Dermal absorption and toxicological risk assessment : pitfalls and promises

Absorption of toxic substances via the skin is an important phenomenon in the assessment of the risk of exposure to these substances. People are exposed to a variety of substances and products via the skin, either directly or indirectly, while at work, at home or in public space. Pesticides, organic solvents and metalworking fluids are seen to be important contributors to adverse health effects due to occupational exposure via the skin. In daily life, cosmetics, clothing and household products are the most relevant commodities with respect to exposure via the skin. Given the importance of skin exposure in the assessment of the risk of toxic substances, the objective of this thesis was to further develop, evaluate and improve methods for including skin absorption data this assessment. In this thesis, four factors influencing dermal absorption, namely dermal loading (chapters 3 and 6), irritative/corrosive potential (chapters 3 and 4), frequency of exposure (chapters 3, 4 and 5) and the vehicle used (chapter 5), were investigated in more detail. Furthermore, a model to extrapolate infinite dose absorption data to finite dose conditions, baptized Dermal Absorption Model for Extrapolation (DAME), was developed and tested. I n <em>chapter 2</em> of this thesis, the relationship between relative dermal absorption and dermal loading was investigated. Hundred-and-thirty-eight dermal publicly available absorption experiments with 98 substances were evaluated. The results obtained revealed that dermal loading ranged mostly between 0.001 and 10 mg/cm2. In 87 experiments (63%), an inverse relationship was observed between relative dermal absorption and dermal loading. On average, relative absorption at high dermal loading was 33 times lower than at low dermal loading. Known skin irritating and volatile substances less frequently showed an inverse relationship between dermal loading and relative absorption. It was concluded that when using relative dermal absorption in regulatory risk assessment, its value should be determined at or extrapolated to dermal loadings relevant for the exposure conditions being evaluated. I n <em>chapter 3</em> of this thesis, a literature search was presented with the aim to investigate whether neglecting the effects of repeated exposure may lead to an incorrect estimate of dermal absorption. The results demonstrated that the effect of repeated versus single exposure does not demonstrate a unique trend. Nevertheless, an increase in daily absorption was frequently observed upon repeated daily exposure. The little information available mostly concerned pharmaceuticals. However, consumers and workers may be repeatedly exposed to other types of chemicals, like disinfectants and cleaning products, which often contain biocidal active substances that may decrease the barrier function of the skin, especially after repeated exposure. These biocidal products, therefore, may present a safety risk that is not covered by the current risk assessment practice since absorption data are usually obtained by single exposure experiments. Consequently, it was decided to investigate the importance of this issue for biocide safety evaluation. As the literature search revealed that hardly any data on absorption upon repeated dermal exposure to biocides are available, it was concluded that data need to be generated by testing. To cover the entire range of biocidal products in such testing, a representative series of biocidal substances should be tested, making <em>in vitro</em> testing of dermal absorption the preferred choice over <em>in vivo</em> testing. Based on an inventory made, it appeared that the 16 product types represented among the biocidal products authorised in the Netherlands could be clustered into 6 more or less homogeneous categories based on similarity in active substances. This result could facilitate experimental testing by providing a basis for selection of a limited number of representative compounds to be evaluated. I n <em>chapter 4</em> of this thesis, the importance of the effect of repeated dermal exposure on skin permeability for biocide safety evaluation was investigated, using a selection of nine representative biocides from the inventory made in chapter 3. The <em>in vitro</em> dermal penetration of tritiated water and [14C]propoxur was chosen as a measure of the permeability and integrity of human abdominal skin after single and repeated exposure. The results indicated that single and repeated exposure to specific biocidal products (e.g. the quaternary ammonium chlorides DDAC and ADBAC) may significantly increase skin permeability, especially when the compounds are applied at high concentrations, while a substance like formaldehyde may reduce skin permeability under specific conditions. I n <em>chapter 5</em> of this thesis, the <em>in vitro</em> dermal absorption kinetics of the quaternary ammonium compound didecyldimethylammonium chloride (DDAC) during single and repeated exposure was studied in more detail. In addition, the influence of biocidal formulations on the absorption of DDAC was investigated, because it was expected that formulation characteristics may be another factor influencing its dermal absorption. The analysis of biocidal products on the Dutch market, reported in chapter 3, indicated that DDAC is often used in combination with other active ingredients. DDAC was most frequently combined with formaldehyde, glutaraldehyde and/or alkyldimethylbenzyl­ammo­nium chloride (ADBAC). Consequently, commercial formulations containing one or more of these additional active ingredients were selected, in addition to one formulation containing only DDAC as an active ingredient. The selected commercial formulations tended to reduce skin penetration of DDAC. This was most pronounced with the formulation containing the highest concentration of formaldehyde (196 mg/mL) and glutaraldehyde (106 mg/mL), which reduced the flux of DDAC across the skin by 95%. The reduction caused by the only tested formulation containing no other active ingredients than DDAC, and thus incorporating no aldehydes, was smallest, and did not reach statistical significance. I n <em>chapter 6</em> of this thesis, a simple <em>in silico</em> model to predict finite dose dermal absorption from infinite dose data (<em>kp</em> and lag time) and the stratum corneum/water partition coefficient (<em>KSC,W</em>) was developed. This model was tentatively called <strong>D</strong>ermal Absorption Model for Extrapolation (DAME). As dermal exposure may occur under a large variety of conditions leading to quite different rates of absorption, such a predictive model using simple experimental or physicochemical inputs provides a cost-effective means to estimate dermal absorption under different conditions. To evaluate the DAME, a series of <em>in vitro</em> dermal absorption experiments was performed under both infinite and finite dose conditions using a variety of different substances. The <em>kp</em>’s and lag times determined in the infinite dose experiments were entered into DAME to predict relative dermal absorption value under finite dose conditions. For six substances, the predicted relative dermal absorption under finite dose conditions was not statistically different from the measured value. For all other substances, measured absorption was overpredicted by DAME, but most of the overpredicted values were still lower than 100%, the European default absorption value for the tested compounds. In conclusion, our finite dose prediction model (DAME) provides a useful and cost-effective estimate of <em>in vitro</em> dermal absorption, to be used in risk assessment for non-volatile substances dissolved in water at non-irritating concentrations. I n <em>chapter 7</em> of this thesis, the results of the research reported in chapters 2 to 6 were put into perspective, the pitfalls and promises emanating from them discussed and general conclusions drawn. The possible influence of vehicles on absorption and the possible impact of irritative or corrosive vehicles or chemicals on the skin barrier have been demonstrated in this thesis. An <em>in silico</em> predictive model tentatively called DAME was developed, which enables the user to evaluate a variety of dermal exposure scenarios with limited experimental data (<em>kp</em> and lag time) and easy to obtain physicochemical properties (MW and log <em>KOW</em>). The predictions of our experiments reported in chapter 6 were compared to those of the Finite Dose Skin Permeation (FDSP) model published on the internet by the US Centers for Disease Control and Prevention (CDC). DAME outperformed FDSP (R2 of the correlation predicted/measured potential absorption 0.64 and 0.12, respectively). At present, the applicability domain of DAME is limited to non-volatile substances dissolved in aqueous solvents. However, in future the model will be adapted to include volatile substances as well. Altogether, it is concluded that dermal exposure can be an important factor in risks posed by chemicals and should be taken into account in risk assessment. The methods to actually do this are still open for further improvement to better account for the various factors influencing skin penetration and to develop adequate combinations of <em>in vitro </em>and <em>in silico</em> models that can accurately predict human dermal absorption.