Androsta-1,4-diene-17-carbothioic acid, 6,9-difluoro-11-hydroxy-16-methyl-3-oxo-17-(1-oxopropoxy)-, (6.alpha.,11.beta.,16.alpha.,17.alpha.)-

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:

15th May 2003 - 30th April 2004

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

other: ISO 14593

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: OPPTS 835.3120

Deviations:

no

GLP compliance:

yes

Specific details on test material used for the study:

Test substance name: Thio Acid Propionate (CCI22363)
Chemical composition: 100%
Empirical formula: C24H30F2O5S
Appearance: White/ off white cystalline solid
Batch number: H030714
Receipt date: 26 April 2003
Storage Conditions: Room temperature (10 to 30°C)

Oxygen conditions:

aerobic

Inoculum or test system:

activated sludge, domestic (adaptation not specified)

Details on inoculum:

The activated sludge inoculum was not deliberately acclimatised or adapted to the test substance before exposure to the test substance before expoisure to the test substance in this study.

Duration of test (contact time):

ca. 28 d

Initial conc.:

1.74 mg/L

Based on:

IC (inorganic carbon)

Parameter followed for biodegradation estimation:

inorg. C analysis

Details on study design:

Sample Preparation:
At each analysis point, duplicate vessels from each treatment group were removed from the orbital shaker. Sodium hydroxide solution (1mL at nominally 7M) was injected into each vessel through the butyl rubber stopper. The vessels were then returned to the orbital shaker and shaken for a period of at least one hour. The purpose of rendering the cultured alkaline in this way, was to fix all of the inorganic carbon present in the vessels into the liquid phase. The vessels were then removed from the shaker and allowed to stand to allow any sediment to settle. Samples were then removed from the liquid phase of each vessel for IC analysis.

Reference substance:

benzoic acid, sodium salt

Test performance:

The mean inorganic carbon yield attributable to Thio Acid Propionate (CCI22363) was less than 1% of the initial organic carbon loading throughout the time course of this study. Although Thio Acid propionate (CCI22363) did not undergo significant mineralisation under the conditions employed in this study, this is not necessarily an indication that Thio Acid Propionate (CCI22363) would persist in the environment.

Key result

Parameter:

% degradation (inorg. C analysis)

Value:

< 1

Sampling time:

3 d

Key result

Parameter:

% degradation (inorg. C analysis)

Value:

< 1

Sampling time:

7 d

Key result

Parameter:

% degradation (inorg. C analysis)

Value:

< 1

Sampling time:

14 d

Key result

Parameter:

% degradation (inorg. C analysis)

Value:

< 1

Sampling time:

21 d

Key result

Parameter:

% degradation (inorg. C analysis)

Value:

ca. 0

Sampling time:

28 d

Details on results:

The ISO standard states that degradation of the reference substance should be at least 60% on the 14th day of incubation. Degradation of sodium benzoate was 90% on Day 14, thus satisfying this requirement, therefore the results of this study may be considered valid.

Inorganic Carbon Analytical Data:

	Level of Inorganic Carbon (ppm) at Time (days)
	3	7	14	21	28
Control #1	1.487	1.749	1.909	2.590	2.618
Control #2	1.565	1.875	1.808	2.674	2.499
Mean Control	1.526	1.812	1.858	2.632	2.558
	Level of Inorganic Carbon (ppm) at Time (days)
	3	7	14	21	28
Test #1	0	0.043	0.152	0	0
Test #2	0.041	0.071	0.080	0.045	0
Reference #1	9.632	11.688	13.431	13.240	13.939
Reference #2	9.325	12.152	13.849	13.210	13.992

Validity criteria fulfilled:

yes

Interpretation of results:

under test conditions no biodegradation observed

Conclusions:

The mean inorganic carbon yield attributable to Thio Acid Propionate (CCI22363) was less than 1% of the initial organic carbon loading throughout the time course of this study. Although Thio Acid Propionate (CCI22363) did not undergo significant mineralisation under the conditions employed in this study, this is not necessarily an indication that Thio Acid Propionate (CCI22363) would persist or accumulate in the environment.

Executive summary:

The ultimate biodegradability of Thio Acid Propionate (CCI22363) was assessed by measuring the yield of inorganic carbon recovered under standard test conditions. The methods used were as described in ISO Standard 14593: Water quality - Evaluation of ultimate aerobic biodegradability of organic compounds in aqueous medium - Method by analysis of inorganic carbon in sealed vessels (CO₂headspace test), and is essentially the same as that described in US EPA Guideline OPPTS 835.3120.

The mean inorganic carbon yield attributable to Thio Acid Propionate (CCI22363) was less than 1% of the intial organic carbon loading throught the time course of this study. Although Thio Acid Propionate (CCI22363) did not undergo significant minerlisation under the conditions employed in this study, this is not necessarily an indication that Thio Acid Propionate (CCI22363) would persist or accumulate in the environment. The mean inorganic carbon yield attributable to the reference substance, sodium benzoate, exceeded 90% of the initial organic carbon loading on Days 14 and 28. These data satisfy the validity criteria stated in the ISO standard, and show that the microbial inoculum employed was exerting normal degradative activity.

