211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

EPA 

API reviewer response (black) 

Lab response (red) 

Plus_ 

API 

verification of 

lab response 

In general, the NHEERL reviewers concluded that the 

studies were conducted in accordance with the testing 

guidelines, once changes detailed in the API peer review 

response are made. 

Additional comment is requested on the following points: 

- Page 37 notes an extended diestrous period in a significant 

number of study animals and abnormal cycle patterns 

across the study. Page 38 notes that both the 20,000 

mg/m3

 and 10,000 mg/m3 exposure groups exceeded 4 

days to mate although significance is attributed only to the 

delay seen in the 20,000 mg/m3

 group. The results from 

the 10,000 mg/m3

 group were discounted due to 

inconsistent cycling of the test group attributed to 

incidental pseudo pregnancy (defined as extended periods 

of diestrous) in 9 of 26 animals and 10,000 mg/m3

 was 

selected as the reproductive NOEL. We are concerned that 

the reduction of the 10,000 mg/m3

 exposure group to 17 

animals will leave insufficient statistical power to allow 

this determination, and would request that the study group 

reconfirm the validity of their conclusion after taking the 

decreased group size into account. Should 17 animals 

prove insufficient we would request that 2000 mg/m3

 be 

designated as the reproductive NOEL. 

-

Note that 22 animals were found pregnant in 

the 10000mg/m3 group 

considered sufficient for statistical evaluation. 

OK. 

Table 14 

shows cycle 

lengths were 

evaluated in 

23-26 rats/ 

group. There 

were a 

similar 

number that 

could be 

evaluated for 

estrus cycles. 

Peer Reviewer – Schlesinger

The study was conducted in a scientifically sound manner 

and followed the appropriate protocols. There were no 

significant deviations from these protocols that would have 

affected the outcome of the study. While the NOEL level for 

reproductive performance seems reasonable, some 

consideration should be given to the conclusion for the 

general NOAEL based upon some of the comments below 

. 

No response needed 

√

Statistical Analysis: 2.15.1. 

Is there any reason why two way ANOVA was not used? 

This would have allowed evaluation of exposure groups at 

all times for each endpoint. This could be followed by some 

multiple comparison test. Furthermore, why were tests 

conducted at both the 1% and 5% level of significance. The 

tests and level to be used should be determined a priori 

before the study begins and only one such level of 

significance should be used. 

The computerized statistical testing package 

employed at the laboratory is designed to 

perform one-way ANOVA at each time 

interval and is used for a range of multiple 

dose studies. 

The 1% and 5% levels of significance 

were pre-selected since they were inherent in 

the laboratory’s computer statistical package. 

Comparison at two levels of significance has 

been common practice in this laboratory. 

OK 

This is similar 

to other 

studies.

1 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

Protocol Deviations: 2.18. 

There appeared to be a number of errors committed by 

technicians. While these may not have affected the outcome 

of the study, they seem to indicate a degree of technical 

"sloppiness" in the conduct of the study. 

Nearly all of the protocol deviations were 

related to isolated errors in data collection. 

None significantly impacted the overall results 

or conclusions of the study. While the 

absolute number of protocol deviations may 

seem excessive, a One-Generation 

Reproduction study is a considerable 

undertaking and involves a large number of 

animals. The fact that these protocol 

deviations were noted and documented 

reflects positively on the Quality Assurance 

practices that have been put in place by API, 

and at the laboratory. Nonetheless, the RG 

brought this issue to the attention of the 

laboratory to strive to minimize protocol 

deviations. 

√

Chamber Monitoring: 3.1. 

It is noted that there were some differences between the 

measured and the nominal concentrations in that they varied 

from the expected ratio. The explanation for this was 

suggested to be a combination of slight inaccuracies in the 

calibration of the chamber airflows and the calibration of the 

IR monitor. This reviewer feels that there should be no 

inaccuracies in either of these, especially in the calibration of 

the monitor. The investigators need to assure that the 

calibration of all monitors and flows are extremely accurate. 

There should be no such inaccuracy of any atmospheric 

monitor. In any case, the target, analytical and nominal 

levels seem to be very consistent. 

The investigators need to explain the source of the particles 

in the exposure atmospheres. Was the incoming chamber air 

filtered appropriately? Any particles could act as adsorption 

sites for vapors in the exposure atmosphere. The statement 

that the "...atmospheres were essentially vapor only" is not 

quite right. While the vapors may not have produced 

particles, there were clearly particles in each of the exposure 

atmospheres. Furthermore, the range of particle sizes was 

quite large, going from less than 1 micrometer to over 5 

micrometers in a number of cases. 

To be precise, the report stated that the ratio 

of the measured to nominal concentration was 

not 1:1, and speculated that the cause MAY

have been due calibration error. However, as 

Dr. Schlesinger also pointed out, the target, 

nominal and measured concentrations were 

close, varying 2% or less. The RG feels 

confident that the reported chamber 

concentrations accurately reflect the 

atmospheres to which the animals were 

exposed. With values so close together, the 

explanation for possible variation from the 

Results and Summary will be deleted. 

Deleted and indicated acceptable differences. 

With regard to the presence of particles, the 

RG notes that the mean particle concentrations 

were comparable between the control and 

treatment group chambers. The primary 

purpose of making these measurements in a 

study of this nature (high vapor 

concentrations) is to make certain that vapor 

condensation is not occurring and to preclude 

the possibility of aerosol formation. The 

results provided in the report confirm that 

gasoline + TBA exposure was to vapors and 

not aerosols. Some particles are invariably 

present in chamber atmospheres, representing 

background levels. In fact, careful analysis of 

the data in the table on page 34 [first draft] 

indicates that the average particle 

concentrations in all of the chambers 

including controls were 5-5.7 micrograms/m3

√

» same page 

in draft 2 

2 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

of air. For comparison, it has been reported in 

EPA's PM 10 Criteria document that annual 

background PM 10 levels in eastern cities 

range from 5 to 11 micrograms / cubic meter 

of air. Thus, the RG believes that the particles 

in the chambers reflect ambient background 

values. 

√

Physical Observation Data: 3.2.3. 

There seems to be an exposure related increase in alopecia. 

Dr. Schlesinger is correct that the incidence of 

alopecia of the extremities/snout of male rats 

increased in an exposure related manner, 

exhibiting maximum incidence in the mid 

dose group, and including 3 control rats, by 

week 8. Similar effects were not seen in 

females where the total reported incidence 

was 1 animal in the mid dose and 2 in the high 

dose. The study director will be instructed to 

add these observations to the appropriate 

Results section 

Added to discussion.

