12-hydroxystearic acid

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:

From Sept. 2010 to Nov. 2010

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: The study was conducted according to standard guidelines in compliance with GLP. Measured concentrations were below the limit of quantification of the analytical method.

Justification for data waiving:

other:

Cross-reference

Cross-reference 1

Reason / purpose for cross-reference:

reference to same study

Cross-reference 2

Reason / purpose for cross-reference:

reference to other study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2010

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

Deviations:

no

Test guideline 3

Qualifier:

according to guideline

Guideline:

other:  OECD Series on testing and assessment No. 23: Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures, 2000

Deviations:

no

Principles of method if other than guideline:

Not applicable

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

12-hydroxystearic acid

EC Number:

203-366-1

EC Name:

12-hydroxystearic acid

Cas Number:

106-14-9

Molecular formula:

C18H36O3

IUPAC Name:

12-hydroxyoctadecanoic acid

Constituent 2

Reference substance name:

12-hydroxystearic aicd

IUPAC Name:

12-hydroxystearic aicd

Details on test material:

- Name of test material (as cited in study report): 12-hydroxystearic acid (CAS N°106-14-9, EC N°203-366-1)
- Substance type: Fatty acid
- Physical state: Solid / white to yellowish
- Storage condition of test material: At room temperature (15 - 25 °C), in the dark
- Expiration date of the lot/batch: 09 April 2011

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

- Duplicate samples from the freshly prepared test media (containing algae) of all test concentrations and from the control were taken at the start of the test.For the determination of the stability of the test item under the test conditions and of the maintenance of the test item concentrations during the test period, duplicate samples from the test media of all test concentrations and the control (containing algae) were taken at the end of the test (after the 72 hours test period). Additional samples were taken in duplicates at the test start and end (containing algae) and diluted in acetonitrile by factor 2 directly after sampling before freezing them.

- The concentrations of the test item were only analysed in the diluted duplicate test media samples from all test concentrations and both sampling times (0 and 72 hours). From the diluted control samples only one of the duplicate samples was analysed from each of both sampling times. The non-diluted test samples were not analysed.

Test solutions

Vehicle:

yes

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
The test item was not well soluble in test water. In this study, only the inhibitory effects of dissolved test item was tested and thus no concentrations above the solubility limit were tested. Therefore, a stock suspension of 100 mg /L was prepared by suspending 99.8 mg test item in 998 mL test water. The stock suspension was stirred for 24 hours at room temperature in the dark to dissolve as much test item as possible. Then, non-dissolved fractions of the test item were separated from the test medium by membrane filtration (0.45 µm Cellulose Nitrat Filter). The pH of the stock suspension was adjusted with 1 M NaOH from 7.8 to 8.0. The solution with dissolved test item was used as the test medium of the highest test concentration and to prepare the desired 1:3, 1:9, 1:27 and 1:81 dilutions. The test media were prepared just before introduction of the algae (= start of the test).

Test organisms

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: Freshwater green algae
- Strain: Strain No. 61.81 SAG
- Source (laboratory, culture collection): Sammlung von Algenkulturen, Pflanzenphysiologisches Institut der Universität Göttingen", 37073 Göttingen, Germany
- Method of cultivation: The algae were cultivated in the laboratories under standardised conditions according to the test guidelines

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

None

Test conditions

Hardness:

0.24 mmol/L (= 24 mg/L) as CaCO3.

Test temperature:

22 -24 °C

pH:

8.0 at the start of the test
9.4 - 9.5 at the end of the test

Nominal and measured concentrations:

Nominal: 0, 100 mg/L and 1:3, 1:9, 1:27 and 1:81 dilutions.

Details on test conditions:

TEST SYSTEM
- Test vessel: Erlenmeyer flasks
- Type (delete if not applicable): Closed
- Material, size, headspace, fill volume: 50 mL volume with 50 mL of test medium
- No. of vessels per concentration (replicates): Six
- No. of vessels per control (replicates): Six


GROWTH MEDIUM
- Standard medium used: yes


TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: Reconstituted water (OECD Medium)

OTHER TEST CONDITIONS
- Adjustment of pH: Yes
- Light intensity and quality: 6120 - 6440 lux

EFFECT PARAMETERS MEASURED (with observation intervals if applicable) :
- Determination of cell concentrations: Spectrophotometer

TEST CONCENTRATIONS
- Range finding study
- Test concentrations: 100 mg/L
- Results used to determine the conditions for the definitive study: Yes

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

>= 100 mg/L

Nominal / measured:

nominal

Conc. based on:

other: filtrate

Basis for effect:

other: growth rate and yield

Effect concentrations 2

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

other: filtrate

Basis for effect:

other: growth rate and yield

Effect concentrations 3

Duration:

72 h

Dose descriptor:

other: ErC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

other: filtrate

Basis for effect:

growth rate

Effect concentrations 4

Duration:

72 h

Dose descriptor:

other: EyC50

Effect conc.:

> 100 mg/L

Nominal / measured:

nominal

Conc. based on:

other: filtrate

Basis for effect:

other: yield

Details on results:

- Exponential growth in the control (for algal test): Yes
- Observation of abnormalities (for algal test): No
- Unusual cell shape: No
- Colour differences: No remarkable observations
- Adherence to test vessels: No

Results with reference substance (positive control):

- Results with reference substance valid: Yes

Reported statistics and error estimates:

None

Any other information on results incl. tables

Determination of the Test Item:	Based on the results of LC measurements the concentration of the test item was determined using a calibration curve.
:	0.1 to 5 mg test item/L
Linearity of Response:	Correlation of peak area of different standard solutions with their corresponding concentrations, using a linear regression
Regression Coefficient:	0.9995
Calibration Curve:	y = 68386 * x + 1979 (see also Figure2)
Limit of Detection:	0.04 mg/L
Limit of Quantification:	0.5 mg test item/L 92 % (n = 4, RSD 4 %)
Mean Recovery in the Fortified Samples:	90 % (n = 8, RSD 4 %) (seeTable12)
Results	The quantification of the the test item was performed using liquid chromatography (LC-MS/MS-method). Due to the low water solubility of the test item all concentrations were below the Limit of Quantification of the analytical method.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

Under the test conditions, the 72 h NOEC and LOEC of the test material were determined to be >=100 and > 100 mg/L (nominal), respectively. The 72 h ErC50 and EyC50 were > 100 mg/L (nominal).

Executive summary:

A study was conducted to determine the inhibitory effect of 12 -hydroxystearic acid on the growth of the freshwater green algae Pseudokirchneriella subcapitata in a static test (OECD Guideline 201 and EU method C.3).

The test item was not well soluble in test water. To avoid physical effects of undissolved test item, no concentration above the solubility limit of the test item in test water was tested. A supersaturated stock solution of 100 mg/L was prepared by suspending the substance in test water. The stock suspension was stirred for 24 hours in the dark , then undissolved substance was separated by filtration.

Algae were exposed to the filtrate at 100 mg/L (nominal) and dilutions of 1:3, 1:9, 1:27 and 1:81 for 72 h. Cell densities were measured at 24, 48 and 72 h. Analytical verification of the test concentrations were conducted by LC-MS/MS on samples collected at start and after 72 h.

Under the test conditions, the 72 h NOEC and LOEC of the test material were determined to be >=100 and > 100 mg/L (nominal), respectively. The 72 h ErC50 and EyC50 were > 100 mg/L (nominal).