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Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2017
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

Referenceopen allclose all

Title:
Metabolism and disposition of [14C]-methylcyclosiloxanes in rats
Author:
Domoradzki JY, Sushynski CM, Sushynski JM, McNett DA, Landingham CV, Plotzke KP
Year:
2017
Bibliographic source:
Toxicology Letters, 279, 98-114
Report Date:
2017
Reference Type:
study report
Title:
Unnamed
Year:
2012
Report Date:
2013

Materials and methods

Objective of study:
metabolism
other: disposition
Test guideline
Qualifier:
equivalent or similar to
Guideline:
EPA OPP 85-1 (Metabolism and Pharmacokinetics)
Version / remarks:
The study was conducted in accordance with EPA Toxic Substance Control Act (EPA-TSCA, 1984)
Good Laboratory Practice Standards.
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Specific details on test material used for the study:
The 14C-decamethylcyclopentasiloxane ([14C]D5) used in the study at 100 mg/kg bw was prepared by the Dow Corning Corporation, Auburn, MI. Chemical identity and radiochemical purity of the labelled test article was determined using Gas Chromatography with Mass Spectrometric detection (GC/MS) and High Performance Liquid Chromatography (HPLC) with a radioactivity flow-through detector (RAD). More than one batch of radiolabelled material was used.

Specific activities (mCi/g) of [14C]D5 were 29.081 mCi/g and 29.453 mCi/g. Radiochemical purities were: D5, 98.4 and 99.8. Non-radiolabelled D5 were supplied by the Dow Corning Corporation, Auburn, MI with purity of D5, 99.0%.
Radiolabelling:
yes

Test animals

Species:
rat
Strain:
Fischer 344
Details on species / strain selection:
CDF®(Fischer344)/CrlBR rats
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Inc
- Age at study initiation: approximately 9 weeks of age minimum at dosing
- Weight at study initiation: The mean body weights of female and male rats from the various groups
ranged from 150 to 160 g and 200–231 g
- Housing: Animals were housed individually in suspended wire-mesh cages except for the mass
balance portion of the study where they were housed in glass Roth-style glass metabolism cages (air
flow 500 to 1000 mL/min, 24–48 h acclimation).
- Diet (e.g. ad libitum): Animals were provided LabDiet® Certified Rodent Diet #5002 (PMI Nutrition
International, St. Louis, MO) ad libitum.
- Water (e.g. ad libitum): municipal water ad libitum.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 18–26 °C
- Humidity: 24–70% relative humidity
- Photoperiod (hrs dark / hrs light): 12-h light/dark cycle
Indwelling jugular vein cannulae, implanted by the animal supplier, were used to facilitate collection of
the multiple blood samples collected from each animal (approx. 0.2 mL per time point).
The studies were approved by the Laboratory Animal Care and Use Committee of the lab, fully
accredited by the Association for the Assessment and Accreditation of Laboratory Animal Care
International.

Administration / exposure

Route of administration:
oral: gavage
Vehicle:
other: D5 administered in dosing solution prepared with a Rodent Liquid Diet (RLD)
Details on exposure:
Single dose exposure
Doses / concentrationsopen allclose all
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
100 mg 14C-D5/kg bw in a rodent liquid diet (RLD) vehicle
Dose / conc.:
100 mg/kg bw/day (nominal)
Remarks:
A group of females in the D5 low study for blood kinetics was administered 14C-D5/kg bw in a corn oil vehicle to measure blood kinetics for comparison of vehicles.
Details on study design:
Studies were conducted with a single oral gavage administration 100 mg 14C-D5/kg bw in a RLD vehicle (D5 low). In addition, a group of females in the D5 low study for blood kinetics was administered 100 mg 14C-D5/kg bw in a corn oil vehicle to measure blood kinetics for comparison of vehicles.
Study groups consisted of female and male Fischer 344 rats designated for determining blood kinetics, mass balance (absorption, distribution, metabolism and excretion) and tissue kinetics. Blood (approximately 0.2 mL) was collected at 15, 30, 60 min, and 2, 4, 8, 24, 48, 72, 96, 120, 144 and 168 h post-dosing via a jugular vein cannula. 14C-Activity and parent were measured in blood and kinetic parameters determined.
Collection intervals for urine, CO2 and faeces were 24 h through 168 h (in addition, urine was collected at 0–12 and 12–24 h) in the mass balance groups. Collection intervals for expired volatiles were: 0–1, 1–2, 2–4, 4–6, 6–12, 12–24, and 24 h intervals through 168 h.
Selected tissues and remaining carcass (including pelt) were collected at 168 h post-dosing. Radioactivity (14C-activity) and parent were measured in collected samples and urine was profiled.
Tissues were collected at 2, 6, 12, 24, 48, 72, 120 and 168 h postdosing in the groups to determine tissue kinetics. Tissues collected at these times were adrenals, digestive tract (without contents), perirenal fat, brown fat (D5 study only), liver, lung, ovaries, spleen, testes and uterus. Radioactivity (14C-activity) and parent were measured in extracted samples.
Details on dosing and sampling:
[14C]D5 was administered in dosing solutions prepared with a Rodent Liquid Diet (RLD) product# F6112SP (Bio-Serv, Frenchtown, NJ). The dosing solutions were prepared in RLD to deliver approximately 0.29 mCi/kg body weight (bw), with a nominal dose of 100 mg D5/kg bw in 5 mL dosing solution/kg bw.

