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Toxicological information

Dermal absorption

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Administrative data

Endpoint:
dermal absorption in vitro / ex vivo
Type of information:
experimental study
Adequacy of study:
key study
Study period:
2012-12-04 to 2013-01-24
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2013

Materials and methods

Test guidelineopen allclose all
Qualifier:
according to
Guideline:
OECD Guideline 428 (Skin Absorption: In Vitro Method)
Qualifier:
according to
Guideline:
other: - OECD Environmental Health and Safety Publications, Series on Testing and Assessment no. 28. Guidance document for the conduct of skin absorption studies (March 2004).
Qualifier:
according to
Guideline:
other: - OECD Guidance Notes on Dermal absorption, Draft 22 October 2010
Qualifier:
according to
Guideline:
other: - European Commission Guidance Document on Dermal Absorption – Sanco/222/2000/Rev. 7 (19 March 2004)
GLP compliance:
yes (incl. certificate)

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
other: liquid
Details on test material:
- Name of test material (as cited in study report): 3-Methoxy-3-methyl-1-butanol
- Molecular formula (if other than submission substance): C6H14O2
- Molecular weight (if other than submission substance): 118.14
- Physical state: Clear colourless liquid (determined at WIL Research Europe B.V.)
- Analytical purity: 99.47%
- Lot/batch No.: 32726
- Expiration date of the lot/batch: 30 June 2013
- Stability under storage conditions: Stable
- Storage condition of test material: At room temperature in the dark
- Other:
Study specific test substance information:
- Purity/composition correction factor required: No
- Hygroscopic: Yes, store in well-sealed container
- Volatile: Yes, vapour pressure: 47 Pa at 293K
- Density: 0.927 g/cm3 (20°C)
- Stability in vehicle: Water: Yes
- Solubility in vehicle: Water: Miscible

3-Methyl-3-methoxy [4-14C]butan-1-ol (substance 204543/A)
- Identification: 3-Methyl-3-methoxy-[4-14C]1-butanol
- Molecular formula: C6H14O2
- Molecular weight: 119.85 (at this specific activity)
- Description: Solution in acetonitrile
- Batch: 7047CDB026-6
- Radiochemical purity: 98.5%
- Chemical purity: Not indicated
- Test substance storage: In freezer (= -15°C) in the dark
- Expiry date: 30 November 2013 (allocated by WIL Research Europe B.V., 1 year after receipt of the test substance)
- Specific activity: 16.10 MBq/mg (1930 MBq/mmol)
- Radioactive concentration: 24.76 MBq/mL
- Total activity: 111.6 MBq
- Supplier: Selcia Limited, Fyfield Business and Research Park, Fyfield Road, Ongar, Essex, CM5 0GS, UK

[3H]-H2O (180603/E):
- Identification: Water [3H] (biological grade)
- Molecular formula: H2O
- Molecular weight: 18
- Description: Clear solution
- Batch: 110511
- Test substance storage: In refrigerator (2-8°C) in the dark
- Expiry date: 21 October 2013 (allocated by WIL Research Europe, 1 year after receipt of the test substance)
- Concentration: 1 mCi/mL (37 MBq/mL)
- Total activity: 5 mCi (185 MBq) (taken from label)
- Supplier: American Radiolabeled Chemicals, Inc., 101 ARC Drive, Saint Louis, MO 63146 USA

Radiolabelling:
yes

Test animals

Species:
human
Strain:
not specified
Sex:
not specified

Administration / exposure

Type of coverage:
occlusive
Vehicle:
water
Duration of exposure:
24 h
Doses:
undiluted (100%) mixed with 3-Methyl-3-methoxy[4-14C]butan-1-ol
aqueous dilution (10%) mixed with 3-Methyl-3-methoxy[4-14C]butan-1-ol
Details on study design:
Human skin was placed into flow-through cells and exposed to 3-Methoxy-3-methyl-1-butanol for 24 hours under occlusion conditions. The receptor fluid was collected at various time points. At the end of the experiments the amount of radioactivity was determined at the donor site, in the skin (tape-stripping and total skin) and in the receptor fluid. Prior to the determination of the dermal absorption of 3-Methoxy-3-methyl-1-butanol, the skin integrity was checked by determination of the permeation of tritiated water.

- Justification of species, anatomical site and preparative technique: Human skin is a recommended test system forin vitroskin absorption studies.
Details on in vitro test system (if applicable):
- Flow-through system: The skin penetration study was performed on a flow-through cell system consisting of :
peristaltic pump; 13 flow-through cells, diameter 9 mm (Permegear, Bethlehem, PA, USA), fraction collector and thermostat.
- Alliance HT Separations module 2795 liquid chromatograph and a
- Radio-Detector:ß-RAM Model 4 (Lablogic, Sheffield, England).
The receptor fluid was pumped at a flow rate of 1.5 mL/h.
- Test temperature: The skin surface temperature was 32±1ºC
- Humidity: ambient humidity (30-70%)
- Occlusion: yes

Results and discussion

Signs and symptoms of toxicity:
not examined
Dermal irritation:
not examined
Absorption in different matrices:
The receptor fluid consisted of saline (0.9% NaCl solution); donor chamber was occluded during the exposure.
A maximum solute flux of 0.055mg/cm²/h for the pure MMB and 0.113 mg/cm²/h for the 10% aqueous solution was determined.
Total recovery:
Undiluted sample (100%): 84 +- 11%
Aqueous dilution (10%) : 80 +-4%

Percutaneous absorptionopen allclose all
Dose:
10%
Parameter:
percentage
Absorption:
33 %
Remarks on result:
other: 4h
Remarks:
lag time 2h
Dose:
10%
Parameter:
percentage
Absorption:
40 %
Remarks on result:
other: 24h
Remarks:
lag time 2h
Dose:
100%
Parameter:
percentage
Absorption:
4 %
Remarks on result:
other: 8h
Remarks:
lag time 2.5h
Dose:
100%
Parameter:
percentage
Absorption:
6.9 %
Remarks on result:
other: 24h
Remarks:
lag time 2.5h

Any other information on results incl. tables

Radiochemical purity:

The radiochemical purity of a stock solution of3-Methyl-3-methoxy[4-14C]butan-1-olin 0.1% TFA in MQ/Acetonitrile (1:1, v/v) was determined at the start of the study. The radiochemical purity was 98.9% and 99.3% (based on two independent injections), which was above the required 97%.

