4-methyl-salicylamide

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

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: According to official guidelines and GLP

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2016

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Test guideline 3

Qualifier:

according to guideline

Guideline:

other: Guidance document on aquatic toxicity testing of difficult items and mixtures, OECD series on testing and assessment number 23, December 14, 2000.

GLP compliance:

yes (incl. QA statement)

Test material

Reference

Name:

Unnamed

Type:

Constituent

Type:

Constituent

Test material form:

solid: crystalline

Details on test material:

Identification: V250208
Appearance: Off-white crystals
Batch: F8A
Purity/Composition: >96 %
<3 % 4-methylsalicylacid
<1 % 4-methylsalicylacid-methylester
<0.5 % water
Test item storage: At room temperature protected from light
Stable under storage conditions until: 03 September 2016 (expiry date)
Molecular formula: C8H9NO2
Molecular weight: 151.16

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Frequency: at t=0 h and t=72 h
Volume: 3.0 mL
Storage: Samples were stored in a freezer until analysis.

Test solutions

Details on test solutions:

The batch of V250208 tested consisted of an off-white crystals with a purity of 96% and the test item was not completely soluble in test medium at the initial loading rate. No correction was made for the purity/composition of the test item.

Preparation of test solutions started with a loading rate of 100 mg/L applying a 3-day period of magnetic stirring to ensure maximum dissolution of the test item in the test medium. The resulting mixture was filtered over a 0.45 μm membrane filter (Whatman; RC55). The obtained Water Soluble Fraction (WSF) was used as the highest test concentration. The lower test concentrations were prepared by subsequent dilutions of the WSF in test medium. All final test solutions were clear and colourless.

Test organisms

Test organisms (species):

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

Details on test organisms:

Species: Pseudokirchneriella subcapitata, strain: NIVA CHL 1
Source: In-house laboratory culture.
Reason for selection: This system is a unicellular algal species sensitive to toxic items in the aquatic ecosystem and has been selected as an internationally accepted species.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test conditions

Test temperature:

During the exposure period, the temperature measured in the incubator was maintained between 22 and 23°C remaining within the limits prescribed by the study plan (21-24°C, constant within 2°C).

pH:

The pH was within the limits prescribed by the study plan (6.0-9.0, preferably not varying by more than 1.5 units).

Nominal and measured concentrations:

At the start of the test, the actual test concentrations were in agreement with the nominal percentages of the WSF prepared at a loading rate of 100 mg/L, i.e. the measured test concentrations were 0.93, 3.0, 8.9, 28 and 90 mg/L in 1.0, 3.2, 10, 32 and 100% of the WSF, respectively. All concentrations remained stable during the test period (94-104% of initial). Given this result, the analytically confirmed nominal concentrations were used to determine the effect parameters.

Details on test conditions:

3 replicates of each test concentration.
6 replicates of the control.
1 replicate of an intermediate test concentration without algae for sampling purposes.
1 replicate of each test concentration without algae, not used during the final test.

Cell density: An initial cell density of 1 x 104 cells/mL.
Illumination: Continuously using TLD-lamps with a light intensity within the range of 87 to 92 μE/m2/s.
Incubation: Capped vessels were distributed at random in the incubator and as such were daily repositioned. During incubation the algal cells were kept in suspension by continuous shaking.

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 2

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

49 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: yield inhibition

Remarks on result:

other: 95% confidence interval ranging from 43 to 55 mg/L

Effect concentrations 3

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

32 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 4

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

10 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

other: yield inhibition

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Under the conditions of the present study with Pseudokirchneriella subcapitata, V250208 reduced the growth rate of this fresh water algae species significantly at nominal concentrations of 100 mg/L and higher.
The EC50 for growth rate reduction (72h-ERC50) was beyond the tested concentration range, i.e. exceeded an analytically confirmed concentration of 100 mg/L.
The EC50 for yield inhibition (72h-EYC50) was 49 mg/L with a 95% confidence interval ranging from 43 to 55 mg/L.
The 72h-NOEC for growth rate inhibition was 32 mg/L and the 72h-NOEC for yield inhibition was 10 mg/L .