BACKGROUND: A major bottleneck in microalgal biodiesel production is lipid content, which is often low in microalgal species. The present study examines Chlorella vulgaris as a potential feedstock for biodiesel by identifying and evaluating the relationships between the critical variables that enhance the lipid yield, and characterizes the biodiesel produced for various properties. RESULTS: Factors affecting lipid accumulation in a green microalga, Chlorella vulgaris were examined. Multifactor optimization raised the lipid pool to 55% dry cell weight against 9% control. When C. vulgaris cells pre-grown in glucose ( 0.7%)-supplemented medium were transferred to the optimized condition at the second stage, the lipid yield was boosted to 1974 mg L(-1), a value almost 20-fold higher than for the control. The transesterified C. vulgaris oil showed the presence of similar to 82% saturated fatty acids, with palmitate and stearate as major components, thus highlighting the oxidative stability of C. vulgaris biodiesel. The fuel properties ( density, viscosity, acid value, iodine value, calorific value, cetane index, ash and water contents) are comparable with the international ( ASTM and EN) and Indian ( IS) biodiesel standards. CONCLUSION: C. vulgaris biomass with 55% lipid content and adequate fuel properties is potentially a renewable feedstock for biodiesel. (C) 2011 Society of Chemical Industry