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Reference
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:
other: Well documented publication which meets basic scientific principles
Objective of study:
absorption
Qualifier:
no guideline followed
Principles of method if other than guideline:
The oral absorption of a mixture of N-methyl-2-pyrrolidinone and 2-pyrrolidinone has been investigated in rats in vivo.
GLP compliance:
no
Radiolabelling:
yes
Remarks:
2-[14C]pyrrolidone
Species:
rat
Strain:
Sprague-Dawley
Sex:
male/female
Details on test animals and environmental conditions:
TEST ANIMALS
- Source: Charles River breeding Laboratories (Portage, MI, USA)
- Age at study initiation:
- Weight at study initiation: 192-239 g
- Fasting period before study:
- Housing: individual
- Individual metabolism cages: yes
- Diet: LAD1 Bioscure Ltd., Manea, UK ad libitum
- Water: ad libitum


ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20-24
- Humidity (%): 30-70
- Air changes (per hr): 15
- Photoperiod (hrs dark / hrs light): 12/12
Route of administration:
oral: gavage
Vehicle:
other: water for oral adminstration and isopropanol for dermal administration
Details on exposure:
PREPARATION OF DOSING SOLUTIONS:
[14C]NMP / [14C]2-P mixure (22.5 mg / 15 mg) was prepared as an aqueous mixture (0.6 ml);

These doses were equivalent to about 112 mg NMP and 75 mg 2-P /kg bw

Duration and frequency of treatment / exposure:
single exposure, up to 120 hrs
Remarks:
Doses / Concentrations:
ca. 112 mg NMP and 75 mg 2-P /kg bw
No. of animals per sex per dose:
33 males and 33 females in total; 3 per sex and time-point
Control animals:
yes, concurrent vehicle
Positive control:
not done
Details on study design:
- Dose selection rationale: no data
Details on dosing and sampling:
PHARMACOKINETIC STUDY (Absorption, distribution, excretion)
- Tissues and body fluids sampled: urine, faeces, blood, plasma, serum, cage washes, bile, CO2
- Time and frequency of sampling: expired air: 24h intervals; other parameters at termination of animals

Sample collection:
Following blood withdrawal by cardiac puncture under halothane anaesthesia, the rats were killed by cervical dislocation in groups of six (three males, three females) per time point at selected times after dosing. Plasma was separated from the blood cells by centrifugation, and stored at about -20°C until analysed. Urine and faeces were collected at suitable intervals from the six rats in each group killed at the final killing time (i.e. 120 hr), and any [14C]CO2 present in the expired air was trapped in a mixture of ethanolamine-2-ethoxyethanol (1:3, v/v) at 24-hr intervals. After these rats were killed, the metabolism cages were washed thoroughly with water, and the washings, excreta and carcasses stored at about -20°C until analysed.

Statistics:
data not available
Preliminary studies:
not applicable
Details on absorption:
Absorption and distribution:
After oral administration, the concentrations of radioactivity were greatest in plasma of male and female rats at 2 hrs after dosing. (mean values 0.463% and 0.562% dose/ml). Radioactivity declined thereafter in a muliphasic manner with a terminal half-life of 29 and 27 hrs for male and female rats. the concentrations were still measureable 5 days after dosing when the mean values corresponded to 0.0006% of the total dose in both sexes.

Quantitative analysis of of plasma by HPLC showed, that at 30 min after dosing the proportions of unchanged NMP and 2-P reflected the 3: 2 ratio, demonstrating little metabolism at this time point. Both parent compounds together accounted for at least 80% of plasma radioactivity until 8 hrs post-administration indicating little first-pass metabolism.
Details on distribution in tissues:
not measured
Details on excretion:
Metabolism and excretion:
Biotransformation became more evident 8 hours post-administration and by 12 hrs virtually all of the plasma radioactivity was found in form of unknown polar metabolites. Plasma levels of the unchanged pyrrolidinones were not measurable during the terminal phase of plasma radioactivity disposition after oral dosing.
Following oral co-administration to male and female rats, 94.8% and 94.5% of the radioactive dose were excreted within 5 days. Radioactivity was mainly excreted through the urine (85-88% of the total dose) mostly within 24 hours. Faecal excretion was of minor importance representing only 1-2% of the total dose. 6-7% of the oral dose was eliminated as CO2 in the expired air, which indicates the extent of metabolic degradation of the pyrrolidinone ring. Only about 1% of the total dose remained in the carcass, demonstrating that the excretion was virtually complete by the end of 5 days post-exposure. Recovery of radioactivity in excreta is presented in Table 2 in "Remarks on results".
Test no.:
#1
Toxicokinetic parameters:
Tmax: 2h after oral route of administration
Test no.:
#2
Toxicokinetic parameters:
Tmax: 6 and 2 h for males and females, respectively after dermal administration
Test no.:
#1
Toxicokinetic parameters:
Cmax: 0.171 and 0.215 % dose/ml for m/f respectively after oral administration
Test no.:
#2
Toxicokinetic parameters:
Cmax: 0.052 and 0.093% for m/frespectively after dermal administration
Test no.:
#1
Toxicokinetic parameters:
AUC: 1.07 and 1.33 % dose/ml.h after oral administration
Test no.:
#2
Toxicokinetic parameters:
AUC: 0.31 and 0.59 % dose/ml.h after dermal administration
Metabolites identified:
not measured
Details on metabolites:
12 hours following oral administration little or none of the intact pyrrolidinones were found in plasma and virtually all of the radioactivity was associated with unknown polar metabolites.