 

√ 

3.2.2_p. 35 

Parturition and Litter Survival Data: 3.2.10. 

Data in Table 16 suggest a treatment related increase in 

number of pups dying, missing or cannibalized at the two 

higher exposure levels that does not seem to be clearly noted 

in the report text. 

The statistically significant reduced pup 

viability indices and increased incidence of 

pups stillborn or dead in the 10000mg/m3

 and 

20000mg/m3

 groups were due to two litters in 

the 10000mg/m3

 group: one dam delivered 10 

stillborn pups, and in the other litter all 12 

pups died by the day 1 weighing., and to one 

litter in the 20000mg/m3

 group in which 7 of 

17 pups died by day 1 weighing. When these 

anomalous litters are removed from 

calculations, no statistically significant effects 

were observed in the number of pups or pup 

viability index. Moreover there was no dose 

response across these levels for pup mortality. 

Section 3.2.9 revised to add information on 

stillborns.

P. 39 

OK 

When we 

publish, we 

should include 

the range for 

controls in the 

total program. 

. 

Postmortem Data: 3.4.1. 

The lung discoloration in males seems to be treatment 

related. Perhaps this is meaningful in spite of the lack of any 

effects on lung weight or histopathology. 

Dr. Schlesinger is correct that the number of 

rats with discolored foci in the lungs was 

observed to increase in an exposure-related 

manner in both sexes at necropsy of parental 

animals. As reported in the text, no 

histopathological effects such as alveolar 

macrophage aggregates consistent with 

discolored foci were seen and no changes in 

lung wt occurred. Therefore, the increased 

frequency of these background lesions is 

likely a marker of substance exposure but is 

not considered to be of adverse biological 

significance. 

HLS_ Consider adding a sentence that these 

OK

p. 40 

3.4.1 

3 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

lesions are likely a marker of substance 

exposure not considered adverse. 

Added to discussion.

2nd

 paragraph

Postmortem Data: 3.4.3. 

It is not clear why the effect on spleen weight clearly 

observed in this study is discounted based upon results of the 

13-wk study. 

Statistically significant decreases in absolute 

and relative spleen wt were seen at all doses in 

parental females and in 10000mg/m3

 and 

20000mg/m3

 parental males. As pointed out 

by Dr. Goldsworthy, the consistency of these 

decreases as well as a similar effect 

considered significant in pups from 

20000mg/m3

 exposed dams make it difficult 

to dismiss these observations in the absence of 

concomitant histopathology. 

Revised the discussion. 

Repro NOEL 

and NOAEL 

p.43 

also 

p.41 

section 3.4.3 

2nd paragraph

ok

Peer Reviewer – Goldsworthy

The study was scientifically sound and was properly 

conducted. Exposures were properly conducted and 

adequate to test the potential reproductive toxicity of 

Gasoline TBA Vapor Condensate. None of the protocol 

deviations noted would be expected to adversely effect study 

results or data integrity. Protocol deviations listed on page 

32 did not note the temperature or humidity excursions 

outside the specified range. 

No response needed 

√

No effect of treatment was noted on survival, clinical 

observations, feed consumption, estrus cycles, mating, 

fertility or gestation indices, or on other reproductive 

parameters up to parturition except for animals in the high 

dose group which took slightly longer to mate as compared 

to other groups. There were exposure related decreases in 

body weights (Table 5) and body weight gain (Table 6) in 

the 20,000 mg/m3 animals during the pre-mating period. 

This is not correctly stated in Summary (page 6) and 

Conclusion (page 42) and should be modified. No treatment 

differences were observed with pregnancy rates, duration of 

gestation and percent of females completing delivery. No 

differences between treated and control groups were noted in 

all parturition parameters, including total number of pups 

delivered, viability and lactation indices, the number of 

implantation sites per litter, the pup sex ratio, and the 

number of live pups per litter. Additionally, sperm count, 

motility, morphology and caudal epididymis weight were not 

altered by treatment. 

Dr. Goldsworthy’s comment is correct. The 

Conclusions and Summary will be revised to 

include decreases in absolute body weight as 

well as decrease in weight gain 

Group 4 absolute body weights were not 

decreased but gained less than the controls. 

Discussion OK as is. 

√

There were no exposure-related differences in macroscopic 

postmortem assessments. Increased kidney weights were 

observed in male rats (parental generation) at the two highest 

Statistically significant decreases in absolute 

and relative spleen wt were seen at all doses in 

parental females and in 10000mg/m3

 and 

4 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

exposure levels; this is consistent with the expected 

induction of hydrocarbon nephropathy. Microscopic 

evaluation was not performed to confirm this observation. 

Hydrocarbon nephropathy is generally not considered to be 

relevant for human risk assessment. Statistically significant 

decreases were seen in spleen weights at all exposure levels 

in both males and females. These decreases were seen in 

absolute values as well as relative values to body and brain 

weight. This finding was dismissed as adverse because 

similar effects were not observed in a previous 13-week 

study. However, data findings must place emphasis within 

the context of current study control values and results. The 

consistency of splenic weight decreases in all dose groups, in 

both sexes as well as observed in pups, suggests that this 

finding may be an adverse finding. Unfortunately, 

histopathology results were not conducted to clarify this 

finding. Without further clarification as to the relevance of 

the data, this report should note the decreased spleen weights 

in Summary and Conclusion sections. These data also will 

affect the NOAELNOEL setting. 

20000mg/m3

 parental males. As pointed out 

by Dr. Goldsworthy, the consistency of these 

decreases as well as a similar effect 

considered significant in pups from 

20000mg/m3

 exposed dams make it difficult 

to dismiss these observations in the absence of 

concomitant histopathology. 

Summary and R&D revised to note this spleen 

weight effect.

Summary p7 

paragraph 1 

p41 

3.4.3 

Paragraph 2 

OK 

As noted above, kidney, liver or spleens were not evaluated 

histopathologically. It is not clear why only these three 

organs (plus brain) were not examined microscopically as 

were all other tissues listed in Table I (page 26); this lessens 

the strength of the report. Both gasoline and TBA have been 

shown to induce a variety of renal and hepatic alterations. 

This study was designed to evaluate 

reproductive effects. The RG decided that 

since extensive histopathology had been 

performed in the 13-week study and results 

documented on a variety of organs, it was not 

necessary to repeat here. 

