Hydrocarbons, C10, aromatics, >1% naphthalene

Toxicity to aquatic algae and cyanobacteria

Link to relevant study record(s)

Reference

Reference 1

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

read-across from supporting substance (structural analogue or surrogate)

Adequacy of study:

key study

Study period:

20-Feb-2001 to 23-Feb-2001

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The reliability rating is a 1 because the study followed a standard guideline, which describes a procedure designed to evaluate this endpoint, and the results were reviewed for reliability and assessed as valid. The study was also conducted under GLP.

Justification for type of information:

A discussion and report on the read across strategy is given as an attachment in IUCLID Section 13

Reason / purpose for cross-reference:

read-across: supporting information

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Details on sampling:

Samples of each treatment WAF and the control were taken on Day 0 prior to the addition of algae and at termination (composite of replicates) with no headspace. The samples were stored under refrigeration until analyzed.

Vehicle:

no

Test organisms (species):

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

Details on test organisms:

Algal cells were obtained from in-house culture.

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Post exposure observation period:

None

Test temperature:

Mean test temperature: 24.2°C.

pH:

The pH was 7.7 in all of the test solutions at test initiation and ranged from 7.8 to 8.8 at test termination.

Dissolved oxygen:

not applicable

Salinity:

freshwater

Nominal and measured concentrations:

Due to the relatively complex nature and limited water solubility of the test substance, the following exceptions to the guideline apply for this study: The concentration of the test substance in solution was not determined prior to use. Test substance analysis was performed on samples of the WAFs at the start of the test (day 0) and at termination (day 3). The initial concentration of the test substance was not maintained at 80% in the 2.2 mg/L and 7.6 mg/L treatments throughout the test (this may be due to biological activity or physical processes in the test chambers). It was deemed more appropriate to prepare individual treatment solutions by adding the test substance to dilution water and removing the WAF of each mixture for testing than to prepare dilutions of a stock solution. None of the above exceptions are believed to have affected the outcome, integrity, or quality of the study.

Details on test conditions:

Individual Water Accommodated Fractions (WAF's) were prepared for each treatment. The test substance was added to 4.4-4.5 L of algal nutrient medium augmented with sodium bicarbonate in glass aspirator bottles (capacity 4.5 L). The solutions were mixed for approximately 23.5 hours using a 6.5% vortex (of the static liquid depth). The test solutions were removed through the outlet at the bottom of each mixing vessel into 12 replicates of 140 mL in 125 mL Erlenmeyer flasks (no headspace) containing two 14mm glass spheres to facilitate mixing. The test chambers were inoculated with algae (1.0 x 104 cells/mL) and were sealed with ground glass stoppers. Cell density was determined using a hemacytometer. Three replicates were sacrificed daily for cell density determination. The test chambers were placed on shaker tables (110 rpm) to keep the algae in suspension. The test was performed under static conditions with no aeration. The algae were cultured in-house from 5 day old stock cultures in log phase growth.

Continuous light: intensity was 7840 to 8498 Lux.

Reference substance (positive control):

no

Key result

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

7.9 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Remarks on result:

other: 95% CI = 4.1 to >25

Key result

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

0.22 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

3.8 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Key result

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

0.22 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Reported statistics and error estimates:

The EbL50, ErL50 and confidence intervals for inhibition of growth/growth rate slope were determined by a probit regression calculation of the probit of the growth inhibition/growth rate slope vs the log of the loading rate and associated confidence intervals based on the methods of Finny (1971). Calculations were based on the PROC PROBIT procedure in SAS (2002). The NOELR for the EbL50 and ErL50 was based on Duncan's (1975) Multiple Range test and Dunnett's (1964) test, determined from the GLM procedure of SAS (2002). The Shapiro-Wilk (1965) test for normality was used to test if the assumption of normality of the residuals was met; since the residuals were normally distributed the NOEC was based on the estimated values.

1. Finney, D.J. 1971. Probit Analysis, 3rd Edition, London: Cambridge University Press.
2. SAS Version 8, SAS Institute, Inc., Cary, NC. 2002.
3. Duncan, D.B. 1975, "t-Tests and Intervals for Comparisons Suggested by the Data", Biometrics, 31, 339-359.
4. Dunnett, C. 1964, "New Tables for Multiple Comparisons With A Control", Biometrics, Vol 20, No. 3, pg 482-491.
5. Shapiro, S.S. and Wilk, M.B. 1965,"n analysis of variance test for normality (complete samples)" Biometrika, 52, pg 591-611.

