N-ethyl-p-methoxy-α-methylphenethylamine

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

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

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

data from handbook or collection of data

Remarks:

for read across substance

Justification for type of information:

Data for the target chemical is summarized based on the structurally similar read across chemicals

Reason / purpose for cross-reference:

read-across source

Reason / purpose for cross-reference:

read-across source

Qualifier:

according to guideline

Guideline:

other: as mentioned below

Principles of method if other than guideline:

WoE report is based on two short term toxicity study of fish for the test chemical :
1. This study was designed to assess the toxic effects of the test compound on the green alga Chlorella vulgaris. Test was conducted in compliance with the OECD guideline 201.
2. The freshwater algal growth inhibition test was carried out on Desmodesmus subspicatus with the test material according to OECD Guideline 201.

Specific details on test material used for the study:

- Name of test material (IUPAC name): 2-(N-ethylamino)-1-(4-methoxyphenil propane)
- IUPAC name: 2-(N-ethylamino)-1-(4-methoxyphenil propane)
- Molecular formula: C12H19NO
- Molecular weight: 193.2881 g/mol
- Smiles notation: CCNC(C)Cc1ccc(OC)cc1
- InChl: 1S/C12H19NO/c1-4-13-10(2)9-11-5-7-12(14-3)8-6-11/h5-8,10,13H,4,9H2,1-3H3
- Substance type: Organic

Analytical monitoring:

yes

Vehicle:

not specified

Details on test solutions:

1.Method: The test solution was prepared in aseptic condition. The test substance was prepared by adding 60 µl of test substance in 300 ml of BBM to get the final concentration of 200 mg/L. The remaining test solutions were prepared by dilution from the above stock solution. To have a better growth and visibility of cells, the initial cell density of the culture was kept 1 X 104 cells/ml. Care was taken to have a homogeneous solution for the experiment.

2. PREPARATION AND APPLICATION OF TEST SOLUTION
The stock solution 150.0 mg/L was prepared by dissolving white powder in OECD growth medium. Test solutions of required concentrations were prepared by mixing the stock solution of the test sample with OECD growth medium and inoculum culture.
- Controls: OECD medium as specified in OECD 201, used as a control and for dilution of the stock solution.

Test organisms (species):

other: 1.Chlorella vulgaris , 2. Desmodesmus subspicatus

Details on test organisms:

1.TEST ORGANISM
- Common name:green alga
- Strain:Chlorella vulgaris
- Source (laboratory, culture collection): National Environmental Engineering Research Institute (NEERI), Nagpur (Laboratory)
- Method of cultivation: Bold’s Basal Medium(BBM)

ACCLIMATION
- Culturing media and conditions (same as test or not):The medium to be used for the growth of algae was Bold’s Basal Medium (BBM). It is a medium composed of macronutrients, micronutrients, alkaline EDTA solution and Iron solution. Stock solution of each of these was prepared separately and then a complete medium was prepared and sterilized. De-ionized water was used to prepare the BBM
- Any deformed or abnormal cells observed:no

2.TEST ORGANISM
- Strain: 86.81 SAG
- Source (laboratory, culture collection): Institute of botany of the ASCR
- Age of inoculum (at test initiation): 5x103 cells/ml
- Method of cultivation: No data

ACCLIMATION
- Acclimation period: No data
- Culturing media and conditions (same as test or not): No data
- Any deformed or abnormal cells observed: No data

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Post exposure observation period:

24, 48, 72 hrs

Test temperature:

1) 24 ° C ± 2°C

pH:

1) 6.8 ± 0.3
2) 7.8

Nominal and measured concentrations:

1) Six test concentration were: 6.25mg/l, 12.5mg/l, 25mg/l, 50mg/l, 100mg/l and 200mg/l.
2) 0, 0, 30.0, 45.0, 67.5, 100.0 and 150.0 mg/L

Details on test conditions:

