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Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1981
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: see 'Remark'
Remarks:
Protocol, study conduct and report were audited (study was conducted prior to the enactment of FDA Good Laboratory Practice Regulations), the study was not conducted according to guideline/s and GLP but the report contains sufficient data for interpretation of study results
Cross-referenceopen allclose all
Reason / purpose:
reference to same study
Reason / purpose:
reference to other study

Data source

Referenceopen allclose all

Reference Type:
publication
Title:
Pharmacokinetics of Inhaled 14C-2-Nitropropane in Male Sprague-Dawley Rats
Author:
Nolan, R.J., Unger, S.M., and Muller, C.J.
Year:
1982
Bibliographic source:
Ecotoxicology and Environmental Safety, 6, 388-397
Reference Type:
study report
Title:
Unnamed
Year:
1981

Materials and methods

Objective of study:
other: The purpose of this study was to provide data on the fate of 2-NP in male sprague-Dawley rats that will aid in interpreting the animal toxicity data and in evaluating hazard.
Test guideline
Qualifier:
no guideline followed
Principles of method if other than guideline:
Toxicokinetic study
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
The test material (2-nitropropane- 1,3 14C Lot No. 863-260) was obtained from New England Nuclear Corp., Boston, Mass. with a specific activity of 4.10 mCi/mmol and a radiochemical purity of 99% as determined by GC.

The unlabeled 2-NP (Lot No. 12577) was obtained from International Minerals and Chemical Corp., Terre Haute, Indiana with a purity >99%.
Radiolabelling:
yes

Test animals

Species:
rat
Strain:
Sprague-Dawley
Sex:
male
Details on test animals and environmental conditions:
Male SD rats were obtained from a commercial supplier, and individually identified by metal ear tag. They were housed in controlled rooms with conditions adequate for the species. They were given a commercial diet and water ad libitum except during inhalation exposures. A minimum of 48 hours prior to exposure, jugular cannulas were implanted.

Administration / exposure

Route of administration:
other: inhalation and intravenous
Vehicle:
unchanged (no vehicle)
Details on exposure:
Inhalation exposures were conducted in 30L whole-body chambers with dynamic airflow conditions. The atmosphere was generated by metering the 2-NP into the airstream. Chamber atmosphere was analyzed every 30 minutes by GC. Rats were exposed for 5.5 hours to 154 ppm 2-NP. Two 6-hour exposures were then conducted at 20ppm (4 rats to determine blood and plasma concentration-time profiles, and 6 other rats to define the excretion and tissue distribution of the radioactivity.

Intravenous dose solutions were prepared by suspending 14C-2-NP and unlabeled 2-NP in fresh heparinized rat plasma or dissolved in Tween 80 and then suspended in 0.9% saline. Each animal received ~5uCi of radioactivity in a dose volume of 3 ml/kg administered via the jugular cannulas. Doses ranged from 13 to 148 mg 14C-2-NP/kg.
Duration and frequency of treatment / exposure:
Inhalation
5.5 hours at 154 ppm (0.59 ug/ml) of 14C-2-NP with a specific activity of 344 dpm/ug
Two 6-hour exposures were conducted at a targeted concentration of 20 ppm

Intravenous
Single administration of 13-148 mg/kg
Doses / concentrations
Remarks:
Doses / Concentrations:
Inhalation
154 ppm
20 ppm: blood 2-NP and plasma 14C-concentration time-profiles and a second 20ppm group for excretion and tissue distribution

Intravenous
13-148 mg/kg
No. of animals per sex per dose:
9 male rats/154 ppm
4 male rats/21 ppm and 6 male rats/23 ppm- future discussions will refer to these as 20 ppm exposures
2 male rats/13.3, 15.4, 103 and 148 mg/kg i.v.
Control animals:
no
Positive control:
None
Details on study design:
The purpose of this study was to provide data on the fate of 2-NP in male Sprague-Dawley rats that will aid in interpreting the animal toxicity data and in evaluating hazard. Rats were given single intravenous doses ranging from 13 to 148 mg 14C-2-NP/kg, or exposed for 6 hr by inhalation to ~20 or 154 ppm 14C-2-NP, and the disposition of radioactivity followed for 48 hours. Blood samples were collected via a jugular cannula and analyzed for 2-NP. Plasma samples were also collected via a jugular cannula and analyzed for 14C. Expired 14C (collected in toluene/methanol to trap expired 2-NP and other volatile organics and ethanolamine:2-methoxyethanol to collect CO2), urine, feces, and tissues were collected at selected intervals and analyzed for radioactivity. The amount of radioactivity incorporated into hepatic macromolecules was determined by Jollow et al. (1973). Selected urine specimens were subjected to ion exchange chromatography.
Details on dosing and sampling:
See above "details on exposure" for dosing.

Blood samples were collected via the jugular cannula while patent, and through the orbital sinus or tail vein thereafter. Blood was analyzed for 2-NP and plasma was separated from blood cells and analyzed for radioactivity.