Test substance administration:
Test article was administered as a single dose by oral gavage in an RLD vehicle. RLD vehicle was chosen since a food matrix was desired to evaluate uptake kinetics especially at lower doses. The corn oil vehicle was chosen for one subset of female rats in the D5 low dose study (blood kinetics) to compare to a previous D5 high dose study where a corn oil vehicle was used.
Vehicle only and test article solutions were administered by oral gavage with a 15 gauge, 100 mm plastic oral feeding tube and syringe at 5 mL/kg bw. Volume was adjusted based upon the most recent individual body weights. The weight of the dose solution administered was determined gravimetrically.
The actual mean dose administered (D5 low) ranged from 91.48–112.94 mg/kg in female rats and 92.20–99.15 mg/kg in male rats. Females received a mean range of 35.06–42.45 μCi/animal and males received a mean range of 46.24–54.76 μCi/animal.
Statistics:
Statistical comparisons between parent and 14C-activity Area-under-the-Curves (AUCs) were conducted using the Bailer method, Nedelman et al. (1995) and the Satterthwaite approximation method, Nedelman and Jia (1998).

Data analysis and quality assurance:
Calculations of mean and standard deviation (or standard error of the mean) of sample concentrations were performed using Microsoft Excel™ v5, v7,v9 and 2007.Calculations of μg eq were performed using Provantis™ Version 8.2 (D4 low dose study).Blood Area-Under the–Curves (AUC)s for parent and radioactivity including statistical analyses were calculated using SAS/STAT software, v8.2 and v9.3. These studies were conducted in accordance with EPA Toxic Substance Control Act (EPATSCA, 1984) Good Laboratory Practice Standards.

Results and discussion

Preliminary studies:
Mass balance: dose recovered and disposition:
Mass balance studies were carried out to investigate the absorption, distribution, metabolism and elimination of D4 and D4 following single low dose oral administration.

The mean percentage of the administered dose recovered ranged from 95.5% and 103.24% in females and males. The predominate route of elimination was in faeces as the radioactivity recovered accounted for 82.52 and 82.99% of the recovered dose (D5 low) in females and males, respectively. Urine contained 8.15% and 8.99% of the recovered dose) in females and males, respectively. Expired volatiles contained 2.01 and 1.26% of the recovered dose (D5 low) in females and males, respectively. The percentages of the dose recovered in the urine, faeces and expired 14CO2 of female and male rats were similar. At 168 h, the remaining carcass (including pelt) accounted for 5.07% and 4.72% of the recovered radioactivity (D5 low) in females and males, respectively. Following administration of 14C-D5 at 100 mg/kg bw in an RLD vehicle, the percentage absorbed was 17.4% and 16.9% in females and males, respectively.

Blood kinetics:
Parent concentrations for D5 at 100 mg/kg bw in a rodent diet vehicle were measurable through 168 h and 72 h for females and males, respectively. Radioactivity concentrations were measurable through 168 h for both females and males. In females and males, the peak blood concentration, Cmax, occurred at 4 h for both D5 and total radioactivity and the concentrations were similar.
As part of the study where 14C-D5 at 100 mg/kg bw was administered in a rodent liquid diet vehicle, a group of females was administered the same dose in a corn oil vehicle. Parent and total radioactivity concentrations were measurable through 168 h.
In this group of females, the peak blood concentration occurred at 4 h for both D5 and total radioactivity. In females dosed with 14C-D5 in a corn oil vehicle, Cmax were higher than those observed in the females that were dosed with 14C-D5 in an RLD vehicle (Table 3). With the high dose female data shown for comparison, Tmax is delayed for both parent and radioactivity as compared to the low dose female data which denotes a delay in gastric emptying.
Calculated AUCs from the blood time-course data (D5 low) indicated that 14C-activity was absorbed (AUC in μg 14C-equivalents D5 × hr/g of blood) and were similar, no statistically significant difference, between females and males; AUCs for parent D5 were also similar between sexes.
There was a statistically significant difference between blood AUCs for parent D5 and 14C-activity for females, males and females (corn oil).
There was not a statistically significant difference in 14C-activity blood AUCs between females dosed with D5 (low) in the RLD vs. the corn oil vehicle; however, the blood parent AUCs were significantly different. Terminal 14C-activity half-lives of elimination were similar for females and males. Parent D5 terminal half-lives were different between females and males with the blood elimination half-life in females slower by approx. 4 fold.

