N-[2-[(2-chloro-4,6-dinitrophenyl)azo]-5-(diallylamino)-4-methoxyphenyl]acetamide

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

key study

Study period:

21-01-1991 to 08-02-1991

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

study well documented, meets generally accepted scientific principles, acceptable for assessment

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

1991

Report date:

1991

Materials and methods

Objective of study:

absorption

distribution

excretion

metabolism

Principles of method if other than guideline:

The test substance was administered to Sprague-Dawley rats via oral gavage in corn oil in order to determine the extent of absorption, distribution,into different tissues and elimination through urine. Urine samples were analysed to determine the levels of the test substance and its primary metabolite.

GLP compliance:

no

Test material

Test material information

Constituent 1

Reference substance name:

2,2'-[[5-acetamido-4-[(2-bromo-4,6-dinitrophenyl)azo]-2-methoxyphenyl]imino]diethyl diacetate

EC Number:

222-813-1

EC Name:

2,2'-[[5-acetamido-4-[(2-bromo-4,6-dinitrophenyl)azo]-2-methoxyphenyl]imino]diethyl diacetate

Cas Number:

3618-72-2

Molecular formula:

C23H25BrN6O10

IUPAC Name:

2-{[2-(acetyloxy)ethyl]({4-[ (E)-2-(2-bromo-4,6-dinitrophenyl)diazen-1-yl]-5acetamido-2-methoxyphenyl})amino}ethyl acetate

Details on test material:

Disperse Blue 79:1 (Br)
CAS No. 3618-72-2

Radiolabelling:

yes

Remarks:

radiochemical purity 99%

Test animals

Species:

rat

Strain:

Sprague-Dawley

Sex:

male/female

Details on test animals or test system and environmental conditions:

TEST ANIMALS
- Source: Charles River Laboratories, Inc., Portage, MI
- Age at study initiation: 7 weeks
- Housing: stainless steel cages
- Diet (e.g. ad libitum): ad libitum
- Water (e.g. ad libitum):ad libitum
- Acclimation period: at least 5 days

ENVIRONMENTAL CONDITIONS
- Photoperiod (hrs dark / hrs light): 12 h light:12 h dark

IN-LIFE DATES: From: 21-01-1991 To: 08-02-1991

Administration / exposure

Route of administration:

oral: gavage

Vehicle:

corn oil

Details on exposure:

PREPARATION OF DOSING SOLUTIONS:

The unlabelled test substance was first dissolved in small volume of acetone and then the radio-labelled test substance was added by volume to it. Corn oil was then weighed in the vial containing both the forms of the test substance. A small stirring bar was placed in the vial and a stream of nitrogen was bubbled into the suspension overnight to evaporate the acetone.

Doses / concentrations

Doses / concentrations 1

Dose / conc.:

50 mg/kg bw/day (actual dose received)

Doses / concentrations 2

Dose / conc.:

500 mg/kg bw/day (actual dose received)

No. of animals per sex per dose / concentration:

four per sex per dose group

Control animals:

no

Details on study design:

The test substance was administered to Sprague-Dawley rats via oral gavage in corn oil in order to determine the extent of absorption, disposition into different tissues and the extent of elimination through urine. In addition, the urine samples were analysed to determine the levels of the test substance and its primary metabolite.

Details on dosing and sampling:

Four rats per sex per dose group, kept individually in Roth-type metabolism cages for acclimation and fasting for 15 h, were administered the test substance dissolved in corn oil at the target concentrations of 50 and 500 mg/kg/day at a dose volume of 4 mL/kg and a target radioactivity of 10-15 µCi.

Urine samples were collected at 6, 12, 24, 48, 72 and 96 h post-dosing under dry ice. Fecal samples were collected at room temperature at same intervals. Room air was trapped through the metabolism cages at approximately 500 mL/min and the expired 14CO2 was trapped at 12, 24, 48, 72 and 96 h post-dosing and stored cold until analysis.Volatile organic metabolites were not collected.

At 96 h post-dosing, the animals were sacrificed by overdose of methoxyflurane and exsanguination. Blood collected at termination was separated into plasma and RBCs for analysis. The metabolism cages were washed with 1% trisodium phosphate and an acetone/water rinse. These wash and rinse samples were analysed for inclusion in the material balance.

The pelt was removed from the carcass, prior to removal of the organs. The liver, kidney, bone marrow from femur, brain, peritoneal fat, heart, lung, urinary bladder, spleen, upper GI tract and its contents, testes, ovaries and uterus were isolated and collected for analysis. The remaining carcass was weighed and stored cold until analysis.

