March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

1

COMMENTS Action / Response Page 

#

Dr. Richard Schlesinger

1 I have reviewed the report and found that the original 

goals of the study have been met. The overall 

conclusions are supported by the reported results. 

Agree

2 There is no statistical evaluation in the Report. 

However, the investigators do compare various 

values between genders and at different times. It 

would be best if there was some level of statistical 

significance associated with their value judgments.

Since this is a pilot study, to be 

repeated, we had avoided statistical 

comparisons. 

3 Section 3.4. What was the time frame for sampling 

DIPE at the tower inlet and outlet, i.e., continuous, 

discontinuous, etc.?

Continuous. Added “The 

concentration of DIPE was monitored 

continuously at the inlet to the tower, 

and at the outlet.” 

5

4 In Section 3.5, it is noted that for Study A, a total of 9 

male and 9 female rats were exposed to labeled 

DIPE. However, the Protocol noted in Section 2.0 

indicates that Study A involved use of non-labeled

DIPE and exposures were to be conducted on only 6 

rats. This needs to be clarified.

This has been corrected. Rats were 

exposed to unlabeled DIPE in study 

A, with 8 animals exposed, and 

samples from 6 analyzed. Changed 

9 to 8 in the text

5

5 Section 3.5.1. It is stated that the port to port 

variability was “within accepted limits.” Please define 

in the report body what these limits actually are.

Added criteria for acceptable 

performance.

6

6 Section 3.6.1. The blood sampling times in the first 

paragraph are unclear. It appears that 3 rats had 

blood drawn from 5 min to 6 hr after exposure began 

and another 3 had blood drawn at 14 min to 18 hr 

after the exposures ended. What happened to the 

other 3 rats in each gender (see #2 above)?

In 4 above, the numbers have been 

clarified. Additional samples were 

collected to cover the possibility of 

loss of an animal. However, the 

duration of sample stability did not 

permit analysis of all of the samples 

collected. 

6

7 Section 4.2 should be moved to the Methods section 

of the Report.

We see this as part of the data 

generated, and is appropriate for 

inclusion in the results. No change.

8 Section 4.3 and Table 4. Why is the exhaust 

concentration for Study C half that of the inlet for this 

protocol only, compared to A or B.

The flow rate of the tower was 

reduced because of the limited 

amount of 13C label available. 

9 Section 4.4. The concentrations are in Figures 1-4, 

not just 1-3 as noted in the text. Furthermore, the first 

paragraph on pages 10-11 seems to have sentences 

that are repeated. Finally, why did the concentration 

of acetone exceed the calibration curve range; i.e., 

why did the investigators not anticipate this potential 

and adjust the range accordingly.

Corrected. Removed duplication. 

Acetone concentrations exceeded 

our expectation, and we did not have 

any previous data to base the 

expected concentration range on.

11
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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10 Section 4.4. In the second paragraph, Figure 4 should 

be Figure 5, and Figure 5 should read Figure 6.

Corrected 12

11 Section 4.5. How was dose estimated? It is based on the total amount of 

radioactivity recovered, divided by 

the specific activity. Added a 

footnote to Table 9 to clarify.

25

12 Section 5.0. The investigators noted that comparison 

of DIPE levels in rats immediately after exposure with 

levels in rats after 7 days of collection of excreta 

indicated that there were no significant losses of 

radioactivity from the sample types during processing. 

However, looking at the data there seems to be about 

a 20% loss between input and output, and to this 

reviewer that percentage can be perhaps considered 

as significant.

See discussion below for comment 

#7 from Dr. James Bond. We have 

1) corrected a calculation error and 2) 

corrected the dose calculated for the 

specific inhaled dose, and added to 

the discussion. When corrected for 

the exposure concentration and 

duration a similar recovery per 

ppm.hr was found with Study B (3529 

ppm x 6 hr = 21174 ppm.hr, 372.99 

mg/21174 ppm.hr = 0.0173 

mg/ppm.hr) and Study C (3334 ppm 

x 5.66 hr = 18870 ppm.hr, 317.89 

mg/18870 ppm.hr = 0.16846 

mg/ppm.hr). Since the recovery of 

Study C was 95.6% of Study B when 

corrected for exposure concentration 

and duration, we think that this 

recovery indicates no significant 

losses occurred

14

Dr. James Bond

1 In general, the report is well-written and thorough and 

the stated purpose of this study has been met.

