cyclohexadecanone

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:

19 Sep - 18 Oct 2016

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:

1992

Qualifier:

according to guideline

Guideline:

EU Method C.4-D (Determination of the "Ready" Biodegradability - Manometric Respirometry Test)

Version / remarks:

2008

Qualifier:

according to guideline

Guideline:

EPA OPPTS 835.3110 (Ready Biodegradability)

Version / remarks:

1998

GLP compliance:

yes (incl. QA statement)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

- Source of inoculum/activated sludge: Abwasserreinigungsanlage Birs (aeration stage), Birsfelden, Switzerland
- Storage conditions: Wet sludge was suspended in mineral medium to obtain a concentration equivalent to 4 g dry material per liter. The sludge was aerated at room temperature.
- Storage length: during 1 d prior to use
- Preparation of inoculum for exposure: The activated sewage sludge was washed three times by centrifugation, decantation of the supernatant liquid phase and resuspension of the solid material in tap water and finally in mineral medium. Prior to use, the dry weight of the sludge was again determined and the sludge was diluted approximately 1:4 with mineral medium.
- Pretreatment: Aliquots of the homogenized final sludge suspension were weighed and dried and the dry weight of the suspended solids was determined.
- Concentration of sludge: Wet sludge was adjusted to 4 g dry material per liter during the holding period. For the test, defined volumes of this diluted activated sludge were added to the mineral medium in the test vessels to obtain a final concentration of 30 mg dry material per liter.
- Initial cell/biomass concentration: 30 mg dry material per liter

Duration of test (contact time):

28 d

Initial conc.:

100 mg/L

Based on:

test mat.

Initial conc.:

309 mg/L

Based on:

ThOD

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium: mineral medium, according to the testing guidelines, composed of analytical grade salts.
- Test temperature: 22 - 23 °C in a thermostatic cabinet
- pH: at test start: 7.4, at test end: 7.3 - 7.7
- pH adjusted: yes, the pH of the mineral medium was adjusted from 7.7 to 7.4 with a diluted hydrochloric acid solution.
- Suspended solids concentration: 30 mg dry material per liter
- Continuous darkness: no, the test vessels were incubated under diffuse light

TEST SYSTEM
- Culturing apparatus: OxiTop
- Number of culture flasks/concentration: 2
- Method used to create aerobic conditions: aeration of activated sludge prior use
- Measuring equipment: OxiTop pressure measuring heads and OxiTop OC110 controller
- Details of trap for CO2: sodium hydroxide pellets (alkali trap)
- Other: the test vessels were incubated in a thermostatic cabinet under continuous stirring.

SAMPLING
- Sampling frequency: pressure values from the measuring heads were recorded every 3 h
- Sampling method: OxiTop pressure measuring heads

CONTROL AND BLANK SYSTEM
- Inoculum blank: yes
- Abiotic sterile control: no
- Toxicity control: yes, 100 mg/L test item and 100 mg/L reference item and inoculum
- Procedure control: yes, 100 mg/L reference item and inoculum

Reference substance:

benzoic acid, sodium salt

Parameter:

% degradation (O2 consumption)

Value:

47

Sampling time:

28 d

Details on results:

The test substance had no inhibitory effect on the activity of activated sludge microorganisms at the tested concentration of 100 mg/L.
Since the test did not meet the 10-day window, the test substance was not readily biodegradable but inherent biodegradability can be indicated.

Results with reference substance:

In the procedure controls, the reference item was degraded by an average of 71% and 92% by Exposure Day 3 and 14, respectively, thus confirming suitability of the activated sludge (≥ 60% degradation by Exposure Day 14). By the end of the test (Day 28), average biodegradation was 95%.

Table 1. Oxygen Consumption in the Test Vessels

	Cumulative oxygen consumption [mg/L]
Time [days]	Test item		Inoculum control		Procedure control		Toxicity control
	Replicate No.		Replicate No.		Replicate No.		Replicate No.
	1	2	1	2	1	2	1
0	0	0	0	0	0	0	0
1	2	5	2	4	42	37	43
2	5	7	5	7	91	80	77
3	25	18	7	7	126	125	128
4	48	39	10	10	142	138	142
5	60	55	10	10	147	144	147
6	69	68	11	11	150	146	156
7	85	83	11	14	155	154	168
8	95	94	11	13	157	157	175
9	108	104	13	13	160	158	181
10	114	112	12	13	162	160	186
11	119	118	12	13	162	161	189
12	121	121	11	13	164	162	193
13	126	126	12	13	165	164	197
14	130	129	13	13	168	165	203
15	132	133	12	14	168	167	205
16	140	140	17	20	172	171	212
17	142	143	17	20	172	171	213
18	146	144	19	21	174	175	218
19	147	145	16	19	175	174	222
20	149	149	18	21	175	175	228
21	153	149	17	21	174	175	231
22	154	151	17	21	176	175	237
23	157	155	18	21	176	178	240
24	158	154	19	21	176	176	243
25	160	156	17	21	175	178	245
26	164	158	18	21	176	176	251
27	165	158	16	19	175	176	251
28	167	161	17	21	176	178	255