√ 

There were no exposure-related differences in clinical 

observations or macroscopic postmortem evaluations in 

lactating pups from air control animals. Table 17 and 18 

demonstrated statistically significant decreased body weight 

and body weight gain at some time intervals in the high dose 

groups. 

Thus, the first sentence in section 3.3.2 and a similar 

sentence in Summary section (page 7) should be revised and 

these findings appropriately noted. Significant decreases in 

spleen weights were seen in the 20,000 mg/m3

 exposure 

level animals as compared to controls. 

The Conclusions and Summary will be revised 

to include decreases in absolute body weight 

as well as decrease in weight gain. 

Summary and R&D sections 3.3.2 and 

Conclusions revised.

√ 

In summary, test animals were properly exposed to 

adequately test the potential reproductive toxicity of 

gasoline-TBA. A NOAEL of 10,000 mg/m3

 was determined; 

however, this NOAEL setting does not take into account the 

decreased spleen weights observed at all exposure levels. 

With respect to reproductive performance, the NOAEL 

setting of 10,000 mg/m3

 is warranted. 

Done to reflect that “...with respect to 

reproductive performance, a no observable 

effect level (NOEL) of 10000 mg/m3 of 

Gasoline TBA Vapor Condensate was also 

concluded with an NOAEL of 20000 mg/m3.”

OK

211(b) Research Group Reviewer (Roberts)

General Comments 

5 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

The report should have sequential page numbering from start 

to finish. 

All of the reports should have an appendix summarizing 

historical control data for reproduction endpoints. 

This should be included in all of the reports. This doesn't 

need to be massive; endpoints such as fertility and gestation 

indices, litter size, sperm counts/motility, etc., are the types 

of endpoints that would be of most value. 

HLS and PAI HCD added to the report as 

Appendix EE. 

 OK

The handling and write-up of all of the estrus (vaginal 

smearing) data was very nice, with good explanation and the 

periods of pseudopregnancy were excluded from 

calculations. This is how all of the reproduction studies in 

the 211(b) program should be handled. 

No response needed 

There are still issues with classification of stillborn and 

found dead pups. Individual data in the appendices and the 

summary tables do not seem to add up with consistency. 

Tables and appendices reviewed and corrected 

as necessary.

OK

Specific Comments 

P. 2, Statement of Compliance. 

"It was the Sponsor's responsibility to maintain the method 

of synthesis, fabrication, or derivation of the test fuel, and 

this has not been completed." Not only should this be 

completed, but most likely it is being included in the SOT 

poster. This needs to be rectified between API and HLS. 

Corrected √

 

P 2

P. 5, 4th paragraph. 

Please replace "reasonably close" with "acceptably close," 

since there should be criteria for satisfactory comparison. 

Revised to “acceptably close, in the opinion of 

the study director,”

P6 1st line 

OK 

Please consider deleting the sentence beginning, "The exact 

cause of the differences.. .." since this is unnecessary. 

Deleted. √ 

P 6, Parental Organ Weights. 

Please see comments for p. 41, parental organ weights. 

Added to R&D but not of value to this 

discussion in the Summary. 

√ 

P. 15, 2.4.1, Supplier. 

Please correct the name to ChevronTexaco Research and 

Technology Company, or ChevronTexaco Global 

Technology Company, whichever will be correct to QA and 

consistent throughout the 211(b) program. 

Revised to: ”Chevron Global Technology 

Services Company “ for consistency with 

other reports.

√ 

P. 16, 2.4.9, Dispensing. 

Revised to: ”Chevron Global Technology 

Services Company “ for consistency with 

√ 

6 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

Please correct the name to ChevronTexaco Research and 

Technology Company, or ChevronTexaco Global 

Technology Company, whichever will be correct to QA and 

consistent throughout the 211(b) program. 

other reports.

2.4.1 0, Archival Sample. 

The wording suggests that the archival sample is stored as 

described in 2.4.8; i.e., in an outdoor solvent shed or indoor 

laboratory. Please clarify. 

Clarified to indoor lab. √ 

P. 19, 2.9.4, Feed Analysis. 

The comma after "feed" is incorrect. 

Corrected. √ 

P. 20, 2.9.9, Environmental Conditions. 

The actual low temperature, 14.3C is very low compared to 

the intended range. Please include when this happened 

(study day/phase) and how long it persisted until the room 

temperature was within the desired temperature range. 

This excursion was on 26Sept02 and 

recovered to 18C within 1 hour. I have added 

to report that “Excursions outside the 

specified range were generally brief in 

duration and considered not to have affected 

the integrity of the study.”

√ 

P. 22, 2.1 1.2, Detailed Physical Examinations. 

The comma after "as well as" is incorrect 

Corrected. √ 

P21 bottom 

P. 32, Protocol Deviations. 

- No. 1: What is meant by "no definite effect?" My 

recollection is that, after this occurred, a number of the 

female rats in estrus persisted in estrus longer than usual. 

Please check. 

- No. 3: If pup No. 5 was not preserved, what was the 

consequence? Were less than 3/sex preserved, or was 

another pup preserved in its place? 

- No. 5: Please reword; the correct wording suggests that 

technician error caused No. 1527 to be found dead. 

- No. 6: Were efforts made to contact the supplier and 

obtain the appropriate bedding analyses? Example 

analyses from 1999 are not very relevant (and if this 

occurred in earlier reports, I apologize for not catching this 

problem). 

No. 1. revised to no ‘apparent’ effect. 

No. 3 No consequence: Only macroscopic 

examination was required for 3/sex at terminal 

sacrifice (Day 28); no preservation was 

required. Only pups with abnormalities were 

required to be preserved. This pup was found 

dead on Day 4. There was no milk in the 

stomach, therefore it should have been 

preserved. 

No. 5. Reworded for clarity. 

No. 6. Further contact with the vendor (Fisher 

Feed) was made on 11Sept09 and they 

confirmed that actual lot numbers for the 

bedding for these studies were not available. 

Therefore, the example analysis already 

included should be sufficient. 

√ 

OK

 

P. 33, 3.1, Chamber monitoring. 

Table 6 in Appendix A is for the testing room, not the 

chambers. Could you please point out where the chamber 

oxygen levels are located? Table 6, week 17, indicates 19% 

oxygen for Group 1, so if this is the table that represents 

chamber oxygen levels, the text on p. 33 needs to be 

corrected from "at least 20%" to "at least 19%." 

