In the pharmaceutical, cosmetic, chemical, and food industries high-pressure homogenization is used for the preparation or stabilization of emulsions and suspensions, or for creating physical changes, such as viscosity changes, in products. Another well-known application is cell disruption of yeasts or bacteria in order to release intracellular products such as recombinant proteins. The development over the last few years of homogenizing equipment that operates at increasingly higher pressures has also stimulated research into the possible application of high-pressure homogenization as a unit process for microbial load reduction of liquid products. Several studies have indicated that gram-negative bacteria are more sensitive to high-pressure homogenization than gram-positive bacteria supporting the widely held belief that high-pressure homogenization kills vegetative bacteria mainly through mechanical disruption. However, controversy exists in the literature regarding the exact cause(s) of cell disruption by high-pressure homogenization. The causes that have been proposed include spatial pressure and velocity gradients, turbulence, cavitation, impact with solid surfaces, and extensional stress. The purpose of this review is to give an overview of the existing literature about microbial inactivation by high-pressure homogenization. Particular attention will be devoted to the different proposed microbial inactivation mechanisms. Further, the different parameters that influence the microbial inactivation by high-pressure homogenization will be scrutinized.