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NCEA Proposed Draft Charge to External Reviewers for the IRIS Toxicological Review 

of 1,2,4-Trimethylbenzene (1,2,4-TMB) and 1,3,5-Trimethylbenzene (1,3,5-TMB) 

January 2012 

Introduction 

The U.S. Environmental Protection Agency (EPA) is seeking an external peer review of the 

scientific basis supporting the draft Toxicological Review of 1,2,4-Trimethylbenzene (1,2,4-

TMB) and 1,3,5-Trimethylbenzene (1,3,5-TMB) that will appear on the Agency’s online 

database, the Integrated Risk Information System (IRIS). IRIS is prepared and maintained by the 

EPA’s National Center for Environmental Assessment (NCEA) within the Office of Research 

and Development (ORD). There is currently no entry on the IRIS database for either 1,2,4-TMB 

or 1,3,5-TMB. 

IRIS is a human health assessment program that evaluates qualitative and quantitative risk 

information on effects that may result from exposure to specific chemical substances found in the 

environment. Through the IRIS Program, EPA provides quality science-based human health 

assessments to support the Agency’s regulatory activities. Combined with specific exposure 

information, government and private entities use IRIS to help characterize public health risks of 

chemical substances in site-specific situations in support of risk management decisions. 

The external review draft Toxicological Review of 1,2,4-TMB and 1,3,5-TMB is based on a 

comprehensive review of the available scientific literature on the human and animal health 

effects of 1,2,4-TMB and 1,3,5-TMB, and was developed according to general guidelines for risk 

assessment set forth by the National Research Council (1983) and guidelines and technical 

reports published by EPA (see Preface). This draft IRIS assessment provides an overview of the 

data regarding the toxicokinetics of 1,2,4-TMB and 1,3,5-TMB in humans and animals and 

characterizes the potential hazard posed by 1,2,4-TMB and 1,3,5-TMB exposure for noncancer 

and cancer health effects, including the derivations of a chronic oral reference dose (RfD) and a 

chronic inhalation reference concentration (RfC) for both isomers. Additionally, the draft IRIS 

assessment includes a qualitative characterization of the human cancer potential for both 

isomers. 

Charge Questions 

Below is a set of charge questions that address scientific issues in the draft IRIS Toxicological 

Review of 1,2,4-TMB and 1,3,5-TMB. Please provide detailed explanations for responses to the 

charge questions. EPA will also consider the Science Advisory Board reviewer panel comments 

on other major scientific issues specific to the hazard identification and dose-response 

assessment of 1,2,4-TMB and 1,3,5-TMB. Please consider the accuracy, objectivity, and 

transparency of EPA’s analyses and conclusions in your review. 
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(A) General Charge Questions: 

1. Is the Toxicological Review logical, clear and concise? Has EPA clearly presented and 

synthesized the scientific evidence for noncancer and cancer effects of 1,2,4-TMB and 

1,3,5-TMB? 

2. Please identify any additional peer-reviewed studies from the primary literature that 

should be considered in the assessment of noncancer and cancer health effects of 1,2,4-

TMB and 1,3,5-TMB. 

(B) Toxicokinetics and Pharmacokinetic (PBPK) Modeling 

Data characterizing the toxicokinetics of 1,2,4-TMB and 1,3,5-TMB following inhalation and 

oral exposures in humans and experimental animals supports the development of 

pharmacokinetic (PBPK) models for 1,2,4-TMB. For the purposes of this assessment, the 

Hissink et al. (2007) model, originally describing 1,2,4-TMB toxicokinetics following exposure 

to white spirit (a complex mixture of volatile organic compounds), was modified to calculate 

internal dose metrics following exposure to 1,2,4-TMB alone. Additionally, the model was 

further modified by the addition of an oral route of exposure for use in route-to-route 

extrapolations for the derivation of an oral RfD. 

1. Does the selected PBPK model with EPA’s modifications adequately describe the 

toxicokinetics of 1,2,4-TMB (Appendix A)? Was the PBPK modeling appropriately 

utilized and clearly described? Are the model assumptions and parameters scientifically 

supported and clearly described? Are the uncertainties in the model structure adequately 

characterized and discussed? 

2. The internal dose metric selected for use in the derivation of the RfC and RfD was the 

steady-state weekly average venous blood concentration (mg/L) of 1,2,4-TMB for rats 

exposed to 1,2,4-TMB for 6 h/day, 5 days/week. Please comment on whether the 

selection of this dose metric is scientifically supported and clearly described. If a 

different dose metric is recommended for deriving the RfC, please identify this metric 

and provide scientific support for this choice. Are the uncertainties in the selected dose 

metric adequately characterized and discussed? 

(B) Inhalation Reference Concentration (RfC) for 1,2,4-TMB 

1. A 90-day inhalation toxicity study of 1,2,4-TMB in male rats was selected as the basis 

for the derivation of the RfC (Korsak and Rydzynski, 1996). Please comment on whether 

the selection of this study is scientifically supported and clearly described. If a different 

study is recommended as the basis for the RfC, please identify this study and provide 

scientific support for this choice. 

2. Decreased pain sensitivity (measured as increased latency to pawlick response after hotplate test) in male Wistar rats was concluded by EPA to be an adverse effect on the 

central nervous system and was selected as the critical effect for the derivation of the 
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RfC. Please comment on whether the selection of this critical effect and its 

characterization is scientifically supported and clearly described. If a different endpoint 

is recommended as the critical effect for deriving the RfC, please identify this effect and 

provide scientific support for this choice. 