Table 6 in Appendix A clarified for the room 

and chambers results. 

Text on p. 33 corrected from "at least 20%" to 

"at least 19%."

√

P173 / 654

7 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

P. 34, 3.1, last paragraph. 

Please revise "reasonably close" to "acceptably close," since 

there must be a criteria for satisfactory comparison. 

Revised to “acceptably close, in the opinion of 

the study director,”

P33 

OK 

Last sentence. 

Grammatical correction: "data was" should be "data were." 

Corrected. √

P. 35, 3.2.1, Mortality. 

Table 1 shows that a Group 2 female (2549) died/sac's 

postpartum and two Group 3 females (3536, 3548) died/sac'd 

postpartum. This is not discussed in this section or tied in 

elsewhere, such as in 3.2.1 0 for litter survival (not tied in for 

the dams). The listings in Table 1 need to be explained, if 

only to clarify why the animals were sacrificed. Also, please 

read comments directly below for 3.2.3. 

Added to section 3.2.1. 

No change to Table 1. This is a summary 

table. Explanations for individual animals can 

be found in Appendix C.

√

 

P35 

3.2.2. 

Doesn't exist. Please correct the section ordering. 

Section ordering corrected. √

3.2.3, Physical observations. P35 

Table 3 does not seem to match Table 1 for numbers of 

animals with terminal sacrifice during gestation. This needs 

to be explained and/or corrected. Also, the row for "found 

dead" in Table 4 does not appear to concur with Table 1. 

The fate of animals that did not survive to scheduled 

termination needs to be explained in a clear manner so that a 

reader will understand what happened without needing to dig 

through the individual data tables. 

With the exception of animals with Total 

Litter Loss, Table 1 does not show Terminal 

Sacrifices. No change to Table 1 with regard 

to Terminal Sacrifices during Gestation (Table 

3). A notation was added to show that 

animals with Total Litter Loss are included in 

the animals that died/were sacrificed postpartum. These are classified as Terminal 

Sacrifices in Table 4 (and Appendix C). 

OK 

3.2.4, Body Weight. 

Didn't males at 20K also have lower body weight gain during 

the week prior to exposure initiation? Also, there were 

additional weeks, beyond what is mentioned in the text, with 

lower body weight gain. 

Section revised. OK

P. 36, 3.2.5, Maternal Body Weights - Gestation and 

Lactation. 

Please consider revising this section. The text should focus 

upon what the findings are. It is apparent from the data that 

there were no differences in gestation body weight gain; this 

should be stated. It isn't clear in this section if there is an 

interpretation that biologically or toxicologically significant 

differences occurred in lactational body weight gain; did a 

prior body weight difference resolve during lactation (i.e., 

the adverse effect ended), or did the dams put on more 

weight at the expense of the pups (separate adverse effect)? 

Section revised.

OK

8 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

3.2.6, Feed Consumption. 

Please see question below related to Table 11. 

See response to Table 11 comment below. OK

P. 37, 3.2.8, Estrous Cycle Data. 

This is a nice write-up and explains the results very well. I 

think that the final sentence, "Adequate assessment of 

cyclicity was possible despite the incidental episodes of 

pseudopregnancy" could be strengthened even more, since 

adequate assessment was not only possible, but occurred. 

Section revised. OK

P. 38, 3.2.9, Mating, Fertility, and Gestation Indices. 

This, also, is a nice write-up. There is still some ambiguity 

about the delay in mating at 20K; although it appears that 

this is attributed to exposure, it isn't clear what the "normal" 

range for days to mate is, or how frequently animals miss 

their first opportunity to mate. Although this was not an 

endpoint for adverse findings in other studies in our 

program, the argument for no effects at 10K (artifact of 

pseudopregnancy) should be consistent with what was seen 

in control groups (i.e., BIMTBE) with pseudopregnancies. 

Section revised. OK

3.2.10, Parturition and Litter Survival Data. 

There are discrepancies between Table 16 and Appendices 

R, S, U and W. Table 16 lists 21 animals with liveborn, but 

Appendix R, p. 1, lists 20. Did this eliminate No. 1527, the 

female that died on LD 1 with one fetus inside? Appendix R 

lists 21 animals with liveborn. The table and the appendix 

need to be consistent, and the situation for No. 1527 should 

be described. Also, Table 16 does not report the same 

numbers as Appendix S. Table 16 reports that the control 

group had one litter with stillborn pups, yet Appendices S, 

U, and W show 2 litters13 pups that were stillborn. To add 

to the confusion, the two stillborn pups for No. 1527 that are 

confirmed stillborn by the lung floatation test were reported 

to have milk in their stomachs. Something is wrong here and 

needs to be straightened out and clarified before the results 

can be published. 

Animal No. 1527 is excluded from Table 16 

and Appendix R (delivery and litter data) and 

Table 18 and Appendix R U (pup weights). 

Notations were added to the appendices 

indicating the exclusion. 

Since No. 1527 is excluded from Table 16, the 

pups (Appendix S, U and X) from that litter 

are also excluded – leaving 1 litter with 1 

stillborn pup. 

Animal No. 1527 was not excluded from 

Table 19 or Appendix W (parental necropsy) 

or Table 20 or Appendix X (pup necropsy). 

Notation added to Appendix X indicating that 

the milk in stomach observation supersedes 

the lung floatation test, therefore the status for 

these pups was corrected to “Dead”.

Now 3.2.9 

Append R 

_p336 

Append U 

p 388 

Append S 

p340 

Append U 

p384 

Append X 

p 435 

append W 

p424 

 Append X 

p435 

OK

P. 39, 3.2.10, 9 Parturition and Litter Survival, cont.. 

It seems appropriate to include the reasons why total and 

partial litter loss at 10K and 20K, respectively, are 

considered incidental ("occurrences that were considered 

incidental because...") 

Section revised. OK

3.3.2, Pup Body Weight. 

9 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

When discussing the slightly greater pup weight gain at 2K, 

please consider revising to include comment about the 

slightly smaller mean litter size that occurred in this group 

compared to control. 

Also, there are an excessive number of weight gain intervals 

shown on Table 18. Please consider revising Table 18 to 

include only those intervals that seem most appropriate, then 

be consistent across all the studies for the 211(b) program. 

Section revised. 

Table 18 - Intervals cannot be changed due to 

TASC program limitations.

