Reaction products of disubstituted alkane, nickel, (Z/E)-substituted alkene, reaction mass of tetraalkyl-3,3’-di(propan-2-yl)-biphenyl-diyl tetrakis(dialkylphenyl) bis(phosphite) and tris(dialkylphenyl) phosphite and zinc halide

Short-term toxicity to fish

Link to relevant study record(s)

Reference

Endpoint:

short-term toxicity to fish

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2018-10-03 to 2018-11-27

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 203 (Fish, Acute Toxicity Test)

Version / remarks:

adopted July 1992

Deviations:

no

Qualifier:

according to guideline

Guideline:

other: OECD 23, Guidance document on aquatic toxicity testing of difficult substances and mixtures

Version / remarks:

2000

Deviations:

not applicable

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

Frequency:
At the start (t=0 h), after 24 h and the end of the test (t= 96 h).
While taking samples for analysis, care was taken not to include any floating layer, test item film or undissolved material in separate vessels.

Number of samples:
Sampling consisted of single samples per treatment.
Volume:
2 mL per sample;
Storage:
Samples were stored in a freezer (=-15°C) until analysis.

Vehicle:

no

Details on test solutions:

Water Accommodated Fractions were prepared:
Individual loading rates were vigorously agitated on a magnetic stirrer for 6 days. On day 3 of stirring, pH was measured in case adjustment would be necessary (no adjustment was necessary:
pH of 8.0 ± 0.3). After 6 days of stirring, the obtained mixtures were allowed to settle for 1 day in order to allow separation of phases. Thereafter, the aqueous Water Accommodated
Fractions (WAFs) were collected by means of siphoning over glasswool (put into a funnel). Glasswool filtering was necessary to avoid a positive Tyndall effect. Glasswool was pre-saturated
with the test item beforehand. No Tyndall effect was observed in any prepared solution, and solutions were clear and colourless at the end of the preparation procedure.
Thereafter, the aqueous WAFs were used as test concentrations.
Based on preliminary results, the period of 6 days of stirring was used to ensure that
a) Concentration of the relevant transition metal stabilizes at the pH 8.0 ± 0.3 used for ecotoxicity testing to prevent any precipitation during the test;
b) Saturation of the aquatic phase with organic hydrolysis products stemming from aromatic phosphites is obtained.

Test organisms (species):

Cyprinus carpio

Details on test organisms:

Species Carp (Cyprinus carpio, Teleostei Cyprinidae) Linnaeus, 1758.
Source: Wageningen University and Research Centre, the Netherlands.
Fish length and weight: A representative number (10) of the batch of fish, from which fish were used for the present study, were weighed and measured prior to the start of the test.
Mean length of fish 3.0 ± 1.0 cm
Characteristics: F1 from a single parent-pair bred in UV-treated water. Healthy fish supplied with a health certificate.
Reason for selection: The system has been selected as an internationally accepted species.

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

96 h

Hardness:

180 mg/L expressed as CaCO3

Test temperature:

22 °C, constant within 2°C

pH:

8.0 ± 0.3 at the start of the test

Dissolved oxygen:

> 60% of air saturation value

Salinity:

Not applicable

Conductivity:

See IUCLID section "Any other information on results incl. tables" below.

Nominal and measured concentrations:

For the initial range-finding and the actual limit test, the following water accommodated fractions (WAF, based on nominal loading rate) were individually prepared:
1.0 mg/L; 10 mg/L; 100 mg/L
Please see IUCLID section "Any other information on results incl. tables" below for information on analytical results.

Details on test conditions:

Please see IUCLID section "Any other information on materials and methods incl. tables" below for details.

Reference substance (positive control):

no

Key result

Duration:

96 h

Dose descriptor:

LL0

Remarks:

Based on Water Accommodated Fraction (WAF) approach

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks:

and sub-lethal effects

Duration:

96 h

Dose descriptor:

LL10

Remarks:

Based on Water Accommodated Fraction (WAF) approach

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks:

and sub-lethal effects

Duration:

96 h

Dose descriptor:

LL50

Remarks:

Based on Water Accommodated Fraction (WAF) approach

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality (fish)

Remarks:

and sub-lethal effects

Details on results:

See IUCLID section "Any other information on results incl. tables" for details.

