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Long-term toxicity to aquatic invertebrates

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Description of key information

Freshwater: The 21d-NOEC for Daphnia magna (reproduction) using semi-static exposure according to OECD TG 202, part II (update June 1993) was 0.111 mg/l based on measured concentrations

Marine water: The 6d-EC10 for Acartia tonsa (larval development), static exposure, according to the draft OECDTG Life cycle test with Acartia tonsa (2004): 0.044 mg/l based on nominal concentrations of labelled HHCB because measured concentrations were in line with nominal)

Key value for chemical safety assessment

Fresh water invertebrates

Fresh water invertebrates
Dose descriptor:
Effect concentration:
0.111 mg/L

Marine water invertebrates

Marine water invertebrates
Dose descriptor:
Effect concentration:
0.044 mg/L

Additional information

Two long-term aquatic toxicity tests are available, one for freshwater and one for marine water. An additional study by Breitholtz (2003) is addressed below, however it was considered unsuitable for use (EU RAR).

Key study fresh water: Wüthrich (1996; RCC 380687), Daphnia magna

For Daphnia magna, a GLP semi-static 21-d toxicity test was carried out similar to OECD TG 211 (but mentioned as OECD TG 202). The test medium was refreshed three times per week. Nominal concentrations ranged from 0.062 to 1.0 mg/l (step size factor 2). Concentrations were measured at the start and end of the first and last exposure period. Results: The validity criteria were met. EC50 immobility was 0.293 mg/l (at 0.205 and 0.419 mg/l no and 100% immobility, respectively was seen). The NOEC for reproduction was 0.111 mg/l. Some decrease of 21% was seen at this concentration but it was not statistically significant. At 0.205 mg/l 26% significant decrease in reproduction was seen, according to the Dunnett test (P0.05). The EC50 of the reproductive output after 21 days was determined as 0.282 mg/l. 

Key study marine water: Bjornestad (2007; DHI 91328/700), Acartia tonsa: The chronic toxicity to marine copepod Acartia tonsa was tested according to OECD Draft Guideline for Testing of Chemicals (2004) describing a life cycle test, with specific adaptations to prevent volatilisation and accumulation of organic debris. The test was carried out under GLP and met the quality criteria. For this test the substance was mixed with 0.1% of radio-labelled [3-14C]-HHCB with a radiochemical purity of 94.8%. Ethanol was used as a solvent. Test concentrations were 37.5, 75, 150, 300 and 600 µg/l, with four replicates per concentration and 6 replicates in the control as well as in the solvent control. Natural seawater, salinity 30.2‰ was used. The test was carried out at 20 ± 1.0°C in a climate room with a daily light/dark period of 16:8 hours. After 5.5 days the number of nauplii, copepodites and non-hatched eggs were counted and the lengths of the larvae was measured. Results: The actual test concentrations were >80% of nominal during the whole study, hence nominal concentrations were used for the calculations. The larval development ratio (LDR) was 73% in the control and 72% in the solvent control. The NOEC (LDR) was 37.5 μg/l with an inhibition of 7.5% as compared to the solvent control. The inhibition was 27% in 75 μg/l (LOEC). At 300 μg/l, the length of the nauplii was significantly smaller than in the controls. Moreover, no copepodites were observed. Up to 150 μg/l, there was no impact on growth. The EC10 (LDR) was 43.8 μg/l (95% confidence interval 30.1 – 55.3 μg/l) and the EC50 was 131 μg/l (95% CI: 115-153 μg/l). 


Non-relevant study: The study below was not used since the test conditions did not ensure a 'water-only' exposure and therefore the results of this test could not be expressed on the basis of the concentration in water (EU RAR, 2008).


: Breitholtz et al. (2003), Marine water, harpactoid copepod, Aquatic Toxicology 63:103-118: A population growth experiment was carried out with the marine harpactoid copepod Nitocra spinipes. After short-term exposure (96 h) the LC50 was 1.9 mg/l (1.40-2.70). To investigate the chronic toxicity, eight replicates of 8-10 nauplii per concentration were tested in 10 ml test medium in concentrations ranging from 0.002 to 0.2 mg substance/l. The test was carried out at 20°C in artificial seawater (low salinity: 6‰). The test included the period between hatching of the F0 to hatching of F1-generation. The larval development ratio was established after 7 to 8.5 days and the exposure for the life-cycle experiment was terminated at day 22, although for ovigerous females the exposure was continued until day 26. Concentration measurements showed that the initial concentrations were variable and that concentrations were not maintained at all after the 2 days interval before renewal. This was most likely attributed by a combination of volatilisation and sorption to organic residues which therefore caused a lower exposure concentration in the water phase and additional exposure by oral uptake.

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