High-Resolution alpha-Amylase Assay Combined with High-Performance Liquid Chromatography Solid-Phase Extraction-Nuclear Magnetic Resonance Spectroscopy for Expedited Identification of alpha-Amylase Inhibitors: Proof of Concept and alpha-Amylase Inhibitor in Cinnamon

Type 2 diabetes affects millions of people worldwide, and new improved drugs or functional foods containing selective alpha-amylase inhibitors are needed for improved management of blood glucose. In this article the development of a microplate-based high-resolution alpha-amylase inhibition assay with direct photometric measurement of alpha-amylase activity is described. The inhibition assay is based on porcine pancreatic alpha-amylase with 2-chloro-4-nitrophenyl-alpha-d-maltotriose as substrate, which this gives a stable, sensitive, and cheap inhibition assay as requested for high-resolution purposes. In combination with HPLCHRMSSPENMR, this provides an analytical platform that allows simultaneous chemical and biological profiling of alpha-amylase inhibitors in plant extracts. Proof-of-concept with an artificial mixture of six compoundsof which three are known alpha-amylase inhibitorsshowed that the high-resolution alpha-amylase inhibition profiles allowed detection of sub-microgram amounts of the ?-amylase inhibitors. Furthermore, the high-resolution alpha-amylase inhibition assay/HPLCHRMSSPENMR platform allowed identification of cinnamaldehyde as the alpha-amylase inhibitor in cinnamon (Cinnamomum verum Presl.).