√

√

P. 40, 3.4.1, Postmortem Data, Terminal Estrous. 

Please consider using a single consistent spelling for 

estrus/estrous. At the end of the first paragraph, please 

consider adding in the lab historical control range for estrus 

at termination; it will no doubt provide ample evidence of 

the variability of the endpoint. 

Spelling OK as is since estrous is adjective 

and estrus is noun. 

Discussion of the estrus HCD added to the 

text.

OK

√

Second paragraph. 

The criteria for discounting lung discoloration as unrelated to 

treatment are not convincing. The justification may be 

adequate for a determination that it isn't adverse, but both 

males and females had a higher incidence of discolored 

lungs, and the data look related to exposure. In males, 

discolored thymus also appears to be increased at 20K; 

perhaps comparison to control ranges would be appropriate. 

Discussion of lung discoloration has been 

expanded. Thymic discoloration (red) is a 

common finding in rats and is due to agonal 

hemorrhage (caused at the time of death by 

the method of euthanasia). It is generally not 

recorded microscopically by the pathologist 

since it is considered a post mortem artifact. 

Since the incidence was so low and only 

present in males, it is not considered necessary 

to discuss the artifactual nature of the finding 

in the text. 

OK

3.4.2, Macroscopic Evaluations, Pups. 

Please see comments below for Table 20 and Appendix X. 

This section should discuss each section (died, stillborn, 

culled, necropsied) separately, even if only as a single 

sentence each. 

Discussion has been revised. OK

P. 41, 3.4.3, Organ Weights, Parents. 

There should be a comment about the increased organ/body 

weight ratios due to reduced body weight. 

Mention added at end of this section. P41 end /p 42 

1st sentence 

OK 

The argument that the decreases in spleen weight are 

unrelated to exposure on the basis of the subtonic results is 

problematic. The subchronic study did not expose the 

animals for as long, and exposed only 5 d/w. 

Discussion has been revised. P41 end 

OK

Table 1, Summary of Survival and Pregnancy. 

This table is confusing, since the total females 

died/sacrificed does not clearly add up to the numbers shown 

above. Please clarify or correct. 

No correction necessary. Total females 

died/sacrificed is a sum of the categories 

presented above it on the table.

OK

Table 3, Terminal Sacrifice. 

The numbers don't seem to agree with Table 1. 

Only unscheduled deaths are included on 

Table 1. No change.

OK

10 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

Table 4, Found Dead. 

Again, the numbers don't readily add up. This might be 

handled by discussion in section 3.2.1 . 

Missing terminal sacrifices added. OK

(died post 

partum)

Table 11, day 56. 

Why is the value for group 2 (X=60, SD = 2.6, N=26) 

statistically significant from control (X=62, SD = 3.5, 

N=26), but the value for group 3 (X=60, SD = 2.3, N=26) is 

not? Please check; perhaps this is due to rounding. 

This is due to rounding. OK

Table 16, Summary of Delivery and Litter. 

Please see comments above for section 3.2.10. In the control 

group, there is one less female with liveborn than females 

pregnant. Yet review of the individual data in the 

appendices (R) clearly shows 21 litters with liveborn. I 

assume that No. 1527 was not included, but she met the 

criteria for inclusion in the gestation index. 

Again in the controls, only one litter is listed as having any 

stillborn, yet there were two litters13 pups that are listed in 

appendices as status stillborn. For one of those litters, the 

two stillborn pups should be classified as stillborn on the 

basis of the lung floatation test, but observations for them 

included milk in stomach (see comments under appendix X). 

This doesn't make sense. These data need to be reviewed and 

the individual data need to tally up to the values presented in 

summary tables. 

See response to section 3.2.10, Parturition and 

Litter Survival Data comment.

P35 

OK 

Table 20. 

This table has four page ones; the pagination should be 

continuous, start to finish. The section of the table that is 

headed, "Summary of Pups Stillborn Observations" includes 

a total of 7 pups that the lung floatation test determined to be 

"found dead," not stillborn. Why are they still on this table? 

For "Summary of Pups Cannibalized," there are 5 pups in 

group 3 for whom milk in stomach was undetermined; does 

this mean that there was one pup for which milk in stomach 

was determined? If so, it should be listed, but isn't. 

Entire report will be paginated beginning to 

end at finalization. 

Statuses for pups corrected based on results of 

lung floatation tests. 

Per Appendix X, no observations for milk 

were made for 3547-2. Table 20 is correct as 

is.

OK

( note: 6 

pups & 

P456 

3547-2 

cannibalised)

Appendix A, p. 7, 2.3.2. 

Please correct the grammatical errors in the second sentence. 

Corrected √

Appendix X. 

Some cannibalized pups are listed as status Z for 

cannibalized, others as stillborn. Why the difference, and 

If stillborn or dead status could be determined 

for a cannibalized pup (i.e. a lung floatation 

test could be performed or observations for 

milk in the stomach could be made), that 

OK

11 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

what is the justification for the difference? status is listed. Otherwise, the pup is listed as 

cannibalized (Z).

For 1527/1&2, both are listed as stillborn, yet have milk in 

stomach. This doesn't seem to be a likely assessment. 

Pups will be considered found dead (milk 

observation overrides lung floatation). 

Animals footnoted in Appendix X.

 

(Note: p435 

Footnoted) 

OK 

For 252612, the status is S for stillborn, yet the observations 

indicate milk in stomach-found dead. Why is this listed in a 

conflicting manner? 

1, 2 were two different pups. One was 

stillborn and one was found dead.

OK 

For 253911, the status is stillborn, but the observations 

indicate that the pup should be classified as found dead on 

the basis of the lung floatation test. The same is true for 

254311, 352811 (which also has milk in stomach), 

353617&10, and 453811. 

2539-1, 2543-1, 3528-1, 3536-7, 10, 4538-1 – 

all were found dead according to data. 

3536 – was partially cannibalized meaning it 

was undetermined. 

Statuses for pups corrected based on results of 

lung floatation tests.

OK

211(b) Research Group Reviewer (Cruzan)

p. 6. 

Summary says there was no effect on body weight - High 

dose males had significantly lower BW except day 7 and 42; 

high dose females except day 7, 28, and 42. 

Discussion of body weights has been revised. √

P6 

paragraph 2 

p. 38, Section 3.2.9. 

Was the difference in mating length statistically significant? 

No √

It seems inconsistent to write-off decreased spleen weights in 

parental animals as not treatment-related, but decreased 

spleen weights in offspring as indicative of toxicity. It 

appears that in both cases, there was no histopathological 

difference in the high-dose. This was used as justification to 

dismiss in parents but not pups. 

