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Toxicological information

Basic toxicokinetics

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Administrative data

Endpoint:
basic toxicokinetics in vivo
Type of information:
experimental study
Adequacy of study:
key study
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
test procedure in accordance with generally accepted scientific standards and described in sufficient detail

Data source

Referenceopen allclose all

Reference Type:
study report
Title:
Unnamed
Year:
2016
Report date:
2016
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report date:
2016
Reference Type:
study report
Title:
Unnamed
Year:
2016
Report date:
2016

Materials and methods

Objective of study:
toxicokinetics
Test guideline
Qualifier:
no guideline followed
Version / remarks:
This study is a pilot pharmacokinetic investigation and as such is not designed to follow specific test guidelines.
Principles of method if other than guideline:
This study was performed to obtain blood samples from rats at selected time points following a single 6-hour exposure of the test atmosphere of Methyisobutylketone at two different concentration levels of Low (500 ppm) and High (1000 ppm) in order to determine the concentrations and the pharmacokinetics parameters of Diacetone alcohol (DAA), Methyl-isobutyl ketone (MIBK) and Methylisobutylcarbinol (MIBC) from the plasma.
GLP compliance:
yes (incl. QA statement)

Test material

Constituent 1
Chemical structure
Reference substance name:
4-methylpentan-2-one
EC Number:
203-550-1
EC Name:
4-methylpentan-2-one
Cas Number:
108-10-1
Molecular formula:
C6H12O
IUPAC Name:
4-methylpentan-2-one
Test material form:
liquid
Radiolabelling:
no

Test animals

Species:
rat
Strain:
Sprague-Dawley
Details on species / strain selection:
SPF colony
Sex:
male
Details on test animals or test system and environmental conditions:
TEST ANIMALS
- Source: Charles River Laboratories, Research Models and Services, Germany GmbH, Sandhofer Weg 7, D-97633
- Age at study initiation: 10-11 weeks old
- Weight at study initiation: 387g-468g
- Housing: three animals per cage
- Diet (ad libitum): ssniff® SM R/M-Z+H "Autoclavable complete feed for rats and mice
- Water (ad libitum): tap water from municipal supply
- Acclimation period: At least 5 days

ENVIRONMENTAL CONDITIONS
- Temperature (°C): 20.1 - 25.6
- Humidity (%): 40 - 62
- Air changes (per hr): 15-20
- Photoperiod (hrs dark / hrs light): 12/12

Administration / exposure

Route of administration:
inhalation: vapour
Vehicle:
unchanged (no vehicle)
Details on exposure:
TYPE OF INHALATION EXPOSURE: nose only

GENERATION OF TEST ATMOSPHERE / CHAMBER DESCRIPTION
- Exposure apparatus: nose only, past flow, dynamic flow exposure unit, consisted of two, concentric anodised aluminium cylinders, the inner plenum and the outer chamber with 20 circularly arranged exposure ports.
- Method of holding animals in test chamber: tube
- Source and rate of air: no data
- Method of conditioning air: oil-free compressor passed through a suitable filter system prior to introduction to the nebuliser
- System of generating particulates/aerosols: The test item was administered as vapours. The test item was aerosolised using a stainless steel concentric jet nebulisers (TSE Systems GmbH, Bad Homburg, Germany) located at the top of the exposure chambers and a separator glass was installed to remove any liquid particles from the atmosphere prior entering into the inhalation tower. The rate of test item use was controlled by a syringe pump.
- Temperature, humidity in air chamber: 21.0-25.5°C, 3.0-4.5%
- Air Flow In (Inner Plenum) (L/min): 29.6-30.8
- Air Flow Out (Outer Cylinder) (L/min): 23.6-25.7
- Air change rate: no data
- Method of particle size determination: not determined since the test item was evaporated and non-condensing.
- Treatment of exhaust air: no data

