3,7-dimethyloct-1-en-3-ol

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:

02.10.2014 - 18.03.2015

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Remarks:

GLP study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2015

Report date:

2015

Materials and methods

Test guideline

Test guideline 1

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Freshwater Alga and Cyanobacteria, Growth Inhibition Test)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.3 (Algal Inhibition test)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

3,7-dimethyloct-1-en-3-ol

EC Number:

242-358-2

EC Name:

3,7-dimethyloct-1-en-3-ol

Cas Number:

18479-49-7

Molecular formula:

C10H20O

IUPAC Name:

3,7-dimethyloct-1-en-3-ol

Test material form:

liquid

Specific details on test material used for the study:

Batch: UU14059249
Expiry date: 31 May 2016

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

For the determination of the actual test item concentrations, duplicate samples were taken from the test media of all test concentrations at the start of the test (without algae) and at the end of the test (containing algae). At the same sampling times, duplicate samples were also taken from the control. For sampling at the end of the test, the test medium of the treatment replicates was pooled.

All samples were stored deep-frozen (at about -20 °C) immediately after sampling. Based on pre-experiments for investigation of the storage stability, the test item was found to be stable in the test water under these storage conditions.

Test solutions

Vehicle:

no

Details on test solutions:

Test Water

Reconstituted test water (AAP Medium) prepared according to the test guidelines was used for algal cultivation and testing. Analytical grade salts were dissolved in sterile purified water to obtain the following concentrations:

Ingredients Concentration
Macro-nutrients NaHCO3 235.0 mg/L
K2HPO4 1.044 mg/L
MgSO4 × 7 H2O 14.6 mg/L
MgCl2 × 6 H2O 12.16 mg/L
CaCl2 × 2 H2O 4.41 mg/L
NaNO3 25.5 mg/L
Trace elements H3BO3 186.0 µg/L
MnCl2 × 4 H2O 415.0 µg/L
ZnCl2 3.27 µg/L
CoCl2 × 6 H2O 1.43 µg/L
CuCl2 × 2 H2O 0.012 µg/L
Na2MoO4 × 2 H2O 7.26 µg/L
FeCl3 × 6 H2O 160.0 µg/L
Na2EDTA × 2 H2O 300.0 µg/L

To keep the pH of the test media as constant as possible 6 mmol/L HEPES-buffer (corresponding to 1430 mg/L) were added to the test water. The pH was 7.5. The water hardness (calculated) of the test water was 0.15 mmol/L (= 15 mg/L as CaCO3).

The following nominal concentrations of the test item were tested: 0.32, 1.0, 3.2, 10, 32 and 100 mg/L. Additionally, a control was tested in parallel (test water without test item).

Dosage

The test medium of the highest nominal concentration of 100 mg/L was prepared in a closed system by dissolving 57.94 mg of the test item completely in 570 mL of test water using intense stirring for three hours at room temperature in the dark. The test medium of the highest test concentration was diluted with test water to prepare the test media of the lower test concentrations. The test media were prepared just before the start of the test.
The test method is based on the OECD series on testing and assessment No. 23 on Aquatic Toxicity Testing of Difficult Substances and Mixtures (2000).

Test organisms

Test organisms (species):

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

Details on test organisms:

The test organism used for the study was Pseudokirchneriella subcapitata (formerly Selenastrum capricornutum), Strain No. 61.81 SAG, supplied by the Collection of Algal Cultures (SAG, Institute for Plant Physiology, University of Göttingen, 37073 Göttingen / Germany). The algae were cultivated at Harlan Laboratories under standardized conditions according to the test guidelines.

Nygaard et al. (1986) recommended describing the taxa within the Genus Raphidocelis HINDAK as:
Raphidocelis subcapitata (KORSIKOV) nov. comb.
Basionym: Ankistrodesmus subcapitatus KORSIKOV
Syn.: Kirchneriella subcapitata KORSIKOV
Syn.: Selenastrum capricornutum PRINTZ
Syn.: NIVA-CHL 1

An inoculum culture was set up four days before the start of the exposure. The algae were cultivated under the test conditions. The inoculum culture was diluted three-fold one day before the start of the test to ensure that the algae were in the exponential growth phase when used to inoculate the test solutions.

Study design

Test type:

static

Water media type:

other: Reconstituted test water (AAP medium)

Limit test:

no

Total exposure duration:

72 h

Test conditions

Hardness:

0.15 mmol/L (= 15 mg/L as CaCO3), calculated

Test temperature:

24 °C

pH:

7.5-7.9

Nominal and measured concentrations:

Nominal: 0.32, 1.0, 3.2, 10, 32 and 100 mg/L
Measured (day 0): 0.333, 1.01, 3.21, 9.66, 30.6 and 95.5 mg/L
Measured (day 3): 0.306, 0.960, 3.13, 9.36, 30.1 and 91.6 mg/L

Details on test conditions:

Experimental Conditions

The test flasks were incubated in a temperature-controlled water bath at a temperature of 24 °C and illuminated by fluorescent tubes (Philips TLD 36W-1/840), installed above the test flasks. The test flasks were positioned randomly and repositioned daily. The mean measured light intensity at the level of the test solutions was approximately 6500 Lux (range: 5700 to 7050 Lux, measured at nine places in the experimental area). The light intensity over the incubation area was within a ±15%-deviation from the average light intensity as recommended by the guideline.