 Table 2. Biodegradation of the Test Item and the Reference Item

	Percentage Biodegradation
	Test item		Procedure control		Toxicity control
Time	Replicate No.		Replicate No.		Replicate No.
[days]	1	2	1	2	1
0	0	0	0	0	0
1	0	1	23	20	8
2	0	0	51	44	15
3	6	3	71	71	25
4	12	10	79	77	28
5	16	15	83	81	29
6	19	18	83	81	31
7	23	23	85	85	33
8	27	27	87	87	34
9	31	29	88	87	35
10	33	32	89	88	36
11	35	34	89	89	37
12	35	35	91	90	38
13	37	37	91	91	39
14	38	37	93	91	40
15	39	39	93	92	40
16	39	39	92	91	41
17	40	40	92	91	41
18	41	40	92	93	41
19	42	41	94	94	43
20	42	42	93	93	44
21	43	42	93	93	45
22	44	43	94	93	46
23	45	44	94	95	46
24	45	43	93	93	47
25	46	44	93	95	47
26	47	45	94	94	49
27	48	46	94	95	49
28	48	46	94	95	50
Mean (Day 28)	47		95		not applicable

Validity criteria fulfilled:

yes

Interpretation of results:

not readily biodegradable

Conclusions:

In conclusion, the test item was not readily biodegradable under the test conditions within 28 days. The obtained biodegradation value of 47% may be used to indicate inherent biodegradability, based on the "Revised Introduction to the OECD Guidelines for Testing of Chemicals, Section 3" (2006).
The test item had no inhibitory effect on the activity of activated sludge microorganisms at the tested concentration of 100 mg/L.

Executive summary:

The test item was investigated for its ready biodegradability in a manometric respirometry test over 28 days according to OECD 301 F (1992). The BOD of the test item in the test media significantly increased from Exposure Day 2 until approximately Day 10. Afterwards the increase continued to a weaker extend until test termination. At the end of the 28-day exposure period, the mean biodegradation of the test item amounted to 47%. However, the pass level for ready biodegradability, i.e. biodegradation of at least 60% of the ThOD in a 10-day window within the 28-day period of the test, was not reached. In conclusion, the test item was not readily biodegradable under the test conditions within 28 days. The obtained biodegradation value of 47% may be used to indicate inherent biodegradability, based on the "Revised Introduction to the OECD Guidelines for Testing of Chemicals, Section 3" (2006).

Description of key information

Inherently, ultimately biodegradable (70% within 14 d, OECD 302 C)

Not readily biodegradable (47% after 28 d, OECD 301 F)

Key value for chemical safety assessment

Biodegradation in water:

inherently biodegradable, fulfilling specific criteria

Additional information

Two experimental studies are available, in which the biodegradability of the substance was assessed according to OECD guidelines. The key study was conducted according to the modified OECD guideline 302 C and demonstrated that the substance is inherently, ultimately biodegradable. The supporting study was performed according to OECD 301 F and showed that the substance does not fulfill the criteria for ready biodegradability.

In the manometric respirometry test of the key study, activated sludge (100 mg dw/L) from a local, domestic wastewater treatment plant was inoculated with 30 mg/L test substance (based on 93 mg O2/L ThOD) for 28 days. The biodegradation process was followed by continuously measuring the biochemical oxygen demand (BOD) using manometric methods and expressed as a percentage of Theoretical Oxygen Demand (ThOD). Inoculum, procedure and toxicity controls were run in parallel.

The BOD of the test media significantly increased in the first two weeks of exposure. On Day 14, a mean biodegradation of 70% was reached, which amounted to 79% at the end of the 28-day exposure period. Since biodegradation was greater than 70% of the ThOD, the substance fulfills the criteria for inherent, ultimate biodegradability according to the “Revised Introduction to the OECD Guidelines for Testing of Chemicals, Section 2 (2006)”. The toxicity control showed that the test item had no inhibitory effect on the activity of activated sludge microorganisms at the tested concentration of 30 mg/L (> 25% within 14 days).

In the manometric respirometry test of the supporting study, activated sludge (30 mg dw/L) from a local sewage treatment plant was inoculated with 100 mg/L test substance for 28 days. The biodegradation process was followed by continuously measuring the consumed dissolved oxygen. Inoculum, procedure and toxicity controls were run in parallel. The mean biodegradation of the test item at the end of the 28-day exposure period was 47%. Thus, the pass level for ready biodegradability, i.e. biodegradation of at least 60% of the ThOD in a 10-day window within the 28 days period of the test, was not reached. By Day 28 average biodegradation of the procedure control was 95%, confirming the suitability of the activated sludge. The toxicity control indicated that the test item had no inhibitory effect on the activity of activated sludge microorganisms at the tested concentration of 100 mg/L.