Artemisia herba-alba, ext.

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:

From 2017-03-23 to 2017-05-23

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

All validity criteria was successful

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Qualifier:

according to guideline

Guideline:

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

Deviations:

no

Principles of method if other than guideline:

Not applicable.

GLP compliance:

yes (incl. QA statement)

Remarks:

2017-03-13

Specific details on test material used for the study:

Not applicable

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic (adaptation not specified)

Details on inoculum:

- Test Species: Aerobic activated sludge (microorganisms from a domestic waste¬water treatment plant) was supplied by the sewage treatment plant of Rossdorf, Germany.

- Conditioning: The aerobic activated sludge used for this study was washed by centrifugation and the supernatant liquid phase was decanted. The solid material was re-suspended in test water and centrifuged again. This procedure was done three times. An aliquot of the final sludge suspension was weighed, dried and the ratio of wet sludge to its dry weight was determined. Based on this ratio, calculated aliquots of washed sludge suspension, corresponding to 3.5 g dry material per litre were mixed with test water (see 6.5) and aerated overnight. This suspension was used for the experiment.

Duration of test (contact time):

28 d

Initial conc.:

102.9 mg/L

Based on:

other: Test Item Loading Rate (initial concentration in medium C0)

Initial conc.:

279.7 mg/L

Based on:

ThOD

Remarks:

NH4

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium: Aliquots of washed sludge suspension were mixed with reconstitued test water.
- Reconstitued pure water: Analytical grade salts were added to pure water to prepare the following stock solutions:
a) 8.5 g KH2PO4, 21.75 g K2HPO4, 33.4 g Na2HPO4 x 2 H2O, 0.5 g NH4Cl filled up with pure water to 1000 mL volume; The pH-value was 7.4.
b) 11.25 g MgSO4 x 7 H2O filled up with pure water to 500 mL volume
c) 18.2 g CaCl2 x 2 H2O filled up with pure water to 500 mL volume
d) 0.125 g FeCl3 x 6 H2O filled up with pure water to 500 mL volume
In order to avoid precipitation of iron hydroxide in the stock solution d), one drop of concentrated HCl per litre was added before storage. 50 mL of stock solution a) and 5 mL of the stock solutions b) to d) were combined and filled up to a final volume of 5000 mL with pure water.
- Solubilising agent: Silicone oil AR20 was used as an emulsifying agent. (1%)
- Test temperature: 22°C ± 1°C
- pH: 7.4 to 8.1
- Continuous darkness: yes


TEST SYSTEM
- Culturing apparatus: The closed test flasks were incubated in a climatised chamber under continuous stirring.
- Number of culture flasks/concentration: 2
The test flasks were closed gas-tight by a measuring head. Potassium hydroxide solution (45%) was used for trapping the produced carbon dioxide. The amount of O2 consumed by the activated sludge was calculated from the decrease of pressure in the reaction vessel.
- Measuring equipment: BSB/BOD-Sensor- System, Aqualytic Dortmund, Germany
- Test performed in closed vessels due to significant volatility of test substance:

SAMPLING
O2 consumed daily by the activated sludge was calculated from the decrease of pressure in the reaction vessel.

CONTROL AND BLANK SYSTEM
- Inoculum blank: 2 (activated sludge + silicone oil + test water)
- Abiotic sterile control: 1
- Toxicity control: 1
- Other: Procedure control : (Ref. Item + activated sludge + silicone oil + test water)

Reference substance:

benzoic acid, sodium salt

Remarks:

Sodium Benzoate

Preliminary study:

No data.

Test performance:

No data.

Parameter:

% degradation (O2 consumption)

Remarks:

ThODNH4

Value:

72

Sampling time:

28 d

Details on results:

The 10-day windows began on day 4 after applica-tion, the mean value was calculated to be 13% bio-degradation. Therefore, the end of the 10-day win-dow was day 14. After correction for the mean bio-chemical oxygen demand of the inoculum controls the mean biodegradation percentage based on ThODNH4 at the end of the 10-day window was 56%; the criterion of the 10 day window was not passed. The mean biodegradation percentage at the end of the 28-day exposure period was 72%.