The only results of measured radioactivity of 2 -pyrrolidone in plasma are presented in "Toxicokinetic parameters". All pharmakokinetic parameters are presented in Table 1 atttached in "Attached background material".

Table 2: Excretion balance of radioactivity following co-administration of 14C -NMP and 2 -P to rats

Oral

 

Dermal

Sample

Time (hr)

M

F

M

F

Urine

0-6

16.0

21.4

5.5

11.0

6-24

65.2

60.9

45.3

46.7

24-72

2.9

4.2

8.1

8.8

72-120*

1.1

1.3

2.4

3.3

 

Total

85.2

87.8

61.3

69.8

Faeces

0-24

1.2

0.6

0.7

0.5

24-72

0.8

0.5

0.8

0.6

72-120

0.1

0.1

0.1

0.2

 

Total

2.1

1.2

1.6

1.3

Expired air

0-24

6.8

4.8

3.5

4.2

24-72

0.6

0.6

2.0

2.0

72-120

0.1

0.1

0.5

0.5

 

Total

7.5

5.5

6.0

6.7

Total excretion

0-120

94.8

94.5

68.9

77.8

Carcass

120

1.1

1.2

1.0

1.1

Dose dressing

120

10.7

11.3

Dose-site wash

120

1.2

0.3

Treated skin

120

11.2

2.0

Untreated skin

120

ND

<0.1

Total recovery

0-120

95.9

95.7

93.0

92.5

Conclusions:
Interpretation of results (migrated information): no bioaccumulation potential based on study results
2-pyrrolidone is considered to be fully excreted within 5 days.
Executive summary:

The percutaneous absorption has been investigated in rats of a mixture (3:2, w/w) of N-methyl-2-pyrrolidinone (NMP) and 2-pyrrolidinone (2-P). Co-administration of the two 14 C-radiolabelled compounds was performed by the dermal and oral routes.

Radioactivity was excreted predominantly through the urine after either route of administration, and comparison of the respective excretion profiles indicated that about three-quarters of the applied dose was absorbed through the skin. NMP appeared to be absorbed through the skin more extensively and at a slightly faster rate than 2-P; total percutaneous absorption tended to be more extensive in female than in male rats.

Only about 1% of the dose was retained in the carcasses, demonstrating that the excretion of radioactivity was virtually complete by the end of the 5-day collection period.

Description of key information

Key value for chemical safety assessment

Bioaccumulation potential:
no bioaccumulation potential
Absorption rate - oral (%):
100
Absorption rate - dermal (%):
100
Absorption rate - inhalation (%):
100

Additional information

2-pyrrolidone is the lactam of gamma-aminobutyric acid. It is widely used in industrial solvents. 2-pyrrolidone is a normal component of certain food items. The natural content of 2-pyrrolidone in plant foodstuffs was reported to be in the range of 0.1-2.2 mg/kg (e.g. prunes, orange juice, tomatoes) and up to 20 mg/kg in certain processed food.

Absorption

Bandle et al. found that 2-pyrrolidone is an endogenously occurring molecule (Bandle et al.,1984). 2-pyrrolidone is readily absorbed via the intestinal tract and in addition percutaneously (Midgley et al., 1992). In this study rats received two radiolabeled compounds N-methyl-2-pyrrolidone (NMP) and 2-pyrrolidinone orally or percutaneously (3:2). About 77.5% of the percutaneous applied dose was absorbed through the skin (male = 73%, female = 82%; mean = 77.5%). Plasma concentrations remained relatively uniform from 1 to 6 hours after dermal dosing suggesting constant percutaneous absorption. NMP appeared to be absorbed more extensively and a slightly faster rate than 2-pyrrolidone. Percutaneous absorption was more extensive in females than in males. Subsequently investigation revealed only a negligible first pass effect for the two co-administered substances (Midgley et al., 1992). After oral administration, maximum plasma concentrations of the parent compound reached levels of 25 to 30 mg/L after 2 hours. Plasma levels declined thereafter but were still measureable 5 days after dosing. The terminal half-lives were calculated as 27 and 29 hours for females and males, respectively.

