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Environmental fate & pathways

Biodegradation in water: screening tests

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Reference
Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1998-07-01 to 1998-10-08
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Study conducted in compliance with agreed protocols, with no or minor deviations from standard test guidelines and/or minor methodological deficiencies, which do not affect the quality of the relevant results. The study report was conclusive, done to a valid guideline and the study was conducted under GLP conditions.
Qualifier:
according to
Guideline:
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Deviations:
no
GLP compliance:
yes
Oxygen conditions:
aerobic
Inoculum or test system:
activated sludge, domestic (adaptation not specified)
Details on inoculum:
Fresh activated sludge from a biological waste water treatment plant treating predominantly domestic sewage (City of Geneva, Aire) was used.

The sludge is collected in the morning, washed three times in the mineral medium (by centrifuging at 1000 g for 10 minutes, discarding the supernatant and resuspending in mineral medium) and kept aerobic until being used on the same day.
Duration of test (contact time):
28 d
Initial conc.:
100 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
METHODS

Principle
A measured volume of inoculated mineral medium, containing a known concentration of test substance (100 mg/l) as the nominal sole source of organic carbon, is stirred in a closed flask at a constant temperature (± 1 °C) for up to 28 days. The consumption of oxygen is determined by measuring the quantity of oxygen (produced electrolytically) required to maintain constant the gas volume in the respirometer flask. Evolved carbon dioxide is absorbed in soda lime pellets. The amount of oxygen taken up by blank inoculum, run in parallel) is expressed as a percentage of ThOD (Theoretical Oxygen Demand, calculated from the elemental composition, assuming that carbon oxidised to carbon dioxide and hydrogen to water).

Water
The water used during this study is deionised water containing less than 10 mg/l dissolved organic carbon.

Stock solutions of mineral components

The following stock solutions were prepared:

Solution A:
KH2PO4 8.5g
K2HPO4 21.75g
Na2HPO4.2H2O 33.40g
NH4Cl 0.50g

dissolved in water and made up to 1 litre.

Solution B:
CaCl2 27.50g

dissolved in water and made up to 1 litre.

Solution C:
MgSO4.7H2O 22.50g

dissolved in water and made up to 1 litre.

Solution D:
FeCl3.6H2O 0.25g
HCL Conc. one drop

dissolved in water and made up to 1 litre.

Mineral Medium

Prepared by mixing 50 ml of solution A and 2000 ml deionised water, adding 5 ml of each of the solutions B, C and D and making up to 5 litres with deionised water. The pH is measured and if necessary adjusted to 7.4 ± 0.2 with phosphoric acid or potassium hydroxide.

Determination of dry weight suspended solids
The dry weight of suspended solids is determined by taking two 50 ml samples of the homogenised sludge, evaporating water on a steam bath, drying in an oven at 105 - 110 °C for two hours and weighing the residue.

Reference substance
Sodium benzoate (Fluka, Buchs, Switzerland, Art No. 71300), purity: min. 99.0 %

Toxicity of the test chemical
As suggested in the OECD 301 F method, the toxicity of the test chemical for the inoculum is checked. Therefore, a pair of flasks of the volumetric respirometer (SAPROMAT) are filled with mineral medium + test chemical (100mg/l + reference substance (100 mg/l) + inoculum and their respirations are recorded as for the other flasks. If they are lower than those of the flasks containing mineral medium + reference substance (100 mg/l) + inoculum, the test chemical can be assumed to be inhibitory to the inoculum used.

Inoculum
Fresh activated sludge from a biological waste water treatment plant treating predominantly domestic sewage (City of Geneva, Aire) was used.

The sludge is collected in the morning, washed three times in the mineral medium (by centrifuging at 1000 g for 10 minutes, discarding the supernatant and resuspending in mineral medium) and kept aerobic until being used on the same day.

Preparation of the flasks
test substance samples (25 mg, corresponding to 100 mg/l in a 250-ml flask) are weighed in small aluminium boats and added directly to the test flasks of the SAPROMAT, whereas reference substance samples (sodium benzoate) are added as 1.0 ml of a 25 mg/ml solution in mineral medium.

