Registration Dossier

Toxicological information

Basic toxicokinetics

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Administrative data

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
1988-11-28 to 1991-04-04
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
other: GLP study conducted according to well described internal methods that were similar to OECD Guidelines.
Cross-reference
Reason / purpose:
reference to same study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
1991
Report Date:
1991

Materials and methods

Objective of study:
toxicokinetics
Test guideline
Qualifier:
equivalent or similar to
Guideline:
OECD Guideline 417 (Toxicokinetics)
Deviations:
yes
Remarks:
Single dose chosen rather than recommended 2 doses
Principles of method if other than guideline:
The study followed a specific protocol and protocol amendments.
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Details on test material:
- Name of test material (as cited in study report): 2-[2-(dimethylamino)ethoxy]ethanol
- Substance type: Pure active substance
- Physical state: liquid (clear to pale-yellow liquid with slight amine odor)
- Analytical purity: 97.7%
- Expiration date of the lot/batch: no data
- Radiochemical purity (if radiolabelling): (target range >= 98%) (actual was 99%)
- Specific activity (if radiolabelling): (target range 1-5 mCi/mmol) (actual was 4.1 mCi/mmol)
- Locations of the label (if radiolabelling): uniform radiolabel
- Stability under test conditions: no data
- Storage condition of test material: most appropriate manner to ensure stability
Radiolabelling:
yes
Remarks:
uniformly 14C-labeled test substance (DMEE)

Test animals

Species:
rat
Strain:
other: CDF Fischer 344
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Harlan Sprague Dawley, Inc., Indianapolis, IN
- Age at study initiation: 10-12 wks
- Weight at study initiation: 188-253 g males, 147-170 g females
- Fasting period before study: two groups in study both male and female rats, one was fasted overnight (15 hrs) and the other was not fasted for comparison purposes.
- Housing: Prior to the study, animals were housed in plastic shoe box cages in groups of 3 or 4. The animals selected for the study were transferred to individual Roth-type metabolism cages (designed for the collection of urine, feces, and expired air).
- Individual metabolism cages: yes
- Diet: Agway Certified Rodent Chow, St. Mary's Ohio; ad libitum (non-fasted group)
- Water: Municipal drinking water (Municipal Authority of Westmoreland County, Greensburg, PA); ad libitum
- Acclimation period: At least 5 days when received and housed in shoe box cages; 12-36 hr acclimation in metabolism cages prior to the study.

ENVIRONMENTAL CONDITIONS
- Temperature (°C): Monitored continuously but not reported
- Humidity (%): Monitored continuously but not reported
- Air changes (per hr): N/A
- Photoperiod (hrs dark / hrs light): 12 hrs dark / 12 hrs light

IN-LIFE DATES: From: 1988-11-28 To: 1988-12-07

Administration / exposure

Route of administration:
dermal
Vehicle:
unchanged (no vehicle)
Details on exposure:
TEST SITE
- Area of exposure: Dorsal surface of truck of the animals in the thoracic region.
- % coverage: 3.24 - 4.0 cm2 area depending on the mean body weight of the animal, females were dosed over 1.8 X 1.8 cm area and males were dosed over 2 X 2 cm area.
- Type of wrap if used: The application site was occluded with polyethylene sheeting held in place by waterproof adhesive tape.
- Time intervals for shavings or clipplings: Prior to adminstration of the dose (amount of time prior not specified). Fur was clipped from the dorsal surface of the trunk of the animals in the thoracic region. Care was taken to avoid abrading the skin and to provide an exposure surface which was of suitable distance from the location of the jugular cannula just behind the head.

REMOVAL OF TEST SUBSTANCE
- Washing (if done): No
- Time after start of exposure: N/A

TEST MATERIAL
- Amount(s) applied (volume or weight with unit): 150 mg/kg of test substance including 10-25 uCi of the 14C test material.

