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Biodegradation in water: screening tests

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Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included under 'Attached justification' in IUCLID section 13 and 'Cross-reference'.
Reason / purpose:
read-across: supporting information
Qualifier:
no guideline followed
Principles of method if other than guideline:
The test was performed following the method of J. Blok, as detailed in the paper A repetitive die-away test combining several biodegradability test procedures, published in the journal Int. Biodeterior. Bull (!SSN 0020-6164) 15 (2) 1979 (57--63), with some modifications
GLP compliance:
not specified
Oxygen conditions:
aerobic
Inoculum or test system:
not specified
Details on inoculum:
Details of the inoculum used are not outlined in the paper, but the test was performed following the previously published Blok method (1979). For the Blok method preparation of the inoculum is reported as follows:

To obtain a polyvalent inoculum, various sources of aerobic organisms are used together. To 70 ml tap water the following are added:
10 ml stock solution A (see below)
10 ml fresh activated sludge
10 ml fresh river sludge
I ml fresh domestic sewage
500 mg fresh compost or garden soil.

Solution A:
KH2P04 2.83 g per litre
K2HP04 7.25 g per litre
Na2HP04.2aq 11.13 g per litre

This mixture is homogenised with an ultrasonic mixer for 10 - 20 seconds and paper-filtered. The filtrate is used as soon as possible, in any event within one hour. 3 ml of this inoculum is added to each 600 mi of mineral medium.

The mineral medium is obtained by diluting 1 : 1000 of equal quantities of the stock solutions A - F of composition given below with distilled water. The constituents suffice for the formation of about 150-200 mg/litre biomass or for the ultimate biodegradation of about 1 g/litre of test compound.

Solution A: KH2P04 = 2.83 g per litre; K2HP04 = 7.25 g per litre; Na2HP04.2aq = 11.13 g per litre
Section B: MgS04.7aq = 75 g per litre; CaCI2 = 9.2 g per litre; FeCI3.6aq = 0.083 g per litre
Solution C: NH4CI = 127 g per litre; Solution D: Allylthiourea = 2 g per litre
Solution E: Yeast extract = 5 g per litre
Solution F: H3B04 = 1.2 g per litre; CuSO4.Saq = 1.2 g per litre; KI = 05 g per litre; MnSO4 = 23 g per litre; NaMoO4 = 23 g per litre; ZnSO4.1aq = 4.7 g per litre
Duration of test (contact time):
28 d
Initial conc.:
20 mg/L
Based on:
other: Carbon
Parameter followed for biodegradation estimation:
DOC removal
Remarks:
Based on Blok method
Parameter followed for biodegradation estimation:
O2 consumption
Remarks:
Based on Blok method
Details on study design:
The test was performed following the previously published Blok method (1979) with the following modifications:
- The test concentration was chosen to correspond to 20 mg C/I.
- The ultrasound treatment was done before the inoculation.
- The inoculum was five times that of the OECD Screening Test.
- The test compound was added only once at the beginning.
- The test was run for 28 days.
- The bottles were fastened lying on the trays of the shaking machine.
- The reairation of the culture medium after each intermediate oxygen determination was accomplished by shaking the open upright bottle for one hour.

Details of the calculation of results are not provided in the paper, but the study followed the Blok method, which details that results should be calculated as follows:
The blank value (DOCtb) is subtracted from the measured amount of dissolved organic carbon (DOCt). The added amount of organic carbon (TOCadded) is calculated; for water-insoluble compounds this is a theoretical amount based on the dry weight addition. The results are expressed as per cent elimination of the dissolved organic carbon, calculated as shown:
per cent eliminated = ((TOCadded - (DOCt - DOCtb))/TOCadded)*100.