Discussion of spleen weights has been 

revised.

P43 

OK

Overall, it seems the lab is really reaching to declare a 

reproductive effect at 20,000 mg/m3

. They cite a slight, but 

not significant, extension of the time to pregnancy (2.7+/-1.6 

vs. 4.0+/-2.7 days) and a decreased spleen weight without 

pathology. 

NOEL of 10000 with NOAEL of 20000 

revised in Summary and R&D conclusion.

P43 

OK 

211(b) Research Group QA/QC Reviewer 

Study Title: Gasoline TBA Vapor Condensate: A OneGeneration Whole-Body Inhalation Reproduction Toxicity 

Study in Rats 

The following items require further consideration: 

A. Draft Final Report and Associated Raw Data

1–Statement of Compliance, Page, 2: The second sentence Corrected OK 

12 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

should read, “It was the Sponsor’s responsibility to maintain 

the method of synthesis, fabrication, or derivation of the test 

fuel, and was not completed prior to study initiation, but has 

subsequently been completed. 

2–Quality Assurance Statement, Page 4: The protocol and 

draft report indicate that the Testing Facility was responsible 

for the GLP compliance of the subcontractor, PAI. The 

inspections/audits conducted to ascertain their compliance 

status should be listed on the Quality Assurance Statement of 

this report. 

QA Statement updated to reflect the PAI 

inspection.

OK

3–Page 6, Summary, second paragraph: It is indicated that 

there were no effects of treatment on body weights during 

the pre-mating period. It appears that 14 out of 18 body 

weight intervals had high-dose male body weights 

statistically significantly reduced when compared to 

controls. Approximately one-third of the female high dose 

body weight values were statistically significantly reduced 

compared to controls. Please clarify and add to ‘conclusion 

in third paragraph’. 

The next sentence indicates that body weight gains were 

decreased in an exposure-related manner, yet, in high-dose 

females only two intervals were statistically significantly 

reduced. This should probably be indicated. 

Discussion of body weights has been revised. OK

4–Page 7, Summary, first paragraph: It is indicated that no 

test substance related microscopic changes were noted in any 

tissues. It seems the kidney findings should be mentioned 

even if considered accepted and not relevant. 

The kidneys were not evaluated in this study; 

mention was only made of findings in the 13 

weeks study.

OK

5–Page 20, Environmental Conditions: After review of the 

data, it is not clear whether indicating that the temperature 

and humidity were maintained within the specified ranges to 

the maximum extent possible is true. There are many 

instances where temperatures far outside of acceptable 

ranges were noted and maintenance was not called (8/25 and 

26 and 9/26; temperatures of 15.8, 15.4 and 14.3, 

respectively)(during 9/02 there were 11 high readings of 

humidity over 90% with no maintenance calls). This appears 

to be an SOP deviation and should be documented as such. 

Per REES database records, the temperature 

alarm on 8/25 and 8/26 was answered by 

maintenance within minutes of the alarm and 

the alarm on 9/26 was answered by ToxOps 

(EW) who performed a calibration of the 

probe. 

The humidity excursions during September 

were all brief and intermittent. None lasted 

long enough to trigger an alarm. 

OK

6–Page 20, Temperature: The actual range is 14.3-26.0 

degrees C, I believe. 

Corrected OK

7–Page 20, Temperature and Humidity: The frequency and 

duration of the excursions from desired levels should be 

given. On most days the average is within the desired range, 

but for extended periods, the highs and/or lows are out of the 

desired range. For example, between 9/1 and 26, the relative 

humidity was above 70% at least once each day for all but 5 

days. 

There are too many brief excursions to list in 

the report but all are in the raw data. I have 

added to report for both temperature and 

humidity that “Excursions outside the 

specified range were generally brief in 

duration and considered not to have affected 

the integrity of the study.”

OK

13 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

8–As well, there are times when calls to maintenance were 

made and these calls were not answered for long periods of 

time (12/24 maintenance called for a Humidity measurement 

out of range; the call was recorded on 1/2/03; the call was 

answered on 1/15/03). 

Per REES database records, all calls were 

answered on the day of the alarm. Paperwork 

was completed at a later date. In the specific 

example, the alarm was triggered and 

answered on 12/24/02, the form was filled out 

on 1/2/03 and signed by the responding 

HVAC technician on 1/15/03. 

OK

9–Page 32, Protocol Deviations; Deviations numbered 3 and 

4 were not found written up as deviations with others in the 

data book. 

Deviation memos are in the data book.

OK

10–Page 33, Chamber Monitoring: The oxygen levels were 

at least 19%. 

Corrected. OK

11–Page 35, Physical Observations; Alopecia might be 

considered treatment-related in the males during the premating period. 

Alopecia added to the discussion. OK

12–Page 35, Body Weights: It is indicated that both males 

and females showed slightly reduced weight gain for the first 

3 weeks of exposure at the high dose level which continued 

until termination. As noted in item 3 of this report, body 

weight measurements as well as body weight gains were 

statistically significantly affected. The text indicating 

slightly reduced weight gain may be misleading as 10 out of 

17 measurements for the males were statistically significant, 

many reaching significance at the p<0.01 level. Females 

didn’t seem to be as affected as the males, so maybe it would 

be more accurate to talk about them separately. As well, the 

high-dose males had statistically significantly reduced body 

weight gain the week before exposures began, so this may 

need to be discussed. 

Revised to emphasize more the difference for 

the males and that the differences were 

statistically significant.

OK

13–Page 36, Maternal Body Weights: The deficit in female 

body weight at the high dose appears to be about 5% at the 

end of the pre-mating period, and then ranges from 3-5 

percent throughout the gestation and lactation periods, 

ending at about 1.5 percent at LD28 (resolving only after LD 

21). 

Section revised. OK

14–Table 11, Mean Feed Consumption: There appear 

several suspicious indications of statistical significance. 

These are – Males, Day 56, Levels 2 and 3; Males, Day 13, 

Level 2; Females, Day 35, levels 2 and 4. Please verify that 

appropriate statistical programs were run and reported. 

Correct as reported. OK

15–Page 32, Deviation number 7: Shouldn’t animal number 

1529 be added to this list not observed on Day 105?? 

No correction necessary. Animal No. 1529 

delivered a litter on Day 99. See lactation 

observations (Appendix F) for subsequent 

observations.