USE OF RESTRAINERS FOR PREVENTING INGESTION: no
Duration and frequency of treatment / exposure:
72 hr period
Doses / concentrations
Remarks:
Doses / Concentrations:
150 mg/kg
No. of animals per sex per dose:
Four/sex
Control animals:
no
Positive control:
N/A
Details on study design:
Dose selection rationale: A determination was made of the LD50 for both male and female sexes following intravenous administration in order to allow two choices of doses for pharmacokinetics assessments.
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: blood, urine, feces and exhaled 14-CO2 were collected.
- Time and frequency of sampling:
-- Blood samples were collected at 0, 0.5, 0.75, 1, 2, 3, 6, 9, 12, 18, 24, 30, 36, 48, and 72 hrs post-dosing. Blood was placed in heparinized capiillary tubes and centrifuged in a microhematocrit centrifuge. The plasma was expressed into a tared scintillation vial and weighed. Aquasol-2 counting scintillant was added to the plasma aliquots with shaking and these were then counted by liquid scintillation spectrometry.
-- Urine was collected at 6, 12, 24, 36, 48, 72 hrs post-dosing under dry ice conditions and the urine samples were assayed for 14C levels after collection and then stored frozen until the preliminary metabolite analysis was conducted.
-- Feces were collected at room temperature (time points not specified) and processed to determine radioactivity levels. Feces were refrigerated until processed/analyzed and then stored in a freezer.
-- Expired 14CO2 was trapped using solutions of 2-methoxyethanol:ethanolamine (7:3) in three 24-hr intervals at room temperature. CO2 samples were refrigerated until processed/analyzed and then stored in a freezer.
-- Volatile fractions were trapped on silica gel sorbent traps in three 24-hr intervals and 14C was eluted from the silica by methanol extraction prior to counting.
--72 hrs after administration of dose, the animals were anesthetized with methoxyflurane and killed by exsanguination via cardiac puncture. Skin sections from each animal were removed in the following manner: the dosed area of skin, as well as the skin around the periphery of this site (approximately 1 cm from edge of dose site), was separated from each animal using blunt dissection techniques. The skin segment was pulverized in a Spex Freezer Mill and two aliquots from each sample were counted directly in a suspension of 10 mL Aquasol-2 and 6 mL water. The liver, kidney, bone marrow, brain, fat, heart, lung, muscle, spleen, and testes/ovaries and uterus were isolated from the carcass.

Radioactivity was determined from tissue oxidation of isolated organs or their homogenates, aliquots of homogenized carcass/pelt and feces (33% w/v in deionized water) from all collection intervals and from red blood cells collected at selected intervals. Radioactivity recovered from the dose site and from the occlusive bandage was also quantified. Tissue oxidation was conducted in a Biological Materials Oxidizer and quantification of the 14CO2 derived from combustion of these tissues was by liquid scintillation spectrometry.



Results and discussion

Preliminary studies:
A determination was made of the LD50 for both male and female sexes following intravenous administration in order to allow two choices of doses for pharmacokinetics assessments.

Additionally, probe intravenous and percutaneous studies were conducted over 48 hr post-dosing using two male rats per route at a dose level of 150 mg/kg; the intravenous dose was administered in saline solution and the percutaneous dose was applied neat to the dorsal skin.

When the 150 mg/kg dose was applied as undiluted test substance to the skin of the percutaneous probe animals, nearly the same disposition pattern as that for the IV doses occurred for radioactivity which penetrated the skin and was excreted in the urine over a 48 hr period. Overall, 89.1% +/- 0.6% of the applied dose was recovered in this segment of the probe studies (67% urine + 5% cage, totaling 72% eliminated in the urine; 3.5% in carcass residue of applied dose and 12.9% as unabsorbed dose from bandages taken from the dose site). Thus, more than 76% of the applied cutaneous dose was accounted for in excreta and tissues. Cutaneous recoveries indicated a substantial absorption over a 48 hr period for an undiluted, occluded dose of the test substance.

Pharmacokinetic parameter estimation for the probe study resulted in terminal (beta) phase t1/2 estimates of 12.9 hr.

Toxicokinetic / pharmacokinetic studies

Details on absorption:
Approximately 72% of the 14C-test substance applied to the non-fasted male rats at this dose level was absorbed through the skin. More than 71% of the applied 14C- test substance penetrated non-fasted female rat skin at 150 mg/kg. The absorption of total radioactivity was relatively rapid at this dose level, with t1/2 values of 1.5 hr for males and 2.4 hrs for females.

