Skin penetration of chemicals: quantitative in vivo ......Quantification of skin penetration of products z • ρ A (z) = RA(z) Rprotein(z). 1 A/protein. ρ protein,skin (z) 1. RA(z)

18 Sept 2019, OEESC, Dublin

Skin penetration of chemicals: quantitative in vivo analysis by Raman spectroscopy

PROTECTION AGAINST

MICROORGANISMS,

CHEMICALS,

ALLERGENS,

POLLUTANTS,…

Gerwin Puppels, Claudio Nico, Johanna de Sterke, Paul Pudney,* Peter Caspers*Unilever Research Port Sunlight, UK

Analysis of skin penetration / permeation


Outline


Raman spectroscopy & in vivo skin analysis

Non-invasive analysis of the penetration of topically applied materials

Quantification Concentration (mg/cm3)

Total uptake (mg/cm2)

Flux (mg/cm2.h)

Example: retinol

Applications

molecular composition of the skin - spatially resolved

SC

living

epidermis

25%

70%

H2O

penetration

of topically

applied

materials

UV, season

nmf

age, body site, response to challenges, ….



O C O O C O



500 1000 1500 2000

lactate

blood

glycogen

Fatty acid

cholesterol

RNA

DNA

elastin

collagen

Dcm-1

20 mm

Proteins

DNA

Sugars

Lipids

RNA



Interpretation of spectra

Raman spectrum of stratum corneum

Signal contributions of:

Proteins

amino acids

sweat constituents

lipids

water



in vivo skin

400 600 800 1000 1200 1400 1600 1800

Inte

nsi

ty

Raman shift, cm-1

fit-result

residual



skin

window

laser

dermis

SC

epidermis

500 1000 1500

distance to skin

surface

co

nce

ntr

ation

skin surface





0 20 40 60 80 100 120 140 160 180 2000

10

20

30

40

50

60

70

80

depth (micron)

wat

er (

mas

s-%

)



skin characterization

• skin typing

• age related effects

• seasonal effects

• skin conditions

• skin lesions

Topically applied products

penetration & permeation of:•drugs

•personal care actives •perfumes (fragrances)

•chemicals


SC ED

0 10 20 30

productconcentration

Distance to skin surface (μm)

product


Skin penetration of chemicals: quantitative in vivo analysis

in vivo skin

400 600 800 1000 1200 1400 1600 1800

Inte

nsi

ty

Raman shift, cm-1

fit-result

residual



skin

400 600 800 1000 1200 1400 1600 1800

Inte

nsi

ty

Raman shift, cm-1

fit-result

residual

SC EDproduct



RA ~ ρA

~ Ilaser

~ f(Ω,V)

• RA: intensity of the Raman signal of a compound A [photons/s]

• gA: concentration of compound A in the measurement volume V [kg/m3]

• Ilaser: laser light intensity [J/s]

• V: measurement volume [m3]

• Ω: measurement configuration

• kA: proportionality constant (property of compound A)

RA = kA . Ilaser . f(Ω,V) . gA

= CA(Ilaser,Ω,V) . ρA

homogeneous medium: compound A in solution



z

Skin: an inhomogeneous medium



z

The skin is an inhomogeneous medium



Quantification of skin penetration of products

z

The skin is an inhomogeneous medium

•RA

RB= CA/B .

gAgB

gA = RA

RB

.1

CA/B. gB

• gprotein,skin(z)

the protein concentration in the skin can be estimated from water concentration profiles in the skin


Eliminating unknowns



z 0 20 40 60 80 100 120 140 160 180 2000

10

20

30

40

50

60

70

80

Distance z to skin surface (µm)

wat

er (

mas

s-%

)

sc epidermis dermis

• mass-fractionwater,skin = 0.25 - 0.65 (SC)

• mass-fractionwater,skin = 0.65 - 0.70 (epidermis)

• mass-fractionwater,skin = 0.70 – 0.75 (dermis)



z 0 20 40 60 80 100 120 140 160 180 2000

10

20

30

40

50

60

70

80

Distance z to skin surface (µm)

dry

mas

s (m

ass-

%)

sc epidermis dermis

density dry SC = 1.15 g/cm3

60% protein;

15% lipids;

20 - 30% natural moisturizing factor




z

• ρA(z) = RA(z)

Rprotein(z).

1CA/protein

. ρprotein,skin(z)

1.RA(z)

Rprotein(z): from skin Raman spectra

2. CA/protein : from calibration experiments

3. ρprotein,skin(z) : from water concentration profiles





Proportionality constants Cmaterial:solvent

Cmaterial:solvent = (Rmaterial / Rsolvent) / (mmaterial / msolvent)

Cretinol:protein = Cretinol:ethanol . Cethanol:water / Cprotein:water





• 0.3% retinol in 70% ethanol – 30% PG

• Volar forearm

• 70 ml on skin area of 4 x 4 cm2

• 10 min

distance to skin surface (mm)

con

cen

trat

ion

(m

g /

cm3)

0 5 10 15 20 250

2

44

6

8

10

12

amo

un

t in

SC

(m

g /

cm2)

1

2

3

4

5

6

7

8

0

time (h)

1 2 3 4 5 6

Flux: 0.6 mg/cm2.h



distance to skin surface (mm)

con

cen

trat

ion

(m

g /

cm3)

0 5 10 15 20 25

amo

un

t in

SC

(m

g /

cm2)

50

100

150

200

250

300

0

time (h)

1 2 3 4 5 6

Flux: 8 mg/cm2.h

50

100

150

200

250

300

0

350

400



• retinol

• max. retinol concentration in SC ≃ retinol concentration in formulation after evaporation of ethanol

• 40% of applied dose detected in the skin

• After 6 hrs: 75% skin

• propylene glycol

• 16% of applied dose detected in the skin

• After 6 hrs: 20% skin


Skin penetration of topically applied materials

• In vivo Raman spectroscopy can provide• Qualitative information

• Did a material penetrate into the skin?

• How far into the skin?

• How fast?

• How long does it stay in the skin?

• Semi-quantitative information

• Comparison of efficiency of delivery of an active for different formulations.

• Comparison of delivery for different application times

• …

• Quantitative information

• How much penetrated into the skin (µg.cm-2)?

• What is the flux into the skin (µg/s-1.cm-2)?


Documents

Skin penetration of chemicals: quantitative in vivo ......Quantification of skin penetration of products z • ρ A (z) = RA(z) Rprotein(z). 1 A/protein. ρ protein,skin (z) 1. RA(z)