Thank you.

2 Page 2 of 337, Summary: There are several places in 

the Summary where there are typographical errors or 

missing text. These are noted as comments on the 

Draft Final Report.

Corrected. 2
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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3 Page 20 of 337, Section 4.4: The final report notes 

that regarding DIPE concentrations in blood “There 

was little difference in blood levels at peak 

concentration between males and females”. However, 

in Table 5 it appears that mean blood levels of DIPE 

in female rats appear to be consistently higher than 

males throughout the exposure period. The errors 

associated with these mean values for both males 

and females may make this difference not statistically 

significant. Has a statistical analysis been conducted 

to demonstrate that the blood levels are/are not 

different?

No statistical analysis was 

conducted, since this was a pilot 

study, and no statistical analysis was 

indicated in the agreed protocol.

4 Page 21 of 337, Section 4.4: The report notes that 

“Acetone levels in both male and female rats rose 

steadily during the exposure, and in males continued 

to rise following exposure until 8 hr following the 

initiation of exposure.” However, if one takes into 

account the errors associated with the mean 

concentrations of acetone in blood of male rats 

between 6 and 8 hours after the start of exposure 

(see relevant portion of Table 7 reproduced below), it 

appears that there is no difference among these 

values. These values appear to remain constant over 

this time period (i.e., 6 to 8 hours), a result similar to 

that found for females.

Male Rats

Time (hr) Acetone μg/mL Mean SD

6 288.42 76.855

6.25 298.9 19.328

6.5 319.38 15.137

7 312.47 22.028

7.33 315.15 19.22

8 301.87 16.539

The text has been changed to 

indictate that the behavior of the 

males and females was similar, with 

concentrations constant between 6 

and 8 hours 
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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5 Page 21 of 337, Section 4.4: The pharmacokinetic 

analysis of the decline in blood concentrations of 

DIPE, isopropanol, and acetone after the end of 

exposure seems problematic. It is not clear why the 

same time points are not used for analysis for male 

and female rats for DIPE, isopropanol, and acetone. 

For example, for acetone, time points from 7 to 24 

hours post exposure are used for males, but time 

points from 6.25 to 24 hours post exposure are used 

for females, with no explanation as to why these 

earlier time points were not used for males also. 

Additionally, it is probably not appropriate to include 

data on acetone concentration in blood from the 16 

and 24 hour time points for females since these 

values are at background acetone levels. Inclusion of 

these values tends to increase the half-time by 

artificially elevating the values for acetone derived 

from DIPE. The fit of the predicted exponential 

equation to the data for blood acetone concentration 

in females in Figure 7 of the report shows that there is 

not a good fit to these later time points. Background 

should be subtracted from all values for acetone in 

order to obtain estimates of the DIPE-derived acetone

which is critical at the lower acetone blood 

concentrations. Since a radiolabel was not used in 

Study A it is not possible to differentiate endogenous 

acetone from DIPE-derived acetone.

For each analysis, WinNonlin was 

used for non-compartmental analysis, 

and the analysis was conducted with 

the software choosing the best fit to 

the data. This avoids the perception 

of the operator choosing the fit that 

they like best, and generating 

operator bias (cherry picking). 

However, the data points used by the 

software may be different for each 

animal. 

6 Page 22 of 337, Section 4.6: The report notes that 

“The total recovery of radioactivity from all of the 

samples collected from rats in Group 3, Study C is 

presented in Table 9, and ranged from approximately 

25 to 28 μCi.” How was this range determined? Table 

9 shows a value for CM-04 that is 6.898. If this rat is 

included in calculations then the mean value and 

range in Table 9 would also change. If this animal 

was not included in the calculations it should be 

specifically noted.

