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UNITED STATES ENVIRONMENTAL PROTECTION AGENCY

WASHINGTON, D.C. 20460 

OFFICE OF PREVENTION, PESTICIDES

 AND TOXIC SUBSTANCES

MEMORANDUM

Date: March 15, 2011

SUBJECT: Bromoxynil. Benchmark Dose Analysis on Supernumerary Ribs in Three 

Developmental Toxicity Studies 

PC Code: 035301, 035302 DP Barcode: D374609

Decision No.: 425581 Registration No.: 264-1023

Petition No.: 9F7678 Regulatory Action: Section 3 Registration

Risk Assessment Type: Single Chemical/

Aggregate

Case No.: 2070

TXR No.: 0055648 CAS No.: 1689-84-5, 1689-99-2

MRID No.: 00116558, 40466802 40 CFR: 180.324

FROM: Myron S. Ottley, Ph.D., Senior Biologist

Risk Assessment Branch III (RAB3)

Health Effects Division (HED) (7509P)

THROUGH: Paula Deschamp, Branch Chief 

Risk Assessment Branch III (RAB3)

Health Effects Division (HED) (7509P)

TO: Bethany Benbow, RM#23

Registration Division (7505P)

I. BACKGROUND

In the previous risk assessment (D246944.mem, B. Madden 6/24/1998), dose-response data on 

supernumerary ribs from three studies were considered together in hazard characterization and 

hazard identification. Specifically, a NOAEL of 4 mg/kg/day from Rubin and Nyska (1982) 

was combined with a LOAEL of 5 mg/kg/day from Baker & Cropping (1981). Based on this 

combined evaluation, the previous assessment described concern over the steepness of the doseresponse curve and as a result the FQPA 10X Safety Factor was retained. In the last few years, 

HED has increased its use of benchmark dose (BMD) analysis in risk assessment. In an effort to 

better characterize the dose response relationship for supernumerary ribs in rat fetuses, a BMD
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analysis was conducted on Studies A, B and C at EPA’s National Center for Environmental 

Assessment by statistician Jeff Gift, Ph.D., using the fetal and/or litter data available from each 

study report using EPA’s Benchmark Dose Software (BMDS, Version 2.). NCEA provided the 

full analysis for each study (including the BMD outputs and plots). Only the plots for the best 

BMDS run for each study presented in this memo. However, the complete results of all analyses 

are presented as attachments to this memo. 

Briefly, because of the nature of the data available from each study, different BMD analyses 

were conducted for each. These are relatively old studies and the individual pup data were not 

available for 2 of 3 studies. In the case of developmental studies, it is typical for the responses of 

pups in the same litter to be more similar to each other than to the responses of pups in different 

litters (“intra-litter correlation”, or “litter-effect”). When conducting BMD analyses of 

developmental studies, nested models are the preferred approach since these models incorporate 

two parameters to address this issue: litter-specific covariate (LSC) and intra-litter correlation 

(ILC). The highest confidence analysis is for the Rubin & Nyska study (study A) where 

individual fetus data were available and a nested analysis was completed. The lack of individual 

animal data for the other two studies (B & C) impacts the degree of confidence on the BMD 

analysis. HED can, however, use the BMD analysis of all three studies to provide an improved 

evaluation of the dose response curve compared with that performed in the previous risk 

assessment, and thus inform the magnitude of the FQPA SF.

II. SUMMARY of RESULTS

Study Analysis BMD 

mg/kg/day

BMDL 

mg/kg/day

Comment

Study A

(Rubin & Nyska

1982)

nested dichotomous data 

analysis, using individual litter 

and fetus data

13.66

8.34

Availability of individual data from 

both litters and fetuses allows 

integration of both into the same 

analysis

Study B

(Baker & Cropping

1981)

dichotomous data analysis, using 

the litter as the subject and # 

litters with anomalies as the 

response

1.06

0.75

Lack of individual litter and fetus 

data disallows any accounting for 

litter variation or inter-litter 

correlation; this is the lowest 

confidence BMD run

Study C

(Rogers et al. 1991)

continuous data analysis, using 

mean percent/litter

9.80

8.86

Lack of individual litter and fetus 

data disallows any accounting for 

litter variation or inter-litter 

correlation. Analysis is based on

mean data only.