Toxicokinetic / pharmacokinetic studies

Details on absorption:
Following administration of 14C-D5 at 100 mg/kg bw in a RLD vehicle, the percentage absorbed was 17.4% and 16.9% in females and males, respectively.
Calculated AUCs from the blood time-course data (D5 low) indicated that 14C-activity was absorbed (AUC in μg 14C-equivalents D5 × hr/g of blood) and were similar, no statistically significant difference, between females and males; AUCs for parent D5 were also similar between sexes.
There was a statistically significant difference between blood AUCs for parent D5 and 14C-activity for females, males and females (corn oil). There was not a statistically significant difference in 14C-activity blood AUCs between females dosed with D5 (low) in the RLD vs. the corn oil vehicle; however, the blood parent AUCs were significantly different.
Details on distribution in tissues:
Tissue distribution and kinetics: parent (D5) and 14C-activity in blood and tissues key tissues described in PBPK models were blood, liver, fat and lung.

Parent D5:
Parent D5 was detected in all tissues with the Cmax in tissues typically observed at 2, 4 or 6 h post-dosing, except in brown fat and perirenal fat where the highest concentrations were observed at 12 and 48 h post-dosing. Parent D5 was measurable in tissues through 168 h post-dosing in all animals. In blood Cmax parent D5 was lower than Cmax levels in tissues. Half-lives of elimination (terminal phase) of parent D5 were fastest in liver (17.62 h) and blood (18.90 h); for females and males, respectively. Slower t1/2s of elimination were in the digestive tract: 441.20 and 538.00 h for females and males, respectively. The half-life of parent in blood for the female corn oil group was longer than for females administered D5 in a rodent diet vehicle; 76 and 72 h, respectively.

14C-Activity D5:
Radioactivity was detected in key model tissues with the highest concentrations at 2, 4, or 6 h post-dosing, except in brown fat and perirenal fat where the highest concentrations were observed at 12 and 48 h post-dosing. Total radioactivity was measurable in tissues through 168 h post-dosing in all animals. The Cmax for total radioactivity in blood was at lower levels than Cmax levels in tissues. The half-live of elimination (terminal phase) of total radioactivity was fastest in liver and lung. The slower t1/2s of elimination were in the digestive tract (386.17 h) and perirenal fat (341.32 h) for females and males, respectively.

Metabolite characterisation studies

Details on metabolites:
The radioactivity eliminated in the urine consisted entirely of polar metabolites of D5. The metabolite peak assignments are based on retention time comparison to urinary metabolite profiles performed in a separate study (Varaprath et al., 2003). No confirmation of identity was conducted within these studies.

The mean percentage of radioactivity that was attributed to individual metabolites from urine at 0–12 and 12–24, hours following oral administration D5 in the low dose study. Dimethylsilanediol represented the greatest percentage of total urinary radioactivity followed by methylsilanetriol. The percentages for dimethylsilanediol for the two collection intervals ranged from 53 to 58% in female animals and 50 to 53% in male animals and the percentages for methylsilanetriol for the three collection intervals ranged from 35 to 36% in female rats and 38 to 42% in male rats. Dimethyldisiloxane-1, 3, 3, 3-tetrol as a percentage of urinary activity for the two collection intervals ranged from 1 to 2% in females and 2% in male animals. The average sum of de-methylated peak percentages ranged from 39 to 41% for females and 44 to 47% for males.
Statistical analysis indicates a difference between gender averaged sums at the 0–12, 12–24 and 48–72 (data not shown) h collection intervals for demethylated metabolites, p < 0.05 using a two-tailed t-test assuming equal variance.

Parent D5 represented the major percentage of radioactivity in the faecal samples analysed from the 0–24 h collection interval for both sexes. Ninety-three and 91% percent of faecal radioactivity was D5 in female and male rats, respectively, for the 0–24 h collection interval. An attempt was made to identify unknown peaks in the HPLC/RAD chromatogram by collecting fractions from 0–24 h faecal extracts. Hydroxylated D5 metabolite (D4D’OH) was observed in extracts by GC/MS; however, peak assignment in the radiochemical profile was inconclusive.

