| The peels of Yam were often discarded as trash during the production and processing of Yam,which not only bring pollution to the environment but also greatly wasted resources.We had carried out experimental studies on XPE in order to properly utilize the resources of Dioscorea opposita Thunb.In this paper,with four antioxidant activity assays as activity orientation,the main components of XPE were systematically studied by using various column chromatographic separation methods,and the chemical structure was identified by NMR and other spectral analytical methods.Then the main chromatographic peaks of XPE were identified by HPLC.Meanwhile,Na NO2-Al(NO3)3-Na OH colorimetric method was used to probe that weather it co uld apply to the determination of the content of total flavonoids in Chinese yam.Over besides,we achieved the enrichment of the main phenols in XPE by alkali extraction and acid precipitation method according to their chemical properties.In addition,the identified compounds were assessed for α-glucosidase inhibitory activity,CCK-8 cell proliferation activity and tyrosinase inhibitory activity.The results are as following:The DPPH、ABTS+ 、hydroxyl radical scavenging ability and reducing power of XPE and XRE were analyzed experimentally,and the results showed that the antioxidant activity of XPE was significantly stronger than that of XRE.The phenol content of XPE and XRE were determined by F-C method,and the results showed that it was about 10 times higher in XPE(3.55 mg/g)than that of XRE(0.36 mg/g),which is consistent with the result of the antioxidant activity.The result indicated that XPE was rich in phenolics.To further clarify that the major compounds in XPE(29 g),XPE was isol ated and identified by multiple column chromatography and spectroscopy,and we obtained 17 compounds at last.Including eight stilbenoids: batatasin I(1),3,5-dimethoxy-2,7-phenanthrenediol(2),isobatatasin I(3),dihydropinosylvin(4),batatasin III(5),batatasin IV(6),demethyl batatasin Ⅳ(7),3,3’,5-trihydroxybi benzyl(8).Five diphenylheptanes:(3R,5R)-3,5-dihydroxy-1-(3,4-dihydroxyphenyl)-7-(4-hydroxyphenyl)heptane(9),(3R,5S)-1,7-bis(4-hydroxyphenyl)heptane-3,5-diol(10),1-(4-hydroxy-3-methoxyphenyl)-7-(4-hydroxyphenyl)-3,5-heptanediol(11),(3S,5S)-3,5-dihydroxy-1,7-bis(4-hydroxy-3-methoxyphenyl)-heptane(12),(3R,5R)-3,5-dihydroxy-1,7-bis(4-hydroxy-3-methoxyphenyl)-hepta-3-O-β-D-glucopyranoside(13).Three N-contained compounds: trans-N-p-coumaroyltyramine(14),N-ferulo yltyramine(15),N-trans-cinnamoyltyramine(16).One fatty acid:(8R*,9R*,10S*,6Z)-trihydroxyoctadec-6-enoic acid(17).All compounds are the known compou nd,while compound 4,7,11,13,16 and 17 were first discovered from Diosc orea opposita Thunb.Meanwhile,the HPLC spectrum of the above compounds were analyzed with XPE under the same chromatographic condition,and the main peaks of XPE were identified,and comparison of the graphs showed that the main peaks in the XPE liquid phase spectrum were compounds 1,2 and 16.Combining literature research with the results of our previous experiments,we found that there were no flavonoids but stilbenes and diphenylheptanes in yam,which was not in conformity with our previous experimental results.In this paper,the color reaction and the full wavelength scan determination of compound 1,2,3,11,12,and 16 performed using Na NO2-Al(NO3)3-Na OH.The result showed that all of them,except the compound 16,were able to react with Na NO2-Al(NO3)3-Na OH reagent,and there was an obvious absorption peak at 510 nm.The experimental results showed that the presence of phenolics would affect the determination of their flavonoid content,and Na NO2-Al(NO3)3-Na OH reagent did not apply to the determination of total flavonoids in yam.We used the chemical characteristic of phenolic to achieve the enrichment by using alkaline extraction and the acid precipitation method.The results showed that the extraction of phenolic using different alkaline solutions did no different significantly,however,the extraction rate increased significantly with the increasing alkalinity of the alkaline solution,and 5% Na OH had the highest extraction rate with 0.44 % phenols,the HPLC analysis spectrum showed that the main peaks of the extracts were compounds 1,2 and 3.The α-glucosidase inhibitory activities of compounds 1、2、3、7、8、10、14 and 16 were investigated,and it was found that compound 8 and 14 were characterized by α-glucosidase promoting effect compared to the positive control,while the rest of the compounds had inhibitory activities,indicating that different small molecules have regulatory effects on blood glucose.Using the CCK-8 method,the cytotoxic activity of compounds 2,3,7,9,12,14,and 16 was assayed for RAW264.7 cell,and it was found that compound 2 showed theactivity of promoting cell proliferation,while compounds 3,9,and 16(> 100 μg/m L),compounds 7,12,and 14(> 200 μg/m L),and compound 2(> 25 μg/m L)exhibited cytotoxic activity certainly when compared to the blank group.Compounds 2、3、7、9、12 and 14 did not showed any significant tyrosinase inhibitory activity. |
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