Approximately 73% of the 14C- test substance applied to the fasted male rats was absorbed following a dose of 150 mg/kg. The total amount of radioactivity absorbed through the skin of fasted female rats was slightly more than 60%. The elimination t1/2 values for fasted males was 18.7 hrs and 17.5 for females.
Details on distribution in tissues:
Minor fractions of 14C-test substance was recovered from the tissues of either sex from the non-fasted and fasted groups dosed at 150 mg/kg (See Table 2).
- Minor fractions of the 14C were also recovered in very minor amounts in tissues (liver: 0.02%, Kidney: 0.01%) of non-fasted male rats. The tissues of non-fasted female rats contained 0.03% of the applied dose (liver: 0.02%; kidney: 0.01%).
- Approximately 0.03% of the applied 14C was recovered from fasted male rats in the tissue (liver: <0.02%; expired 14CO2: 0.2%; and volatile 14C-test substance: 0.3%). In the fasted female group, very little of the dose (0.04%) was associated with the organs and tissues collected, with 0.26% found in the liver.
Details on excretion:
Detailed results are provided in Table 1 below.

Non-Fasted Male Rats:
Most of the radioactivity was recovered from non-fasted male rats in the urine and cage wash (65.5%) with the largest fraction of urinary radioactivity (31.4% of the total applied 14C dose) being voided over the first 12 hours after the dose application. By 48 hrs post dose, the majority of the urinary 14C had been excreted (56.8%). The major remaining fraction of the applied radioactivity was recovered as unabsorbed dose from the cutaneous appliances used to occlude the dose site (18.2%). Skin at the dose site contained another 5% while the carcass/pelt retained at additional 4.0 % of the dose. Minor fractions of the 14C dose were also recovered int he feces (1.7%), 14CO2 (0.2%), volatile 14C fractions (0.3%) and in very minor amounts in tissues (see Table X).

Non-Fasted Female Rats:
An average of 97.97 +/- 2.48% of the applied radioactivity was recovered. Urine was the primary excretion route. The largest 14C fraction in the urine was voided in the first 12 hr post-dosing (27.0% of the total applied 14C dose). by 48 hr post-dosing, a total of 49.7% of the 14C dose was recovered in the urine. While there was an apparent lower urine output compared to males, the recovery of radioactivity in cage washes (8.0%) accounted for much of the difference from analogous male animal urinary excretion of 14C. The remaining major fraction of radioactivity was recovered as unabsorbed dose in the cutaneous appliances which occluded the dosing site (22.0%). The skin at the dose site contained 4.0% of applied dose and another 3.8% of the radioactivity was found in the carcass/pelt. Lesser percentages of radioactivity were recovered in feces (5.6%) and as expired 14CO2 (0.2%) and volatile 14C activity (0.4%).

Fasted Male Rats:
An average of 95.96 +/- 1.04% of the applied radioactivity was recovered following a 150 mg/kg percutaneous dose. Urine was the primary route of excretion. The cumulative urinary total radioactivity time-course indicates that more than 65% of the applied radioactivity was absorbed and excreted in the urine over the 72-hr duration of the experiment (28.8% of the 14C dose was recovered in the urine during the first 12 hr post-dosing period). Additional fractions were found in the cage wash (4.4%), feces (3.1%), carcass/pelt (4.2%) and the skin at the dose site (5.8%). Only 0.2% of the 14C dose was found as expired 14CO2 and 0.3% was recovered as a volatile 14C fraction when trapped on silica gel sorbent.

Fasted Female Rats:
An average of 80.90 +/- 1.58% of the applied radioactivity was recovered following a 150 mg/kg percutaneous dose. Urine was the primary route of excretion. Cumulative urinary total radioactivity time-course indicates that 54.0% of the applied radioactivity was absorbed and excreted in the urine over the 72-hr duration of the experiment (21.4% during the first 12 hr post-dosing period). By 24 hr post-dosing, most of the urinary 14C eliminated by this route had been excreted. The remaining recovered dose was found in the unabsorbed dose fractions (rinses; occlusive appliances: 17.2%), the 14C recovered from the skin at or adjacent to the dosing site (3.6%), the cage wash (8.2%) in feces (1.4%), and the carcass/pelt (4.0%). Very little of the dose was found in 14CO2 traps (0.2%) or volatile traps (0.5%).
Toxicokinetic parameters
Test no.:
#1
Toxicokinetic parameters:
other: See Any other information on results incl. tables for toxicokinetic parameters for Total Radioactivity Mean Data and Pooled Plasma data.