Results with reference substance:

The reference item sodium benzoate was sufficient-ly degraded to 77% after 14 days and to 77% after 28 days of incubation.
The percentage biodegradation of the reference item confirms the suitability of the used aerobic activated sludge inoculum.

Cumulative Biochemical Oxygen Demand (mg O2/L) in Test Flasks during the Test Period of 28 Days

Time	Flask No.
(days)	1	2	3	4	5	6	7
1	0	0	0	0	45	0	55
2	0	0	5	5	85	0	85
3	15	15	5	5	105	0	100
4	45	45	5	10	115	0	120
5	60	60	10	10	125	0	125
6	80	80	10	10	135	0	135
7	95	100	10	15	140	0	140
8	115	120	10	15	145	0	150
9	130	135	15	20	150	0	160
10	145	145	15	20	150	0	165
11	155	160	15	20	150	0	170
12	165	170	15	20	155	0	180
13	170	180	15	20	155	0	185
14	175	180	20	25	155	0	195
15	180	185	20	25	155	0	205
16	185	185	20	25	160	0	215
17	195	190	20	25	160	0	220
18	205	190	20	25	160	0	225
19	210	195	25	25	160	0	235
20	215	200	25	25	160	0	240
21	220	205	25	25	160	0	245
22	220	205	25	25	160	0	255
23	225	210	25	25	160	0	260
24	225	215	25	30	160	0	265
25	230	220	25	30	160	0	270
26	230	220	25	30	160	0	275
27	235	225	25	30	160	0	280
28	235	225	25	30	160	0	285

Flasks 1 and 2: Artemisia herba-alba oil (thuyone type)

Flasks 3 and 4: inoculum control

Flask 5:reference (procedure control)

Flask 6: abiotic control

Flask 7: toxicity control

Percentage Biodegradation of Test Item, of Sodium Benzoate and of the Toxicity Control based on ThOD_NH4:

Time	Percentage Biodegradation^1
(Days)	Artemisia herba-alba oil (thuyone type)^1		Sodium Benzoate^2	Toxicity control^1, 2
	Flask 1 [%]	Flask 2 [%]	Flask 5 [%]	Flask 7 [%]
1	0	0	26	12
2	0	0	46	18
3	4	4	58	21
4	13	13	62	25
5	18	18	67	25
6	25	25	73	28
7	30	31	74	28
8	37	38	77	30
9	40	42	77	32
10	46	45	77	33
11	49	51	77	34
12	53	54	80	36
13	55	58	80	37
14	55	56	77	38
15	57	58	77	40
16	58	58	80	43
17	62	60	80	44
18	66	60	80	45
19	66	61	78	47
20	68	62	78	48
21	70	64	78	49
22	70	64	78	51
23	72	66	78	52
24	71	67	77	53
25	73	69	77	54
26	73	69	77	55
27	74	70	77	56
28	74	70	77	57

            ¹ThOD_NH4of Artemisia herba-alba oil (thuyone type): 2.719 mg O₂/mg test item
            ²ThOD_NH4of sodium benzoate: 1.666 mg O₂/mg reference item

Biodegradation in the Toxicity Control

In the toxicity control containing both, the test item and the reference item sodium benzoate, 38% biodegradation was noted within 14 days and 57% biodegradation after 28 days of incubation. According to the test guidelines, the test item can be assumed to be not inhibitory to the aerobic activated sludge microorganisms because degradation was >25% within 14 days.

According to the test guidelines the test item can be assumed to be not inhibitory to the aerobic activated sludge microorganisms because degradation was >25% within 14 days.

Validity Criteria of the Study

- Inoculum Control: The oxygen demand of the inoculum control (medium and inoculum) was 27.5 mg O2/L and thus not greater than 60 mg O2/L within 28 days as required by the test guideline.

