Registration Dossier

Administrative data

toxicity to microorganisms
Type of information:
experimental study
Adequacy of study:
key study
Study period:
Feb 1996
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
other: According to DIN guideline under GLP
Cross-referenceopen allclose all
Reason / purpose for cross-reference:
reference to same study
Reason / purpose for cross-reference:
reference to other study

Data source

Reference Type:
study report
Report date:

Materials and methods

Test guideline
according to guideline
DIN 38412-8 (Pseudomonas Zellvermehrungshemmtest)
Principles of method if other than guideline:
Not relevant
GLP compliance:

Test material

Constituent 1
Chemical structure
Reference substance name:
EC Number:
EC Name:
Cas Number:
Molecular formula:
13-ethyl-3-methoxy-4,6,7,8,9,11,12,13,14,15,16,17-dodecahydro-1H-cyclopenta[a]phenanthren-17-ol (non-preferred name)
Details on test material:
- Name of test material (as cited in study report): Ethyldienol
- Analytical purity: 80.9 - 93.9%
- Lot/batch No.: 15003801-07 (mix of 7 batches)

Sampling and analysis

Analytical monitoring:

Test solutions


Test organisms

Test organisms (species):
Pseudomonas putida

Study design

Test type:
Water media type:
Limit test:
Total exposure duration:
16 h

Results and discussion

Effect concentrations
16 h
Dose descriptor:
Effect conc.:
> 200 mg/L
Nominal / measured:
Conc. based on:
test mat.
Basis for effect:
growth inhibition
Remarks on result:
other: saturated solution

Any other information on results incl. tables

Table 1: Mean values of turbidity (expressed as TE/F) and growth inhibition of Pseudomonas putida as a function of the

concentration (n=3) of ZK 47569

 Dilution of ZK 47569    TE/F [mean value (16 hours)]    Growth inhibition (%)  
 Control   93 0
 1:160   98 -5
 1:80   91 2
 1:40   76 18
 1:20   55 41
 1:10   64 31
 1 :5   53 43
 1:2,5   43 54
 Saturated   45 52

Applicant's summary and conclusion

Validity criteria fulfilled:
Since the growth inhibition was not concentration-related at dilutions of 1:10 and the higher
concentration, it was difficult to interpret the results. Therefore, another test with ZK 47569 was
performed. No inhibition of growth was observed in that study (Schering report X 131) which was
considered valid. Therefore, the results of this test are not further evaluated.
Executive summary:

The purpose of this study was to determine the growth inhibition of Pseudomonas putida under

exposure to Ethyldienol. The test was conducted in agreement with the DIN standard 38412, L8,

"Wachstumshemmtest mit dem Bakterium Pseudomonas putida".

200 mg of the test substance were suspended in 1000 ml demineralized water under constant stirring tor

24 hours. In order to prepare the test solutions, this suspension was filtered. This solution was

used as the highest test concentration and was further diluted 1 :2.5, 1 :5, 1: 10, 1 :20, 1 :40, 1 :80

and 1 :160. Additionally, one control without the test substance was used. All test solutions including the control were incubated in triplicate. Furthermore, for each test concentration one test vessel was incubated without addition of the

inoculum in order to analyse the inherent turbidity of the test compound.

The concentrationof the saturated, filtered solution was estimated by a TOC-analysis and

subsequent calculation based on the molecular formula.

According to the results of a TOC-analysis, the saturated, filtered solution has a concentration of < 1 mg/L.

As a parameter for the growth of the bacterial population, theturbidity of the test and control

solutions was analysed photometrically at a wave-Iength of 436 nm.

The measurements showed a growth decrease in all solutions except the 1 :160 dilution in

comparison with the control group.

However, the control growth exceeded the required minimum of the DIN standard only by

3% and therefore, it is doubtful, whether the observed reduction of growth was compound related,

since it was not concentration-related at dilutions of 1:10 and lower.