OK

16–Page 37, Estrous Cycle Data: The second paragraph 

indicates that there were 3 instances of pseudopregnancy in 

Discussion added for Animal 1529 not 1528. OK

14 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

the control group. In Appendix P, only two instances are 

noted and excluded from the cycle calculation. Should the 

extended period of diestrous for Animal 1528 also be 

excluded? 

17–Page 37, Estrous Cycle Data: The second paragraph 

indicates that there were 9 instances of pseudopregnancy in 

group 3, yet Appendix P only footnotes 6 extended periods 

of diestrous that are excluded from the cycling calculation. 

Shouldn’t animals 3541, 3542 and 3550 have the periods of 

pseudopregnancy excluded from the cycling calculation? 

Table 14 will change if these cycle lengths are changed. 

Discussion revised to 8 instances in Group 3 

and 3541 and 3550 excluded and Table 14 and 

Appendix P updated.

OK

18–Page 38, Mating, Fertility, and Gestation Indices: Please 

verify the group 4 data presented in the text box. I cannot 

verify the ‘Number missing first opportunity to mate or 

missing 2 or more opportunities’. 

Upon further review, we corrected the 

‘Number missing first opportunity to mate’ to 

7 and concur with 1 for ‘missing 2 or more 

opportunities’.

OK

19–Page 40, Macroscopic Evaluations: Lung observations 

were not really scattered as indicated in this section. They 

were actually dose related. 

See previous discussion of this topic. OK

20–Page 41, Histopathological Evaluations, Parental 

Generation: Chronic inflammation of the lung appeared 

slightly increased in the high-dose male animals. 

Minimal subacute/chronic inflammation is a 

common finding in lungs of rats on inhalation 

studies. In the absence of any other findings, 

this is not considered treatment related.

OK

21–Table 1, No. of Females Cohabited without evidence of 

mating, for Group 1 should be 3 (#1529, 1548, 1050), with 

one being pregnant. Please verify. 

Corrected. OK

22–Table 1, No of Females Cohabited without evidence of 

mating, for Group 4 should be 2 (4541, 4547), with one 

being pregnant. Please verify. 

Corrected. OK

23–Table 1, Pregnant, died/sacrificed: shouldn’t this be 0 as 

the dam that died, did so on LD1 and should be included in 

the category, ‘died/sacrificed post partum. 

Status corrected. OK

24–Table 3, Summary of Clinical Observations During 

Gestation: Terminal Sacrifice. It is unclear what animals 

were part of a Terminal Sac during Gestation. No animals 

died or were sacrificed during Gestation. 

Animals that did not deliver a litter were 

euthanized on GD 25 as Terminal Sacrifice. 

OK

25–Table 4, Terminal Sacrifice: For group 1, shouldn’t this 

be 20/20? 

Missing terminal sacrifices added. OK

26–Table15, Summary of Cohabitation Data: Females with 

defined day 0 of Gestation: These values include females for 

which there is a day 0 of gestation but were not pregnant. 

Should that be the case? 

Yes, that is correct. OK

27–Table 16: The category ‘Females Completing Delivery Statuses corrected. OK

15 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

with Stillborn’ will need to be revised after all of the 

stillborn/dead/cannibalized/ autolyzed are sorted out. 

28–Table 16, Duration of Gestation: Appendix R indicates 

an N of 20 for Group 1 while this table indicates 19. Please 

clarify. 

Corrected in a previous comment. OK

29–Table 16: It appears that the female that died after 

parturition (in Group 1) is excluded from the ‘females with 

liveborn gestation index’, ‘females completing delivery’, 

‘litters with liveborn pups’, ‘implantation sites’, and ‘live 

pups/litter’ . It seems that if she lived through delivery, she

and her pups could be included in these parameters.

No change to table. Exclusion from this table 

is “all or nothing”.

OK

30–Table 16, Pups dying, Missing and/or Cannibalized’ on 

Day 0 will change when these are sorted out. 

Statuses for pups corrected based on results of 

lung floatation tests.

OK

31–Table 16, Page 2, Pups Liveborn, not culled prior to Day 

21: I could not verify these values and I am not sure what 

they represent. I see 199, 196, 194 and 219 pups for this 

parameter in the Group 1, 2, 3, and 4’s respectively.) 

Values appear to be correct as is: Liveborn 

minus those culled on Day 4 = Liveborn, not 

culled prior to Day 21. 

Values cited are the Pups Surviving 21 Days: 

Liveborn, not culled prior to Day 21 minus 

pups dying, missing, and/or cannibalized days 

0-21. OK

OK

32–Table 16, Page 2, Pups dying, missing, and/or 

cannibalized: For group 4, Appendix 4 indicates 13, and 12. 

Please clarify. 

No change. Per Appendix S, 12 pups died or 

were missing on Days 0 to 4 and one on Day 

5. Table 16 indicates 12 pups during Days 0-4 

and 13 pups for Days 0-21.

OK

33–Table 20: All pages of this table are numbered 1. All of 

these tables listed as Table 20 have different headings. They 

should be separated. 

All of Table 20 is pup necropsy findings, with 

the table further broken down by type of death 

(dead, stillborn, culled, etc), therefore this 

should remain as one table. Consecutive 

pagination will be applied at report 

finalization.

OK

34–Table 20, Stillborn vs. Dead Pups: According to 

Appendix S, there are several dead pups that are being 

designated as still born. These are: Group 1 #1527-1 and 2, 

lungs do not float, but there was milk in the stomach. Group 

2 #2526-2, lungs float and there was milk in the stomach; 

#2539-1, lungs float; #2543-1, lungs float; #3528-1, lungs 

float; #3536-7,10, lungs float; #4538-1, lungs float. 

See response to section 3.2.10, Parturition and 

Litter Survival Data comment. 

Statuses for other pups corrected based on 

results of lung floatation tests.

OK

35–Since all pups dying or stillborn were reported as 

stillborn, it is unclear whether the designation of 

cannibalized, autolyzed and dead pups is consistent. 

Presently all cannibalized and autolyzed pups on Day 0 are 

reported as Stillborn. Is this how they should be reported? 

For example, Animals #2540-1, 3536-1, -2, were listed as 

stillborn, but were partially cannibalized, so no lung 

floatation was done, and it was unknown whether there was 

milk in the stomach. Should these be designated as Z, 

cannibalized, or Dead? Animal #2528-1, 3533-1, and 3536-

Statuses for 2540-1 and 3536-1 & 2 changed 

to Z-cannibalized. Dead or stillborn could not 

be determined. 