Effects on growth rate (r) based upon actual loading rates:
72 hr ErL50 (loading rate) = 7.9 mg/L (4.1 - >25* mg/L)
72 hr NOELR (loading rate) = 0.22 mg/L

Effects on biomass (b) based upon actual loading rates:
72 hr EbL50 (loading rate) = 3.8 mg/L (CNC)
72 hr NOELR (loading rate) = 0.22 mg/L

Effects on growth rate (r) based upon measured concentrations:
72 hr ErC50 (measured conc.) = 0.42 mg/L (0.38 - 48 mg/L)
72 hr NOEC (measured conc.) = 0.07 mg/L

Effects on biomass (b) based upon measured concentrations:
72 hr EbC50 (measured conc.) = 0.29 mg/L (<0.07* - >0.81* mg/L)
72 hr NOEC (measured conc.) = 0.07 mg/L

Values in parenthesis are 95% confidence intervals.
* Confidence interval exceeded the highest or lowest loading rate or concentration tested.
CNC = Could Not Calculate 95% confidence intervals.

The analytical method used was headspace gas chromatography with flame ionization detection.

Summary of In-Life observations - % Inhibition

Loading Rate (mg/L)	Control	0.22	0.69	2.2	7.6	25
Meas. Conc.  (mg/L)	0	0.07	0.18	0.32	0.43	0.81
Based on Growth Rate
72 hours	N/A	0.7	11	29	48	99
Based on Biomass
72 hours	N/A	0.9	35	62	80	98

Validity criteria fulfilled:

yes

Conclusions:

Water accommodated fractions of hydrocarbons, C10-C13, aromatics, >1% naphthalene, produced a freshwater 72-hour EL50 value for the green alga (Pseudokirchneriella subcapitata formerly Selenastrum capricornutum) of 3.8 mg/l and 7.9 mg/l, based on reduction in biomass and growth rate, respectively. The 72-hour NOELR (No Observed Effect Loading Rate) value for Pseudokirchneriella subcapitata is 0.22 mg/l based on both growth rate and biomass.

Executive summary:

Water accommodated fractions of hydrocarbons, C10-C13, aromatics, >1% naphthalene, produced a freshwater 72-hour EL50 value for the green alga (Pseudokirchneriella subcapitata formerly Selenastrum capricornutum) of 3.8 mg/l and 7.9 mg/l, based on reduction in biomass and growth rate, respectively. The 72-hour NOELR (No Observed Effect Loading Rate) value for Pseudokirchneriella subcapitata is 0.22 mg/l based on both growth rate and biomass.

Reference 2

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

experimental study

Adequacy of study:

key study

Study period:

1993

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: see 'Remark'

Remarks:

The study was given a reliability rating of 1 because it followed a standard test guideline, which describes a procedure to evaluate this endpoint, and the results were reviewed for reliability and assessed as valid. The study was also conducted under GLP.

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes

Analytical monitoring:

yes

Vehicle:

no

Details on test solutions:

Individual treatment concentrations were prepared as water accommodated fractions (WAF). Control and dilution water were laboratory prepared algal nutrient medium. Test substance was mixed with dilution water in sealed vessels for approximately 24 hours, and the mixtures were allowed to settle for approximately 1 hour prior to drawing off the aqueous phase for testing.

Test organisms (species):

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

Details on test organisms:

Algal cells obtained from testing laboratory cultures. Original algal cultures obtained from American Type Culture Collection (ATCC 22662).

Test type:

static

Water media type:

freshwater

Limit test:

no

Total exposure duration:

72 h

Test temperature:

Temperature was 22 to 24 Deg C.

pH:

The pH ranged from 7.2 to 7.7 at test initiation and 7.2 to 10.2 at test termination.

Dissolved oxygen:

Not applicable

Salinity:

freshwater

Nominal and measured concentrations:

Nominal loading rates were 0.1, 0.4, 1.0, 3.0, and 10 mg/L. Only the 0 and 72 hours samples from the 3.0 and 10 mg/L treatments were analyzed yielding mean measured concentrations of 3.2 and 8.2 mg/L, respectively.

Details on test conditions:

Lighting was continuous at approximately 3000 lux.

Test vessels were sealed 300 ml conical flasks that were completely filled with test solution. The treatment and control flasks were inoculated with algal cells to yield initial concentrations ranging from 3200 to 5000 cells/ml. Uninoculated flasks were used to determine particle counts without algal cells. Biomass was calculated as area under the growth curve.

Reference substance (positive control):

no

Key result

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

> 1 - < 3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Key result

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

> 1 - < 3 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Key result

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

ca. 1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Key result

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

ca. 1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Details on results:

See Tables Below

Reported statistics and error estimates:

EL50 values estimated by visual inspection of data to determine range of loading rates, which bracketed a 50% reduction in biomass and growth rate. ANOVA and Williams test used to determine NOELs.

Nominal Loading	72 h
(mg/L)	% Inhibition
Control	N/A
0.1	-14.0
0.4	-27.0
1.0	-8.0
3.0	97.0
10.0	100.0

Nominal Loading	Conc.	Conc.
(mg/L)	0 hr (cells/mL)	72 hr (cells/mL)
Control	3.2xE3	3.7xE5
0.1	4.6xE3	4.2xE5
0.4	5.3xE3	4.7xE5
1.0	5.4xE3	4.0xE5
3.0	5.2xE3	9.3xE5
10.0	3.5xE3	0.0

Validity criteria fulfilled:

yes

Conclusions:

The hydrocarbons, C10, aromatics, >1% naphthalene, freshwater 72-hr EL50 value for Pseudokirchneriella subcapitata (formerly Selenastrum capricornutum) is 1.0 to 3.0 mg/l based on both growth rate and biomass. The hydrocarbons, C10, aromatics, >1% naphthalene, freshwater 72-hr NOELR (No Observed Effect Loading Rate) value for Pseudokirchneriella subcapitata is approximately 1.0 mg/l based on both growth rate and biomass.

