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Sediment toxicity

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sediment toxicity: short-term
Type of information:
experimental study
Adequacy of study:
other information
Study period:
19.01.1996 - 09.02.1996
4 (not assignable)
Rationale for reliability incl. deficiencies:
other: Study in accordance with national standard methods, but only aquatic compartment was spiked. Hence, the study cannot be used for assessment of the sediment compartment

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
other: Preliminary BBA- guideline : "Effects of plant protection products on the development of sediment-dwelling larvae of Chironomus ripuarius in a water-sediment system"
Version / remarks:
as of 1995
some minor modifications regarding the applicability towards general chemicals were taken into account
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
2-phosphonobutane-1,2,4-tricarboxylic acid
EC Number:
EC Name:
2-phosphonobutane-1,2,4-tricarboxylic acid
Cas Number:
Molecular formula:
2-phosphonobutane-1,2,4-tricarboxylic acid
Test material form:
Details on test material:
Water solubility: completely soluble

Sampling and analysis

Analytical monitoring:
Details on sampling:
Sampling was performed immediately at the beginning of the study, once a week during the study and at the end of the study. Analytical samples were determined via ionchromatography.

Test substrate

Details on sediment and application:
Sediment is added to the test vessel up to 2 cm and has a wet weight of 540 g. The test matrix is M4 medium and the height of the liquid phase is 16 cm.

The test substance is used in the appropriate amount for each test batch. The amount substance is weighed, dissolved in 10 ml test medium (from the corresponding test vessel previously removed) and added using a measuring pipette applied approx. 2 cm below the water surface. After that the test medium is carefully stirred, without stirring up the bottom sediment, using a glass rod.

Test organisms

Test organisms (species):
Chironomus riparius
Details on test organisms:
- Common name: Chironomus riparius L.
- Strain/clone: Laborstamm Bayer AG, PF-E, Ökonbiologie
- Details on collection: Breeding of the larvae takes place in developer dishes, with aereated M4-medium (according to Elendt) and diatomaceous earth as substrate. The colony is fed using Tetramin pet food and Tetraphyll (1:1) as food substrate (ad libitum). Emergence, nuptial flight and oviposition of the imagines take place in a gauze-covered flight cage. The colony is held at 20±2 °C, with a photoperiod of 16h brightness (light intensity~1000 lux) and 8 h darkness.

- Age of animals at beginning of exposure: newly hatched
- Feeding during test: 3 times a week (mondays, wednesdays, and fridays) 1mg food susbtrate / larva/day,i.e. 7 mg per larva a week, amounting to 175 mg per test vessel and week. From week 2 on, the amount of food substrate is increased by 50 %.
- Food type: Tetramin pet food and Tetraphyll (1:1) as food substrate

Study design

Study type:
laboratory study
Test type:
Water media type:
Type of sediment:
artificial sediment
10 % peat 20 % kieselguhr 70 % sand, industrial grade maximum WHC = 41.7% (referenced to dry weight)
Limit test:
Exposure duration
21 d
Exposure phase:
total exposure duration
Post exposure observation period:
21 d (28d planned, test was finished after 21 d as no effect was observed)

Test conditions

2.6 mmol/L
Test temperature:
19.6 °C - 22.2 °C
Dissolved oxygen:
8.9 mg/L
not applicable
not applicable
700 mSv/cm
Nominal and measured concentrations:
nominal concentrations: 100 mg/L, 50 mg/L, 25 mg/L, and 12.5 mg/L measured concentrations (arithmetic mean of initial concentrations): 96.3 mg/L, 47.8 mg/L, 19.0 mg/L, 11.1 mg/L
Details on test conditions:
Glass vessels with a height of 24 cm and a diameter of 13.5 cm were used. In each vessel the artificial medium was filled up to about 2 cm height. then 2.3 L testmedium (aereated) were applied. Into each vessel 25 young larvae were placed. Four nominal concentrations and the blank were tested. The test was performed in quadruplicate.

Results and discussion

Effect concentrationsopen allclose all
Key result
21 d
Dose descriptor:
Effect conc.:
> 96.3 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
act. ingr.
Basis for effect:
development rate
Key result
21 d
Dose descriptor:
Effect conc.:
> 96.3 mg/L
Nominal / measured:
meas. (initial)
Conc. based on:
act. ingr.
Basis for effect:
emergence rate
Results with reference substance (positive control):
not performed
Reported statistics and error estimates:
Standard deviations were calculated with the statistics program of the pocket calculator HP 48SX.
Multiple testing of the means using Dunnett's test versus 'control', with a = 0.05; unilateral (smaller), were performed.

Any other information on results incl. tables

nominal concentration


measured concentration (arith. mean) [mg/L]

measured concentration (arith. mean)

[% of nominal concnetration]

emergence rate 

emergence rate

[deviation in % of control]

mean of developement ratemean of development rate [deviation in % of control]
control<1 0.84100.00.072100.0

The pH of bottom sediment was not at target of 6.0 ± 0.5 at the start of the test but adjusted to pH 7.2. This deviation from
the target range was not relevant to the result.

At the start of the test, the pH value of the 100 mg/L test vessels was not in the proposed range (pH 6.0 - 9.0) . This is a minor deviation and not relevant to the outcome of the test. In the control and in the test batches 25 mg/L and 12.5 mg/L the temperature range was slightly exceeded on day 14 of the test in some parallels.

The test was terminated prematurely on the 21st day of the test, when it was evident that a further extension of the test could have no influence on the result.

Applicant's summary and conclusion

Validity criteria fulfilled:
not applicable
An ecologically risk potential for aquatic developmental stages of midges was not observed. A 21-d NOEC of > 96.3 mg/L (emergence) and a 21-d NOEC of > 96.3 mg/L (development rate) were derived. 
Executive summary:

In a study regarding the toxicity of 2-phosphonobutane-1,2,4-tricarboxylic acid towards sediment-dwelling midges, Chironomus ripuarius,  according to preliminary BBA- guideline : "Effects of plant protection products on the development of sediment-dwelling larvae of Chironomus ripuarius in a water-sediment system" the effect regarding emergence and development rate were determined over 21 days. 

The test was performed on nominal concentrations of 100 mg/L, 50 mg/L, 25 mg/L, and 12.5 mg/L as well as a control with no substance, each in quadruple. Sampling was perfromed at the start of the test, twice a week and upon finalisation. Analytical monitoring was performed using ion chromatography. 

No effect on emergence or development rate could be observed. Therefore, the test was terminated prematurely on the 21st day, as a prolongation would have no effect on the outcome of the test. 

A 21-d NOEC of > 96.3 mg/L (emergence) and a 21-d NOEC of > 96.3 mg/L (development rate) were derived.