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Cosmetic ingredients are often complex mixtures from natural sources such as botanical extracts that might contain minute amounts of constituents with sensitizing potential. The sensitivity of in vitro skin sensitization test methods such as KeratinoSens TM and h-CLAT for the detection of minute amounts of sensitizer in mixtures remains unclear. In this study, we assessed the detection sensitivity of the binary test battery comprising KeratinoSens TM and h-CLAT for minute amounts of sensitizers by comparing the LLNA EC3 (estimated concentration of a substance expected to produce a stimulation index of 3) values to the minimum detection concentrations (MDCs) exceeding the positive criteria for each of the two in vitro test methods. 146 sensitizers with both sets of in vitro data and LLNA data were used. MDC values for KeratinoSens TM and h-CLAT were calculated from exposure concentrations exceeding positive criteria for each in vitro test method (EC1.5 and minimum induction thresholds, respectively). The dilution rate used to expose culture medium was also considered. For 86% of analyzed sensitizers, the in vitro test methods showed MDC values lower than LLNA EC3 values, suggesting that the binary test battery with KeratinoSens TM and h-CLAT have greater sensitivity for detection of minute amounts of sensitizer than LLNA. These results suggest the high applicability of KeratinoSens TM and h-CLAT for detecting skin sensitizing constituents present in botanical extract.

Sensitivity of KeratinoSens<sup>TM</sup> and h-CLAT for detecting minute amounts of sensitizers to evaluate botanical extract