Human metmyoglobin was separated electrophoretically into four subfractions: Mb(1), Mb(2), Mb(3), and Mb(4), which divide into at least two biochemically independent groups: Mb(1) and Mb(2), and Mb(3), and Mb(4). In normal subjects, Mb(1) constituted the predominant component; Mb(2), Mb(3), and Mb(4) were the minor components in this descending order. In the Duchenne type of progressive muscular dystrophy, on the contrary, a remarkable decrease in Mb(1) and a concomitant increase in Mb(3) were observed. This unique abnormality in the relative distribution of myoglobin subfractions was recognized only in the Duchenne type and not in other types of progressive muscular dystrophy or in other myopathies.