Executive summary:

The hydrocarbons, C10, aromatics, >1% naphthalene, freshwater 72-hr EL50 value for Pseudokirchneriella subcapitata (formerly Selenastrum capricornutum) is 1.0 to 3.0 mg/l based on both growth rate and biomass. The hydrocarbons, C10, aromatics, >1% naphthalene, freshwater 72-hr NOELR (No Observed Effect Loading Rate) value for Pseudokirchneriella subcapitata is approximately 1.0 mg/l based on both growth rate and biomass.

Description of key information

There is data available for this substance. Additionally, key data is available for the structural analogue Hydrocarbons, C10-C13, aromatics, >1% naphthalene. The data for this substance is presented in the dossier. The data is read across to this substance based on analogue read across and a discussion and report on the read across strategy is provided as an attachment in IUCLID Section 13.

Water accommodated fractions of hydrocarbons, C10-C13, aromatics, >1% naphthalene, presented a 72-hour EL50 value for Pseudokirchneriella subcapitata of 3.8 mg/l and 7.9 mg/l based on biomass and growth rate, respectively. The 72-hour NOELR (No Observed Effect Loading Rate) value for Pseudokirchneriella subcapitata is 0.22 mg/l based on both growth rate and biomass.

 

Water accommodated fractions of Hydrocarbons, C10, aromatics, >1% naphthalene, presented a 72-hr EL50 value for Pseudokirchneriella subcapitata of between 1.0 to 3.0 mg/l based on both growth rate and biomass. The 72-hr NOELR (No Observed Effect Loading Rate) value for Pseudokirchneriella subcapitata is approximately 1.0 mg/l based on both growth rate and biomass (Shell, 1993c).

 

Water accommodated fractions of Hydrocarbons, C10, aromatics, >1% naphthalene, presented a 72-hr EL50 value for Pseudokirchneriella subcapitata of 11 mg/l and 8.5 mg/l based on growth rate and biomass, respectively. The 72-hr NOELR (No observed Effect Loading Rate) value for Pseudokirchneriella subcapitata is 2.5 mg/l based on both growth rate and biomass (Exxon, 2006d).

Additional information

Three study reports were available and input as endpoint records.

The study from Exxon (2006) examined the toxicity of the test substance Hydrocarbons, C10-C13, aromatics, >1% naphthalene, to Pseudokirchneriella subcapitata. Algal cells from in-house culture were exposed to nominal treatment levels of control, 0.22, 0.69. 2.2, 7.6 and 25 mg/l for a total exposure duration of 72 hours. Water accommodated fractions of hydrocarbons, C10-C13, aromatics, >1% naphthalene, presented a 72-hour EL50 value for Pseudokirchneriella subcapitata of 3.8 mg/l and 7.9 mg/l based on biomass and growth rate, respectively. The 72-hour NOELR (No Observed Effect Loading Rate) value for Pseudokirchneriella subcapitata is 0.22 mg/l based on both growth rate and biomass.

 

The study from Shell (1993c) examined the toxicity of the test substance Hydrocarbons, C10, aromatics, >1% naphthalene, to Pseudokirchneriella subcapitata. Algal cells obtained from testing laboratory cultures were exposed to nominal treatment levels of control, 0.1, 0.4, 1.0, 3.0, and 10 mg/l for a total exposure duration of 72 hours. Water accommodated fractions of Hydrocarbons, C10, aromatics, >1% naphthalene, presented a 72-hr EL50 value for Pseudokirchneriella subcapitata of between 1.0 to 3.0 mg/l based on both growth rate and biomass. The 72-hr NOELR (No Observed Effect Loading Rate) value for Pseudokirchneriella subcapitata is approximately 1.0 mg/l based on both growth rate and biomass.

 

The study from Exxon (2006d) examined the toxicity of the test substance Hydrocarbons, C10, aromatics, >1% naphthalene, to Pseudokirchneriella subcapitata. Algal cells from in-house culture were exposed to nominal treatment levels of control, 0.9, 2.5, 4.9, 12.0 and 30.0 mg/l for a total exposure duration of 72 hours. Water accommodated fractions of Hydrocarbons, C10, aromatics, >1% naphthalene, presented a 72-hr EL50 value for Pseudokirchneriella subcapitata of 11 mg/l and 8.5 mg/l based on growth rate and biomass, respectively. The 72-hr NOELR (No Observed Effect Loading Rate) value for Pseudokirchneriella subcapitata is 2.5 mg/l based on both growth rate and biomass.