5H-Dicyclopropa[6,7:15,16]cyclopenta[a]phenanthrene-3,5,17-triol, octadecahydro-17-(3-hydroxypropyl)-10,13-dimethyl-, (3S,5R,6R,7R,8R,9S,10R,13S,14S,15S,16S,17S)-

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:

Feb - Nov 2003

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2005

Report date:

2005

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

17 alpha-(3-Hydroxypropyl)-6 beta,7 beta;15 beta,16 beta-dimethylene-5 beta-androstane-3 beta,5,17 beta-triol

EC Number:

617-350-0

Cas Number:

82543-18-8

Molecular formula:

C24 H38 O4

IUPAC Name:

17 alpha-(3-Hydroxypropyl)-6 beta,7 beta;15 beta,16 beta-dimethylene-5 beta-androstane-3 beta,5,17 beta-triol

Test material form:

solid

Sampling and analysis

Analytical monitoring:

yes

Test solutions

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: For the stock solution 51 mg of ZK 92836 were suspended in 500 mL demineralized water ultrasonified for approximately 30 min and constantly stirred for approximately 24 hours at room temperature. Thereafter, the suspension was filtered through a glassfibre filter. The resulting solution was considered to be a saturated solution, which is the maximum achievable concentration to be tested according to the EC guideline
- Differential loading: yes

Test organisms

Test organisms (species):

Desmodesmus subspicatus (previous name: Scenedesmus subspicatus)

Details on test organisms:

TEST ORGANISM
- Common name: green algae
- Strain: Collection of Algae Cultures, University Goettingen kept on slant agar containing a nutrient solution under standardized conditions (10-15°C, diffuse light [light/dark rhythm 12 h/12 h]).
- Age of inoculum (at test initiation): 3 hours

ACCLIMATION
- Any deformed or abnormal cells observed: no

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test conditions

Test temperature:

23 °C

pH:

9.7 - 10.3

Nominal and measured concentrations:

nominal: 100 mg/L nominal
measured saturated solution: 64.3 mg/L
Dilutions extrapolated: 0.6 , 1.3, 2.6, 6.4, 12.9, 51.5 mg/L

Details on test conditions:

TEST SYSTEM
- Initial cells density: 10000 cells/mL
- Control end cells density: approx. 958800 cells/mL
- No. of vessels per concentration (replicates): 3
- No. of vessels per control (replicates): 6

GROWTH MEDIUM
- Standard medium used: yes

TEST MEDIUM / WATER PARAMETERS
- Source/preparation of dilution water: according to guideline

OTHER TEST CONDITIONS
- Photoperiod: continuous
- Light intensity and quality: approx. 6000 - 8000 lux

EFFECT PARAMETERS MEASURED
- Determination of cell concentrations: Fluorescence measurement (wavelength of 320 and 800 nm for agitation and measurement, respectively)
- Other: The algae were microscopically examined at the end of exposure.

CULTURING APPARATUS
-Details on culturing apparatus used: Abimed AlgenTest XT

Reference substance (positive control):

no

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

51.5 mg/L

95% CI:

>= 34.9 - <= 110.5

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Effect concentrations 2

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 51.5 mg/L

Nominal / measured:

meas. (arithm. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

- Exponential growth in the control (for algal test): yes
- Observation of abnormalities (for algal test): At the concentration of 6.4 mg/L oversized cells were noted, which was not considered to be an adverse effect. At the two highest concentrations damaged cells were observed.

Any other information on results incl. tables

Inhibition of biomass and the growth rate (%) of Desmodesmus subspicatus after 72 hours ex posure to ZK92836

Dilution of	Concentration of	Inhibition in percent
ZK 92836	ZK 92836 (mg/L)	Biomass (integral)	Growth rate
0	0,0 (control)	0	0
1:100	0,6	4,5	-5,6
1:50	1,3	-5,2	8,1
1:25	2,6	5,7	3,5
1:10	6,4	7,8	0,1
1:5	12,9	10,6	4,5
1:1,25	51,5	50,7	23,4

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

ErC50 (growth rate): >51.5 mg/L

Executive summary:

The purpose of this study was to determine the effects of the test compound Dimethylenpropanol (ZK 92836) on the growth of the green algae Desmodesmus subspicatus. ZK 92836 is an intermediate of the synthesis of Drospirenone. The study was conducted in agreement with the test guideline OECD no. 201.

The test substance ZK 92836 was incubated in an aqueous solution inc1uding nutrients with an algae population of Desmodesmus subspicatus for a test duration of approximately 72 hours in an electronically controlled dosing and incubation apparatus.

For the preparation of the test solutions a suspension with a nominalloading of approximately 100 mg/L, ZK 92836 was suspended in water, ultrasonified for approximately 30 min and constantly stirred for approximately 24 hours. The organic carbon concentration of the stock solution was analyzed with a TOC analyzer in a sampie taken at the start of the incubation. The substance concentration was calculated on the basis of the molecular formula. It was 64.3 mg/L.

The biomass and growth rate was relevantly (>10%) inhibited at the dilution of 1:5 and 1:1.25, respectively. This was considered to be a compound related effect. ZK 92836 had a concentration-depended inhibitory effect on the growth of Desmodesmus subspicatus. The EbC50 (biomass) was 51.5 mg/L [95% confidence limits: 34.9-110.5], the ErC50 (growth rate) was > 51.5 mg/L on the basis of measured concentrations. Consequently, ZK 92836 was considered to be harmful to the green algae Desmodesmus subspicatus, since the ECso (biomass) was between 10 mg/L and 100 mg/L.