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EC number: 226-939-8 | CAS number: 5567-15-7
Mortality of Adult Earthworms in [%] after 28 Days of Exposure
[mg test item/kg soil dry weight]
Earthworm Behaviour and Pathological Symptoms after 28 Days of Exposure
A)= No observation of symptoms or behavioural changes; H)= dead earthworm
Body Weight Changes of the Adult Earthworms
Application rate[mg test item/ kg soil dry weight]
Mean body weights per replicate
Mean body weight change of earthworms
Mean ± SD
0.23 ± 0.04
47.8 ± 7.81
0.25 ± 0.04
53.0 ± 8.34
0.27 ± 0.04
56.9 ± 9.15
Rep.= replicate; Sign. = significance compared to pooled control;
Reproduction Rate (Number of Juveniles after 8 Weeks)
Number of juveniles
of pooled control
Significance compared to pooled control
304 ± 41.9
291 ± 44.9
329 ± 32.5
Effects of the test item on mortality, biomass and the reproductive potential of the earthworm species Eisenia fetida(Savigny) were determined according to the guidelines DIN ISO 11268-2 (1998), OECD 222 (2004) and BBA (1994) from November 19th, 2009 to January 16th, 2010 at Dr.U.Noack-Laboratorienin 31157 Sarstedt, Germany. The study was conducted under static conditions over 8 weeks with a limit test item concentration of 1000 mg test item/kg soil dry weight, which was mixed into artificial soil containing 10 % peat. A control using untreated artificial soil and a solvent control with the solvent mixed into the artificial soil were set up. 80 test organisms, with an individual weight of 0.30 to 0.60 g, were divided into 8 replicate groups with 10 test organisms per replicate.
After 28 days no evident mortality as well as no pathological symptoms or changes in the behaviour of adult earthworms were seen in the control as well as at the limit test item concentration of 1000 mg test item/kg soil dry weight (Table 1). The earthworm biomass increased in the controls and at the limit test item concentration of 1000 mg test item/kg soil dry weight over 28 days without any statistically significant differences between pooled control and limit test item concentration.
After further four weeks the reproduction rate (average number of juveniles) was 297 in the pooled control and 329 at the limit test item concentration. Compared to the pooled control, the reproduction rate was not statistically significantly reduced at 1000 mg test item/kg soil dry weight (Table 1).
Overall, the NOEC of the test item concerning mortality, biomass and reproduction of earthworms was determined to be at 1000 mg test item/kg soil dry weight (Table 1).
All validity criteria recommended by the test guidelines were fulfilled.
Summary of All Observed Effects in the Tested Dose
Mortality of adult earthworms
Reduction of body weight
Reduction of reproduction rate
(Number of juveniles)
Summary of all effects
NOEC and LOEC for Reproduction, Mortality and Body Weight
mg test item/kg soil dry weight
NOECreproduction, mortality, body weight
LOECreproduction, mortality, body weight
Under the conditions of this study, the test item did not induce evident mortality or pathological symptoms of adult earthworms after an exposure to the test item concentration of 1000 mg test item/kg soil dry weight for 28 days. Also, the biomass increase of the adult earthworms after 28 days was not statistically significantly reduced compared to the pooled control at the test item concentration of 1000 mg test item/kg soil dry weight. After eight weeks of exposure the earthworm reproduction was not statistically significantly reduced at the limit test item concentration of 1000 mg test item/kg soil dry weight compared to the pooled control.
Thus, the No Observed Effect Concentration (NOEC) with respect to mortality, body weight and reproduction of Eisenia fetida fetida after 8 weeks of exposure in artificial soil was determined to be at 1000 mg test item/ kg soil dry weight.
The results from the two long term toxicity tests on Eisenia fetida with Pigments Yellow 83 and its structural analogue, Pigment Yellow 12, conclusively show absence of toxicity to earth worms, neither in regard to reproduction, nor in regard to mortality and body weight.
As long term toxicity tests towards soil macro-organisms except arthropods (earth worm Eisenia fetida) for two different pigments of the Diarylide Yellow group had been conducted with test substance concentrations far above the solubility limit in water, from acute tests no additional information could be expected. Therefore, no acute toxicity tests were performed.
Judging from these long-term toxicity tests with two different pigments, the Diarylide Yellow Pigments are to be regarded as nontoxic to soil macroorganisms.
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