N1,N3-diallylpropane-1,3-diamine dihydrochloride

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

31 March 2020 to 29 June 2020

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Version / remarks:

23 March 2006

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

- Concentrations: 1.23 mg/L, 3.70 mg/L, 11.11 mg/L, 33.33 mg/L, 100 mg/L
- Sampling method: after 0h, 24h, 48h and 72h 200 µl test solution were transferred into a 96-well microplate

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION
- Method: stock solution was prepared by adding 111.1 mg test item to 1000 mL test medium and stirring for 5 min.:
- Controls: negative control

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

TEST ORGANISM
- Strain: 61.81 SAG
- Source (laboratory, culture collection): Culture Collection of Algae at the University of Goettingen
- Method of cultivation: twice a week the stock suspension is inoculated into fresh Holm-Hansen medium under axenic conditions to keep it in exponential growth.
ACCLIMATION
- Culturing media and conditions: Holm-Hansen medium for culturing, OECD 201 medium for testing
- Any deformed or abnormal cells observed: none by microscopy

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

21-24 °C

pH:

7.7-8.2

Nominal and measured concentrations:

Nominal concetrations: 1.23 mg/L, 3.70 mg/L, 11.11 mg/L, 33.33 mg/L, 100 mg/L
Measured concentrations are within 20% deviation from the nominal concentrations.

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer glass flasks 100 ml, Schott, Mainz
- Initial cells density: 7 x 10^4 cells/ml
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates):6


GROWTH MEDIUM
- Standard medium used: yes

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: 24h
- Light intensity and quality: 80.5 µE m/s +/- 9.8%


EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: cell counter
- Chlorophyll measurement: with fluorescence microplate reader (excitation at 465 nm and emission at 670 nm)

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 3
- Test concentrations: 1.23 mg/L, 3.70 mg/L, 11.11 mg/L, 33.33 mg/L, 100 mg/L
- Results used to determine the conditions for the definitive study: range finding study

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

33.66 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

6 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

3.7 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

10.38 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

3.79 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

3.7 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Details on results:

- Exponential growth in the control: yes
- Observation of abnormalities: no
- Any stimulation of growth found in any treatment: yes, up to 11.11 mg/L after 24h, up tp 3.70 mg/L after 48h and at 1.23 mg/L after 72h

Results with reference substance (positive control):

- Results with reference substance valid? yes (potassium dichromate)
- EC50: 1.19 mg/L

Reported statistics and error estimates:

ECx and LOEC/NOEC for yield and growth rate were determined with the statistical software ToxRat 3.3.0 (ToxRat Solutions GmbH, Alsdorf).

Table 1: Inhibition of yield after 24h, 48h and 72h exposure

Nominal test item concentration (mg/L)	Inhibition of yield (%)
	24h	48h	72h
NC	--	--	--
1.23	1.3	-17.9	-20.0
3.70	-30.7	-10.8	4.2
11.11	-6.9	15.7	50.9
33.33	38.5	83.4	91.6
100	56.8	89.6	97.2

Table 2: Inhibition of growth rate after 24h, 48h and 72h exposure

Nominal test item concentration (mg/L)	Inhibition of growth rate (%)
	24h	48h	72h
NC	--	--	--
1.23	3.1	-4.7	-3.6
3.70	-17.9	-3.4	1.0
11.11	-5.2	6.2	16.2
33.33	25.7	55.8	54.4
100	40.1	65.5	74.1

Table 3: Effect concentrations for yield and growth rate at the end of the test

(Lowest/No Observed) Effect Concentration	Nominal test item concentration (mg/L)
95 % Confidence Limits (CL)	Yield (72h)	Growth rate (72h)
EC50	10.38	33.66
CL	4.53 – 23.32	16.69-66.69
EC20	5.36	10.84
CL	2.75 – 10.50	3.3 – 10.90
EC10	3.79	6.00
CL	1.87 – 7.70	3.30-10.90
LOEC	33.33	11.11
NOEC	11.11	3.70

Validity criteria fulfilled:

yes

Conclusions:

N1,N3-diallylpropane-1,3-diamine dihydrochloride has inhibitory effects on the growth rate of freshwater algae above concentrations of 3.70 mg/L (=NOEC). The 72-hr ErC50 is 33.66 mg/L.

Executive summary:

In a 72 hour acute toxicity study, the cultures of Raphidocelis subcapitata strain 61.81 SAG were exposed to N1,N3-diallylpropane-1,3-diamine dihydrochloride at nominal concentrations of 0, 1.23, 3.70, 11.11, 33.33 and 100 mg/L under static conditions in accordance with the OECD 201 guideline (23 March 2006).  The NOEC and EC₅₀ values based on growth rate were 3.70 and 33.66 mg/L, respectively.  The % growth inhibition in the treated algal culture as compared to the control ranged from 1 to 74 .

 

Abnormalities were not observed.

 

This toxicity study is classified as acceptable and satisfies the guideline requirements for algae growth inhihition test toxicity study.

 

Results Synopsis

Test Organism: Raphidocelis subcapitata strain 61.81 SAG

Test Type: static

 

72 hr EC₁₀:  6.00 mg /L                       95% C.I.:  3.59 to 9.96 mg/L

72 hr EC₅₀:  33.66 mg/L                95% C.I.:  26.25 to 43.18 mg/L

72 hr NOEC:  3.70 mg/L                   

Endpoint(s) Effected:  growth rate

Description of key information

In a single key study on the acute toxicity of N1,N3-diallylpropane-1,3-diamine dihydrochloride to Raphidocelis subcapitata according to OECD guideline 202, an 72-hr EC₅₀ of 33.66 mg/L based on growth rate was reported. The test was conducted under static conditions and fulfilled all criteria for a valid test.

Key value for chemical safety assessment

EC50 for freshwater algae:

33.66 mg/L

EC10 or NOEC for freshwater algae:

3.7 mg/L

Additional information

Cultures of Raphidocelis subcapitata strain 61.81 SAG were exposed to N1,N3-diallylpropane-1,3-diamine dihydrochloride at nominal concentrations of 0, 1.23, 3.70, 11.11, 33.33 and 100 mg/L under static conditions in accordance with the OECD 201 guideline (23 March 2006).  The NOEC and EC₅₀ values based on growth rate were 3.70 and 33.66 mg/L, respectively. The % growth inhibition in the treated algal culture as compared to the control ranged from 1 to 74.

Abnormalities were not observed.