- pH-Value: The pH-value of the test item flasks at day 32 of the test was 7.4 and therefore within the range of pH 6.0 to 8.5 as required by the test guideline.

- Reference Item: The percentage degradation of the reference item should reach the level for ready biodegradability (>60%) within 14 days as required by the test guideline. The reference item sodium benzoate was degraded to more than 60% after 4 days of incubation.

- Test Item: The difference of duplicate values for the degradation of the test item at the plateau, at the end of the test and at the end of the 10-day window was less than 20%. The difference of duplicate values at day 28 differed by 4%. The validity criterion was fulfilled.

- Toxicity Control: If in a toxicity test, containing both the test substance and a reference compound less than 25% biodegradation (based on total ThOD) occurred within 14 days, the test substance can be assumed to be inhibitory. The biodegradation was 38% at day 14; the test item was not inhibitory.

All validity criteria are fulfilled.

Validity criteria fulfilled:

yes

Interpretation of results:

readily biodegradable

Conclusions:

Artemisia herba-alba oil (thuyone type) is considered to be readily biodegradable.

Executive summary:

The test item Artemisia herba-alba oil (thuyone type) was investigated for its ready biodegradability in a manometric respirometry test over a period of 28 days, according to the OECD 301F guideline. The biodegradation was followed by the oxygen uptake of the microorganisms during exposure. As a reference item sodium benzoate was tested simultaneously under the same conditions as the test item, and functioned as a procedure control.

The degradation rate of Artemisia herba-alba oil (thuyone type) reached 72% at 28 days and was therefore greater than 60% after 28 days.

Due to the fact that the substance is identified as an UVCB or as a complex, multi-constituent substance with structurally similar constituents, the 10-day window condition may be waived and the pass level applied at 28 days according to section 4.1.2.9.5 of CLP COMMISSION REGULATION (EU) No 286/2011 of 10 March 2011.

Therefore, Artemisia herba-alba oil (thuyone type) is considered to be readily biodegradable.

The reference item sodium benzoate was sufficiently degraded, thus confirming the suitability of the aerobic activated sludge inoculum used.

According to the test guidelines, the test item can be assumed to be not inhibitory to the aerobic activated sludge microorganisms because degradation was >25% within 14 days in the toxicity control.

The oxygen demand of the inoculum control (medium and inoculum) was not greater than 60 mg O₂/L within 28 days as required by the test guideline.

pH values were in the range required in the guideline. Therefore, all validity criteria of the study were fulfilled.

Therefore, this study is considered valid for that endpoint.

Results synopsys:

Test item: Artemisia herba-alba oil (thuyone type)

Biodegradation endpoint:Readily biodegradable based on the results at 28 days

Description of key information

Artemisia herba-alba oil (thuyone type) is considered to be readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

readily biodegradable

Additional information

A reliable experimental study was available for that endpoint.

The test item Artemisia herba-alba oil (thuyone type) was investigated for its ready biodegradability in a manometric respirometry test over a period of 28 days, according to the OECD 301F guideline.

The degradation rate of Artemisia herba-alba oil (thuyone type) reached 72% at 28 days and was therefore greater than 60% after 28 days.

Due to the fact that the substance is identified as an UVCB or as a complex, multi-constituent substance with structurally similar constituents, the 10-day window condition may be waived and the pass level applied at 28 days according to section 4.1.2.9.5 of CLP COMMISSION REGULATION (EU) No 286/2011 of 10 March 2011.

All validity criteria were fulfilled. Moreover, the reference item sodium benzoate was sufficiently degraded, thus confirming the suitability of the aerobic activated sludge inoculum used.

In addition, the test item can be assumed to be not inhibitory to the aerobic activated sludge microorganisms because degradation was >25% within 14 days in the toxicity control.

Therefore, this study was compliant with the guideline requirements. The study is considered valid for that endpoint.