Anti-hyperglycemic activity of HPLC-fractionated Momordica charantia seed extract enriched in a novel napin-like protein in experimental diabetic rats and its validation with recombinant napin-like protein

Momordica charantia, a tropical plant, is used in traditional medicine to treat multiple ailments, including diabetes. Several studies have identified compounds with anti-hyperglycemic activity in crude or partially purified M. charantia extracts. However, systematic identification and purification studies validating the bioactivity of active components are unavailable to date. The present study aimed to use an extensive bioactivity guided purification strategy to identify and characterize the proteinaceous active components of M. charantia. The purification scheme, involving acid–ethanol extraction of M. charantia seeds, fractionation by isoelectric precipitation, gel filtration, and reverse-phase HPLC, yielded an anti-hyperglycemic protein fraction consisting of two heterodimeric proteins with molecular masses of 11.70 kDa and 11.47 kDa. Further analyses confirmed that both proteins had characteristics of napin-like proteins. Administration of the purified protein fraction to diabetic animals significantly decreased blood glucose. Acute toxicity studies involving repeated administration of the active peak fraction to experimental animals did not indicate any hepatotoxicity or nephrotoxicity. Furthermore, we evaluated the anti-hyperglycemic activity of a recombinant napin-like protein of M. charantia in diabetic animals, thus confirming our findings that purified napin-like proteins have anti-hyperglycemic activity.