211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

1 5/4/2010 

Comments Action/Response Page 

EPA 

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: 

The peer reviewers were concerned that statistically 

significant increases in testis, seminal vesicle, and ovarian 

weights relative to total body weight in the high exposure 

group were considered spurious. It appears that this poor 

correlation was related to the significant weight loss reported 

in the parental animals in the 20,000 mg/m3

 exposure group, 

since the application of covariance analysis eliminated the 

earlier statistical significance of the individual organ 

weights. We would request that the study group perform a 

statistical evaluation to confirm that the observed 20% 

increase in seminal vesicle weight (relative to body weight) 

does not reach significance when compared to brain weight. 

Additional discussion added to end of section 

3.4.2. 

Ok 

pp 41-42 OK

 

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. However, there does seem 

to be some indication that the 2,000 mg/m3

 exposure level is 

not a NOAEL, as noted below. 

Dr. Schlesinger’s comment concerning the 

NOAEL = 2,000 mg/m3 seems to refer to 

lower absolute body weights in low dose 

female rats premating. However, the effect 

was statistically significant in the 2000mg/m3

females at Day 63 (p<0.05) only; on Day 69, 

the average female body weight in the low 

dose group was within 10g of control value 

compared to weights lower by 17 and 19g 

respectively in the 10000 and 20000mg/m3

groups. In addition Table 6 demonstrated that 

over the entire period, body weight gain was 

not significantly lower in 2000mg/m3

 females. 

The RG agrees with the study director’s 

evaluation of the absence of biological 

significance for decreased body wt at the low 

dose. 

Ok 

Specific comments are as follows: 

There should be consistency in the Group numbers. In some 

places they are noted as I, 11, III, and IV, while in others 

they are noted as 1,2, 3, and 4. 

This inconsistency has been brought to the 

attention of the study director and, where 

practical, adjustments have been made. 

However, in computer generated tables and 

compilation of raw data the number format is 

predetermined. This observation does not 

affect the validity of the data 

Ok 

p. 29 Statistical Analysis: 2.15.1. 

It is not clear why a one-way ANOVA was performed at 

each time interval, resulting in the need to perform multiple 

ANOVAs. 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 an appropriate multiple comparison test. 

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. 

Ok 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

2 5/4/2010 

Comments Action/Response Page 

What is the rationale for having a separate discussion of the 

method of analysis of the organ weight data when these data 

also used the ANOVA procedure? 

Furthermore, if Bartlett's test showed there to be unequal 

variances, why was a parametric test still used? 

Organ weight statistics were discussed 

separately because, unlike other parameters, 

Bartlett’s test was used here to assess 

variance. Parametric tests were used only 

with homogeneous data. If data were nonhomogeneous, the non-parametric Cochran & 

Cox’s modified t-test was used to evaluate 

significance. RG will discuss rewording of 

the first part of this paragraph for clarity with 

the study director. 

We have reviewed and not made any changes 

because: 

1. 1st paragraph describes TASC procedures 

(all parameters except parental organ 

weights). 

2. 2nd paragraph describes Xybion procedures 

(parental organ weights). Xybion’s stats are 

wholly parametric (no non-parametric), so the 

statement about Cochran & Cox above is not 

correct. 

If the 2nd paragraph refers only to Xybion 

statistics why is Cochran and Cox modified Ttest mentioned in that paragraph (see p. 30). If 

entry is incorrect should it be deleted from 

text or explained differently? 

Text is correct per Xybion stat package 

3. 3rd paragraph describes Startox procedures 

(analysis of co-variance for selected organs).

Ok 

Provide the rationale for the ANOVA procedure with certain 

organ weights using terminal body weight as a single 

covariant. Which organ weights were evaluated using this 

procedure? 

Absolute organ weights that fall within normal 

range may become significantly different from 

controls in the context of terminal body 

weights that are lower or higher than controls. 

Where significance in relative organ/body wt 

seems skewed due to body weight variation, 

the analysis of covariance is used to assess the 

statistical significance of these values. 

Organs evaluated by covariance were 

epididymis, seminal vesicle, testes and ovary 

weights in the 20000mg/m3

 group to 

determine if statistically significant increases 

relative to body weight identified by 

Dunnett’s test were accurate. Analysis of 

covariance demonstrated that the earlier 

statistical significance was spurious for all 

organs. In addition there were no statistically 

significant increases in absolute organ weights 

or organ weights relative to brain weight. 

Ok 

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. 

Both 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 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

3 5/4/2010 

Comments Action/Response Page 

It is noted that, "Dose groups were eliminated from statistical 

analysis if their standard deviation was 0 and/ or N...in the 

group was less than or equal to two." How many such cases 

were encountered in the study? 

This is a standard statement for the statistical 

methods section. In this study, Table 1 

Summary of Survival and Pregnancy provides 

examples of where the n is 0 or 1 and this 

statement is relevant. For example, No 

animals died pre-mating in controls and high 

dose groups but one each died in the low and 

mid dose groups. No statistical analysis was 

applied. 

Ok 

p. 35 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 correspondence between the 

measured and nominal levels is very good, so it is not clear 

why any explanation in this regard is warranted. 

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 to calibration error. In any 

event, the inaccuracies are believed to be 

slight. The RG feels confident that the 

reported chamber concentrations accurately 

reflect the atmospheres to which the animals 

were exposed. 

HLS – will you want to delete the calibration 

error explanation 

Yes and deleted. OK

The investigators need to explain the source of the particles 

in the exposure atmospheres. Was the incoming chamber air 

filtered appropriately? Any particles could ad as adsorption 

sites for vapors in the exposure atmosphere. 

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 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 36 indicates that 

the particle concentrations in all of the 

chambers including controls ranged from 

about 7 to 9 micrograms/m3

 per cubic meter 

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. 

ok 

p. 37 Bodv Weight: 3.2.3. 