TEST ATMOSPHERE
- Brief description of analytical method used: the concentration of the test item during the exposure was measured by gas chromatography (GC/FID). Samples for analytical determination were taken once an hour during the exposure into an impinger containing ethanol as a solvent.
- Samples taken from breathing zone: yes
Duration and frequency of treatment / exposure:
single 6-hour exposure
Doses / concentrationsopen allclose all
Dose / conc.:
500 other: ppm (target)
Dose / conc.:
531 ppm (analytical)
Remarks:
2.18 mg/L
Dose / conc.:
1 000 other: ppm (target)
Dose / conc.:
1 022 ppm (analytical)
Remarks:
4.19 mg/L
No. of animals per sex per dose / concentration:
9
Control animals:
no
Details on dosing and sampling:
TOXICOKINETIC STUDY
- Tissues and body fluids sampled: plasma
- Time and frequency of sampling: 0 (on day -1 or -3), 0.5, 1, 1.5, 3, 4.5, 6, 8, 18 and 24 hours post exposure

METABOLITE CHARACTERISATION STUDIES
- Tissues and body fluids sampled: plasma
- Time and frequency of sampling: 0 (on day -1 or -3), 0.5, 1, 1.5, 3, 4.5, 6, 8, 18 and 24 hours post exposure
- From how many animals: 3
- Method type(s) for identification: Gas Chromatography coupled with MS detection after electron impact ionization (GC-MS)
- Limits of quantification: DAA < 0.5 µg/mL; MIBC < 0.1 µg/mL; MIBK < 0.2 µg/mL

TREATMENT FOR CLEAVAGE OF CONJUGATES (if applicable): no

Results and discussion

Toxicokinetic / pharmacokinetic studies

Toxicokinetic parametersopen allclose all
Key result
Toxicokinetic parameters:
other: Cmax (µg/mL)/500 ppm: DAA 45.1, MIBC 0.66, MIBK 23.5
Key result
Toxicokinetic parameters:
other: Tmax (h)/500 ppm: DAA 1.5, MIBC 0.5, MIBK 0.5
Key result
Toxicokinetic parameters:
other: t1/2 (h)/500 ppm: DAA 2.4, MIBC nc, MIBK 0.581
Key result
Toxicokinetic parameters:
other: AUC0.5-24h (h*µg/mL)/500 ppm: DAA 266, MIBC 0.499, MIBK 20.1
Key result
Toxicokinetic parameters:
other: Cmax (µg/mL)/1000 ppm: DAA 82.7, MIBC 1.36, MIBK 56.8
Key result
Toxicokinetic parameters:
other: Tmax (h)/1000 ppm: DAA 1.5, MIBC 0.5, MIBK 0.5
Key result
Toxicokinetic parameters:
other: t1/2 (h)/1000 ppm: DAA 4.91, MIBC nc, MIBK nc
Key result
Toxicokinetic parameters:
other: AUC0.5-24h (h*µg/mL)/1000 ppm: DAA 522, MIBC 1.82, MIBK 70.2

Metabolite characterisation studies

Metabolites identified:
yes
Details on metabolites:
Diacetone alcohol (DAA), Methyl-isobutyl ketone (MIBK) and Methylisobutyl carbinol (MIBC)

Any other information on results incl. tables

Mean plasma concentrations of DAA, MIBK and MIBC after an inhalation exposure to MIBK at 500 and 1000 ppm

Exposure

Analyte (µg/mL)

Sampling time

Pre-dose

0.5 h

1 h

1.5 h

3 h

4.5 h

6 h

8 h

18 h

24 h


MIBK

500 ppm

 

DAA

BLQ

41.7

44.3

45.1

43.1

29.0

15.8

12.9

0.559

BLQ

MIBK

BLQ

23.5

13.7

5.18

2.32

0.15

BLQ

BLQ

BLQ

BLQ

MIBC

BLQ

0.660

0.356

0.170

BLQ

BLQ

BLQ

BLQ

BLQ

BLQ

MIBK
1000 ppm

 

DAA

BLQ

62.6

58.7

82.7

70.8

49.9

52.8

32.4

0.84

0.73

MIBK

BLQ

56.8

33.1

23.6

12.5

0.633

0.267

0.193

BLQ

BLQ

MIBC

BLQ

1.36

0.852

0.698

0.280

BLQ

BLQ

BLQ

BLQ

BLQ

BLQ: Below Limit of Quantification.

