Azar, C.G., McDonnell, G.E., Pretzer, D. Steris Corp. P.O.Box 147, St. Louis, MO 63166

The development of an in vitro irritation assay producing results which correlate well with those obtained from in vivo animal studies would be a valuable tool for the evaluation of topical products. Human skin equivalent cultures are currently used as in vitro models to predict the topical irritation potential of actives, excipients, and formulations. These models consist of normal, neonatal human-derived epidermal keratinocytes (HNEK), grown under conditions and on various substrates which allow the keratinocytes to differentiate into a model of the human epidermis. The model used for this work (EpiDerm™ from MatTek) consists of keratinocytes growing on a porous synthetic membrane in a tissue culture insert. This model is easy to handle, relatively inexpensive, and produces low levels of lot to lot variability. In response to exposure to irritants, keratinocytes have the ability to produce and release pro-inflammatory cytokines. These factors are responsible for the primary epidermal response to irritants and sensitizers which consists of erythema (redness), edema (swelling) and immune cell infiltration at the affected area of skin (2). Levels of Interleukin-1α (IL-1α), a pro-inflammatory factor released by keratinocytes which plays a major role in the initiation and maintenance of inflammation and the irritation response (2), were quantitated. Our studies also included the in vitro assessment of cell viability as a marker for irritation because some irritants can also cause cell death. MTT is a dye which changes color in the presence of active mitochondria and the MTT assay, which assesses cell viability, can be conveniently performed using in vitro skin models. Chlorhexidene Gluconate (CHG) and iodine are antimicrobial actives widely used in the health care industry. They are present in surgical preps, scrubs and personnel handwashes. The goal of this study was to compare in vitro human skin model and in vivo animal irritation testing results for these two antimicrobial actives. Results show that CHG, although irritating in vitro, is not irritating by it self in vivo. Results of in vitro and in vivo testing of iodine products correlate well and indicate that iodine by itself is generally not irritating. Other ingredients typically present in these health care products do cause irritation in both in vitro and in vivo tests.


Anti-microbial, Anti-microbial actives, Antimicrobial, Antimicrobial actives, Antiseptics, CHG, Chlorhexidene Gloconate, Cytokines, Draize, EpiDerm, IL-1a, Interleukin (IL), Irritants, MTT, MTT ET-50 tissue viability assay, MTT assay, Pre-validation, Prevalidation, Serine protease inhibitor, Serine protease sensitizers, Validation

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