The oxygen uptake is corrected for that in the blank and related to a measured chemical oxygen demand or to a theoretical oxygen demand (TOD) of the amount added. The maximum oxygen uptake per litre of flask volume is 2/3 of this TOD. The total oxygen per litre of flask volume at 20°C and 76 cm Hg before the incubation is:
- in air 1/3x 1/22.4x (273/293) x32 x 0.21 = 93mg O2
- in water 2/3 x 8.8 = 6mg O2
Therefore the total = 99 mg O2
This figure has a standard variation of + 2 mg because of variations in the atmospheric pressure.

The per cent degradation follows from:
% degradtion = (((DOtb -DOt)*99) /(DOst*2/3TOD))*100
where:
DOtb = dissolved oxygen in blank after incubation
DOt = dissolved oxygen in test after incubation
DOst = dissolved oxygen at saturation (known from the temperature) after incubation.
TOD = Total (or Theoretical) oxygen demand of the substance added at the beginning of incubation (mg/litre water).
Reference substance:
not specified
Parameter:
other: % ThOD
Value:
84
Sampling time:
28 d
Details on results:
The study was also performed following the same method for C8/18 fatty acid, C18 -stearic acid and C22 -behenic acid and the biodegradation was assessed to be 100 %, 79 % and 69 %, respectively.
Validity criteria fulfilled:
not specified
Interpretation of results:
readily biodegradable
Conclusions:
The test item was shown to have degraded by 84% over 28 days by the modified Blok method.
Executive summary:

The biodegradability of the test item was assessed using a modified version of the Blok method (Blok, 1979) conducted over 28 days. Biodegradation was assessed based on theoretical O2 demand. After 28 days it was determined that the test item had degraded 84% from the starting concentration.

The publication is a near guideline study suitable for assessment of this endpoint with restrictions.

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included under 'Attached justification' in IUCLID section 13 and 'Cross-reference'.
Reason / purpose:
read-across: supporting information
Qualifier:
no guideline followed
Principles of method if other than guideline:
The test was performed following the method of J. Blok, as detailed in the paper A repetitive die-away test combining several biodegradability test procedures, published in the journal Int. Biodeterior. Bull (!SSN 0020-6164) 15 (2) 1979 (57--63), with some modifications
GLP compliance:
not specified
Specific details on test material used for the study:
None reported
Oxygen conditions:
aerobic
Inoculum or test system:
not specified
Details on inoculum:
Details of the inoculum used are not outlined in the paper, but the test was performed following the previously published Blok method (1979). For the Blok method preparation of the inoculum is reported as follows;

To obtain a polyvalent inoculum, various sources of aerobic organisms are used together. To 70 ml tap water the following are added:
10 ml stock solution A (see below)
10 ml fresh activited sludge
10 ml fresh river sludge
I ml fresh domestic sewage
500 mg fresh compost or garden soil.

Solution A:
KH2P04 2.83 g per litre
K2HP04 7.25 g per litre
Na2HP04.2aq 11.13 g per litre

This mixture is homogenised with an ultrasonic mixer for 10 - 20 seconds and paper-filtered. The filtrate is used as soon as possible, in any event within one hour. 3 ml of this inoculum is added to each 600 mi of mineral medium.

The mineral medium is obtained by diluting 1 : 1000 of equal quantities of the stock solutions A - F of composition given below with distilled water. The constituents suffice for the formation of about 150-200 mg/litre biomass or for the ultimate biodegradation of about 1 g/litre of test compound.

