(1R,2R)-1-Amino-2-(difluoromethyl)-N-(1-methylcyclopropylsulfonyl)cyclopropanecarboxamide hydrochloride

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

15 August - 18 August, 2017

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2017

Report date:

2017

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

(1R,2R)-1-amino-2-difluoromethyl)-N- (1-methylcyclopropylsulfonyl)cyclo propanecarboxyamide hydrochloride

EC Number:

943-460-6

Cas Number:

1360828-80-3

Molecular formula:

C9H15CIF2N203S

IUPAC Name:

(1R,2R)-1-amino-2-difluoromethyl)-N- (1-methylcyclopropylsulfonyl)cyclo propanecarboxyamide hydrochloride

Test material form:

solid: particulate/powder

Details on test material:

White solid

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

A range-finding test:
The results of a non-GLP preliminary test showed that the 72-hour EC50 was located between 100-500 mg/L (nominal concentration), and NOEC may be
located in the range of 10-50 mg/L.

Exposure concentrations and grouping in the definitive test:
According to the results of the range-finding test, the definitive test was carried out as a static-exposure test with the concentration (with series of 1.875) of 40, 75, 141, 266 and 500 mg/L (nominal concentration, not be adjusted for purity).

The algal biomass in each flask was determined daily (at 24h, 48h and 72h) during the test period. The biomass was measured by manual cell counting by microscope. Small volumes were pipetted from the test solution to determine the biomass and then the residual solution was discarded. Microscopic observation was performed to verify a normal and healthy appearance of the inoculum culture and to observe any abnormal appearance of the algae. Any abnormal symptom was recorded.

The pH values of the solutions were measured at the start and the end of the test. The pH of the control medium were not to increase by more than 1.5 units during the test.

The light intensity was measured at the beginning and end of the test, and 5 points were selected and measured on the test vessel level. The light intensity was maintined within 15% from the average light intensity over the incubation area.

The average specific growth rate for a specific period was calculated as the logarithmic increase in the biomass from the equation for each single vessel of controls and treatments.

Test solutions

Vehicle:

not specified

Details on test solutions:

The definitive test was carried out as a static-exposure test with the concentrations of 40, 75, 141, 266 and 500 mg/L.

Each solution was divided 3 replicates with 100 ml solution, and the control was with 100 ml alga medium for each replicate.

The biomass of the pre-cultured alga solution was counted. And the mean concentration was 1.91x10^6 cell/mL, and then 0.260 mL volume pre-cultured alga solution was added into each replicate vessel, were mixed well to make the initial biomass to be 5 x 10^3 cell/ml. The residual solution was used for analysis of initial concentrations. All test flasks were capped with air-permeable membrane. Then the test vessels of each batch were put to an incubator respectively.

Test organisms

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

Test alga was Desmodesmus subspicatus, which is a freshwater alga species, and is one of the recommended species of OECD Guidelines for the Testing of Chemicals. The stock alga was originally introduced from Institute of Hydrobiology, Chinese Academy of Sciences and cultured continually in the facility.

To prepare the test solutions, 0.02021 g, 0.03766 g, 0.07072 g, 0.13321 g and 0.25022 g of the test item were weighed and dissolved into 500 ml volumetric flask with alga medium using a stirring rate at 320- 370 rpm 25 min.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Test conditions

Hardness:

Not specified

Test temperature:

21 - 24 °C, maintained at + 2 °C ( the actual monitored temperature was in the range of22.8 - 23.9°C)

pH:

The values of pH determined at the beginning and the end of the test were within the range of 3.01 - 8.64. The pH of the control medium increased 0.58
units maximally during the test. The concentrations group pH of adjusted were 3.10- 8.10 before, and adjusted to 7.98 - 8.09 at the beginning. The values of pH atthe end of the test were within the range of 7 .88 -8.57. The pH of the control medium increased 0.47 units maximally during the test.

Dissolved oxygen:

Not specified

Salinity:

Not specified

Conductivity:

Not specified

Nominal and measured concentrations:

Nominal test item concentrations of 40, 75, 141, 266 and 500 mg/L. Determined concentrations in 0-hour media were 41.6, 67.7, 128.3, 255.5, 478.3 mg/L. determined concentrations in 72-hour media were 32.9, 52.5, 116.8, 269.4, 507.0 mg/L. The overall determined concentrations were 37.3, 60.1, 122.5, 262.4, 492.6 mg/L. The determined concentration was 80.2 - 98.7% of the nominal concentration.

Details on test conditions:

Test growth media: Alga growth medium recommended by OECD was used and sterilized by Autoclave

Test duration: 72 hours

Initial biomass concentration: 5xl0^3 cell/mL

Temperature: 21 - 24 °C, maintained at + 2 °C ( the actual monitored temperature was in the range of22.8 - 23.9°C)

Oscillation rates: 100 Revolutions per minute

Illumination for Innova 43R incubator: The continuous light was used; the measured light intensity was 4956 lux - 5912 lux for cool white light at the beginning and the end of the test. The light intensity was in the range of -9.7% - 7.8% from the average light intensity, i.e. 5486 lux over the incubation area.

Illumination for QHZ-98B incubator: The continuous light was used; the measured light intensity was 5038 lux - 5987 lux for cool white light at the beginning and the end of the test. The light intensity was in the range of -8.2% - 9.1 % from the average light intensity, i.e. 5488 lux over the incubation area. All test vessels were capped with an air-permeable membrane.

Reference substance (positive control):

yes

Remarks:

3,5-dichlorophenol (3,5-DCP)

Results and discussion

Effect concentrations

Effect concentrations 1

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

86.15 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: adjusted for pH

Effect concentrations 2

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

45.07 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: yield

Remarks on result:

other: adjusted for pH

Effect concentrations 3

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

261.59 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: adjusted for pH

Effect concentrations 4

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

118.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: yield

Remarks on result:

other: adjusted for pH

Details on results:

During the 72-hour test duration, there was no any abnormal appearance of the algae observed in control and test replicates. After 72 hours exposure, there were apparent inhibition effects upon growth at the test concentration comparing with the control group. Inhibition rate increased with an increase in concentration.

Results with reference substance (positive control):

72 hour EyC50 of 3,5-DCP to Desmodesmus subspicatus was 1.81 mg/L with the 95% confidence limits 1.45 - 2.26 mg/L and 72 hour ErC50 was 2.00 mg/L with the 95% confidence limits 1.56 - 2.57 mg/L.

Reported statistics and error estimates:

The ErC10, EyC10, ErCso, EyCso and their 95% confidence limits were calculated using excel for pro bit method.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

According to the results, the 72-hour median effect concentration of average specific growth rate (ErC10 ErC50) and yield (EyC10 EyC50) and 95% confidence limit of Difluorosulfonamide HCl to Desmodesmus subspicatus were list as below:

Item EC10 or EC 50 95% confidence limit
ErC10 91.43 mg/L 0.65-166.82 mg/L
EyC10 48.49 mg/L 2.00-87.13 mg/L
ErC50 207.78 mg/L 82.03-990.87 mg/L
EyC50 116.77 mg/L 51.52-306.33 mg/L