Metabolite characterisation studies

Metabolites identified:
yes
Details on metabolites:
Data were consistent between treatment groups at each of the six intervals measured and the amount of metabolite(s) detected was similar for each sex (e.g., non-fasted males vs. fasted males) for a given interval. Female animals produced slightly less metabolite(s) than their male counterparts.

For male rats during the first 6 hr interval, 83-87% of the 14C was excreted as unchanged test substance and 12-16% of the radioactivity excreted as apparent metabolite(s). Female urinary data for unchanged test substance showed similar results between treatment groups for this 0-6 hr interval, with 90-92% as unchanged test substance and 8-9% as labeled apparent metabolite(s). The level of unchanged test substance decreased slightly (to 81-84%) for males over the 6-12 hr interval for both the fasted and non-fasted groups but was steady for females in this interval (87.9-88.2%). The apparent radioactivity metabolite(s) in this 6-12 hr interval amounted to 14-17% of the 14C excreted by males and 10-11% for females.

For the next 12-72 hr, the level of unchanged test substance for all of the 150 mg/kg cutaneous doses remained relatively constant in the urine, comprising 80 - 88% of the total excreted by males and 86 - 87% of that excreted by females; about 15 - 18% of that excreted by male animals was apparent 14C metabolite(s) during this 12 - 72 hr period while about 11 - 13% of that excreted by the females comprised a similar apparent metabolite fraction.

Any other information on results incl. tables

Table 1. Disposition of Radioactivity Following Cutaneous Administration of 150 mg/kg to Non-fasted and Fasted Fischer 344 Rats

Non-Fasted Rats Fasted Rats
% Recovery % Recovery
Males Females Males Females
A. Tissues & Excreta
Urine (0 - 12 hr) 31.36 +/- 8.50 27.03 +/- 8.72 28.80 +/- 5.03 21.36 +/- 6.37
Urine (12 - 24 hr) 15.33 +/- 6.84 13.12 +/- 1.78 14.25 +/- 1.91 11.57 +/- 0.90
Urine (24 - 72 hr) 14.34 +/- 3.82 13.65 +/- 3.46 17.65 +/- 6.14 12.85 +/- 2.58
Cage Wash 4.43 +/- 1.58 7.96 +/- 1.43 4.43 +/- 1.62 8.24 +/- 1.29
Subtotals 65.46 +/- 9.08 61.77 +/- 11.95 65.13 +/- 2.74 54.02 +/- 3.39
Feces (0 - 24 hr) 0.97 +/- 0.85 4.42 +/- 6.96 2.43 +/- 2.40 0.46 +/- 0.24
Feces (24 - 72 hr) 0.70 +/- 0.30 1.14 +/- 0.64 0.66 +/- 0.39 0.97 +/- 0.79
Subtotals 1.66 +/- 1.02 5.56 +/- 6.95 3.09 +/- 2.14 1.44 +/- 1.01
Expired14CO2(0 - 24 hr) 0.11 +/- 0.02 0.10 +/- 0.01 0.12 +/- 0.01 0.09 +/- 0.01
Expired14CO2(24 - 72 hr) 0.07 +/- 0.01 0.06 +/- 0.01 0.06 +/- 0.00 0.07 +/- 0.02
Subtotals 0.18 +/- 0.02 0.16 +/- 0.02 0.18 +/- 0.01 0.16 +/- 0.02
Volatiles (0 - 72 hr) 0.31 +/- 0.22 0.35 +/- 0.24 0.30 +/- 0.22 0.50 +/- 0.17
Tissues 0.04 +/- 0.01 0.03 +/- 0.01 0.03 +/- 0.01 0.04 +/- 0.01
Carcass/Pelt 3.96 +/- 1.02 3.76 +/- 1.61 4.23 +/- 0.89 4.01 +/- 1.28
Section A (Absorbed) Totals 71.61 +/- 8.34 71.62 +/- 4.90 72.96 +/- 3.70 60.17 +/- 1.45
B. Occlusion Devices and Skin
Tape, bandage & plastic rinses 18.16 +/- 4.69 21.97 +/- 5.00 17.01 +/- 4.26 16.74 +/- 1.24
Skin rinses 0.18 +/- 0.13 0.31 +/- 0.12 0.14 +/- 0.06 0.43 +/- 0.48
Recovery in skin at dose site 5.48 +/- 1.47 4.04 +/- 0.99 5.84 +/- 1.42 3.55 +/- 1.36
Section B Totals 23.82 +/- 5.66 26.33 +/- 5.62 22.98 +/- 3.74 20.71 +/- 0.66
Total % Recovered Dose 95.44 +/- 4.78 97.97 +/- 2.48 95.96 +/- 1.04 80.90 +/- 1.58

