This research focused on optimizing the upstream process time for production of polyhydroxybutyrate (PHB) from sucrose by two-stage batch and fed-batch fermentation with Alcaligenes latus ATCC 29714. The study included selection of strain, two-stage batch fermentations with different time points for switching to nitrogen limited media (14, 16 or 18 h) and fed-batch fermentations with varied time points (similar to two stage) for introducing nitrogen limited media. The optimal strain to produce PHB using sucrose as carbon source was A. latus ATCC 29714 with maximum-specific growth rate of 0.38 ± 0.01 h(-1) and doubling time of 1.80 ± 0.05 h. Inducing nitrogen limitation at 16 h and ending second stage at 26 h gave optimal performance for PHB production, resulting in a PHB content of 46.7 ± 12.2 % (g PHB per g dry cell weight) at the end of fermentation. This was significantly higher (P ≤ 0.05) (approximately 7 %) than the corresponding fed batch run in which nitrogen limitation was initiated at 16 h.