This study investigated two different strategies for enrichment using crude glycerol and mixed culture. Coupled feeding allowed simultaneous supplies of glycerol and nutrients at start of feast, whereas uncoupled feeding postponed nutrient supply until start of famine. Adaptation of culture enriched by the uncoupled feeding strategy was faster with higher operational stability. This however resulted in low PHB content of 20% and yield of 0.137 mg PHB/mg glycerol in feast phase, cell growth at yield of 0.298 mgX/mg PHB in famine. With more abundance (29.2%) of PHA storing bacteria in culture enriched by the coupled feeding strategy as revealed from 16 S rRNA analysis, results showed that polymer storage was dominated in feast phase (max. PHB content of 68%, yield of 0.563 mg PHB/mg glycerol), leading to high cell growth yield of 0.745 mg X/mg PHB. PHB accumulation by the enriched culture from coupled feeding has achieved 82% PHB content with 1094 mg PHB/L.h volumetric productivity. Thus, the coupled feeding was more effective to enrich PHA storing culture for PHA production using crude glycerol via two-step process.