1-(5,6,7,8-tetrahydro-3,5,5,6,8,8-hexamethyl-2-naphthyl)ethan-1-one

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:

14 September 1994 - 5 January 1998

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study conducted by GLP accredited laboratory and according to OECD guideline.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1998

Report date:

1998

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

yes

Remarks:

- the pH increased from 8.0 to 10.0. This slightly exceeds the tolerance of 1.5 pH units given by the guidelines. Because rapid growth and hence the large CO2 consumption, this could not be avoided, despite stirring at 500 rpm.

GLP compliance:

yes

Test material

Test material information

Constituent 1

Reference substance name:

1-(5,6,7,8-tetrahydro-3,5,5,6,8,8-hexamethyl-2-naphthyl)ethan-1-one

EC Number:

244-240-6

EC Name:

1-(5,6,7,8-tetrahydro-3,5,5,6,8,8-hexamethyl-2-naphthyl)ethan-1-one

Cas Number:

21145-77-7

Molecular formula:

C18H26O

IUPAC Name:

1-(3,5,5,6,8,8-hexamethyl-5,6,7,8-tetrahydronaphthalen-2-yl)ethan-1-one

Details on test material:

AHTN provided by the Research Institute of Fragrance Materials, Inc., USA

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Samples of 3.5 ml (main test 1) or 1-2 ml (main test 2) were taken from the test solutions containing algae after 24, 48, 72 and 96 hours of exposure.

Test solutions

Vehicle:

yes

Details on test solutions:

Nominal concentrations were prepared by dissolving AHTN in 5 ml of dimethylformamide (DMF). This was mixed with Tween 80, followed by a subsequent dilution with aqueous test medium, to form a stock solution.

Test organisms

Test organisms (species):

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

Details on test organisms:

The test was run with the recommended species Pseudokirchneriella subcapitata (strain no. 61.81 SAG). These test organisms, formerly known as Selenastrum capricornutum were suppiled by “Pflanzenphysiologisches Institut der Universitât Göttingen, D-37073 Göttingen). The algae were grown in the laboratories of RCC under standardized conditions.
Three days prior to the test a pre-culture of algae was inoculated under the same conditions as in the test. From this pre-culture the algae cells were taken to prepare an algae suspension in test medium. Aliquots of this algae suspension (corresponding to a biomass of 10,000 cells per ml of nutrient solution) were mixed with the respective test concentrations in test medium.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test conditions

Hardness:

The hardness of the test medium corresponded to about 24 mg CaCO3/l.

Test temperature:

Room temperature

pH:

The pH for all test concentrations was 8.0 at the start of the test after equilibration with air and ranged from 9.0 to 10.1 at the end of the test.

Dissolved oxygen:

Solution contibuously stirred at 500 rpm.

Nominal and measured concentrations:

See table 1.

Reference substance (positive control):

yes

Remarks:

Potassium Dichromate

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

612 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Effect concentrations 2

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

605 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Effect concentrations 3

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

278 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

biomass

Effect concentrations 4

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

> 816 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 5

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

816 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 6

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

409 µg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Results with reference substance (positive control):

The EbC50 and ErC50 for Potassium Dichromate after 72 hours of exposure was found to be at 0.53 mg/l (0.20 to 0.75 mg/l), and 0.84 mg/l (0.60 to 1.03 mg/l), respectively. Therefore, the algal cultures used by RCC were demonstrated to be adequately sensitive.

Reported statistics and error estimates:

As far as possible, the EbC50 and ErC and there confidence limits were estimated using the Logit model. The NOEC and LOEC were determined by applying Dunnett’s test.

Any other information on results incl. tables

Potassium dichromate as reference compound showed after 72 hours of exposure an EbC50 of 0.53mg/l and ErC50 of 0.84mg/l. It means that the test is sufficient sensitive. The pH was initially 8 at the start of the experiment and increased to 9 -10 at the end.