Results with reference substance (positive control):

Not applicable

Reported statistics and error estimates:

Not applicable - no effects observed.

Sublethal observations / clinical signs:

These data are based on results from draft report and will be amended as soon as the final report will have been issued.

Details on Biological Data:

No mortality or any other sub-lethal symptoms were observed in the control and at any test concentrations of the range finding as well as the actual limit test (see table below).

Step 2 Catalyst (Stripped) Loading Rate  (mg/L)	Initial number of fish	Cumulative mortality					Total Mortality (%)
		2¼h	24h	48h	72h	96h
1.0 (range finding)	3	0	0	0	0	0	0
10 (range finding)	3	0	0	0	0	0	0
100 (range finding)	3	0	0	0	0	0	0
100 (limit test)	7	0	0	0	0	0	0
Control (limit test)	7	0	0	0	0	0	0

Details on Analytical Data:

At this time, analytical data are available for the range-finding test, only. Data on the final limit test will be amended as soon as they will become available.

1) Analysis of the relevant transition metal by ICP-MS

Time of sampling hours	Date of sampling	Date of analysis	Loading rate (WAF) [mg/L]	Concentration test item analysed [mg/L]	Relative to initial [%]	Transition metal concentration calculated from elemental fraction (0.8%) determined in study number 519586 [mg/L]
0	15 Oct 2018	24 Oct 2018	100	20.3		0.162
24	16 Oct 2018	24 Oct 2018	100	20.9	103	0.167
96	19 Oct 2018	24 Oct 2018	100	20.9	103	0.167

2) Analysis of the organics fraction by LC-UV at 210 nm

Time of sampling [hours]	Date of sampling	Date ofanalysis	Loading rate WAF [mg/L]	Concentration analysed [mg/L]	Relative to initial[%]
0	15 Oct 2018	31 Oct 2018	100	142	0
24	16 Oct 2018	31 Oct 2018	100	176	124
96	19 Oct 2018	31 Oct 2018	100	208	146

Validity criteria fulfilled:

yes

Conclusions:

The acute toxicity to freshwater fish was tested according to OECD 203 compliant with GLP. Due to the nature of Step 2 catalyst (UVCB), according to OECD 23 water accommodated fractions (WAF) were prepared.
Result:
LL0 (Cyprinus carpio; 96 h; static) = 100 mg/L;
LL10 (Cyprinus carpio; 96 h; static) > 100 mg/L
LL50 (Cyprinus carpio; 96 h; static) > 100 mg/L

Executive summary:

The acute toxicity to freshwater fish was tested according to OECD 203 compliant with GLP. The test organism was Cyprinus carpio (carp). Due to the nature of Step 2 catalyst (UVCB), according to OECD 23 water accommodated fractions (WAF) were prepared and based on the results of a range finding tests (neither mortalities nor any sublethal effects observed up to WAF 100 mg/L) a limit test was conducted. WAFs were prepared at 1, 10 and 100 mg/L nominal loading (range finding) and 100 mg/L nominal loading (limit test): individual loading rates were vigorously agitated on a magnetic stirrer for 6 days. Based on preliminary results, the period of 6 days of stirring was used to ensure that

a) Concentration of the relevant transition metal stabilizes at the pH 8.0 ± 0.3 used for ecotoxicity testing to prevent any precipitation during the test;

b) Saturation of the aquatic phase with organic hydrolysis products stemming from aromatic phosphites is obtained.

The presence of any non-dissolved test item fraction could be excluded before testing by measuring the Tyndall effect.

Analytical monitoring was performed for the relevant transition metal (ICP-MS) as well as the organic fraction (UPLC-UV at 210 nm). At the limit concentration of WAF 100 mg/L transition metal concentration was stable over the 96 hours of the test. Step 2 catalyst concentration equivalent of 20.3 mg/L was found in freshly prepared WAF (after 6 days of stirring and one day settling period) and 103% of this initial concentration was recovered after 24 and 96 hours (20.9 mg/L). This is equivalent to transition metal concentrations at 0, 24 and 96 hours of 162 µg/L, 167 and 167 µg/L, respectively.

