SRC TR-10-067

HPV1 TEST RULE STUDY – DATA ADEQUACY ASSESSMENT

Dibromomethane, CAS No. 74-95-3

Acute Toxicity to Daphnia

Prepared by:

Ashlee Aldridge, Justine Von Runnen, and Joan D. Garey

Environmental Science Center
SRC, Inc.

7502 Round Pond Road North Syracuse, NY 13212

Contract No. EP-W-09-027

Task FG007.0.009.004

Submitted to:

OPPT/RAD/HPVCB (7403M) U.S. Environmental Protection Agency 1200 Pennsylvania Avenue

Washington, D.C. 20460

Elizabeth Margosches, Project Officer Meena Sonawane, Work Assignment Manager David Brooks, Alternate Wo rk Assignment Manager

March 5, 2010HPV1 Test Rule – Data Adequacy Assessment

Dibromomethane, CAS No. 74-95-3

Acute Toxicity to Daphnia

March 2010

Submitted to: High Production Volume Chemicals Branch

Risk Assessment Division

Office of Pollution Prevention and Toxics Environmental Protection Agency

WAM: Meena Sonawane

Alternate WAM: David Brooks 1200 Pennsylvania Avenue, NW

Washington, DC 20460-0001

Prepared By:

SRC, Inc.

Environmental Science Center 2451 Crystal Drive, Suite 804 Arlington, VA 22202Draft HPV1 Test Rule – Data Adequacy – Acute Toxicity to Daphnia CAS No. 74-95-3

PREPARATION OF THE DAT A ADEQUACY ASSESSMENT

This document contains the review of a toxi city study conducted in response to the first

HPV TSCA Section 4 Test Rule (71 FR 13708, March 16, 2006). Using the applicable test guideline and professional judgment as guide s, the study was reviewed for adequacy and evaluated for compliance with the test guideline. (T he applicable test guid eline is not always the test guideline required by the test rule, because some test sponsors requested and received approval from EPA to use a different guideline .) The study was summarized according to a standard format, assessed for adequacy, and assi gned a Klimisch reliability code. If the study was determined to be inadequate, a basis for th at conclusion is supplie d. For studies already assigned Klimisch codes, an explanation is prov ided if the reviewer’s assessment of data adequacy differed from the pre-assigned Klimisch code.

A detailed evaluation of the test report a nd study conduct is provided at the end of the summary document in a section entitled “Revie w of TSCA Section 4 Final Study Report”. Any deviations from the applicable test guideline are described. The study is also evaluated for compliance with Standard Operating Procedur es, Good Laboratory Prac tice Standards, and Quality Assurance procedures, as well as the use of appropriate controls and test substances. Any deficiencies in the descri ptions of the study method, test system, animals studied, physical conditions, dosage information, indi vidual animal data, calculations, and statistical analyses are described, and internal inconsistencies within the study report are identified. An overall assessment is provided for each of the following: study conduct, data reliability, and data utility.

1Draft HPV1 Test Rule – Data Adequacy – Acute Toxicity to Daphnia CAS No. 74-95-3

ACUTE TOXICITY TO DAPHNIA

Title: Dibromomethane: Acute Toxicity to Daphnia magna

This study is: Adequate Not Adequate

TEST SUBSTANCE

Identity: Dibromomethane, CAS No. 74-95-3

Remarks: Clear colorless liquid; Pu rity: 99.4%; individual samples of dibromomethane were supplied by both co-sponsors. These two test ma terials were then mixed in equal proportions and the resulting mixture anal yzed to determine its purity.

METHOD

Method/guideline followed: US CFR Title 40 Part 797 Section 1300: US EPA Pesticide Assessment Guideline, Sub-Division E Section 72 -2, US EPA Draft Ecol ogical Effects Test Guideline OPPTS 850.1010, ASTM E 729-96 and th e OECD Guidelines for Testing of Chemicals (April 2004) No 202, “ Daphnia sp, Acute Immobilization Te st” referenced as Method

C.2 of Commission Directive 92/ 69/EEC (which constitutes Annex V of Council Directive 67/548/EEC).

