The isolation and identification of 1,6-phenazinediol-5-oxide from Microbispora aerata, Pseudomonas iodina, and Streptomyces thioluteus is reported. The title compound was also synthesized by the partial reduction of 1,6-phenazinediol-5,10-dioxide (iodinin) with sodium hydrosulfite. The antimicrobial activity of 1,6-phenazinediol-5-oxide is intermediate between that of iodinin and phenazinediol. Experiments with cell suspensions indicate that 1,6-phenazinediol-5-oxide is an intermediate both in the biosynthesis of iodinin from 1,6-phenazinediol and in the microbial reduction of iodinin.