Table 2. Distributuion of Radioactivity in Tissues from Non-Fasted and Fasted 344 Rats Following Cutaneous Exposure to 150 mg/kg Test Substancea

Non-Fasted
Male Female
Tissue % of Dose µg Equiv /g % of Dose µg Equiv /g
Liver 0.022 +/- 0.004 1.007 +/- 0.257 0.020 +/- 0.004 0.726 +/- 0.152
Kidney 0.006 +/- 0.002 1.315 +/- 0.385 0.005 +/- 0.001 0.815 +/- 0.160
Bone Marrow 0.000 +/- 0.000b 7.253 +/- 8.834 0.000 +/- 0.000b 0.347 +/- 0.694
Spleen 0.002 +/- 0.000 0.646 +/- 0.122 0.003 +/- 0.001 0.700 +/- 0.214
Brain 0.000 +/- 0.000 0.000 +/- 0.000 0.000 +/- 0.000 0.000 +/- 0.000
Heart 0.001 +/- 0.000 0.567 +/- 0.053 0.001 +/- 0.000 0.565 +/- 0.094
Lung 0.002 +/- 0.001 0.805 +/- 0.192 0.003 +/- 0.001 0.836 +/- 0.216
Muscle 0.000 +/- 0.000b 0.369 +/- 0.106 0.001 +/- 0.000b 0.455 +/- 0.358
Fat 0.000 +/- 0.000b 0.000 +/- 0.000 0.000 +/- 0.000b 0.093 +/- 0.186
Ovaries  ---  --- 0.000 +/- 0.000 0.716 +/- 0.314
Uterus  ---  --- 0.001 +/- 0.000 1.063 +/- 0.807
Testes 0.003 +/- 0.002 0.361 +/- 0.243  ---  ---
Carcass/Pelt 3.964 +/- 1.019 8.540 +/- 2.143 3.756 +/- 1.611 7.541 +/- 3.278
Fasted
Male Female
Tissue % of Dose µg Equiv /g % of Dose µg Equiv /g
Liver 0.019 +/- 0.004 0.645 +/- 0.106 0.026 +/- 0.007 1.020 +/- 0.311
Kidney 0.006 +/- 0.002 1.026 +/- 0.237 0.007 +/- 0.001 1.277 +/- 0.174
Bone Marrow 0.001 +/- 0.001b 5.359 +/- 6.644 0.001 +/- 0.001b 9.845 +/- 8.682
Spleen 0.001 +/- 0.000 0.316 +/- 0.125 0.002 +/- 0.001 0.566 +/- 0.139
Brain 0.000 +/- 0.000 0.000 +/- 0.000 0.001 +/- 0.002 0.150 +/- 0.300
Heart 0.002 +/- 0.000 0.925 +/- 0.206 0.001 +/- 0.000 0.461 +/- 0.142
Lung 0.001 +/- 0.001 0.459 +/- 0.324 0.003 +/- 0.001 0.772 +/- 0.283
Muscle 0.000 +/- 0.000b 0.457 +/- 0.173 0.001 +/- 0.000b 0.436 +/- 0.106
Fat 0.000 +/- 0.000b 0.369 +/- 0.454 0.001 +/- 0.000b 1.124 +/- 0.993
Ovaries  ---  --- 0.000 +/- 0.000 1.014 +/- 0.196
Uterus  ---  --- 0.001 +/- 0.000 0.976 +/- 0.230
Testes 0.001 +/- 0.002 0.108 +/- 0.217  ---  ---
Carcass/Pelt 4.232+/- 0.894 8.439 +/- 2.096 4.011 +/- 1.284 7.654 +/- 2.288

a Means were computed from groups of four animals per sex.

b Percent of dose expressed for sample assayed but tissue was only partially sampled (entire tissue not collectable).