Test type: Acute toxicity to Daphnia

- Static

GLP compliant?: Yes

Year study was performed: 2006

Test species: Daphnia magna ; derived from in-house laboratory cultures; 1 st instar

Analytical monitoring: Yes

Exposure duration: 48 hours

Concentrations tested: Preliminary range-finding test: 0 (control), 1.0, 10 and 100 mg/L (nominal)

Definitive test : 0 (control), 10, 18, 32, 56 and 100 mg/L (nominal)

Test method/conditions remarks: The test concentrations to be used in the definitive test were determined by a preliminary range-finding test. Based on the results of the range-finding test, the following test concentrati ons were assigned to the definitive test: 10, 18, 32, 56 and 100 mg/L. A range of five test concentrations was used as it was considered that both a 48-hour EC 50 and a No Observed Effect Concentration (NOEC) could be achieved. At the start of the study, 10 daphnids were placed in each test and contro l vessel at random, in the test preparations. Duplicate test vessels were used for each test and control group.

2Draft HPV1 Test Rule – Data Adequacy – Acute Toxicity to Daphnia CAS No. 74-95-3

The test was conducted using glass-stoppered coni cal flasks with minimal headspace due to the

volatile nature of the test material. Due to th e volatile nature of the test material, it was considered that the use of dynamic, continuous fl ow test conditions was unsuitable as significant losses due to volatility may occur.

Statistical Analysis: The EC 50 values and associated confiden ce limits at 48 hours and the slope of the response curve and its standard error were calculated by the maximum-likelihood probit method. Probit analysis is used where two or more partial responses to exposure are shown.

Culture conditions: dechlori nated laboratory tap water Water temperature: 18.5 – 18.8 oC Diet: algae ( Chlorella sp.)

Test water: dechlorinated labor atory tap water prepared by passa ge through an activated carbon filter and partly softened, giving wate r with a total hardness of 140 mg CaCO 3 /L.

Test water ready for use:

Particulate matter conc entration: 0.50 mg/L Total water hardness: 114 mg CaCO 3 /L Alkalinity: 87 mg CaCO 3 /L Conductivity: 428 μS/cm Total organic carbon content: 2.5 mg/L pH: 7.7

Test chambers: 300 mL glass-stoppered conical flasks Photoperiod: 16:8 hours light-dar k regime with 20-minute dawn and dusk transition periods

Volume of water: 300 mL

Aeration: None Test temperature: 19.6 – 20.4 oC pH during the test: 7.7-7.9

Dissolved O 2 during test: 8.4-8.9 mg/L Loading rate: ~33 daphnids/L

Observation time: 24 and 48 hours Endpoints investigat ed: immobilization

Preparation of the test item: the test material wa s dissolved directly in dechlorinated tap water.

RESULTS

Endpoint value(s): The 48-hour EC 50 = 66 mg/L; the NOEC was 32 mg/L.

Results remarks : The 48-hour EC 50 value is based on nominal test concentrations. The slope

and standard error of the response curve at 48 hours was 9.4 (SE = 2.1). Due to the unsuitable nature of the data it was not possible to calcu late the slope and error of response curve at 24

3Draft HPV1 Test Rule – Data Adequacy – Acute Toxicity to Daphnia CAS No. 74-95-3

hours. Test concentrations in excess of 100 mg/L were not used for the test as this is not a requirement of the test guidelines. It wa s considered that the absence of an EC 50 value at 24 hours would not affect the integrity of the study as it is the 48-hour EC 50 value which is used for classification and labelling purposes.

Range-finding Test: No immobilization was observed at the test concentrations of 1.0 and 10 mg/L after 24 and 48 hours. However, immobilization was observed at 100 mg/L. Based on this information test concentrations of 10, 18, 32, 56 and 100 mg/L were selected for the definitive

test.

The results of the definitive test showed the lowe st test concentration resulting in 100% mortality to be greater than 100 mg/L. The No Observed Effect Concentra tion after 24 and 48 hours exposure was 32 mg/L. The No Observed Effect Concentration is based upon zero

immobilization at this concentration.

The test media preparations were observed to be clear colorless soluti ons throughout the duration of the test. Analysis of the test preparat ions at 0 and 48 hours showed measured test concentrations to range from 83 to 115% of nomi nal values and it was considered justifiable to

calculate the EC 50 values in terms of the nominal test concentrations only.

