cis-4-[3-(p-tert-butylphenyl)-2-methylpropyl]-2,6-dimethylmorpholine

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

27.10.2003 - 23.07.2005

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Cross-reference

Reason / purpose for cross-reference:

reference to same study

Reference

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

27.01.2004 - 16.08.2005

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Reason / purpose for cross-reference:

reference to same study

Objective of study:

absorption

distribution

excretion

toxicokinetics

Qualifier:

according to guideline

Guideline:

OECD Guideline 417 (Toxicokinetics)

Version / remarks:

4.4.1984

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Radiolabelling:

yes

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: RCC Ltd, Füllinsdorf, Schweiz (HanBrl:WIST (SPF) rats), Charles River Laboratories, Sulzfeld (Germany) (Crl:WI (Han) rats - used for tissue distribution experiments)
- Age at study initiation: at least 9 weeks
- Weight at study initiation: not specified
- Housing: individually during the experiments
- Diet ad libitum: "Maus, Ratte, Haltung GLP" either pelleted (e.g. during
acclimatization and in steel wire mesh cages) or granulated (e.g. in metabolism cages)
- Water (ad libitum): tap water
- Acclimation period: at least 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24
- Humidity (%): 30 - 70
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): not specified

IN-LIFE DATES: not specified

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Remarks:

(0.5% CMC in water)

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
- respective amounts of non-radiolabelled material were added to an aliquot of the stock solution of the radiolabelled material and the mixture was evaporated to dryness
- the mixture was added to the respective volume of 0.5 % CMC in water and was filled up to the final volume
- prior to administration the preparation was sonicated and stirred to produce a homogeneous preparation
- for experiments on the biliary excretion, 12C-test substance was mixed with 13C-test substance in a 1:1 ratio
- quantity of radioactivity per animal: about 0.5 - 2 MBq

Duration and frequency of treatment / exposure:

one single administration (treatment with radiolabelled test substance) or one daily treatment for 14 days (treatment with non-radiolabelled test substance)

Dose / conc.:

1 mg/kg bw (total dose)

Dose / conc.:

15 mg/kg bw (total dose)

No. of animals per sex per dose / concentration:

- Balance/excretion - experiments: 4/sex/treatment group (only high dose tested);
- Blood/Plasma level - experiments: 4/sex/treatment group;
- Tissue distribution - experiments: 12/sex/treatment group;
- Excretion via bile - experiments: 4/sex/treatment group

Control animals:

no

Positive control reference chemical:

No

Details on study design:

- Dose selection rationale: according to the relevant guideline; doses were selected based on the results from previously performed studies on the chronic toxicity of the test substance in rats
- Rationale for animal assignment (if not random): Animals of similar age were ordered sequentially in portions prior to the experiments. Therefore the conventional randomization and assignment to groups was not possible.

Details on dosing and sampling:

TOXICOKINETIC / PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: urine, faeces, blood, plasma, tissues (heart, liver, spleen, bone, skin, lung, ovaries, carcass, muscle, kidney, brain, pancreas, uterus, testes, adipose tissue, stomach & stomach contents, thyroid gland, adrenal glands, gut and gut contents, bone marrow), cage washes, bile
- Time and frequency of sampling:
Balance/Excretion - experiments: Excreta were collected after 6, 12 and 24 h and subsequently in time intervals of usually 24 h for up to 120 h. After 120 h, animals were sacrificed for tissue sampling.

Blood/Plasma level - experiments: Blood samples were taken 1, 2, 4, 8, 24, 48, 72, 96, and 120 h after administration.

Tissue distribution - experiments: Each 3 animals per sex were killed at 4 different time points after dosing (corresponding to the time points of maximum plasma concentration MPC, 1/2 MPC (or alternatively the time when a second maximum occurred), 1/4 MPC and 1/8 MPC) for tissue sampling.

Excretion via bile - experiments: Bile was collected at time intervals of 3 hours for up to 48 hours depending on the health state of the animals and the bile flow.

Statistics:

not specified

Type:

other: excretion (bile); dose 1 mg/kg bw

Results:

phenyl-label: 34.73 (males) and 20.35% (females) within 48 h; morpholine-label: 38.94 (males) and 57.41% (females) within 48 h

Type:

other: excretion (bile); dose 15 mg/kg bw

Results:

phenyl-label: 64.21 (males) and 60.99% (females) within 48 h; morpholine-label: 46.70 (males) and 42.57% (females) within 48 h

Type:

other: excretion (feces); dose 1 mg/kg bw

Results:

31.40 (males) and 34.74% (females) after 120 h

Type:

other: excretion (urine); dose 1 mg/kg bw

Results:

54.22 (males) and 49.32% (females) within 120 h

Type:

other: excretion (feces); dose 15 mg/kg bw

Results:

phenyl-label: 35.89 (males) and 33.52% (females) within 120 h; morpholine-label: 41.52 (males) and 35.33% (females) within 120 h