Toxicokinetic parameters of DAA, MIBC and MIBK after a single 6-hour inhalation
exposure of male Sprague-Dawley rats to 500 ppm and 1000 ppm of DAA

 

Concentration

Analytes

Number of point used to calculatelz

lz

t1/2

Tmax

Cmax

Cmax/
concentration#

AUC0.5-24h

AUC0.5-24h/
concentration#

Metabolite/
Administered compound

1/h

h

h

µg/mL

h*µg/mL

Cmax

AUC0.5-24h

500 ppm
(2.38 mg/L)

DAA

0.994

5

0.289

2.40

1.5

45.1

22.0

266

130

1.92

13.20

MIBC

na

na

nc

nc

0.5

0.66

0.322

0.499

0.243

0.0281

0.0248

MIBK

0.944

4

1.19

0.581

0.5

23.5

11.5

20.1

9.82

na

na

1000 ppm
(4.75 mg/L)

DAA

0.948

6

0.248

2.79

1.5

82.7

20.2

522

127

1.45

7.44

MIBC

0.995

3

0.569

1.22

0.5

1.36

0.331

1.82

0.445

0.0239

0.0260

MIBK

0.912

6

0.832

0.833

0.5

56.8

13.9

70.2

17.1

na

na

na: not applicable.

nc: not calculated as not enough quantifiable points were available.

#:Cmaxand AUC0.5-24hwere divided by the exposure concentrations of DAA (in mg/L).

 

Applicant's summary and conclusion

Executive summary:

A study was performed to obtain blood samples from rats at selected time points following a single 6-hour exposure of the test atmosphere of Methylisobutylketone at target concentration levels of 500 and 1000 ppm in order to determine the concentrations and the pharmacokinetics parameters of Diacetone alcohol (DAA), Methyl-isobutyl ketone (MIBK) and Methyl-isobutyl carbinol (MIBC) from the plasma. The animals were exposed to the test atmosphere (in the form of a vapour) using a nose-only exposure system. Analytical concentrations of 531 ppm, 1022 ppm were achieved in the respective groups. No control animals were used in the study. The test atmosphere concentration was monitored based on a validated GC method (by trapping the vapors in an impinger containing ethanol). The results of the test atmosphere characterization were considered suitable for the study purposes. Heighteen male Sprague-Dawley rats were involved in the study, 9 animals in each treatment groups. Plasma samples were prepared from male rats following the end of the exposure at nine time-points (three animals at each sampling occasion) and the plasma level of DAA, MIBK and MIBC was determined. The analytical method consisted of a liquid/liquid extraction of the three analytes from plasma with Methyl tert-Butyl Ether (MtBE) followed by analysis of supernatant by Gas Chromatography coupled with MS detection after electron impact ionization (GC-MS). The toxicokinetic evaluation was performed using non-compartmental analysis on Phoenix WinNonlin software (Pharsight Corporation, Mountain View, California 94040/USA). DAA, MIBC and MIBK toxicokinetic parameters were determined from the mean plasma concentration collected from each animal/group, at each post-exposure time-point. A separate analysis was performed for each dose-level. The following toxicokinetic parameters were calculated t½, ¿z, AUC0.5-24h, Tmaxand Cmax.

The mean actual achieved concentration of the test item were:

Target Concentration

(ppm)* [mg/L]

Achieved Concentration

(ppm)* [mg/L]

Nominal Concentration

 mg/L

500 [2.05]

531 [2.18]

3.69

1000 [4.10]

1022 [4.19]

6.17

For the three analytes (DAA, MIBK and MIBC), the standard acceptance criteria for the calibration lines and Quality Control samples were met for a successful analysis of the study samples.

Study samples with concentrations above the calibration ranges were repeated diluted (for DAA principally and MIBK). DAA, MIBC and MIBK were not quantifiable in pre-dose samples in any group.