There is a footnote in the table noting 

that rat CM-04 was not included in 

the total analysis of samples, with the 

exhaled VOC traps reserved for 

possible later analysis. A note has 

been added to the text.
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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7 Page 23 of 337, Discussion: The report notes the 

following “Comparison of the exposure of rats to 3600 

ppm [2- 14C]DIPE/DIPE euthanized immediately

following exposure (372 ± 8 mg/kg) with that of rats 

euthanized following 7 days of collection of excreta 

and exhaled breath (299 ± 7 mg/kg) yielded values 

that were very similar. This suggested that there were 

no significant losses of radioactivity from the various 

sample types during processing of radioactivity.”

However, the value of 299 is about 20% less than the 

value of 372 [calculated as follows: (372-

299/372)*100]. This suggests that 20% of the 

radioactivity is unaccounted for when all routes of 

elimination and the material remaining in the carcass 

are quantitated. On the surface these data seem to 

suggest that about 20% of the radioactivity was lost 

during processing. However, there may be other 

factors that would result in a lower dose in the Group 

C rats. Since the Group B and Group C rats were not 

exposed to DIPE at the same time, differences in the 

exposures may explain part of the difference in the 

body burdens of the two groups. The report should 

address the impact that a shorter exposure duration 

and a lower mean DIPE exposure concentration for 

Study C would have on the dose of DIPE that was 

absorbed by these rats compared with rats in Group 

B. These two factors can be, and should be, 

quantitated.

We did discover an error in the 

calculation of recovery for Group C, 

which increased the dose for each 

animal. The effects of the exposure 

duration and exposure concentration

on the dose have been calculated. 

Group B had an exposure duration of 

6 hr, with a mean concentration of 

3529 ppm, giving 21174 ppm.hr of 

exposure. Group C had an exposure

duration of 5.66 hr, with a mean 

concentration of 3334 ppm, giving 

18870 ppm.hr of exposure. 

Comparing the total radioactivity 

recovered normalized by exposure 

yields 0.0176 mg/ppm.h for Group B, 

and 0.0168 mg/ppm.h for Group C. 

The normalized values for Group C is 

approximately 95.6% of that obtained 

for Group B. This has been added to 

the Conclusion.
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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8 Page 209 of 337, Appendix E, Pre-Study Inhalation 

Report, Table 3: This study shows that the distribution 

of DIPE throughout the inhalation chamber is uniform. 

However, this table does not show that the 

concentration AT THE BREATHING ZONE of the rats 

is equivalent to the target concentration or to the 

DIPE concentration measured at the inlet. 

Demonstration that the concentration of the exposure 

atmosphere at the port is equivalent to the 

concentration at the inlet to the chamber is critical 

since it is the inlet concentration that is reported as 

the chamber concentration. One cannot assume that 

the concentration at the breathing zone of the rat is 

equal to the concentration of DIPE at the inlet to the 

chamber. The fact that these concentrations are 

about 48% of the target concentration of 3600 is 

problematic. The report notes that 200 ml/min of test 

atmosphere was sampled from each port and was 

diluted with 200 ml/min of air before the diluted test 

atmosphere entered the MIRAN cell. This would 

suggest that the concentration of the test atmosphere 

at the port was twice the concentration listed in Table 

3 unless some adjustment, which is not described in 

the report, was used to account for this

dilution. The quantitative relationship between the port 

concentrations and the inlet concentration, and the 

consequences of the port/inlet concentrations not

being equal should be addressed.

The reviewer correctly notes below 

that the reduction in concentrations 

measured and concentrations at the 

inlet result from additional air added 

to make up the flow rate into the 

Miran detector from the ports. The 

port measurements were made to 

establish port to port variability and 

were diluted to enable more rapid 

sampling at a number of ports. This 

has been clarified in the final report. 

