| In recent years,consumers are more and more interested in using natural fruits or food to improve their health.Fruits and vegetables contain natural antioxidants,which are very important for human health.Apple(Malus × domestic Borkh.)is one of the most popular fruits in the world,which is rich in nutrients and bioactive substances and has been widely used in health-related research around the world.The phytochemicals in apples have beneficial effects on antioxidation,prevention of inflammation,obesity,hyperlipidemia,cancer,diabetes,atherosclerosis and coronary heart disease.To prevent disease and improve health through diet is an effective way of health care.In the past ten years,our team has constructed a series of hybrid populations with M.sieversii f.niedzwetzkyana and two excellent white-fleshed apple varieties(‘Fuji' and ‘Gala'),among which there are some red-fleshed apples with red peel and red flesh.These red-fleshed apple are rich in anthocyanins and other phenolic substances,with high nutritional value,which lays a good foundation for the processing and utilization of apple resources,and has great prospects for development and utilization.Therefore,in this paper,the F2 hybridpopulations of M.sieversii f.niedzwetzkyana was used as raw material,the content of phenols,individual phenolics,antioxidant and antiproliferative activities capacity of red-fleshed apple were systematically studied;‘Meihong' was selected as the research object to conduct in-depth study,to estimate the effects of in vitro digestion on total phenols,flavonoids,flavanols,anthocyanins and phenolic components,intracellular/extracellular antioxidation,antiproliferation and cytotoxicity of red-fleshed apple.The effects of polyphenol extracts from different parts of red-flesed apple on the antiproliferation of breast cancer cells were studied.The effects of processing on the chemical compositions,phenolic compositions and antioxidant activities of red-fleshed apple cider were studied to determine the best fermentation mode of red-fleshed apple cider.The main results are as follows:1.In this study,compared with the traditional white-fleshed apple variety ‘Fuji',the phenolic characteristics,antioxidant and antiproliferative activities of the four hybrids of M.sieversii f.niedzwetzkyana were analyzed.In addition,the antioxidant and anti-proliferation properties of these apples were measured.Compared to ‘Fuji' apples,the red-fleshed apples were rich in phenolics and flavonoids,ranging from 1.5-to 2.6-and 1.4-to 2.4-fold,respectively.Among them,the contents of total phenolics,flavonoids,flavanol and anthocyanin in ‘A38' were the highest,which were 2945.03±42.02 mg GAE/kg FW,2191.02±58.07 mg RE/kg FW,1354.22±35.74 mg CE/kg FW ?282.88±8.11 mg C3GE/kg FW,respectively.A total of 13 major phenolic components in different apple varieties were identified and quantified by using UPLC-MS / MS,including 5 flavanols,4 flavonols,2 anthocyanins,1 hydroxycinnamic acid and 1 dihydrochalcone.The total individual phenolics of UPLC-MS/MS was consistent with that of TPC.In all antioxidant methods [DPPH radical-scavenging capacity,ABTS radical scavenging activity,ferric reducing antioxidant power(FRAP)and cell antioxidant capacity(CAA)],‘A38' obtained the highest antioxidant value(DPPH: 22.00±0.89 TE ?M/g FW,ABTS: 31.88±0.53 TE ?M/g FW FRAP: 12.30±0.24 TE ?M/g FW),while ‘Fuji' got the lowest antioxidant value(DPPH: 7.19±0.69 TE ?M/g FW,ABTS: 10.19±0.49 TE ?M/g FW,FRAP: 3.75±0.19 TE ?M/g FW).The results of CCK8 showed that the IC50 values of different apples for MCF-7 ranged from 33.44 mg/m L to 73.36 mg/m L,‘A38' had the lowest IC50 value,which was 33.44 mg/m L,and ‘Fuji' had the highest IC50 value,which was 73.36 mg/m L.However,the IC50 value for MAD-MB-231 cells ranged from 20.94 mg/m L to 39.39 mg/m L,‘A38' obtained the lowest IC50 value,which was 20.94 mg/m L,and ‘Fuji' had the highest IC50 value,which was 39.39 mg/m L.The lower the IC50 value obtained from apple extract,the stronger the antiproliferation ability to cancer cells.The red-fleshed ‘A38' and ‘Meihong' exhibited higher