Studies On Isolation, Purification And Antioxidant Activities Of Polyphenols In Leaf Of Lonicera Japonica

Polyphenols in leaf of Lonicera Japonica possess broad physiological activities, such as antioxidation, inhibition of cardiovascular and cerebrovascular diseases, antimicrobial, antitumor, protect the liver and so on. Leaf of Lonicera Japonica was found to be a rich source of phenolic compounds. In order to better exploit and utilize the rich Lonicera Japonica leaf resource, this paper studies the HPLC detection conditions and fingerprint characteristics of Lonicera Japonica leaf active substances from different acquisition months and different regions (Jiangxi, Shandong and Henan), the isolation of polyphenols into phenolic acids and flavonoids and antioxidant activity of total polyphenols, chlorogenic acids and flavonoids.1The chromatographic separations of active substances in leaf of Lonicera Japonica were obtained by Alltima C18column (250mm×4.6mm,5μm) using gradient elution with acetonitrile and0.4%phosphoric acid aqueous solution. The temperature of column was at40degrees. Detection wavelength was set at238nm. The flow rate was0.7mL/min. The injection volume of sample was10μL. Detection time was55minutes. The extraction method of Lonicera Japonica leaf affects the content of its active substances. Extraction efficiency of the sample extracted with50%methanol in an ultrasonic water bath for30min is best. Optimization method is precise, stable and reproducible enough for quantitative evaluation.2The contents of active substances of Lonicera Japonica leaf can vary depending on geographical source, cultivation conditions, harvest time. With chlorogenic acid as the reference peak, the contents of active substances of Lonicera Japonica. leaf in different growth periods are some different and present some rules. The similarity values of the chromatogram of May, June and July with the contrast are all below0.90and other months’similarity values are all higher and above0.90. The peak2-chlorogenic acids and peak6are chosen as the standard with a consistently high content, the best harvest time of Lonicera Japonica leaives is July. HPLC chromatogram of Lonicera Japonica leaf polyphenols from Shandong Pingyi, Henan Mixian and Jiangxi Nanchang shows that Shandong Pingyi and Henan Mixian have a high similarity values of0.929each other; And at the same time, the contents of their peak3and peak4are more. While, the content of peak2-chlorogenic acids of Lonicera Japonica leaf from Jiangxi is the most.3Taking polyphenols in leaf of Lonicera Japonica as raw material, its purity is71.24%. It studies NKA-Ⅱ resin of the separation of phenolic components and flavonoid components and HPD500macroporous resin and Sephadex LH-20gel resin for further separation and purification of chlorogenic acid. In the initial separation of NKA-Ⅱ resin, it eluted phenolic acid with20%ethanol and its elution chlorogenic acid content was52.16%; It eluted flavonoid fraction with50%ethanol and adopted concentration and crystallization to deal with50%ethanol fraction and its precipitation was91.64%. It used HPD500macroporous resin to separate chlorogenic acid with30%ethanol and its content was up to77.25%; Further, using Sephadex LH-20gel resin to purify chlorogenic acid with20%ethanol to gain pure chlorogenic acid and its purity was95.01%and it had a better purification effect.4It studies on the external antioxidant activities of Lonicera Japonica leaf polyphenols and tests capacities of Lonicera Japonica leaf polyphenols. flavonoids and pure chlorogenic acid of reducing power and scavenging hydroxyl radicals. DPPH· radicals and ABTS·+radicals. The results of tests show that total polyphenols in leaf of Lonicera Japonica, flavonoids and pure chlorogenic acid have good antioxidant activities. It shows that pure chlorogenic acid has the best effect on reducing power and removing the DPPH· radicals and ABTS·+radicals, flavonoids second and total polyphenols last. The abilities of removing·OH radicals show that flavonoids ability of removing·OH radicals is strongest, pure chlorogenic acid second and total polyphenols last.