Solution A: KH2P04 = 2.83 g per litre; K2HP04 = 7.25 g per litre; Na2HP04.2aq = 11.13 g per litre
Section B: MgS04.7aq = 75 g per litre; CaCI2 = 9.2 g per litre; FeCI3.6aq = 0.083 g per litre
Solution C: NH4CI = 127 g per litre; Solution D: Allylthiourea = 2 g per litre
Solution E: Yeast extract = 5 g per litre
Solution F: H3B04 = 1.2 g per litre; CuSO4.Saq = 1.2 g per litre; KI = 05 g per litre; MnSO4 = 23 g per litre; NaMoO4 = 23 g per litre; ZnSO4.1aq = 4.7 g per litre
Duration of test (contact time):
28 d
Initial conc.:
20 mg/L
Based on:
other: Carbon
Parameter followed for biodegradation estimation:
DOC removal
Remarks:
Based on Blok method
Parameter followed for biodegradation estimation:
O2 consumption
Remarks:
Based on Blok method
Details on study design:
The test was performed following the previously published Blok method (1979) with the following modifications:
- The test concentration was chosen to correspond to 20 mg C/I.
- The ultrasound treatment was done before the inoculation.
- The inoculum was five times that of the OECD Screening Test.
- The test compound was added only once at the beginning.
- The test was run for 28 days.
- The bottles were fastened lying on the trays of the shaking machine.
- The re-aeration of the culture medium after each intermediate oxygen determination was accomplished by shaking the open upright bottle for one hour.

Details of the calculation of results are not provided in the paper, but the study followed the Blok method, which details that results should be calculated as follows:
The blank value (DOCtb) is subtracted from the measured amount of dissolved organic carbon (DOCt). The added amount of organic carbon (TOCadded) is calculated; for water-insoluble compounds this is a theoretical amount based on the dry weight addition. The results are expressed as per cent elimination of the dissolved organic carbon, calculated as shown:
per cent eliminated = ((TOCadded - (DOCt - DOCtb))/TOCadded)*100.
The oxygen uptake is corrected for that in the blank and related to a measured chemical oxygen demand or to a theoretical oxygen demand (TOD) of the amount added. The maximum oxygen uptake per litre of flask volume is 2/3 of this TOD. The total oxygen per litre of flask volume at 20°C and 76 cm Hg before the incubation is:
- in air 1/3x 1/22.4x (273/293) x32 x 0.21 = 93mg O2
- in water 2/3 x 8.8 = 6mg O2
Therefore the total = 99 mg O2
This figure has a standard variation of + 2 mg because of variations in the atmospheric pressure.
The per cent degradation follows from:
% degradation = (((DOtb -DOt)*99) /(DOst*2/3TOD))*100
where:
DOtb = dissolved oxygen in blank after incubation
DOt = dissolved oxygen in test after incubation
DOst = dissolved oxygen at saturation (known from the temperature) after incubation.
TOD = Total (or Theoretical) oxygen demand of the substance added at the beginning of incubation (mg/litre water).
Reference substance:
not specified
Parameter:
other: % ThOD
Value:
79
Sampling time:
28 d
Details on results:
The study was also performed following the same method for C8/18 fatty acid, C16 -palmitic acid and C22 -behenic acid and the biodegradation was assessed to be 100 %, 84 % and 69 %, respectively.
Validity criteria fulfilled:
not specified
Interpretation of results:
readily biodegradable
Conclusions:
The test item was shown to have degraded by 79% over 28 days by the modified Blok method.
Executive summary:

The biodegradability of the test item was assessed using a modified version of the Blok method (Blok, 1979) conducted over 28 days. Biodegradation was assessed based on theoretical O2 demand. After 28 days it was determined that the test item had degraded 79% from the starting concentration.

The publication is a near guideline study suitable for assessment of this endpoint, with some restrictions (Klimisch 2).

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included under 'Attached justification' in IUCLID section 13 and 'Cross-reference'.
Reason / purpose:
read-across: supporting information
Qualifier:
equivalent or similar to
Guideline:
EPA OTS 796.3260 (Ready Biodegradability: Modified Sturm Test)
GLP compliance:
not specified
Specific details on test material used for the study:
Not reported
Oxygen conditions:
aerobic
Inoculum or test system:
water (not specified)
Duration of test (contact time):
21 d
Initial conc.:
58 other: mg
Based on:
other: Theoretical amount of initial CO2
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
TEST CONDITIONS
- Composition of medium: Not stated in paper but test method followed a modification of the Sturm test (Sturm, 1973). The Sturm test uses raw sewage, river water and BOD water
- Test temperature: Not stated in paper but test method followed a modification of the Sturm test (Sturm, 1973). 22 - 24 °C for the Sturm test.
- Aeration of dilution water: Yes, via peristaltic pump