Any other information on results incl. tables

Tissue kinetics and metabolism:

Total radioactivity concentration, 14C-activity, is composed of both parent D5 and metabolites of D5. Blood and tissues had measurable concentrations of both parent D5 and metabolites. The ratio of the AUCs for D5 in tissues to blood was greater than 1 for all tissues. The highest tissue-to-blood AUC ratios were observed in brown fat, 100.9 and 72.9, for females and males, respectively. The percentage of the total radioactivity attributed to metabolites in tissues and blood from females ranged from 7.74% to 56.09% with liver (56.09%) and blood (47.81%) having the highest percentages. The and blood from males ranged from 5.12% to 71.38% with liver (71.38%) and blood (46.24%) having the highest percentages.

The percentage of total radioactivity in the blood attributed to metabolites following administration of 14C-D5 (low) in a corn oil vehicle to a subset of females was 27.95% which was a lower percentage of metabolites than observed in the female group where administration of D5 was in a rodent diet vehicle, 47.81%. There was a statistically significant difference in mean blood parent AUCs between females that were dosed with D4 in RLD vs. dosed with D4 in the corn oil vehicle.

Elimination

Radioactivity was measurable through 168 h in faeces, expired volatiles, urine, and as 14CO2 following single oral gavage administration of D5. A total of 2.0 and 1.3% of the recovered dose was accounted for in expired volatiles in females and males, respectively. The highest amounts of radioactivity measured in expired volatiles was during the 4–6 h collection interval for females (17.1 μg eq/h) and males (15.8 μg eq/h, data not shown), respectively. The highest amounts of parent D5 measured in expired volatiles was during the 4–6 h collection intervals for females (15.2 μg/h) and males (15.7 μg/h, data not shown), respectively. A total of 8.2 and 9.0% of the recovered dose was accounted for in the urine in females and males, respectively. The highest concentration in urine was measured at the 12–24 h collection interval, 84 and 144 μg eq/g in female and male rats (data not shown), respectively. A total of 83% of recovered radioactivity was found in faeces. The largest percentage of elimination of radioactivity excreted in faeces occurred during the first collection interval, 0–24 h. The highest concentration in faeces was measured at the 0–24 h collection interval, 1618 and 1890 μg eq/g in female and male rats (data not shown), respectively. Concentrations of parent D5 were measurable through 96 h in faeces and through 168 h in expired volatiles following single oral gavage administration of 14C-D5. The highest concentration of parent in faeces was measured in the 0–24 h collection interval with 1786 and 1513 μg/g in female and male rats (data not shown), respectively.

Terminal half-lives of elimination for radioactivity were shorter in male animals for expired volatiles, urine, faeces, and CO2. Half-lives ranged from 28.36 to 69.77 h in females and 25.72 to 49.39 h in males. The AUC for total radioactivity is composed of both parent D5 and labelled metabolites. Comparison to the AUC derived from parent D5 demonstrates the percentage of the 14C-pool that is composed of metabolites. Only faeces (males) contained both parent and metabolites.

No parent D5 was found in urine samples; only metabolites were present. The percentage of the total radioactivity attributed to metabolites in excreta from females following administration was: urine (100.00%) and in males it was: faeces (19.40%) and urine (100.00%).

Applicant's summary and conclusion

Conclusions:
This study demonstrated a comparison of the low vs. high dose oral gavage administration of D5 demonstrated dose-dependent kinetic behaviour. Data suggest differences in metabolism between the low and high dose administration indicating high dose administration results in or approaches non-linear saturated metabolism. In addition, studies with high doses suggests vehicle my influence absorption.

The study showed that a high mean percentage of the administered dose recovered ranged from 95.5% and 103.24% in females and males. The predominate route of elimination was in faeces as the radioactivity recovered accounted for 82.52 and 82.99% of the recovered dose (D5 low) in females and males, respectively. Urine contained 8.15% and 8.99% of the recovered dose) in females and males, respectively. Expired volatiles contained 2.01 and 1.26% of the recovered dose (D5 low) in females and males, respectively. The percentages of the dose recovered in the urine, faeces and expired 14CO2 of female and male rats were similar. At 168 h, the remaining carcass (including pelt) accounted for 5.07% and 4.72% of the recovered radioactivity (D5 low) in females and males, respectively. Following administration of 14C-D5 at 100 mg/ kg bw in a RLD vehicle, the percentage absorbed was 17.4% and 16.9% in females and males, respectively.