Description of key information

Test vessels were filled with inoculated mineral salts medium, with or without the test refernce substances, to provide a liquid-to-headspace ratio of 2:1. The vessels were sealed and incubated with shaking for 28 days in darkness. At intervals replicate vessels from each treatment group were rendered alkaline to fix inorganic carbonates and to absorb CO₂from the headspace into the medium. The inorganic carbon yield in each vessel was then determined by carbon analysis.

Inorganic carbon (IC) yields for vessels in the test and reference groups were corrected for the IC content of control vessels containing unamended mineral medium. The extent of biodegradation was calculated by expressing the blank corrected IC yield in test vessels as a percentage of the organic carbon loading from the test substance added at the start of the test. The procedure and the activity of the inoculum were checked by a set of test vessels containing the readily degradable reference substance sodium benzoate.

The study was initialted (protocol issued) on 15 May 2003, and completed (final report issued) on 30 April 2004. It was conducted in the Ecotoxicological Section of the Department of Environmental Sciences, Covance, Harrogate, North Yorkshire, England. The test subtance was dosed to the test system on 28 August 2003, and the practical phase was completed on 16 October 2003.

Test Medium:

The test was conducted in an aqueous synthetic mineral salts medium. On Day 0 of the test, test medium was prepared so that the final concentration established in test systems would be equivalent to 10mL/L solution (a) and 1mL/L of each of solutions (b), (c) and (d). Solutions (a) to (d) were nominally composed as follows:

(a):8.50g potassium dihydrogen phophate (Fisher); 21.75g dipotassium hydrogen phosphate (BDH); 33.4g disodium hydrogen phosphate dihydrate (Fisher); 0.50g ammonium chloride (Sigma), all dissolved in and made up to 1L with reverse-osmosis water.

(b):36.4g calcium dissolved in and made up to 1L with reverse-osmosis water.

(c):22.5g magnesium sulphate heptahydrate (Fisher), dissolved in and made up to 1L with reverse-osmosis water.

(d):0.25g ferric chloride hexahydrate (Aldrich) plus one drop concentrated hydrochloric acid (BDH); dissolved in and made up to 1L with reverse-osmosis water.

On the basis of the suspended solids determination described above, the medium was inoculated with activated sludge to provide a nominal final solids concentration of 4mg/L.

The medium was dispensed into "125"mL glass serum bottles, which have an approximate total volume of 160mL. Aliquots of 100ml were initially dispensed into each bottle. The remaining 7mL to be added comprised reference substance stock solution and/or reverse-osmosis water, depending on the treatment group.

Dosing the Reference Substance:

Based on the molecular formula of sodium benzoate, its carbon content expressed as a percentage by weight is 58.3%. This value was used in setting the concentration of sodium benzoate to be applied in this study. A level of 15mgC/L is achieved from a stock sodium benzoate concentration of 0.5506g/L dosed at a dilution of 5mL in 107mL. The stock was prepared by dissolving 55.1mg of sodium benzoate in 100mL of reverse-osmosis water, and this stock was added (5ml) to vessels in that refernce group, to give a nominal reference substance concentration corresponding to 15mgC/L.

Dosing the Test Substance:

The test substance was sufficiently soluble to permit dosing of the test system with a concentration aqueous stock solution. The test substance was therefore added directly to vessels in the test group. Based on the molecular formula of the test substance provided by the Sponsor, the carbon content of Thio Acid Propionate (CCI22363) [1.74mg +/- 0.03mg] was added to vessels in the test group to achieve a final nominal concentration of 10mgC/L.

Incubation Conditions:

The incubation and test measurements were conducted in a facility where the temperature was maintained at a nominal 21°C. Temperatures recorded during the study fell between 19.8°C and 20.8°C, and were essentially comliant with the test Guideline requirement that the termperature be maintained in the range 20 to 25°C, with the overall variation not exceeding +/- 2°C.

Test substance name: Thio Acid Propionate (CCI22363)

Chemical composition: 100%

Empirical formula: C24H30F2O5S

Appearance: White/ off white cystalline solid

Batch number: H030714 Receipt date: 26 April 2003

Storage Conditions: Room temperature (10 to 30°C)

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed

Additional information

The ultimate biodegradability of Thio Acid Propionate (CCI22363) was assessed by measuring the yield of inorganic carbon recovered under standard test conditions. The methods used were as described in ISO Standard 14593: Water quality - Evaluation of ultimate aerobic biodegradability of organic compounds in aqueous medium - Method by analysis of inorganic carbon in sealed vessels (CO₂headspace test), and is essentially the same as that described in US EPA Guideline OPPTS 835.3120.

The mean inorganic carbon yield attributable to Thio Acid Propionate (CCI22363) was less than 1% of the intial organic carbon loading throught the time course of this study. Although Thio Acid Propionate (CCI22363) did not undergo significant minerlisation under the conditions employed in this study, this is not necessarily an indication that Thio Acid Propionate (CCI22363) would persist or accumulate in the environment. The mean inorganic carbon yield attributable to the reference substance, sodium benzoate, exceeded 90% of the initial organic carbon loading on Days 14 and 28. These data satisfy the validity criteria stated in the ISO standard, and show that the microbial inoculum employed was exerting normal degradative activity.