Understanding the respective roles of UV photons and radicals in cold plasma sterilization

Summary form only given. Low-temperature plasma sterilization can be achieved under direct contact with the discharge or in its afterglow. The results to be presented were obtained with B. Subtilis spores subjected to the flowing afterglow of a microwave discharge, at pressures typically below 10 Torrs. As a rule, the sterilization cycle, characterized by the corresponding survival curve (logarithm of the number of living spores as a function of exposure time), takes place in three distinct steps (called phases), each presenting a specific kinetics of inactivation. The analysis of these diagrams as functions of the operating conditions enables one to identify the two principal physico-chemical processes leading to plasma sterilization, namely irradiation of the spore DNA by UV photons and spore erosion by free radicals (e.g., oxygen atoms). The UV photons are emitted by atoms or molecules excited in the afterglow through collisions. Among those directed toward the spore, only a fraction actually penetrates into the spore up to its genetic material, participating in its alteration. A spore is ultimately inactivated when the number of lesions caused to its DNA strands is sufficient to prevent the spore from germinating when put back in a favorable environment. During the first phase, almost all of the isolated spores are inactivated by UV photons, usually in a relatively short period of time (=10 minutes). However, statistically, one spore in a thousand presents a particular resistance to UV irradiation, leading to a considerable slowing down of the inactivation process, well reflected by the longer characteristic time of the second phase of the cycle. For example, spores that are partially or totally covered by other spores or debris of all kinds present a "thickness" superior to that of a single average microorganism. In this case, the erosion process, induced by oxygen atoms forming volatile compounds with the spore material, is of particular importance. It reduces progressively the distance the photons have to travel within the spore to reach DNA, therefore contributing to an increase of the number of UV photons hitting the genetic material per unit time. Most of our previous work has been achieved with an N/sub 2/-O/sub 2/ post-discharge, where the UV photons mainly result from excited NO molecules, whose population is strongly dependent on the O/sub 2/ percentage in the N/sub 2/-O/sub 2/ mixture. In such a case, it is difficult to fully discriminate damage to the spore due to UV irradiation from that resulting from its erosion (etching) due to oxygen atoms. To separate these two effects, we have used noble gas/O/sub 2/ mixtures, where no O/sub 2/-containing molecules are formed with the carrier gas that could further emit UV photons. Survival curves obtained with the noble gas alone and in the presence of O/sub 2/ seem to indicate that oxygen atoms actually play a secondary role in the spore inactivation, in accordance with our observations in N/sub 2/-O/sub 2/ mixtures.