Hexyl benzoate

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:

15 December 2011 to 6 January 2012

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

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Remarks:

Swiss Federal Office of Public Health. Consumer protection directorate. Notification authority of chemicals. CH-3003 Bern (d.d. 2011-02-08)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic (adaptation not specified)

Details on inoculum:

- Source: Fresh activated sludge from a biological waste water treatment plant treating predominantly domestic sewage (Bois-de-Bay, Satigny, Switzerland) was used.
- Pre-treatment: The sludge is collected in the morning, washed three times in the mineral medium (by centrifuging at 1000 g for 10 minutes, discarding the supernatant and resuspending in mineral medium) and kept aerobic until being used on the same day.
- Dry weight of suspended solids: The dry weight of suspended solids was determined by taking two 50 mL samples of the homogenised sludge, evaporating water on a steam bath, drying in an oven at 105 - 110 °C for two hours and weighing the residue. The dry weight of suspended solids was determined at 6.41 g/L and further diluted to 1.53 g/L. To obtain a concentration of 30 mg/L (dry weight) in 103 mL total volume (reference test flasks), 2.00 mL of sludge was added (inoculum). To obtain a concentration of 30 mg/L (dry weight) in 255 mL total volume (blank control and test flasks), 5.00 mL of sludge was added (inoculum).

Duration of test (contact time):

32 d

Initial conc.:

30 mg/L

Based on:

test mat.

Initial conc.:

76.8 mg/L

Based on:

ThOD

Parameter followed for biodegradation estimation:

O2 consumption

Details on study design:

TEST CONDITIONS
- Composition of medium: The water used during this study is deionised water containing less than 10 mg/L dissolved organic carbon (DOC). The following stock solutions were prepared: Solution A: 8.5 g KH2PO4, 21.75 g K2HPO4, 33.4 g Na2HPO4·2H2O and 0.5 g NH4Cl dissolved in water and made up to 1 litre. Solution B: 27.5 g CaCl dissolved in water and made up to 1 litre. Solution C: 22.5 g MgSO4·7H2O dissolved in water and made up to 1 litre. Solution D: 0.25 g FeCl3·6H2O and one drop of concentrated HCl dissolved in water and made up to 1 litre. Mineral medium was then prepared by mixing 50 mL of solution A and 2000 mL deionised water, adding 5 mL of each of the solutions B, C and D and making up to 5 litres with deionised water. The pH was measured and if necessary adjusted to 7.4 ± 0.2 with phosphoric acid or potassium hydroxide.
- Test temperature: 22.1 °C to 22.9 °C.
- Preparation of test samples: Test substance samples (7.65 mg, corresponding to 30.0 mg/L in 255 mL of test medium) were weighed in small aluminium boats and added directly to the test flasks of the Oxitop, whereas reference the substance (sodium benzoate) was added as 1.00 mL of a 10.2 g/L solution in mineral medium, to give a total volume of 103 mL.
- Preparation of flasks: Flasks were filled with 250 mL of mineral medium (flasks containing reference substance: 100 mL). Samples of test or reference substance were added. Then suspended sludge diluted to a concentration of 1.53 g/L dry matter was added. Except when the test substance has an acid or alkaline character, the pH of each flask is not measured but assumed to be the same as the mineral medium, in order not to remove any floating undissolved test substance from the test medium by dipping a glass electrode in it. Neutral test substances, even sodium benzoate, were shown not to affect the pH of the medium by more than 0.1 pH unit. Two sodium hydroxide pellets were placed in the quivers on top of the bottle, and the flasks were closed tightly with the measuring heads. The flasks were allowed to equilibrate to the test temperature. The measurement was started by programming the measuring unit of the Oxitop test flasks, and the test flasks were placed in the temperature controlled cupboard of the Oxitop system. After temperature equilibration, the controller of the instrument started the data acquisition (time zero of the experiment).

PERFORMANCE OF THE TEST
Everyday the oxygen consumption of each flask was recorded and correct temperature and stirring were checked. At the end of the test period, the pH of each flask was measured again.

Reference substance:

benzoic acid, sodium salt

Remarks:

100 mg/L based on test. mat, 167 mg/L based on ThOD.

Key result

Parameter:

% degradation (O2 consumption)

Value:

61

Sampling time:

28 d

Parameter:

% degradation (O2 consumption)

Value:

63

Sampling time:

32 d

Details on results:

- The 10-day window criterion is not fulfilled (11 % biodegradation on day 1 and 47 % on day 11).
- Hexyl Benzoate did not inhibit the intrinsic respiration of the inoculum at the test concentration and was therefore considered to be non-toxic to the inoculum at the test concentration.

