1,3-Diazaspiro[4.5]decane-2,4-dione, 8-methoxy-, sodium salt (1:1), cis-

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2007

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2007

Report date:

2007

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

sodium (5s,8s)-8-methoxy-2,4-dioxo-1,3-diazaspiro[4.5]decan-3-ide

EC Number:

929-513-6

Cas Number:

1400584-86-2

Molecular formula:

C9H13N2O3 · Na

IUPAC Name:

sodium (5s,8s)-8-methoxy-2,4-dioxo-1,3-diazaspiro[4.5]decan-3-ide

Test material form:

solid: bulk

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

49.0 mg of the test item were dissolved in Milliporewater and made up with Milliporewater to the mark in a 100 mL volumetric flask to prepare a stock solution of 490 mg/L. Defined volumes of this stock solution were diluted with Milliporewater to obtain standard solutions in the range of 0.025 to 4.90 mg/L.

Sample storage conditions before analysis:
Routinely, the samples are analysed immediately. Only in exceptional cases, they are stored overnight deeply frozen and protected from light.

Test solutions

Vehicle:

no

Details on test solutions:

To produce the only test concentration 125.0 mg test item were added to 1 litre of dilution water. The pH was adjusted to pH 7.9.

Test organisms

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

Common name: green algae
Source: Non-axenic strain of the test species obtained from "The Collection of Algal Cultures" of the Institute of Plant Physiology at the University of Gottingen (Germany).

Maintenance of stock cultures:
Exponentially-growing stock cultures are maintained in the test facility under constant temperature conditions (23 +/- 2°C) at a light intensity in the range 60 - 120 µE x m-2 x s-1 (measured in the range 400 to 700 nm using a spherical quantum flux meter). The nutrient medium (according to BRINGMANN & KUHN (1977) is renewed once a week. Cell density measurements are made using a microcell counter.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Test conditions

Hardness:

Water hardness of the final nutrient medium is 1.3°dH, corresponding to 22.5 mg/L CaCO3.

Test temperature:

21 - 25 °C

pH:

8.1 - 9.6

Nominal and measured concentrations:

nominal: 100 mg/L
meassured: 99.5 mg/L (0 h), 99.9 mg/L (48 h)

Details on test conditions:

Exposure conditions
Test vessels: 300 mL Erlenmeyer flasks with cotton stoppers

Culturing apparatus:
Light chamber in which a temperature in the range 2 °C to 25 °C can be maintained at +/- 2 °C, and continuous uniform illumination is provided in the spectral range 400 to 700 nm.

Light intensity:
At the average of the test solutions, a light intensity in the range 60 to 120 µE x m-2 x s-1, or in equivalent range of 4000 to 8000 lx, is recommended to use.

Cell density measurements:
Cell densities are measured in a microcell counter or, alternatively, are determined by means of a microscopic counting chamber.

Experimental design:
1 test concentration plus 1 control
3 replicates per concentration, 6 replicates per control
initial cell density in the test cultures approximately 5000 cells per millilitre

Results and discussion

Effect concentrations

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

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

No toxic effects against algae at a limit test concentration of 100 mg/L.

Executive summary:

A study was performed to assess adverse effects on the growth (= increase in cell density) and the growth rate (= rate of increase in cell density with time) of the planktonic freshwater algal species Desmodesmus subspicatus over several generations.

The study was conducted in accordance with EEC Methods for Determination of Ecotoxicity Annex to Directive 92/69/EEC (OJ. No. L383A, 29.12.92) Part C, Method 3 'Algal inhibition test' which is in most parts equivalent to the OECD Guideline for Testing of Chemicals No. 201 'Alga, Growth Inhibition Test'.

Exponentially growing algal cells were exposed for a period of 72 hours to a limit test concentration of nominally 100 mg/L dissolved in water.

The cell densities were measured at 24-hour intervals. Inhibition of the algal population was measured as reduction in growth (index b) and growth rate (index r), relative to control cultures grown under identical conditions. The following values were determined:

Results	[mg/L]
ErC 50 (0-72 h)	> 100
EbC 50 (0-72 h)	> 100

No toxic effects against algae at a limit test concentration of 100 mg/L.

All results are expressed in terms of nominal concentrations. Recovery rates correspond to 99.5 % of nominal values at 0 hours, and to 99.9 % of nominal values at 48 hours, respectively.