N-(2-ethylhexyl)-8,9,10-trinorborn-5-ene-2,3-dicarboximide

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:

7 March 2002 to 26 June 2002

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 301 B (Ready Biodegradability: CO2 Evolution Test)

Qualifier:

according to guideline

Guideline:

EPA OPPTS 835.3110 (Ready Biodegradability)

Version / remarks:

January 1998

GLP compliance:

yes (incl. QA statement)

Specific details on test material used for the study:

Identity: MGK 264
Lot number: 7437
Expiry date: 22 January 2003
Purity: 94.5%

Oxygen conditions:

aerobic

Remarks:

Test, control and reference mixtures were aerated for 29 days with air that had been treated to remove carbon dioxide (CO2)

Inoculum or test system:

sewage, predominantly domestic (adaptation not specified)

Details on inoculum:

A sample of activated sludge was collected from Oakley sewage treatment works, which treats predominantly domestic waste. Aliquots (25 ml) of a homogenised sample were filtered through dried (approximately 105°C) and pre-weighed Whatman GFC filter papers. The filters were dried for at least one hour, allowed to cool and re-weighed. The solids level in the sludge was determined and then an appropriate volume used to inoculate control and test vessels to give a final suspended solids concentration of 30 mg/l.

Duration of test (contact time):

29 d

Initial conc.:

10 other: mgCarbon/l (mgC/l)

Based on:

other: Carbon Content of MGK264

Parameter followed for biodegradation estimation:

CO2 evolution

Details on study design:

Air-saturated ultrapure water was added to each of six, five litre amber glass culture bottles followed by the volumes of each of the stock solutions required to prepare three litres of mineral salts medium (MSM, Appendix 1 ). Each culture bottle was then inoculated with activated sludge (30 mg solids/I)
and a magnetic stirring bar was added. The bottles were stoppered, their contents were mixed by swirling then each was placed on an electrically operated magnetic stirrer. The mixtures were aerated overnight with a supply of air that had been treated to remove carbon dioxide by passing it through cylinders containing fused calcium chloride, Carbosorb AS and a trap containing 1M sodium hydroxide solution.

The reference substance sodium benzoate was added as an aqueous stock solution (1.72 g/l) to one bottle containing the test substance and to one containing inoculated mineral salts medium alone. Ultrapure water (200 ml) was also added to the control cultures on Day 0 in order to ensure similarity in volume additions to those containing the test substance. Since identical volumes were added to test and control cultures on Day 0 of the test, the precision of the assay was not affected by the addition of the suspension containing the test substance to the test cultures. Therefore, it was not necessary to measure the total inorganic and organic carbon content of the test substance suspension, as the inclusion of similar traces of inorganic carbon would have occurred in the test and the blank control cultures.

The mixtures prepared for the test are summarised in the following table:
Bottle No.- Contents
1, 2 - Controls - mineral salts medium plus inoculum (30 mg solids/I)
3 - Reference - inoculated mineral salts medium plus sodium benzoate ( 10 mgC/l)
4,5 - Test substance (10 mgC/l) plus inoculated mineral salts medium
6 - Sodium benzoate (10 mgC/l) plus test substance (10 mgC/l) plus inoculated mineral salts medium

The pH of each culture was determined in situ. Each vessel was then fitted with a stopper holding an air inlet tube reaching approximately 10 cm below the liquid surface and an air outlet just below the stopper.
The vessels contents were continuously flushed for 29 days with treated air. The air outlet from each vessel was connected to three Dreschel bottles in series, each containing 0.025N, nominal barium hydroxide (100 ml).
The biodegradation of the reference substance in the mixture containing the test and reference substance was calculated to confirm that the test substance was not inhibitory to the activity of the microbial inoculum. When the level of biodegradation of sodium benzoate achieved the pass level for ready biodegradability(≥ 60%), the pH was measured and the treatment was terminated.

Reference substance:

benzoic acid, sodium salt

Preliminary study:

N/A

Test performance:

Cumulative CO2 production in the controls (80.3 and 78.7 mgCO2) was within the acceptable range for this assay system (recommended maximum for a three litre culture = 120 mgC02). The degradation of sodium benzoate was rapid and had achieved 67% of its TCO2 after 7 days and 84% after 29 days.
The degradation of sodium benzoate was also rapid in the presence of MGK 264 and had achieved 61 % of its TCO2 after 8 days. This test vessel was terminated on Day 8 of the study; the final pH of the culture was 7.5.

Key result

Parameter:

% degradation (CO2 evolution)

Value:

3

Sampling time:

29 d

Remarks on result:

other: MGK 264 cannot be considered to be readily degradable.

Details on results:

The blank-corrected C02 production and percentage degradation in test and reference mixtures at intervals during the test are given in Table 1. Degradation of the test and reference substances is illustrated graphically in Figure 1.

Cumulative CO2 production in the controls (80.3 and 78.7 mgCO2) was within the acceptable range for this assay system (recommended maximum for a three litre culture = 120 mgC02). The degradation of sodium benzoate was rapid and had achieved 67% of its TCO2 after 7 days and 84% after 29 days.

The degradation of sodium benzoate was also rapid in the presence of MGK 264 and had achieved 61 % of its TCO2 after 8 days. This test vessel was terminated on Day 8 of the study; the final pH of the culture was 7.5.