Results and discussion

Main ADME results

Main ADME results 1

Type:

other: Material balance

Results:

The overall recovery of the administered dose was 98+/-2.1% for males and 92.5+/-2.6% for female animals.

Main ADME results 2

Type:

absorption

Results:

The test substance was not absorbed substantially through oral gavage administration.

Main ADME results 3

Type:

distribution

Results:

The test substance was not found to be distributed to the tissues to a major extent following oral administration.

Main ADME results 4

Type:

metabolism

Results:

The test substance was substantially cleared from the GI tract and does not appear to be extensively metabolised following oral administration.

Main ADME results 5

Type:

excretion

Results:

The majority (greater than 85%) of the oral dose of the test substance was excreted in the faeces and less than 6% was recovered in the urine during the 96 h post dose period.

Toxicokinetic / pharmacokinetic studies

Details on absorption:

By 24 h post dosing, approximately 86% (males) and 74% (females) of the administered dose was excreted in the faeces. Additionally, approximately 3% of the dose was recovered in urine in the initial 24 h for either sex. Therefore, it is evident that the test substance was not substantially absorbed when orally administered to rats.

Details on distribution in tissues:

Less than 0.15% (males) and 0.04% (females) of the test substance was detected in the tissues 96 h post administration. Liver was the only tissue that had greater than 0.02% of the administered dose for either sex. Less than 0.01% of the administered dose was recovered from the femur bone marrow. From these results, it is evident that, the test substance was not distributed significantly into different tissues.

Details on excretion:

By 24 h post dosing, approximately 86% (males) and 74% (females) of the administered dose was excreted in the faeces. Additionally, approximately 3% of the dose was recovered in the urine in the initial 24 h for either sex. The majority of the elimination of the radioactivity was virtually complete by 48 h post-dosing. Expired CO2 was less than 0.1% for males and 0.1% for females.

Metabolite characterisation studies

Metabolites identified:

no

Details on metabolites:

Analysis of pooled samples from animals of each sex in each dose group showed an unidentified metabolite which effectively accounted for all (>90%) of the detectable radioactivity in the urine samples. The suspected metabolite BDNA did not appear in any of the urine samples.

Applicant's summary and conclusion

Conclusions:

Under the conditions of the study, the test substance was not substantially absorbed through oral gavage. Majority of the administered dose was excreted through feces and only a minor proportion of the dose was excreted through urine. The test substance was not distributed to different tissues and most of the test substance was eliminated unchanged.

Executive summary:

A study was conducted to determine the toxicokinetic behaviour of the test substance in rats according to an internal protocol of the test facility. The test substance (dissolved in corn oil) was administered to groups of fasted rats at 50 and 500 mg/kg/day through oral gavage at a dose volume of 4 mL/kg and a target radioactivity of 10-15 µCi. Urine and faecal samples were collected at regular intervals until 96 h post dosing. At 96 h post-dosing, the animals were sacrificed by overdose of methoxyflurane and exsanguination. Blood collected at termination was separated into plasma and RBCs for analysis. The liver, kidney, bone marrow from femur, brain, peritoneal fat, heart, lung, urinary bladder, spleen, upper GI tract and its contents, testes, ovaries and uterus were isolated and collected for analysis. The remaining carcass was weighed and stored cold until analysis. By 24 h post dosing, approximately 86% (males) and 74% (females) of the administered dose was excreted in the faeces. Additionally, approximately 3% of the dose was recovered in the urine in the initial 24 h for either sex. The majority of the elimination of the radioactivity was virtually complete by 48 h post-dosing. Expired CO2 was less than 0.1% for males and 0.1% for females. Less than 0.15% (males) and 0.04% (females) were detected in the tissues 96 h post administration. Liver was the only tissue that had greater than 0.02% of the administered dose for either sex. Less than 0.01% of the administered dose was recovered from the femur bone marrow. Analysis of pooled samples from animals of each sex in each dose group showed an unidentified metabolite which effectively accounted for all (>90%) of the detectable radioactivity in the urine samples. The suspected metabolite BDNA did not appear in any of the urine samples. Under the conditions of the study, the test substance was not substantially absorbed through oral gavage. The majority of the administered dose was excreted through feces and only a minor proportion of the dose was excreted through urine. The test substance was not distributed to different tissues and most of the test substance was eliminated unchanged (Frantz, 1991).