1)TEST SYSTEM
- Test vessel: Conical flasks
- Type (delete if not applicable): No data available
- Material, size, headspace, fill volume: Conical flasks of 100 ml size filled with 60 ml was used for the study.
- Aeration: No data available
- Type of flow-through (e.g. peristaltic or proportional diluter): No data available
- Renewal rate of test solution (frequency/flow rate): No data available
- Initial cells density: 10000 cells/mL
- Control end cells density: No data available
- No. of organisms per vessel: 10000cells/ml
- No. of vessels per concentration (replicates): Two replicates for each test concentration
- No. of vessels per control (replicates): Three replicates for Control
- No. of vessels per vehicle control (replicates): No data available

GROWTH MEDIUM
- Standard medium used: No data available
- Detailed composition if non-standard medium was used: The medium to be used for the growth of algae was Bold’s Basal Medium (BBM). It is a medium composed of macronutrients, micronutrients, alkaline EDTA solution and Iron solution. Stock solution of each of these was prepared separately and then a complete medium was prepared and sterilized. De-ionized water was used to prepare the BBM.


OTHER TEST CONDITIONS
- Sterile test conditions: Yes
- Adjustment of pH: No data available
- Photoperiod: 16 Hour Light Period : 8 Hour Dark Period
- Light intensity and quality: continuous, uniform fluorescent illumination(1500Lux)
- Salinity (for marine algae): No data available

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Spectrophotometer - The absorbance values of each test vessel and control vessel was noted at 680nm.The BBM was taken as blank for both control and test vessels. The absorbance value of each vessel was in line with the average specific growth rate.
- Chlorophyll measurement: No data available
- Other: Microscopic observations: The cultures were observed daily with the help of a microscope to verify a normal and healthy appearance of the algal culture and also to observe any abnormal appearance of the algae (as may be caused by the exposure of the test item). Apart from this, the cell count of each test vessel was also noted with the help of a microscope and haemocytometer.

TEST CONCENTRATIONS
- Spacing factor for test concentrations: All the six concentrations were in geometric series spaced by a factor of 2.
- Justification for using less concentrations than requested by guideline: No data available
- Range finding study: No data available
- Test concentrations: Six test concentration were: 6.25mg/l, 12.5mg/l, 25mg/l, 50mg/l,100mg/l and 200mg/l (Nominal concentrations)
- Results used to determine the conditions for the definitive study: No data available

6.1. Test solution: The test solution was prepared in aseptic condition. The test item4-amino-5-hydroxynaphthalene-1,7-disulphonic acid was prepared by adding 500 mg of test item in 250ml of BBM to get the final concentration of 200mg/L. This stock solution was kept for stirring for 30minutes to obtain a homogenous solution for the experiment. The test concentrations were chosen according to the available data of the test item.

The remaining test solutions were prepared by dilution from the above stock solution. To have a better growth and visibility of cells, the initial cell density of the culture was kept 1 X 104cells/ml. Care was taken to have a homogeneous solution for the experiment.

6.2. Test vessels: All the tests were carried out in 100mL conical flasks which were carefully autoclaved and sterilized. The test solution in each of these test vessels was kept constant which is 60ml so that a sufficient amount of head space was left.

6.3. Replicates: For the assessment of algal growth, the study was conducted in replicates. The control vessel was maintained in triplicates as recommended in the OECD guideline and the test concentrations were selected in geometric series which were maintained in duplicates.

6.4. Incubation:
i) The temperature of the orbital shaking incubator was kept constant throughout the period of exposure of the experiment. The temperature was maintained at 24°C ± 2°C.
ii) The test vessels were incubated with a continuous, uniform fluorescent illumination (1500Lux).
iii) The pH of the control cultures needs to be noted during the study and the pH of the control medium should not increase by more than 1.5 units during the test.
iv) The orbital shaking incubator was set at a speed of 120 revolutions per minute throughout the study period. This is to provide constant shaking to the algal cells to keep them in suspension and to ensure that they do not settle down on the bottom of the test vessel.

