Using volatile fatty acids (VFA) from the anaerobic digestion liquor of sewage sludge, up to 9.2 mM 5-aminolevulinic acid (ALA) could be produced by Rhodobacter sphaeroides under anaerobic-light (5 kLux) conditions with repeated addition of levulinic acid (LA) and glycine and using a large inoculum (approx. 2 g/l of cells, initially from glutamate/malate medium). As the VFA medium also contained organic nitrogen sources such as glutamic acid, the cells were later grown up to about 2 g/l in the VFA medium instead of the glutamate/malate medium. ALA production was then again promoted by adding LA and glycine. Using this improved method, up to 9.3 mM ALA was produced by feeding propionate and acetate together with LA and glycine, indicating that VFA medium formed from sewage sludge could be useful for ALA production.