Statuses for 2528-1, 3533-1 and 3536-9 are 

OK as reported. Default is stillborn. Since no 

determination could be made due to autolysis, 

pup status remains as stillborn.

OK

16 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

9 were autolyzed with no lung floatation or stomach 

observations done. Should these be designated as stillborn 

or dead 

36–Animal #3547-1 is designated as stillborn in the data, 

indicating that lungs did not float, and there was no milk in 

the stomach, but then there is a designation of partially 

cannibalized with only the head present on Day 1 of 

Lactation (do I have this confused with pup 2?) Please 

clarify. 

3547-1 = still born on LD0 

3547-2 = partially cannibalized on LD1

OK

37–Animal 4532-8 is listed as D, but partially cannibalized 

on Day 1 of lactation. No lung floatation could be done, 

should it be classified as Z, cannibalized for consistency? 

There should be some kind of explanation (preface page) 

of how all of these pup deaths are going to be 

+summarized so the reader understands how the 

designations were assigned. 

The pup was cannibalized and autolyzed so no 

exam performed. 

Per litter observations, (Appendix R), this pup 

was alive on Day 0, so lung floatation was not 

necessary on Day 1. Status of D-dead is 

correct. 

Designation of pup death status seems to be 

self-evident: D = found dead, S = stillborn, Z 

= cannibalized (dead/stillborn determination 

not possible) or U = undetermined.

OK

38–Table 22, Summary of Pup Organ Weight Data: For the 

tissue measurements of the combined sexes, shouldn’t the N 

be 40, not 20, for controls for example? This table probably 

took a mean of the male and female weights and then 

averaged them? 

No change to Table 22. Data presented are 

litter averages.

OK

39–Appendix A, page 46, Injection Volume 86 ul: 

Acceptable Absorbance Range is 0.279 not 0.278. 

Corrected. OK

40–Appendix B, page 6, last sentence: The extraction 

efficiency evaluation data and results from 00-6131 should 

be included here. 

Added. OK

41–Appendix B, page 25, TMStd2(Reinj): the reason for 

reinjection should be given. 

Injection error reason added to footnote. OK

42–Appendix K, Computer correction data sheet indicates 

that there should be a change for Animal 2546, LD ? BW. It 

is LD 7 and this should be added to the correction sheet. The 

incorrect value was corrected. 

The data sheet has been updated to indicate 

that it was LD7.

OK

43–Appendix K, Individual Body Weight Gain Data was not 

presented for the males. 

Added. OK

44–Appendix M, Ind. Feed Consumption: Animal 1534 and 

2540 had ‘invalid feeder weights’ recorded in the data for 

Study Day 69, yet there is a food consumption for these 

animals on this day in Appendix M. Could these animals 

have just been found pregnant and this ‘invalid feeder’ 

weight was not needed anyway? Please clarify. 

The TASC data has been edited to reflect the 

documentation of ‘spilled feeder’ for this 

study day (as per invalid feeder record). 

Table 11/Appendix M reprinted.

OK

17 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

Comments Action/Response Page 

45–Appendix P, see item numbers 16 and 17 for additional 

pseudopregnancies. 

Appendix P revised. OK

46–Appendix U: Once the S, D, Z designations are sorted 

out, this appendix will need to reflect corrections. 

Appendix U re-printed. OK

47–Appendix V, Individual Terminal Estrous 

Determinations: Animals 1535 and 1536 were both 

designated in the data as Diestrous at termination, not 

estrous. 

Corrected. OK

48–Appendix W, Animal 4549, print-out dated 4/4/03: 

There was a correction page dated 4/30/03 that indicates the 

organ was supposed to be designated ‘pituitary’ and not 

‘gross exam’. As well, the finding of incisors broken/ 

missing is supposed to be an external exam finding. I 

believe these corrections were made but the latest computer 

version might not have been used for reporting. Actually 

corrections were made 5/1 and 6/16 as well. Please verify 

that the latest version is reported. 

Table and Appendix re-printed. OK

49–Appendix X will need to be updated when Stillborn and 

Dead designations are resolved. 

Statuses for other pups corrected based on 

results of lung floatation tests.

OK

50–The protocol didn’t require that culled pups be 

necropsied, but only observed externally for abnormalities. 

There is a data sheet with culled pups listed indicating ‘no 

gross observations’. Appendix X indicates culled pups with 

‘no remarkable observations’. Were these pups necropsied? 

Please clarify what no remarkable observations refers to, 

either internal gross or external exams? 

External exam only performed by the 

necropsy department.

OK

51–Appendix X, page 14: Data for animal 3528 indicates, 

‘exam not completed’. Was it completed later? What does 

this mean and why are no remarkable observations reported? 

Raw data states that the macro exam is 

complete as does the appendix for this dam.

OK

52–Appendix Y, page 7: For animal 3029, it should indicate 

‘see deviation 4’ in the spaces where there are no organ 

weight/ratios. 

Notation added to bottom of page 8 in 

Appendix Y.

OK

53–Appendix Y, page 24: For animal 4532, there was 

documentation to indicate that the left ovary weight and 

ovaries weight were to be deleted due to tech. error in 

weighing. In a memo of 1/7/03 it is unclear whether these 

data were deleted and erroneously added back in or were 

these data just to be deleted? Please clarify. 

No changes necessary. Per memo, the tech 

error was the erroneous deletion of the 

weights. Raw data listings were present in the 

data file, therefore the weights were re-input.

OK

54–Appendix CC: The certificate for Jun 20 02 3B is 

included twice, but the certificate for Jul 01 02 2A is 

missing. 

Correct certificate added to report and 

duplicate certificate deleted.

OK

18 5/25/2010 
211(b) Toxicology Research Group 

TBA One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist 

19 5/25/2010 

Comments Action/Response Page 

Data Issues: 

Individual Estrous Cycle Data: For animal 1531, for 

intervals 7, 8, and 9, the stage of cycle was changed on 

10/28. The original data was collected on 10/14, 15, and 16, 

and reviewed on 10/17. How could these data be changed on 

10/28? 

The slides for each animal for every day of 

smearing are retained until approval is given 

by study director to discard. As the slides 

(original raw data) are kept, the vaginal 

smears on days 7, 8 and 9 for animal 1531 

were re-read by a technician on 10/28. Based 

on the final reading of the smears, a decision 

was made to edit the cycle to what was read 

on 10/28. 

OK