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Basic toxicokinetics

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Administrative data

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2017
Report Date:
2017

Materials and methods

Objective of study:
absorption
distribution
excretion
toxicokinetics
Test guideline
Qualifier:
no guideline available

Test material

Reference
Name:
Unnamed
Type:
Constituent
Radiolabelling:
yes
Remarks:
[14C]-4,5 Dimethyl-1-hexanol was synthesized for use as representative radiolabeled tracer molecule of Isooctanol.

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on species / strain selection:
Sprague Dawley (Crl:CD[SD]) rats were used as the test system on this study. This species and breed of animal is recognized to be appropriate for this type of metabolism study. The rat was utilized because it is a widely used species for which significant historical control data are available and because it has been determined to be a pharmacologically responsive species.
Sex:
male/female
Details on test animals and environmental conditions:
Test animals:
- Source: Charles River (Ashland, OH, US; Raleigh, NC, US)
- Age at study initiation: approximately 8 to 12 weeks of age
- Weight at study initiation: between 195 g and 305 g at the time of dosing
- Housing: individually in clean, suspended wire-mesh cages; wire-mesh caging was utilized to acclimate the animals to the wire-mesh flooring utilized in the glass metabolism cages utilized during excreta collection. After dosing, animals used in the pilot and mass balance phases were placed into glass metabolism cages for up to 168 hours
- Diet (e.g. ad libitum): PMI Nutrition International, LLC Certified Rodent LabDiet® 5002 (block), ad libitum
- Water (e.g. ad libitum): Reverse osmosis-treated water, ad libitum
- Acclimation period: approximately 1 week

ENVIRONMENTAL CONDITIONS:
- Temperature: 71.8°F to 72.8°F (22.1°C to 22.7°C)
- Humidity: 43.2% to 53.0%
- Air changes (per hr): 10 air changes per hour, 100% fresh air
- Photoperiod: 12 hours fluorescent light followed by 12 hours of darkness; the light/dark cycle was interrupted for protocol-specified activities as needed

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
corn oil
Duration and frequency of treatment / exposure:
All animals received a single oral or IV dose of [14C]-4,5 Dimethyl-1-hexanol at 10, 150, 450, or 1000 mg/kg and a target radioactivity of 40, 100, or 400 μCi/kg.
Doses / concentrationsopen allclose all
Dose / conc.:
150 mg/kg bw (total dose)
Remarks:
Oral gavage, [14C]-4,5 Dimethyl-1-hexanol (isooctanol equivalent)
Dose / conc.:
450 mg/kg bw (total dose)
Remarks:
Oral gavage, [14C]-4,5 Dimethyl-1-hexanol (isooctanol equivalent)
Dose / conc.:
1 000 mg/kg bw (total dose)
Remarks:
Oral gavage, [14C]-4,5 Dimethyl-1-hexanol (isooctanol equivalent)
Dose / conc.:
10 mg/kg bw (total dose)
Remarks:
IV bolus dose, [14C]-4,5 Dimethyl-1-hexanol (isooctanol equivalent)
No. of animals per sex per dose:
Pilot mass balance phase (1000mg/kg total oral gavage dose): 2M
Pharmacokinetic phase (150mg/kg total dose, 450mg/kg total dose, 1000mg/kg total dose, all oral gavage; 10mg/kg total dose IV bolus): 6M/6F
Mass balance phase (450mg/kg total dose, 1000mg/kg total dose, oral gavage): 3M/3F
Tissue distribution phase (450mg/kg total dose, 1000mg/kg total dose, oral gavage): 6M/6F
Control animals:
other: One animal not dosed with test material was used for matrix.
Details on study design:
[14C]-4,5 Dimethyl-1-hexanol was synthesized for use as representative radiolabeled tracer molecules of isooctanol. Concentration data were calculated from the radioactivity in each sample.

A pilot mass balance phase was conducted to determine if [14C]-isotridecanol equivalents-derived radioactivity was present in expired air. The pilot study demonstrated that up to approximately 25% of study radioactivity could be present in expired air; therefore, the collection of expired air was necessary for the calculation of mass balance in the definitive excretion mass balance study phase.

Three dose groups consisting of 6 male and 6 female Crl:CD(SD) rats each received a single oral dose of [14C]-isotridecanol equivalents at 150 mg/kg, 450 mg/kg, or 1000 mg/kg and a target radioactivity of 40 μCi/kg. A single dose group consisting of 6 male and 6 female Crl:CD(SD) rats received a single IV (bolus) dose of [14C]-isotridecanol equivalents at 10 mg/kg and a target radioactivity of 40 μCi/kg. Blood samples were collected from 3 animals/sex/time point/group at approximately 0.083, 0.25, 0.5, 1, 2, 4, 8, 24, 48, and 72 hours postdose.

