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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
Reliability:
1 (reliable without restriction)
Rationale for reliability incl. deficiencies:
guideline study

Data source

Reference
Reference Type:
study report
Title:
Unnamed
Year:
2008
Report Date:
2008

Materials and methods

Test guideline
Qualifier:
according to
Guideline:
OECD Guideline 428 (Skin Absorption: In Vitro Method)
Version / remarks:
2004
Deviations:
no
GLP compliance:
yes

Test material

Reference
Name:
Unnamed
Type:
Constituent
Type:
Constituent
Test material form:
solid - liquid: aqueous solution
Details on test material:
- Name of test material: DOPA-Glycinate
Radiolabelling:
yes

Test animals

Species:
human

Administration / exposure

Type of coverage:
open
Vehicle:
other: tap water
Duration of exposure:
6 hours
Termination by washing skin sample surface with commercial soap concentrate followed by rinsing with a dilute soap solution and drying with tissue swabs.
Doses:
Test preparation 1: 14C DOPA-Glycinate (21.094 % a.i.) determined by radioactivity
Test preparation 2: 14C DOPA-Glycinate (0.228 % a.i.) determined by radioactivity (diluted with tap water)
Details on in vitro test system (if applicable):
SKIN PREPARATION
- Skin samples: Split-thickness human skin membrane samples

APPARATUS
Automated flow-through diffusion cell apparatus (Scott/Dick, University of Newcastle-upon-Tyne, UK), placed in a steel manifold heated via a circulating water bath to maintain the skin surface temperature. The cells were connected to multi-channel peristaltic pumps from their afferent ports with the receptor fluid effluent dropping via fine bore tubing into scintillation vials on a fraction collector.

PRINCIPLES OF ASSAY
- Volume applied: 10 µL/cm²; the surface area of exposed skin within the cells was 0.64 cm².
- Receptor fluid: Tissue culture medium containing bovine serum albumin (ca 5%, w/v), glucose (ca 1%, w/v), streptomycin (0.1 mg/mL) and penicillin G (100 units/mL). The receptor chamber volume was 0.25 mL, the peristaltic pumps were adjusted to maintain a flow-rate of ca 1.5 mL/h.

Results and discussion

Signs and symptoms of toxicity:
not examined
Dermal irritation:
no effects
Total recovery:
Test preparation 1, 20 % (w/w): 101.69 % (SD = 3.04%)
Test preparation 2, 0.2 % (w/w): 97.10 % (SD = 2.99%)

Percutaneous absorptionopen allclose all
Time point:
24 h
Dose:
20%
Parameter:
percentage
Remarks:
potentially absorbable dose (sum of the absorbed dose, exposed skin and stratum corneum tape strips 3–20)
Absorption:
1.1 %
Time point:
24 h
Dose:
0.2%
Parameter:
percentage
Remarks:
potentially absorbable dose (sum of the absorbed dose, exposed skin and stratum corneum tape strips 3–20)
Absorption:
15.1 %
Time point:
24 h
Dose:
20%
Parameter:
percentage
Remarks:
Dermal delivery (exposed skin + absorbed dose)
Absorption:
0.6 %
Time point:
24 h
Dose:
0.2%
Parameter:
percentage
Remarks:
Dermal delivery (exposed skin + absorbed dose)
Absorption:
5 %

Any other information on results incl. tables

Test preparation 1, 20 % (w/w):99.76 % of the applied dose was removed by washing at 6 h post application. At 24 h post application, the total dislodgeable dose was 100.35 % of the applied dose. The stratum corneum retained 0.72 % of the applied dose, 0.37 % was removed with the first 5 tape strips.

Absorbed dose: 0.01 % (0.18 ng equivalent/cm²)

Dermal delivery: 0.62 % (13.17 ng equivalent /cm²)

Potentially absorbable dose: 0.97 % (20.53 ng equivalent /cm²)

Test preparation 2, 0.2 % (w/w):73.66 % of the applied dose was removed by washing at 6 h post application. At 24 h post application, the total dislodgeable dose was 75.30 % of the applied dose. The stratum corneum retained 16.08 % of the applied dose, 9.71 % was removed with the first 5 tape strips.