The investigators state that the only exposure related effects 

were in males and females at the highest concentration, and 

females at 10,000 mg/m3

. However, data in Table 5 show 

Overall gasoline +ETBE vapor exposed 

animals showed slight to significant lower 

absolute body weights during pre-mating. 

ok 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

4 5/4/2010 

Comments Action/Response Page 

significant difference at Day 63 in females exposed at 2,000 

mg/m3

 and a trend upward in the same exposure group at 

Day 69. Thus, it does appear that, with increasing time of 

exposure, there is a real effect on this parameter in the lowest 

exposure level group. 

However, the effect was statistically 

significant in the 2000mg/m3 females at Day 

63 (p<0.05) only; on Day 69, the average 

female body weight in the low dose group was 

within 10g of control value compared to 

weights lower by 17 and 19g respectively in 

the 10000 and 20000mg/m3

 groups. In 

addition Table 6 demonstrated that over the 

entire period, body weight gain was not 

significantly lower in 2000mg/m3

 females. 

The RG agrees with the study director’s 

evaluation of the absence of biological 

significance for decreased body wt at the low 

dose. 

p. 38 Maternal Body Weight: 3.2.4. 

Data in Table 7 and 9 indicate what appears to be an early 

decrease in weight gain at 20,000 mg/m3

. Furthermore, 

clarification is needed concerning the last sentence in this 

section regarding differences in the premating period. 

Table 7 and 9 show absolute body weights 

throughout gestation and lactation, 

respectively, not weight gain. Wt gain data 

during gestation is provided on Table 8, and 

there are no statistically significant differences 

in wt gain during this period. Although the 

high dose females showed significantly lower 

absolute weights during gestation and 

lactation, they started gestation weighing less 

than controls due to low pre-mating weight. 

The weight gains during gestation and 

lactation were comparable to controls. The 

RG accepts the evaluation of the study 

director. No changes in text have been made. 

Ok 

p. 40. Postmortem Data: 3.4.1. 

Data in Table 19 appear to indicate a dose-related trend of 

increasing lung discoloration. 

The incidence of lung discoloration in each 

dose group (sexes combined) does show a 

general increasing trend in the mid and high 

dose groups [control 9.8%; low 8.2%; mid 

17.6% and high 16.7%]. The report text (p. 

40) indicates that lung discoloration was noted 

but in the absence of any effect on lung wt or 

histopathology was not considered treatment 

related.” Occurrence of discolored foci is 

considered a background finding which may 

occur non-specifically with a variety of 

inhaled materials, and sometimes is correlated 

with histologic identification of alveolar 

macrophage aggregates. In this study, the 

pathologist did not report any remarkable 

changes in lungs of treated animals to support 

a significant correlation with grossly observed 

lung discoloration. The RG accepts the 

evaluation of the study director. No changes 

in text will be made 

OK

p. 41. Organ Weight: 3.4.2. 

The investigators note that there is an effect on kidney 

weights at all exposure levels in the males. Table 21 seems 

Table 21 evaluates relative organ/body weight 

data. Two statistical methods are used, one 

Ok 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

5 5/4/2010 

Comments Action/Response Page 

to indicate such an effect only at the two highest levels of 

exposure. 

for homogeneous data and one for nonhomogeneous data. The statistical significance 

at the two highest doses was identified using 

Dunnett’s test, and the significance at 

2000mg/m3 was determined using the 

Modified t-test for non-homogeneous data. 

The symbols used to designate significance 

are different depending on the test employed 

(see footnote on Table 21). 

It is not clear whether the lack of any histopathologic 

correlation with the change in liver weight is meant to 

suggest that the noted effect on weight is of no biological 

significance. 

As pointed out by Dr. Goldsworthy, the 

statement on p. 41 indicating the absence of 

histological changes in livers is incorrect. 

Table 1 in Methods (p.26) indicates that no 

microscopic evaluation was performed on the 

livers of any animals. This is confirmed by 

the RG review of Appendix Z, Individual 

Animal Histopathology, in which “no 

micropathologic observations on tissue” was 

listed whenever necropsy data on livers was 

cited in any group. The study director will 

correct this statement. 

Statement has been corrected and considered 

in the NOAEL of 2000mg/m3

. OK

Concerning the biological significance of the 

increased liver weight in mid and high dose 

male rats, the effect was considered 

significant enough to be used as a factor in 

setting the parental NOAEL at 2000mg/m3

. 

What is the basis for the conclusion that changes in 

epididymis, seminal vesicle and ovary weight are considered 

to be spurious? 

See response on top of p. 2 addressing “Use of 

analysis of variance for certain organs using 

terminal body weight as a single covariant. 

Discussion has also been expanded on pp 41-

42. 

Ok 

Table 21 

Statistical results are indicated for two different tests. Why 

were two tests on the same dataset, and is the reader to 

accept a conclusion of significance if either test indicates 

this? Some rationale for this type unorthodox statistical 

procedure needs to be provided. 

The statistical routine is a conditional regimen 

with different tests required for data sets 

depending on whether they are homogeneous 

[Dunnett’s test] or non-homogeneous 

[Modified T-test]. This is typical for 

statistical evaluations of toxicology data. The 

reader should never assume that statistical 

significance with either test equates to 

biological significance without reviewing the 

data set carefully including the pattern of 

response, the direction of response, the degree 

of response, etc. 

Ok 

Peer Reviewer – Goldsworthy

The study was scientifically sound and was properly 

conducted. Exposures were properly conducted and adequate 

toe test the potential reproductive toxicity of Gasoline ETBE 

Vapor Condensate. None of the protocol deviations noted 

No response needed 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

6 5/4/2010 

Comments Action/Response Page 

would be expected to adversely affect study results, data 

integrity or conclusions. 

