Essential oil of Blue tansy obtained from the flowers and leaves of Tanacetum annuum (Asteraceae) by steam distillation

Biodegradation in water: screening tests

Administrative data

Endpoint:

biodegradation in water: ready biodegradability

Type of information:

experimental study

Adequacy of study:

key study

Study period:

15 May 2019 - 21 June 2019

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2019

Report date:

2019

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 D (Ready Biodegradability: Closed Bottle Test)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Essential oil of Tanacetum annuum

IUPAC Name:

Essential oil of Tanacetum annuum

Test material form:

liquid

Study design

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic, non-adapted

Details on inoculum:

Secondary activated sludge (09-05-2019) was obtained from the wastewater treatment plant Nieuwgraaf in Duiven, The Netherlands. This plant is an activated sludge treatment plant treating predominantly domestic wastewater. The activated sludge was preconditioned to reduce the endogenous respiration rates. To this end, 0.4 g Dry weight (DW)/L of activated sludge was aerated for one week. The sludge was diluted in the BOD bottles (van Ginkel and Stroo, 1992).

Duration of test (contact time):

28 d

Initial test substance concentration

Initial conc.:

2 mg/L

Based on:

test mat.

Parameter followed for biodegradation estimation

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

Test bottles:
The test was performed in 0.30 L BOD (biological oxygen demand) bottles with glass stoppers.

Deionized water:
Deionized water containing < 1.0 mg/L of organic carbon was prepared in a water purification system.

Nutrients, and stocks:
Deionized water used in the Closed Bottle test contained per liter of water 8.5 mg KH2PO4, 21.7 mg K2HPO4, 26.7 mg Na2HPO4, 22.5 mg MgSO4·7H2O, 36.3 mg CaCl2 2H2O, 0.25 mg FeCl3·6H2O. Ammonium chloride was omitted from the medium to prevent nitrification.
Accurate administering of the test substance was accomplished by preparing a solid stock of 3.0 mg of test substance per g of silica gel in a 50-mL serum flask. Only part of the top layer of the silica gel was brought into contact with the test substance. The serum flask was closed with a screw top and the content was mixed vigorously. Subsequently, 0.20 g of silica gel with the test substance was added to the test bottles. The resulting concentration of test substance in the bottles was 2.0 mg/L. Next the bottles were filled with nutrient medium with inoculum and closed. Sodium acetate was added to the bottles using aqueous stock solution of 1.0 g/L.

Test procedure:
Use was made of 10 bottles containing only inoculum, 10 bottles containing inoculum and silica gel, 10 bottles containing inoculum and silica gel with test substance, 6 bottles containing inoculum and sodium acetate. The concentrations of the test substance, and sodium acetate in the bottles were 2.0 mg/L and 6.7 mg/L, respectively. Each of the prepared solutions was dispensed into the respective group of BOD bottles so that all bottles were completely filled without air bubbles. The zero time bottles were immediately analyzed for dissolved oxygen using an oxygen electrode. The remaining bottles were closed and incubated in the dark. Two duplicate bottles of all series were withdrawn for analyses of the dissolved oxygen concentration at day 7, 14, 21, and 28.

Test conditions:
The pH of the media for the control, control with silica gel, reference and test substance was 7.4 at the start of the test. The pH of the medium at day 28 was 7.2, 7.0, and 7.1 for the control, test substance and control with silica gel, respectively. The temperature ranged from 22.6 to 22.9 °C which is within the prescribed temperature range of 22 to 24°C.

Calculation of the results:

- Calculation of endogenous respiration
The endogenous respiration (oxygen depletion in the control) was calculated as follows;
Oxygen depletion (endogenous respiration) (mg/L) = Mc (day 0) - Mc (day 28)
Mc is the mean oxygen level in the control bottle inoculated with activated sludge.

- Calculation of the theoretical oxygen demand (ThOD)
The ThODs of the test item and sodium acetate were calculated from their molecular formulae and molecular weights as follows.

-Calculation of the biochemical oxygen demand (BOD)
Provided that the oxygen concentrations in all bottles at the start of the test were equal, the amounts of oxygen consumed in test and reference compound bottles were calculated as follows:
Oxygen consumptionn (mg/L) by test substance = Mcs - Mt
Oxygen consumptionn (mg/L) by reference compound = Mc - Ma
Mc or cs is the mean oxygen level in the control bottles with and without silica gel n-days after the start of the test.
Mt or a is the mean oxygen concentration in the bottles containing the test substance (t) or the reference compound, sodium acetate (a), n-days after the start of the test.
The biological oxygen demand (BOD) mg/mg of the test substance and sodium acetate was calculated by dividing the oxygen consumption by the concentration of the test substance and sodium acetate in the closed bottle, respectively.

