The objective of this study was to compare and assess various thermal sterilization methods of environment based on the inactivation of Geobacillus stearothermophilus spores present therein. Gebacillus stearothermophilus ATCC 10149 spores were suspended in an water tryptone solution (5 g / 100 mL) and in a citrate buffer. The spores studied were thermally treated in glass capillary tubes; two variants of this thermal treatment were considered: one-stage and two-stage variants. Under the onestage variant, a sterilization process was applied at 1250 C. Under the two-stage variant, a preliminary thermal treatment (115 °C for 6 minutes) was added and preceded the sterilization process at 125 °C. During the one-stage variant of the research project, it was proven that at a higher temperature (125 °C), the application of low pH (4.0) of the medium under sterilization always accelerated the inactivation process of the spores suspended therein; however, a survival curve tailing might occur, thus, forcing the considerable extension of the duration time of sterilization. Based on the progressing of spore inactivation processes during the sterilization carried out using different methods, it was shown that the use of the preliminary thermal treatment always eliminated a shoulder occurring on the survival curve at the initial stage of the process. Then again, no event was reported where the tailing appearing on those survival curves was fully eliminated. It was found that the introduction of a preliminary thermal treatment prior to the main sterilization process was not beneficial since it did not cause the duration time of sterilizing medium at 125 °C to be reduced. An obstacle to hinder any significant sterilization time reduction are the extraordinarily heat-resistant cells present in the population of spores. Even after the application of preliminary thermal treatment, a higher temperature should be applied for a longer time in order to activate those cells compared to other spores present in the population.