The sampling was conducted to show 

port to port variability, and 

9. Page 210 of 337, Appendix E, Pre-Study Inhalation 

Report, Table 4: The daily mean concentration for the 

Inlet in Table 4 suggests that the concentration at the 

breathing zone (1720 ppm for the 17 open ports taken 

from Table 3) is from 56 to 50% of the inlet 

concentration depending on when that concentration 

was measured (averaged over the whole exposure or 

during the last 2.5 hours). This comparison raises 

several important issues. First, why is the 

concentration at the breathing zone so much lower 

than the inlet concentration? Second, why is the 

concentration measured at the Inlet so much higher 

during the last 2.5 or 3.5 hours of the exposure? 

Third, why is the exhaust concentration higher than 

the inlet concentration? These are important issues, 

critical to the demonstration that the inhalation 

exposure system can deliver the target concentration 

to the breathing zone of the rats, and should be 

addressed.

As noted in 8 above, the 

concentrations were determined after 

dilution with an equal volume of air, 

as noted in the report. The 

concentration is not lower in the 

breathing zone. The concentration 

measured at the inlet was obtained 

by passing 200 mL/min into the inlet 

Miran, whereas the concentration at 

the outlet was measured with the 

entire effluent from the tower. This 

was modified during the definitive 

studies so that the entire flow onto 

the tower passed through the flow 

cell. 
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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10 Page 21 of 337, Appendix E, Pre-Study Inhalation 

Report, Study A, Table 5a: The data in this table 

suggests that it took 4.4 hours for the exposure 

system to reach a stable concentration. This is 

problematic if this inhalation methodology is used in 

the animal studies because in Study A, for example, 

blood samples are being taken from rats starting at 5 

minutes into the exposure. Table 5a would then 

suggest that in Study A the lower blood levels of the 

rats during the early times in the exposure could be a 

combination of short exposure duration and lower 

exposure concentration. It would be preferable to start 

the exposure of the animals (i.e., plug the tubes 

holding the animals into the ports) AFTER the 

chamber had reach a constant concentration.

If you examine the data in Table 4 for 

Study A, the inlet concentration was 

approximately 99% of the maximum 

concentration by 75 minutes. The 

inlet sampling was modified for the 

later definitive studies.

11 Page 231 of 337, Appendix F, Inhalation Reports, 

Study A, Table 4: The data in this table is problematic 

for a number of reasons. First, it does not appear that 

DIPE concentrations reached steady-state until 105 

minutes into the exposure. Does this mean that the 

rats were initially exposed to a lower concentration of 

DIPE for the first 100 minutes? This is problematic 

because blood samples were taken from the rats as 

soon as 5 minutes into the exposure where they 

would be exposed to concentrations of DIPE that 

were only 82% of the concentrations from 105

minutes until 405 minutes. However, the exhaust 

concentration appears to be stable from the first 

sampling point (15 min) until the end of the exposure, 

suggesting that it is the sampling methodology, not 

the generation of the exposure atmosphere that is an 

issue. It appears that the problem is the time that it 

takes to equilibrate the MIRAN (>60 min) that is 

analyzing the air sample taken from the inlet. Time to 

equilibrate is not a problem for analysis of the exhaust 

since 100% of the exhaust enters the MIRAN that is 

analyzing that concentration. It is likely that the “real” 

exposure concentration in this study is approximately 

3700 ppm, and the exposure atmosphere samples 

taken up to and including 75 minutes are not 

representative of the chamber concentration, but 

rather representative of the time it takes to equilibrate 

the MIRAN. Inclusion of these low values in the 

calculation of the mean chamber concentration is 

misleading an artificially lowers the average for the 

exposure concentration.

If you examine the data in Table 4 for 

Study A, the inlet concentration was 

approximately 99% of the maximum 

concentration by 75 minutes. The 

inlet sampling was modified for the 

later definitive studies.
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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12 Page 247 of 337, Appendix E, Pre-Study Inhalation 

Report, Study B, Table 4: The data for the exposure 

concentration at the inlet indicate that for Study B the 

exposure concentrations had reached steady-state by 

the first sample time, 10 minutes into the exposure. 