TEST SYSTEM
- Number of culture flasks/concentration: Performed in duplicate
- Method used to create aerobic conditions: Perstitalic pump and magentic stirrer
- Measuring equipment: CO2 absorbers containing 100 mL Ba(OH)2
- Test performed in closed vessels due to significant volatility of test substance: No
- Test performed in open system: Not reported
- Details of trap for CO2 and volatile organics if used: CO2 absorbers containing 100 mL Ba(OH)2

SAMPLING
- Sampling frequency: Days 1, 3, 5, 7, 9,12, 14, 16, 19 and 21
- Sampling method: Not reported
- Sample storage before analysis: Not reported

CONTROL AND BLANK SYSTEM
- Other: Theoretical amount of initial CO2 was calculated as 58 mg

STATISTICAL METHODS: Calculated based on the average of two experiments
Reference substance:
not specified
Parameter:
% degradation (CO2 evolution)
Value:
2
Sampling time:
1 d
Parameter:
% degradation (CO2 evolution)
Value:
9
Sampling time:
3 d
Parameter:
% degradation (CO2 evolution)
Value:
24
Sampling time:
5 d
Parameter:
% degradation (CO2 evolution)
Value:
36
Sampling time:
7 d
Parameter:
% degradation (CO2 evolution)
Value:
53
Sampling time:
9 d
Parameter:
% degradation (CO2 evolution)
Value:
69
Sampling time:
12 d
Parameter:
% degradation (CO2 evolution)
Value:
76
Sampling time:
14 d
Parameter:
% degradation (CO2 evolution)
Value:
83
Sampling time:
16 d
Parameter:
% degradation (CO2 evolution)
Value:
90
Sampling time:
19 d
Parameter:
% degradation (CO2 evolution)
Value:
95
Sampling time:
21 d
Details on results:
The results obtained in the biodegradability determination of stearic acid show that this compound is degradable after an initial lag phase, reaching the “pass level” of biodegradation, corresponding to 60%, within 10 days.

Table 1: Results observed in the Sturm Test with Stearic Acida

 Days  CO2 produced (mg)  % Biodegradability
 1  1  2
 3  5  9
 5  14  24
 7  21  36
 9  31  53
 12  40  69
 14  44  76
 16  48  83
 19  52  90
 21  55  95

a The theoretical amount of initial CO2 was 58 mg. Indicated values are means of two experiments.

Validity criteria fulfilled:
not specified
Interpretation of results:
readily biodegradable
Conclusions:
The test item was shown to have degraded by 95% at day 21 in the Modified Sturm test.
Executive summary:

The biodegradability of the test item was assessed in a modified Sturm test conducted over 21 days. Biodegradation was assessed based on CO2 production. The test item is degradable after an initial lag phase, reaching the “pass level” of biodegradation, corresponding to 60%, within 10 days and after 21 days the test item showed 95% biodegradation.

The publication is a near guideline study suitable for assessment of this endpoint, with some restrictions (Klimisch 2).

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included under 'Attached justification' in IUCLID section 13 and 'Cross-reference'.
Reason / purpose:
read-across source
Qualifier:
no guideline followed
Principles of method if other than guideline:
The test was performed following the method of J. Blok, as detailed in the paper A repetitive die-away test combining several biodegradability test procedures, published in the journal Int. Biodeterior. Bull (!SSN 0020-6164) 15 (2) 1979 (57--63), with some modifications
GLP compliance:
not specified
Oxygen conditions:
aerobic
Inoculum or test system:
not specified
Details on inoculum:
Details of the inoculum used are not outlined in the paper, but the test was performed following the previously published Blok method (1979). For the Blok method preparation of the inoculum is reported as follows:

To obtain a polyvalent inoculum, various sources of aerobic organisms are used together. To 70 ml tap water the following are added:
10 ml stock solution A (see below)
10 ml fresh activated sludge
10 ml fresh river sludge
I ml fresh domestic sewage
500 mg fresh compost or garden soil.

