Perhydroimidazo[4,5-d]imidazole-2,5-dione

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:

2002

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Test substance: Tetrahydro-imidazo-(4,5.d-)-imidazol-2,5-dion.
Batch number: Fixapretl4o Nr. 766.
Purity: 96.3g/100g; C=33.5, H4.1, N38.9, O~23.0 g/100g.
Impurities: possible rest of urea and sulfuric acid (according to the sponsor), Glyoxal<0,1%.

Analytical monitoring:

yes

Details on sampling:

Samples were directly used for HPLC-analysis.

Vehicle:

yes

Details on test solutions:

The test medium was prepared according to guidelines under OECD-Medium part 9 (single concentrated).

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

Test organism
Strain: Desmodesmus subspicatus CHODAT SAG 86.81.
Source: SAG (collection of algal cultures in Göttingen).
Pre-culture techniques: Seed-culture; seed culture incubated for 7 days at 23°C ± 2°C. The final cell density was 391x10 4 cells/mL. Pre-culture: seed culture was taken to inoculate a pre-culture (initial cell density; 1x10 4 cells/mL). The pre-culture was incubated for 3 days at 23°C ± 2°C. Final cell density: 26x10 4 cells/mL.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Remarks on exposure duration:

Exposure was carried out in Erlenmeyer flasks (nominal volume 250mL) plugged with gas permeable silicon sponge caps.

Post exposure observation period:

Observations were taken 0, 24, 48 and 72 hours after exposure to the test substance.

Hardness:

Not specified.

Test temperature:

23°C ± 2°C; measured continously during the whole test period in a climate chamber.

pH:

8; measured after 0 and 72hrs in an additional uninoculated replicate and after 72hrs in the inocculated replicate No.1 of each concentration.

Dissolved oxygen:

Not specified.

Salinity:

Not specified.

Conductivity:

Not specified.

Nominal and measured concentrations:

The stock solution was diluted to obtain the following nominal concentrations (mg/L): 3.13, 6.25, 12.5, 25, 50, 100.
A negative control (0mg/L) were no test substance was present was also tested.

Details on test conditions:

Test temperature: 23°C ± 2°C.
Test vessel: Exposure was carried out in Erlenmeyer flasks (nominal volume 250mL) plugged with gas permeable silicon sponge caps.
Inoculation density: 1x10 4
Number of replicates per concentration: 3.
Number of replicates per control: 3.
Illumination: artificial light, type universal white (e.g OSRAM L 25), permanent illumination.
Light intensity: about 60 - 120µEI(m2s) at a wave length of 400 - 700 nm.
Test parameter: in vivo chlorophyll-a-fluorescence (pulsed excitation with light flashes having a wavelength of 435 nm).
Cell counting: after 72 h in a counting chamber (Neubauer improved) in replicate No. 2 of the inoculated control.

Reference substance (positive control):

not required

Key result

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Key result

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Duration:

72 h

Effect conc.:

ca. 0 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 0

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 3.13 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 0.3

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 6.25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 2.6

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 12.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: -0.9

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 1.1

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 1.4

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 3.6

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 0 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 0

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 3.13 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 1

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 6.25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 1.5

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 12.5 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 0.2

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 25 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 1.2

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 50 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 1.5

Remarks:

Inhibition (% of the control)

Duration:

72 h

Effect conc.:

ca. 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 0.4

Remarks:

Inhibition (% of the control)

Details on results:

The results show that there were some slight variation in the values for inhibition in relation to the percentage of the control for both the growth rate and biomass.

Reported statistics and error estimates:

Determination of the mean fluorescence after 0, 24, 48, 72 h.
Calculation of the integral of biomass growth over the total duration of the test for each concentration level and comparison of values of treated samples in relation to untreated samples. Calculation of the growth rate over the total duration of the study for each concentration level and comparison of values of treated samples in relation to untreated samples. Due to the results of the test the EC values are not calculated (linear regression analysis) from the concentration-
response relationship.

TEST RESULT

 

Effect on the development of biomass:

              E_bC₁₀(72 h)        >            100 mg/L

              E_bC5₀(72 h)       >            100 mg/L

              E_bC₉₀(72 h)        >            100 mg/L

 

Effect on growth rate:

              E_bC₁₀(72 h)        >            100 mg/L

              E_bC5₀(72 h)       >            100 mg/L

              E_bC₉₀(72 h)        >            100 mg/L

 

No Observed effect concentration

              NOEC (72 h)      ≥            100 mg/L

 

 

Lowest Observed Effect Concentration

              LOEC (72 h)       >            100 mg/L

Validity criteria fulfilled:

yes

Conclusions:

The results show that there were some slight variation in the values for inhibition in relation to the percentage of the control for both the growth rate and biomass however overall there was no effect on the growth rate and biomass.

Executive summary:

A study was performed to determine the acute toxicity of the test substance to the algae Desmodesmus subspicatus Chodat Sag 86.81 in a 72 hour static test according to; EEC Directive 92/69/EEC, Annex V, Part C.3 (December 1992). Algal inhibition test, OECD 201 (June 1984), and EPA OPPTS 850.5400 (April 1996). The test substance was tested in the concentration range between 100 and 3,13 mg/L. The dilution factor was 2. The analytical verifications of the test substance were investigated at different concentrations in OECD medium. The analytical results yielded 80% or higher recoveries. These varied between 95.1 and 104.8% of the nominal concentrations at test initiation, and between 95.3 to 101.9% at test termination. Therefore, all biological results are related to the nominal concentrations of the test item. Since the results yielded >80% recovery rates, it can be concluded that based on the experimental conditions the test substance is non-toxic to algae under EC 1272/2008 and the EC50 value is >100mg/L based on analytically confirmed nominal concentrations.

Description of key information

A study was performed to assess the acute toxicity of the test substance to the algae Desmodesmus subspicatus Chodat Sag 86.81 in a 72 hour static test according to; EEC Directive 92/69/EEC, Annex V, Part C.3 (December 1992). Algal inhibition test, OECD 201 (June 1984), and EPA OPPTS 850.5400 (April 1996). The test substance was tested in the following concentrations; 3.13, 6.25, 12.5, 25, 50, 100mg/L. Two parameters were examined for the effect of the test substance; the development of biomass (Eb50) and growth rate (Eg50). Test results show that the Eb50 and Eg50 was >100mg/L. The calculated no observed effect and lowest observed effect concentration levels after 72 hours was >100mg/L.

Based on the conditions and the results of the study, it can be concluded that the test substance is non-toxic to algae under EC 1272/2008, since the effect concentration value (EC50) was >100mg/L.

Key value for chemical safety assessment

EC10 or NOEC for freshwater algae:

100 mg/L