A Comparative Study on the Inhibitory Effects of Different Parts and Chemical Constituents of Pomegranate on a-Amylase and a-Glucosidase Antony Kam, 1 Kong M. Li, 2 Valentina Razmovski-Naumovski, 1,3,4 Srinivas Nammi, 1,3,4 Jeffrey Shi, 5 Kelvin Chan 1,3,4 and George Q. Li 1 * 1 Faculty of Pharmacy, University of Sydney, NSW 2006, Australia 2 Discipline of Pharmacology, Sydney Medical School, University of Sydney, NSW 2006, Australia 3 Centre for Complementary Medicine Research, University of Western Sydney, NSW 2560, Australia 4 School of Science and Health, University of Western Sydney, NSW 2560, Australia 5 School of Chemical and Biomolecular Engineering, University of Sydney, NSW 2006, Australia Pomegranate has been documented for the management of diabetes in Unani and Chinese medicine. This study compared the effects of the extracts of different pomegranate parts, including juice, peels, seeds and flowers, on carbohydrate digestive enzymes (a-amylase and a-glucosidase) in vitro. The methanolic flower extract inhibited a-amylase and a-glucosidase, while the methanolic peel extract inhibited a-glucosidase selectively. The most active flower extract was subjected to water-ethyl acetate partition. The ethyl acetate fraction was more potent than the water fraction in inhibiting both enzymes. Gallic acid and ellagic acid also showed selective inhibition against a-glucosidase, and their presence in the ethyl acetate fraction was confirmed by HPLC-DAD and HPLC-HESI-MS. Our findings suggest that the inhibition of carbohydrate digestive enzymes and their phenolic content may contribute to the anti-hyperglycaemic effects of pomegranate flower and peel, and support their claims in diabetes. Copyright © 2012 John Wiley & Sons, Ltd. Keywords: a-amylase; a-glucosidase; diabetes; pomegranate; Punica granatum. INTRODUCTION Diabetes mellitus is a major health problem globally. More than 171 million people worldwide have diabetes, and the prevalence is expected to increase dramatically to approximately 366 million by 2030 (Wild et al. 2004). Diabetes is commonly characterised by hypergly- caemia in both postprandial and fasting state, as a con- sequence of impaired insulin secretion with or without insulin resistance. A therapeutic approach to attenuate postprandial spikes is to inhibit carbohydrate digestive enzymes. This delays the digestion of non-absorbable polysaccharides or disaccharides to absorbable mono- saccharide, subsequently reducing their gastrointestinal absorption, and hence, glycaemic variability is minimised (Tadera et al. 2006). a-Amylase and a-glucosidase are the two commonly targeted carbohydrate digestive enzymes in human. a-Amylase is produced from the salivary glands and pancreas. The enzyme catalyses the hydrolysis of polysaccharides (e.g. starch) to disaccharides (e.g. dextrin and maltose) from the mouth to the small intes- tine. a-Glucosidase, which is predominantly located in the epithelial cells within the small intestine brush-border membrane, can further hydrolyse disaccharides (e.g. sucrose and maltose) to glucose and other monosaccharides for gastrointestinal absorption (Li et al. 2005). Punica granatum Linn., commonly known as pome- granate, belongs to the Punicacae family (Li et al. 2005). Pomegranate, particularly the flower and peel, has been traditionally consumed for diabetes mellitus according to Unani literature, and is also used in some parts of China (Li et al. 2004; Li et al. 2008). Recent studies demonstrate that the flower, peel and seeds of pomegranate exhibit hypoglycaemic activities in several rodent models (Das et al. 2001; Huang et al. 2005; Jafri et al. 2000; Katz et al. 2007; Li et al. 2005; Parmar and Kar 2007). In particular, the flowering part showed positive effects against postpran- dial hyperglycaemia, potentially linked with a-glucosidase inhibition (Li et al. 2005). However, the effects of pome- granate flower and other pomegranate parts (e.g. peel, juice and seeds) on mammalian-type a-glucosidase and a-amylase have not been examined. In addition, the chemical constituents in pomegranate responsible for the inhibitory effects are still unknown. Recent studies demonstrated a positive correlation of the phenolic content from apples and Gymnema montanum leaves with the inhibitory activities against a-glucosidase and a-amylase (Barbosa et al. 2010; Ramkumar et al. 2010). Since pomegranate contains an exceptionally high phenolic content, the present study evaluates the effects of the extracts of different pomegranate parts, including the juice, peels, seeds and flowers, on porcine pancreatic a-amylase and rat intestinal a-glucosidase enzyme activities in vitro. MATERIALS AND METHODS Plant materials. Pomegranate flowers were collected in Maharashtra, India, while pomegranate fruits were * Correspondence to: Dr George Q. Li, Faculty of Pharmacy, University of Sydney, NSW 2006, Australia. E-mail: george.li@sydney.edu.au PHYTOTHERAPY RESEARCH Phytother. Res. (2012) Published online in Wiley Online Library (wileyonlinelibrary.com) DOI: 10.1002/ptr.4913 Copyright © 2012 John Wiley & Sons, Ltd. Received 19 August 2011 Revised 15 November 2012 Accepted 25 November 2012