O-phosphoserine

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:

from 2017-08-21 to 2017-09-13

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)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.5400 (Algal Toxicity, Tiers I and II) (January 2012)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

TOXI-COOP ZRT., 8230 Balatonfüred, Arácsi út 97., Hungary

Analytical monitoring:

yes

Details on sampling:

For determination of the test item concentrations, six replicate samples (5 mL per replicate) were taken from the test concentration and from the control at the start and at the end of the test. The samples were prepared immediately after sampling. Formulation samples were diluted with Ultrapure Water and analysed after derivatization.

Vehicle:

no

Details on test solutions:

The test solutions used in the test was prepared by mechanical dispersion without using any solubilising agents. A stock solution was first prepared by dissolving an amount of 0.08 g test item in 800 mL dilution water (OECD Medium) to obtain the nominal concentration of 100 mg/L. pH of this test solution was adjusted to that of the culture medium using 1N NaOH.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: green algae
- Strain: 61.81 SAG
- Source: Collection of Algal Cultures, Inst. Plant Physiology, University of Göttingen, Untere Karspüle 2, D-37073 Göttingen, Germany
- Age of inoculum (at test initiation): 4 days

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Test temperature:

22.7 - 23.2 °C

pH:

7.96 - 9.37

Salinity:

NA

Nominal and measured concentrations:

Nominal concentrations: 0 and 100 mg/L
Measured concentrations: in the range of 101 - 103 % of the nominal concentration

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer flasks
- Material, size, headspace, fill volume: glass, ~250 mL volume, 100 mL test medium
- Initial cells density: 1 x 10^4 cells/mL
- Control end cells density: 126.50 x 10^4 cells/mL
- No. of organisms per vessel: 0.1 mL algal biomass (10^7 algal cells per mL)
- No. of vessels per concentration (replicates): 6
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: OECD Medium prepared according to OECD 201

OTHER TEST CONDITIONS
- Adjustment of pH: yes, using 1N NaOH
- Photoperiod: continuously illuminated
- Light intensity and quality: 8038 lux

TEST CONCENTRATIONS
- Range finding study; yes
- Test concentrations: 0.1, 1, 10, 100 mg/L

EFFECT PARAMETERS MEASURED :
- Determination of cell concentrations: microscope with counting chambe at 24, 48 and 72 hours

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks:

and yield

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks:

and yield

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks:

and yield

Details on results:

- Abnormal appearance of the algal cells was not observed during the experiment.
- The growth inhibition calculated for the 72 hours exposure in treatment group compared to the control was 0.7 % for the average specific growth rate and 2.7 % for the yield. The results of the statistical evaluation showed that the 0-72 h average specific growth rate and yield at the treatment group (100 mg/L nominal concentration) was not statistically significantly different from the untreated control .

Results with reference substance (positive control):

The date of the last study with the reference item Potassium dichromate was: 07 – 10 March 2017. The following results were obtained:
The 72h ErC50: 0.99 mg/L, (95 % confidence limits: 0.68 – 1.49 mg/L)
The 72h EyC50: 0.59 mg/L, (95 % confidence limits: 0.47 – 0.75 mg/L)

Pre-experiments

In the 1st preliminary test up to 10 mg/L no effect was observed. At concentration level 100 mg/L 100% mortality was observed. Since toxicity was observed in the first preliminary test at the 100 mg/L concentration, a second preliminary test was performed using pH adjustment to determine the approximate toxicity of the test item independent of the pH. After adjustment of the pH of the stock solution no effect was observed also at the highest test concentration. Therefore, adjustment of the pH was performed also in the main test.

Validity criteria

- Cell density in the control cultures increased by a factor of more than 16 (by a factor of 126.50) within 72 hours. This corresponds to a specific growth rate of 1.61 day^-1.

- The mean coefficient of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3, for 72 h-tests) in the control cultures did not exceed 35 %.

- CV for section-by-section growth rate day 0-1: 12.68 %

- CV for section-by-section growth rate day 1-2: 9.93 %

- CV for section-by-section growth rate day 2-3: 6.94 %

The mean coefficient of variation for section-by-section specific growth rates: 9.85 %.

