In the present study, efforts have been made to optimize the three physical process variables viz; pH, temperature and agitation speed for enhanced polyhydroxybutyrate (PHB) production in batch cultivation by Alcaligenes sp. which serves as precursor for bioplastic (PHB) production. Strain selection was done by viable staining method using nile blue A dye. Agro-industrial by products; cane molasses and urea were used as carbon and nitrogen source for PHB production. Optimization of physical process variables was done by central composite rotatable design (CCRD) using design expert (DX 8.0.6) software. Shake flask cultivation performed under optimum physical condition viz; 34.5 degrees C temperature, 6.54 pH and agitation speed of 3.13 Hz, gave PHB mass fraction yield of 76.80% on dry molasses substrate and showed 98.0% resemblance with the predicted percentage yield of 77.78%. Batch cultivation further performed in 7.5 L lab scale bioreactor (working volume: 5.6 L) under optimized condition gave maximum cell biomass of 11 +/- 0.5 g L (1) with a PHB content of 8.8 +/- 0.4 g L-1 after 48.0 h of fermentation. Scale up study on bioreactor gave maximum PHB yield (Y-P/X) and productivity of 0.78 and 0.19 g L-1 h, which are higher than previous reports under similar condition. Characterization of PHB was done by FTIR. (C) 2013 Elsevier Ltd. All rights reserved.