Solution A:
KH2P04 2.83 g per litre
K2HP04 7.25 g per litre
Na2HP04.2aq 11.13 g per litre

This mixture is homogenised with an ultrasonic mixer for 10 - 20 seconds and paper-filtered. The filtrate is used as soon as possible, in any event within one hour. 3 ml of this inoculum is added to each 600 mi of mineral medium.

The mineral medium is obtained by diluting 1 : 1000 of equal quantities of the stock solutions A - F of composition given below with distilled water. The constituents suffice for the formation of about 150-200 mg/litre biomass or for the ultimate biodegradation of about 1 g/litre of test compound.

Solution A: KH2P04 = 2.83 g per litre; K2HP04 = 7.25 g per litre; Na2HP04.2aq = 11.13 g per litre
Section B: MgS04.7aq = 75 g per litre; CaCI2 = 9.2 g per litre; FeCI3.6aq = 0.083 g per litre
Solution C: NH4CI = 127 g per litre; Solution D: Allylthiourea = 2 g per litre
Solution E: Yeast extract = 5 g per litre
Solution F: H3B04 = 1.2 g per litre; CuSO4.Saq = 1.2 g per litre; KI = 05 g per litre; MnSO4 = 23 g per litre; NaMoO4 = 23 g per litre; ZnSO4.1aq = 4.7 g per litre
Duration of test (contact time):
28 d
Initial conc.:
20 mg/L
Based on:
other: Carbon
Parameter followed for biodegradation estimation:
DOC removal
Remarks:
Based on Blok method
Parameter followed for biodegradation estimation:
O2 consumption
Remarks:
Based on Blok method
Details on study design:
The test was performed following the previously published Blok method (1979) with the following modifications:
- The test concentration was chosen to correspond to 20 mg C/I.
- The ultrasound treatment was done before the inoculation.
- The inoculum was five times that of the OECD Screening Test.
- The test compound was added only once at the beginning.
- The test was run for 28 days.
- The bottles were fastened lying on the trays of the shaking machine.
- The reairation of the culture medium after each intermediate oxygen determination was accomplished by shaking the open upright bottle for one hour.

Details of the calculation of results are not provided in the paper, but the study followed the Blok method, which details that results should be calculated as follows:
The blank value (DOCtb) is subtracted from the measured amount of dissolved organic carbon (DOCt). The added amount of organic carbon (TOCadded) is calculated; for water-insoluble compounds this is a theoretical amount based on the dry weight addition. The results are expressed as per cent elimination of the dissolved organic carbon, calculated as shown:
per cent eliminated = ((TOCadded - (DOCt - DOCtb))/TOCadded)*100.

The oxygen uptake is corrected for that in the blank and related to a measured chemical oxygen demand or to a theoretical oxygen demand (TOD) of the amount added. The maximum oxygen uptake per litre of flask volume is 2/3 of this TOD. The total oxygen per litre of flask volume at 20°C and 76 cm Hg before the incubation is:
- in air 1/3x 1/22.4x (273/293) x32 x 0.21 = 93mg O2
- in water 2/3 x 8.8 = 6mg O2
Therefore the total = 99 mg O2
This figure has a standard variation of + 2 mg because of variations in the atmospheric pressure.