For the excretion mass balance phase, 2 dose groups, each consisting of 3 male and 3 female Crl:CD(SD) rats, received a single oral dose of [14C]-isotridecanol equivalents at 450 mg/kg or 1000 mg/kg and a target radioactivity of 400 μCi/kg. Following dosing, animals were placed into glass metabolism cages for separate collection of expired air through 72 hours and urine and feces through 168 hours.

For the quantitative whole body autoradiography phase, 2 dose groups, each consisting of 6 male and 6 female Crl:CD(SD) rats, received a single oral dose of [14C]-isotridecanol equivalents at 450 mg/kg or 1000 mg/kg and a target radioactivity of 100 μCi/kg. Following dosing, blood samples were collected from 1 animal/time point at approximately 0.5, 1, 4, 8, 24, and 168 hours. Following blood collection, animals were processed for quantitative whole body autoradiography.

Whole blood, plasma, urine, expired air, activated carbon, cage rinse, feces, and cage wash samples were analyzed by liquid scintillation counting.
Details on dosing and sampling:
Male and female Crl:CD(SD) rats received a single oral dose of [14C]-4,5 Dimethyl-1-hexanol administered via oral gavage at 150 mg/kg, 450 mg/kg, or 1000 mg/kg, or a single IV (bolus) dose of [14C]-4,5 Dimethyl-1-hexanol at 10 mg/kg. Based on body weights, the dose to be administered was calculated on a mg/kg body weight basis using a dosage volume of 2 or 10mL/kg.

Results and discussion

Preliminary studies:
RADIOLABELED DOSE FORMULATION ANALYSES:
The predose and postdose analyses established homogeneity of the solutions as demonstrated by the low %CV, < 4% across triplicate analyses at each interval for each formulation. The radio-HPLC analyses of the dosing solutions performed prior to and after completion of dosing demonstrated radiochemical purity of 100%; therefore, stability over the dosing period was demonstrated.

ACTUAL DOSE ADMINISTERED:
The amount of Isooctanol to be administered was based on the body weights of the animals on the day of dosing. Each group received 101% to 106% of the target mass (mg/kg) and 97% to 111% of the target radioactivity (μCi/kg). For all other animals, the approximate amount delivered was calculated based on the volume administered to each animal.
Main ADME resultsopen allclose all
Type:
absorption
Results:
At 150mg/kg: plasma Cmax (66.3ug/g 0.5h post-dose males, 66.4ug/g 1h post-dose females); AUClast (474 h*ug/g males, 610 h*ug/g females); T1/2 (6.13h males, 7.05h females)
Type:
absorption
Results:
At 450mg/kg: plasma Cmax (216ug/g 1h post-dose males, 220ug/g 2h post-dose females); AUClast (2110 h*ug/g males, 3290 h*ug/g females); T1/2 (3.59h males, 4.02h females)
Type:
absorption
Results:
At 450mg/kg (QWBA phase): plasma Cmax (238ug/g 1h post-dose males, 297ug/g 8h post-dose females); AUClast (2370 h*ug/g males, 4370 h*ug/g females); T1/2 (3.57h males, not calculable for females)
Type:
absorption
Results:
At 1000mg/kg: plasma Cmax (369ug/g 4h post-dose males, 396ug/g 4h post-dose females); AUClast (6030 h*ug/g males, 8100 h*ug/g females); T1/2 (7.77h males, 46.8h females)
Type:
absorption
Results:
At 1000mg/kg (QWBA): plasma Cmax (533ug/g 4h post-dose males, 459ug/g 8h post-dose females); AUClast (8080 h*ug/g males, 7550 h*ug/g females); T1/2 not calculable
Type:
absorption
Results:
At 10mg/kg (IV dose): plasma Cmax (11.3ug/g at T0 males, 13.4ug/g at T0 females); AUClast (29.7 h*ug/g males, 32.0 h*ug/g females); T1/2 (10.3h males, 9.52h females)
Type:
absorption
Results:
At 450mg/kg: whole blood Cmax (172ug/g 4h post-dose males, 220ug/g 8h post-dose females); AUClast (2120 h*ug/g males, 3580 h*ug/g females)
Type:
absorption
Results:
At 1000mg/kg: whole blood Cmax (406ug/g 4h post-dose males, 350ug/g 8h post-dose females); AUClast (7770 h*ug/g males, 11500 h*ug/g females)
Type:
excretion
Results:
At 1000mg/kg (pilot phase): 94% radioactivity recovery from excreta in males; of this total percentage, 79% was recovered in urine, 9.4% in feces, 1.5% expired air
Type:
excretion
Results:
At 1000mg/kg (excretion phase): 93.7% radioactivity recovered in excreta in males (91.0-96.7% range); 92.8% radioactivity recovered in excreta in females (87.6-98.6% range); of this total percentage of 92.8%, 87% was recovered in urine, 6% in feces
Type:
excretion
Results:
At 450mg/kg (excretion mass balance phase): 92.7% radioactivity recovered in excreta in males; (91.7-93.4% range); 93.4% radioactivity recovered in excreta in females (90.9-95.5% range); of this total, 83-85% recovered in urine, 8-11% recovered in feces

