Registration Dossier

Ecotoxicological information

Short-term toxicity to aquatic invertebrates

Currently viewing:

Administrative data

Link to relevant study record(s)

Reference
Endpoint:
short-term toxicity to aquatic invertebrates
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
comparable to guideline study with acceptable restrictions
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)
Version / remarks:
OECD Guideline for Testing of Chemicals, No. 202, Daphnia sp., Acute Immobilization Test and Reproduction Test, Part 1,1984
Deviations:
not specified
Qualifier:
equivalent or similar to
Guideline:
EU Method C.2 (Acute Toxicity for Daphnia)
Version / remarks:
EU Commission Directive 92/69/EEC, C.2, Acute Toxicity for Daphnia, 1992
Deviations:
not specified
GLP compliance:
not specified
Remarks:
no information on GLP compliance available in this publication
Specific details on test material used for the study:
Diameter (nm) - (no information)
Length (µm) - (no information)
Lateral size (µm) 0.5 – 2.0
Purity (%) 99
Company where purchased Nanjing XFNANO Materials Tech Co., Ltd.
Analytical monitoring:
yes
Vehicle:
yes
Details on test solutions:
The graphene (GN) suspension was prepared using a simplified SM7 medium, a mixture solution of CaCl2 (293.8 mg /L), MgSO4 (123.3 mg/L), K2HPO4 [0.184 mg/L), NaNO3 (0.274 mg/L), NaHCO3 (64.8 mg/L), Na2SiO3 (10 mg/L), H3BO3 (0.175 mg/L), KCl (5.8 mg/L), and KH2PO4 (0.143 mg/L), with a water hardness of 250 mg CaCO3 per L and pH of 7.8 - 8.2 as recommended in the Organization for Economic Cooperation and Development (OECD) Guideline 211.
PVP was selected as a non-covalent dispersant. In the preliminary experiment, it was shown that PVP at a concentration as high as 10 mg/L was not toxic and did not affect the growth and reproduction of 24 h old daphnia compared with controls after 21 days of chronic exposure.
The GN stock suspension was obtained by adding 10 mg of GN to 100 mL of a 1 g/L PVP solution (PVP dissolved in SM7 medium) followed by sonication using a sonication bath (ultrasonic bath KQ-500DB, Kun Shan, China) for 2 h at room temperature to obtain optimal particle dispersion. Most of the GN were suspended and no precipitate was observed after sonication. The nominal concentration (100 mg/L) was considered as the concentration of the GN stock suspension, although it was difficult to reach the full extent of exfoliation by sonication. The GN suspension with different concentrations used for acute and chronic tests was prepared by diluting the GN stock suspension followed by sonication for 20 min before exposure.
Test organisms (species):
Daphnia magna
Details on test organisms:
TEST ORGANISM
- Common name: Daphnia magna, neonates less than 24 h old cultured in freshwater under normal environmental conditions under a 16:8 light-dark cycle at 23.5 °C. The freshwater sample was obtained from the Huo Qi Ying Bridge area, China (latitude 39°58'5.58", longitude 116° 16'53.3"), filtered through glass fiber filters before use, and renewed every 2 d.
- Feeding during test: not specified
- Food type: green alga Chlamydomonas reinhardtii at a cell density of 5 x 10,000 cells per mL (for daphnids <3 days old) and 100,000 cells per mL (for daphnids >3 days old) every day.

To check whether the sensitivity of the daphnia was within the limits (EC50, 24 h = 0.5 - 1.2 mg/L) as specified in the GB/T 13266 guideline (Water quality-determination of the acute toxicity of substance to Daphnia issued by China's health ministry) and reduce the risk of experimental failure, the 24 h acute toxicity test of the daphnids was performed every month using the reference toxicant K2Cr2O7.
Test type:
static
Water media type:
freshwater
Limit test:
no
Total exposure duration:
48 h
Post exposure observation period:
none
Hardness:
no details given
Test temperature:
no details given
pH:
no details given
Dissolved oxygen:
no details given
Salinity:
no details given
Conductivity:
no details given
Nominal and measured concentrations:
0.2 - 16 mg/L (nominal)
Details on test conditions:
TEST SYSTEM
- Test vessel: beaker
- No. of organisms per vessel: 10
- No. of vessels per concentration (replicates): 4

OTHER TEST CONDITIONS
- Photoperiod: 16-hour light to 8-hour darkness photoperiod (with a 30 minute transition period).

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
The mortality of the daphnids were observed after 48 hours of exposure to the test material

VEHICLE CONTROL PERFORMED: yes
Reference substance (positive control):
no
Key result
Duration:
48 h
Dose descriptor:
LC50
Effect conc.:
> 16 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: mortality = 40% derived from graph
Duration:
48 h
Dose descriptor:
other: LC20
Effect conc.:
> 4 mg/L
Nominal / measured:
nominal
Conc. based on:
test mat.
Basis for effect:
mortality
Remarks on result:
other: derived from graph
Details on results:
- Behavioural abnormalities: none stated
- Mortality of control: none
- Other adverse effects control: none stated
- Abnormal responses: none stated
- Any observations (e.g. precipitation) that might cause a difference between measured and nominal values: none stated
Reported statistics and error estimates:
All data were represented as mean values +/- standard deviation. All statistics were performed using SPSS Statistics 19.0 for Windows. Values were considered to be significant if p < 0.05.
Validity criteria fulfilled:
not specified
Conclusions:
The LC 50 was determined as > 16 mg/l (derived from graph).
Executive summary:

The acute toxicity of graphene (GN) to Daphnia magna was determined in a 48-hour static test similar to the EU Commission Directive 92/69/EEC, Part C.2 (1992), and the OECD Guideline for Testing of Chemicals, No. 202, Part I (1984). This study is classified as reliable with restrictions (Klimisch 2) and satisfies the guideline requirements for an acute toxicity study with freshwater invertebrates.

Application of up to 16 mg/L graphene induced <50% daphnid mortality, therefore the LC 50 was determined as >16 mg/L.

Description of key information

The acute toxicity of the test item graphene towards aquatic invertebrates was assessed in a study conducted by Fan et al., 2016. In this study, the acute toxicity of graphene to Daphnia magna was determined in a 48-hour static test similar to the EU Commission Directive 92/69/EEC, Part C.2 (1992), and the OECD TG 202.

Application of up to 16 mg/L graphene induced <50% daphnid mortality, therefore the LC50 was determined as >16 mg/L.

Key value for chemical safety assessment

EC50/LC50 for freshwater invertebrates:
16 mg/L

Additional information

The results reported in the key study are supported by a study conducted by Guo et al., 2013. Here, no Daphnia were immobilized after being exposed for 48 h to a graphene concentration of 250 µg/L, the highest graphene concentration in this study. Thus, this study underlines the absence of acute toxicity of graphene to Daphnia magna at low concentrations.