Study Design

In the definitive test, the enlarged spacing factor of 3.2 between the test concentrations was chosen based on the results of the range-finding test that documented a rather flat concentration-effect relationship; therefore, a large concentration range had to be tested.
The test design included three replicates per test concentration and six replicates of the control.
The test was started using a nominal algal cell density of 5000 cells/mL. The initial cell density was selected according to the recommendations of the OECD test guideline. The algal cell density in the pre-culture was determined using an electronic particle counter (Coulter Counter®, Model Z2).

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

64 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% confidence interval 61-66

Effect concentrations 2

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

28 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 95% confidence interval 23-33

Effect concentrations 3

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

27 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% confidence interval 24-30

Effect concentrations 4

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

4.9 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Remarks on result:

other: 95% confidence interval 2.6-7.3

Details on results:

The measured concentrations of 3,7-dimethyloct-1-en-3-ol in the test media of all test concentrations were between 96 and 104% of the nominal values at the start of the test and between 92 and 98% at the end of the test. Thus, the correct dosing of the test item 3,7-dimethyloct-1-en-3-ol was confirmed. The test item was stable in the test media over the test period of 72 hours. The biological results were related to the nominal concentrations of the test item.

The microscopic examination of the algal cells at the end of the test showed no difference between the algae growing at the nominal test concentration of 32 mg/L and the algal cells in the control. The shape and size of the algal cells were obviously not affected by the test item up to at least this concentration.

Results with reference substance (positive control):

For evaluation of the algal quality and experimental conditions, potassium dichromate is tested as a positive control twice a year to demonstrate satisfactory test conditions. The result of the latest positive control test performed in August 2014 (72-hour EC50 for the growth rate: 1.1 mg/L) showed that the sensitivity of the test system was within the internal historical range of the 72-hour EC50 for the growth rate from 2000 to 2014: 0.71-1.7 mg/L.

Reported statistics and error estimates:

The 72-hour EC10 and EC50 values for the inhibition of average growth rate and yield and their 95% confidence intervals were calculated by Weibull Analysis. For the determination of the NOEC, the average growth rate and yield at the test concentrations were compared to the control values by Dunnett t-test or Williams t-test, where appropriate.

Any other information on results incl. tables

The biological results of the study were as follows:

Parameter (0 -72 h):	Growth rate	Yield
EC50 [mg/L]	64	28
95% confidence interval [mg/L]	61 -66	23 -33
EC10 [mg/L]	27	4.9
95% confidence interval [mg/L]	24 -30	2.6 -7.3

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Conclusions:

The impact of the test item 3,7-dimethyloct-1-en-3-ol on the growth of the freshwater green algal species Pseudokirchneriella subcapitata can be summarized as follows (based on nominal test item concentrations):

Growth rate:
EC50 (95% CI) [mg/L]: 64 (61-66)
EC10 (95% CI) [mg/L]: 27 (24-30)

Yield (biomass):
EC50 (95% CI) [mg/L]: 28 (23-33)
EC10 (95% CI) [mg/L]: 4.9 (2.6-7.3)
 

Executive summary:

The impact of the test item 3,7-dimethyloct-1-en-3-ol on the growth of the freshwater green algal species Pseudokirchneriella subcapitata was investigated in a 72-hour static test according to the OECD Guideline 201 (adopted 2006, corrected 2011) and the Commission Regulation (EC) No 761/2009, C.3.

The nominal test item concentrations tested were 0.32, 1.0, 3.2, 10, 32 and 100 mg/L. Additionally, a control group was tested in parallel.

As the test item is a volatile substance, the test was performed using glass vessels completely filled with test medium, minimizing the headspace, that were tightly sealed with glass stoppers to avoid losses of test item. The preparation of the test media was based on the OECD Guidance

Document on Aquatic Toxicity Testing of Difficult Substances and Mixtures, 2000.

The measured concentrations of 3,7-dimethyloct-1-en-3-ol in the test media of all test concentrations were between 96 and 104% of the nominal values at the start of the test and between 92 and 98% at the end of the test. Thus, the correct dosing of the test item 3,7-dimethyloct-1-en-3-ol was confirmed. The test item was stable in the test media over the test period of 72 hours. The biological results were related to the nominal concentrations of the test item.

The biological results of the study were as follows:

Parameter (0 -72 h):	Growth rate	Yield
EC50 [mg/L]	64	28
95% confidence interval [mg/L]	61 -66	23 -33
EC10 [mg/L]	27	4.9
95% confidence interval [mg/L]	24 -30	2.6 -7.3