Type:

other: excretion (urine); dose 15 mg/kg bw

Results:

phenyl-label: 52.59 (males) and 48.44% (females) after 120 h; morpholine-label: 47.88 (males) and 45.51% (females) after 120 h

Type:

other: excretion (exhaled air)

Results:

morpholine-label: 8.88% (males) and 9.28% (females); phenyl-label: <2%

Details on distribution in tissues:

- Tissue radioactivity levels in both sexes were in the same range at the respective time points and dose levels.
- The pattern of distribution in the various organs and tissues was also similar.
- Tissue radioactivity concentrations generally declined with time parallel to plasma concentrations.
- Throughout the time course of the experiments, highest radioactivity concentrations were found in the GI tract, liver, kidney, pancreas and adrenal glands whereas radioactivity levels were lowest in brain, bone and muscle.

Details on excretion:

The time course of the amount of radioactivity found in urine, feces and CO2 indicates very rapid excretion.

- There was no sex- and dose difference with respect to the excretion pattern at both dose levels and radiolabels.

Toxicokinetic parameters:

AUC: 326.28 µg Eq*h/g

Remarks:

females; dose: 15 mg/kg bw

Toxicokinetic parameters:

AUC: 349.79 µg Eq*h/g

Remarks:

males; dose: 15 mg/kg bw

Toxicokinetic parameters:

AUC: 19.57 µg Eq*h/g

Remarks:

females; dose: 1 mg/kg bw

Toxicokinetic parameters:

AUC: 18.68 µg Eq*h/g

Remarks:

males; dose: 1 mg/kg bw

Toxicokinetic parameters:

half-life 1st: 21.39 h

Remarks:

females; dose: 15 mg/kg bw

Toxicokinetic parameters:

half-life 1st: 13.99 h

Remarks:

males; dose: 15 mg/kg bw

Toxicokinetic parameters:

half-life 1st: 13.51 h

Remarks:

females; dose: 1 mg/kg bw

Toxicokinetic parameters:

half-life 1st: 3.57 h

Remarks:

males; dose: 1 mg/kg bw

Toxicokinetic parameters:

half-life 2nd: 49.73 h

Remarks:

females; dose: 1 mg/kg bw

Toxicokinetic parameters:

half-life 2nd: 64.37 h

Remarks:

males; dose: 1 mg/kg bw

Toxicokinetic parameters:

half-life 3rd: 34.84 h

Remarks:

females; dose: 15 mg/kg bw

Toxicokinetic parameters:

half-life 3rd: 40.88 h

Remarks:

males; dose: 15 mg/kg bw

Toxicokinetic parameters:

half-life 3rd: 29.66 h

Remarks:

females; dose: 1 mg/kg bw

Toxicokinetic parameters:

half-life 3rd: 22.75 h

Remarks:

males; dose: 1 mg/kg bw

Toxicokinetic parameters:

Tmax: 1 and 8 h

Remarks:

both sexes; dose: 15 mg/kg bw

Toxicokinetic parameters:

Tmax: 1 h

Remarks:

females; dose: 1 mg/kg bw

Toxicokinetic parameters:

Tmax: 4 h

Remarks:

males; dose: 1 mg/kg bw

Toxicokinetic parameters:

Cmax: 7.84 and 7.92 µg Eq/g

Remarks:

females; dose: 15 mg/kg bw

Toxicokinetic parameters:

Cmax: 9.42 and 11.32 µg Eq/g

Remarks:

males; dose: 15 mg/kg bw

Toxicokinetic parameters:

Cmax: 0.41 µg Eq/g

Remarks:

females; dose: 1 mg/kg bw

Toxicokinetic parameters:

Cmax: 0.63 µg Eq/g

Remarks:

males; dose: 1 mg/kg bw

- The second maximum observed is assessed to be a consequence of an enterohepatic circulation.

- During the complete experimental period post dosing (120 hours), lower concentrations of radioactivity were generally found in blood at both dose levels indicating that major parts of the radioactivity were in plasma and not bound to cellular blood constituents.

- After a single oral administration of 15 mg/kg bw, mean total recoveries of radioactivity were 94.66 and 93.45% in males and 95.17 and 94.80% in females for the phenyl- and the morpholine-labelled test substance, respectively. After single oral administration of 1 mg/kg bw phenyl-labelled test substance, mean total recoveries of radioactivity were 90.07% in males and 89.90% in females.

