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Biodegradation in water: screening tests

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Reference
Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study
Reference:
Composition 0
Qualifier:
according to
Guideline:
OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)
Principles of method if other than guideline:
The method used is basically the one described under no. 301F in the OECD Guidelines for Testing of chemicals. A measured volume of inoculated mineral medium, containing a known concentration of test substance (100 mg/l) as the nominal sole source of organic carbon, is stirred in a closed flask at a constant temperature (+/- 1 deg C.) for up to 28 days. The consumption of oxygen is determined by measuring the quantity of oxygen (produced electrolytically) required to maintain constant the gas volume in the respirometer flask. Evolved carbon dioxide is absorbed in soda lime pellets. The Biological Oxygen Demand (BOD), amount of oxygen taken up by the microbial population during biodegradation of the test chemical (corrected for uptake by blank inoculum, run in parallel) is expressed as a percentage of ThOD (Theoretical Oxygen Demand, calculated from the elemental composition, assuming that carbon is oxidized to carbon dioxide and hydrogen to water).
GLP compliance:
yes
Test material information:
Composition 1
Specific details on test material used for the study:
- IUPAC name: 2-phenylethyl 2-hydroxybenzoate
- Common name: Phenethyl Salicylate
- Molecular formula: C15H14O3
- Molecular weight : 242.2726 g/mol
- Smiles notation : c1ccc(cc1)CCOC(=O)c2ccccc2O
- InChl: 1S/C15H14O3/c16-14-9-5-4-8-13(14)15(17)18-11-10-12-6-2-1-3-7-12/h1-9,16H,10-11H2
- Substance type:Organic
- Physical state: Solid
- Physical apperance: white crystals
- Melting point: 41 deg C
- Lot No: 9000348758
- Purity: 100%
Oxygen conditions:
not specified
Inoculum or test system:
activated sludge (adaptation not specified)
Details on inoculum:
- Source of inoculum/activated sludge (e.g. location, sampling depth, contamination history, procedure): Fresh activated sludge from a biological waste water treatment plant treating predominantly domestic sewage was used.
- Preparation of inoculum for exposure: The sludge is collected in the morning, washed three times in the mineral medium (by centrifuging at 1000 g for 10 minutes, discarding the supernant and resuspending in mineral medium) and kept aerobic until being used on the same day.
- Cocnentration of inoculum: To obtain a concentration of 30 mg/l (dry weight) in a 250 ml flask, 2.28 ml of sludge is needed (inoculum).

Duration of test (contact time):
28 d
Initial conc.:
100 mg/L
Based on:
test mat.
Parameter followed for biodegradation estimation:
O2 consumption
Details on study design:
TEST CONDITIONS
- Composition of medium: Mineral medium was 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 is measured and if necessary adjusted to 7.4 +/- 0.2 with phosphoric acid or potassium hydroxide.
- Test temperature: 22 deg C.
- pH: 7.4 +/- 0.2
- pH adjusted: yes, if necessary pH was adjusted to 7.4 +/- 0.2 with phosphoric acid or potassium hydroxide.
- Suspended solids concentration: 3.289 g/l (dry weight).
- Other: The water used during this study is deionised water containing less than 10 mg/l dissolved organic carbon.

TEST SYSTEM
- Measuring equipment: The respirometer used during this study is an SAPROMAT D 12, made by J. M. VOITH GmbH, Heidenheim, Germany.
Reference substance:
other: Sodium benzoate (100 mg/l) (purity min. 99.0%) was used as a reference substance for the study.
Key result
Parameter:
% degradation (O2 consumption)
Value:
93
Sampling time:
28 d
Remarks on result:
other: Readily biodegradable
Details on results:
Oxygen uptakes, as read on the SAPROMAT meters, are corrected by deducing the basic oxygen uptake of sludge (flask 1/7 and 1/8) and proportionally to account for the small differences between actual and nominal concentrations of test and reference substances. Phenylethyl salicylate undergoes 93% biodegradation after 28 days in the test conditions. Biodegradation starts on day 3 and reached 80% at the end of the 10-day window (day 3 to 13).
Results with reference substance:
Degradation of sodium benzoate exceeded 40% after 7 days and 65% after 14 days,: the activity of the inoculum was thus verified and the test is considered as valid.

Biodegradability: Respirometric method

Test substance: Phenylethyl Salicylate

Study no.:99-E106

1.      Theoretical Demand

Formula: C15H14N0O3

Mol. Wt.: 242.27 g/mol

Theoretical Oxygen Demand (ThOD): 2.25 mg O2/mg

2.      Biodegradability

Biological Oxygen Demand (BOD, mg O2/l, adjusted to nominal concentrations):

 

 

 

Days

3

7

13

14

21

28

BOD sludge

1stflask     B1

2ndflask    B2

Mean        B

6.0

16.0

16.0

17.0

19.0

21.0

6.0

16.0

16.0

18.0

21.0

21.0

6.0

16.0

16.0

17.0

20.0

21.0

BOD Test subs.

