(benzothiazol-2-ylthio)acetic acid

Short-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

short-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Study period:

2015-03-31 to 2015-04-02

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

(benzothiazol-2-ylthio)acetic acid

EC Number:

228-565-0

EC Name:

(benzothiazol-2-ylthio)acetic acid

Cas Number:

6295-57-4

Molecular formula:

C9H7NO2S2

IUPAC Name:

2-(1,3-benzothiazol-2-ylsulfanyl)acetic acid

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

-Test item (Benzothiazol-2-ylthio)acetic acid
-CAS No. 6295-57-4
-EC No. 228-565-0
-Molecular formula C9H7NO2S2
-Molecular weight 225.29
-Active content (certified) 97.58 [GC area-%]

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

The limit concentration and the control were analytically verified via HPLC-DAD at the start of the exposure (0 hours) and at the end of the
exposure (48 hours) as specified below.
At the start of the exposure (0 hours), samples were taken after preparation of the limit concentration and analysed.
At the end of the exposure (48 hours), samples were taken from additional replicates with test media, but without daphnids, which were incubated under test conditions until sampling, and analysed.
The method was validated according to SANCO 3029/99 rev.4 (2000).

Recoveries of the test item should be within ± 20 % of the nominal or initially measured concentration. The effect levels can be given based on the
nominal concentrations, if this quality criteria is fulfilled.

Test solutions

Vehicle:

no

Details on test solutions:

Stock solution The stock solution (100 mg/L of the test item were weighed out) was freshly prepared with ISO test water (see below) before the
start of the exposure (0 hours).

Treatment The stock solution was treated with ultrasound for 1 minute at room temperature and stirred with approximately 1100 rpm for
90 minutes at room temperature.

Limit concentration A limit concentration of the test item of 100 mg/L was tested. The limit concentration was selected based on the results of a
non GLP preliminary range finding test.

Control Dilution water without test item tested under the same conditions as the test groups.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM
- Common name: Daphnia magna
- Strain: Clone 5
- Source:
Origin: Institut für Wasser-, Boden- und Lufthygiene (WaBoLu)
Breeder: DR.U.NOACK-LABORATORIEN, Käthe-Paulus-Str. 1, 31157 Sarstedt, Germany
- Age at study initiation (mean and range, SD): 2 to 24 h old daphnids from a healthy stock were used for the study
- Method of breeding:
Culture: In glass vessels (2 - 3 L capacity) with approximately 1.8 L culture medium, at 20 +- 2°C, in an incubator, 16 h illumination; light intensity
of max. 20 µE x m-2 x s-1
Culture medium: Elendt M4, according to ELENDT (1990), modified to a total hardness of 160 to 180 mg CaCO3/L
Cultur feeding: At least 5 times per week ad libitum with a mix of unicellular green algae, e.g. Pseudokirchneriella subcapitata and
Desmodesmus subspicatus, with an algae cell density of > 106 cells/mL
- Feeding during test: No feeding

ACCLIMATION
- Acclimation period: At least 2 h in dilution water.
- Acclimation conditions (same as test or not): Same as test conditions.
- Health during acclimation (any mortality observed): No mortality observed.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

48 h

Test conditions

Hardness:

269 mgCaCO3/L

Test temperature:

18 - 22°C, constant within ± 1°C
At start of exposure: 20.8 °C

pH:

pH at the Start of the Exposure (0 hours)
(measured in one additional replicate of the limit concentration and the control)

Nominal test item
concentration
[mg/L] pH-value

100 6.10
Control 7.61


pH at the End of the Exposure (48 hours)
(measured in all replicates)

Nominal test item
concentration
[mg/L] pH-values
Replicates
1 2 3 4
100 7.43 7.36 7.34 7.34
Control 7.65 7.65 7.66 7.65

Dissolved oxygen:

O2-concentration at the Start of the Exposure (0 hours)
(measured in one additional replicate of the limit concentration and the control)

Nominal test item
concentration
[mg/L]
Dissolved
O2-concentration
[mg/L]
100 8.34
Control 8.70



O2-concentration at the End of the Exposure (48 hours)
(measured in all replicates)

Nominal test item
concentration
[mg/L] Dissolved O2 -concentration [mg/L]
Replicates
1 2 3 4
100 8.18 8.10 8.22 7.89
Control 7.99 7.68 7.78 7.75

Nominal and measured concentrations:

A limit concentration of the test item of 100 mg/L was tested.

Details on test conditions:

TEST SYSTEM
- Test vessel: Glass beakers (4 cm ID x 7 cm H), 50 mL capacity, loosely covered with watch glasses
- Test volume: 20 mL
- Aeration: None
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: ISO Test water, according to OECD 202, Annex 3
Component Concentration [mg/L]
KCl 5.75
NaHCO3 64.8
CaCl2 x 2 H2O 294
MgSO4 x 7 H2O 123

- Water Quality Parameters of the Dilution Water prior to the Start of the Exposure (0 h):
pH-value Dissolved O2 concentration [mg/L] Temperature [°C] Conductivity [µS/cm] Total hardness [mg CaCO3/L]
7.61 8.70 20.8 680 269

- Culture medium different from test medium: Elendt M4, according to ELENDT (1990), modified to a total hardness of 160 to 180 mg CaCO3/L
- Intervals of water quality measurement: test start (0 h) and test end (48 h)

OTHER TEST CONDITIONS
- Photoperiod: 16/8 h light/dark cycle
- Light intensity: Diffuse light, light intensity of max. 20 µE x m-2 x s-1 (<1340 lx)

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Immobilisation, 24 h interval

TEST CONCENTRATIONS
A limit concentration of the test item of 100 mg/L was tested.

