Clinical Trials A Suction Blister Experiment

Aesthetic Dermatology Cosmeceuticals

Topical Retinoid Penetrates Intact Human Epidermis

A report by Peter Bjerring, Henrik Egekvist, Preben Jacobsen and Jørgen Serup:

  • Aarhus University
  • Copenhagen University

More than 90% of dietary retinol is in the form of esters, usually retinyl palmitate. Intestinal absorption following ingestions approximating daily requirements is complete. Largeamounts are stored in the liver and gradually metabolised or released into the plasma. Endogenous retinoids eventually reach the skin and exert a number of hormonal effects, maintaining normal skin functions. Retinoid metabolism and effects on skin are multiple.

One effect of natural and synthetic retinoids of major importance in cosmetic dermatology is a beneficial effect on skin ageing. However, the skin target may also be reached from outside using retinoids formulated as creams.’ With this approach, first-pass metabolism in the liver is avoided and the target in skin treated directly. Retinoids can actually penetrate the stratum corneum barrier quite well. Retinoids are local irritants, and penetration may be a result of their potentially damaging effect on the penetration barrier.

In our study, human skin penetration by natural retinoids from a cosmetic formulation was studied using the suction blister experiment. It was monitored using non-invasive biophysical methods to see whether any irritant effect on the skin occurred during the experiment. The aim was to study epidermal penetration of retinoids in intact human skin under conditions simulating normal usage.


Material and Methods


Fifteen healthy women aged 45.2+-9.1 years (mean +- standard deviation (SD]) participated in the study. The study was in accordance with the Declaration of Helsinki.

Test Substances arid Study Procedure

The subject were treated with a retinoid cream (Créme Mètamorfique, Beauté Pacifique, Hadsund, Denmark) and a placebo cream on the inside of the upper arms applied once daily for five consecutive days. A small amount (0.3ml) of the cream was applied onto a test area covering 36cm2 of the medial side of each upper arm. The retinoid cream contained 0.3mg retinoid per gram; 60% was formulated as retinyl palmitate and 40% as retinyl acetate. Applications were blinded. The application of active and placebo was randonmised for application of the right and left arms, respectively. The volunteers were instructed to wash their hands between applications No use of retinoid-containing capsules or creams was allowed before or during the study. Sun exposure on the upper arms was also avoided.

At the end of the study four suction blisters were applied, producing a vacuum of 25OmmHg for 85+-23 minutes. Suction cups were 6mm in diameter. The 200-350ml suction blister liquid was harvested with a thin syringe, and the fluid mixed with 25ul ascorbic acid 25mM and 25ul ethylene diamine tetraacetic acid (EDTA) 10mM. Analysis by high-performance liquid chromatography (HPLC) was performed and concentrations of retinyl palmitate, retinyl acetate and retinol were determined.

Clinical Scoring

Objective and subjective clinical changes were recorded. Symptoms and signs were categorised as burning or stinging skin sensation, local redness, local inflammation and local scaling measured as 0 (nil), 1 (minimal), 2 (moderate), or 3 (severe).

High Frequency, Ultrasound

Ultrasound examination was performed with a high-frequency 20Mhz ultrasound device (Dermascan C, Cortex Technology, Denmark) The axial resolution is 6Oum, and the lateral resolution 130um. The maximum viewing depth is 30mm The intensity of each image element. i.e. pixel in the dermis, was evaluated on an arbitrary scale (0-255), and the mean density of the dermal echo was measured. The percentage of pixels within the dermal region with the intensity of 0-30 on the intensity scale was calculated to measure dermal water and development of inflammatory oedema due to irritation.


Redness of the application sites was examined before and after cream application with a spectrophotometer (Dermaspectrometer, Cortex Technology, Hadsund, Denmark). The erythema index was measured.

Trans-epidermal Water Loss

Trans-epidermal water loss (TEWL) (g/m2h) was measured (Dermalab, Cortex Technology,Hadsund, Denmark) before and after the cream application period. Measurements were performed when the readings were stabilised after a period of 30-45 seconds. TEWL is a direct measure of the efficacy of the stratum corneum water barrier as a marker of barrier integrity to chemical penetrants.


Comparison of application sides treated with retinoid and placebo creams was performed using the Student’s t-test for paired samples. The values of retinyl acetate and retinyl palmitate concentrations in suction blister fluid did not follow a normal distribution. Final analysis was performed with the Wilcoxon signed rank test. The level of statistical significance was p‹0.05.



Suction Blister Fluid

Retinoids including vitamin A, precursors and derivatives were significantly higher in the suction blister fluid harvested from sites.

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