The antimicrobial and sporicidal activities in vitro and in vivo of 1% peroxygen (Virkon) and 2% alkaline glutaraldehyde (Asporin) were evaluated on dental instruments before and after cleaning. The in vitro antimicrobial activity against vegetative bacteria, bacterial spores and fungi indicated that glutaraldehyde is more active against these organisms than peroxygen. Asporin killed all vegetative bacteria within 1 min after cleaning, whereas Virkon was active, in the majority of cases, within 15 min and obtained a greater than 105-fold reduction in count before killing for the vast majority of instruments, and for all micro-organisms. The spores of Bacillus subtilis were killed by Asporin within 4-5 h after cleaning, whereas Virkon required almost 20 h. A meticulous instrument cleaning process followed by an appropriate disinfection treatment assures a shorter disinfection time. Asporin should be recommended for chemical sterilization or high-level disinfection of dental instruments, and Virkon, if only disinfection is required, would seem to be a possible alternative, even if used with a higher exposure time.