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Environmental fate & pathways

Biodegradation in water: screening tests

Administrative data

biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
EU Method C.4-E (Determination of the "Ready" Biodegradability - Closed Bottle Test)
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
1,3-dioxane, 2-(3,3-dimethyl-1-cyclohexen-1-yl)-2,5,5-trimethyl
Cas Number:
Molecular formula:
1,3-dioxane, 2-(3,3-dimethyl-1-cyclohexen-1-yl)-2,5,5-trimethyl
Specific details on test material used for the study:
Maderal, reference ZA1076
Batch n° 100026324
Received the 08/07/2015 from the sponsor
ThOD for Maderal = 2.76 g/g
Presented as a colourless translucent liquid
Certified as 97.5% for isomer 1 of Maderal
Stored at room temperature protected from direct sun light

Study design

Oxygen conditions:
Inoculum or test system:
sewage, predominantly domestic (adaptation not specified)
Details on inoculum:
Surface water collected near area of Pau, France, and receiving predominantly domestic sewage.
Pre-conditioned to the experimental conditions under aerobic conditions for 6 days in the mineral medium at the test temperature: 250 mL of surface water were added with 0.25 mL of the stock solution, the volume was then made to 500 mL. Resulting solution was maintained under strong aeration and agitation at 20°C (+/-2°C) for 5 days. The preconditioned inoculum was further used at a rate of 2 mL/L of medium
Duration of test (contact time):
ca. 33 d
Initial test substance concentration
Initial conc.:
ca. 6.7 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
Medium Preparation:
Kh2PO4 : 8.50 mg/L
K2HPO4: 21.75 mg/L
Na2HPO4-2H2O: 33.40 mg/L
NH4Cl: 0.50 mg/L
CaCl2 : 27.50 mg/L
CaCl2-2H2O: 36.40 mg/L
MgSO4-7H2O: 22.50 mg/L
FeCl3-6H2O: 0.25 mg/L
The mineral medium was strongly aerated for at least 20min, and then allowed to stand for approximately 20h at the test temperature before use.

Number of flask:
Blank groupe: mineral medium+ inoculum (x2)
Test group: 6.7 mg/L test solution+mineral medium+inconulum (x2)
Reference group: 2.0 mg/L aniline+mineral medium+inoculum (x2)
Toxic control group: 6.7 mg/L of test substance+2.0 mg/L aniline+mineral medium+inoculum (x1)
Reference substance
Reference substance:
(>99%), batch N°034K0126

Results and discussion

% Degradationopen allclose all
Key result
% degradation (O2 consumption)
ca. 10.6
Sampling time:
23 d
Key result
% degradation (O2 consumption)
ca. 13
Sampling time:
33 d

BOD5 / COD results

Results with reference substance:
Aniline :
Beginning of the 10-day windows on day 9.
Pass-level reached on day 12 with 71.6% of biodegradation.
Total biodegradation observed on day 16

Toxic series
the biodegradation based on the total ThOD was 22.4% on day 16.
At the end of the test, the biodegradation reached 30% : The oxygen uptake corresponded to the biodegradation of the sole aniline, without inhibition.

Any other information on results incl. tables

The beginning of the 10 days windows was reached on day 23 for the Maderal, consequently, the incubation was prolonged up to day 33.

Primary degradation: Analytical determinations proved that recovery of Maderal was 17.7% after 23 days, and 4.2% after 33 days.

Applicant's summary and conclusion

Validity criteria fulfilled:
Interpretation of results:
not readily biodegradable
The 10-day windows was reached on day 23 (10.6% biodegradation)
At the end of the 10-day window the biodegradation of Maderal was only 13.0%
Maderal was considered as not easily biodegradable

It was thus concluded that Maderal was subjected to primary degradation.
Executive summary:

The biodegradation in water was performed according to the EU Method C.4-E (Determination of the "Ready" Biodegradability - Closed Bottle Test). The substance is not easily biodegradable