3. Benchmark dose (BMD) modeling was used in conjunction with dosimetric adjustments 

for calculating the human equivalent concentration (HEC) from a rat and human PBPK 

model (Hissink et al., 2007) to estimate the point of departure (POD) for derivation of the 

RfC. Please comment on whether this approach is scientifically supported and clearly 

described. 

a) Has the modeling been appropriately conducted and clearly described, based on 

EPA’s draft Benchmark Dose Technical Guidance Document (U.S. EPA, 2000b)? 

Has the choice of the benchmark response (BMR) for use in deriving the POD 

(i.e. a change equal to 1 SD of the estimated control mean latency to pawlick 

response) been supported and clearly described? 

4. Please comment on the rationale for the selection of the uncertainty factors (UFs) applied 

to the POD for the derivation of the RfC for 1,2,4-TMB. Are the UFs appropriate and 

clearly described based on the recommendations described in A Review of the Reference 

Dose and Reference Concentration Processes (U.S. EPA, 2002)? If changes to the 

selected UFs are proposed, please identify and provide scientific support for the proposed 

changes. 

(C) Inhalation Reference Concentration (RfC) for 1,3,5-TMB 

Two short-term neurotoxicity studies (Gralewicz and Wiaderna, 2001; Wiaderna et al., 2002) 

and one developmental toxicity study (Saillenfait et al., 2005) following inhalation exposure to 

1,3,5-TMB were identified in the literature and were considered as potential principal studies for 

the derivation of the RfC of 1,3,5-TMB. However, an RfC was not derived for 1,3,5-TMB based 

on these data for reasons described in the Toxicological Review. The available toxicokinetic and 

toxicological databases for 1,2,4-TMB and 1,3,5-TMB indicate several similarities between the 

two isomers. Thus, EPA concluded that given the similarities, including chemical properties, 

toxicokinetics, and potential toxicity between the two isomers, there was sufficient similarity to 

support adopting the RfC for 1,2,4-TMB for both isomers. 

1. Has the rationale for using the RfC for 1,2,4-TMB for both isomers been clearly 

presented? Please comment on whether the approach is scientifically supported. Is the 

scientific justification for not deriving an RfC based on the available data for 1,3,5-TMB 

supported and has it been clearly described? 

(D) Oral Reference Dose (RfD) for 1,2,4-TMB

A route to route extrapolation from inhalation to oral exposure using the modified Hissink et al. 

(2007) PBPK model has been used to derive a chronic oral RfD for 1,2,4-TMB. In order to 
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perform the route-to-route extrapolation, an oral component was added to the model, assuming a 

constant infusion rate into the liver. Specifically, in the absence of chemical-specific 

information an assumption was made that 100% of the ingested 1,2,4-TMB was absorbed by 

constant infusion of the oral dose into the liver compartment. The contribution of first-pass 

metabolism was also evaluated. 

1. The available oral studies for 1,2,4-TMB were determined to be inadequate for derivation 

of an RfD. Please comment on whether the justification for the determination that the 

oral database is insufficient to support the derivation of an RfD for 1,2,4-TMB is clearly 

described in the document and whether there are available data to support the derivation 

of an RfD. 

2. Has the PBPK modeling been appropriately utilized and clearly described? Are the 

model assumptions and parameters scientifically supported and clearly described? Are 

the uncertainties in the model structure adequately characterized and discussed? Please 

comment on whether this approach is scientifically supported and clearly described in the 

document. 

3. Please comment on the rationale for the selection of the uncertainty factors (UFs) applied 

to the POD for the derivation of the RfD for 1,2,4-TMB. Are the UFs appropriate and 

clearly described based on the recommendations described in A Review of the Reference 

Dose and Reference Concentration Processes (U.S. EPA, 2002)? If changes to the 

selected UFs are proposed, please identify and provide scientific support for the proposed 

changes. 

(E) Oral Reference Dose (RfD) for 1,3,5-TMB 

The oral database for 1,3,5-TMB was determined to be inadequate for derivation of an RfD. 

EPA concluded that given the similarities, including chemical properties, toxicokinetics, and 

toxicity between the two isomers, there was sufficient similarity to support adopting the RfD for 

1,2,4-TMB for both isomers. 

1. Has the rationale for using the RfD for 1,2,4-TMB for both isomers been appropriately 

and clearly presented? Please comment on whether the current is scientifically supported 

and clearly described in the document and whether there are available data to support the 

derivation of a 1,3,5-TMB-specific RfD. 

 (F) Carcinogenicity of 1,2,4-TMB and 1,3,5-TMB 

1. Under EPA’s Guidelines for Carcinogen Risk Assessment (U.S. EPA, 2005a; 

www.epa.gov/iris/backgrd.html), the draft Toxicological Review of 1,2,4-TMB and 

1,3,5-TMB concludes that there is “inadequate information to assess the carcinogenic 

potential” of 1,2,4-TMB and 1,3,5-TMB. Please comment on whether this 

characterization of the human cancer potential for 1,2,4-TMB and 1,3,5-TMB is 

scientifically supported and clearly described. 
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2. The draft Toxicological Review of 1,2,4-TMB and 1,3,5-TMB did not derive a 

quantitative cancer risk estimate for 1,2,4-TMB and 1,3,5-TMB due to lack of available 

studies. Are there available data to support the derivation of a quantitative cancer risk 

estimate for 1,2,4-TMB and 1,3,5-TMB? If so, please identify these data.