However, the same methodology for generating and 

sampling the exposure atmosphere that was used in 

Study B was used in Study A where it took 

approximately 100 minutes to reach steady-state.

In fact, the exact same Miran IR spectrometer (Serial 

number 4121) was used in both studies. Thus, with 

the same methodology and sampling system, it is not 

clear how the laboratory was able to generate 

analytical data to suggest that steady-state 

concentrations were achieved in 10 minutes in one 

study, but not until 100 minutes in the other study. 

Some explanation as to the differences between the 

two exposures in sampling or analysis is required.

There were no differences in the 

exposure sampling or analysis 

between Study A and Study B. In 

Study B, the raw data indicates that 

the quantity of DIPE displaced once 

the syringe was loaded was greater 

than 0.18 mL. This caused the 

concentration on the nose only tower 

to go higher than the target 

concentration of 3600 ppm. The 

syringe pump was turned off and the 

excess test material was allowed to 

evaporate until the concentration 

value at the inlet Miran and the outlet 

Miran were within 10% of the 

exposure target concentration. The 

differences in the sample times 

between Study A and Study B can be 

attributed to technician error.
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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13 Page 263 of 337, Appendix E, Pre-Study Inhalation 

Report, Study C, Table 4: Similar to Study B the data 

for exposure concentration at the inlet indicate that for 

Study C high concentrations of exposure atmosphere 

were attained by the first sampling point, in this case 

20 minutes. In fact, this 20 minute sample is the 

highest concentration of the exposure atmosphere 

recorded over the entire 335 minutes of sampling. 

Again, the same methodology for generating and 

sampling the exposure atmosphere was used in 

Study C as was used in Study A and again the exact 

same Miran IR spectrometer (Serial number 4121) 

was used. Thus, with the same methodology and 

sampling system, it is not clear how the laboratory 

was able to generate analytical data to suggest that

the maximum exposure concentration was achieved 

by 20 minutes into the exposure. Some explanation 

as to the differences between Study A and Study C in 

sampling or analysis is required.

There were no differences in the 

exposure sampling or analysis 

between Study A and Study C. At 

the time the first animal was loaded 

the inlet concentration was within 

20% of the exposure target 

concentration of 3600 ppm and the 

raw data indicates the concentration 

value measure by the inlet Miran was 

still increasing and had not reached a 

steady-state. (NOTE: The amount of 

time needed to fill the inlet Miran cell 

is 29 minutes – the system had been 

running for 35 minutes when the first 

animal was loaded.) In Study C, there 

was a limited amount of 13C6 test 

material available. Calculations on 

the exposure day indicated that the 

amount of test material remaining in 

the syringe had reached a critical 

point and the decision was made to 

start loading animals onto the nose 

only tower, otherwise the entire 

exposure would compromized. Study 

records indicate the actual exposure 

length was ca. five hours and forty 

minutes. A Protocol Deviation Report 

documents the Deviation, Reason, 

Study Director Recommendations 

and indicates that the Sponsor was 

contacted.

QA Comments

1 Summary: Second paragraph ends in mid-sentence. Missing text has been added 1

2 Summary: Last paragraph says “...for inhalation 

exposure to DIPE and rr achieved.” Please correct.

Corrected 2

3 Need to complete QA statement Will complete when finished Iii

4 Need to add API signature block for GLP statement. Iv

5 3.1.2 Cannot find supporting documentation for the 

purities reported here.

Added 2
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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6 3.3 Diet. Protocol indicates that waters samples 

should have also been checked for the presence of 

TBA. Where are the results of the water sample 

tests? 

