For the in vitro toxicological assessment of current and next generation products, including e-vapour products, the standard in vitro test battery developed for conventional combustible products needs further refinement in terms of sensitivity to ensure that the toxicological potential of such products can be reliably compared and is not underestimated. In particular, the trapping of any particulate matter from such products is more difficult due to increasing trapping times, concomitant ageing and increased water levels trapped. Consequently direct exposure-to aerosol technologies for different biological test systems have been developed.

Here we present a methodology developed to enhance the sensitivity of the OECD recommended Salmonella typhimurium TA100 reverse mutation assay (Ames test) for the assessment of e-vapour aerosols but also applicable to fresh cigarette smoke. In our standard exposure protocol the bacteria suspension is exposed directly to the aerosol which is lead through the bacteria suspension located in an impinger. Following the logic of former Ames test developments (e.g. micro-suspension version or liquid pre-incubation assay protocols), the probability of bacteria / fresh whole smoke contact was increased by increased cell density during exposure.

Using the CORESTA Monitor test piece CM7 and the Kentucky Reference cigarette 3R4F a correlation of bacteria cell density and the test sensitivity, as measured by the slope of the linear part of the dose response curve, could be shown. For example, for CM7 we found that a five-fold increase in cell density led to a 18-fold slope increase (from 0.2 rev./puff up to 3.7 rev./puff).

Due to low or missing responses to e-vapour products under standard Ames test conditions, a vapour-specific positive control was developed and tested. Hence, we present an adjusted product assessment for bacterial mutagenicity with increased test sensitivity for aerosol products.