All flasks are filled with 250 ml mineral medium. Samples of test or reference substance, or both, are added. Then, a volume of suspended sludge corresponding to 7.5 mg dry weight ( as determined from 3.6, generally 1 to 3 ml) is added. Except when the test substance has an acid or alkaline character, the pH of each flask is not measured but assumed to be the same as the mineral medium , in order not to remove any floating undissolved test substance from the test medium by dipping a glass electrode in it. Neutral test substances, even sodium benzoate, were shown not to affect the pH of the medium by more than 0.1 pH unit. About 2 g of soda lime is placed in an attachment of the stopper, the flasks are closed and placed in the water bath of the SAPROMAT. After temperature and pressure equilibration, the oxygen meters of the instrument are set to zero (time zero of the experiment).

TEST CONDITIONS FOR THE PRESENT STUDY

Activated sludge
Dry weight of suspended solids: 2.186 g/l

To obtain a concentration of 30 mg/l (dry weight) in a 250 ml flask, 343 ml of sludge is needed ( inoculum).

Test and Reference substances

Nominal concentrations:

-test substance: 100 mg/l

-Reference substance: 100 mg/l
Reference substance:
other: Sodium Benzoate
Preliminary study:
Not applicable
Test performance:
Everyday the oxygen consumption of each flask is recorded and corrected temperature and stirring are checked. At the end of the test period (normally 28 days), the pH of each flask is measured again.
Key result
Parameter:
% degradation (O2 consumption)
Value:
93
Sampling time:
28 d
Details on results:
Benzyl salicylate undergoes 93 % biodegradation after 28 days in the test conditions. Biodegradation starts on day 1 and reaches 87 % at the end of the 10 -day window (days 1 to 11) - See attachment "Benzyl Salicylate_Ready biodegradability.pdf" for the graph showing % biodegradation over time.

Thus, Benzyl Salicylate should be regarded as readily biodegradable according to this test.

At the concentration used in the test (100 mg/l), Benzyl Salicylate is not inhibitory to the micro-organisms.
Results with reference substance:
Degradation of sodium benzoate exceeds 40 % after 7 days and 65 % after 14 days: the activity of the inoculum is thus verified and the test is considered as valid.
Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
The test material undergoes 93 % degradation after 28 days in the test conditions. Biodegradation starts on day 1 and reaches 87 % at the end of the 10-day window (days 1 to 11).

Thus, Benzyl Salicylate should be regarded as readily biodegradable under the strict terms and conditions of OECD Guideline No 301F.
Executive summary:

The Ready biodegradability of Benzyl Salicylate was determined by the Manometric Respiratory Test according to the OECD Guidelines for Testing of Chemicals, Method No. 301 F.

Benzyl salicylate undergoes 93 % biodegradation after 28 days in the test conditions. Biodegradation starts on day 1 and reaches 87 % at the end of the 10 -day window (days 1 to 11).

Thus, Benzyl Salicylate should be regarded as readily biodegradable according to this test.

At the concentration used in the test (100 mg/L), Benzyl Salicylate is not inhibitory to the micro-organisms.

Description of key information

Benzyl salicylate attained 93% degradation after 28-days in an OECD 301F ready biodegradation study, and fulfilling the 10-day window criterion, thus fulfilling the criteria as 'readily biodegradable'.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information

The Ready biodegradability of Benzyl Salicylate was determined by the Manometric Respiratory Test according to the OECD Guidelines for Testing of Chemicals, Method No. 301 F.

Benzyl salicylate undergoes 93 % biodegradation after 28 days in the test conditions. Biodegradation starts on day 1 and reaches 87 % at the end of the 10 -day window (days 1 to 11).

Thus, Benzyl Salicylate should be regarded as readily biodegradable according to this test.

At the concentration used in the test (100 mg/L), Benzyl Salicylate is not inhibitory to the micro-organisms.