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Biodegradation in water: screening tests

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biodegradation in water: ready biodegradability
Type of information:
migrated information: read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Study period:
From 19 June 2012 to 14 December 2012
1 (reliable without restriction)

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)
GLP compliance:

Test material

Constituent 1
Chemical structure
Reference substance name:
Sodium diisopropylnaphthalenesulphonate
EC Number:
EC Name:
Sodium diisopropylnaphthalenesulphonate
Cas Number:
Molecular formula:
sodium 2,3-diisopropylnaphthalene-1-sulfonate
Test material form:
solid: particulate/powder
migrated information: powder

Study design

Oxygen conditions:
Inoculum or test system:
activated sludge, domestic, non-adapted
Details on inoculum:
A sample of activated sludge was collected from one of the return lines at Burley Menston sewage treatment works (West Yorkshire, UK), which has a predominantly domestic waste-water catchment.
On arrival, the sample was aerated by means of a compressed air supply.
The suspended solids concentration of the activated sludge was determined by filtering a subsample (25 mL) through a pre-dried and pre-weighed glass microfibre filter (Whatman GF/C). The filter and retained solids were then dried in an oven and re-weighed. The weight of the sludge solids was determined from the difference in the weights before and after drying.
The activated sludge used in this study was not deliberately acclimatised or adapted to Supragil WP before exposure under test conditions.
Concentration of sludge: 30mg suspended solid / L
Duration of test (contact time):
28 d
Initial test substance concentration
Initial conc.:
15 mg/L
Based on:
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
CO2 evolution
Reference substance
Reference substance:
benzoic acid, sodium salt

Results and discussion

Test performance:
Validity Criteria
The following criteria were required for the study to be considered valid:
 the IC content of the mineral medium was less than 5% of the total carbon content at the beginning of the test (See note 1)
 degradation of the reference material reached 60% by Day 14
 duplicate percentage biodegradation values for vessels containing the test substance differed from one another by less than 20%
 total CO2 production from each blank control vessel did not exceed 120 mg.

Note 1: At a dose level of 15 mg carbon/L, the 5% equivalent would be 15/100 x 5 = 0.75 mg carbon/L. However, because the measurement is performed on the mineral medium concentrate which is diluted by a factor of three in the final culture, the concentrate may be three times as concentrated in terms of IC, therefore the maximum permitted IC concentration is 2.25 mg carbon/L.
% Degradation
% degradation (CO2 evolution)
Sampling time:
29 d

Any other information on results incl. tables


Inorganic Carbon Content of the Test Medium

The IC concentration of the test inoculated mineral salts medium concentrate was 0 mg carbon/L. The validity criterion of less than 5% carbon loading was therefore met.

Measured CO2 Yields as a Percentage of Theoretical

Carbon dioxide evolution and percentage biodegradation data are presented in Table 2, Table 3 and Figure 1. The total carbon dioxide production in the blank control vessels was 101 mg for each of the two vessels, satisfying the validity criterion of less than 120 mg.

Supragil WP

To be considered readily biodegradable, a test substance must achieve 60% biodegradation by the end of the test. Additionally, the rate of biodegradation must be sufficient to reach 60% within ten days of reaching 10%. Since no biodegradation was observed, Supragil WP cannot be considered readily biodegradable. Percent biodegradation values at each sampling interval, for the two replicates containing Supragil WP, did not vary, therefore satisfying the validity criterion of less than 20% difference.

Sodium Benzoate

Rapid CO2 generation commenced immediately and declined to a more gradual rate over the period of the incubation as shown in Figure 1. The mean percentage biodegradation had reached 82% by Day 14 and 97% by Day 28. The validity criterion of 60% biodegradation at 14 days was therefore met.

Toxicity Control

Assessment of biodegradation in the toxicity control was calculated for the sodium benzoate fraction. The rate of biodegradation of the reference substance in the presence of Supragil WP (75% at 14 days and 84% at 28 days) was slightly lower than that of the reference substance alone, suggesting that Supragil WP may have had a slight inhibitory effect on the sludge microorganisms under the test conditions.

Applicant's summary and conclusion

Validity criteria fulfilled:
Interpretation of results:
under test conditions no biodegradation observed
All validity criteria were satisfied and the results of this study are therefore consideredto be valid.

Mean CO2 evolution from Reaction product of naphthalene, propan-2-ol, sulfonated and neutralized by caustic soda remained at 0% of the theoretical CO2 yield over the 28 days. The level of biodegradation failed to meet the requirements for ready biodegradability and Reaction product of naphthalene, propan-2-ol, sulfonated and neutralized by caustic soda cannot, therefore, be classified as readily biodegradable. However, the results of this study do not necessarily suggest that Reaction product of naphthalene, propan-2-ol, sulfonated and neutralized by caustic soda would persist or accumulate in the aerobic environment.
Executive summary:

A study was performed to assess the ready biodegradability of the test item in an aerobic aqueous medium. The method was designed to be compatible with the OECD Guidelines for Testing of Chemicals (1992) No 301B, "CO2 evolution (Modified Sturm test).

The test substance, at a concentration of 15.0 mg organic carbon /L, was exposed to sewage treatment micro-organisms with mineral medium in sealed culture vessels in the dark at temperatures of 22 ± 2 °C for 28 days.

The degradation of the test item was assessed by the determination of the evolution CO2. Control solutions with inoculum and the reference item, sodium benzoate, together with a toxicity control were used for validation purposes.

The test item attained 0% degradation after 28 days and therefore cannot be considered to be readily biodegradable under the strict terms and conditions of OECD Guideline No 301B.

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