Isolation, Structure Elucidation And Structure Activity Relationship Of Antioxidant Compounds From Zanthoxylum Bungeanum Leaves

The leaves of Zanthoxylum bungeanum are commonly consumed as condiments andvegetables in China for their special flavor properties. In spite of its long history ofconsumption, this material has neither been fully investigated nor developed and there stilllack information for consumers to understand its phytochemical composition and nutritivevalues.1. The total phenolic, flavonoid content and antioxidant activities of ethanol crudeextracts (ECE) and five sub fractions (petroleum ether fractions (PEF), chloroform fraction(CF), ethyl acetate ftractions (EAF), acetone fraction (AF) and methanol fraction (MF)) of Z.bungeanum leaves were evaluated. The results indicated that EAF had the highest totalflavonoid and phenolic content, which were in good correlation with its significantantioxidant activities. AF is not significantly different with EAF in phenolic content andantioxidant activity. Thus, EAF and AF were screened as the active parts for further isolationand purification.2. Nine compounds were isolated and identified from the EAF and AF using silica gelcolumn chromatography, gel filtration chromatography, preparative high performance liquidchromatography for isolation. All the compounds were identified on the basis of spectralanalysis (UV, IR, El-MS,1H-NMR,13C-NMR,2D-NMR) and their phytochemical properties.Nine compounds were quercetin (1), afzelin (2), quercitrin (3), trifolin (4), isoquercitrin (5),isorhamnetin3-O-α-L-rhamnoside (6), hyperoside (7), vitexin (8) and rutin (9). Allcompounds were isolated from the leaves of Z. bungeanum for the first time. Five compounds(2,4,5,6and8) were found for the first time in the genus Zanthoxylum. Furthermore, the2D-NMR data of the five known compounds (2,3,6–8) are given for the first time in thispaper.3. The antioxidant activities of compounds were evaluated by in vitro and in vivoantioxidant assays. Quercetin and quercetin glycosides showed the highest antioxidant activity.Structure-activity relationships indicated that, the-OH in4’ position on the B ring and the-OH in7position on the A ring possessed high antioxidant activity; B ring and/or A ring withadjacent-OH groups could greatly increase their antioxidant ability.4. The content of eight phenolic compounds (chlorogenic acid, epicatechin, rutin, hyperoside, trifolin, quercitrin, afzelin and quercetin) content of various fractions of Z.bungeanum leaves were evaluated by HPLC method. Morever, the possible mechanismresponsible for the significant antioxidant activities of the obtained fractions (E-4, E-5, E-7E-2-4and E-3-3) were investigated by HPLC analysis. It is demonstrated that chlorogenicacid, epicatechin, rutin, hyperoside, trifolin, quercitrin, afzelin and quercetin were the majorantioxidants in Z. bungeanum leaves. The HPLC profiles also demonstrated that hyperosideand quercitrin exist as the major components in Z. bungeanum leaves.5. The results firstly revealed the chemical composition and biological activity of Z.bungeanum leaves. The content of those isolated flavonoids could be used as a marker for thequality assessment of Z. bungeanum leaves and its bioactivity products. Our present workprovides a theoretical foundment and technical support for better development and utilizationof Z. bungeanum leaves.