Serine, 2-[(2,3,4-trihydroxyphenyl)methylene]hydrazide, hydrochloride (1:1)

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: This study was selected as the key study because the information provided for the hazard endpoint is sufficient for the purpose of classification and labelling and/or risk assessment.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 202 (Daphnia sp. Acute Immobilisation Test)

Deviations:

no

Remarks:

The study was conducted according to guideline in effect at time of study conduct.

Test guideline 2

Qualifier:

according to guideline

Guideline:

EPA OPPTS 850.1010 (Aquatic Invertebrate Acute Toxicity Test, Freshwater Daphnids)

Deviations:

no

Remarks:

The study was conducted according to guideline in effect at time of study conduct.

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

2-amino-3-hydroxy-N'-[(E)-(2,3,4-trihydroxyphenyl)methylidene]propanehydrazide hydrochloride

EC Number:

940-411-0

Cas Number:

1353749-74-2

Molecular formula:

C10H14ClN3O5

IUPAC Name:

2-amino-3-hydroxy-N'-[(E)-(2,3,4-trihydroxyphenyl)methylidene]propanehydrazide hydrochloride

Details on test material:

- Purity: 100%

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Duplicate samples were collected from each batch of prepared test solution at the beginning of the test, and from two of the four replicate test chambers at 48 hours (± 1 hour) to measure concentrations of the test item. All samples were collected at mid-depth from the batch solutions or the test chambers, placed in glass scintillation vials, and acidified with 1.0 mL of 1% phosphoric acid in methanol. One set of samples was processed immediately for analysis while the second set was stored refrigerated for possible future analysis.

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION:
The highest concentration test solution was prepared by mixing 0.2000 g of test substance in 2 L of UV sterilized dilution water at a nominal concentration of 100 mg/L. The 100 mg/L test solution was sonicated for approximately 45 minutes then stirred for 90 minutes. The 100 mg/L test solution also served as the primary stock solution for preparing the remaining test solutions. Aliquots of the 100 mg/L test solution were proportionally diluted with dilution water to prepare four additional test solutions at nominal concentrations of 6.3, 13, 25 and 50 mg/L. The test solutions were stirred for 30 minutes. Approximately 210 mL of solution was placed in each of four replicate test chambers per treatment group. The negative control solution was dilution water only.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

TEST ORGANISM:
- Strain: Daphnia magna (obtained from cultures maintained by Wildlife International, Easton, Maryland)
- Age at study initiation: < 24 hours
- Feeding during test: None

ACCLIMATION
- Acclimation period: The three adult daphnids used to supply neonates for the test were held for 17 days prior to collection of the juveniles for testing and had each produced at least three previous broods.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

48 h

Test conditions

Hardness:

140 mg/L as CaCO3

Test temperature:

20 ± 1°C

pH:

7.4 to 8.7

Dissolved oxygen:

At test initiation, dissolved oxygen concentrations fell below 3.0 mg/L in the 19.2 mg/L treatment groups, so gentle aeration was added to all treatment groups. For the remainder of the test, dissolved oxygen measurements were > 8.6 mg/L (>96% saturation).

Salinity:

freshwater

Nominal and measured concentrations:

Nominal concentrations: 0, 6.3, 13, 25, 50 and 100 mg/L
Measured concentrations (Centrifuged): 0, 0.044, 0.75, 2.09, 6.35 and 19.2 mg/L

Details on test conditions:

TEST SYSTEM
- Test vessel: 250-mL glass beakers
- No. of organisms per vessel: 5
- No. of vessels per concentration (replicates): 4
- No. of vessels per control (replicates): 4


TEST MEDIUM / WATER PARAMETERS
- Conductivity: 362 μS/cm

OTHER TEST CONDITIONS
- Adjustment of pH: no
- Photoperiod: A photoperiod of 16 hours of light and 8 hours of darkness was controlled with an automatic timer. A 30-minute transition period of low light intensity was provided when lights went on and off to avoid sudden changes in lighting.
- Light intensity: 415 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable): All organisms were observed periodically to determine the number of immobile organisms in each treatment group. Immobility was defined as a lack of movement by the organism except for minor activity of the appendages. The numbers of individuals exhibiting signs of toxicity or abnormal behaviour also were evaluated. Observations were made approximately 3.5, 24 and 48 hours after test initiation.

