Reaction mass of 1-hydroxypentan-2-yl formate and 2-hydroxypentyl formate and pentane-1,2-diol and pentane-1,2-diyl diformate

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Version / remarks:

2011

GLP compliance:

yes (incl. QA statement)

Remarks:

Federal Office for the Environment (FOEN)

Analytical monitoring:

yes

Details on sampling:

- Concentrations: the test item was determined in one of the duplicate test medium samples from the control and the test concentrations of 46 and 100 mg/L, since the remaining concentrations were below the NOEC determined in this test.
-Sampling method: duplicate samples were taken from the test media of all test concentrations and the control at the start of the test (without algae) and at the end of the test (containing algae). Test media of the replicates were pooled before sampling at the end of the test
- Sample storage conditions before analysis: after sampling, samples were stabilized with acetonitrile (1 mL acetonitrile per 10 mL sample volume) and deep-frozen (at about -20 °C) (method based on pre-experiments)

Vehicle:

no

Details on test solutions:

PREPARATION AND APPLICATION OF TEST SOLUTION (especially for difficult test substances)
- Method: a 100 mg/L solution was prepared by dissolving the test item in 1 L test water using intense stirring for 10 min at room temperature. This test concentration was further diluted with test water to prepare the remaining test concentrations.

Test organisms (species):

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

Details on test organisms:

TEST ORGANISM
- Common name: green algae
- Strain: No. 61.81 SAG
- Age of inoculum (at test initiation): 4 d (exponential growth phase by inoculation time)
- Method of cultivation: same as test

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Hardness:

-

Test temperature:

23 °C

pH:

Test item vessels: 7.1 - 8.6
Control: 7.3 - 8.4

Dissolved oxygen:

-

Salinity:

-

Conductivity:

-

Nominal and measured concentrations:

Nominal: 4.6, 10, 22, 46 and 100 mg/L
Mean measured: n.d., n.d., n.d, 27 and 85 mg/L (geom. mean)

Details on test conditions:

TEST SYSTEM
- Test vessel: 75 mL Erlenmeyer flasks (covered with a glass dish)
- Fill volume: 30 mL
- Initial cells density: 5000 cells/mL (determined using an electronic particle counter)
- Control end cells density: 235.5 x 10^4 relative fluorescence units
- 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
- Intervals of water quality measurement: pH was recorded in all treatments at the start and end of the test. The temperature in the incubator was monitored continuously. The appearance of the test media was visually controlled daily.

OTHER TEST CONDITIONS
- Photoperiod: continuously illuminated
- Light intensity and quality: LED light, 77 μE s^-1 m^-2

EFFECT PARAMETERS MEASURED
- Determination of cell concentrations: 100 μL algal suspension was withdrawn daily from each test flask and biomass was determined by duplicate fluorescence
- Other: at the end of the test, a sample was taken from the control and from the concentration of 46 mg/L and inspected for shape and size of the algal cells. This test concentration was chosen since the algal cell density was too low for an examination at the concentration of 100 mg/L.

TEST CONCENTRATIONS
- Range finding study: yes
- Test concentrations: 1, 10, 100 mg/L
- Results used to determine the conditions for the definitive study: 2.1, 1.8 and 5.9 % inhibition of the growth rate at concentrations of 1, 10 and 100 respectively after 72 h

Reference substance (positive control):

yes

Remarks:

potassium dichromate

Key result

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

77 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI: 68 – 81 mg/L

Key result

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

42 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI: 19 – 54 mg/L

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

27 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

85 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

growth rate

Duration:

72 h

Dose descriptor:

EC50

Effect conc.:

46 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Remarks on result:

other: 95% CI: 44 - 47 mg/L

Duration:

72 h

Dose descriptor:

EC10

Effect conc.:

29 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Remarks on result:

other: 95% CI: 28 - 30 mg/L

Duration:

72 h

Dose descriptor:

NOEC

Effect conc.:

27 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Duration:

72 h

Dose descriptor:

LOEC

Effect conc.:

85 mg/L

Nominal / measured:

meas. (geom. mean)

Conc. based on:

test mat.

Basis for effect:

other: yield

Details on results:

- Exponential growth in the control (for algal test): yes (biomass increased by a factor of 222)
- Observation of abnormalities (for algal test): no difference between the algae growing at the nominal test concentration of 46 mg/L and the algal cells in the control at the end of the test (microscopic examination)
- Uncommon observations concerning the appearance of the media: no

Results with reference substance (positive control):

- Results with reference substance valid? yes
- ErC50 (72 h): 0.74 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 Probit Analysis using linear maximum likelihood regression.
For the determination of the LOEC and NOEC, the average growth rate and yield at the test concentrations were compared to the control values by Dunnett t-test, Williams t-test or Welch t-test, where appropriate.
Statistical analysis was performed using ToxRat Professional®.
The values for the validity criteria of the test were also calculated by the statistic program ToxRat Professional®

87 and 91% of the nominal values were recovered for the test item concentrations of 46 and 100 mg/L at the start of the test, respectively. At the end of the test, 39 and 79% of the nominal values were recovered, respectively.

Table 1: Analytical results.

Nominal test concentration [mg/L]	Mean measured concentration of the test item (geometric mean)
	[mg/L]	% of nominal
4.6	--	--
10	--	--
22	--	--
46	27	58
100	85	85

Table 2: Effect concentrations (0 -72 h) in mg/L.

	Growth Rate	Yield
EC10 95% CI	42 19 -54	29 28 -30
EC50 95% CI	77 68 -81	46 44 -47
NOEC	27	27
LOEC	85	85

Table 3: % Inhibition of growth rate and yield.

Nominal concentration [mg/L]	24 h	48 h	72 h
	% inhibition of growth rate
Control	0	0	0
4.6	-6.4	-0.7	-0.2
10	-4	-0.8	0
22	-2.2	0.3	2.5
46	-3.3	0.8	1.2
100	9.4	27.8	58.4
	% inhibition of yield
Control	0	0	0
4.6	-14.6	-2.9	-0.9
10	-9	-3.1	0
22	-4.6	1.1	12.8
46	-7.3	2.9	6.2
100	19.7	66.7	96.1

Validity criteria fulfilled:

yes

Conclusions:

Growth inhibition of the freshwater green algal species Pseudokirchneriella subcapitata was observed in a 72 h test (OECD 201). The ErC50 (72 h) was 77 mg/L and ErC10 (72 h) = 42 mg/L.

Description of key information

ErC50 (72 h) = 77 mg/L and ErC10 (72 h) = 42 mg/L, based on growth rate of Pseudokirchneriella subcapitata (OECD 201, GLP).

Key value for chemical safety assessment

EC50 for freshwater algae:

77 mg/L

EC10 or NOEC for freshwater algae:

42 mg/L

Additional information

One experimental study is available investigating the toxicity of the substance to aquatic freshwater algae. The study was conducted according to OECD 201 and GLP using Pseudokirchneriella subcapitata as the test species. The test organism was exposed at nominal test item concentrations of 4.6, 10, 22, 46 and 100 mg/L for 72 h. Only the test concentrations 46 and 100 mg/L were analyzed at the beginning and at the end of the test by HPLC/UV, since the remaining concentrations were below the NOEC determined in this test. The analysis indicated that the test item concentrations were within 87 - 91% of nominal at the beginning of the test and dropped to 39 – 79% after 72 h. The biological results were related to the mean measured test item concentrations (geometric mean of concentrations measured at the start and end of the test). Based on growth rate inhibition the test resulted in an ErC50 (72 h) of 77 mg/L and an ErC10 (72 h) of 42 mg/L.