The mean plasma concentrations of DAA, MIBK and MIBC after an inhalation exposure to MIBK at 500 and 1000 ppm were:

Exposure

Analyte (µg/mL)

Sampling time

Pre-dose

0.5 h

1 h

1.5 h

3 h

4.5 h

6 h

8 h

18 h

24 h


MIBK

500 ppm

 

DAA

BLQ

41.7

44.3

45.1

43.1

29.0

15.8

12.9

0.559

BLQ

MIBK

BLQ

23.5

13.7

5.18

2.32

0.15

BLQ

BLQ

BLQ

BLQ

MIBC

BLQ

0.660

0.356

0.170

BLQ

BLQ

BLQ

BLQ

BLQ

BLQ

MIBK
1000 ppm

 

DAA

BLQ

62.6

58.7

82.7

70.8

49.9

52.8

32.4

0.84

0.73

MIBK

BLQ

56.8

33.1

23.6

12.5

0.633

0.267

0.193

BLQ

BLQ

MIBC

BLQ

1.36

0.852

0.698

0.280

BLQ

BLQ

BLQ

BLQ

BLQ

BLQ: Below Limit of Quantification.

The toxicokinetic parameters of DAA, MIBC and MIBK after a single 6-hour inhalation exposure of male Sprague-Dawley rats to 500 ppm and 1000 ppm of MIBK were:

 

Concentration

Analytes

Number of point used to calculatelz

lz

t1/2

Tmax

Cmax

Cmax/
concentration#

AUC0.5-24h

AUC0.5-24h/
concentration#

Metabolite/
Administered compound

1/h

h

h

µg/mL

h*µg/mL

Cmax

AUC0.5-24h

500 ppm
(2.38 mg/L)

DAA

0.994

5

0.289

2.40

1.5

45.1

22.0

266

130

1.92

13.20

MIBC

na

na

nc

nc

0.5

0.66

0.322

0.499

0.243

0.0281

0.0248

MIBK

0.944

4

1.19

0.581

0.5

23.5

11.5

20.1

9.82

na

na

1000 ppm
(4.75 mg/L)

DAA

0.948

6

0.248

2.79

1.5

82.7

20.2

522

127

1.45

7.44

MIBC

0.995

3

0.569

1.22

0.5

1.36

0.331

1.82

0.445

0.0239

0.0260

MIBK

0.912

6

0.832

0.833

0.5

56.8

13.9

70.2

17.1

na

na

na: not applicable.

nc: not calculated as not enough quantifiable points were available.

#:Cmaxand AUC0.5-24hwere divided by the exposure concentrations of DAA (in mg/L).

After a 6-hour inhalation exposure to MIBK:

·        MIBK was on average quantifiable from 0.5 to 4.5 h after the end of exposure to 500 ppm and for up to 8 h at 1000 ppm. Maximum concentrations of MIBK were at 0.5 h after the end of exposure for both MIBK concentrations,

·        DAA was on average quantifiable from 0.5 to 18 h after the end of exposure to MIBK at 500 ppm and for up to 24 h at 1000 ppm. Maximum concentrations of DAA were at 1.5 h after the end of exposure for both MIBK concentrations,

·        MIBC plasma concentrations were very low but nevertheless quantifiable from 0.5 to 1.5 h after the end of exposure to 500 ppm of MIBK and for up to 3 h at 1000 ppm. Maximal plasma concentrations of MIBC were at 0.5 h after the end of exposure for both MIBK concentrations.

·        the elimination rates and half-live of DAA and MIBK were not significantly changed between both exposure concentrations, indicating that the excretion pathway was not saturated.

The plasma concentration values of DAA were higher than those of MIBC and MIBK after administration of MIBK, this indicates that DAA may represent a major metabolite of MIBK in rats. MIBC was considered to be a minor metabolite as it represented approximately 2.5% of the plasma MIBK exposure.

Based on AUC0-24hand Cmax(both normalized by exposure concentration), plasma DAA, MIBC and MIBK exposure increased dose proportionally, between 500 and 1000 ppm of MIBK.

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