Mechanistic investigation of skin barrier perturbation induced by surfactants in the presence of humectants

Abstract

The stratum corneum (SC) of the skin functions as a barrier between the body and the environment. Surfactants such as Sodium Dodecyl Sulfate (SDS) are used in skin cleansers and in skin-care formulations because of their ability to stabilize oil-water emulsions and clean the surface of the skin. However, they also have adverse effects on the skin barrier, including enhancing skin barrier perturbation which may lead to a disruption of the protective functions carried out by the skin barrier. On the other hand, humectants, such as Glycerol, which maintain the natural water content of the skin and preserve the skin barrier, have been shown to mitigate surfactant-induced skin barrier perturbation. The primary objective of this thesis was to develop a mechanistic understanding, including visualization and quantification, of: (i) how aqueous surfactant solutions, once in contact with the skin, can induce skin barrier perturbation, and (ii) how surfactant-induced skin barrier perturbation can be effectively mitigated through the addition of humectants to the aqueous surfactant contacting solutions. SDS monomers self-assemble to form micelles at a SDS concentration above the Critical Micelle Concentration (CMC). The SDS skin penetration and associated skin barrier perturbation is dose-dependent, that is, it increases with an increase in the total SDS concentration above the CMC of SDS.

(cont.) However, when Glycerol was added to the aqueous SDS contacting solution, through in vitro quantitative skin radioactivity assays using 14C radiolabeled SDS, I found that the dose-dependence in SDS skin penetration was almost completely eliminated. This is because the addition of Glycerol hinders the ability of the SDS micelles to penetrate into the skin barrier through aqueous pores that exist in the SC. In vitro Mannitol skin permeability and average skin electrical resistivity measurements, in the context of a hindered-transport aqueous porous pathway model of the SC, demonstrated that the addition of 10 wt% Glycerol: (1) reduces the average aqueous pore radius resulting from exposure of the skin to the aqueous SDS contacting solution from 33±5A to 20+5A, such that a SDS micelle of radius 18.5±1 A (as determined using dynamic light scattering (DLS) measurements) experiences significant steric hindrance and cannot penetrate into the SC, and (2) reduces the porosity-to-tortuosity ratio of the aqueous pores in the SC by more than 50%, thereby further reducing the ability of the SDS micelles to penetrate into the SC and perturb the skin barrier.

(cont.) In vitro skin electrical current measurements can be used effectively to rank aqueous contacting solutions containing surfactants and humectants (the enhancer), relative to a PBS aqueous contacting solution (the control), based on their ability to perturb the skin aqueous pores. Specifically, an in vitro ranking metric was introduced using the enhancement in the skin electrical current induced by an enhancer relative to the control. For this study, I considered aqueous contacting solutions of the following chemicals: (1) humectants - Glycerol and Propylene Glycol, (2) surfactants - SDS and C12E6 (Dodecyl Hexa (Ethylene Oxide)), and (3) a control - PBS. Utilizing the in vitro ranking metric, the aqueous solutions above contacting the skin were ranked as follows (from the mildest to the harshest): Glycerol < Propylene Glycol < PBS < C12E6 < SDS. In order to further develop this ranking methodology, which can potentially lead to the reduction of several costly operations associated with identifying surfactant/humectant systems which are mild to the skin, such as, in vivo clinical testing and trial-and-error screening, it was important to correlate the in vitro ranking metric findings with direct in vivo skin barrier measurements.

(cont.) For this purpose, in vivo soap chamber measurements were carried out on human subjects, using the aqueous surfactant/humectant solutions described above. The results of these in vivo measurements of skin barrier perturbation were found to be consistent with the ranking results obtained using the in vitro ranking metric for the aqueous surfactant and humectant contacting solutions considered. In addition, in vivo soap chamber measurements were carried out for aqueous SDS+Glycerol contacting solutions. These in vivo measurements indicated that adding Glycerol to a SDS aqueous contacting solution significantly mitigates SDS-induced in vivo skin barrier perturbation, which is consistent with the results of my in vitro skin electrical current and Mannitol skin permeability measurements. In order to visualize the effects of aqueous surfactant/humectant systems on the skin barrier, an in vitro dual-channel two-photon fluorescence microscopy (TPM) visualization study was carried out. TPM is a non-invasive imaging technique based on two-photon induced nonlinear excitations of fluorophores, with the capability for deep-tissue imaging (up to several hundred micrometers).

(cont.) The following aqueous surfactant and humectant contacting solutions were studied: (i) SDS, (ii) SDS+Glycerol, (iii) SCI (a mild surfactant), (iv) PBS control, and (v) Glycerol. Sulforhodamine B (SRB), which is a hydrophilic fluorescent probe, was used to probe the effect of aqueous contacting solutions (i)-(v) on the skin barrier morphology. The results of this TPM visualization study revealed that SDS induces corneocyte damage by denaturing keratins and creating intra-corneocyte penetration pathways. On the other hand, SDS+Glycerol did not significantly induce corneocyte damage. The dual-channel TPM images corresponding to aqueous contacting solutions (iii)-(v) showed low SRB penetration into the corneocytes, as well as localization of the SRB probe within the lipid bilayers surrounding the corneocytes of the SC. Through a quantification of the amount of SRB that penetrated into the skin as a function of the skin depth, I found that adding Glycerol to SDS could significantly reduce the SDS-induced penetration depth of SRB, which provides evidence of the ability of Glycerol to mitigate SDS-induced skin barrier perturbation.

(cont.) The fundamental understanding of surfactant-induced skin barrier perturbation in the presence of humectants developed in this thesis is of particular relevance to the cosmetic industry in enabling the formulation of mild, non-drying, skin-care products that contain surfactants and humectants. The novel TPM studies that visualize, as well as quantify, skin morphology upon exposure of the skin to surfactant/humectant systems, has the potential to be developed into a high-throughput imaging tool for the screening of new skin-care formulations. Such a strategy can simultaneously screen the skin-mildness potential of many skin-care formulations, thereby significantly speeding up the effort and time required to bring new skin-care formulations to the market. In addition to the practical impact on the formulation of mild skin-care products, this thesis has also advanced fundamental research carried out in the investigative dermatology and related health disciplines.