The per cent degradation follows from:
% degradtion = (((DOtb -DOt)*99) /(DOst*2/3TOD))*100
where:
DOtb = dissolved oxygen in blank after incubation
DOt = dissolved oxygen in test after incubation
DOst = dissolved oxygen at saturation (known from the temperature) after incubation.
TOD = Total (or Theoretical) oxygen demand of the substance added at the beginning of incubation (mg/litre water).
Reference substance:
not specified
Parameter:
other: % ThOD
Value:
84
Sampling time:
28 d
Details on results:
The study was also performed following the same method for C8/18 fatty acid, C18 -stearic acid and C22 -behenic acid and the biodegradation was assessed to be 100 %, 79 % and 69 %, respectively.
Validity criteria fulfilled:
not specified
Interpretation of results:
readily biodegradable
Conclusions:
The test item was shown to have degraded by 84% over 28 days by the modified Blok method.
Executive summary:

The biodegradability of the test item was assessed using a modified version of the Blok method (Blok, 1979) conducted over 28 days. Biodegradation was assessed based on theoretical O2 demand. After 28 days it was determined that the test item had degraded 84% from the starting concentration.

The publication is a near guideline study suitable for assessment of this endpoint with restrictions.

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included under 'Attached justification' in IUCLID section 13 and 'Cross-reference'.
Reason / purpose:
read-across source
Qualifier:
no guideline followed
Principles of method if other than guideline:
The test was performed following the method of J. Blok, as detailed in the paper A repetitive die-away test combining several biodegradability test procedures, published in the journal Int. Biodeterior. Bull (!SSN 0020-6164) 15 (2) 1979 (57--63), with some modifications
GLP compliance:
not specified
Specific details on test material used for the study:
None reported
Oxygen conditions:
aerobic
Inoculum or test system:
not specified
Details on inoculum:
Details of the inoculum used are not outlined in the paper, but the test was performed following the previously published Blok method (1979). For the Blok method preparation of the inoculum is reported as follows;

To obtain a polyvalent inoculum, various sources of aerobic organisms are used together. To 70 ml tap water the following are added:
10 ml stock solution A (see below)
10 ml fresh activited sludge
10 ml fresh river sludge
I ml fresh domestic sewage
500 mg fresh compost or garden soil.

Solution A:
KH2P04 2.83 g per litre
K2HP04 7.25 g per litre
Na2HP04.2aq 11.13 g per litre

This mixture is homogenised with an ultrasonic mixer for 10 - 20 seconds and paper-filtered. The filtrate is used as soon as possible, in any event within one hour. 3 ml of this inoculum is added to each 600 mi of mineral medium.

The mineral medium is obtained by diluting 1 : 1000 of equal quantities of the stock solutions A - F of composition given below with distilled water. The constituents suffice for the formation of about 150-200 mg/litre biomass or for the ultimate biodegradation of about 1 g/litre of test compound.

Solution A: KH2P04 = 2.83 g per litre; K2HP04 = 7.25 g per litre; Na2HP04.2aq = 11.13 g per litre
Section B: MgS04.7aq = 75 g per litre; CaCI2 = 9.2 g per litre; FeCI3.6aq = 0.083 g per litre
Solution C: NH4CI = 127 g per litre; Solution D: Allylthiourea = 2 g per litre
Solution E: Yeast extract = 5 g per litre
Solution F: H3B04 = 1.2 g per litre; CuSO4.Saq = 1.2 g per litre; KI = 05 g per litre; MnSO4 = 23 g per litre; NaMoO4 = 23 g per litre; ZnSO4.1aq = 4.7 g per litre
Duration of test (contact time):
28 d
Initial conc.:
20 mg/L
Based on:
other: Carbon
Parameter followed for biodegradation estimation:
DOC removal
Remarks:
Based on Blok method
Parameter followed for biodegradation estimation:
O2 consumption
Remarks:
Based on Blok method
Details on study design:
The test was performed following the previously published Blok method (1979) with the following modifications:
- The test concentration was chosen to correspond to 20 mg C/I.
- The ultrasound treatment was done before the inoculation.
- The inoculum was five times that of the OECD Screening Test.
- The test compound was added only once at the beginning.
- The test was run for 28 days.
- The bottles were fastened lying on the trays of the shaking machine.
- The re-aeration of the culture medium after each intermediate oxygen determination was accomplished by shaking the open upright bottle for one hour.