 

Homogeneity of test substance samples mixed with 3-Methyl-3-methoxy[4-14C]butan-1-ol:

The results of the homogeneity check of the formulations mixed with 3-Methyl-3-methoxy[4-14C]butan-1-olare presented inTable 2. The relative standard deviations from samples taken from the top, middle and bottom of the vial were below 3% and thus all formulations were homogeneous.

 

Table 2:  Results of homogeneity check of formulations

Formulation

Activity

(MBq/mL)

Subsample

counted

(mL)

Activity (dpm)

MBq/mL

Undiluted (100%)

2.1

0.005

T: 631078

M: 623856

B: 606871

2.104

2.080

2.023

 

 

 

Average

RSD

2.1

2.0%

 

2.0

0.010

T: 1207028

M: 1197346

B: 1203662

2.012

1.996

2.006

 

 

 

Average

RSD

2.0

0.4%

Aqueous dilution (10%)

1.9

0.005

T: 580451

M: 572013

B: 551076

1.935

1.907

1.837

 

 

 

Average

RSD

1.9

2.7 %

 

2.0

0.010

T: 1198504

M: 1213581

B: 1231184

1.998

2.023

2.052

 

 

 

Average

RSD

2.0

1.3%

RSD: relative standard deviation

Table 3:   Results of the solubility test in the receptor fluid

Formulation

Expected activity in receptor fluid

Actual activity in receptor fluid

Dose

(g/L)

Activity

(MBq/mL)

Volume formulation applied (µL)

Volume receptor fluid (mL)

Activity (kBq/mL)

Subsample counted (mL)

(DPM)

(kBq/mL)

Recovery (%)1)

0.922

2.1

6.4

36

0.368

1.0

23365

0.389

106

 

 

 

1.0

23353

0.389

106

 

 

 

1.0

23266

0.388

105

 

 

 

 

 

Average

0.389

106

 

 

 

 

SD

0.0009

 

 

 

 

 

RSD

0.23%

 

1) ratio of activity in receptor fluid solution and the activity of formulation in receptor fluid, multiplied by 100

Skin integrity:

The permeability coefficients for tritiated water for the skin discs that have been used for this study are presented inTable 4. The integrity of the reported skin discs was within the acceptability criteria (Kp< 4.5 x 10-3cm/h).

Table 4: Skin integrity values

Test substance sample

Skin disc (donor number-disc number)

Kp value for tritiated water (*10-3cm/h)

undiluted

(100%)

2796-03

2824-08

2796-04

2824-11

2825-04

2830-04

3.80

1.93

4.02

1.90

2.15

3.13

Aqueous dilution

(10%)

2825-01

2830-01

2832-05

2830-05

 

 

2.75

3.58

3.49

2.97

 

 

Table 5: Dermal absorption parameters and percentage absorption of 3-Methoxy-3-methyl-1-butanol in human skinin vitro

Formulation

Undiluted 100%

(0.922 g/mL)

Aqueous dilution 10%

(0.092 g/mL)

Skin samples

human (n=6)

human (n=4)

% RAD1)

SD2)

% RAD1)

SD2)

DERMAL ABSORPTION PARAMETERS

 

 

 

Lag time (h)

1-3

 

1-2

 

Maximum flux (µg/cm2/h)

0.055

0.018

0.113

0.020

SURFACE COMPARTMENT

 

 

 

 

Total skin swabs 24 h

0.16

0.08

0.73

0.55

Material remaining in donor chamber

77

12

39

4

Total % non-absorbed3)

78

12

40

4

SKIN COMPARTMENT

 

 

 

 

Skin

0.09

0.04

0.35

0.06

Tape strips 1&2

0.01

0.01

0.005

0.004

Stratum corneum

(tape strips excluding 1&2)

0.03

0.01

0.03

0.01

Total % at dose site

(without tape strips 1&2)

0.13

0.04

0.38

0.06

RECEPTOR COMPARTMENT

 

 

 

 

Receptor fluid (collected over 24 h)

7

2

40

1

Receptor fluid terminal

0.03

0.01

0.04

0.03

Receptor chamber

0.03

0.02

0.07

0.07

Total % directly absorbed4)

7

2

40

1

OVERALL ABSORPTION

 

 

 

 

Total % potentially absorbable

(without tape strips 1&2)5)

7

2

40

1

Total % recovery

84

11

80

4

1)RAD = radioactivity

2) SD= Standard deviation

3)Total % non-absorbed = % in total skin swabs 24 h + % material remaining in donor chamber

4)Total % directly absorbed = % receptor fluid + % receptor fluid terminal + % receptor chamber

5)Total % potentially absorbable (without tape strips 1&2) = Total % at dose site (without tape strips 1&2) + total % directly absorbed

Applicant's summary and conclusion

Conclusions:
The in vitro dermal absorption of human skin with 3-Methoxy-3-methyl-1-butanol was 7 ± 2% for the undiluted substance and 40 ± 1%.for the 10% aqueous solution over a penetration time of 24 hours. A maximum solute flux of 0.055mg/cm²/h for the pure MMB and 0.113 mg/cm²/h for the 10% aqueous solution was determined.