All were below the reported limit of 

detection and text has been added:

All of the analytes were below the 

reporting limit for the method in the 

samples obtained before and after 

the study (tertiary butanol 1 μg/L, 

tertiary amyl methyl ether 0.1 μg/L, 

ethyl tertiary butyl ether 0.1 μg/L, 

diisopropyl ether 0.1 μg/L, and methyl 

tertiary butyl ether 0.1 μg/L).

3

7 3.4 Where is the stability reported? It has been added 5

8 4.1 99.71 ± 0.01% Where is this purity reported? 

Cannot verify.

Reported in revised Appendix B, 

Table 1. 

9

9 4.1 Cannot find results of NMR spectroscopy 

analysis.

Added Appendix containing the data 10

10 4.3 Appendix F indicates the name of the report is, 

Inhalation Summary Report:DIPE, 14C-DIPE/DIPE, 

and 14C-DIPE/13C-DIPE.

Changed 10

11 4.4 Please indicate that “In the male rats, blood 

concentrations of isopropanol were below LOQ...”

Added 10

12 4.4 top of page 11. Please indicate that “In the 

female rats, isopropanol concentrations were below 

LOQ....”

Added 11

13 4.4 last paragraph. Please recheck all the Figures 

indicated. It appears that they off by 1 (i.e. 5 should 

be 6)

Changed 11

14 4.5 “... approximately 33 to 35 uCi.” Changed 12

15 4.6 3rd paragraph. “...at 8 and 24 hours, respectively. 

Less than or equal to 0.2% of the radioactivity...”

Changed 12

16 5.0 “...3600 ppm DIPE.” Changed 13

17 5.0 3rd paragraph. Please recheck the percent 

14CO2 recovery and the percent in the carcass 

recovered. 

Corrected. 20.2 % for CO2, and 

2.6% for carcass

14

18 5.0 Last paragraph. “... 10 hours after initiation of 

exposure.” “... radioactivity in the exhaled breath and

as 14CO2.”

Added “initiation of” exposure. 

Actually meant both the exhaled 

breath volatiles and CO2. Added 

clarification.

14

19 b Animals used in Group 3 and Group 4 ......” Deleted 17

20 Table 2. add footnote from Table 3 Added. 18

21 b Two animals with measurements less that the limit of 

quantitation. 

Changed 23

22 Table 10. The results for the second two animals in 

Group 2 Study B/Exp. Urine Sample 2 should be N/A

Added NS. 26
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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23 Table 11. Is N.S. not sampled? Where are the 

results of the polyethylene bag that held the animals 

during transfer? 

N.S. is no sample. The animals were 

transferred inside the nose only tube 

into the Tedlar bag for euthanasia. 

Any radioactivity that was in the nose 

only tube or spilled out into the Tedlar 

bag was included in the exposure 

urine, exposure feces, or nose only 

tube rinse.

26

24 Table 18. Where does data from “0” time point come 

from?

This comes from the materials 

collected as the animals were placed 

in metabolism cages.

33

25 The protocol indicates that the exposure atmosphere 

generation design was to be developed at CIIT.

It further indicates that the design technology will be 

reproduced at RTI and that verification studies will be 

conducted without animals to assure that exposure 

atmospheres can be produced in the setup at RTI. It 

implies that the distribution studies will be done when 

the verification studies are done, at RTI. The 

distribution study; however, was performed at CIIT 

during the development and testing of the system.

Work performed at CIIT is indicated to not be 

incompliance with GLP regulations. The distribution 

study is a GLP requirement and should have been 

conducted in compliance with GLPs. This will need to 

be indicated in the GLP Compliance Statement of 

both the main study and the Inhalation report. 

This will be indicated in the 

compliance report for the main study.

26 Appendix E and F are signed as final. If any changes 

are made to these, they should be done as 

amendments to the report.

Noted.

27 Table 9 indicates that the data reported does not 

include exhaled breath traps, which were saved for 

other possible analysis, yet in the raw data there 

appear to be data for exhaled CO2 traps, urine, feces, 

and carcass. Should these values be reported? 

