Viral clearance is a primary concern for parenteral protein biopharmaceuticals. Low pH, detergent/solvent wash, or heating, called pasteurization, has been the main process for virus inactivation. Detergent/solvent wash is also used to treat superficial infectious diseases, including herpes simplex virus (HSV) infections. Herein we examined virus inactivation effects of acidic arginine on HSV type 2 (HSV-2) as a function of pH and temperature in an attempt to find solvent conditions that are effective for virus inactivation, yet are compatible with in vivo applications. Aqueous arginine at 0.7 M was highly effective on HSV-2, more so at lower pH and higher temperature. Its effects were stronger than 0.1 M citrate, 0.1 M citrate/0.6 M NaCl or 0.7 M citrate at any pH and temperature. This demonstrates that strong virus inactivation effects of arginine are not simply due to ionic strength or high concentration and arginine possesses a unique property that results in irreversible damage in virus particles. Such strong virus inactivation effects can be used in vivo for certain superficial infectious diseases, such as genital infections.