No effect of treatment was noted on survival, clinical 

observations, feed consumption, estrus cycles, mating, 

fertility or gestation indices, or no other reproductive 

parameters up to parturition. An exposure-related decrease 

in body weight gain was observed in high dosed male and 

female rats, and the mid-dosed female rats during the premating interval. Text (on page 37) notes the decrease in 

body weights in female rats exposed to the 10,000mg/m3

gasoline-ETBE vapor condensate; however, neither the 

summary (pages 5-7) nor the conclusion (page 43) note this 

adverse finding. This should be noted in both the summary 

and the conclusion. 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. Additional sperm count, motility and morphology 

was not altered by the treatment. 

Dr. Goldsworthy’s comment is correct. The 

Conclusions and Summary will be revised to 

include decreases in body weight for 

10,000mg/m3

 females and cite the criteria in 

setting the NOAEL where appropriate. 

Done. 

Checked both locations Summary entry OK 

Conclusion p. 43 As presented in Summary, 

add “females only” in parentheses after 10000 

to read “ decreased body weight gain in the 

10000 (females only) and 20000mg/m3

exposed parental animals” 

Done 

ok 

There were no exposure-related differences in macroscopic 

postmortem assessments. Increased kidney weights were 

observed in male rats (parental generation) at all exposure 

levels; this is consistent with the expected induction of 

hydrocarbon necropathy. Microscopic evaluation was not 

performed to confirm this observation. Liver weights were 

increased at the two higher exposure levels in male rats. 

Text on page 41 notes that this alteration "were not 

correlated with any histopathological finding". However, the 

tissue list on page 26 notes that liver was not examined 

microscopically; thus text should be modified. It is not clear 

why, in this study, brains, kidneys, liver and spleen were not 

examined microscopically when they were examined in the 

gasoline-DIPE one generation reproductive study. There 

were statistical differences noted in a number of organ 

weights in treatment groups, especially the high dose group; 

these differences, which included alterations in 

epididymides, testis, ovaries, seminal vesicle and spleen 

weights, were considered not attributed to the test article 

because of the absolute differences noted, the lack of a 

treatment-related patter, and the lack of correlation to body 

weights. Whereas it may be correct to not attribute these 

changes to treatment, these differences are significant in 

nature to note in summary section. There were no exposurerelated difference in body weights, macroscopic postmortem 

evaluations and organ weights in lactating pups from Air 

Control animals. 

Liver weights: This statement will be 

corrected. See response above to Dr. 

Schlesinger comments (p. 4, second paragraph 

under Organ wts). 

Brain, kidney, liver and spleen were not 

examined histopathologically because it was 

decided by the RG that sufficient 

histopathology data on these organs was 

available from the 90-day BG+ETBE study in 

which no significant effects were observed, 

and by read-across from other gasoline+ 

oxygenate blend studies. In actuality, the 

histopathology performed in the DIPE 1-

generation reproduction study was a deviation 

from the standard protocol for the 1-

generation studies. 

The statistically significant increase in relative 

weights of epididymis, testes, ovaries and 

seminal vesicles at 20000mg/m3

, were also 

cited by Dr. Schlesinger. Dr Goldsworthy did 

not disagree with the conclusion that the 

significance of these changes as treatment 

induced events was spurious, but as he did 

recommend that these effects be included in 

the summary section with appropriate 

explanation, the RG requests the study 

director to provide a brief comment on these 

findings indicating they are not treatmentrelated events 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

7 5/4/2010 

Comments Action/Response Page 

Additional discussion added to end of section 

3.4.2. OK

In summary, test animals were properly exposed to 

adequately test the potential reproductive toxicity of 

gasoline-ETBE. A NOAEL of 2,000 mg/m3

 was 

determined. This NOEL setting is based on decreased body 

weight gains and increased liver weights in parental animals 

in the 10,000 mg/m3 and 20,000 mg/m3

 groups [note text in 

summary and conclusion sections do not mention the 

decreased body weight gain with regard to the NOAEL 

setting]. The NOAEL setting of 2,000 mg/m3

 does not take 

into account the increased renal weights observed at all 

exposure levels; this finding is consistent with hydrocarbon 

nephropathy observed in similar animals exposed to test 

substance and generally regarded as not to be relevant to 

human risk assessment. There was no effect at any exposure 

level on reproductive performance, including mating, 

fertility, parturition, lactation, and offspring survival. Thus, 

with respect to reproduction performance, a NOAEL of 

20,000 mg/m3

 was appropriately concluded. 

Dr. Goldsworthy pointed out that the parental 

NOAEL of 2000mg/m3

 did not take into 

account the renal changes in male rat and 

perhaps this should be stated. 

HLS please include a statement in the 

Conclusion and Summary that the NOAEL 

was determined exclusive of the renal effects 

due to light hydrocarbon nephropathy seen in 

male rats, since these effects are not relevant 

to humans. 

We have been consistently stating in all of the 

211(b) reports that the male renal effects are 

not considered in the NOAEL setting. OK

 

211 (b) Research Group Reviewer 

General Comments: 

The report should have sequential numbering from start to 

finish. 

This will be done at report finalization. OK

The numbers for cannibalized/stillborn/missing/found dead 

are confusing; please see comments related to T. 20. 

No change to report. OK as is per Robert 

Parker’s review. OK

Historical control data is listed with released pups as part of 

App. A, but also appears in the Table of Contents as App. 

DD. 

TOC corrected. OK

However, App. DD is the Protocol and Amendments, and the 

subsequent appendices are also in error. Please straighten 

this up. 

TOC corrected. OK

Specific Comments: 

P. 5, Summary, 4th paragraph 

Please consider changing the word, "reasonably" to 

"acceptably," since there should be an SOP or protocolspecific criteria for concordance of nominal and analytical 

concentrations. The same change should be made later in the 

same paragraph on p. 6. 

Revised and also in the R&D. OK

Also, please consider deleting the sentence, "The exact cause 

of the differences ....." as unnecessary. 

Deleted. OK

P. 6, 2nd paragraph 

Female premating body weight gain was also decreased at 

10,000 mg/m3 (p. 37). 