2)TEST SYSTEM
- Test vessel: 50ml glass vessel
- Type : closed (with air-permeable stopper)
- Material, size, headspace, fill volume: 15 ml
- Aeration: No data
- Type of flow-through (e.g. peristaltic or proportional diluter): No data
- Renewal rate of test solution (frequency/flow rate): No data
- Initial cells density: 5x103 cells/mL
- Control end cells density: No data
- No. of organisms per vessel: No data
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): No data
- No. of vessels per vehicle control (replicates): No data

GROWTH MEDIUM
- Standard medium used: Yes

OTHER TEST CONDITIONS
- Adjustment of pH: Without adjustment
- Photoperiod: Continuous
- Light intensity and quality: 6000 lx - 8000 lx

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell counting: microscope with counting chamber Cyrus I or electronic particle counter.
- Chlorophyll measurement: No
- Other: ErC50 was calculated using non-linear regression by the software Prism 4.0

Reference substance (positive control):

yes

Remarks:

Potassium dichromate (K2Cr2O7) for read across 2

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 200 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Read across 1

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

106.9 mg/L

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: Read across 2

Results with reference substance (positive control):

2) - Results with reference substance valid
ErC50: 0.75 mg/L K2Cr2O7

Validity criteria fulfilled:

not specified

Conclusions:

The test chemical 2-(N-ethylamino)-1-(4-methoxyphenil propane) is not likely to be toxic to fish atleast in the dose range of 106-200 mg/l

Executive summary:

Data available for the structurally and functionally similar read across chemicals has been reviewed to determine the toxicity to aquatic algae and cynobacteria of the test chemical 2-(N-ethylamino)-1-(4-methoxyphenil propane) (14367 -46 -4).The studies are as mentioned below:

1.This study was designed to assess the toxic effects of the test compound on the growth of green alga Chlorella vulgaris.

The study was conducted following OECD guideline 201- Alga, growth inhibition test. The test concentration chosen for the study were 6.25mg/l, 12.5mg/l, 25mg/l, 50mg/l,100mg/l and 200mg/l.

All the tests were carried out in 100mL conical flasks which were carefully autoclaved and sterilized. The test solution in each of these test vessels was kept constant which is 60 ml so that a sufficient amount of head space was left. The test solution was prepared in aseptic condition. The test substance was prepared by adding 60 µl of test substance in 300 ml of BBM to get the final concentration of 200 mg/L. The remaining test solutions were prepared by dilution from the above stock solution. To have a better growth and visibility of cells, the initial cell density of the culture was kept 1 X 104 cells/ml. Care was taken to have a homogeneous solution for the experiment.

For the assessment of algal growth, the test was conducted in replicates. The control flask was maintained in triplicates as recommended in the OECD guideline and the test concentration were selected in geometric series which were maintained in duplicates. To obtain a quantitative concentration-response relationship by regression analysis, a linearizing transformation of the response data into probit was performed. Using the same, effective concentration (EC) were determined.

After 72 hours of exposure to test item to various nominal test concentrations, EC50 was determine to be >200mg/l and the EC10 was determine to be 158.49 mg/l graphically and through probit analysis.

2.The freshwater algal growth inhibition test was carried out on Desmodesmus subspicatus with the test material according to OECD Guideline 201.

The stock solution 150.0 mg/L was prepared by dissolving white powder in OECD growth medium. Test solutions of required concentrations were prepared by mixing the stock solution of the test sample with OECD growth medium and inoculum culture and tested at the concentrations 0, 0, 30.0, 45.0, 67.5, 100.0 and 150.0 mg/L.Effects on the growth rate of the organism were studied. The test was performed under static conditions in a static fresh water system at a temperature of 23± 2°C. Initial cell density of test organism used was 5x10(3) cells/ml. Determination of cell counting involve the use of microscope with counting chamber Cyrus I or electronic particle counter. ErC50 was calculated using non-linear regression by the software Prism 4.0.