Mean cumulative CO2 production by the mixtures containing MGK 264 was negligible and had achieved, at most, 3% of the TCO2 by the end of the test on Day 29. The pH of all test and control mixtures was 7.6 at the start of the test and ranged from 7.4 to 7.6 at the end.

The typical rate of air flow during the test ranged from 30 to 90 mL/minute. Levels lower than 30 mL/minute were recorded in single vessels on four occasions during the test. The temperature of a 3L volume of water held under test conditions ranged from 20.8°C to 22.3°C over the test period.

Validity criteria fulfilled:

yes

Interpretation of results:

not readily biodegradable

Conclusions:

The biodegradation of sodium benzoate in the presence of the test substance was monitored in order to assess whether any inhibitory effects were exerted on the activity of the microbial inoculum. None were observed.

The mean cumulative CO2 production by mixtures containing MGK 264 Insecticide Synergist at 10 mgC/l was negligible and had achieved, at most, 3% of the TC02 by the end of the test on Day 29.
Substances are considered to be readily biodegradable in this test if CO2 production is equal to or greater than 60% of the theoretical value within ten days of the level achieving 10%. MGK 264 Insecticide Synergist cannot, therefore, be considered to be readily degradable.

Because of the stringency of this type of test, poorly soluble substances that fail to show the required rate of biodegradation are not necessarily poorly degradable under environmental conditions. Biodegradability may progress at a higher rate where the substance, at a concentration within its aqueous solubility, is exposed to a larger and more diverse microbial population.

The results obtained for the degradation of sodium benzoate ( 67% of its TCO2 after 7 days and 84% after 29 days) and for cumulative CO2 production by the control mixtures (80.3 and 78.7 mgCO2) fulfil the validity criteria for this test.

Executive summary:

The ready biodegradability of MGK 264 Insecticide Synergist was assessed in the CO2 Evolution test (Modified Sturm test, Procedure C.4-C of the Annex to Directive 92/69/EEC; OECD Procedure 301B; US Environmental Protection Agency (EPA), Office of Prevention, Pesticides and Toxic Substances, (OPPTS) Method 835.3110 (m) Carbon Dioxide Evolution test, adopted January 1998).

 

MGK 264 Insecticide Synergist was added to two vessels containing mineral salts medium inoculated with activated sludge (30 mg solids/I) to give a nominal test concentration of 10 mgCarbon[C]/l. Two control vessels contained inoculated mineral salts medium alone, and one contained inoculated mineral salts medium plus the reference substance sodium benzoate (10 mgC/l).

 

An additional mixture containing sodium benzoate (10 mgC/l) and MGK 264 Insecticide Synergist (10 mgC/l) was established in order to assess the potential inhibitory effects of the test substance on the activity of the microbial inoculum.

 

Test, control and reference mixtures were aerated for 29 days with air that had been treated to remove carbon dioxide (CO2).The CO2 produced by each culture was trapped in a series of Dreschel bottles containing barium hydroxide, which were connected to the outlet from each test vessel. The residual barium hydroxide was determined at intervals by titration.

 

The pH of control, reference and test mixtures was measured at the start of the test and after 28 days. The pH of the test plus reference mixture was measured at the start of the test and on the day of its termination (Day 8).

 

Sodium benzoate had been biodegraded by 67% after 7 days and 84% after 29 days in the absence of MGK 264 Insecticide Synergist, and by 61 % after 8 days in its presence which confirmed that MGK 264 Insecticide Synergist was not inhibitory to the activity of the microbial inoculum. Cumulative levels of CO2production in the controls after 29 days (80.3 and 78.7 mgCO2)were within the acceptable range for this assay system (recommended maximum=120 mgCO2for a three-litre culture). These results confirm that the inoculum was viable and that the test was valid.

 

Mean cumulative CO2 production by mixtures containing MGK 264 Insecticide Synergist was negligible and had achieved, at most, 3% of the theoretical value (TCO2,110.1 mg CO2) by the end of the test on Day 29.

 

Substances are considered to be readily biodegradable in this test if CO2 production is equal to or greater than 60% of the theoretical value within 10 days of the level achieving 10%. MGK 264 Insecticide Synergist cannot, therefore, be considered to be readily biodegradable.

Description of key information

The ready biodegradability of MGK 264 was assessed in the CO₂ Evolution test (Modified Sturm test, Procedure C.4 -C of the Annex to Directive 92/69/EEC; OECD Procedure 301B; US Environmental Protection Agency (EPA), Office of Prevention, Pesticides and Toxic Substances, (OPPTS) Method 835.3110 (m) Carbon Dioxide Evolution test, adopted January 1998).

 

Mean cumulative CO₂ production by mixtures containing MGK 264 was negligible and had achieved, at most, 3% of the theoretical value (TCO₂,110.1 mg CO₂) by the end of the test on Day 29.

 

Substances are considered to be readily biodegradable in this test if CO₂ production is equal to or greater than 60% of the theoretical value within 10 days of the level achieving 10%. MGK 264 cannot, therefore, be considered to be readily biodegradable.

Key value for chemical safety assessment

Biodegradation in water:

under test conditions no biodegradation observed