RESULTS - PHARMACOKINETIC STUDIES: TOTAL RADIOACTIVITY

Non-Fasted Male and Female Animals

The uptake and disappearance of test substance-derived radioactivity in the plasma was well-described by a triexponential, first-order equation following a percutaneous application of a 150 mg/kg dose to non-fasted animals. The mean pharmacokinetic parameters are summarized in Table 3 below. The absorption of total radioactivity was relatively rapid, with t1/2 values of 1.5 hr for males and 2.4 hr for females. The maximum plasma concentrations (Cmax) were of similar magnitude between the sexes but the time to Cmax, or tmax, was slightly different for males (3.3 hr) and females (5.6 hr). There was also an initial disposition phase (t1/2 -alpha = 2.2 hr, males; 6.3 hr, females) which was consistent with the rapid penetration of skin by radioactivity. A terminal t1/2 value was derived for males (18.7 hr) but a longer 1/2 -beta value for females (31.8 hr) was obtained from RSTRIP analysis for the disappearance of total 14C from the plasma after a 150 mg/kg cutaneous application. The reason for this difference was seen in the variation in the pharmocokinetic parameter values from the individual animal RSTRIP analyses, which was probably related to the variation often seen after continuous contact (72 hr) cutaneous dosing.

Fasted Male and Female Animals

Pharmacokinetic analyses of the data from fasted animals (see Table 3 below) also yielded biexponential time-course plots for absorption and elimination of total 14C in plasma. The RSTRIP analysis revealed similar absorption t1/2 values to those observed for the non-fasted group and Cmax values which were of similar magnitude to the corresponding non-fasted group values. There was a slight difference in tmax values between the sexes (males, 4.1 hr; females, 3.2 hr). The elimination t1/2 values were also similar between males (18.7 hr) and females (17.5 hr).

Table 3. 150 mg/kg Cutaneous 14C-DMEE in Un-Fasted and Fasted Fischer 344 Rats: Summary of Pharmacokinetic parameter Estimates for Total Radioactivity Mean with a Two-Exponential Fit

Non-Fasted Fasted
Parametera Symbol Male Female Male Female
Absorption rate constant (min-1) ka 0.007539 0.004776 0.014742 0.021607
Half-life of absorption (min) t1/2abs 91.937 (1.53 hr) 145.12 (2.42 hr) 47.02 (0.78 hr) 32.08 (0.53 hr)
Initial disposition rate constant (min-1) a 0.005180 0.001821
Initial disposition half-life (min) t1/2a 133.83 (2.23 hr) 380.61 (6.34 hr)
Terminal rate constant (min-1) ß 0.000617 0.000364 0.000618 0.000661
Terminal half-life (min) t1/2ß 1,123.6 (18.73 hr) 1,905.9 (31.76 hr) 1,121.0 (18.68 hr) 1,049.4 (17.50 hr)
Maximal plasma concentration (µg/g) Cmax 6.1851 4.9187 4.8071 5.0048
Time to Cmax (min) tmax 200.67 (3.34 hr) 338.80 (5.65 hr) 245.70 (4.10 hr) 192.16 (3.20 hr)
AUC through 72 hrs. (µg eq/g*min) AUC72 5,739.4 5,843.1 8,279.0 7,946.3
AUC to infinite time (µg eq/g*min) AUC8 6,052.4 6,267.7 8,932.4 8,457.8
Total body clearance (ml/min*kg) Cltotal 24.8 23.9 16.8 17.7
Mean residence time (min) MRT8 1,289.6 (21.49hr) 1,417.0 (23.62 hr) 1,685.1 (28.08 hr) 1,560.2 (26.00 hr)

a Parameters were estimated from plasma concentration data using RSTRIP computer modeling software.

RESULTS - PLASMA PHARMACOKINETICS OF UNCHANGED TEST SUBSTANCE

Non-Fasted Male and Female Animals

Unchanged test substance was very readily absorbed through dorsal rat skin over the 72 -hr exposure period evaluated. Plasma concentrations after the 150 mg/kg percutaneous dose application to non-fasted animals were analyzed up to 30 hr post-dosing for males but only up to 24 hr post-dosing for females (see Parts A and B, Table 4), based on diminishing 14C levels. This dose was rapidly absorbed (t1/2 = 0.3 hr, males; 0.5 hr, females) and the observed absorption rate constants were larger (Table 5) than those for total radioactivity concentrations (Table 3) for both sexes. Maximum plasma concentrations (Cmax) were somewhat lower than those calculated using 14C data and were achieved at tmax values of 2.1 hr for males and 2.4 hr for females, which were similar to tmax values observed for total radioactivity at this dose. The terminal rate constants were larger when compared to the corresponding values for 14C disappearance from plasma, resulting in terminal t1/2 values (10.7 hr, males; 4.7 hr, females - see Table 5) which were much shorter than the half-life values for elimination of 14C (18.7 hr, males; 31.8 hr, females - see Table 3). Overall, the uptake and disappearance of unchanged DMEE paralleled total radioactivity plasma profiles and were consistent with HPLC results for the appearance of unchanged DMEE in the urine.