antioxidant and antiproliferative activities in vitro because of the higher levels of phenolics,and the higher potential for development and utilization value.2.Simulated digestion experiments in vitro were carried out on the peel and pulp of ‘Meihong' to assess the effects of simulated digestion on the content and composition of phenols in different parts of red-fleshed apple,the antiproliferative activities and cytotoxicity,including three stages of oral,gastric and intestinal digestion.The results showed that compared with chemical extraction,the TPC,TFC,TFAC and TAC in peel and pulp after digestion decreased significantly,but the contents of free phenolic acid increased significantly.UPLC-MS/MS and HPLC were used to characterize the main flavonoids and phenolic acids in the extracts of different treatments,including 13 flavonoids and 12 phenolic acids.An analysis of the in vitro antioxidant activities(including ABTS,DPPH,FRAP,and CAA)revealed that the digestion decreased the ABTS,DPPH,and FRAP values,but increased the CAA values,relative to the corresponding values for the chemical extracts.These results suggest that the digestion improved the effectiveness of the phenolic substances.Moreover,our findings implied that the digestion promoted the antiproliferative activities of red-fleshed apple peel and flesh relative to the effects of the chemical extracts.Future in vivo investigations are warranted based on the results of the current study.3.The TPC and TFC in APP was significantly higher than that in AFP.The qualitative and quantitative analysis of 14 major phenolic components in APP and AFP was carried out by UPLC-MS/MS,including 5 flavanols,5 flavonols,2 anthocyanins,1 hydroxycinnamic acid and 1 dihydrochalcone.The results showed that the contents of different phenolic substances in APP and AFP were significantly different.The results of CCK8 showed that: APP and AFP inhibited cell proliferation in a time and dose-dependent manner with an IC50 of 662.97 ?g/m L and 860.24 mg/m L for 48 h expose time,respectively,suggesting that APP was more effective than AFP in inhibiting MDA-MB-231 cells proliferation.Besides,the significantly inhibition effect was achieved through Akt pathway by inducing apoptosis(significantly up-regulating reactive oxygen species(ROS),Bax levels,and down-regulating expression of p Akt,p Bad,Bcl-2,promoting Cytochrome c release,activating Cle-Caspase 9,and inducing expressions of Cle-Caspase 3 and Cle-PARP),and inducing G0/G1 cell cycle arrest(increased expressions of p-p53 and p21 and decreased expressions of PCNA and Cyclin D1).These results suggest that the peel and pulp are the active parts of red-fleshed apple,which may be an excellent raw materials of natural chemicals for the treatment of MDA-MB-231 cells.4.The processing quality of red-fleshed apple cider was studied by different fermentation modes.The fermentation process of apple cider was monitored by the determination of cell activity and organic acid metabolism.SPME/GC-MS,UPLC-MS/MS and HPLC were used to analyze the main volatile components,flavonoids and phenolic acids in apple cider,including volatile components,flavonoids and phenolic acids.And sensory analysis is used to evaluate the quality of ciders.Our findings suggest that Cider obtained after co-inoculation with oeni PG-16(SIM/PG-16)had the highest antioxidant activity and a sensory evaluation profile dominated by fruits and floral notes linked to esters,and to butter and creamy notes associated with ethyl lactate and 2,3-butanediol that enhanced the global aromatic intensity.Thus,SIM/PG-16 is an effective method for preparing excellence red-fleshed apple cider.Then the main chemical compositions and nutritional quality of ‘Fuji' apple cider were evaluated by SIM/PG-16.All results showed that:compared with ‘Fuji',the red-fleshed apple cider obtained by ‘Meihong' had higher nutritional value,full body,strong structure,strong fruit fragrance and flower fragrance,high quality,which fully showed the brewing characteristics of ‘Meihong'. |
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