SRC TR - 12 - 033

HPV3 T OXICITY TEST PLAN REVIEW (DRAFT) Propanoic acid, 2 - methyl - , 3 - (benzo yl ox y) - 2,2,4 - trimethylpent ylester, CASRN 22527 - 63 - 5 Prepared b y: William L . Richards , Teresa Manyin , and Laura Morlacci Chemical, Biological and Environmental Center

SRC, Inc. 7502 Round Pond Road North Sy racuse, NY 13212 Contract No. EP - W - 09 - 027 Task FG007. 2 .009.006 Submitted to: OCSPP / OPPT/RAD (7403M) U.S. Environmental Protection Agency 1200 Pennsylvania Avenue Washington, D.C. 20460 Debra Millig an, Project Officer Meena Sonawane, Work Assignment Manager David Brooks, Alternate Work Assignment Manager

March 9 , 2012Final HPV3 Toxicity Tes t Plan Review CASRN 22527 - 63 - 5

1 **HPV3 Toxicity Test Plan Review Propanoic acid, 2**  **- methyl**  **- , 3**  **- (benzoyloxy)**  **- 2,2,4**  **- trimethylpentyl ester, CASRN 22527**  **- 63**  **- 5**  The test rul e, “Testing of Certain High Pro duction Volume Chemicals; Third Group of Chemicals ” ( 76 FR 65385, October 21 , 2011 ), requires the following toxicity testing for p ropanoic acid, 2 - methyl - , 3 - (benzo yl ox y) - 2,2,4 - trimethylpent ylester, CASRN 22527 - 63 - 5 : C1 (aqu atic toxicity), D (mammalian toxicity

– acute), E1 (bacterial reverse mutation test) , E2 (chromosomal aberration or micronucleus test) and F1 ( mammalian toxicity

– repeated dose/reproduction/developmental) . Eastman Chemical Company ( Eastman ) submitted a t est plan on February

20, 2011 to address the endpoints required by the test rule. The adequacy of the test plan is evaluated below. **1. General Comments:**  The test rule classifies propanoic acid, 2 - methyl - , 3 - (benzoyloxy) - 2,2,4 - trimethylpent ylester, CAS RN 22527 - 63 - 5 as a Class 1 Chemical Substance for which testing must be conducted with material having a purity

of 99% or greater. However, f or toxicity

testing, Eastman proposes the use of a bulk reaction product of the following purity : 60 - 65% propanoic acid, 2 - methyl - , 3 - (benzoyloxy) - 2,2,4 - trimethylpent ylester ( CASRN 22527 - 63 - 5 ), 20 - 25% 1,3 - pentanediol, 2,2,4 - trimethyl - 1,3 - dibenzoate ( CASRN 68052 - 23 - 3 ), and 10 - 15% 2,2,4 - trimethyl - 1,3 - pentanediol diisobuty rate ( CASRN 6846 - 50 - 0 ) . EPA approves Eastman’s plan to conduct a quatic and mammalian toxicity

tests of this mixture (“the bulk reaction product”) based on the following : (1) Eastman states that “The manufacture of benzoate ester for commercial use has been as a bulk reaction and there is no readil y available process to s ynthesize the chemical substance b y direct reaction to achieve the purity required for Class 1 substances. Eastman believes that use of the bulk reaction product which is formulated into finished products for commercial sale will more accurately represent exposure and use of the benzoate ester and avoid a costly and technically

uncertain purification procedure.” (2 ) In commerce, the bulk reaction product that is formed through an esterification reaction between benzoic acid (CASRN 65 - 85 - 0) and 2,2,4 - trimethyl - 1,3 - pentanediol diisobuty rate (CASRN 6846 - 50 - 0) results in three che micals (CASRNs 22527 - 63 - 5, 68052 - 23 - 3 , and 6846 - 50 - 0) . These chemicals, the diesters of 2,2,4 - trimethyl - 1,3 - pentanediol with benzoic acid and 2 - methylpropanoic a cid (isobut yric acid) , can be considered close analogs. (3) T he major constituent of the bulk reaction product is the test rule substance, p ropanoic acid, 2 - methyl - , 3 - (benzo yl ox y) - 2,2,4 - trimethylpent ylester, CASRN 22527 - 63 - 5 . For all toxicity

tests, the final study

report should include the anal yz ed composition of the test substance, including identities and concentrations of major ingredients and of impurities.Final HPV3 Toxicity Tes t Plan Review CASRN 22527 - 63 - 5