Table 1 Nominal and measured AHTN concentrations in the samples

Nominal concentration of test article (µg/l)	Time hours	% of nomal found	Mean measured concentration of test article (µg/l)	% of nomal found	Mean measured concentration of test article (µg/l)
62.5	0	82.6		77.4
	72	30.8		140
	mean	56.7	35.4	108.7	67.9
125	0	80.7		81.4
	72	60.2		142
	mean	70.5	88.1	111.7	139.6
250	0	90.1		83.5
	72	72.8		52.7
	mean	81.5	203.8	68.1	170.3
500	0	89.7		84.4
	72	85.3		67.8
	mean	87.5	437.5	76.1	380.7
1000	0	84.7		90.4
	72	74.7		76.6
	mean	79.7	797	83.5	835.0

The results of the inhibition test and algal growth can be found in table 2.

Table 2 AHTN-related inhibition of algal growth after 72 hours of exposure

Concentration of test article (µg/l)		% Inhibition
Nominal	Mean measured	Biomass (AUC) (0-72 hours)		Growth rate (µ)
Test 1	Test 2	Test 1	Test 2	Test 1	Test 2
35.4	67.9	17.8	-4	3	1.6
88.1	139.6	25.4	-11.8	6	-3.1
203.8	170.3	17.4	-20.9	-3	-2.1
437.5	380.7	36.0	-12.4	8.4	0.5
797.0	835.0	81.6	54.0	33.1	16.4

AUC: Areau Under Curve

The effect concentrations for the biomass and algal growth is shown in table 3

Table 3 NOEC, LOEC and EC50 for biomass and growth rate

Biomass	Test 1	Test 2
NOEC:	204 µg/l	381 µg/l
LOEC:	438 µg/l	835 µg/l
EbC50:	468 µg/l	801 µg/l (extrapolated value)
Growth rate (µ):	Test 1	Test 2
NOEC:	438 µg/l	381 µg/l
LOEC:	797 µg/l	835 µg/l
ErC50:	>797 µg/l	>835 µg/I

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

See overall remarks.

Conclusions:

The highest NOEC the biomass (AUC) is 381 µg/l, the lowest LOEC value is 438 µg/l and the lowest EbC50 is 468 µg/l. The highest NOEC for growth rate (µ) is 438 µg/l, the lowest LOEC is 797 µg/l and the ErC50 is above 835 µg/l.

Executive summary:

The toxicity to algae was studied in a static test on Pseudokirchneriella subcapitata. Nominal concentrations ranged from 0.0625 to 1.0 mg/l (step size 2). Concentrations were measured at the start and end of the test by HPLC analysis. Maintenance of the concentrations proved to be difficult and in some cases pH-values rose to above 10 indicating an insufficient buffering capacity of the medium. The loss of test material in the algal test was studied for a related polycyclic musk and it was shown that sorption to algal cells was the most relevant ‘sink’, whereas sorption to glass might play a role as well as volatilisation. The results were expressed as the mean concentration over the exposure time. The test was carried out twice (test A and B) due to the sharp concentration decrease in the first study with the goal to confirm the observed toxicity levels in a second study. Growth rate (μ) was the less sensitive parameter. In test (B) the growth rate reduction at 0.835 mg/l was 16% (LOECr) with a NOECr of 0.374 mg/l. In test (A), the NOECr for effects on growth rate was slightly higher, 0.438 mg/l, whereas the inhibition at 0.797 mg/l (LOECr) was 33%. The geometric mean NOECr for both tests was 0.40 mg/l. The 72-h ErC50 was > 0.797 and > 0.835 in test (A) and (B) respectively. Growth measured as Area Under the Curve was not significantly inhibited in concentrations up to and including (A) 0.204 and (B) 0.374 mg/l. In the next higher concentration, the LOECb, (A) 0.438 and (B) 0.835 mg/l, the biomass production was inhibited by (A) 36% and (B) 54%.