Results and discussion

Effect concentrations

Duration:

48 h

Dose descriptor:

EC50

Effect conc.:

0.96 mg/L

Nominal / measured:

meas. (not specified)

Conc. based on:

test mat.

Basis for effect:

mobility

Remarks on result:

other: 0.73 – 1.24 mg/L

Details on results:

Daphnids in the negative control and the 0.044 mg/L treatment group appeared normal throughout the test with no immobility or overt signs of toxicity observed. Percent immobility at test termination in the 0.044, 0.75, 2.09, 6.35 and 19.2 mg/L treatment groups was 0, 30, 95, 100 and 100%, respectively. The sublethal effects of lethargy were observed in the 0.75 and 2.09 mg/L treatment groups.

Reported statistics and error estimates:

The immobility data were analysed using the computer program of C. E. Stephan. The program was designed to calculate the EC50 value and the 95% confidence interval by probit analysis, the moving average method, and binomial probability with nonlinear interpolation. In this study, probit analysis was used to calculate the 24 and 48-hour EC50 values and the 95% fiducial intervals. The highest mean measured test concentration causing no immobility at test end and the lowest mean measured test concentration causing 100% immobility at test end were assessed by visual observation of the immobility data.

Any other information on results incl. tables

Table 1- Mobility Results

Mean Measured Concentration mg/L	~3.5 Hours				24 Hours		48 Hours
	Rep	Daphnia/ Replicate	Number Immobile 1	Observations 2	Number Immobile 1	Observations 2	Number Immobile  1	Observations 2	Percent Immobile
Negative Control	A	5	0	5 AN	0	5 AN	0	5 AN	0
	B	5	0	5 AN	0	5 AN	0	5 AN
	C	5	0	5 AN	0	5 AN	0	5 AN
	D	5	0	5 AN	0	5 AN	0	5 AN
0.044	A	5	0	5 AN	0	5 AN	0	5 AN	0
	B	5	0	5 AN	0	5 AN	0	5 AN
	C	5	0	5 AN	0	5 AN	0	5 AN
	D	5	0	5 AN	0	5 AN	0	5 AN
0.75	A	5	0	5 AN	1	1 C; 3 AN	3	2 AN	30
	B	5	0	5 AN	0	5 AN	0	5 AN
	C	5	0	5 AN	0	2 C; 3 AN	2	3 C
	D	5	0	5 AN	0	2 C; 3 AN	1	2 C, 2 AN
1 Cumulative number of immobile daphnids. 2 Observed effects: AN = appear normal; C = lethargy.

Table 1 continued

Mean Measured Concentration (mg/L)	~3.5 Hours				24 Hours		48 Hours
	Rep.	Daphnia/ Replicate	Number Immobile 1	Observations 2	Number Immobile 1	Observations 2	Number Immobile  1	Observations 2	Percent Immobile
2.09	A	5	0	5 AN	4	1 AN	5	--	95
	B	5	0	5 AN	4	1 AN	4	1 C
	C	5	0	5 AN	4	1 AN	5	--
	D	5	0	5 AN	2	3 AN	5	--
6.35	A	5	0	5 AN	5	--	5	--	100
	B	5	0	5 AN	5	--	5	--
	C	5	0	5 AN	5	--	5	--
	D	5	0	5 AN	5	--	5	--
19.2	A	5	0	5 AN	5	--	5	--	100
	B	5	0	5 AN	5	--	5	--
	C	5	0	5 AN	5	--	5	--
	D	5	0	5 AN	5	--	5	--
1 Cumulative number of immobile daphnids. 2 Observed effects: AN = appear normal; C = lethargy.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

This study and the conclusions which are drawn from it fulfil the quality criteria (validity, reliability, repeatability).
48-hour EC50 = 0.96 mg/L

Executive summary:

The cladoceran (Daphnia magna) was exposed to five concentrations of the test item, ranging from 0.044 to 19.2 mg/L, based on mean measured concentrations, for 48 hours under static conditions according to OECD Guideline 202. The highest mean measured test concentration causing no immobility at test end was 0.044 mg/L. The lowest mean measured test concentration causing 100% immobility at test end was 6.35 mg/L. The 48-hour EC₅₀ value was 0.96 mg/L based on mean measured test concentrations of centrifuged test solutions.