Cocoa, powd., alkalized

Toxicity to aquatic algae and cyanobacteria

Administrative data

Endpoint:

toxicity to aquatic algae and cyanobacteria

Type of information:

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

Adequacy of study:

key study

Study period:

December, 2012 to April, 2013

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

other: Study was conducted by a GLP accredited laboratory using OECD Testing Guideline 201.

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2013

Report date:

2013

Materials and methods

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 201 (Alga, Growth Inhibition Test)

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

Cocoa Powder (High Fat)

IUPAC Name:

Cocoa Powder (High Fat)

Test material form:

solid: particulate/powder

Remarks:

migrated information: powder

Details on test material:

Test substance name: Cocoa Powder (22.5% lipid)
Batch number: BHF092012
Description: Brown solid
Purity: Not applicable – complex mixture
Storage: Room temperature (15 to 30oC), desiccated and protected from light

Test solutions

Vehicle:

yes

Test organisms

Test organisms (species):

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

Details on test organisms:

The test organism, Pseudokirchneriella subcapitata (Strain 278/4), was originally obtained from the Culture Collection of Algae and Protozoa (CCAP) and is a representative species of the freshwater aquatic phytoplankton. This species is recommended for testing in accordance with the OECD regulatory guidelines.
Prior to testing, duplicate starter cultures were prepared and incubated under test conditions to obtain sufficient algal cells in exponential growth and to achieve a starting alga cell density of 1 x 104 cells/mL.

Study design

Test type:

static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

72 h

Test conditions

Test temperature:

The test incubator temperature was set within the range 21 to 24°C and maintained within ± 2°C for the duration of the test. The incubator temperature was recorded continuously using a digital temperature logger.

pH:

At the start of the test, the pH of freshly prepared test media, before and after alga cell inoculation was determined. The pH in each test vessel was also determined at the end of the test.
The OECD guideline states that the pH in the control should not vary by more than 1.5 units during the test. At the end of the test elevated pH values were observed in the control and 1.0 – 3.2 mg/L loading rate WAF test groups. This is an indicator of good algal cell growth, whereby the release of oxygen from the algal cells during normal photosynthetic activity promotes an alkali pH in the surrounding test media. Given that the pH values in the non-inoculated vessels showed similar pH values to that at the start of the test, it was considered that the elevated pH levels in the inoculated vessels was due to good algal cell growth and therefore considered not to affect the validity of the test.

Nominal and measured concentrations:

The definitive test was conducted at nominal test concentrations of 1.0, 3.2, 10, 32, 50 and 100 mg/L loading rate Water Accomodated Fraction (WAF).

Details on test conditions:

All environmental conditions were within those specified in the study protocol.

Reference substance (positive control):

yes

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

27 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

cell number

Effect concentrations 2

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

30 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Effect concentrations 3

Duration:

72 h

Dose descriptor:

EL50

Effect conc.:

88 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Effect concentrations 4

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

cell number

Effect concentrations 5

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

biomass

Effect concentrations 6

Duration:

72 h

Dose descriptor:

NOELR

Effect conc.:

3.2 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

growth rate

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

All validity criteria were met, therefore the test was considered to be valid.

Conclusions:

Based on nominal loading rates, the 72-hour EyL50 (yield) and the 72 hour EbL50 (biomass) and ErL50 (growth rate) values were calculated to be 27, 30 and 88 mg/l loading rate WAF, respectively. The corresponding NOELR values for yield, biomass and specific growth rate after 72 hours were 1.0, 1.0 and 3.2 mg/l loading rate WAF, respectively. Under the conditions of this test, Cocoa Powder (22.5% lipid) was considered to inhibit the growth of Pseudokirchneriella subcapitata at nominal loading rates >1.0 mg/L loading rate WAF. Under the conditions of this test, Cocoa Powder (22.5% lipid) was considered to inhibit the growth of Pseudokirchneriella subcapitata at nominal loading rates >1.0 mg/l loading rate WAF.

Executive summary:

The potential of the test substance to inhibit the growth of algae was determined in accordance with the OECD Guideline for Testing of Chemicals 201. The definitive test was conducted at nominal test concentrations of 1.0, 3.2, 10, 32, 50 and 100 mg/l loading rate Water Accomodated Fraction (WAF). The concentrations used were based on the results of a range finding test, which are not fully reported. A control group was also included. Based on nominal loading rates, the 72-hour EyL₅₀ (yield) and the 72 hour EbL₅₀ (biomass) and ErL₅₀ (growth rate) values were calculated to be 27, 30 and 88 mg/l loading rate WAF, respectively. The corresponding NOELR values for yield, biomass and specific growth rate after 72 hours were 1.0, 1.0 and 3.2 mg/l loading rate WAF, respectively. Under the conditions of this test, Cocoa Powder (22.5% lipid) was considered to inhibit the growth of Pseudokirchneriella subcapitata at nominal loading rates >1.0 mg/l loading rate WAF. The EL₅₀ is considered comparable to the EC₅₀ value calculated from the mass of test substance in the WAF.