2 **2 . Aquatic Toxicity**  Under testing requirement C1 of the test rule, a determination of whether to perform aquatic toxicity testing under Test Group 1 or Test Group 2 is based on a consideration of the measured log Kow for the test substance. EPA approves Eastman ’s plan to condu ct a Chronic T oxicity to Daphnia test of the bulk reaction produ ct using ASTM E 1193 - 97 (Reapproved 2004) if the measured log K ow is ≥ 4.2 or an Acute T oxicity to Daphnia test using ASTM E 729 - 96 (Reapproved 2007) if the measured log K ow i s < 4.2. In addition to daphnid testing , the following studies of the bulk reaction product must be conducted to fulfill testing requirement C1 of the test rule: Toxicity to Plants (Algae) using ASTM E 1218 - 04 (require d for all chemicals regardless of log K ow value) ; and Acute Toxicity to Fish using ASTM E 729 - 96 (Reapproved 2007) if the measured log K ow is < 4.2. Note that Eastman failed to inclu de these required tests in the submitted test plan. **3. Mammalian Toxicity –**  **Acute** EPA approves Eastman ’s p lan to conduct an A cute Oral Toxicity

– Up - and - Down Procedure test of the bulk reaction product using OECD TG 425 to fulfill testing requirement D. **4. Bacterial Reverse Mutation Test** EPA approves Eastman ’s p lan to conduct a Bacterial R everse Mutation T est using 40 CFR 799.9510 of the bulk reaction product to fulfill testing requirement E1 . **5. Chromosomal Aberration/Micronucleus Test** EPA approve s Eastman ’s p lan to conduct an I n vitro Mammalian Chromosomal Aberration T est of the bulk reaction product using 40 CFR 799.9537 to fulfill testing requirement E2. **6. Mammalian Toxicity –**  **Repeated Dose/Reproduction/Developmental** For the following reason s, EPA does not approve Eastman ’s request to fulfill testing requirement F1 “through the use of data already available on the three primary metabolites of the test substance: namel y benz oic acid, 2,2,4 - trimethyl - 1,3 - pentanediol (TMPD) and isobuty rate.”: (1) For the diesters of 2,2,4 - trimethyl - 1,3 - pentanediol (CASRNs 22527 - 63 - 5, 68052 - 23 - 3 and 6846 - 50 - 0) that are present in the bulk reaction product that Eastman proposes for testing for the phy sical/chemical properties, environmental fate, aquatic toxic ity , acute mammalian toxicity

and genotoxicity

endpoints, Eastman ha s not provided data to show that non - enzy

matic or enzy

matic hydrol ys is of these esters will occur fast enough to justify testing of only

the presumptive hydroly sis products/m etabolites for addressing the mammalian repeated - dose, reproductive , and developmental toxicity endpoints . Eastman cites an oral gavage study of diethyleneglycol dibenzoate, which showed “that the ester linkages are completely hydrol yz ed with no evidence of parent compound in theFinal HPV3 Toxicity Tes t Plan Review CASRN 22527 - 63 - 5

3 urine”. However, the rate and extent of non - enz ymatic or enzy matic h ydroly sis of the three diesters present in Eastman ’s bulk reaction product ( CASRNs 22527 - 63 - 5, 68052 - 23 - 3 and 6846 - 50 - 0) may be inhibited compared with diethylenegl yc ol diben zoate because of greater steric hindrance b y the branched 2,2,4 - trimethylpent ylmoiety than b y the diethylene gl ycol moiety. (2) Eastman does not present a rationale to show that data from separate repeated - dose, reproductive , and developmental toxicity s tudies of individual hydrol ys is products/metabolit es of the bulk reaction product would be adequate to characterize toxicity of the mixture of th ese substances expected to occur in vivo . Although data for the individual hydrol ys is products/metabolites are useful to address the overall toxicity of the bulk reaction product, the data for these individual hydroly sis products/metabolites would not address the potential for interactions (e.g., additivity , s yn ergism or antagonism) among these degradat ation produ cts, mono - ester intermediates, and the residual diesters of the bulk reaction product . The likelihood for such interactions could vary with exposure route since the acidic environment of the stomach during oral exposure could result in higher rates of non enzy matic ester hydrol ys is than for exposures by the inhalation or dermal routes. Based on the preceding discussion , testing requirement F1 should be fulfilled by conduc ting a Combined Repeated Dose Toxicity Study with the Reproduction/Developmental Toxic ity Screening T est (40 CFR 799.9365) of the bulk reaction product ; OR the R eproducti on/Developmental Toxicity Screening T est (40 CFR 799.9355) AND the Repeated Dose 28 - Day

Oral Toxicity

S tudy in R odents T est (40 CFR 799.9305) of the bulk reaction product . **Conclusion:** The submitted test plan for toxicity test ing is adequate to satisfy test rule requirement s C1, D, E1, E2, and F 1 with the exceptions and stipulations listed above.