The median effective concentration (ErC50) for the test material, in a freshwater algae Desmodesmus subspicatus was determined to be 106.9 mg/L on the basis of effects on growth rate in a 72 hour study with 95% Cl of 83.2 - 137.3 mg/L.

Thus, based on the above summarised studies, 2-(N-ethylamino)-1-(4-methoxyphenil propane) and it’s structurally and functionally similar read across substance, it can be concluded that effect concetration value is in the range 106- 200 mg/l. Thus, comparing this value with the criteria of CLP regulation , 2-(N-ethylamino)-1-(4-methoxyphenil propane) cannot be classified for toxicity to aquatic algae and cynobaccteria .Hence,based on the data available for the structurally and functionally similar read across, test chemical 2-(N-ethylamino)-1-(4-methoxyphenil propane) is not likely to be toxic atleast in the concentration range of 106 - 200 mg/L

Description of key information

Toxicity to algae and cynobacteria :

Data available for the structurally and functionally similar read across chemicals has been reviewed to determine the toxicity to aquatic algae and cynobacteria of the test chemical 2-(N-ethylamino)-1-(4-methoxyphenil propane) (14367 -46 -4).The studies are as mentioned below:

1.This study was designed to assess the toxic effects of the test compound on the growth of green alga Chlorella vulgaris.

The study was conducted following OECD guideline 201- Alga, growth inhibition test. The test concentration chosen for the study were 6.25mg/l, 12.5mg/l, 25mg/l, 50mg/l,100mg/l and 200mg/l.

All the tests were carried out in 100mL conical flasks which were carefully autoclaved and sterilized. The test solution in each of these test vessels was kept constant which is 60 ml so that a sufficient amount of head space was left. The test solution was prepared in aseptic condition. The test substance 4-methoxybenzyl alcohol was prepared by adding 60 µl of test substance in 300 ml of BBM to get the final concentration of 200 mg/L. The remaining test solutions were prepared by dilution from the above stock solution. To have a better growth and visibility of cells, the initial cell density of the culture was kept 1 X 104 cells/ml. Care was taken to have a homogeneous solution for the experiment.

For the assessment of algal growth, the test was conducted in replicates. The control flask was maintained in triplicates as recommended in the OECD guideline and the test concentration were selected in geometric series which were maintained in duplicates. To obtain a quantitative concentration-response relationship by regression analysis, a linearizing transformation of the response data into probit was performed. Using the same, effective concentration (EC) were determined.

After 72 hours of exposure to test item to various nominal test concentrations, EC50 was determine to be >200mg/l and the EC10 was determine to be 158.49 mg/l graphically and through probit analysis.

2.The freshwater algal growth inhibition test was carried out on Desmodesmus subspicatus with the test material according to OECD Guideline 201.

The stock solution 150.0 mg/L was prepared by dissolving white powder in OECD growth medium. Test solutions of required concentrations were prepared by mixing the stock solution of the test sample with OECD growth medium and inoculum culture and tested at the concentrations 0, 0, 30.0, 45.0, 67.5, 100.0 and 150.0 mg/L.Effects on the growth rate of the organism were studied. The test was performed under static conditions in a static fresh water system at a temperature of 23± 2°C. Initial cell density of test organism used was 5x10(3) cells/ml. Determination of cell counting involve the use of microscope with counting chamber Cyrus I or electronic particle counter. ErC50 was calculated using non-linear regression by the software Prism 4.0.

The median effective concentration (ErC50) for the test material, in a freshwater algae Desmodesmus subspicatus was determined to be 106.9 mg/L on the basis of effects on growth rate in a 72 hour study with 95% Cl of 83.2 - 137.3 mg/L.