Toxicokinetic / pharmacokinetic studies

Details on absorption:
CONCENTRATION AND KINETICS IN PLASMA:
The AUClast increased in a linear, but greater than dose-proportional manner for males, as shown by the increasing dose-normalized AUClast from 3.2 (h·μg/g)/(mg/kg) at 150 mg/kg in males to 6.0 (h·μg/g)/(mg/kg) at 1000 mg/kg; a similar increase was seen in females. Concurrent with the observation in the PK phase, plasma AUClast in the QWBA phase increased 3.4-fold over a 2-fold increase in dose for males and 2-fold for females, further indicating that exposure to [14C]-isooctanol equivalents is nearly dose proportional or slightly greater than dose-proportional over the dose range examined.

Using the IV AUClast to calculate bioavailability for each group receiving an oral dose demonstrated bioavailability > 100% at all dose levels, from a minimum of 105% for males receiving 150 mg/kg to 251% for females receiving 1000 mg/kg. Greater than dose-proportional AUClast and bioavailability >100% are consistent with an extended absorption phase following oral administration at increasing doses.

CONCENTRATION AND KINETICS IN WHOLE BLOOD:
Approximately 2.7% of the dose was circulating in whole blood at Tmax following a single oral dose of 450mg/kg [14C]-isooctanol equivalents. The mean ratio of whole blood to plasma exposure to [14C]-isooctanol-equivalents was approximately 0.86 in males and females, demonstrating approximately equal distribution of isooctanol in plasma and blood cells.

Approximately 3.0% of the dose was circulating in whole blood at Tmax following a single oral dose of 1000mg/kg [14C]-isooctanol equivalents. The ratio of whole blood to plasma exposure to [14C]-isooctanol-equivalents was approximately 0.95 in males and 1.52 in females, demonstrating approximately equal distribution of Isooctanol in plasma and blood cells. While the dose increased 2-fold, the whole blood AUClast increased 3.6-fold for males and 3.2-fold for females. The data indicate that exposure to [14C]-isooctanol equivalents is slightly greater than dose-proportional over the 2-fold dose range examined.
Details on distribution in tissues:
After a single oral dose of [14C]-isooctanol equivalents at 450 mg/kg, [14C]-isooctanol and/or its metabolites were broadly distributed and detected by QWBA in all tissues for at least 24 hours postdose, except for the eye, brain, and uveal tract of both males and females, and the muscle (femoral), pituitary gland, and testes of males (8h). The [14C]-isooctanol equivalents were measured in the fat (yellow) and prostate of male rats and the adrenal gland, fat (yellow), skin, and uterus of female rats up to 168 hours postdose.

In males, Cmax for [14C]-isooctanol-derived radioactivity was highest in the tissues expected from extensive absorption following oral administration: primarily the liver and the small intestine, followed by the kidney (all sections), stomach, harderian gland, and large intestine. The lowest radioactivity was observed in the bone marrow, muscle, prostate, testes, thymus, uveal tract, pituitary gland, eye (lens), and brain. The Tmax was generally 1 hour postdose. Exposure, as measured by AUClast, was highest in the fat. Otherwise, AUClast reflected the similar ranking of tissues as was observed in Cmax, i.e. highest in liver, kidney, harderian gland, stomach, and large intestine and lowest in the eye (lens), pituitary gland, testes, uveal tract, and brain. As calculated using AUClast, the tissue:plasma ratios were < 1.7 demonstrating limited affinity for all tissue types except for fat and liver, whose ratios were still nominal at 3.0 and 2.1, respectively.

