daily in an Erlenmeyer flask filled with water and incubated under the same conditions as the test flaks. The behavior of the test item in the medium was determined daily in all test concentrations.

OTHER TEST CONDITIONS
- Photoperiod: continuous illumination
- Light intensity and quality: 7470 - 7990 Lux (fluorescent tubes)
- Other: continuous stirring

EFFECT PARAMETERS MEASURED (with observation intervals if applicable):
- Determination of cell concentrations: Determiend by spectrophotometrical measurement. The cell density in one control was counted by microscope after 72 h. Based on the determined absorption of the control and five dilutions of the control, a linear regression was performed for the calculation of the cell densities in all other samples measured spectrophotometrically during the test.
- Other: One sample was taken after 72 h and the shape of the algae compared to the control was examined microscopically.

RANGE-FINDING STUDY
- A range finding test was performed in order to determine the test concentrations. This test was not performed according to GLP regulations and data are excluded from the final report.

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

40.71 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI: 31.36 - 53.60 mg/L

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

16.88 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI: 10.24 - 22.88 mg/L

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): No microscopically visual abnormalities up to the highest test concentration.
- Effect concentrations exceeding solubility of substance in test medium: At the two highest test concentrations of nominal 28.57 and 100 mg/L the test medium was turbid after 72 h.

Reported statistics and error estimates:

Results of Dunnett-test (a=0.05, one sided) with biomass:
Calculated t-Values showed a significant difference of the biomass
- between the control and the concentrations of 28.57 and 100 mg/L (nominal) at 0-24 h, 0-48 h and 0-72 h.

Results of the Dunnett-test (a=0.05, one sided) with the growth rate:
Calculated t-Values showed significant differences of the growth rate:
- between the concentration of 28.57 mg/L and the control at 0-24 h and 0-72 h
- between the control and the 100 mg/L concentration at 0-24 h, 0-48 h and 0-72 h

ANALYTICAL RESULTS:

86% of the nominal test item concentrations were found in the analyzed test media at the start of the test while 87% of the nominal values were recovered after 72h incubation. Therefore, the reported results are expressed as nominal values.

Table 1: pH values in the test media at the start and at the end of the test

Concentration [mg/L]	pH values
	start	end
control	8.2	10.3
0.05	8	10.2
0.19	8.2	10.5
0.67	8.1	9.7
2.33	8.1	9.7
8.16	8.1	9.6
28.57	8.2	8.9
100	8.2	8.3

The pH increase during the test was caused by CO2 consumption of the algae due to their rapid growth (although the test media have been stirred).

BIOLOGICAL RESULTS:

Table 2: Algal cell densities during the test period of 72 h

Concentration [mg/L]	Density of algal cells (x 10 000/mL)
	24 h	48 h	72 h
control	3.3	18.6	76
0.05	3.6	20.2	72.6
0.19	3.7	19.3	73.9
0.67	3.4	20.5	78.4
2.33	5	19.7	72.6
8.16	3.6	18.2	63.4
28.57	1.8	8	31.5
100	-1.6	-0.4	-0.6

Table 3: % Inhibition of growth rate between 0 and 72 h.

Concentration [mg/L]	% inhibition of growth rate 0-72 h
control	0
0.05	1.1
0.19	0.6
0.67	-0.7
2.33	1.1
8.16	4.2
28.57	20.5
100	100

Further effect concentrations mentioned in the report:

- NOEC (72 h) = 2.33 mg/L

- LOEC (72 h) = 8.16 mg/L

VALIDITY CRITERIA:

Table 1: Validity criteria for OECD 201

Criterion from the guideline	Outcome	Validity criterion fulfilled
The biomass in the control cultures should have increased exponentially by a factor of at least 16 within the 72-hour test period.	The cell density in the control increased by a factor of 76 within 72 h	yes
The mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3, for 72-hour tests) in the control cultures must not exceed 35%	The coefficient of variation of the sectional growth rate in the control was 19.2	yes
The coefficient of variation of average specific growth rates during the whole test period in replicate control cultures must not exceed 7% in tests with Pseudokirchneriella subcapitata and Desmodesmus subspicatus. For other less frequently tested species, the value should not exceed 10%.	The coefficient of variation within the control after 72 h was 4.4	yes

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Conclusions:

An ErC10 (72 h) of 16.88 mg/L and an ErC50 (72 h) of 40.71 mg/L were determined in the study testing the toxicity of the substance on Desmodesmus subspicatus according to the OECD guideline 201.

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

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

Adequacy of study:

key study

Justification for type of information:

Please refer to the attached justification below and the overall justification for grouping of substances attached in IUCLID Section 13.

Reason / purpose for cross-reference:

read-across source

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

40.71 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI: 31.36 - 53.60 mg/L

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

16.88 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI: 10.24 - 22.88 mg/L

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): no microscopically visual abnormalities up to the highest test concentration
- Effect concentrations exceeding solubility of substance in test medium: at the two highest test concentrations of nominal 28.57 and 100 mg/L the test medium was turbid after 72 h

Description of key information

ErC50 (72 h) = 40.71 mg/L and ErC10 (72 h) = 16.88 mg/L (nominal, OECD 201) based on read across from CAS 27445-54-1.

Key value for chemical safety assessment

EC50 for freshwater algae:

40.71 mg/L

EC10 or NOEC for freshwater algae:

16.88 mg/L

Additional information

Experimental data on the toxicity of N-[3-(trimethoxysilyl)propylcyclohexylamine] (CAS 3068-78-8) towards algae are not available. Therefore, in accordance with the Regulation (EC) No. 1907/2006 Annex XI, 1.5 “Grouping of substances and read across” and the read across assessment framework (RAAF, ECHA 2017) a read across from the analogue substance N-{[Diethoxy(methyl)silyl]methyl}cyclohexanamine (CAS 27445-54-1) has been performed. Details on the read-across justification can be found in the attached justification in the respective target entry and in the overall justification for grouping of substances attached in IUCLID Section 13.

The influence of the analogue substance N-{[Diethoxy(methyl)silyl]methyl}cyclohexanamine (CAS 27445-54-1) on the growth of the green algal species Scenedesmus subspicatus was investigated in a 72-hour static test according to the OECD guideline 201 and GLP standards (2004). The nominal test item concentrations used were 0.05, 0.19, 0.67, 2.33, 8.16, 28.57, and 100 mg /L and a control. The test item concentrations were analytically monitored at the beginning of the test and at 24, 48 and 72 h via TOC measurements. The analytically determined mean test item concentrations were between 86 and 87% of the nominal values. Since static conditions were used for the test and the hydrolysis of the substance at neutral pH is rapid (half-life = 4.1 min at pH 7), it is assumed that algae were exposed to both, the parent compound and the hydrolysis products. The study resulted in an ErC50 (72 h) of = 40.71 mg/L and an ErC10 (72 h) of 16.88 mg/L (nominal).

Based on the results of the analogues substance and the attached read across justification, similar effects are expected by the target substance N-[3-(trimethoxysilyl)propylcyclohexylamine] (3068-78-8).