Table: Biodegradability of Test substance 

O2 uptake	Replicate	Day
		1	7	11	14	21	28	32
Sludge (inoculum blank)	1	10.8	25.6	29.6	32.3	36.3	39.0	40.4
	2	8.1	24.2	28.3	32.3	36.3	39.0	41.7
	Mean	9.5	24.9	29.0	32.3	36.3	39.0	41.1
Test substance + sludge	1	17.5	55.2	67.3	72.7	83.5	88.8	94.2
	2	18.8	52.6	63.4	67.4	75.5	82.2	84.9
Test substance	1	8.1	30.3	38.4	40.4	47.2	49.8	53.2
	2	9.4	27.7	34.4	35.1	39.2	43.2	43.9
% Biodegradation	Replicate	Day
		1	7	11	14	21	28	32
% Biodegradation of test substance	1	10.0	39.0	50.0	53.0	61.0	65.0	69.0
	2	12.0	36.0	45.0	46.0	51.0	56.0	57.0
	mean	11.0	38.0	47.0	49.0	56.0	61.0	63.0

 

Table: Biodegradability of Reference substance

O2 uptake	Replicate	Day
		5	7	14	21	28
Sludge (inoculum blank)	1	22.9	25.6	32.3	36.3	39.0
	2	21.5	24.2	32.3	36.3	39.0
	Mean	22.2	24.9	32.3	36.3	39.0
Reference substance + sludge	1	141.2	152.2	172.4	178.4	188.5
	2	146.2	152.2	178.4	188.5	193.5
Reference substance	1	119.0	127.3	140.1	142.1	149.5
	2	124.0	127.3	146.1	152.2	154.5
% Biodegradation	Replicate	Day
		5	7	14	21	28
Reference substance	1	71	76	84	85	90
	2	74	76	88	91	93
	mean	73	76	86	88	91

 

Validity criteria fulfilled:

yes

Remarks:

see 'Any other information on materials and methods incl. tables'.

Interpretation of results:

readily biodegradable

Conclusions:

The substance is readily biodegradable.

Executive summary:

The biodegradation potential of the substance in water was determined in a screening study according to OECD TG 301F (Manometric Respirometry) and in compliance with GLP criteria. In this study, 30 mg/L test substance (76.8 mg/L based on ThOD) was inoculated with activated sludge from a municipal wastewater treatment plant for 32 days under aerobic conditions. During the incubation period the biological oxygen demand (BOD) was measured and biodegradation expressed as percentage of the theoretical uptake (ThOD). All validity criteria were met. No toxicity control was performed but as Hexyl benzoate did not inhibit the intrinsic respiration of the inoculum at the tested concentration, it was considered to be non-toxic to the inoculum at this concentration.

After 28 days incubation 61% of the substance was biodegraded. The 10-day window criterion was not fulfilled with observed biodegradation of 11% on day 1 and 47% on day 11. This is considered to be attributable to the fact that esters are usually not viewed as a single substance by microorganisms. Ester hydrolysis products are sequentially degraded, often by different microorganisms. The 10-day window criterion should therefore not be used as a pass/fail criterion and the substance should be regarded as readily biodegradable according to this test.

Description of key information

The biodegradation potential of Hexyl benzoate in water was determined in a screening study according to OECD TG 301F (Manometric Respirometry) and in compliance with GLP criteria. In this study, 30 mg/L test substance (76.8 mg/L based on ThOD) was inoculated with activated sludge from a municipal wastewater treatment plant for 32 days under aerobic conditions. During the incubation period the biological oxygen demand (BOD) was measured and biodegradation expressed as percentage of the theoretical uptake (ThOD). All validity criteria were met. No toxicity control was performed but as Hexyl benzoate did not inhibit the intrinsic respiration of the inoculum at the tested concentration, it was considered to be non-toxic to the inoculum at this concentration.

After 28 days incubation, 61% of the substance was biodegraded. The 10-day window criterion was not fulfilled with observed biodegradation of 11% on day 1 and 47% on day 11. This is considered to be attributable to the fact that esters are usually not viewed as a single substance by microorganisms. Ester hydrolysis products are sequentially degraded, often by different microorganisms. The 10-day window criterion should therefore not be used as a pass/fail criterion and the substance should be regarded as readily biodegradable according to this test.

Key value for chemical safety assessment

Biodegradation in water:

readily biodegradable