Cyclooctapentylose

Toxicity to aquatic plants other than algae

Administrative data

Endpoint:

toxicity to aquatic plants other than algae

Type of information:

experimental study

Adequacy of study:

supporting study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2009

Report date:

2009

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 221 (Lemna sp. Growth Inhibition Test)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Cyclooctapentylose

EC Number:

241-482-4

EC Name:

Cyclooctapentylose

Cas Number:

17465-86-0

Molecular formula:

C48H80O40

IUPAC Name:

cyclooctapentylose

Details on test material:

The test item and the information concerning the test item were provided by the sponsor.
Name: .gamma.-cyclodextrin
CAS No.: 17465-86-0
Chemical Name(s): cyclooctapentylose
Batch No.: 80P230
Certificate of Analysis Date: April 14, 2008
Aggregate State at Room Temperature: solid
Colour: white
Solubility: soluble in water up to 232 g/L (25°C)
Expiry Date: November 2010
Stability: In water: test item is considered to be stable under test conditions
Storage: In original container, at room temperature (10 - 30 °C), in the dark.

Specific details on test material used for the study:

Details on properties of test surrogate or analogue material (migrated information):
Species: Lemna gibba G 3
Origin: The fronds introduced in the test were taken from our in-house laboratory culture.
Breeding Conditions: The plants are cultured under standardised conditions according to the test guidelines.
Pre-Culture: For 14 days under test conditions.
Reference Item: For the evaluation of the quality of the Lemna gibba and the experimental conditions the substance potassium dichromate p.a. is tested at least twice a year to demonstrate satisfactory test conditions.

Type and Size: Glass flasks of 250 mL volume with approximately 200 mL test medium, covered with glass dishes.
Identification: Each test unit was uniquely identified with the serial number, treatment and replicate number.

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Storage of Samples: The samples were analysed directly. The rest of the samples was stored in the freezer (2 - 10 °C), protected from light and will be kept stored up to the date of the final report.
Fortified Samples: Approximately 50 mg test item were dissolved in 50 mL test water to obtain a stock solution of 1 g test item/L. Appropriate amounts of the stock solution were diluted with test water to obtain fortified samples at concentrations of 50 and 250 mg test item/L. Exact values were documented in the raw data.

Test solutions

Vehicle:

no

Details on test solutions:

Culture Medium:

20X AAP-Growth Medium: Analytical grade salts were added at the following nominal concentrations in deionised water (conductivity < 5 µS/cm):
Macro-nutrients:
NaHCO3 300 mg/L
K2HPO4 x 3 H2O 30 mg/L
MgSO4 x 7 H2O 290 mg/L
NaNO3 510 mg/L
MgCl2 x 6 H2O 240 mg/L
CaCl2 x 2 H2O 90 mg/L
Micro-nutrients:
H3B03 3.7 mg/L
MnCl2 x 4 H2O 8.3 mg/L
ZnCl2 0.066 mg/L
CoCl2 x 6 H2O 0.029 mg/L
CuCl2 x 2 H2O 0.00024 mg/L
Na2MoO4 x 2 H2O 0.145 mg/L
FeCl3 x 6 H2O 3.2 mg/L
Na2EDTA x 2 H2O 6.0 mg/L

The pH was adjusted with 1 M HCl to 7.4. The culture medium was prepared 1 day before test start to allow pH to stabilize.

Test organisms

Test organisms (species):

Lemna gibba

Details on test organisms:

The fronds introduced in the test were taken from our in-house laboratory culture.
The plants are cultured under standardised conditions according to the test guidelines.
For the evaluation of the quality of the Lemna gibba and the experimental conditions the substance potassium dichromate p.a. is tested at least twice a year to demonstrate satisfactory test conditions.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

7 d

Test conditions

Hardness:

no data available

Test temperature:

24 °C with a maximum deviation of +/- 2°C

pH:

7.5 +/- 0.1 at test start

Dissolved oxygen:

no data available

Salinity:

no data available

Nominal and measured concentrations:

The only concentration tested was nominal 250 mg test item/L. Additionally, a control was tested in parallel (test water without addition of the test item). Thus a limit test was performed in accordance with the Commission Directive 92/69/EEC to demonstrate that the test item has no inhibitory effect on the growth of the test organism up to this concentration.

