Sterilization is a process that eliminates or kills all microbes, including transmissible agents present on a surface, contained in a fluid, medication or biological culture media (UCLA Department Epidemiology). Dry heat sterilization is one of the important methods of eliminating microbes, as it coagulates the proteins in an organism, causing oxidative free radical damage thus leading to cell death. Heat is lethal to micro organisms, but each species has its own particular heat tolerance. This study is based on effect of sterilization, at varied time periods as well as constant temperature and pressure conditions, on sporulating and non-sporulating bacteria. Bacillus subtilis and Escherichia coli cultures were inoculated in nutrient media and incubated. After incubation, the concentration of bacterial cells in the media, before and after sterilization at 80°C in hot air oven, was found out. Bacterial cultures were subjected to sterilization and OD values of both the cultures were taken at an interval of 10 minutes, using UV Spectrophotometer. The reaction constant (K) and sterilization reaction rates were calculated from the obtained data, for both cultures and an increased sterilization rate was observed for E.coli. A reaction equation was also designed for sterilization reaction for both cultures. This study would help us understand the dynamics of sterilization and methods of varying specified conditions in order to increase the rate of sterilization of pathogenic, sporulating bacteria.