- The coefficient of variation of average specific growth rates during the whole test period in replicate control cultures did not exceed 7 % in test.

- CV for average specific growth rate day 0-3: 3.39 %

All validity criteria were met, therefore, the study is considered as valid.

Analytical results

Six samples were analysed from the test concentration with two parallel injections of each and six samples were analysed from the control solutions. The measured concentration of the test item was 103 % of the nominal at the start and 101 % at the end of the test.

Table 1. Test item concentrations

Sampling date	Nominal concentration (mg Test Item/L)	Mean of the measured concentrations (mg Test Item/L)	Measured concentration in percentage of the nominal	Relative Standard Deviation % (n=6)
22 August 2017	100	103.3	103	0
25 August 2017	100	101.0	101	1

Biological endpoints

Table 2. Growth Rates (r) and percentage inhibition of r during the test period

Test Group	Growth Rates (r)and % Inhibition ofr
	0–24 h		0–48 h		0–72 h
	r	%	r	%	r	%
Control	0.0518	–	0.0629	–	0.0671	–
100 mg/L(nominal)	0.0593	-14.6	0.0628	0.2	0.0666	0.7

Table 3. Yield (y) and percentage inhibition of y during the test period

Test Group	Yield y and % Inhibition of y
	0–72 h
	y	%
Control	125.5	–
100 mg/L(nominal)	122.2	2.7

Table 4. The section-by-section specific growth rates in the control cultures

Section-by-Section Growth Rate
Expressed in hours	0–24 h	24-48 h	48-72 h
	0.0518	0.0740	0.0755
Expressed in days	0-1 day	1-2 day	2-3 day
	1.2425	1.7755	1.8110

Validity criteria fulfilled:

yes

Conclusions:

In a 72-h algal growth inhibition test the EC50 based on growth rate was determined to be above 100 mg/L, the EC10 was >100 mg/L and the NOEC was determined to be 100 mg/L, respectively.

Executive summary:

The effect of the test item on the growth of a unicellular green algal species Raphidocelis subcapitata was determined according to OECD 201, Regulation (EC) No. 226/2016 Method C.3 and EPA OPPTS 850.5400 and under GLP. The study was conducted under static conditions with an initial cell density of 1x10^4 cells/mL for a period of 72 hours. A limit test with one test concentration - 100 mg/L, and control was performed. The measured concentrations of the test item at the start and at the end of the test were in the range of 101 - 103 % of the nominal concentration. The measured concentrations deviated not more than 20 % from the nominal concentration, therefore, the nominal concentration was used to determine exposure concentrations.The EC50-values for inhibition of growth rate (ErC50) and for inhibition of yield (EyC50) were >100 mg/L. The ErC10 and EyC10 values were determined to be >100 mg/L. The NOEC was determined to be 100 mg/L.

Description of key information

In a 72-h algal growth inhibition test the EC50 based on growth rate was determined to be above 100 mg/L, the EC10 was >100 mg/L and the NOEC was determined to be 100 mg/L, respectively.

Key value for chemical safety assessment

EC10 or NOEC for freshwater algae:

100 mg/L

Additional information

The effect of the test item on the growth of a unicellular green algal species Raphidocelis subcapitata was determined according to OECD 201, Regulation (EC) No. 226/2016 Method C.3 and EPA OPPTS 850.5400 and under GLP (reference 6.1.5-1). The study was conducted under static conditions with an initial cell density of 1x10^4 cells/mL for a period of 72 hours. A limit test with one test concentration - 100 mg/L, and control was performed. The measured concentrations of the test item at the start and at the end of the test were in the range of 101 - 103 % of the nominal concentration. The measured concentrations deviated not more than 20 % from the nominal concentration, therefore, the nominal concentration was used to determine exposure concentrations.The EC50-values for inhibition of growth rate (ErC50) and for inhibition of yield (EyC50) were >100 mg/L. The ErC10 and EyC10 values were determined to be >100 mg/L. The NOEC was determined to be 100 mg/L.