Di-tert-dodecyl disulphide

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

20-01-2017 to 10-02-2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Analytical monitoring:

yes

Details on sampling:

Algae were exposed to a serie of dilutions (1.0, 3.0, 10.0, 31.0 and 100% v/v) of the saturated stock solution, prepared using slow-stirring conditions described in the OECD 123 guideline, in dilution water.
The incubation was conducted in a phytoculture cabinet that allows test flasks to be incubated under precise conditions: temperature was set to 23 ± 1°C, flasks were continuously stirred at around 100 rpm and constantly illuminated by fluorescent tubes between 6,000 and 10,000 lux.
Test vessels were made of glass bottles (120 mL capacity) capped with cellulose bungs. They were filled with a volume of 50 mL.
An inoculated control flask was prepared and incubated under the same conditions, with no test item. Three vessels were prepared at each test concentration and six vessels for the control group, each vessel was inoculated with ca 10000 cells P. subcapitata /mL.

Vehicle:

no

Details on test solutions:

A saturated solution was prepared using slow-stirring conditions described in the OECD 123 guideline. The method used a special temperature controlled glass-jacketed test vessel (around 1000 mL of capacity) with a sampling tap at the lower end and a magnetic stir bar at its bottom. A known volume of test medium (1000 mL) was first poured into the flask, then a known quantity of test item (110 µL corresponding to a loading rate of 100 mg/L) was gently added at the surface of medium. The solution was stirred at room temperature with a magnetic stir bar at a speed avoiding formation of an emulsion and set up in order to create a vortex depth of around 0.5 cm: the aqueous phase was kept under stirring during approximately 72 hours, and then drawn off as a clear solution after a rest phase of around 45 minutes without agitation. Tests solutions (1.0, 3.0, 10.0, 31.0 and 100% v/v) were then prepared from dilutions of this saturated solution.

Test organisms (species):

Raphidocelis subcapitata (previous names: Pseudokirchneriella subcapitata, Selenastrum capricornutum)

Details on test organisms:

The test organism used for the study was Pseudokirchneriella subcapitata, Strain No. CCAP 278/4, supplied by the Culture Centre of Algae and Protozoa (Ambleside, UK).

Transfers of P. subcapitata were made regularly to provide suitable subcultures. The algae were cultivated under standardized conditions as described in Annex 2 of the OECD 201 guideline.

The quality of the stock culture was checked for the absence of micro-organisms and deformed or abnormal cells under microscopic observation before use.

Three days before the start of the exposure, two pre-cultures were prepared by inoculating sufficient cells from the algal stock culture to the growth medium to obtain a low cell density (104 cells/mL) for pre-culturing , in order to maintain exponential growth until the start of the test. The pre-cultures were incubated under the same conditions as those used for the test cultures. Only one of the two pre-cultures was used to inoculate the test flasks for the study. The second one was only used if the first one was damaged.

At the beginning of the test, the cell density of the pre-culture was determined. The result was used to calculate the volume to be introduced into each test flask in order to obtain an initial cell concentration of 104 cells/mL as recommended in the OECD 201 guideline.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

Temperature was maintained at 23 ± 1°C: range from 22.8 to 23.3°C (mean: 22.9°C).

pH:

7.9 - 8.0

Dissolved oxygen:

8.7 - 9.0

Nominal and measured concentrations:

Nominal concentrations : 1.0-3.1-10.0-31.0-100.0% v/v saturated solution

Reference substance (positive control):

yes

Key result

Duration:

72 h

Basis for effect:

growth rate

Remarks on result:

other: No effect up to the solubility limit.

Results with reference substance (positive control):

The sensitivity of the test system and the methodology are evaluated every two months by performing an algal growth inhibition test on potassium dichromate. The latest value of ErC50 obtained in March 2017 was 0.63 mg/ L.

For information, according to ISO 8692 standard, 72h-ErC50 obtained from an inter-laboratory exercise on potassium dichromate was in the range 0.65 to 1.73 mg/L.

Validity criteria fulfilled:

yes

Conclusions:

This study was designed to determine the effects of the test item on the growth of Pseudokirchneriella subcapitata in a 72-hour test according to the OECD 201 Guideline. It was demonstrated that algal growth was not inhibited at all tested concentrations (slight negative inhibitions were calculated) indicating that the test item has no effect on algae up to its water solubility limit.

Executive summary:

This study was designed to determine the effects of the test item on the growth of Pseudokirchneriella subcapitata in a 72-hour test according to the OECD 201 Guideline.

The algae test met the validity criteria of the test guideline detailed as follows:

- The biomass in the control cultures increased exponentially by a factor of 48.6 within the 72-hour test period (required: at last 16-fold)

- The mean coefficients of variation for section-by-section specific growth rates (days 0-1, 1-2 and 2-3) in the control cultures were max 21.1% (required: < 35%)

- The coefficient of variation of average specific growth rates during the whole test period in the control cultures was 0.9% (required: < 7%)

 

Tests solutions were prepared from dilutions of a saturated solution prepared using slow-stirring conditions described in the OECD 123 guideline.

Chemical analysis of test samples taken at 0 indicated that the test item was not detected (all results are below the detection limit of the analytical method, i.e. 0.036 mg/L). The exposure concentrations were thus based on the loading rates.

It was demonstrated that algal growth was not inhibited at all tested concentrations (slight negative inhibitions were calculated) indicating that the test item has no effect on algae up to its water solubility limit.

Description of key information

The toxicity to algae has been measured according to OECD 201. No significant inhibition was recorded up to the solubility limit of DTDDS after 72 h.

This result is in line with the weight of evidence appraoch developped demontrated that DTDDS is unlikely to cross biological membrane. See also "additional information".

Additional information

Thiebaud (2000) is a GLP-compliant guideline study showing the growth inhibition effect of polysulfides, di-tert-dodecyl on Pseudokirchneriella subcapitata over 72 hours. The maximum level of exposure, the saturation concentration (0.080 mg/L), resulted in no inhibition of the growth of Pseudokirchneriella subcapitata. The study is considered reliable and use as supportive data for this endpoint.

This is supported by Hoffmann (2010), a GLP compliant, algae toxicity study following OECD guideline 201 and OECD guideline 23 on the testing of difficult substances. Due to the low water solubility of the substance, the test solutions were based on Water Accommodated Fractions with no analytical monitoring, therefore only nominal concentrations are used. The only deviations to the study plan are the range in temperatures of the storage of the substance and this is not expected to affect the outcome of the experiment.

A weight of evidence has demonstrated that di-tert-dodecyl polysulfide and di-tert-dodecyl disulfide are almost perfect structural analogues with similar physico-chemical bioavailability driving properties; the only structural difference is the slight difference in the number of atoms in the polysulfide chain. This does not affect log Kow, i.e. lipophilicity. Di-tert-dodecyl-polysulfide is unable to enter the fish body via the diet as shown by an OECD TG 305 (dietary) study. Shorter substances similar to both di-tert-dodecyl disulfide and Di-tert-dodecyl-polysulfide are unable to enter the fish body via the gills as shown by OECD TG 305 (flow-through) studies. Di-tert-dodecyl-polysulfide has no effect on aquatic life in laboratory tests. Di-tert-dodecyl disulfide has no effect on aquatic plants in laboratory test. No effect was also observed in the human health endpoints.

Taken all together, all these data form a weight of evidence where no diakyl disulfide or dialkyl polysulfide (alkyl > 9 carbons) is likely to cross biological membranes and exert toxicity on any organisms.