Fasted Male and Female Animals

The plasma unchanged DMEE levels after a 150 mg/kg dose application to fasted animals were quantifiable to 48 hr for males but only up to 36 hr post-dosing for females (Parts C and D, Table 4). However, the 36- and 48 -hr points for males and the 30- and 36 -hr points for the female data had to be omitted before these data could be fit using RSTRIP. It was concluded that this was due to a second increase in plasma DMEE concentrations which was possibly related to the long (72 hr) contact period for the dose. The absorption phase data points on the plasma curves for unchanged DMEE with either sex were lower in concentration values than their total radioactivity counterparts but closely paralleled the uptake of 14C into the blood (See Table 5). The half-life values for absorption in male animals were similar (0.8 hr, total 14C - see Table 3; 0.7 hr, unchanged DMEE - see Table 5), and the terminal half-life value of 24.9 hr for the unchanged DMEE is similar in magnitude to that for the plasma disappearance of 14C for this treatment group (18.7 hr). For females, the absorption t1/2 value for unchanged DMEE was quite similar to the total 14C t1/2 value (0.45 hr and 0.53 hr, respectively). While the terminal phase t1/2 value (12.4 hr) agreed well with the 14C half-life value (17.5 hr), it is only based on three data points. Consequently, this agreement may only be coincidence.

Table 4. Concentrations of Unchanged DMEE vs. Radioactivity in Plasma of Fischer 344 Rats Following Percutaneous Administration

Collection Interval Plasma DMEEa(µg DMEE/g) RadioactivitybConcentration (µg Equiv./g) Plasma DMEE as Percent of Radioactivity
A. Non-Fasted Males
45 min 1.155 2.492 46.35%
1 hr 2.334 3.373 69.20%
2 hr 2.802 5.481 51.12%
3 hr 3.326 5.603 59.36%
6 hr 2.597 4.754 54.63%
9 hr 2.239 3.932 56.94%
12 hr 1.634 2.488 65.68%
18 hr 1.091 1.538 70.94%
24 hr 0.731 1.094 66.82%
30 hrc 0.581 0.915 63.50%
B. Non-Fasted Females
45 min 1.845 2.052 89.91%
1 hr 3.128 3.696 84.63%
2 hr 4.798 5.900 81.32%
3 hr 5.568 6.264 88.89%
6 hr 3.281 4.880 67.23%
9 hr 2.142 4.355 49.18%
12 hr 1.031 2.987 34.52%
18 hr 1.013 2.341 43.27%
24 hr 2.174 2.878 75.54%
C. Fasted Males
45 min 1.736 1.650 105.21%
1 hr 1.631 2.229 73.17%
2 hr 2.784 4.240 65.66%
3 hr 2.772 4.834 57.34%
6 hrd 3.090 7.707 40.09%
9 hr 2.416 4.420 54.66%
12 hr 1.818 2.927 62.11%
18 hr 2.003 2.539 78.89%
24 hr 2.821 2.923 96.51%
30 hr 1.230 1.655 74.32%
36 hr 2.149 2.459 87.39%
48 hrc 2.463e 3.603 68.36%
D. Fasted Females
45 min 1.564 1.994 78.44%
1 hr 2.086 2.903 71.86%
2 hr 3.167 4.294 73.75%
3 hr 3.372 4.861 69.37%
6 hr 4.275 5.289 80.83%
9 hr 2.945 4.023 73.20%
12 hrd 1.298 4.311 30.11%
18 hr 2.119 2.957 71.66%
24 hr 1.075 1.735 61.96%
30 hr 2.383 2.834 84.09%
36 hrc 2.016 2.779 72.54%

a Equivolume pooled samples from four animals per group unless otherwise noted.

b Expressed as the group mean (n=4) unless otherwise noted.

c Subsequent interval samples were not analyzed due to low radioactivity levels.

d Sample was clotted and eluted slowly from the extraction column, may have lowered recovery of test chemical.

e Value is the mean of two analyses.