Revised. OK

P. 6, 3rd paragraph: Please spell "2". Done 

P. 7, 3rd paragraph 

Again, female premating body weight gain was also 

decreased at 10,000 mg/m3. 

Revised. OK 

P. 16, Dispensing 

Chevron Research and Technology Company. The supplier 

Revised to Chevron Global Technology 

Services Company (per other reports). OK
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

8 5/4/2010 

Comments Action/Response Page 

can be described either as Chevron Global Technology 

Company, which is the name on the contract, or as Chevron 

Research and Technology Company, that provided the 

sample, whichever QA considers to be most appropriate. 

P. 22, Estrous Cycling 

Please note the study days for the start of the daily vaginal 

smears. 

Added. OK

P. 23, 2.12.4, Litter Observations 

Please make note of the lung floatation test to distinguish 

stillborn and found dead pups, also that autolyzed remains 

found on the day of parturition were presumed to be 

stillborn. 

Added. OK 

P. 32, Protocol Deviation 1 

Please note the sex and exposure group of the two animals 

not exposed on Day 97. 

Added. OK 

P. 33, Protocol Deviation 8 

Thank you for describing the incident clearly and 

thoroughly. Could you also please add the phase of the 

lactation period when the incident occurred? I.e., most or all 

of the pups were LD > 25. 

Added. OK LD was 25-27. 

P. 35, final paragraph 

Please consider changing the word "reasonably" to 

"acceptably" as requested above. 

Also, please consider deleting the sentence, "The exact 

cause.. ." as unnecessary. 

Revised. OK

P. 37, Mortality/Physical Observation data 

The two animals killed accidentally are not described 

similarly in Table 2. 2073 is listed as found dead. 4553 is 

listed as accidental death. Shouldn't these be described in the 

same manner if the same thing happened to each of them? 

Revised. OK

Body Weights- Premating Period 

Please see note below regarding App. H. 

See response below. 

P. 38-39, Estrous Cycle Data 

The data suggest that pseudopregnancies were included in 

the group - summaries and comparisons, which shouldn't 

happen. Please check 1559, a control animal with a single 18 

day cycle, appears to be included in the control group 

summary. This value is suggestive of an artifactual factor 

and diminishes the ability to detect intergroup differences. 

We have reviewed the data and the tabulation 

and confirm that pseudopregnancies were not 

included (including for #1559) in the group - 

summaries and comparisons. OK

P. 40, Pup Body Weight Data: 

Please include an explanation for the multiple age ranges for 

which weight gain was evaluated. 

Pup weight changes are preselected by the 

TASC system and there is no way to change 

which day ranges to include/exclude. OK

P. 41, Organ Weights, 2nd paragraph 

- Please spell "2". 

- Also, liver weights were only affected in males; please 

include gender as a qualifier in the first sentence. 

-

- Done OK

- Done OK

3rd paragraph 

It's been my understanding that testes and ovaries aren't 

usually well-correlated to body weight, and that absolute 

organ weight is usually the preferred endpoint for their 

evaluation. Are there other reasons to consider these 

statistically significant differences to be spurious that could 

We have reviewed and confirm that the 

offered explanation is sufficient. 

[See expanded discussion on pp 42-43] 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

9 5/4/2010 

Comments Action/Response Page 

be cited? 

P. 43, Conclusion 

Decreased body weight gains were also noted in the 10K 

females (p. 37), and so should be noted here as well. 

Added OK

Tbl. 14 

For the control group, please footnote that the value for the 

number of estrous cycles includes one animal with no 

complete cycle. Please see above, 1;..38-39, regarding 

concern about the control group cycle length value. 

Animal was pseudopregnant and reformatted 

table excludes this animal. OK

Tbl. 19 

I am disappointed that GD 25 animals were terminated 

without necropsies; please explain. 

This was done as per protocol amendment#1, 

point#2. OK

Tbl. 20, p. 1 

Please indicate if cannibalized should be interpreted as 

missing. 

No: cannibalized means that some tissue was 

still present. OK

Tbl. 20, p. 2 Lung Floatation Test - Found Dead 

The number for the controls should be three, not two. Please 

see comment below. 

Corrected. OK

Tbl. 20, p. 4: Lung Floatation Test - Found Dead 

App. W, p. 3, 

Indicates that #1561/14F should not be listed as a stillborn 

based upon the results of the lung floatation test and milk in 

stomach. Please correct Tbl. 20. 

Corrected. OK

Tbl. 20, p. 5: Lung Floatation Test – Stillborn 

The numbers match up if the autolyzed and cannibalized 

pups are included. This may need footnoting for 

clarification. 

Numbers for Lung Floatation Test – Stillborn 

match Appendix W. Pups/Litter Evaluated 

includes cannibalized/autolytic pups for which 

the floatation test could not be performed. 

Footnote added to pages 4 & 5. OK

App. H. p. 5- 8: 

Individual body weight for the Day 0-69 interval should be 

shown as in Table 6. 

Added. OK 

App. S, p. 1 

1561114F is listed as stillborn, but according to App. W 

individual pup necropsy observations, this pup should be 

classified as found dead. Please correct. 

Corrected. OK 

App. U, p. 1 

Please make the same correction for 156111 4F 

Corrected. OK 

App. AA 

There are multiple page 1's. This seems like a bad idea and 

could be confusing. This appendix includes historical control 

data for pups; should this be in App. DD? 

Pagination will be corrected in the final report. 

HCD is in Appendix AA (TOC corrected). 

OK

App. DD: The historical control data appendix is missing. TOC corrected. OK

App. EE - GG: All are mislabeled according to the Table of 

Contents. Please correct. 

