3,3'-thiodi(propionic acid)

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

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Guideline study with acceptable restrictions

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

At the start of exposure (0 hr) 2.0 mL each of the test liquid was sampled from the 3 test containers in each test group and mixed together to use as the analysis sample. When exposure was completed (72 hr), 2.0 mL each of the test culture liquid was sampled from the 3 test containers in each test group and mixed together before centrifuging the algae (3,000 rpm, 10 min) and using the clear liquid at the top as the analysis sample.
Equal amounts of acetonitrile was added and mixed into 0.75 mL of each analysis sample and then these were respectively analyzed using HPLC. The test material concentration in each test liquid and each test culture liquid were quantified using the ratio with standard solution peak area.

Vehicle:

no

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Japanese name: Muremikazukimo (monocellular green algae)
- Stock name: ATCC22662
- Source (laboratory, culture collection): American Type Culture Collection

ACCLIMATION
- Acclimation period: Pre-culture period: March 19, 2004 to March 23, 2004
- Culturing media and conditions: The environmental conditions were the same as for the test

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

23 +/- 2°C

pH:

The test liquid pH was not adjusted. The pH of the test liquid was 4.7 to 7.8 at the start of exposure, but the pH declined as the concentration increased. This is thought to be due to the test material. The test culture liquid pH at the end of exposure was 4.7 to 9.1. In the control group and the 20.0 mg/L concentration group the pH increased by 1 or more, but if the carbon dioxide assimilation progresses and the algae growth rate is high, it can increase by 1 or more, so this is not thought to be a problem. In addition, in the 45.0 mg/L concentration group, the pH decreased by 1, but this is thought to be due to the result of mutual action between the test material and the algae in the growth inhibition concentration range.

Nominal and measured concentrations:

Nominal concentrations: 20, 30, 45, 67 and 100 mg/L
Measured concentrations at 0 Hour: 19.4, 28.6, 43.5, 64.4, 96.5 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel:
- Type: open (with air permeable silicone stopper), continuously shaking culture (100 rpm)
- Material, size, headspace, fill volume: 300 mL glass triangular flasks
- Initial cells density: Pre-culture algae 1 x 10^4 cells/mL
- Control end cells density: 289.6x 10^4 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 3

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Culture medium different from test medium: no

OTHER TEST CONDITIONS
- Sterile test conditions: yes
- Adjustment of pH: no
- Photoperiod: continuous
- Light intensity and quality: 4000 lux

EFFECT PARAMETERS MEASURED: growth rate and biomass after 24, 48 and 72 hours
- Determination of cell concentrations: particle counter (CDA-500), blood cell counting chamber, Optical microscope: Model ECLIPSE TE300 made by Nikon Corporation

TEST CONCENTRATIONS
- Spacing factor for test concentrations: 1.5
- Range finding study
- Test concentrations: 0.1, 1, 10, 100 mg/L and 5, 9, 20, 50, 90, 100 mg/L in range finding studies

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

43.5 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 95% CL 33.7-56.2

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

28.6 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

biomass

Duration:

24 h

Dose descriptor:

EC50

Effect conc.:

52.9 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 24-48 hrs

Duration:

24 h

Dose descriptor:

NOEC

Effect conc.:

43.5 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 24-48 hrs

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

48.8 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 24-72 hrs

Duration:

48 h

Dose descriptor:

NOEC

Effect conc.:

19.4 mg/L

Nominal / measured:

meas. (initial)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 24-72 hrs

Details on results:

- Exponential growth in the control: yes
- Observation of abnormalities:
- Aggregation of algal cells: Under a microscope at the end of exposure it was found that some of the cells in the 45.0 mg/L or higher concentration groups had aggregated. A change in cell morphology (contraction, expansion, rupture, etc.) or cell aggregation was not observed in concentration groups of 30.0 mg/L or less, and that there was no difference from the control group.

Results with reference substance (positive control):

A growth inhibition test was regularly (approx. every 6 months) conducted using the standard material (potassium dichromate, special grade chemicals) to investigate the specimen organism sensitivity. The 72-hour 50% growth inhibition concentration (EbC50) since November 1996 is shown:
Average ± standard deviation = 0.432 ± 0.0740 mg/L, n = 14 (Lowest value to highest value = 0.285 to 0.543 mg/L)

Reported statistics and error estimates:

EC50 Values: The EC50 values and associated 95% confidence limits were determined by least squares linear regression analysis of the logarithm of test concentration against the growth inhibition (%) relative to control.
NOEC determination:
A Bartlett equal variance test (α = 0.01) was performed, and after equal variance was confirmed, a one-way analysis of variance (1-way ANOVA, α = 0.05) and a Williams multiple comparison test (α = 0.05, both sides) were performed, a comparison with the control group was made, and the highest test concentration for which no significant difference was found was used as the highest non-action concentration (NOEC). At this time, NOECb (0 to 72 hrs) was used when found using the area method, and NOECr (24 to 48 hrs) or NOECr (24 to 72 hrs) was used when found using the speed method.
The Yukms software Statlight “#4 Multigroup Comparison” (Yukms Corp., Tokyo) was used for the statistical analysis.

EC50 and NOEC values for growth rate were only calculated for 24 - 48 and 24 - 72 hours. In the report it was not measured why 72 hours values were not calculated for growth rate, but for biomass.

Table 1: Measured Concentration of the test substance in test cultures

Nominal Concentration (mg/L)	Measured concentration (mg/L)
	0 Hour	Percent of Nominal	72 Hours	Percent of Nominal
Control	< 0.08	--	< 0.08	--
20.0	19.4	97	19.5	98
30.0	28.6	95	29.0	97
45.0	43.5	97	43.8	97
67.0	64.4	96	64.4	96
100	96.5	97	97.1	97

Description of key information

The NOEC (growth rate) is  19.4 mg/L.

The ErC50 is 48.8 mg/L.

Key value for chemical safety assessment

EC50 for freshwater algae:

48.8 mg/L

EC10 or NOEC for freshwater algae:

19.4 mg/L

Additional information

During the static Algal Growth Inhibition test with Selenastrum capricornutum according to OECD 201 no effects on algae growth rate were observed at 19.4 mg/L (initial measured concentration). The ErC50 was determined to be 48.8 mg/L (initial measured concentration). Thus, the test substance shows a toxicity to Selenastrum capricornutum under conditions tested.