Details of the calculation of results are not provided in the paper, but the study followed the Blok method, which details that results should be calculated as follows:
The blank value (DOCtb) is subtracted from the measured amount of dissolved organic carbon (DOCt). The added amount of organic carbon (TOCadded) is calculated; for water-insoluble compounds this is a theoretical amount based on the dry weight addition. The results are expressed as per cent elimination of the dissolved organic carbon, calculated as shown:
per cent eliminated = ((TOCadded - (DOCt - DOCtb))/TOCadded)*100.
The oxygen uptake is corrected for that in the blank and related to a measured chemical oxygen demand or to a theoretical oxygen demand (TOD) of the amount added. The maximum oxygen uptake per litre of flask volume is 2/3 of this TOD. The total oxygen per litre of flask volume at 20°C and 76 cm Hg before the incubation is:
- in air 1/3x 1/22.4x (273/293) x32 x 0.21 = 93mg O2
- in water 2/3 x 8.8 = 6mg O2
Therefore the total = 99 mg O2
This figure has a standard variation of + 2 mg because of variations in the atmospheric pressure.
The per cent degradation follows from:
% degradation = (((DOtb -DOt)*99) /(DOst*2/3TOD))*100
where:
DOtb = dissolved oxygen in blank after incubation
DOt = dissolved oxygen in test after incubation
DOst = dissolved oxygen at saturation (known from the temperature) after incubation.
TOD = Total (or Theoretical) oxygen demand of the substance added at the beginning of incubation (mg/litre water).
Reference substance:
not specified
Parameter:
other: % ThOD
Value:
79
Sampling time:
28 d
Details on results:
The study was also performed following the same method for C8/18 fatty acid, C16 -palmitic acid and C22 -behenic acid and the biodegradation was assessed to be 100 %, 84 % and 69 %, respectively.
Validity criteria fulfilled:
not specified
Interpretation of results:
readily biodegradable
Conclusions:
The test item was shown to have degraded by 79% over 28 days by the modified Blok method.
Executive summary:

The biodegradability of the test item was assessed using a modified version of the Blok method (Blok, 1979) conducted over 28 days. Biodegradation was assessed based on theoretical O2 demand. After 28 days it was determined that the test item had degraded 79% from the starting concentration.

The publication is a near guideline study suitable for assessment of this endpoint, with some restrictions (Klimisch 2).

Endpoint:
biodegradation in water: ready biodegradability
Type of information:
read-across from supporting substance (structural analogue or surrogate)
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
study well documented, meets generally accepted scientific principles, acceptable for assessment
Justification for type of information:
REPORTING FORMAT FOR THE ANALOGUE APPROACH
Further information is included under 'Attached justification' in IUCLID section 13 and 'Cross-reference'.
Reason / purpose:
read-across source
Qualifier:
equivalent or similar to
Guideline:
EPA OTS 796.3260 (Ready Biodegradability: Modified Sturm Test)
GLP compliance:
not specified
Specific details on test material used for the study:
Not reported
Oxygen conditions:
aerobic
Inoculum or test system:
water (not specified)
Duration of test (contact time):
21 d
Initial conc.:
58 other: mg
Based on:
other: Theoretical amount of initial CO2
Parameter followed for biodegradation estimation:
CO2 evolution
Details on study design:
TEST CONDITIONS
- Composition of medium: Not stated in paper but test method followed a modification of the Sturm test (Sturm, 1973). The Sturm test uses raw sewage, river water and BOD water
- Test temperature: Not stated in paper but test method followed a modification of the Sturm test (Sturm, 1973). 22 - 24 °C for the Sturm test.
- Aeration of dilution water: Yes, via peristaltic pump


