Registration Dossier

Diss Factsheets

Administrative data

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
2 (reliable with restrictions)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Data source

Reference
Reference Type:
publication
Title:
Metabolism of vinyltoluene in the rat: effect of induction and inhibition of the cytochrome P-450
Author:
Heinonen THH
Year:
1984
Bibliographic source:
Biochem. Pharmacol. 33(10):1585-1593

Materials and methods

Objective of study:
metabolism
Principles of method if other than guideline:
Metabolism of the test substance in rats was studied by investigating urinary metabolites after injection of different doses. The urinary metabolites analysed were: thioesters, p-methylmandelic acid, p-methylphenylglyoxylic acid, the glycine conjugates of p-methylbenzoic acid, p-methylphenylacetic acid and p-vinylbenzoic acid. The role of cytochrome P-450 in the formation of the metabolites was studied by inhibiting its catalytic activity.
GLP compliance:
no

Test material

Constituent 1
Chemical structure
Reference substance name:
Vinyltoluene
EC Number:
246-562-2
EC Name:
Vinyltoluene
Cas Number:
25013-15-4
Molecular formula:
C9H10
IUPAC Name:
1-methyl-2-vinylbenzene
Test material form:
liquid
Details on test material:
- Mix of isomers: 65-71% s-isomer, 32-35% p-isomer
Radiolabelling:
no

Test animals

Species:
rat
Strain:
Wistar
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Weight at study initiation: 350-400 g
- Diet (e.g. ad libitum): commercial pellet diet from Astra-Ewos
- Water (e.g. ad libitum): tap water

Administration / exposure

Route of administration:
intraperitoneal
Vehicle:
olive oil
Duration and frequency of treatment / exposure:
Single intraperitoneal dose
Doses / concentrationsopen allclose all
Dose / conc.:
50 mg/kg bw/day (actual dose received)
Remarks:
First experiment
Dose / conc.:
250 mg/kg bw/day (actual dose received)
Remarks:
First experiment
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Remarks:
First experiment
Dose / conc.:
1 000 mg/kg bw/day (actual dose received)
Remarks:
First experiment
Dose / conc.:
500 mg/kg bw/day (actual dose received)
Remarks:
Second and third experiments
Control animals:
yes, concurrent vehicle

Results and discussion

Metabolite characterisation studies

Metabolites identified:
yes
Details on metabolites:
After rats received a single intraperitoneal injection of 50 mg/kg, 55% of the dose was found as urinary metabolites, mainly in the first 6 h; at higher doses, slightly smaller percentages were found (Heinonen, 1984). The principal urinary metabolites were thioethers (25%), p-methylmandelic acid (5.7%), p-methylphenylglyoxylic acid (11.9%), p-methylbenzoyl glycine (9.3%), p-methylphenylacetyl glycine (2.5%), and p-vinylbenzoyl glycine (1%). The excretion of these metabolites was prevented by pretreatment with an inhibitor of the cytochrome P450 monoxygenases. Further, the test substance was found to bind to hepatic cytochrome P450, and the reduced glutathione content of the liver and kidney was decreased in rats after a single intraperitoneal injection (Heinonen and Vainio, 1980). These findings suggest that metabolism of the test substance is catalyzed by cytochrome P450, producing vinyl toluene-7,8-oxide as the main reactive intermediate, with subsequent conjugation to glutathione or hydration to diols (Heinonen, 1984).

Applicant's summary and conclusion

Conclusions:
Under the study conditions, following intraperitoneal injection to the rat, metabolism is catalysed by cytochrome P450 producing vinyl toluene-7,8-oxide as the main reactive intermediate, with subsequent conjugation to glutathione or hydration to diols. The main route of excretion of metabolites was via the urine (55% within first 6 h).
Executive summary:

A study was conducted to determine the metabolism of the test substance in rats by investigating urinary metabolites after injection of different doses. Male Wistar rats received the test substance by single intraperitoneal injection in three different experiments. In the first, the test substance dissolved in olive oil was administered at 50, 250, 500 and 1000 mg/kg bw. Four rats were sacrificed after 12 h at each dose level and three rats at the dose levels of 50, 250 or 500 mg/kg bw after 23 h. In the second experiment, rats were given the test substance (500 mg/kg bw in olive oil), 1-phenylimidazole (50 mg/kg bw in DMSO) or both. The control rats received olive oil or DMSO alone. 1-Phenylimidazole and DMSO were administered 1.5 h before the test substance. Four rats of each group were sacrificed 12 h after injection of the test substance and 3 in each group after 23 h. In the third experiment, rats were exposed to PCBs (500 mg/kg bw in olive oil), test substance (500 mg/kg bw in olive oil) or both Controls received olive oil only. The dose of PCB was given 5 d before the test substance. Three animals in each group were sacrificed 23 h after injection of the test substance. Urine was collected during the exposure from all rats in all groups. The urinary metabolites analysed were: thioesters, p-methylmandelic acid, p-methylphenylglyoxylic acid, the glycine conjugates of p-methylbenzoic acid, p-methylphenylacetic acid and p-vinylbenzoic acid. The role of cytochrome P-450 in the formation of the metabolites was studied by inhibiting its catalytic activity.The highest excretion rate was obtained with doses of 50, 250 and 500 mg/kg bw already within the first 6 h. However, the dose of 500 mg/kg bw did not increase the excretion rates of these metabolites compared to the dose of 250 mg/kg bw, suggesting that the metabolic pathways begin to be saturated with the amount of 250 mg/kg bw. At 50 mg/kg bw, 55% of the dose was detected as urinary metabolites within 23 hr, mainly within the first 6 h. The amounts of the excreted metabolites expressed as % of injected dose (250 or 500 mg/kg bw) were lower than that caused by 50 mg/kg bw, and a noticeable amount of the total sums were excreted within 11–23 h, suggesting that the excretion was still continued with the doses of 250 and 500 mg/kg bw 23 h after injection. The excretion of all analyzed metabolites was prevented by the pre-treatment of the rats with 1-phenylimidazole, an inhibitor of cytochrome P-450 monooxygenases. This indicates that these metabolites were formed as catalyzed by cytochrome P-450. The structures of the analyzed metabolites suggest that the main reactive intermediate of the test substance is vinyltoluene-7,8-oxide. Furthermore, the amounts of the excreted metabolites showed that the main detoxification pathways of vinyltoluene-7,8-oxide were the conjugation with reduced glutathione and hydration to diols. Pre-treatment of the rats with PCBs increased the excretion rates of the metabolites. However, the PCB pre-treated rats excreted less thioethers (62%) compared to the rats treated only with the same amount of test substance, whereas the total sum of the other metabolites was about the same in these both groups. This result suggests that PCBs change the metabolism of the test substance to some other pathway which could be glucuronide conjugation because PCBs increased the activity of UDP-glucuronosyltransferase in a dose-dependent manner (Heinonen, 1984).