The production of biodiesel from edible oils may cause negative impact to any country through food crisis which may lead to economic imbalance. Hence, this study focuses on viability of extracting the oil from the Citrus limetta seeds for biodiesel production for the first time. Composition of C. limetta oil was determined by gas chromatography. C. limetta biodiesel was produced by simple transesterification process, and further physiochemical properties were analyzed as per the standards. This study also describes the suitable characterization and optimization parameters used for conversion of C. limetta seed oil into biodiesel.