1. Study A: Rubin & Nyska (1982)

The current DER states that the NOAEL and LOAEL for this study are 4 mg/kg/day and 

12.5 mg/kg/day, respectively. The data-set for this study included both individual litter 
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and individual fetus data, which allows for the “nested dichotomous” analysis. This 

analysis produced high confidence results. The benchmark response of 5% ‘extra risk’ 

(i.e., incidence above the background) was used here. The use of a BMR5 is normally

used for developmental endpoints. It is notable that the BMD5 and BMDL5 are both 

higher than the NOAEL of 4 mg/kg/day. 

 0

 0.05

 0.1

 0.15

 0.2

 0.25

 0.3

 0.35

 0 5 10 15 20 25 30 35 40

Fraction Affected

dose

Nested Logistic Model with 0.95 Confidence Level

14:44 12/02 2010

BMDL BMD

 

Nested Logistic

2. Study B: Baker and Cropping (1981)

There is low confidence in the results of the Study B analysis because the data needed for 

meaningful and reliable analysis, such as individual animal and litter variation, were not 

available from the study, rendering the data less reliable. However, visual inspection of 

the graph shows large variability of the data, suggesting that there is no meaningful 

difference between the control group and the low dose level (5 mg/kg/day).
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 0

 0.2

 0.4

 0.6

 0.8

 1

 0 5 10 15 20 25 30 35

Fraction Affected

dose

Quantal Linear Model with 0.95 Confidence Level

13:50 12/02 2010

BMDL BMD

 

Quantal Linear

3. Study C: Rogers et al (1991) 

Similar to Baker and Cropping (1981), the individual pup data are not available, thus 

limiting the utility of the BMD values. However, visual inspection of the plots shows a 

shallow dose-response relationship at the low and mid dose such that the effects at the 

mid dose are not meaningfully different from the control or low dose.

 0

 10

 20

 30

 40

 50

 60

 70

 80

 0 2 4 6 8 10 12 14

Mean Response

dose

Exponential Model 2 with 0.95 Confidence Level

14:47 12/02 2010

BMDL BMD

 

Exponential
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III. DISCUSSION

The previous risk assessment used study A and study B as co-critical studies and established the 

NOAEL of 4 mg/kg/day based on study A, and the LOAEL of 5 mg/kg/day based on Study B. 

These values suggest a very steep dose-response curve, which provided the basis for the 

retaining of the 10X FQPA factor.

Since that time, use of BMD analyses in HED assessment has accelerated, and the RAB3 Risk 

Assessment Team considered this as a good opportunity to use a more refined dose-response 

approach. BMD modeling uses the entire dose-response curve, considers the variability of the

entire data-set, and takes sample size into account instead of just looking at a single point 

(NOAEL or LOAEL) for establishing a point of departure. Thus, BMD modeling provides an 

objective re-consideration of the available dose-response data.

Study A provided the individual fetal and litter data needed to assess changes in incidence/litter 

of supernumerary ribs with dose level, allowing a nested approach to analysis which integrates 

litter and fetal data. For the two studies without individual animal data (B and C), visual 

inspection of the graphs suggest that the dose-response curves are more shallow than previously

interpreted. As such, this analysis reduces, if not alleviates, the previous concern over the 

steepness of the dose-response curve related to the supernumerary ribs and retention of the 

FQPA 10X. At this time, the RAB3 Team has concluded that HED can have confidence in the 

NOAEL of 4 mg/kg/day and use it as the point of departure for supernumerary ribs, without the 

need to retain a 10X FQPA factor. 

It is important to note that this analysis is limited to a single endpoint and was conducted for the 

purpose of evaluating the dose-response data only from the supernumerary ribs and thus may not 

appropriate for use in setting other points of departure. Specifically, the BMDL5 of 8.34 

mg/kg/day it is not protective of the decreased body weight gain observed at 5 mg/kg/day in F1 

and F2 parents in a multi-generational reproduction study, or the decreased body weight gain, 

panting and liquid feces in female dogs at 5 mg/kg/day in a 90-day oral toxicity study.

IV. ATTACHMENTS 

1. Jeff Gift cover email

2. BandC 1981 -- BMD analysis of Baker & Cropping (1981) study data

3. Rand1982-BMR05 – BMD analysis of Rubin & Naska (1982) study data

4. Rand1982-BMR10 – BMD analysis of Rubin & Naska (1982) study data

5. Rogers1981-1Std – BMD analysis of Rogers, Francis et al. (1991) study data

6. Data set MRID 00116558 Baker & Cropping 1981

7. Data set MRID 40466802 Rubin & Nyska 1982

8. Data set Rogers Francis et al 1981