Absorbed dose: 0.11 % (0.02 ng equivalent /cm²)

Dermal delivery: 5.04 % (1.01 ng equivalent /cm²)

Potentially absorbable dose: 11.42 % (2.28 ng equivalent /cm²)

 

 

Table A6.2-4:Details on human skin.

Donor no.

Sex/age of donor

Site of sampling

Supplier

Membrane full-thickness (µm)

Membrane split-thickness (µm)

0164

F/28Y

Breast

St. Johns Hospital

1010–1150

380–390

0161

F/36Y

Abdomen

BUPA

1020–1080

390–400

0162

F/35Y

Abdomen

BUPA

700–1190

390–400

0179

F/84Y

Breast

Nottingham

1130

400

0169

F/82Y

Breast

Nottingham

1000

400

0180

F/74Y

Breast

Nottingham

940–1000

400

0166

F/33Y

Abdomen

Transkin

1240

400

 

 

Table A6.2-5:Summary of the results (mean values).

Test preparation

1

2

Target concentration of active substance

20 % (w/w)

0.2 % (w/w)

Active substance concentration in test preparation by radioactivity

21.016 % (w/w)

0.199 % (w/w)

Application rate of Test preparation

10.06 mg/cm²

10 µl/cm²

Application rate of test item

2114 µg equivalent/cm²

19.94 µg equivalent/cm²

Distribution

% applied dose

ng equivalent /cm²

% applied dose

ng equivalent /cm²

Dislodgeable dose 6 h
(skin wash+tissue swab+pipette tips)

99.76

2108.47

73.66

14.69

Total dislodgeable dose
(dislodgeable dose 6 h +stratum corneum + unexposed skin + cell wash)

100.35

2120.83

75.30

15.02

Unabsorbed dose

101.07

2136.17

92.06

18.36

Absorbed dose
(cumulative receptor fluid + receptor rinse)

0.01

0.18

0.11

0.02

Dermal delivery
(exposed skin + absorbed dose)

0.62

13.17

5.04

1.01

Potentially absorbable dose
(dermal delivery + stratum corneum tape strips)

0.97

20.53

11.42

2.28

Mass balance
(unabsorbed dose + dermal delivery)

101.69

2149.34

97.10

19.36

 

According to EFSA Guidance on Dermal Absorption (2012) as well as the EU Guidance Document

on Dermal Absorption (2004) only the first 2 tape strips should be discarded. On that basis dermal absorption for both low and high doses have been re-calculated.  

 

Recalculation disregarding 2 rather than 5 tape strips:

 

Prep 1 (20% a.i.)

 

Cell 1

Cell 2

Cell 3

Cell 4

Cell 8

Cell 9

Cell 10

Cell 12

Cell 13

Cell 14

Mean

STDEV

1

0.008

0.019

0.08

0.068

0.18

 

0.282

0.12

0.023

0.144

0.103

0.090

2

0.003

0.011

0.141

0.099

0.093

 

0.295

0.063

0.01

0.201

0.102

0.098

Stratum total

0.04

0.1

0.99

0.69

0.95

 

1.58

0.62

0.16

1.33

0.718

0.549

minus 1+2

0.029

0.07

0.769

0.523

0.677

 

1.003

0.437

0.127

0.985

0.513

0.378

Dermal delivery

0.02

0.09

0.77

0.27

1.35

 

1.73

0.82

0.15

0.41

0.623

0.597

Absorbable Dose

0.049

0.16

1.539

0.793

2.027

 

2.733

1.257

0.277

1.395

1.137

0.907

 

Prep 2 (0.2% a.i.) 