TEST SYSTEM
- Number of culture flasks/concentration: Performed in duplicate
- Method used to create aerobic conditions: Perstitalic pump and magentic stirrer
- Measuring equipment: CO2 absorbers containing 100 mL Ba(OH)2
- Test performed in closed vessels due to significant volatility of test substance: No
- Test performed in open system: Not reported
- Details of trap for CO2 and volatile organics if used: CO2 absorbers containing 100 mL Ba(OH)2

SAMPLING
- Sampling frequency: Days 1, 3, 5, 7, 9,12, 14, 16, 19 and 21
- Sampling method: Not reported
- Sample storage before analysis: Not reported

CONTROL AND BLANK SYSTEM
- Other: Theoretical amount of initial CO2 was calculated as 58 mg

STATISTICAL METHODS: Calculated based on the average of two experiments
Reference substance:
not specified
Parameter:
% degradation (CO2 evolution)
Value:
2
Sampling time:
1 d
Parameter:
% degradation (CO2 evolution)
Value:
9
Sampling time:
3 d
Parameter:
% degradation (CO2 evolution)
Value:
24
Sampling time:
5 d
Parameter:
% degradation (CO2 evolution)
Value:
36
Sampling time:
7 d
Parameter:
% degradation (CO2 evolution)
Value:
53
Sampling time:
9 d
Parameter:
% degradation (CO2 evolution)
Value:
69
Sampling time:
12 d
Parameter:
% degradation (CO2 evolution)
Value:
76
Sampling time:
14 d
Parameter:
% degradation (CO2 evolution)
Value:
83
Sampling time:
16 d
Parameter:
% degradation (CO2 evolution)
Value:
90
Sampling time:
19 d
Parameter:
% degradation (CO2 evolution)
Value:
95
Sampling time:
21 d
Details on results:
The results obtained in the biodegradability determination of stearic acid show that this compound is degradable after an initial lag phase, reaching the “pass level” of biodegradation, corresponding to 60%, within 10 days.

Table 1: Results observed in the Sturm Test with Stearic Acida

 Days  CO2 produced (mg)  % Biodegradability
 1  1  2
 3  5  9
 5  14  24
 7  21  36
 9  31  53
 12  40  69
 14  44  76
 16  48  83
 19  52  90
 21  55  95

a The theoretical amount of initial CO2 was 58 mg. Indicated values are means of two experiments.

Validity criteria fulfilled:
not specified
Interpretation of results:
readily biodegradable
Conclusions:
The test item was shown to have degraded by 95% at day 21 in the Modified Sturm test.
Executive summary:

The biodegradability of the test item was assessed in a modified Sturm test conducted over 21 days. Biodegradation was assessed based on CO2 production. The test item is degradable after an initial lag phase, reaching the “pass level” of biodegradation, corresponding to 60%, within 10 days and after 21 days the test item showed 95% biodegradation.

The publication is a near guideline study suitable for assessment of this endpoint, with some restrictions (Klimisch 2).

Description of key information

Biodegradation data are not available for triglycerides, C16 and C18 (unsaturated) but data has been read across from fatty acids with the same chain lengths. The fatty acids stearic acid and palmitic acid are readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:
readily biodegradable
Type of water:
freshwater

Additional information

In a modified Sturm test conducted over 21 days, with biodegradation assessed on the basis of CO2 production, it was observed that stearic acid is biodegradable after an initial lag phase, reaching the “pass level” of biodegradation, corresponding to 60%, within 10 days. After 21 days the test item showed 95% biodegradation (Ruffo et al. 1983). Biodegradation of stearic acid was also assessed using a modified version of the Blok method (Blok 1979) conducted over 28 days with biodegradation assessed based on theoretical O2 demand. After 28 days it was determined that the test item had degraded 79% from the starting concentration (Gerike 1984). The biodegradability of palmitic acid was also assessed in Gerike (1984), following the same method as for stearic acid, and after 28 days the test item had degraded 84% from the starting concentration, and therefore is readily biodegradable.

These studies are published in peer-reviewed journals and are well documented, meeting generally accepted scientific principles, and therefore are considered to be suitable for read-across purposes.