TOC corrected. OK

 

211(b) Research Group QA/QC Reviewer

The following items are comments or require further 

consideration: 

 

Compliance Statement 

The sponsor also needs to sign a compliance statement. It No change. Sponsor signature line is already 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

10 5/4/2010 

Comments Action/Response Page 

can be a separate one from the Testing Facility’s, but there 

must be one signed by the sponsor. 

included on the Compliance Statement. OK 

Page 2, Compliance Statement 

Since it was the sponsor’s responsibility to maintain the 

method of synthesis, fabrication, or derivation of the test 

fuel, and this had not been completed, it must be included in 

the sponsor’s compliance statement. 

Exception added. OK 

Page 4, Quality Assurance Statement: 

Since HLS has inspected the sperm evaluation subcontractor 

(PAI) and is responsible for ensuring their GLP-compliant 

status, these facility inspections should be listed on the QA 

statement for this study. 

Added. OK 

Draft Report, Page 3 

Should the signature of the reproductive toxicologist who 

reviewed this report also be added to the signature page? 

Signature page revised OK 

Draft Report, Page 13, Section 2.2.6 Terminal Sacrifice 

The dates for sacrifice for the nonpregnant females have not 

been included (June 22, 23, 25 and July 5). 

Added OK 

Draft Report, Page 19, Section 2.9.8 Bedding Analysis 

States that analyses for bedding can be found in Appendix 

CC. Analyses are available, but are from 1999. Are there any 

more recent/representative analyses? 

These are the analyses that were received with 

the lots used on this study. OK 

Draft Report, Page 32, Protocol Deviations 

All adult animals were to receive gross necropsies. Since the 

non-pregnant females were not given a necropsy and tissues 

were not saved, this is a deviation and should be included in 

this section. Protocol Amendment No. 1 indicates they will 

be euthanized, but doesn’t indicate a change with respect to 

necropsies and tissue harvest. 

Deviation added to report and data OK 

Draft Report, Page 38, Section 3.2.4 Maternal Body 

Weights, Gestation and Lactation Periods 

The absolute body weights for the females in the highest 

level were statistically significant during lactation and 

gestation. Should this fact be discussed in this section? 

Additional discussion added to this section. 

OK 

Draft Report, Page 37, Section 3.2.1 Mortality – The raw 

data records that male rat #2703 was found dead, not 

“accidentally killed” as the text on Page 37 states. 

Corrected. OK 

Draft Report, Page 42, Section 3.4.3, Released Pup Data, 

Appendix AA: It may not be accurate to indicate that this Air 

Control set of pups (released) was comparable to the 

historical control pups of similar age as in Appendix AA, as 

it appears that Day 21 values are used for Study 4202, 

making the mean much lower than it should be. The 

historical values range from 73g to 82g, where these values 

are around 66g. The nonreleased pups are even a little 

smaller than the historical control. Were statistics run on 

these to verify that these animals were similar to historical 

controls? 

Appendix AA, Study 4202 terminal body 

weights corrected to Day 28 values. OK 

Draft Report, Table 16 – The indices should be calculated to 

include survival to Day 28. 

All calculated indices stop with Day 21. 

No change possible due to system limitations. 

OK 

Draft Report, Table 16, Groups I and III – The Number of 

pups dying, missing and/or cannibalized Days 5-21 and 0-21 

should be checked. This data also changes the pups surviving 

Numbers reported on Table 16 appear to be 

correct and match data in Appendices R and 

S. Still cannot verify some values, but they 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

11 5/4/2010 

Comments Action/Response Page 

21 days and the lactation index for these two levels. 

Appendices S and R do not agree with the Table 16 

tabulations. 

are close. 

Groups I and III on Table 16 checked against 

Appendix R and Appendix S. No changes 

apparent: 

Group I – 1558-11 (Day 9), 1562-2 (Day 15) 

and 1562-5 (Day 16) = 3 pups 

dying/missing/cannibalized Days 5-21. 

6 pups on Days 0-4 + 3 pups on Days 5-21 = 9 

pups Days 0-21. 

Group III – 3555-4 (Day 21) = 1 pup 

dying/missing/cannibalized Days 5-21. 

4 pups on Days 0-4 + 1 pup on Days 5-21 = 

5 pups Days 0-21. OK 

Draft Report, Table 17 – The observation of laceration mid 

abdomen has not been reported (Animal #355-9). Also the 

observation for tip of tail missing should be 2/2 for Group 

IV. Animals #4561-2 and 4561-7 both had the tip of the tail 

missing for one observation. 

Laceration observation added. ok 

Tip of tail observation was seen in 2 pups in 1 

litter. Frequency/animal is the number of 

times an observation occurs/number of litters 

it was seen in. ok 

Draft Report, Table 18, Summary of Pup Body Weight Gain 

– This table does not report the weight gain for Day 4-14 and 

Days 7-21. The only weight gain report from Day 1 was 

Days 1-4. Should the other time periods also be reported for 

weight change (Days 1-7, 1-14, 1-21, 1-28)? 

Not available based on TASC program hardcoded output. Ok 

Draft Report, Table 19, Gross Observations Preface: Animal 

number 2559 is mentioned twice when I believe number 

2553 should be referred to. Please verify. It also indicates 

that observations for this animal are presented in Appendix 

V. Appendix V is Estrous Cycle determinations and does not 

contain gross observations. 

Animal number corrected. ok 

Reference to Appendix V removed. ok 

Draft Report, Table 20, GROSS EXAM: I cannot verify the 

“Litter Incidence or Pup Incidence” values for this table. 

What do they mean? It is not the litter incidence of dead 

pups as that would be a much higher value. 

The table is divided into Pups Died, Pups 

Stillborn, Pups Culled and Weanlings. Litter 

incidence and pup incidence are dependent on 

these sub-sets. OK 

For instance, Pups Died in Group 1: Three 

pups have gross exams (1562-2, 1562-5 and 

1573-10). The Litter Incidence for Gross 

Exams is 2 litters and the Pup Incidence for 

Gross Exams is 3 pups. Since all three pups 

were found to be autolytic, the same Litter and 

Pup Incidences can be found for Autolysis.

