Reaction mass of 1,2,3,4,4a,5,6,7-octahydro-2,5,5-trimethyl-2-naphthol and 4-(2,6,6-trimethyl-2-cyclohexen-1-yl)butan-2-one

Biodegradation in water: screening tests

Link to relevant study record(s)

Reference

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

22 Jun - 20 Jul 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

Version / remarks:

17 Jul 1992

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Staatliches Gewerbeaufsichtsamt Hildesheim, Germany (03 Jan 2017)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge: Aqueous phase of non-adapted activated sludge from the Municipal sewage treatment plant, Hildesheim, Germany
- Storage conditions: Aerobic conditions by aeration with CO2-free air
- Storage length: 7 d (15 - 22 Jun 2017)
- Pretreatment: The activated sludge was washed twice with chlorine-free tap water. After the 2nd washing the settled sludge was resuspended in mineral salts medium and was maintained in aerobic conditions for 2 h. Then the sludge was homogenized with a blender. After sedimentation, the supernatant was decanted and stored.
- Concentration of sludge: 10 mL/L were used to initiate inoculation
- Initial cell concentration (colony forming units, CFU): 1.03 * 10^9 CFU/L (sludge); 1.03 * 10^7 CFU/L (test vessels)
- Other: The colony forming units (CFU) of the inoculum were determined before test start by standard dilution plate count.

Duration of test (contact time):

28 d

Initial conc.:

17 mg/L

Based on:

test mat.

Initial conc.:

50.3 other: mg O2/L

Based on:

ThOD

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium: Mineral salts medium, according to OECD 301 F
- Test temperature: 20.0 - 20.6 °C (occasionally below 20 °C, i.e. 19.9 °C, for short periods)
- pH: 7.73 (test start), 7.68/7.69 (test end)
- Continuous darkness: Yes
- Other: Continuous stirring in an incubator

TEST SYSTEM
- Culturing apparatus: 500 mL brown glass bottles filled with 250 mL test volume
- Number of culture flasks/concentration: 2 each for inoculum control and test item, 1 each for functional control and toxicity control
- Method used to create aerobic conditions: Aeration of inoculum prior to test
- Measuring equipment: OxiTop measuring heads (WTW) and OC110 controller with software (Achat OC, WTW)
- Details of trap for CO2: A rubber sleeve filled with soda lime was hung into the opening of the bottles to absorb the evolved CO2.
- Other: Separate replicates of test item and toxicity control were prepared for measurement of the pH value at test start.

SAMPLING
- Sampling frequency: Oxygen consumption was determined directly in the incubation vessels by the OxiTop measuring system at 360 measuring points (every 112 min).

CONTROL AND BLANK SYSTEM
- Inoculum blank: 250 mL inoculated test medium consisting of mineral medium stock solutions, ultrapure water and inoculum (2 replicates)
- Functional control: Reference item (45 mg/L; 75.2 mg O2/L) in 250 mL inoculated test medium (1 replicate)
- Toxicity control: 250 mL reference item medium consisting of reference item dissolved in inoculated test medium + 4.5 µL test item (1 replicate)

Reference substance:

benzoic acid, sodium salt

Key result

Parameter:

% degradation (O2 consumption)

Value:

39

Sampling time:

28 d

Details on results:

The mean of replicates reached the 10% level (beginning of biodegradation) on Day 11. The 60% pass level was not reached within the 28 d period of the study.

Results with reference substance:

The pass level for ready biodegradation (≥ 60% degradation) was reached within 3 d. The validity criterion that degradation should be ≥ 60% after 14 d was fulfilled.
Stages of biodegradation: The adaptation phase changed to degradation phase within 2 d (degradation > 10%) and biodegradation came to a maximum of 97% on Day 28.

VALIDITY CRITERIA

The study fulfilled the validity criteria set out by the guideline (Table 1).

 

Table 1: Validity criteria for OECD 301 F.