With regard to the organics fraction monitored via UPLC-UV at 210 nm, an initial Step 2 catalyst concentration equivalent of 142 mg/L was measured at the limit concentration of WAF 100 mg/L. This increased during 96 hours of incubation by 46%. The reason for this increase is not clear. It could be due to further abiotic transformations (hydrolysis / oxidation) during the test. The results confirm in any case that WAF preparation succeeded in a relevant organic fraction present at the beginning and during the test.  

For all concentrations tested (including the limit concentration of WAF 100 mg/L) neither in control groups nor in treatment groups any mortality or any other effects could be observed.

Result:

LL0 (Cyprinus carpio; 96 h; static) = 100 mg/L;

LL10 (Cyprinus carpio; 96 h; static) > 100 mg/L

LL50 (Cyprinus carpio; 96 h; static) > 100 mg/L

Description of key information

The acute toxicity to freshwater fish was tested according to OECD 203 compliant with GLP. Due to the nature of Step 2 catalyst (UVCB), according to OECD 23 water accommodated fractions (WAF) were prepared.

Result:

LL0 (Cyprinus carpio; 96 h; static) = 100 mg/L;

LL10 (Cyprinus carpio; 96 h; static) > 100 mg/L

LL50 (Cyprinus carpio; 96 h; static) > 100 mg/L

Additional information

The acute toxicity to freshwater fish was tested according to OECD 203 compliant with GLP. The test organism was Cyprinus carpio (carp). Due to the nature of Step 2 catalyst (UVCB), according to OECD 23 water accommodated fractions (WAF) were prepared and based on the results of a range finding tests (neither mortalities nor any sublethal effects observed up to WAF 100 mg/L) a limit test was conducted. WAFs were prepared at 1, 10 and 100 mg/L nominal loading (range finding) and 100 mg/L nominal loading (limit test): individual loading rates were vigorously agitated on a magnetic stirrer for 6 days. Based on preliminary results, the period of 6 days of stirring was used to ensure that

a) Concentration of the relevant transition metal stabilizes at the pH 8.0 ± 0.3 used for ecotoxicity testing to prevent any precipitation during the test;

b) Saturation of the aquatic phase with organic hydrolysis products stemming from aromatic phosphites is obtained.

The presence of any non-dissolved test item fraction could be excluded before testing by measuring the Tyndall effect.

Analytical monitoring was performed for the relevant transition metal (ICP-MS) as well as the organic fraction (UPLC-UV at 210 nm). At the limit concentration of WAF 100 mg/L transition metal concentration was stable over the 96 hours of the test. Step 2 catalyst concentration equivalent of 20.3 mg/L was found in freshly prepared WAF (after 6 days of stirring and one day settling period) and 103% of this initial concentration was recovered after 24 and 96 hours (20.9 mg/L). This is equivalent to transition metal concentrations at 0, 24 and 96 hours of 162 µg/L, 167 and 167 µg/L, respectively.

With regard to the organics fraction monitored via UPLC-UV at 210 nm, an initial Step 2 catalyst concentration equivalent of 142 mg/L was measured at the limit concentration of WAF 100 mg/L. This increased during 96 hours of incubation by 46%. The reason for this increase is not clear. It could be due to further abiotic transformations (hydrolysis / oxidation) during the test. The results confirm in any case that WAF preparation succeeded in a relevant organic fraction present at the beginning and during the test.  

For all concentrations tested (including the limit concentration of WAF 100 mg/L) neither in control groups nor in treatment groups any mortality or any other effects could be observed.

Result:

LL0 (Cyprinus carpio; 96 h; static) = 100 mg/L;

LL10 (Cyprinus carpio; 96 h; static) > 100 mg/L

LL50 (Cyprinus carpio; 96 h; static) > 100 mg/L