Retrograded starch is known to be resistant to digestion. We used enzyme kinetic experiments to examine how retrogradation of starch affects amylolysis catalysed by porcine pancreatic amylase. Parallel studies employing differential scanning calorimetry, infra red spectroscopy, X-ray diffraction and NMR spectroscopy were performed to monitor changes in supramolecular structure of gelatinised starch as it becomes retrograded. The total digestible starch and the catalytic efficiency of amylase were both decreased with increasing evidence of retrogradation. A purified sample of retrograded high amylose starch inhibited amylase directly. These new findings demonstrate that amylase binds to retrograded starch. Therefore consumption of retrograded starch may not only be beneficial to health through depletion of total digestible starch, and therefore the metabolisable energy, but may also slow the rate of intestinal digestion through direct inhibition of alpha-amylase. Such physiological effects have important implications for the prevention and management of type 2 diabetes and cardiovascular disease. (C) 2017 The Authors. Published by Elsevier Ltd.