Draft Report, Table 21, Parental Organ Weights: Should the 

animals (dams) from the litters on the rack that fell be 

included with others for terminal body and organ weights? 

Some of these animals were lactating 2 or 3 days fewer than 

others. The same question could be raised for Appendix K 

where lactation day 28 weights are reported for all animals 

(except not pregnant). Lactation may be at a minimum at this 

point, but some body weight difference with these animals 

may be realized. 

We have reviewed and do not feel it is 

necessary to make any changes since weaning 

occurs around LD21. Any differences these 2 

to 3 days make would be minimal. Weight 

changes from Days 21 to 28 were comparable 

for all groups. OK 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

12 5/4/2010 

Comments Action/Response Page 

Draft Report, Table 23 

The Table number should appear on the Table itself and not 

just the cover page. 

No change possible due to Xybion limitations. 

OK 

Appendix A, Page 2 and 3 

The duration of the temperature and humidity measurements 

that were out of range should be indicated. 

No temperature excursions. 

Incidence of humidity excursions noted. OK 

Appendix A, Page 4, Frequency of Administration 

Typo (2 periods) in the middle of the paragraph. 

Corrected. OK 

Appendix A, Page 7, Section 2.3.1: “Concentration”. Corrected ok 

Appendix A, Page 20 

The GSD value for April 17 is 1.663, I believe. Please 

verify. 

Corrected. OK 

Appendix A, Page 26 

The nominal value for March 20 is 2155 and should 

probably round to 2200. Please verify. 

No change. 

Data indicates nominal value is 2150. ok

Appendix A, Page 35 

The MMAD for May 22 should be 12.27. Please verify. 

Corrected. OK

Appendix A, Page 37 

Please verify the nominal value for July 15. 

Nominal value reviewed and corrected. ok

Appendix A, Page 40 

The temperature for May 16 should be 26. Please verify. 

Corrected. OK

Appendix A, Page 31, July 3, Individual Analytical Chamber 

Concentration 

The last value for this day is actually a mean of 2 values 

taken during a 7- minute span. Usually when a low or high 

value is noted, adjustments are made and additional assays 

are performed to verify that concentrations are indeed closer 

to target, and the original value is reported as it is the most 

representative for the time interval. Why was an average 

reported and why were all values not used? Please clarify. 

All values were reported as per laboratory 

SOP which states that when a sample is not 

within 20% of the target concentration, a 

repeat sample is taken. If the repeat sample is 

within 20% of the original sample, the original 

sample is reported. If the repeat sample is not 

within 20% of the original sample, a third 

sample is taken. All three samples are 

evaluated and the 2 values closest in 

arithmetic agreement (providing the higher 

value is within 25% of the lower value) are 

averaged and this average is reported. 

ok 

No adjustments are made to the system 

between the original and repeat samples.

Appendix A, Page 47, Group 2 

The H-1 value is not reported. It was run after the second H3 measurement and was 2000 mg/m3. 

Corrected. ok

Appendix A, Page 49, Methodology 

The pathlength should be 3.39, I believe. The first setting 

was 7.00, but it was reset to 3.39 on 6/2/02. Please clarify. 

Corrected. OK

Appendix A, Page 49, Calibrations 

The calibration from 10/10/01 was used for this study as 

well. 

Per the data, only 1 calibration (10/10/01) is 

included in the data and report. OK

Appendix A, Page 50, Calibration Checks 

None of the absorbance values fell outside of the 15% range, 

so the rest of this paragraph is not needed. 

Deleted. OK

Draft Report, Appendix B, Page 25 

The value reported for benzene in the control sample should 

be < LOQ and the value for Toluene should be ND to agree 

with the raw data. 

Corrected. OK

Draft Report, Appendix B, Page 26 Corrected. OK
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

13 5/4/2010 

Comments Action/Response Page 

The value reported for the control sample benzene should be 

ND and the toluene value should be < LOQ to agree with the 

raw data. 

Draft Report, Appendix B, Pages 27 through 30 

Should state that these gas chromatograms were obtained 

from the Day 21 exposure samples. 

Indicated that the Gas Chromatograms were 

obtained from Exposure 21 Samples on Pages 

27 through 30. OK

Draft Report, Appendix B, Page 28 

The retention time value for 3-Methyl-1-butane should be 

reported as 19.6 not 19.80. The retention times for trans-2-

pentene, 2 methylpentane, 2,4 dimethylpentane and 2,3-

dimethylpentane are not represented on the chromatograms. 

(These four compounds are also missing from the 

chromatograms on Pages 29 and 30.) 

No correction is made. Due to the limited 

space on the chromatograms, the retention 

times for trans-2-pentene, 2 methylpentane, 

2,4 dimethylpentane and 2,3-dimethylpentane 

were not shown properly on the 

chromatograms. OK

Draft Report, Appendix B, Page 30 

The retention times for N-pentane and 2,3 dimethylbutane do 

not agree with the raw data values (23.08 report vs. 23.19 

data and 32.99 report vs. 33.11 data). 

Same as the above. OK

Draft Report, Appendix D, Page 15 

Female #1555 is missing the Clinical Observation of 

Terminal Sacrifice (also Females #2559 and 2533; on Page 

16 Females #4552 and #4562). 

Animal Nos. 1555, 2553, 4552 and 4562 were 

found mated and gestation physical 

observations were collected. Appendix D 

only includes non-mated females during the 

mating and post-mating periods. 

Terminal Sacrifice for female No. 2559 added 

to Appendix D. OK

Draft Report, Appendix D, Page 21 

Females # 4553 observation states “accidental” should state 

“accidental death”. 

Corrected (also Appendix C and Table 2). 

OK

Draft Report, Appendix D and Table 2 

The observation for Male Animal #3055, “humane sacrifice, 

black/brown stains on snout and grasping” are reported in 

Appendix D as the 56th day of the study, but Table 3 has 

these observations reported as Day 63. Also on Day 56 the 

observation of “within normal limits” is reported. This data 

should be reviewed and appropriate changes made to Table 

2. 