 

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2006

Report date:

2006

Materials and methods

Objective of study:

metabolism

Test guideline

Qualifier:

according to guideline

Guideline:

OECD Guideline 417 (Toxicokinetics)

Version / remarks:

not specified

Deviations:

no

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

cis-4-[3-(p-tert-butylphenyl)-2-methylpropyl]-2,6-dimethylmorpholine

EC Number:

266-719-9

EC Name:

cis-4-[3-(p-tert-butylphenyl)-2-methylpropyl]-2,6-dimethylmorpholine

Cas Number:

67564-91-4

Molecular formula:

C20H33NO

IUPAC Name:

(2R,6S)-4-[3-(4-tert-butylphenyl)-2-methylpropyl]-2,6-dimethylmorpholine

Radiolabelling:

yes

Test animals

Species:

rat

Strain:

Wistar

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River, Sulzfeld
- Age at study initiation: at least 9 weeks
- Weight at study initiation: not specified
- Housing: individually during the experiments
- Diet ad libitum: "Maus, Ratte, Haltung GLP" either pelleted (e.g. during acclimatization and in steel wire mesh cages) or granulated (e.g. in metabolism cages)
- Water (ad libitum): tap water
- Acclimation period: at least 6 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20 - 24
- Humidity (%): 30 - 70
- Air changes (per hr): not specified
- Photoperiod (hrs dark / hrs light): not specified

IN-LIFE DATES: not specified

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

CMC (carboxymethyl cellulose)

Remarks:

(0.5% CMC in water)

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:
- respective amounts of non-radiolabelled material were added to an aliquot of the stock solution of the radiolabelled material and the mixture was evaporated to dryness
- the mixture was added to the respective volume of 0.5 % CMC in water and was filled up to the final volume
- prior to administration the preparation was sonicated and stirred to produce a homogeneous preparation
- for experiments on the biliary excretion, 12C-test substance was mixed with 13C-test substance in a 1:1 ratio
- quantity of radioactivity per animal: about 0.5 - 2 MBq
- about 1 ml of the test item preparation/100 g bw were applied

Duration and frequency of treatment / exposure:

see Table 1 in section "Any other information on materials and methods incl. tables"

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

1 mg/kg bw (total dose)

Doses / concentrations 2

Dose / conc.:

15 mg/kg bw (total dose)

Doses / concentrations 3

Dose / conc.:

100 mg/kg bw (total dose)

No. of animals per sex per dose / concentration:

see Table 1 in section "Any other information on materials and methods incl. tables"

Control animals:

no

Positive control reference chemical:

no

Details on study design:

- Dose selection rationale: according to the relevant guideline; doses were selected based on the results from previously performed studies on the chronic toxicity of the test substance in rats
- Rationale for animal assignment (if not random): Animals of similar age were ordered sequentially in portions prior to the experiments. Therefore the conventional randomization and assignment to groups was not possible.

Details on dosing and sampling:

METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled: urine, feces, tissues (liver, kidney, fat), plasma, bile
- Time and frequency of sampling:
urine and feces: after 6, 12, 24 h and subsequently in time intervals of usually 24 h for up to 168 h;
bile: 0 - 48 h post dose in sampling periods of 3 h (0 - 3 h, 3 - 6 h, 6 - 9 h, ...);
tissues and plasma: 2 and 8 h post dose

- From how many animals: (samples pooled)
urine and feces: 4/sex/treatment group (dosed with 1 and 15 mg/kg bw), 10/sex/treatment group (dosed with 100 mg/kg bw);
bile: 4/sex/treatment group;
tissues and plasma: 4/sex/treatment group

- Method type(s) for identification: Liquid scintillation counting (LSC), HPLC, LC/MS
- Limits of detection and quantification: LSC - approx. 50 dpm; HPLC - different limits for an individual metabolite in each chromatogram

Statistics:

not specified

Results and discussion

Metabolite characterisation studies

Metabolites identified:

yes

Details on metabolites:

- In urine: 3 main metabolites: metabolite 1 (2.7 - 5.8% of the administered dose) originated from oxidation of the side chain of the parent compound, the other 2 metabolites (sum: 7.6 - 27.9% of the administered dose) from further hydroxylation and cleavage of the morpholine-ring-system; additionally a sulfoxylated product was identified (0.6 - 1.1% of the dose in males and 1.6 - 3.8% in females); the amount of the parent compound was
within the range of 2.25 - 6.37 % of the dose;
- In feces: 8 main metabolites: 4 metabolites (sum: 10 - 24% of the dose) were constituted by hydroxylation, oxidation, sulfoxylation of the side chains and of the phenyl- and morpholine-ring-systems, combined with cleavage of the morpholine ring in the end; additionally the sulfate of one metabolite was identified (1.3 - 8.2% of the dose in females and 1.1% of the dose in males); the amount of the parent compound was within the range of 1.8 - 7.2% of the dose with higher amounts at the high dose levels;
- In bile: 6 main metabolites (sum: 10 - 37% of the dose): 2 metabolites were formed by conjugation with glucoronic acid, 1 metabolite derived from another metabolite by shortening of the side chain (demethylation), 2 of the metabolites were only found in bile; the sulfate of one metabolite was identified (3.2 - 9.8% of the dose in females and 0.6 - 0.8% of the dose in males); the parent compound was found in the range of 1.0 - 1.1% of the dose;
- Some of these metabolites were also detected in liver, kidney, plasma and fat