Study On The Compositions, Antioxidant Capacity And Fingerprint Of Polyphenols From Lycium Ruthenicum Murr

Lycium ruthenicum Murr. is a newly discovered wild resource. It is rich in anthocyanins and other polyphenols. The main distribution areas of Lycium ruthenicum Murr., across different ecological conditions, are the northwest region of China, including Xinjiang Uygur Autonomous region, QingHai province, NingXia Hui Autonomous region and GanSu province. However, there was little information related to the anthocyanins of samples from different biogeocoenosis and various species. The composition, structure and structure-activity relationship of its effectiveness is not clear. In the project, compared with lycum barbarum, different tissues of lycum barbarum, different fruits and vegetables, the phytochemical, in vitro antioxidant activity of Lycium ruthenicum Murr. were evaluated. We also study the compositions, contents and effect of anthocyanins on the viability of oxLDL-induced vascular endothelial cells from Lycium ruthenicum Murr., compared with different fruits and vegetables. Isolation, purification, and identification anthocyanins and other phenolic compounds from Lycium ruthenicum Murr. were by high-performance liquid chromatography (HPLC), preparative chromatograph, mass spectrometer (MS). The HPLC chemical fingerprints of polyphenols from Lycium ruthenicum Murr. were established using the materials from 26 main production sites and from 17 different germplasm resources and analyzed by principal component analysis (PCA), hierarchical cluster analysis (HCA) and Chromatographic Fingerprint of Traditional Chinese Medicine composed by Chinese Pharmacopoeia Committee (Version 2004 A). Partial least squares rregression(PLSR) are adopted to establish speturum-effect quality evaluation system of in vitro antioxidant activity of Lycium ruthenicum Murr., as well as to study correlation between polyphenols composion and the efficacy of polyphenols and reveal the material basis of its effectiveness. The aim of this project is to provide basis for strategies breeding, quality cultivating, assessment the quality and spreading of Lycium ruthenicum Murr..1. Phytochemical and Antioxidant Characteristics of Lycium ruthenicum Murr.The contents of moisture, protein, carbohydrate and lipid were 9.51%,11.26%, 34.52% and 5.10% respectively in Lycium ruthenicum Murr.. Sixteen amino acids were found in Lycium ruthenicum Murr. protein. The predominant fatty acids found in Lycium ruthenicum Murr. were linoleic acid (61.49%), followed by oleic acid (16.69%). Among the microelements, the concentrations of Zn (27.4 mg kg-1) and Fe (405.3 mg kg-1) were considerably higher in Lycium ruthenicum Murr.. The major phytochemicals of anthocyanins (29.70 g kg-1), total polyphenols (41.50 g kg-1), ascorbic acid (1961.2 mg kg-1) and tocopherols (21.3 mg kg-1) were found in Lycium ruthenicum Murr.. The ethanol extracts of Lycium ruthenicum Murr. showed high antioxidant activities using DPPH radical scavenging capacities, ferric reducing power and hydroxyl radical scavenging capacities assays with EC50 values of 61.28 μg/mL,51.21 ug/mL and 48.69 μg/mL. The data clearly indicate that Black Goji could be good reservoir of nutrients, phenolic compounds (main group is anthocyanins) and potent antioxidant activity that might favor them use as natural pigment source and functional food.2. The compositions, contents and effect of anthocyanins on the viability of oxLDL-induced vascular endothelial cells from Lycium ruthenicum Murr., different fruits and vegetablesThe total contents, composisions and effect on the viability of oxLDL-induced vascular endothelial cells of anthocyanins from 6 kinds of different fruits and vegetables, including Lycium ruthenicum Murr.. The results showed that Total anthocyanins of the 6 different fruits and vegetables were 0.21-2.26 mg/g fresh fruits. Comparatively, the anthocyanins compositions of Lycium ruthenicum Murr. are different from the composions of the other 5 kinds of fruits and vegetables, and Lycium ruthenicum Murr. had the highest value of total anthocyanins content and largest effect on oxidative damage of vascular endothelial cells in vitro. It is a rich source of anthocyanins, and one kind of good resource for development of healthy food.3. Isolation, purification and chemical characterization of polyphenols from Lycium ruthenicum Murr.In the present study, through HPLC-ESI-MS analysis,7 acylated anthocyanins and 6 non-anthocyanin polyphenols in Lycium ruthenicum Murr. fruit were identified by HPLC-ESI-MS. Among them, one