Appears to be reported correctly. Animal was 

sacrificed on Day 59. When the death of an 

animal occurs between scheduled observation 

intervals, the individual animal data 

(Appendix D) is presented at the previous 

interval (Day 56), but the summary table 

(Table 2) presents the data at the next interval 

(Day 63). OK

Within Normal Limits was noted for Animal 

No. 3055 during the scheduled observations 

on Day 56. Black/Brown Stains on Snout and 

Gasping were noted on Day 59 prior to 

sacrifice for humane reasons (Room Check 

obs). Notation added to preface page to Table 

2; Preface page with explanatory notes added 

to Appendix D. OK

Draft Report, Appendix D, Page 10 

Male Animal #4055 “decreased food consumption and 

decreased fecal volume” are reported twice for this animal. 

The total days for this observation are not listed correctly in 

Table 2. This data for all observations for this animal needs 

to be re-evaluated and tabulated correctly. 

Reported correctly in Appendix D and Table 

2. Observations were noted at both the 

weekly physical examination and during the 

daily viability check (room check) and are 

categorized separately. The duplication in 

Appendix D indicates these observations were 

seen at both types of exam. Preface page 

added to appendix explaining this apparent 

duplication. OK
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

14 5/4/2010 

Comments Action/Response Page 

In Table 2, one set of these observations is 

summarized in the Miscellaneous category 

(physical examination), and the other in the 

Room Check category (viability check). A 

notation has been added to the preface page. 

OK

Comment: Draft Report, Appendix G 

When an animal is no longer on study, a footnote should be 

added to clarify the missing weights – i.e. Males #2073 and 

3055. 

Footnotes added to Appendices G, H and M. 

OK

Draft Report, Appendix P 

The calculations for number of days in stage, number of 

cycles and mean number of days of cycle are calculated 

using the data obtained from Day 49 up to Day 69. The data 

from Days 70 to 83 has not been included in these 

calculations. If this data is included, the summary data will 

change (i.e. female #1559, the number of cycles changes 

from 1 to 4 and the mean number of days of cycle changes 

from 18.0 to 7.8). 

Appears to be reported correctly. Data is 

summarized for the 21 day pre-mating 

evaluation period only. Since the length of 

the co-habitation period varied from 1 to 14 

days, including these data would skew the 

results. 

Appendix P was regenerated to reflect the day 

on which estrus cycles were monitored within 

the exposure period. Estrus cycle monitoring 

started on test day 49 and continued for 21 

days before mating was started. Preface 

indicates estrus cycling was evaluated for 21 

days prior to mating and then up until 

successful mating. Only the 21 days prior to 

mating is used in the assessment. 

Agreed OK

Draft Report, Appendix Q 

Should NA be added to the entry date of insemination, 

instead of leaving a blank space for the animals that were 

pregnant and a sperm/plug was not observed? 

It would seem that adding NA would imply 

the animal was not pregnant. Insemination 

DID occur; the laboratory staff just missed 

seeing any evidence (sperm/plug). 

Ok

Draft Report, Appendices R and S, Group 1 

The values for Day 28 should be corrected to reflect the 

litters that were sacrificed early due to the rack overturning. 

These pups did not survive to Day 28. Also for Appendices 

T and U the litters should be labeled that did not survive to 

Day 28. 

Appendix R – Litter Data. Notation added to 

Day 28. No other change necessary. ok

Appendix S – Sex and Status. Day of 

sacrifice corrected for pups sac’d early. No 

other changes necessary. ok

Appendix T – Pup Clin Obs. Notation added. 

No other changes necessary. ok

Appendix U – Pup Body Wts. Notation added 

to Day 28. No other changes necessary. Ok

Data, Draft Report, Appendix Y 

Some raw data printouts record the organ weights to three 

decimal places but Appendix Y reports the data to four 

places. Is the computer file the raw data or the computer 

printout? 

Per SOP, auto-print from the TASC data 

collection system is raw data. For this study, 

data collected on 16 and 17 Jul 02 was printed 

to 3 decimals and data collected thereafter (18 

to 29 Jul 02) was collected to 4 decimals. 

Appendix Y was re-printed to 3 decimals to 

match data from 16 and 17 Jul 02. ok

Draft Report, Appendix Y Corrected. Ok
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

15 5/4/2010 

Comments Action/Response Page 

The final body weight used for the organ weight to body 

weight ratios was the Day 21 body weight not the Day 28 

weight. These tables need to be recalculated using the Day 

28 weight. 

Draft Report, Appendix Z 

Animal number 3055 is reported as “elective sacrifice” when 

it was supposed to be change to “human sacrifice”. It was 

changed in the TASC system to “humane sacrifice”, but not 

in ZYBION. Please clarify. As well, shouldn’t a gross 

observation of “snout injury” have been recorded? 

Type of death corrected for Animal No. 3055. 

“Snout injury” was not recorded by the 

Necropsy Dept at the time of sacrifice, 

therefore this cannot be added at this time. ok

Appendix Z, Animal number 4553 

The fracture of the “head” was supposed to be changed to 

“skull” but was not. 

Corrected. Ok

Appendix AA, Released Pup and Historical Control Data 

It would be helpful to indicate when the necropsies occurred 

for the 106 pups. It may also be helpful to indicate that dams 

were in the inhalation chambers at the time of the accident. 

Lactation day of sacrifice for each litter is 

indicated on the preface. 

Notation added that the dams were in the 

exposure chamber. Ok

Appendix AA, Release Pup Data 

Is there any way of indicating on this printed out data that 

pups were at from day 25 to 28 of lactation instead of just 

indicating day 25? Also, as noted before, the historical data 

for pup body weights from study 4202 has used the Day 21 

body weights, I believe. 

We cannot make this a range of days.ok 

Terminal body weights for 4202 corrected to 

Day 28. Ok

Appendix DD: There were two Protocol Amendments, I 

believe. The second should be added. 

