In photosynthetic bacteria, after transition to light-anaerobic and nitrogen-deficient conditions, hydrogen evolution starts with expression of nitrogenase activity. Until the expression of enough activity, Rhodovulum sulfidophilum consumed substrates and converted them to poly(3-hydroxybutyrate) (PHB), resulting in a decrease in the proportion of substrate converted into hydrogen gas. To prevent conversion to PHB during the period when nitrogenase activity is derepressed, the authors employed a cultivation method consisting of three stages : cell growth, nitrogenase derepression, and hydrogen production. Cells cultivated by this method exhibited no lag time before the commencement of hydrogen evolution and gave an improved yield of hydrogen from the algal fermentative products.