Traps from one animal were 

reserved. The other samples were 

analyzed for that animal. They were 

included in the calculation in Table 9, 

but excluded from the group mean 

and SD. The data for recovery

expressed as a percentage of the 

total recovered would give an 

inappropriate comparison with the 

other animals, so the data has not 

been included in Tables 11 – 18. 
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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28 Table 18: This table is to include recovery of 

radioactivity in excreta and carcass, yet there is a 0 

hour timepoint. Wasn't only blood was taken at 0 

hours? Where could this data come from? 

Table 18 includes the samples 

collected at each timepoint. 0 h

included the nose only tube wash, 

and any urine or feces collected prior 

to the animals being placed in the 

metabolism cage. 

29 Appendix E, Table 3: What are the units of measure 

for the reported values? Since the test atmosphere 

was further diluted to reduce sampling time, 3600 

ppm wasn't really the Target concentration. 

The variability was what was being 

measured here. 

30 Appendix B, Page B-3, 5th paragraph: Is the word 

'signal' correct, or is 'singlet' correct?

They are both correct. However 

“singal” has been corrected. 

31 Appendix B, Page B-5, Peroxide Check: Last 

sentence might be better as 'A white color indicated 

the peroxide value was 0 mg/L, through the storage 

period.'

No change

32 Appendix B, Page B-5, Conclusion, last sentence: .... 

FID was 99.63 %....

Corrected.

33 Appendix B, Page B-11: Was a stability test done 

after the study? If so, it should be included here.

Yes. It has been added.

34 Appendix C, Page C-4, Conclusion: The purity of the 

test substance was 97.23% (?) by HPLC... 

This has been corrected

35 Appendix F, Study A, page 10 of 16 (F-11), Nose 

Only Exposure System Distribution: The distribution 

study was performed at CIIT, not RTI.

The title of the report in Appendix E is 

as shown. 

36 The same is true for Study B, page 10 of 16 (F-27), 

and Study C, page 10 of 16 (F-43). 

The title of the report in Appendix E is 

as shown. 

37 Appendix F, Studies B and C, Generation and 

Chamber Concentration, Pages 10 of 16 (F-27 and F43): In the second paragraphs of the 'Generation and 

Chamber Concentration' sections indicate the actual 

DIPE that was used. For Study B, it was 14CDIPE/DIPE and for Study C, 14C-DIPE/13C-DIPE.

The test chemicals used is clearly 

indicated throughout the text with this 

one exception. No change. 

38 Appendix F: Throughout this appendix it is indicated 

that the stability of the test atmosphere was checked 

twice during the exposure by RTI (Page 7 of 16 

in Study A, B, and C reports). Where are 

these analyses reported? 

These have been included
March 24, 2013 Draft Report - METABOLISM AND PHARMACOKINETICS OF DIISOPROPYL ETHER IN 

MALE AND FEMALE RATS: PILOT STUDY

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39 Appendix F: For Study C, the exhaust concentrations 

appear to be about half of the inlet concentrations. In 

the preliminary work done with 7 ports open and all of 

the other studies, the inlet and exhaust concentrations 

appear more similar. Any reason for this difference 

in Study C? Was the exhaust Miran sample further

diluted for some 

reason? 

 

In the preliminary work, there are no 

animals to take up the DIPE. This is 

a reflection of the uptake of DIPE. 

40 Appendix G: Page numbers for this appendix appear 

as H-#s. 

Corrected

41 Appendix H, Raw Data Tables, Table 7: The header 

on these tables indicates ETBE not DIPE. Please 

verify that the data reported are actually from the 

DIPE study.

Corrected

42 Appendix H, Raw Data Tables, Table 2, Blood 

Sample weights for Animal AM-06: Data for this 

animal appears twice. Could there be a mix-up or just 

clerical error? 

Corrected

43 Appendix J: The note on the cover page indicates 

that this appendix will be updated with 'final version'. 

What will change?

The analysis was redone with a 

validated version of WinNonlin

following completion of review.