N,N'-bis(2,4,6-triisopropylphenyl)methanediimine

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Data waiving:

study scientifically not necessary / other information available

Justification for data waiving:

the study does not need to be conducted because the substance is highly insoluble in water, hence indicating that aquatic toxicity is unlikely to occur

Justification for type of information:

JUSTIFICATION FOR DATA WAIVING
According to REACH Annex VII column 1, 9.1.1. Short-term toxicity testing on invertebrates (preferred species Daphnia) The registrant may consider long-term toxicity testing instead of short-term. According to column 2, 9.1.1. The study does not need to be conducted if:
— there are mitigating factors indicating that aquatic toxicity is unlikely to occur, for instance if the substance is highly insoluble in water or the substance is unlikely to cross biological membranes, or
— a long-term aquatic toxicity study on invertebrates is available, or
— adequate information for environmental classification and labelling is available.
The long-term aquatic toxicity study on Daphnia (Annex IX, section 9.1.5) shall be considered if the substance is poorly water soluble.

In general, there is no legal limit value available defining a substance as poorly water soluble or insoluble under REACH. However, in ECHA’s Guidance document Chapter R.7b: Endpoint specific guidance Version 3.0 – February 2016, it is stated: „poorly water soluble substances (e.g. water solubility below 1 mg/L or below the detection limit of the analytical method of the test substance)“. Further, poorly soluble substances are defined by OECD (2000 OECD SERIES ON TESTING AND ASSESSMENT, Number 23, GUIDANCE DOCUMENT ON AQUATIC TOXICITY TESTING OF DIFFICULT SUBSTANCES AND MIXTURES, ENV/JM/MONO(2000)6) as substances with a limit of solubility <100 mg/l although technical problems are more likely to occur at <1mg/l as defined in TGD (1996). Very low water solubility (i.e. in the low μg/l range) could be used as a reason to significantly modify a standard test or to test non-pelagic organisms preferentially.
ECHA’s Guidance document Chapter R.7c: Endpoint specific guidance Version 3.0 – June 2017 further says: „As indicated in the OECD TG 305, for strongly hydrophobic substances (log Kow > 5 and a water solubility below ~ 0.01-0.1 mg/L), testing via aqueous exposure may become increasingly difficult. However, an aqueous exposure test is preferred for substances that have a high log Kow but still appreciable water solubility with respect to the sensitivity of available analytical techniques, and for which the maintenance of the aqueous concentration as well as the analysis of these concentrations do not pose any constraints.

In the available water solubility study according to OECD 105 (shake flask method), in the first run, no reproducible results could have been obtained, so a second run was performed. Therein, approx. 93.2 mg/L test item were weighed into three flask which were stirred for 48, 72 or 96 hours. The water solubility of Bis(2,4,6-triisopropylphenyl)carbodiimide was determined to be 2.3 µg/L after 48 h shaking, and it was determined to be below the LOQ of 1.7 µg/L after 72 h resp. 96 h shaking. This fact and visualisation of the chromatograms indicate a time-dependent decrease in substance concentration. This first shows that the test item is practically insoluble in water, and second, that it is not stable over time in water.
Although the observed water solubility values are already in the range of trace analysis, attempts were made to lower the LOQ. However, the determined LOQ was already associated with a certain standard deviation. The target concentration of 1.638 µg/L resulted in measured values of 1.240 resp. 1.412 µg/L. Further dilutions of this concentration resulted in the following findings:

Dilution Target conc. [µg/L] Measured conc. [µg/L]
1:1000 0.002 0.662
0.659
1:100 0.016 0.714
0.718
1:10 0.164 1.042
1.013

As obvious, already the 1:10 dilution resulted in a deviation of one magnitude, clearly showing that the measured concentrations are not reliable any more. So the lowest achievable calibration standard was determined to be 1.638 μg/L as it was the lowest concentration resulting in acceptable deviations of the measured value from the target one.
So no analytical method with appropriate sensitivity is available or can be developed, because longer stirring times or exposure times as required for both short- and long-term aquatic toxicity would result in a decrease in the analyte concentration, which is below the possibly achievable LOQ.

