SRC TR-10-061 

HPV1 TEST RULE STUDY – DATA ADEQUACY ASSESSMENT 

Methyl Heptanone, CAS No. 409-02-9 

Acute Fish Toxicity 

Prepared by: 

Kendra Moran, Ashlee Aldridge, 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 Work Assignment Manager 

March 5, 2010 

 
Draft HPV1 Test Rule – Data Adequacy – Acute Toxicity to Fish CAS No. 409-02-9

HPV1 Test Rule – Data Adequacy Assessment 

Methyl Heptanone, CAS No. 409-02-9 

Acute Toxicity to Fish 

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 22202 
Draft HPV1 Test Rule – Data Adequacy – Acute Toxicity to Fish CAS No. 409-02-9

PREPARATION OF THE DATA ADEQUACY ASSESSMENT

This document contains the review of a toxicity 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 guides, the study was reviewed for adequacy and 

evaluated for compliance with the test guideline. (The applicable test guideline 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 assigned a Klimisch reliability code. If the study 

was determined to be inadequate, a basis for that conclusion is supplied. For studies already 

assigned Klimisch codes, an explanation is provided if the reviewer’s assessment of data 

adequacy differed from the pre-assigned Klimisch code. 

A detailed evaluation of the test report and study conduct is provided at the end of the 

summary document in a section entitled “Review 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 Procedures, Good Laboratory Practice Standards, and 

Quality Assurance procedures, as well as the use of appropriate controls and test substances. 

Any deficiencies in the descriptions of the study method, test system, animals studied, physical 

conditions, dosage information, individual 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. 

1 
Draft HPV1 Test Rule – Data Adequacy – Acute Toxicity to Fish CAS No. 409-02-9

ACUTE TOXICITY TO FISH 

Title: Fish Acute Toxicity Test 

This study is: Adequate Not Adequate 

TEST SUBSTANCE 

 

Identity: Methyl Heptanone, CAS No. 409-02-9 

Remarks: The test substance, Baton Rouge MEK Heavy Ketone Heart Cut C8 Ketone Fraction, 

was characterized as a complex mixture. 

METHOD

Method/guideline followed: OECD TG 203 (1992) 

Test type: Static Renewal (24-hr renewal period) 

GLP compliant?: Yes 

Year study was performed: 2007 

Test species: Oncorhynchus mykiss. Rainbow trout used in this study were obtained from the 

Thomas Fish Company (Anderson, CA Lot 512, Receipt date: 31-Oct-07). A test initiation fish 

were approximately 56 days post hatch, fish mean weight was 0.749 g, and mean total length 

was 4.5 cm. Total loading was 0.666 g of fish/L test solution. 

Analytical monitoring: Yes 

Exposure duration: 96-hr

Concentrations tested: 0 (control), 4.2, 8.5, 20, 45, and 101 mg/L test substance water 

accomadated fractions (WAF), which corresponded to mean measured concentrations of 0 

(control), 3.2, 5.4, 13, 36, and 103 mg/L.

Test method/conditions remarks: Individual Water Accommodated Fractions (WAF's) were 

prepared for each treatment. The test substance was added to 12 L of reconstituted water in glass 

aspirator bottles (capacity 13.5 L). The solutions were mixed for 23 to 24 hours using a 5% 

vortex (of the static liquid depth). The test solution was removed through the outlet at the 

bottom of each mixing vessel (following a one hour settling period) into two replicates of 

approximately 4.5 L in 4.5 L size aspirator bottles (no headspace). Four fish were added to each 

replicate and the replicates were closed with foil covered stoppers. Daily renewals were 

performed by removing approximately 90% of the test solution through the outlet at the bottom 

of the aspirator bottle and refilling with fresh solution. The fish were not fed during the study. 

Mean test temperature ranged from 13.8 - 14.9°C. Diurnal light was approximately 16 hours 

2 
Draft HPV1 Test Rule – Data Adequacy – Acute Toxicity to Fish CAS No. 409-02-9

light and 8 hours dark with 433 to 438 Lux during full daylight periods. Dissolved oxygen 

ranged from 4.5 to 9.3 mg/L and pH ranged from 6.3 to 7.9 during the study. Water hardness was 

92 mg/L as CaCO3. 

Due to the complex nature and limited water solubility of the test substance, the following 

exceptions to the guideline apply for this study: 1) The concentration of the test substance in 

solution was not determined prior to use; 2) 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; 3) The fish used in 

the study were held/acclimated for 10 days instead of at least 12 days as stated in the protocol; 

and 4) There were no culls or mortality during acclimation and the fish appeared healthy and 

normal. This is not believed to have adversely impacted the study. 

RESULTS

Endpoint value(s): The 96-hour LC50 value for rainbow trout was 68 mg/L based on WAF 

loading rates and 61 mg/L based on mean measured concentrations. The maximum measured 

concentration causing no mortality after 96-hours was 36 mg/L, which corresponded to a 

maximum WAF loading rate of 45 mg/L. The minimum measured concentration causing 100% 

mortality after 96-hours was 103 mg/L, which corresponded to a minimum WAF loading rate of 

101 mg/L. 

Results remarks: Sub-lethal effects (e.g., loss of equilibrium and dark pigmentation) were 

observed in all fish exposed to 36 mg/L test substance (mean measured) by the 96-hour 

observation. No sub-lethal effects were observed in lower treatment concentrations, and 100% 

mortality was observed in fish exposed to 103 mg/L test substance (mean measured). 

CONCLUSIONS

After Oncorhynchus mykiss were exposed to WAFs prepared from Baton Rouge MEK Heavy 

Ketone Heart Cut - C8 Ketone Fraction for 96-hours, the LL50 based on WAF loading rates was 

68 mg/L and the LC50 based on mean measured concentrations was 61 mg/L. 

STUDY RELIABILITY

[1] Reliable without restrictions. 

REFERENCE

ExxonMobil Biomedical Sciences, Inc. 2008. FISH, ACUTE TOXICITY TEST on Baton Rouge 

MEK Heavy Ketone Heart Cut - C8 Ketone Fraction. Study # 0700858. 

3 
Review of TSCA Section 4 Final Study Report 

CITB Project Manager: Catherine Roman 

Chemical Name: Methyl Heptanone 

CAS Registry Number: 409-02-9 

Report Title: Fish Acute Toxicity Test 

Test Sponsor: ExxonMobil Chemical Company 

Testing Lab: ExxonMobil Biomedical Sciences Inc. 

Test Standard: ASTM E 729

 

Date Study Received by EPA: 06/23/09 

Date Study Due to EPA: 05/17/07 

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. 

 [D] 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 inadequately describes the test system. 

 
[D] 11. The final study report inadequately 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 summary 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 risk 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/Branch/Division/Office: Kendra Moran/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 met, 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).