Table 5. 150 mg/kg Cutaneous 14 -C-DMEE in Non-Fasted and Fasted Fischer 344 Rats: Summary of Pharmacokinetic Parameter Estimates for Unchanged DMEE in Pooled Plasma Samples with a Two-Exponential Fit

Parametera Symbol Non-Fasted Fasted
Male Female Male Female
Absorption rate constant (min-1) ka 0.040025 0.022480 0.015560 0.025705
Half-life of absorption (min) t1/2abs 17.32 (0.29 hr) 30.83 (0.51 hr) 44.55 (0.74 hr) 26.96 (0.45 hr)
Terminal rate constant (min-1) ß 0.0010804 0.0024752 0.00046467 0.00093542
Terminal half-life (min) t1/2ß 641.58 (10.69 hr) 280.03 (4.67 hr) 1,491.7 (24.86 hr) 741.0 (12.35 hr)
Maximal plasma concentration - Calculated (µg/g) Cmax 3.1396 5.1268 2.7513 3.1788
Maximal plasma concentration - Observed (µg/g) Cmax 3.3260 5.5680 3.0900 4.2750
Time to Cmax (min) tmax 127.37 (2.12 hr) 140.83 (2.35 hr) 230.83 (3.85 hr) 156.80 (2.61 hr)
AUC through 30 hrs. (µg eq/g*min)b AUC30 2,722.1 2,493.7b 3,653.0 2,789.4b
AUC to infinite time (µg eq/g*min) AUC8 3,212.4 2,721.4 6,596.8 3,851.3
Total body clearance (ml/min*kg)c Cltotal 46.7 55.1 22.7 38.9
Mean residence time (min) MRT8 950.59 (15.84 hr) 448.49 (7.47 hr) 2,214.6 (36.91 hr) 1,107.9 (18.46 hr)
Volume of distribution at steady state (L/kg)d Vdss 44.387 24.72 50.356 43.150

a Parameters were estimated from plasma concentration data using RSTRIP computer modeling software.

b AUC0 -24 hr for female data; data at 36 and 48 hr for males and 30 and 36 hr for females were omitted from these fits because they presumably represented points on a second "input" phase of the plasma curves.

c Cltotal = Dose (in ug/kg) + AUC to infinite time (in ug/g*min).

d Vdss = Cltotal * MRT, corrected to L/kg.

RESULTS - PHARMACOKINETIC ANALYSIS OF URINE DATA

Sigma minus analysis results show that the amount of unchanged DMEE excreted into urine over the entire collection period were 15.89 mg for non-fasted males and 11.5 mg for non-fasted females. These amounts represented about 69.6% for males and 65.4% for females of the total number of mg of radioequivalents absorbed and excreted in the urine. The approximate values for the terminal t1/2 which were calculated from this sigma minus plot of the data for beta half-life estimates were 12.22 hr for males and 13.31 hr for females. These values virtually matched the t1/2 values which were obtained from sigma minus evaluation for total radioactivity after a 150 mg/kg percutaneous dose (12.16 hr males; 13.29 hr females). Overall, the estimates obtained for beta from the urine data were in general agreement with the terminal rate constants which could be obtained from the cutaneous plasma curves for total 14C.

Applicant's summary and conclusion

Conclusions:
Interpretation of results (migrated information): no data
Cutaneous application of 14C-test substance penetrated skin rapidly and as a substantial fraction of the applied dose (72-73% of the applied 14C, except fasted females 60%) was excreted predominantly as unchanged test substance in the urine (65-70% of the absorbed dose), for both fasted and non-fasted rats. The absorption half-life values which can be calculated for each of the two 150 mg/kg percutaneous experiments conducted were 0.3 - 0.7 hr for males and 0.4 - 0.5 hr for females. These rapid absorption half-life values are consistent with the fact that as much as 73% of the 14C-DMEE applied was absorbed in the early intervals of the 72 hr of contact. However, the high concentrations of unchanged DMEE in the plasma are quickly cleared and excreted in the urine, indicating that doses within this range will be rapidly eliminated from the plasma and excreted principally in the urine. In addition, the analysis of urine by HPLC indicates that most of the DMEE which penetrates skin is excreted in its unchanged form and that the circulating plasma chemical levels observed following cutaneous exposure are largely the unchanged DMEE.