1stflask     C1

2ndflask    C2

1stfl. Corr. C1-B

2ndfl. Corr. C2-B

75.0

144.2

189.9

198.6

224.9

229.8

72.2

149.4

203.9

209.8

225.4

229.8

69.0

128.2

173.9

181.1

204.9

208.8

66.2

133.4

187.9

192.3

205.4

208.3

% degr.

1stflask     D1

2ndflask    D2

Mean        D

31

57

77

81

91

93

29

59

84

85

91

93

30

58

80

83

91

93

B = (B1 + B2)/2

D1 = 100*(C1-B)/ThOD*[S]

D2 = 100*(C2-B)/ThOD*[S]

D = (D1 + D2)/2

[S] = initial test substance concentration (mg/l)

Biodegradability: Respirometric method

Reference substance:Sodium benzoate 

Study no.:12-E023

1.      Theoretical Demand

Formula: C7        H5          O2         Na1

Mol. Wt.: 144.1 g/mol

Theoretical Oxygen Demand (ThOD):1.67 mg O2/mg

2.      Biodegradability

Biological Oxygen Demand (BOD, mg O2/l, adjusted to nominal concentrations):

 

 

 

Days

3

7

13

14

21

28

BOD sludge

1stflask     B1

2ndflask    B2

Mean        B

14.0

16.0

16.0

17.0

19.0

21.0

13.0

16.0

16.0

18.0

21.0

21.0

13.5

16.0

16.0

17.5

20.0

21.0

BOD Refer. subs.

1stflask     A1

2ndflask    A2

1stfl. Corr. A1-B

2ndfl. Corr. A2-B

134.0

150.0

162.0

168.0

174.0

176.0

131.0

146.0

160.0

170.0

176.0

179.0

120.5

134.0

146.0

150.5

154.0

155.0

117.5

130.0

144.0

152.5

156.0

158.0

% degr.

1stflask     D1

2ndflask    D2

Mean        D

72

80

88

90

92

93

71

78

86

92

94

95

71

79

87

91

93

94

B = (B1 + B2)/2

D1 = 100*(A1-B)/ThOD*[S]

D2 = 100*(A2-B)/ThOD*[S]

D = (D1 + D2)/2

[S] = initial test substance concentration (mg/l)

Validity criteria fulfilled:
yes
Interpretation of results:
readily biodegradable
Conclusions:
Phenylethyl salicylate undergoes 93% biodegradation after 28 days in the test conditions. Biodegradation starts on day 3 and reached 80% at the end of the 10-day window (day 3 to 13). Thus, Phenylethyl salicylate should be regarded as readily biodegradable according to this test. The curves obtained with the reference substance alone and with Phenylethyl salicylate plus reference substance show no toxic effect of Phenylethyl salicylate to the micro-organisms at the test concentration.
Executive summary:

The ready biodegradability of Phenylethyl salicylat (CAS no. 87 -22 -9) was determined by the OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test).

Fresh activated sludge from a biological waste water treatment plant treating predominantly domestic sewage was used. The sludge is collected in the morning, washed three times in the mineral medium (by centrifuging at 1000 g for 10 minutes, discarding the supernant and resuspending in mineral medium) and kept aerobic until being used on the same day. Initial test substance concentration used in the study was 100 mg/l. Mineral medium was 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 is measured and if necessary adjusted to 7.4 +/- 0.2 with phosphoric acid or potassium hydroxide. The water used during this study is deionised water containing less than 10 mg/l dissolved organic carbon. Suspended solids concentration used in the study was 3.289 g/l. To obtain a concentration of inoculum of 30 mg/l (dry weight) in a 250 ml flask, 2.28 ml of sludge is needed (inoculum). Sodium benzoate (100 mg/l) (purity min. 99.0%) was used as a reference substance for the study. Degradation of sodium benzoate exceeded 40% after 7 days and 65% after 14 days, the activity of the inoculum was thus verified and the test is considered as valid. Phenylethyl salicylate undergoes 93% biodegradation after 28 days in the test conditions. Biodegradation starts on day 3 and reached 80% at the end of the 10-day window (day 3 to 13). The curves obtained with the reference substance alone and with Phenylethyl salicylate plus reference substance show no toxic effect of Phenylethyl salicylate to the micro-organisms at the test concentration (100 mg/l). Thus, Phenylethyl salicylate should be regarded as readily biodegradable according to this test.