- Calculation of the biodegradation percentages
The biodegradation was calculated as the ratio of the biochemical oxygen demand (BOD) to the theoretical oxygen demand (ThOD).
 

Reference substance

Reference substance:

acetic acid, sodium salt

Remarks:

Purity: >99%

Results and discussion

Test performance:

The validity of the test is demonstrated by an endogenous respiration of 1.15 mg/L at day 28. Furthermore, the differences of the replicate values at day 28 were less than 20%. The biodegradation percentage of the reference compound, sodium acetate, at day 14 was 90%. Finally, the validity of the test is shown by oxygen concentrations >0.5 mg/L in all bottles during the test period.

% Degradation

% Degradation 1

Key result

Parameter:

% degradation (O2 consumption)

Value:

49

Sampling time:

28 d

% Degradation 2

Parameter:

% degradation (O2 consumption)

Value:

49

Sampling time:

21 d

% Degradation 3

Parameter:

% degradation (O2 consumption)

Value:

41

Sampling time:

14 d

% Degradation 4

Parameter:

% degradation (O2 consumption)

Value:

35

Sampling time:

7 d

% Degradation 5

Parameter:

% degradation (O2 consumption)

Value:

0

Sampling time:

0 d

Details on results:

Blue Tansy oil was biodegraded by 49% at day 28 in the Closed Bottle test.

BOD5 / COD results

Results with reference substance:

The biodegradation percentage of the reference compound, sodium acetate, at day 14 was 90 %

Any other information on results incl. tables

Toxicity

Inhibition of the degradation of a well-degradable compound, sodium acetate by the test substance in the Closed Bottle test was not determined because possible toxicity of the test substances to microorganisms degrading acetate is not relevant. Inhibition of the endogenous respiration of the inoculum by the test substance at day 7 was not detected. Therefore, no inhibition of the biodegradation due to the "high" initial test substance concentration is expected.

Dissolved oxygen concentrations (mg/L) in the closed bottles.

Time (days)	Oxygen concentration (mg/L)
	Oc	Oa	Ocs	Ot
0	9.0	9.0	9.0	9.0
	9.0	9.0	9.0	9.0
Mean (M)	9.00	9.00	9.00	9.00
7	8.4	4.0	8.2	5.8
	8.4	4.0	8.3	6.2
Mean (M)	8.40	4.00	8.25	6.00
14	8.2	3.5	8.1	5.5
	8.2	3.5	8.1	5.5
Mean (M)	8.20	3.50	8.10	5.50
21	8.0		7.9	4.7
	8.0		7.9	4.9
Mean (M)	8.00		7.90	4.80
28	7.9		7.8	4.7
	7.8		7.8	4.7
Mean (M)	7.85		7.80	4.70

O_c     Mineral nutrient solution with only inoculum.

O_cs    Mineral nutrient solution with inoculum and silica gel

O_t     Mineral nutrient solution with inoculum, test substance (2.0 mg/L) and silica gel

O_a      Mineral nutrient solution with inoculum and sodium acetate (6.7 mg/L).

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Interpretation of results:

not readily biodegradable

Conclusions:

Blue Tansy oil was biodegraded by 49% at day 28 in the Closed Bottle test. The test substance should therefore not be classified as readily biodegradable.

Executive summary:

In order to assess the biodegradation of Blue Tansy oil (UVCB), a screening test was performed according to OECD TG 301D (Closed Bottle test) and under GLP conditions. In this study, the test item (2 mg/L) was exposed to activated sludge which was spiked to a mineral nutrient solution, dosed in closed bottles and incubated in the dark at 22-24°C for 28 days. The degradation of the test item was assessed by the measurement of oxygen consumption. According to the results of this study, the test item did not cause a reduction in the endogenous respiration at day 7. The test substance is therefore considered to be non-inhibitory to the inoculum. Blue Tansy oil was biodegraded by 49% at day 28 in the OECD 301D Closed Bottle test and should therefore not be classified as readily biodegradable.