Separation Of Anti-hyperglycemia And Antioxidant Active Ingredients From Purple Sweet Potato

With the change of lifestyle and dietary constituents, more and more people are suffering from diabetes mellitus. It is well known that the long term oxidative stress can lead to insulin resistance, which in turn can amplify the original damaging effect of hyperglycemia. Improving the impaired antioxidant status in patients is not only the key factor to preventing diabetes mellitus but also to reversing other disease, such as arteriosclerosis, alzheimer’s disease and inflammation. A recent study have reported that normalizing mitochondrial superoxide production blocks three pathways of hyperglycaemic damage, including:glucose-induced activation of protein kinase C isoforms; increased formation of glucose-derived advanced glycation end-products and increased glucose flux through the aldose reductase pathway. Pharmacological study have demonstrated that administration of hiamine derivative-benfotiamine can inhibit these three pathways, as well as hyperglycemia-associated NF-κB activation, by activating the pentose phosphate pathway enzyme transketolas, and other antioxidants alsopossess promising antidiabetic activity. Purple sweet potato, originated from Japan, is now widely grown in the world. There is a high content of anthocyanin pigments (mainly are peonidin and cyanidin) in the tuber of purple sweet potato. As one of the most widely used foods throughout the world, the the functional properties of purple sweet potato attracted the most interest. Many studies mainly focused on only separation and in vitro study of anthocyanins in purple sweet potato. And it is reported that anthocyanins exhibited various biological activities such as scavenging free radicals, antioxidant, anti-tumor, anti-hyperglycemia effect. However, in both animals and humans, the efficiency of anthocyanins absorption is relatively poor due to their rapid absorption and elimination. Additionally, epidemiological studies have not revealed protective effects of anthocyanin consumption on cancer risk in humans. Previous studies have suggested that other constituents except anthocyanins in purple sweet potato also possess pharmacological properties. So, it is necessary to separate and identify other bioactive compounds in purple sweet potato。In this study, the anti-hyperglycemia activity of purple sweet potato was evaluated using a-glucosidase and cell assay. The active fractions were further separated by column chromatography and high speed counter-current chromatography (HSCCC).The main contents of this study are as follows:(1) The a-glucosidase inhibitory activity of different tissues (root, stem, leaves) of purple sweet potato of four varieties were analyzed. Results showed that all the extracts of different parts of purple sweet potato have certain a-glucosidase inhibitory activity, and the a-glucosidase inhibitory activity vary with tissues and varieties. The root and stem extract of Zishul5-6have shown higher inhibitory activity against a-glucosidase than acarbose. The glucose consumption promoting activities of purple sweet potato extracts were evaluated by cell assay. The results indicated that the ethyl acetate fraction and N-butyl alcohol fraction showed strong glucose consumption promoting activity.(2) The n-buthanol fraction of purple sweet potato was further purified by D101macroporous resin column, and separated by high speed counter-current chromatography (HSCCC) using chloroform-methanol-water (10:8:3, v/v/v) as solvent system. Peonidin was successfully separated from80%and100%ethanol eluted fraction.(3)The thin layer chromatograph coupling with fluorometric (TLC-F) method was used to determine the partition coefficient of target compounds in HSCCC solvent system. Two components,6,7-dimethoxycoumarin and5-hydroxymethyl-2-furfural were successfully separated from purple sweet potato extracts by successive sample injection for the first time, using n-hexane-ethyl acetate-methanol-water (1:2:1:1, v/v/v/v) as the solvent system.(4). Mitochondria in mammalian cells were used as reactive oxygen species initiator and the inhibitory degree of DCFH oxidation by antioxidants scavenging ROS, generated in mitochondria, was used as the basis for calculating antioxidant activity. The results suggested that this method can be used to assess the pro-and antioxidant properties of compounds and mixtures from fruit and herb extracts. And the assay has more physiological relevance and acceptable accuracy, precision, and reproducibility. Unlike ABTS assay, the antioxidant activity obtained by mitochondria and cell-based assay are strongly positively correlated.