Exhaled air was trapped by pulling room air through a series of absorbant tubes (collected in toluene/methanol to trap expired 2-NP and other volatile organics and ethanolamine:2-methoxyethanol to collect CO2) and analyzed. Urine was collected at 6, 12, 24, 36 and 48 hour intervals and analyzed for radioacitivity. Portions of the urine specimens collected at 0-6 hr following the 15.4 and 103 mg/kg i.v. doses and inhalation exposures were subjected to ion exchange chromatography. Feces were collected at 24-hour intervals and also analyzed for radioactivity.

Animals were sacrificed, and the liver, kidney, lung, brain, heart, skeletal muscle, fat, and skin, and carcass were oxidized and analyzed by liquid scintillation counting for remaining radioactivity.

The amount of radioactivity incorporated into hepatic macromolecules was determined by Jollow et al. (1973).

Time-weighted average exposure concentrations were calculated. Half-lives were estimated by least-squares analysis.

Statistics:
Where appropriate, the groups were compared using a two-tailed Students' t-test with P<0.05 selected for the criterion of statistical significance.

Results and discussion

Preliminary studies:
None

Toxicokinetic / pharmacokinetic studies

Details on absorption:
Data indicate that 40% of the inhaled dose was absorbed.
Details on distribution in tissues:
The relative distribution of 2-NP in the tissues and the tissue/blood ratios were similar regardless of the dose delivered. The excretory organs (liver, kidney and lungs) generally contained the highest concentrations of radioactivity. During the 48 hr following the two inhalation exposures a 2 - 5 fold decrease in tissue radioactivity occurred and could account for the slow terminal clearance phase observed for plasma radioactivity. The lowest tissue/plasma 14C-ratios (0.2-1.8) were observed immediately following the two inhalation exposures, and the largest tissue/plasma 14C ratios (1.8 - 8.4) were found in rats sacrificed 48 hr after the 103 mg/kg i.v. dose.

Only a small fraction of the radioactivity in the liver (
Differences in the blood 2-NP concentration time profiles observed following i.v. administration of 13.3 and 148 mg 14C-2-NP/kg dissolved in Tween 80~3, and 37.6 and 104 mg 14C-2-NP/kg suspended in rat plasma during the first 20 to 60 min after dosing were clearly related to the vehicle used to prepare the dosing solution. In contrast the change in the half lives for the apparent first-order clearance of 2-NP from the blood, starting about 1 hr after dosing, were not related to the vehicle. The half lives for the clearance of 2-NP from the blood following the 13.3, 37.6, 104 and
148 mg/kg i.v. doses were 25, 29, 39 and 70 min, respectively. This consistent increase in half lives with increasing dose is a classical indicator of dose-dependent or non-linear kinetics.

Following the 20 ppm exposure, blood 2-NP concentrations were close to the analytical detection limit and extremely variable and the half-life for the clearance of 2-NP from the blood could not be reliably estimated. Individual data for the 3 rats exposed to 154 pprn were plotted, and these data suggest that 2-NP was cleared from the blood in a first-order manner (t 1/2 = 48 min). The concentrations of 2-NP in the blood immediately following
the 20 and 154 pprn were not proportional to the exposure concentrations. The 7 fold difference in exposure concentrations resulted in a 12 fold difference in blood 2-NP concentrations (0.6 versus 7.1 ug/ml).

The concentration of radioactivity in the plasma following the 37.6 and 104 mg/kg i.v..doses was initially proportional to the dose and decreased in a biphasic manner. During the first 12 hr after dosing the radioactivity was cleared from the plasma faster following the 37.6 mg/kg dose (t 1/2= 98 min) than after the 104 mg/kg dose (t 1/2 = 156 min) . During the 24 to 48 hr post-dosing interval radioactivity was cleared from the plasma with a half life of 36 hr following both i.v. doses.

Differences were observed in both the concentration and clearance of radioactivity from the plasma following the two inhalation exposures. The average concentrations of radioactivity in the plasma immediately following the 20 and 154 ppm exposures were 3.6 and 33.3 ug eq 14C-2-NP/g. Thus the 7 fold difference in exposure concentrations resulted in a 9 fold difference in plasma radioactivity. The radioactivity was cleared from the plasma faster during the first 12 hr after the 154 ppm exposure (t 1/2 = 172 min) than after the 20 ppm exposure (t 1/2 = 354 min) . In contrast, the clearance of radioactivity from the plasma 12 to 48 hr after the 20 and 154 ppm exposure was similar (t 1/2 = 35 and 36 hr, respectively).
Details on excretion:
The major route of excretion was the lungs (55% of the radioactivity was excreted as 14CO2 and 14C-2-NP). Rats given 103 mg/kg i.v. dose excreted a larger percentage of the radioactivity as 14C-2-NP (35.4 versus 13.4%) and a smaller percentage as 14CO2 (45.1 versus 59.1%) than those given 15.4 mg/kg i.v. dose. A larger percentage of the radioactivity recovered following the 154 ppm inhalation exposure was excreted exhaled as unchanged 14C-2-NP (21.7 versus 3.7%) although following both the 20 and 154 ppm exposures about 51% of the recovered radioactivity was excreted as 14CO2. Urine and feces represented minor routes of excretion. Following both i.v. dose and inhalation exposure concentrations about 10% of the recovered radioactivity was excreted in the urine. At 103 mg/kg i.v. and 154 ppm exposure concentrations there was a trend toward decreased fecal excretion of radioactivity, and a smaller percentage of the radioactivity was found in the tissues. The total amount of radioactivity recovered following the two i.v. doses was proportional to the dose and represented 130% of that administered. Nearly identical amounts of radioactivity were recovered from rats sacrificed 0 and 48 hours after the 20 ppm exposure. In contrast 2.4 times more radioactivity was recovered from rats sacrificed 48 hr after the 154 ppm exposure than was recovered from animals sacrificed immediately following this exposure. This difference is believed to be due to the loss of volatile radioactivity (i.e. unchanged 14C-2-NP) during preparation of the tissues for analysis and is consistent with the difference in amount of unchanged 14C-2-NP excreted via the lungs following the 20 and 154 ppm.