Reference substance (positive control):

yes

Remarks:

Potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

24 h

Dose descriptor:

other: EL10,50,100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other:

Effect concentrations 2

Duration:

48 h

Dose descriptor:

other: EL10,50,100

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Effect concentrations 3

Duration:

48 h

Dose descriptor:

other: NOEL

Effect conc.:

100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Effect concentrations 4

Duration:

48 h

Dose descriptor:

other: LOEL

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: immobilisation

Results with reference substance (positive control):

The percentage immobility for the reference item was determined after 24 hours. The EC50-value with 95 % confidence limits (CI) was calculated by
sigmoidal dose-response regression. The EC50-value for the most recent of the monthly performed reference tests was:
EC50: 1.51 mg/L (CI 1.08 - 2.10 mg/L)
The EC50-value of the reference item potassium dichromate after 24 h is within the prescribed concentration range of 0.6 - 2.4 mg/L of quality
criteria according to AQS P 9/2 (02/2000) for daphnids clone 5 cultured in Elendt M4 medium. The EC50-value of the reference item is also within the recommended range of 0.6 - 2.1 mg/L according to OECD-Guideline 202.

Any other information on results incl. tables

Biological Data

 The percentage immobility, determined in the limit concentration and the control after 24 and 48 hours under static conditions, is given in the table below. 

Immobilisation Rates after 24 and 48 hours of Exposure in the Definitive Test

  (n = 20, divided into 4 replicates with 5 daphnids each)

(Benzothiazol-2-ylthio) acetic acid	IMMOBILISATION [%]
Nominal test item concentration   [mg/L]	24 hours					48 hours
	Replicates					Replicates
	1	2	3	4	MV	1	2	3	4	MV
100	0	0	0	0	0	0	0	0	0	0
Control	0	0	0	0	0	0	0	0	0	0

The limit concentration was visually clear throughout exposure.

 

Measured Exposure Concentrations during the Definitive Test

 

The limit concentration of the test item (Benzothiazol-2-ylthio) acetic acid and the control were analytically verified by HPLC-DAD at the start of the exposure (0 hours) and at the end of the exposure (48 hours).
The measured concentration of the test item was 101 mg/L at the start of the exposure (0 hours) and also 101 mg/L at the end of

the exposure (48 hours).
The analytical results arepresented in the table below.

The recoveries of the test item were within ± 20 % of the nominal value. This indicates that the test item was successfully maintained for the duration of the test.

  Measured Concentrations and Percent of the Nominal Concentration of the Test Item(Benzothiazol-2-ylthio)acetic acidin the Definitive Test

Sampling date	2015-03-31 Start of the exposure, 0 hours		2015-04-02 End of the exposure, 48 hours
Date of analysis	2015-03-31		2015-04-02
Nominal test item concentration [mg/L]	(Benzothiazol-2-ylthio)acetic acid
	Meas. conc. [mg/L]	%	Meas. conc. [mg/L]	%
100	101	101	101	101
Control	< LOQ		< LOQ

Meas. conc.= measured concentration of the test item, mean value of 2 injections, dilution factor taken

into account

%                  = percent, related to the nominal test item concentration of the test item

LOQ             = limit of quantification of theanalytical method(0.2 mg/L of the test item)

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

At the limit concentration of 100 mg/L of the test item (Benzothiazol-2-ylthio)acetic acid, no effects were observed on Daphnia magna.

Executive summary:

In the acute immobilisation test with Daphnia magna (STRAUS), the effects of the limit concentration of 100 mg/L of the test item (Benzothiazol-2-ylthio)acetic acid (batch number:41273) was determined according to OECD 202 (2004). The limit test was conducted under static conditions over a period of 48 hours from 2015-03-31 to 2015-04-02, at Dr.U.Noack-Laboratorien, Käthe-Paulus-Str.1, 31157 Sarstedt, Germany.

The test item is a light yellow to white powder and has a water solubility of 133 mg/L at room temperature. The limit concentration of 100 mg/L was visually clear throughout the exposure.

Twenty daphnids were exposed to the limit concentration and the control.

The limit concentration of the test item (Benzothiazol-2-ylthio)acetic acid and the control were analytically verified by HPLC-DAD at the start of the exposure (0 hours) and at the end of the exposure (48 hours).
The measured concentration of the test item was 101 mg/L at the start of the exposure (0 hours) and also 101 mg/L at the end of the exposure

(48 hours).

The recoveries of the test item were within±20 % of the nominal value. This indicates that the test item was successfully maintained for the duration of the test. Therefore, the effect levels can be based on the nominal concentrations of the test item (Benzothiazol-2-ylthio)acetic acid.

The water quality parameters (i.e. pH-value and dissolved oxygen concentration), measured at the start (0 hours) and at the end of the exposure (48 hours), were within the acceptable limits. The validity criteria of the test guideline were fulfilled.

At the limit concentration of 100 mg/Lof the test item
(Benzothiazol-2-ylthio)acetic acid,

no effects were observed on Daphnia magna.

 EC-Values, NOEC and LOEC (based on the nominal concentration of the test item)

Effect levels	Test duration [h]	(Benzothiazol-2-ylthio)acetic acid
		Nominal test item concentration [mg/L]
EC10 /50 / 100	24	> 100
	48	> 100
NOEC	48	100
LOEC	48	> 100