Research On Polyphenols From Apple: Preparation, Identification And Antioxidant Activity

China is the country with largest apple planting area and output in the world.Fuji is themain apple varieties in China, which accounting for65%of apple’s total production. In applemain origin-shandong, Fuji respectively accounts for70.2%and76.2%of total area and yield.Apple polyphenols—secondary metabolites of apple, have important physiological functionsand development prospect. In order to realize the integrated application of apple polyphenols,this article took shandong Fuji apple as the research object, with extraction by ultrasonicmethod and isolation and purification by macroporous resin and Sephadex LH-20. Thenmethod of high performance liquid chromatography (HPLC), ultra performance liquidchromatography-quadrupole time of flight-mass spectrometry (UPLC-QTOF-MS), nuclearmagnetic resonance (NMR), Fourier transform infrared spectrometry (FT-IR), ionchromatography(IC)were applied to analysis the structures of the apple polyphenols. Finallyantioxidant activity of apple polyphenols was evaluated. The main results were as follows:First, extraction of apple polyphenols was studied.Apple polyphenols were extracted byultrasonic method. The optimum extraction conditions were got by single factor andorthogonal experiments: extractant ethanol, pH3.5, concentration65%, ultrasonic extractiontime25min, solid-liquid ratio1:17. Under the optimum conditions, extraction ratio was11.5mg/g(dry weight). According to the optimum conditions, the expanded experiment wasconducted with100kg apples, getting crude extract solid powder1069g, which thepolyphenols content was10.1%.Second, isolation and purification of apple polyphenols crude extraction werestudied.XAD7HP macroporous resin showed the best adsorption and desorption capacityamong six kinds of macroporous resins. Its static adsorption capacity was14.3mg/g, staticdesorption ratio was98%. The optimal purification conditions were sample concentration of1.46mg/mL, sample pH of5.50, the flow rate of sample of1.25mL/min, eluant of90%acetone, the flow rate of eluent of1mL/min. In this condition, purity could be up to80.1%,which was more than8times than before, and the recovery ratio was as high as90%.According to the optimum conditions, the expanded experiment was conducted with300gapple polyphenol crude extract. Through purification and separation by XAD7HPmacroporous resin,ultimately getting FA(1.38g),FB(7.50g) and FC(9.75g). FAwas mainlyphenolic acids, FCwas mainly flavonoids, FBwas a mixture of phenolic acids and flavonoids.On Sephadex LH-20column1g of FAwas taken isolation process, then obtained compoundFA1(480.2mg). On Sephadex LH-20column5g of FCwas taken isolation process, thenobtained compounds FC1(1079.0mg), FC2(182mg), FC3(273.6mg), FC4(123.6mg).Third, identification of the structures of apple polyphenols was studied. Integrated HPLC,UPLC-MS, FT-IR, NMR and method of hydrolysis analysis analysis, Fuji apple containedchlorogenic acid(3-Caffeoylquinic acid), epicatechin, phloretin, quercetin-rhamnoside,phloretin-glucoside. Fuji apple maybe contained procyanidin B2, quercetin-glucoside,quercetin-xyloside, Phloretin-xylosyl-glucoside,etc.Last, antioxidant ability of apple polyphenols was studied. DPPH scavenging activity of apple polyphenols were established by HPLC. Half maximal inhibitory concentration (IC50)of DPPH was calculated. So the DPPH scavenging activity of six kinds of polyphenols wasthat epicatechin (37.44μg/mL)>chlorogenic acid (40.09μg/mL)> phloretin-glucoside (41.43μg/mL)> mixed polyphenols (45.32μg/mL)>quercetin-rhamnoside (50.76μg/mL)>phloretin(56.57μg/mL). The effect of apple polyphenols on oxidation stability of soybean oil wasstudied by rancimat method, finding that six kinds of apple polyphenols could significantlyimprove the oxidation stability of soybean oil,that chlorogenic acid> phloretin-glucoside>quercetin-rhamnoside> mixed polyphenols>epicatechin>phloretin. Antioxidant activity ofsynthetic antioxidant TBHQ was slightly better than the polyphenols. Antioxidant activity ofVc is lower than chlorogenic acid and phlorizin, slightly higher than quercetin-rhamnoside,etc.We concluded that antioxidant effect of five kinds of homemade apple polyphenols wasgood, there was no advantage of mixed polyphenols.