Thus, based on the above summarised studies, 2-(N-ethylamino)-1-(4-methoxyphenil propane) and it’s structurally and functionally similar read across substance, it can be concluded that effect concetration value is in the range 106- 200 mg/l. Thus, comparing this value with the criteria of CLP regulation , 2-(N-ethylamino)-1-(4-methoxyphenil propane) cannot be classified for toxicity to aquatic algae and cynobaccteria .Hence,based on the data available for the structurally and functionally similar read across, test chemical 2-(N-ethylamino)-1-(4-methoxyphenil propane) is not likely to be toxic atleast in the concentration range of 106 - 200 mg/L

Key value for chemical safety assessment

EC50 for freshwater algae:

200 mg/L

Additional information

Toxicity to algae and cynobacteria :

Data available for the structurally and functionally similar read across chemicals has been reviewed to determine the toxicity to aquatic algae and cynobacteria of the test chemical 2-(N-ethylamino)-1-(4-methoxyphenil propane) (14367 -46 -4).The studies are as mentioned below:

1.This study was designed to assess the toxic effects of the test compound on the growth of green alga Chlorella vulgaris.

The study was conducted following OECD guideline 201- Alga, growth inhibition test. The test concentration chosen for the study were 6.25mg/l, 12.5mg/l, 25mg/l, 50mg/l,100mg/l and 200mg/l.

All the tests were carried out in 100mL conical flasks which were carefully autoclaved and sterilized. The test solution in each of these test vessels was kept constant which is 60 ml so that a sufficient amount of head space was left. The test solution was prepared in aseptic condition. The test substance 4-methoxybenzyl alcohol was prepared by adding 60 µl of test substance in 300 ml of BBM to get the final concentration of 200 mg/L. The remaining test solutions were prepared by dilution from the above stock solution. To have a better growth and visibility of cells, the initial cell density of the culture was kept 1 X 104 cells/ml. Care was taken to have a homogeneous solution for the experiment.

For the assessment of algal growth, the test was conducted in replicates. The control flask was maintained in triplicates as recommended in the OECD guideline and the test concentration were selected in geometric series which were maintained in duplicates. To obtain a quantitative concentration-response relationship by regression analysis, a linearizing transformation of the response data into probit was performed. Using the same, effective concentration (EC) were determined.

After 72 hours of exposure to test item to various nominal test concentrations, EC50 was determine to be >200mg/l and the EC10 was determine to be 158.49 mg/l graphically and through probit analysis.

2.The freshwater algal growth inhibition test was carried out on Desmodesmus subspicatus with the test material according to OECD Guideline 201.

The stock solution 150.0 mg/L was prepared by dissolving white powder in OECD growth medium. Test solutions of required concentrations were prepared by mixing the stock solution of the test sample with OECD growth medium and inoculum culture and tested at the concentrations 0, 0, 30.0, 45.0, 67.5, 100.0 and 150.0 mg/L.Effects on the growth rate of the organism were studied. The test was performed under static conditions in a static fresh water system at a temperature of 23± 2°C. Initial cell density of test organism used was 5x10(3) cells/ml. Determination of cell counting involve the use of microscope with counting chamber Cyrus I or electronic particle counter. ErC50 was calculated using non-linear regression by the software Prism 4.0.

The median effective concentration (ErC50) for the test material, in a freshwater algae Desmodesmus subspicatus was determined to be 106.9 mg/L on the basis of effects on growth rate in a 72 hour study with 95% Cl of 83.2 - 137.3 mg/L.

Thus, based on the above summarised studies, 2-(N-ethylamino)-1-(4-methoxyphenil propane) and it’s structurally and functionally similar read across substance, it can be concluded that effect concetration value is in the range 106- 200 mg/l. Thus, comparing this value with the criteria of CLP regulation , 2-(N-ethylamino)-1-(4-methoxyphenil propane) cannot be classified for toxicity to aquatic algae and cynobaccteria .Hence,based on the data available for the structurally and functionally similar read across, test chemical 2-(N-ethylamino)-1-(4-methoxyphenil propane) is not likely to be toxic atleast in the concentration range of 106 - 200 mg/L