Pulmonary 14C-2-NP Elimination : All radioactivity trapped by the to1uene:methanol was considered to represent unchanged 14C- 2-NP. The amount of unchanged 14C-2-NP trapped during the initial 0.5 hr collection interval was proportional to the i.v. dose. Pulmonary 14C-2-NP elimination was biphasic following both i.v. dose. However, after the 15.4 mg/kg dose the rapid initial elimination phase lasted about 7 hr and had a half life of 46 min. In contrast, after the 103 mg/kg dose the rapid initial elimination phase lasted 24 hr and had a half life of 103 min. During the second 24 hr post-dosing interval, pulmonary 14C-2-NP elimination appeared to be faster after the 103 mg/kg dose than after the 15.4 mg/kg dose (t 1/2 = 7.5 versus 8 .7 hr) ; however, less than 0.2% of the 14C-2-NP recovered was excreted during this interval.
Over 75% of the radioactivity in the urine was excreted during the first 12 hr after the two i.v. doses and 20 ppm exposure.

Pulmonary elimination of 14C-2-NP following the 20 and 154 ppm exposures was biphasic and exhibited the same exposure dependent characteristics as the blood 2-NP concentrations. Fifty times more 14C-2-NP was excreted via the lungs during the first hour after the 154 ppm exposure than after the 20 ppm exposure (985 versus 17 ug eq 14C-2-NP) . The rapid initial phase for pulmonary 14C-2-NP elimination after the 20 ppm exposure was of shorter duration (7 versus 12 hr) and had a slightly smaller half life (64 versus 71 min) than after the 154 ppm exposure. During the 12 - 48 hr post-exposure interval 14C-2-NP was eliminated slightly faster after the 154 ppm exposure than following the 20 ppm exposure (t
1/2 = 13.2 versus 16.4 hr).

Pulmonary 14CO2-Excretion- Pulmonary excretion of 14CO2 during successive 12 hr intervals following i.v. administration and
inhalation of 14C-2-NP indicate over 80% of the 14CO2 recovered was excreted during the initial 12 collection interval. A slightly larger percentage of the 14CO2 was excreted during the first 12 hr after the 15.4 mg/kg dose than after the 103 mg/kg dose (91.4 versus 80.5%) while the reverse was true for the second 12 hr collection interval (5.7 versus 15.9%) . Although disproportionately more 14CO2 was excreted following the 154-ppm exposure than after the 20 ppm exposure, the excretion profile (% of total 14CO2 excreted during each interval) were similar following the two exposure concentrations.
Toxicokinetic parameters
Test no.:
#1
Toxicokinetic parameters:
half-life 1st: <2 hours following inhalation exposure, Note: 2-nitropropane has a low potential to accumulate in the rat. The kinetics of 2-NP are nonlinear in rats when the exposure concentration is less than 154ppm.

Metabolite characterisation studies

Metabolites identified:
no
Details on metabolites:
Most (88 +/- 4%) of the radioactivity applied to the column was recovered, and 85 - 87% of the recovered radioactivity could be assigned to 6 or possibly 7 distinct peaks, i.e., metabolites. The minor differences in retention volumes at the two dose levels were believed to be due-to small differences in chromatographic conditions. However, difference in chromatographic conditions do not account for the difference in the relative size of these peaks at the two dose levels. Thus these chromatograms indicate that the same metabolites were excreted at the two dose levels but that relative amounts of each metabolite excreted depends on the dose. No attempt was made to identify the urinary metabolites other than determining that authentic 14C-2-NP took 26 to 31 min to elute from the column.

Any other information on results incl. tables

None

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): low bioaccumulation potential based on study results
These data indicate that -40% of the inhaled 2-NP was absorbed. 2-NP was rapidly metabolized and eliminated, and thus has a low potential to accumulate during prolonged or repeated exposures.The estimated half-life for 2-NP in the rat is less than 2 hours. The kinetics of 2-NP are nonlinear in rats when the exposure concentration is > or = to 154 ppm.
Executive summary:

None