Following intramuscular injection of 25 mg/kg bw labelled 2-pyrrolidone to rats rapid absorption and distribution with peak plasma levels of 33.1 mg/L at 1 to 3 hours were observed. This concentration declined rapidly within 8 hours to about 4 mg/L (EMEA summary report on 2-pyrrolidone, July 1998). 

 

Distribution

The distribution of 2-pyrrolidone includes the interesting information that its metabolite 2-pyrrolidinone is able to cross the blood-brain-barrier, and is metabolized in the brain to gamma-aminobutyric acid (Callery et al., 1979).

Metabolism

The metabolism of 2-pyrrolidone occurs via two main pathways, with one leading to the formation of gamma-aminobutyric acid (Callery et al., 1979), an inhibitory transmitter in the central nervous system. The distribution of a radioactive labeled metabolite of 2-pyrrolidone, namely 2-pyrrolidinone administered intravenously to mice was investigated by Callery et al. 1979. They proved that 2-pyrrolidone can cross the blood-brain barrier and can be classified as a precursor of gamma-aminobutyric acid, due to the fact that they detected 2-pyrrolidinone and its hydrolysation to gamma-aminobutyric acid in the brain (see also Roberts et al. 1974, Roberts et al. 1976). The steady state level of gamma-aminobutyric acid in the brain was not altered after 2-pyrrolidinone application, and due to this, the question arises, if 2-pyrrolidone possesses relevance by possible influence on the physiological processes of gamma-aminobutyric acid in the brain. The other main pathway of metabolism leads with various intermediates finally to succinimide (Bandle et al.,1984). They showed that 2-pyrrolidinone, a derivate of 2-pyrrolidone can be found in plasma of the three following species: human, rat and mouse, but in dog plasma, the amount of 2-pyrrolidinone was below the detection limit. The assumption was made that 2-pyrrolidinone can be metabolized first to 5-hydroxy-2-pyrrolidinone, which is further metabolized to succinimide. They have proven that the conversion of 2-pyrrolidinone to 5-hydroxy-2-pyrrolidinone can only be conducted by rat liver but not by rat brain homogenates.

An investigation of the fate of putrescine in vitro was conducted by Lundgren and Fales in 1980. They have proven that 2-[14C]-pyrrolidone, a metabolite of putrescine is metabolized in rat liver slices in a considerable amount to 5-hydroxy-2-pyrrolidone (Lundgren and Fales, 1980). Additionally there is considerable evidence that the conversion of putrescine to 2-pyrrolidone can be performed by diamin oxidases and that the conversion of both substances can be mediated by monoaminooxidases (Lundgren and Fales, 1980). In rats, 8 hours after oral co-administration of 112 mg/kg bw labelled 2-pyrrolidone and 75 mg/kg bw labelled N-methyl-2-pyrrolidone (2:3) in isopropanol, the parent compounds represented 80 % of the total plasma radioactivity indicating low first pass metabolism (Midgley et al., 1991). By 12 hours after dosing virtually all radioactivity was found as unknown polar metabolites. In an other study metabolism of putrescine was investigated in different organs slices in vitro. The authors demonstrated the formation of 2-pyrrolidone after a 4 h incubation of liver, spleen and lung slices with putrescine, whereas brain, kidney, heart and brain slices were found to be unable to metabolise 2-pyrrolidone (Lundgren and Hankins, 1978).

Excretion

Its elimination is primarily conducted via urine and as minor routes of elimination feces (1-2 %) and exhalation (6-7 %) were identified in rats (Midgley et al., 1992). Already 12 hours after oral intake nearly complete conversion of radiolabeled N-methyl-2-pyrrolidone (NMP) and 2-pyrrolidinone to unknown polar metabolites had taken place. A near complete excretion (85-88 % of the total radioactive dose) of mother compounds and metabolites was realized after 5 days in rats (Midgley et al., 1991, cited in Midgley et al., 1992). In rats, 61-70 % of a dermal dose of 2.5 mg/cm² N-methyl-2-pyrrolidone and 1.67/cm² of 2-pyrrolidone was excreted urinary, faecal excretion represented 1-2 % and 6-7 % were exhaled as CO2within 5 days. About 1 % of the total dose remained in the carcass (Midgley et al, 1991). Approximately 40 % of an administered intramuscular injection of 25 mg/kg bw labelled 2-pyrrolidone to rats was recovered in the urine within the first day and only 1 % on the second day. About 10% of the dose were exhaled in form of CO2 within 8 hours and only negligible amounts were found in the faeces. In a second study urinary elimination amounted for about 60 % of the total dose within 24 hours following intramuscular injection of 23 mg/kg bw 2-pyrrolidone (EMEA summary report on 2-pyrrolidone, 1998).