2,2,6,6-tetramethyl-4-oxopiperidinooxy

Basic toxicokinetics

Administrative data

Endpoint:

basic toxicokinetics in vivo

Type of information:

experimental study

Adequacy of study:

weight of evidence

Reliability:

2 (reliable with restrictions)

Rationale for reliability incl. deficiencies:

other: Acceptable, well documented publication report which meets basic scientific principles.

Data source

Reference

Reference Type:

publication

Title:

Metabolism of the stable nitroxyl radical 4-oxo-2,2,6,6-tetramethylpiperidine-N-oxyl (TEMPONE)

Author:

Kroll. C, Borchert. H.H

Year:

1998

Bibliographic source:

European Journal of Pharmaceutical Sciences, 8 (1999) 5-9

Materials and methods

Objective of study:

metabolism

Test guideline

Qualifier:

no guideline followed

Principles of method if other than guideline:

Perfused rat liver was flushed with a solution of the test substance. The composition of the test solution was monitored with time over a period of 3 hours.

GLP compliance:

not specified

Test material

Test material information

Constituent 1

Reference substance name:

4-oxo-2,2,6,6,-tetramethylpiperidine-N-oxyl

IUPAC Name:

4-oxo-2,2,6,6,-tetramethylpiperidine-N-oxyl

Constituent 2

Reference substance name:

TEMPONE

IUPAC Name:

TEMPONE

Test material form:

not specified

Details on test material:

- Name of test material (as cited in study report): TEMPONE (4-oxo-2,2,6,6-tetramethylpiperidine-N-oxyl)
-Other: Supplied by Sigma

Radiolabelling:

no

Test animals

Species:

rat

Strain:

Wistar

Sex:

male

Administration / exposure

Duration and frequency of treatment / exposure:

3 hours

Doses / concentrations

Remarks:

Doses / Concentrations:
Nominal: 1 mmol/L in 100 mls Dulbeccos buffer
Measured : 195 and 190 micrograms /ml

No. of animals per sex per dose / concentration:

Not specified

Control animals:

not specified

Results and discussion

Main ADME results

Type:

metabolism

Results:

t ½ = 47 -70 minutes,refer to results section for details of metabolites.

Metabolite characterisation studies

Metabolites identified:

yes

Details on metabolites:

Rapid decay of TEMPONE was observed during the liver perfusion. After 3 hours, 29.7% of the initially applied TEMPONE was detected. Five ‘new’ peaks were detected in the gas chromatogram, when compared to a blank sample.
After 3 hours, 80.5 µg/ml TEMPOL (4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl) and 43.4 µg/ml 1-hydroxy-TEMPOL (1,4-dihydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl), equivalent to 41.3 and 22.3% of the initial applied TEMPONE were detected. These form the main products of hepatic biotransformation of TEMPONE.
Two further metabolic reactions occur: keto reduction in the 4-position, resulting in the hydroxyl adduct at the 4-position and hydroxylamine formation in the 1-position.
The remaining two previously unknown peaks were identified as the secondary amines of the TEMPONE and TEMPOL hydroxylamines.
Please refer to the attached metabolic pathway for details, fig 1.
The exclusion of Cytochrome P450 demonstrated no detectable effect on the hepatic metabolism of TEMPONE.
Results generated from GC chromatography gave t ½ = 70 minutes. Results generated using ESR determined the half life to be t ½ = 47 minutes. The difference was thought to be due to partial re-oxidation of the formed hydroxylamines due to increasing the concentration of the sample for analysis.
Experiments using TEMPOL at a similar concentration to TEMPONE (190-195 µg/ml) showed rapid formation of the 1-hydroxy derivative and no re-oxidation of the 4-hydroxy TEMPOL to the 4-oxo derivative TEMPONE.

Applicant's summary and conclusion

Conclusions:

Interpretation of results: low bioaccumulation potential based on study results
TEMPONE has been shown to be rapidly metabolised in perfused rat liver, the major metabolites being identified as TEMPOL (4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl) , 1-hydroxy-TEMPOL (1,4-dihydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl). 4-oxo-1-hydroxy-2,2,6,6-tetramethylpiperidine-N-pxyl was also identified as a metabolite, the secondary amine derivatives of both TEMPONE and TEMPOL were identified as being present in the analysis.
No re-oxidation of the hydroxyl group in TEMPOL was observed.

Executive summary:

The formation of the metabolites of the stable free radical 4-oxo-2,2,6,6-tetramethylpipreridine-N-oxyl (TEMPONE) was examined in perfused rat livers. The 4-hydroxy derivative (TEMPOL) and it’s 1,4-dihydroxy derivative were found to be the major metabolites, in addition to the corresponding hydroxylamine derivative of TEMPONE.

No re-oxidation of the hydroxyl group in the 4-position was observed. The secondary amines of the nitroxides were detected during the analysis, but the mechanism of their formation remains speculative.

Analysis of the perfusion solution was performed by GC and GC-MS and also ESR, data from both methods of analysis showed the half –life of TEMPONE in perfused rat livers to be t_½= 47 -70 minutes.