Description of key information

The ready biodegradability of Phenylethyl salicylat (CAS no. 87 -22 -9) was determined by the OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test)

(SSS (Sustainability Support Services) has a letter of access, 2000). Fresh activated sludge from a biological waste water treatment plant treating predominantly domestic sewage was used. The sludge is collected in the morning, washed three times in the mineral medium (by centrifuging at 1000 g for 10 minutes, discarding the supernant and resuspending in mineral medium) and kept aerobic until being used on the same day. Initial test substance concentration used in the study was 100 mg/l. Mineral medium was 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 is measured and if necessary adjusted to 7.4 +/- 0.2 with phosphoric acid or potassium hydroxide. The water used during this study is deionised water containing less than 10 mg/l dissolved organic carbon. Suspended solids concentration used in the study was 3.289 g/l. To obtain a concentration of inoculum of 30 mg/l (dry weight) in a 250 ml flask, 2.28 ml of sludge is needed (inoculum). Sodium benzoate (100 mg/l) (purity min. 99.0%) was used as a reference substance for the study. Degradation of sodium benzoate exceeded 40% after 7 days and 65% after 14 days, the activity of the inoculum was thus verified and the test is considered as valid. Phenylethyl salicylate undergoes 93% biodegradation after 28 days in the test conditions. Biodegradation starts on day 3 and reached 80% at the end of the 10-day window (day 3 to 13). The curves obtained with the reference substance alone and with Phenylethyl salicylate plus reference substance show no toxic effect of Phenylethyl salicylate to the micro-organisms at the test concentration (100 mg/l). Thus, Phenylethyl salicylate should be regarded as readily biodegradable according to this test.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable

Additional information

Experimental key study for the target compound 2-phenylethyl 2-hydroxybenzoate (CAS No. 87-22-9) and supporting study for its read across substance were reviewed for the biodegradation end point which are summarized as below:

 

The ready biodegradability of test chemical Phenylethyl salicylat (CAS no. 87 -22 -9) was determined by the OECD Guideline 301 F (Ready Biodegradability: Manometric Respirometry Test) at Givuadan Roure SA Ecotoxicological Laboratory, Switzerland (SSS (Sustainability Support Services) has a letter of access, 2000). Fresh activated sludge from a biological waste water treatment plant treating predominantly domestic sewage was used. The sludge is collected in the morning, washed three times in the mineral medium (by centrifuging at 1000 g for 10 minutes, discarding the supernant and resuspending in mineral medium) and kept aerobic until being used on the same day. Initial test substance concentration used in the study was 100 mg/l. Mineral medium was 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 is measured and if necessary adjusted to 7.4 +/- 0.2 with phosphoric acid or potassium hydroxide. The water used during this study is deionised water containing less than 10 mg/l dissolved organic carbon. Suspended solids concentration used in the study was 3.289 g/l. To obtain a concentration of inoculum of 30 mg/l (dry weight) in a 250 ml flask, 2.28 ml of sludge is needed (inoculum). Sodium benzoate (100 mg/l) (purity min. 99.0%) was used as a reference substance for the study. Degradation of sodium benzoate exceeded 40% after 7 days and 65% after 14 days, the activity of the inoculum was thus verified and the test is considered as valid. Phenylethyl salicylate undergoes 93% biodegradation after 28 days in the test conditions. Biodegradation starts on day 3 and reached 80% at the end of the 10-day window (day 3 to 13). The curves obtained with the reference substance alone and with Phenylethyl salicylate plus reference substance show no toxic effect of Phenylethyl salicylate to the micro-organisms at the test concentration (100 mg/l). Thus, Phenylethyl salicylate should be regarded as readily biodegradable according to this test.

 

In a supporting weight of evidence study from authoritative database (J-CHECK, HSDB 2017 and Envichem, 2014) for the read across chemical Acetic acid, phenylmethyl ester (CAS no. 140-11-4), biodegradation experiment was conducted for 28 days for evaluating the percentage biodegradability of read across substance Acetic acid, phenylmethyl ester. The study was performed according to OECD Guideline 301 C (Ready Biodegradability: Modified MITI Test (I)).Concentration of inoculum i.e, sludge used was 30 mg/l and initial test substance conc. used in the study was 100 mg/l, respectively. The percentage degradation of read across substance Acetic acid, phenylmethyl ester was determined to be 95, 98 and 100% by BOD, TOC removal and GC parameter in 28 days. The test substance was partially hydrolyzed in water to form Benzyl alcohol and acetic acid. Thus, based on percentage degradation, Acetic acid, phenylmethyl ester is considered to be readily biodegradable in nature.

On the basis of above results for target chemical 2 -phenylethyl 2 -hydroxybenzoate (from experimental study report, 2000) and for its read across substance (from authoritative database J-CHECK, HSDB and Envichem), it can be concluded that the test substance 2 -phenylethyl 2 -hydroxybenzoate can be expected to be readily biodegradable in nature.