 

Cell 16

Cell 17

Cell 18

Cell 19

Cell 24

Cell 25

Cell 26

Cell 28

Cell 29

Cell 30

Mean

STDEV

1

3.11

2.24

2.84

4.42

3.36

3.89

4.82

3.78

4.87

6.06

3.939

1.131

2

1.66

1.8

1.71

2.33

1.73

2.53

2.19

2.26

2.5

2.26

2.097

0.338

Stratum total

12.02

13.59

14.46

19.48

15.39

15.49

20.79

16.73

14.73

18.17

16.085

2.718

minus 1+2

7.25

9.55

9.91

12.73

10.3

9.07

13.78

10.69

7.36

9.85

10.049

2.052

Dermal delivery

2.56

2.52

3.84

3.15

8.21

9.37

6.12

7.47

3.15

4.02

5.041

2.538

Absorbable Dose

9.81

12.07

13.75

15.88

18.51

18.44

19.9

18.16

10.51

13.87

15.090

3.611

Applicant's summary and conclusion

Conclusions:
The systemically available dose of DOPA-Glycinate (= dermal delivery: exposed skin + absorbed dose) was 0.6±0.6% for the high exposure scenario (20% a.i.), and 5.0±2.5% for the low exposure scenario (0.2% a.i.), respectively.
The potentially absorbable dose of DOPA-Glycinate was 1.1±0.9% for the high exposure scenario (20% a.i.), and 15.1±3.6% for the low exposure scenario (0.2% a.i.), respectively.
Executive summary:

The percutaneous absorption of [14C]-DOPA-Glycinate was tested according to OECD 428: Skin Absorption: In Vitro Method (2004).

Split-thickness human skin membranes were mounted into flow-through diffusion cells. Receptor fluid, consisting of bovine serum albumin (ca. 5%), glucose (ca 1%), streptomycin (0.1 mg/mL), and penicillin G (100 U/mL), was pumped underneath the skin at a flow rate of ca 1.5 mL/h. Receptor fluid was collected in hourly fractions and analysed by liquid scintillation counting.

Test preparation 1, 20 % (w/w): Mass balance 101.69 % (SD = 3.04%), thus considered to be complete.

Absorbed dose: 0.01 % (0.18 ng equivalent/cm²)

Dermal delivery: 0.62 % (13.17 ng equivalent /cm²)

Potentially absorbable dose(sum of absorbed dose, exposed skin and stratum corneum tape strips 6–20): 0.97 % (20.53 ng equivalent /cm²)

Test preparation 2, 0.2 % (w/w): Mass balance 97.10 % (SD = 2.99%), thus considered to be complete.

Absorbed dose: 0.11 % (0.02 ng equivalent /cm²)

Dermal delivery: 5.04 % (1.01 ng equivalent /cm²)

Potentially absorbable dose (sum of absorbed dose, exposed skin and stratum corneum tape strips 6–20): 11.42 % (2.28 ng equivalent /cm²)

 

However, according to EFSA Guidance on Dermal Absorption (2012) as well as the EU Guidance Document on Dermal Absorption (2004) only the first 2 tape strips should be discarded. On that basis the potentially absorbable dose for both low and high doses has been re-calculated.  

The potentially absorbable dose (the sum of the absorbed dose, exposed skin and stratum corneum tape strips 3–20 may be used for risk assessment; this corresponds to 1.1 ±0.9% for the high exposure (20% a.i.), and 15.1±3.6% for the low exposure scenario (0.2 % a.i.), respectively.

For REACH-purposes the

"Dermal absorption represents the amount of topically applied test substance that is found in the epidermis (stratum corneum excluded) and in the dermis, and this quantity is therefore taken as systemically available. Dermal absorption is influenced by many factors, e.g. physico-chemical properties of the substance, its vehicle and concentration, and the exposure pattern (e.g. occlusion of the application site) as well as the skin site of the body (for review see ECETOC, 1993; Howes et al, 1996; Schaefer and Redelmaier, 1996). The term percutaneous penetration refers to in vitro experiments and represents the amount of topically applied test substance that is found in the receptor fluid – this quantity is taken as systemically available. " (Guidance on Information requirements and chemical safety assessment, R.7c).