CONCLUSIONS

The acute toxicity of the test materi al to the freshwater invertebrate Daphnia magna has been investigated and gave a 48-hour EC 50 value of 66 mg/L with 95% confidence limits of 58 – 76 mg/L. The No Observed Effect Concentration at 48 hours was 32 mg/L.

STUDY RELIABILITY

[1] Reliable without restrictions

REFERENCE

EPA-HQ-OPPT-2005-0033-0259.2, Durward et al . (2007) Dibromomethane: Acute Toxicity to Daphnia magna , SPL Project Number 0466/0264, 1-34 pgs . Sponsor: Bromine Compounds Ltd. and Albemarle Corporation Hea lth, Safety and Environment.

4Review of TSCA Section 4 Final Study Report

CITB Project Manager: Catherine Roman

Chemical Name: Dibromomethane

CAS Registry Number: 74-95-3

Report Title: Acute Toxicity to Daphnia magna

Test Sponsor: Bromine Compounds Ltd. and Albemarle Corporation

Testing Lab: Safepharm Laboratories Limited

Test Standard: ASTM E 729

Date Study Received by EPA: July 12, 2007 Date Study Due to EPA: May 17, 2007

Special Instructions (e.g., regarding approved protocol modifications, etc.):

Part I. Test Report and Study Conduct

Enter one of the following codes into each of the checkboxes 1 through 13: Agree [ A ], Disagree [ D ], Not Applicable [ NA ], or Cannot Determine [ C ], and add comment, below, as needed

[D] 1. The final study report indicates evidence of gross, unauthorized or undocumented deviation from the EPA test standard requirements, including “shall,” “must,” or “will” violations. List significant test standard deviations and “shall,” “must,” or “will” violations in comments below.

[D] 2. The final study report indicates evidence of deviations from the “should” statements in the study protocol that may have a bearing on the study conclusions. List these in comments below.

[NA] 3. Approved modifications of test standard or testing protocol (CITB PM describe or attach) were not followed or are inadequately documented.

[D] 4. There is evidence that laboratory Standard Operating Procedures (SOPs) were inadequately followed.

[D] 5. The Statement of Quality Assurance (QA) inspections, their dates, and findings were missing or not signed. There is evidence that QA procedures may be inadequate.

[D] 6. There is evidence that Good Laboratory Practice Standards (GLPS) were inadequately followed.

[D] 7. The test, control, or reference substances are different from that specified in the section 4 test rule or

Enforceable Consent Agreement (ECA).

[D] 8. Animal identification is inadequate or unclear; use of unhealthy animals or other factors (e.g., deviations from temperature, lighting, dosing) are apparent.

[D] 9. The final study report inadequately describes the study method.

[D] 10. The final study report in adequately describes the test system.

[D] 11. The final study report inadeq uately describes the dosage information.[D] 12. The final study report does not accurately reflect the individual animal data (or that of other discrete observations pertinent to the type of study conducted), or individual data were not provided.

[D] 13. The final study report inadequately describes the calculations, interpolations, data transformations, statistical analyses, or modeling parameters used.

Part II. Conclusions

Place a check mark beside the summar y statement(s) that apply to this review and add comments below as appropriate

Part A. To be filled out by Reviewer

Study Conduct 1. The study was well-conducted. [X] 2. The study was generally well-conducted, with some minor [ ] deficiencies (list in notes below). 3. The study conduct was significantly deficient or questionable, (give [ ] reasons in notes below).

Data Reliability 4. The study results are precise and accurate. [X] 5. Appropriate methods were used to determine [X] statistical significance. 6. If repeated, the test results would be expected to be the [X]

same.

Data Utility 7. The data are useful for ri sk assessment for the endpoint tested. [X] 8. The data are useful for some other purpose (e.g., dose-setting). [ ] 9. The data are not useful for any purpose (describe in notes below). [ ]

Reviewer Comments (continue on separate page, if needed):

Reviewer Name/Title/Bran ch/Division/Office: Ashlee Aldridge/SRC, Inc.

Reviewer Signature: David Brooks/HPVCB/RAD/OPPTS

Part B. To be filled out by CITB PM

Tiered Testing Decision 1. The tiered testing trigger was me t, additional testing is indicated. [ ] (when applicable) 2. The trigger for additional (tiered) testing was not met. [ ]

Study Acceptability 3. The study is acceptable. [ ] 4. The study may not be acceptable [ ] (provide rationale in memo to CITB chief).