Protocol amendment #2 has not been signed 

as of yet and will be resubmitted to API with 

the 2nd draft report. Ok...have now? 

Protocol Amendment 2: Please verify #3 / 

second paragraph 

Author references match those in report 

(Dunlap and Duffy 1975 is listed in 

amendment versus___ Armitage 1971 is in the 

report on p.30) 

Amendment corrected to match the report 

which has been modified for the ANCOVA 

analysis. Ok

DATA ISSUES 

Data 

The animals for this study were housed in Room I-809 from 

June 17 through July 8 according to the data. The air change 

data is present for Rooms V- 708 and I-810. Air change data 

for Room I-809 should also be available in the records for 

this study or a note should be in the data to explain the 

situation. 

Please see protocol deviation dated 23 Jan 

2004 which addresses this situation. ok

Analytical Data 

The GC Chromatogram printouts for Exposures 21 through 

56 list the study number as 00-6129 instead of 00-4204. 

A study clarification note for the error was 

added to the analytical raw data. ok

Test Material receipt records 

Four, 50 cc cylinders were received on 11/6/01. These 

records are included with this study data. Are we certain 

these were filled with ETBE? Are more notations written on 

the original documents? Please verify that these are the 

correct receipt documents for this test material. 

The label for the contents of the archived 

cylinder is marked that it is G/ETBE. The 

disposition of the other 3 cylinders received 

that same day was also HLS archives for 

G/Baseline, G/ETOH and G/MTBE.ok 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

16 5/4/2010 

Comments Action/Response Page 

Test Material Usage Records 

It is indicated that large Container Number 1 was returned to 

sponsor, Chevron on 12/7/01 and Number 2 on 8/5/02. 

Please correct this to indicate exactly where these tanks were 

sent. API is the sponsor. The location of Chevron should be 

given. As well, it is indicated that 1 cylinder was sent to the 

“archives” on 12/6/01. Which archives, HLS or EPL? Please 

clarify in the data. 

Test cylinder returns were made directly to 

Universal Environmental (trucking company 

hired by Chevron) and then they delivered to 

Chevron. We do not have a receipt showing 

exactly when and where the cylinders were 

then delivered but this was recently requested 

from API and Chevron. 

The 1 cylinder that was sent to the “archives” 

on 12/6/01 was at HLS as per the data. 

EPL archives received one cylinder retain 

from Chevron_same lot as used in study 

ok 

Test Material Usage Records 

 An ending measurement of the small tanks needs to be 

taken. 

The weights were transferred from the TMAS 

sheets to complete this data. ok

Parental Organ Weight Data 

The lung data for animal number 4563 was originally 

recorded as 15.5467g, then was changed to 0, then to **, 

then to 0 again on the computer. None of these changes were 

recorded on the hand recorded edit sheet. The last change 

seen during the audit was to 0 and was done 1 month after 

the sacrifice. A value of 15.5460 g is reported, but it is kept 

from mean calculation. No explanation for this weight that is 

10 times that of other animals is given in the report in 

Appendix X. Please add an explanation to the report, 

comment on the data trial, and make certain that data are 

trackable to that reported. 

The original weight recorded was a combined 

liver and lung weight and the edit to 0.0000 

was intended to exclude the erroneous weight 

in the TASC system. It appears that the edit 

from 0.000 to ****** was made in error and 

was therefore corrected back to 0.0000. All 

organ weights were transcribed to the Xybion 

data collection system for reporting, where the 

lung weight for Animal 4563 was input as 

15.5467. During editing to exclude this 

weight from calculations, the weight was 

inadvertently changed to 15.5460. On further 

review, it was determined that, since a valid 

liver weight was obtained, the lung weight 

could be calculated from the combined weight 

and reported. An additional edit was 

performed and an explanatory memo added to 

the file. OK Where did separate liver weight 

come from? 

The reported liver weight was recorded 

directly into the TASC system at necropsy 

OK

Apparently, Animal number 2070 was “sacrificed” on the 

computer system too early. The animal was “resurrected” 

and data was re-entered according to the computer data 

correction sheets. It then appears that data for number 2070 

was removed because the wrong animal number was selected 

when data was added. The data may have belonged to 

animal 3074? A necropsy sheet was found for number 2070, 

but no print-out was found with 2070’s observations that had 

been deleted. As well, no findings for number 3074 were 

found. Please verify that the data trail is complete for these 

errors and corrections and that it is clear to a reviewer what 

happened. 

This is incorrect. According to the Computer 

Raw Data Change and Addition form, 

necropsy observation data for Animal No. 

3074 was entered as Animal No. 2070. In 

order to remove this data, Animal No. 2070 

was “resurrected”. This single edit removed 

all necropsy observations as well as the 

terminal body weight. The next entry on the 

CRDCA form was to re-enter the terminal 

body weight for Animal No. 2070. 

Necropsy data for Animal No. 2070 was 

collected manually (on a Necropsy Record 

Sheet) and entered into the Xybion system 

once the animal was “resurrected”. 
211(b) Toxicology Research Group 

ETBE One-Generation Reproduction Toxicity Study Report 

Reviewer Checklist

17 5/4/2010 

Comments Action/Response Page 

Data for Animal No. 3074 was captured 

directly into the Xybion system (albeit as 

Animal No. 2070) therefore no Necropsy 

Record Sheet exists for this animal. A printout dated 10 Jul 02 shows data for Animal No. 

2070 (actually No. 3074) collected on and 

entered on 9 Jul 02. The data was input under 

the correct animal number when 2070 was 

edited. 

Print-outs showing the corrected necropsy 

observation data for Animal Nos. 2070 and 

3074 and the audit trail “resurrecting” Animal 

No. 2070 have been added to the data. OK

Data 

The printout for the weight of food left is missing for June 

20, 2002. 

Food left weights for this date are located in 

data records Volume II Book 5. ok