Details on test conditions:

Test environment: Controlled environment room
Water temperature: 24 °C with a maximum deviation of ± 2 °C. The temperature was measured daily in a test vessel filled with culture medium and incubated under the same conditions as the test flasks.
pH-Value: 7.5 ± 0.1 at test start, the pH-value of the culture medium was measured in the culture medium of all test concentrations and the control at test start, on each observation day and at test end.
Light regime: Continuous illumination
Light intensity: 6500 - 10000 Lux; the illumination was achieved by fluorescent tubes, installed above the test flasks. Measurements were performed once during the test at 6 places distributed over the experimental area at the surface of the test media. Differences in light intensity over the test area did not exceed ± 15 %.

Reference substance (positive control):

yes

Remarks:

Control: Reconstituted water. In the control, test medium was used without addition of test item.

Results and discussion

Details on results:

Biological Results:
Growth Inhibition: The 7-day NOEµC and the LOEµC were determined to be < 250 and 250 mg test item/L, respectively.
The 7-day NOEµC and the LOEµC were determined to be < 250 and 250 mg test item/L, respectively.
However, due to the problems described in the conclusion, these values can not be taken as representative.
Shape of Fronds: In the only test item concentrations of nominal 250 mg/L the fronds showed slight chlorosis from day 5 on and gibbous growth on day 7. This supports the assumption that nutrition depletion occurred during the test.

Results with reference substance (positive control):

For the evaluation of the quality of the Lemna gibba and the experimental conditions the substance potassium dichromate p.a. is tested at least twice a year to demonstrate satisfactory test conditions.

Applicant's summary and conclusion

Validity criteria fulfilled:

not applicable

Conclusions:

The influence of .gamma.-cyclodextrin on the growth of the freshwater plant Lemna gibba was assessed in a static limit test.
The 7-day NOEµC and the LOEµC were determined to be < 250 and 250 mg test item/L, respectively.
The 7-day NOEµC and the LOEµC were determined to be < 250 and 250 mg test item/L, respectively.
In the course of the test, a strong turbidity occurred in the only test concentration of nominal 250 mg/L. Since the Lemna cultures are not axenic, this leads to the implication that turbidity was caused by growth of bacteria introduced with the Lemna culture. Similar findings in the previously performed algae growth inhibition test support the thesis that turbidity was caused by bacterial growth. Therefore, the given NOEC and LOEC values can not be considered representative.
Summing up, the test is technically not feasible under the given test conditions, due to the properties of the test item.

Executive summary:

Title: Toxicity of .gamma.-cyclodextrin to the Aquatic Plant Lemna gibba in a Static Growth Inhibition Limit-Test

- Guidelines/Recommendations: OECD Guideline 221: "Lemna sp. Growth Inhibition Test", adopted March 23, 2006. OECD Series on testing and assessment No. 23: Guidance Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures, December 2000

Purpose: The purpose of this test was to determine the inhibitory effect of the test item .gamma.-cyclodextrin on the growth of the freshwater aquatic plant Lemna gibba. Cultures of Lemna gibba were exposed to the only test concentration of 250 mg test item/L under defined conditions. The inhibition of growth in relation to control cultures was determined over a test period of 7 days. The test method of application and the test system are recommended by the test guidelines and Lemna gibba is one of the recommended test species.

Test Concentrations: The only concentration tested was nominal 250 mg test item/L. Additionally, a control was tested in parallel.

Validity of theTest: The doubling time of the frond number in the Control was 1.9 days. The test was therefore considered valid as the validity criterion of achieving a doubling time < 2.5 days was met.

Analytical Method: The quantification of the test item was performed using liquid chromatography (TOC-method).

Analytical Results: In the freshly prepared test media 83 % of the nominal concentration was determined, whereas in the aged test media 15 % of the nominal test concentration was measured. Therefore, all reported biological results are related to the mean measured concentration of the test item.

The influence of .gamma.-cyclodextrin on the growth of the freshwater plant Lemna gibba was assessed in a static limit test.

The 7-day NOEµC and the LOEµC were determined to be < 250 and 250 mg test item/L, respectively.

The 7-day NOEµC and the LOEµC were determined to be < 250 and 250 mg test item/L, respectively.

In the course of the test, a strong turbidity occurred in the only test concentration of nominal 250 mg/L. Since the Lemna cultures are not axenic, this leads to the implication that turbidity was caused by growth of bacteria introduced with the Lemna culture. Similar findings in the previously performed algae growth inhibition test support the thesis that turbidity was caused by bacterial growth. Therefore, the given NOEC and LOEC values can not be considered representative.

Summing up, the toxicity of .gamma.-cyclodextrin to Lemna gibba in OECD 221 study was technically not feasible under given test conditions due to occurring turbiditiy in the course of the test.