Criterion from the guideline	Outcome	Validity criterion fulfilled
Difference of extremes of replicate values of the removal of the test chemical at the plateau, at the end of the test or at the end of the 10-d window, as appropriate, is less than 20%.	2% difference on Day 28.	Yes
Percentage degradation of the reference compound has reached the pass level of 25% by day 14.	The toxicity control reached the pass level of 25% within 2 d.	Yes
The oxygen uptake of the inoculum blank is normally 20-30 mg O2/L and should not be greater than 60 mg/L in 28 days.	The mean oxygen depletion in the inoculum control was 10.6 mg O2/L on Day 28 (< 60 mg oxygen/L)	Yes

 

RESULTS

Biodegradation (Table 2) was expressed as the percentage of the measured biological oxygen demand (BOD) to the theoretical oxygen demand (ThOD).

Table 2: Biodegradation [%] of the test item, functional control and toxicity control.

Day [d]	Biodegradation [%]
	Functional control	Test item		Toxicity control (test item + reference item)
	R1	P1	P2	T1
1	6	0	0	7
2	51	0	0	33
3	65	0	0	39
4	72	0	0	46
5	76	1	1	49
6	78	1	1	50
7	80	3	3	53
8	82	4	4	56
9	84	5	6	58
10	87	6	9	60
11	88	9	11	61
12	90	12	13	62
13	91	16	17	64
14	91	18	19	65
15	92	21	21	66
16	93	22	22	66
17	94	24	25	67
18	94	26	26	68
19	94	27	28	68
20	95	28	30	69
21	96	29	32	70
22	96	31	33	71
23	95	33	35	71
24	96	34	36	71
25	96	35	38	72
26	96	36	38	72
27	96	37	39	73
28	97	38	40	73

Validity criteria fulfilled:

yes

Remarks:

For further details please refer to “Any other information on results incl. tables”.

Interpretation of results:

not readily biodegradable

Conclusions:

The substance is not readily biodegradable according to the available guideline study (39% in 28 d, OECD 301 F).

Executive summary:

The ready biodegradability of the test item was assessed according to OECD 301 F and GLP. In the Manometric Respirometry Test, 17 mg/L test item (corresponding to 50.3 mg O2/L) was inoculated with non-adapted activated sludge for 28 d in the dark. Degradation was followed by measuring the evolution of oxygen concentrations and expressed as % biological oxygen demand (BOD, measured) of theoretical oxygen demand (ThOD, calculated). The mean biodegradation of the test item reached the 10% level (beginning of biodegradation) on Day 11 and peaked at 39% on Day 28. The pass level of 60% biodegradation was not reached within the 28 d period. Therefore, the test item is classified as not readily biodegradable within 28 d. The functional control reached the pass level (> 60%) within 3 d, confirming the suitability of the inoculum. The toxicity control reached 65% degradation within 14 d, indicating that the activity of the activated sludge (i.e. degradation) was not inhibited by the test item at the tested concentration of 17 mg/L (nominal).

Description of key information

Not readily biodegradable (39% after 28 d, OECD 301 F)

Key value for chemical safety assessment

Biodegradation in water:

inherently biodegradable

Additional information

There is one study available, in which the ready biodegradability of the test item was assessed according to OECD 301 F and GLP.

In the Manometric Respirometry Test, 17 mg/L test item (corresponding to 50.3 mg O2/L) was inoculated with non-adapted activated sludge (1.03 * 10^7 CFU/L) for 28 d in the dark. Degradation was followed by measuring the evolution of oxygen concentrations and expressed as % biological oxygen demand (BOD, measured) of theoretical oxygen demand (ThOD, calculated).

The mean biodegradation of the test item reached the 10% level (beginning of biodegradation) on Day 11 and peaked at 39% on Day 28. The pass level of 60% biodegradation was not reached within the 28 d period. Therefore, the test item is classified as not readily biodegradable within 28 d. The functional control reached the pass level (> 60%) within 3 d, confirming the suitability of the inoculum. The toxicity control reached 65% degradation within 14 d, indicating that the activity of the activated sludge (i.e. degradation) was not inhibited by the test item at the tested concentration of 17 mg/L (nominal).