In females, Cmax for [14C]-isooctanol-derived radioactivity was highest in the tissues expected from extensive absorption following oral administration: primarily the kidney (all sections), liver, small intestine, and stomach. The thyroid concentration was also high relative to the other tissues, but rapidly decreased after Tmax at 1 hour postdose and the resultant AUClast was moderate. The lowest radioactivity relative to other tissues (<100,000ng/g) was observed in the eye (lens), skin, large intestine, thymus, pituitary gland, muscle (femoral), fat, and brain, in decreasing order of concentration. The Tmax was generally 8 hours postdose. Exposure, as measured by AUClast, was highest in the kidney, small intestine, liver, adrenal gland, uterus, fat, and stomach, and lowest in the uveal tract, eye (lens), and brain. Adrenal gland, fat, skin, and uterus exposures were all impacted by concentrations above the limit of quantitation at 168 hours postdose. As calculated using AUClast, the tissue:plasma ratios were < 1.0 demonstrating limited affinity for all tissue types except for liver, small intestine, and kidney, whose ratios only slightly exceeded 1.0 with a maximum ratio of 1.2. For both males and females, where half-lives could be calculated with reasonable certainty, they were < 10 hours.



After a single oral dose of [14C]-isooctanol equivalents at 1000 mg/kg, [14C]-isooctanol and/or its metabolites were detected in all tissues for at least 24 hours postdose, except for brain and uveal tract in males (8h) and from 1 to 8 hours postdose in the brain and 4 to 24 hours postdose in the uveal tract in females.

In males, Cmax for [14C]-isooctanol-derived radioactivity was highest in the tissues expected from extensive absorption following oral administration, plus others that were also identified in the low (450 mg/kg) dose group: primarily the small intestine, large intestine, liver, stomach, pancreas, fat, harderian gland, and adrenal gland. The lowest radioactivity (equivalent to < 235,000 ng/g) was observed in the muscle (femoral), bone marrow, skin, uveal tract, prostate, thymus, pituitary gland, eye (lens), and brain. The Tmax was generally 8 hours postdose. The AUClast was highest in harderian gland, fat, liver, large intestine, adrenal gland, and stomach, with tissue:plasma ratio ranging from 0.05 to 5.22. The AUClast was lowest in the brain and uveal tract.

In females, Cmax for [14C]-isooctanol-derived radioactivity was highest in the stomach, liver, harderian gland, adrenal gland, large intestine, and small intestine. The lowest radioactivity (equivalent to < 220,000 ng/g) was observed in the fat, skin, thymus, pituitary gland, muscle (femoral), eye (lens), and brain, in decreasing order of concentration. The Tmax was generally 8 hours postdose. Exposure, as measured by AUClast, was highest in the fat, kidney (cortex), adrenal gland, large intestine, stomach, uterus, and ovary, and lowest in the brain. Adrenal gland, fat, large intestine, ovary, and uterus exposures were all impacted by concentrations above the limit of quantitation at 168 hours postdose. As calculated using AUClast, the tissue:plasma ratios were < 2.0 for all tissues except stomach, large intestine, kidney (cortex), fat, and adrenal gland. Where half-lives could be calculated with reasonable certainty, they ranged from 7.8 to 10 hours in males.

Tissue exposure to [14C]-isooctanol equivalents increased an average of 2- to 3-fold for both males and females with increasing dose from 450 mg/kg to 1000 mg/kg when evaluated from AUC0-24 values.
Details on excretion:
In the pilot mass balance phase, after a single oral dose of [14C]-isooctanol equivalents to Crl:CD(SD) rats at 1000 mg/kg, a total of approximately 94% of the administered radioactivity was recovered in the excreta and expired air collected from 2 male rats over 48 hours postdose. Because the small amount recovered in the expired air does not significantly impact mass balance, expired air collection was discontinued for the remaining mass balance phases.

In the mass balance phase, after a single oral dose of [14C]-isooctanol equivalents to Crl:CD(SD) rats at both 450 mg/kg and 1000mg/kg, the majority of the elimination occurred in the first 24 hours postdose in males and females.

Metabolite characterisation studies

Metabolites identified:
not measured
Remarks:
Metabolites are being identified in a follow-up study (2018).

Applicant's summary and conclusion

Executive summary:

In a study to determine the plasma pharmacokinetics of [14C]-isooctanol equivalents-derived radioactivity in male and female Sprague Dawley rats following a single oral (gavage) or intravenous (bolus) administration, to determine the routes of elimination and excretion mass balance, and tissue distribution and tissue pharmacokinetics of [14C]-isooctanol equivalents-derived radioactivity in male and female Sprague Dawley rats following a single oral (gavage) administration.

Isooctanol is nearly completely absorbed following oral administration and excreted primarily in the urine. Accordingly, tissue concentrations are highest in the tissues associated with oral administration and urinary excretion, but [14C]-isooctanol and/or its metabolites were detected in all tissues, usually at low or equivalent concentrations relative to the plasma. There appeared to be little affinity for whole blood cells or any tissues types, and the majority of the [14C]-isooctanol derived radioactivity was excreted in the first 24 hours postdose. There was no notable difference in the absorption, distribution, or excretion of [14C]-isooctanol equivalents with respect to the sex of the rats.