Based on the available information it can be stated that, as outlined in the first waiving possibility under REACH, the substance is highly insoluble in water, and hence, aquatic toxicity is unlikely to occur. So short-term toxicity testing does not need to be conducted. Long-term testing does also not need to be conducted, as very low water solubility can be assumed if the substance is soluble only in the low μg/l range, and the actual water solubility after a certain stirring / exposure time is even below this range. Further, the registrant demonstrated that the analysis of concentrations in water poses constraints, the sensitivity of available analytical techniques is not sufficient to analyse concentrations in the required water solubility range.

In consequence, testing can be omitted.

Cross-reference

Reason / purpose for cross-reference:

data waiving: supporting information

Remarks:

water solubility

Reference

Endpoint:

water solubility

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2018-05-08 - 2018-11-12

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

under GLP

Qualifier:

according to guideline

Guideline:

EPA OPPTS 830.7840 (Water Solubility)

Version / remarks:

United States Environmental Protection Agency, OPPTS 830.7840 - Water Solubility: Column Elution Method; Shake Flask Method, 1988.

Deviations:

yes

Remarks:

In the second attempt, all test solutions were stirred completely at 20 °C without previously being stirred at 30 °C.

Qualifier:

according to guideline

Guideline:

OECD Guideline 105 (Water Solubility)

Version / remarks:

OECD - Organisation for Economic Co-operation and Development, OECD Guideline 105: Water Solubility, 1995

Deviations:

yes

Remarks:

In the second attempt, all test solutions were stirred completely at 20 °C without previously being stirred at 30 °C.

GLP compliance:

yes (incl. QA statement)

Type of method:

flask method

Water solubility:

2.3 µg/L

Conc. based on:

test mat.

Loading of aqueous phase:

93 mg/L

Incubation duration:

48 h

Temp.:

20 °C

pH:

5

Water solubility:

< 1.7 µg/L

Conc. based on:

test mat.

Loading of aqueous phase:

94.5 mg/L

Incubation duration:

72 h

Temp.:

20 °C

pH:

5

Remarks on result:

other: It was determined to be below the LOQ of 1.7 μg/L

Water solubility:

< 1.7 µg/L

Conc. based on:

test mat.

Loading of aqueous phase:

92 mg/L

Incubation duration:

96 h

Temp.:

20 °C

pH:

5

Remarks on result:

other: It was determined to be below the LOQ of 1.7 μg/L

Conclusions:

The water solubility of the test item Bis(2,4,6-triisopropylphenyl)carbodiimide was determined according to OECD 105, using the flask method, under GLP with acceptable modifications, which may be considered as a scientifically reasonable approach. Hence, the observations and results can be considered reliable.
The water solubility of Bis(2,4,6-triisopropylphenyl)carbodiimide was determined to be 2.3 µg/L after 48 h shaking, and it was determined to be below the LOQ of 1.7 µg/L after 72 h resp. 96 h shaking. This fact and visualisation of the chromatograms indicate a time-dependent decrease in substance concentration. This first shows that the test item is practically insoluble in water, and second, that it is not stable over time in water.

Executive summary:

The water solubility of the test item Bis(2,4,6-triisopropylphenyl)carbodiimide in demineralized water was determined with the shake flask method at 20 °C according to OECD 105 and OPPTS 830.7840 under GLP. According to the results of a preliminary test the column elution method should be carried out but the test item is a liquid and therefore the flask method was applied.

The concentration of the test item in the aqueous test solutions was determined by HPLC analysis with MS detection.

In a first attempt the results showed a large deviation. Therefore a second attempt with deviations from OECD 105 was carried out. All test solutions were stirred completely at 20 °C without previously being stirred at 30 °C.

 

Table Results of individual determinations and mean value

Water solubility test at 20 °C	Flask A [µg/L]	Flask B [µg/L]	Flask C [µg/L]
Stirring time (at 20 °C)	96 h	72 h	48 h
Concentration of Bis(2,4,6-triisopropylphenyl)carbodiimide	< 1.7	< 1.7	2.3
pH-value	5	5	5

 

Water solubility of Bis(2,4,6-triisopropylphenyl)carbodiimide at 20 °C after shaking for 48 hours: 2.3 µg/L