anthocyanin,4 naringenin derivatives and 2 chrysin derivatives were reported for the first time and petunidin-3-O-rutinoside (cis-p-coumaroyl)-5-O-glucoside was found to be the main anthocyanin in Lycium ruthenicum Murr.. Feasibility of producing high quality of Lycium ruthenicum Murr. several anthocyanins from the extract of Lycium ruthenicum Murr. by macroporous resin column chromatography was investigated. The purity of polyphenols was 89.66%, passed through columns of XAD-7 macroporous resin and Sephadex LH-20 column chromatography respectively, Lycium ruthenicum Murr. polyphenols-enriched fraction was then purified by semi-preparative HPLC, and one anthocyanin of high purity were obtained. Through the analysis of ESI-MS, the unknown compounds were elucidated as petunidin-3-O-rutinoside (cis-p-coumaroyl)-5-O-glucoside.4. The polyphenols fingerprints of Lycium ruthenicum Murr. from different placesThe HPLC chemical fingerprints of polyphenols from Lycium ruthenicum Murr. in Ningxia, Xinjiang, Gansu and Qinghai were established using the materials from 26 main production sites. The results of the analasis for the fingerprints are as follows:1) The compositons and total polyphenols contents in fruits of Lycium ruthenicum Murr. from different places are different. The samples from Wutumeiren Gomud of Qinghai, Ganhezi of Xinjiang and Helan 12-01 of Ningxia had the higher value of Total polyphenols, total flavones and total anthocyanins;2) A total of 12 phenolic compounds were detected across all 26 samples, with Petunidin-3-O-rutinoside (cis-p-coumaroyl)-5-O-glucoside being the most abundant; 3) The polyphenols fingerprints of Lycium ruthenicum Murr. from 26 different places were similar, excluding a few of samples; 4) All the the polyphenols results of Lycium ruthenicum Murr. from different places were clustered into 7 kinds according to different polyphenols indexes. There were litte differences according different polyphenols indexes; 5) the results of PCA showed that the quality differences of the polyphenols contents in Lycium ruthenicum Murr. from different places were determined by the mainly phenolic compounds:naringenin, delphinidin-3-O-rutinoside (cis-p-coumaroyl)-5-O-glucoside, petunidin-3-O-rutinoside (cis-p-coumaroyl)-5-O-glucoside chrysin-p-ferulic acid-hexoside and chrysin-p-ferulic acid, identified by HPLC-ESI-MS tentatively.5. The polyphenols fingerprints of Lycium ruthenicum Murr. from different germplasm resourcesThe HPLC chemical fingerprints of polyphenols from Lycium ruthenicum Murr. was established using the materials from 17 different germplasm resources. The results of the analasis for the fingerprints are as follows:1) The compositons and total polyphenols contents in fruits of Lycium ruthenicum Murr. from different germplasm resources are different. Samples No.9-11 showed relatively high contents of total phenols, total flavonoids and total anthocyanins. In contrast, the phenol contents of samples No.4 and No. 12 were comparatively very low. Samples No.8, No.10 and No.11 showed high contents of total anthocyanins and monomers;2) A total of 8 phenolic compounds were detected across all germplasm materials, with petunidin-3-O-rutinoside (cis-p-coumaroyl)-5-O-glucoside being the most abundant; 3) The polyphenols fingerprints of Lycium ruthenicum Murr. from 17 different germplasm resources were different, the similarities are between 0.325 to 1.000; 4). Different polyphenol parameters gave rise to slightly different clustering results, but the 17 materials always clustered into six classes.5) The results of PC A showed that the quality differences of the polyphenols contents in Lycium ruthenicum Murr. from different germplasm resources were determined by the mainly phenolic compounds: naringenin, petunidin-3-O-rutinoside (cis-p-coumaroyl)-5-O-glucoside, chrysin-p-ferulic acid-hexoside, chrysin-p-ferulic acid, and 2 unkwown compounds, identified by HPLC-ESI-MS tentertively.6. The spectrum-effect relationship of in vitro antioxidant activity of polyphenols from Lycium ruthenicum Murr.Five different assays, including FRAP values, hydroxyl radical scavenging capacities, DPPH radical scavenging capacities, reduce power and ABTS radical scavenging capacities were used to study the antioxidant activity of the polyphenols of Lycium ruthenicum Murr. from 26 different places. The results indicated that in vitro antioxidant activity of polyphenols from Lycium ruthenicum Murr. from different places are differents, and the results of the same sample are different from the different assays. The samples from 12-01 Helan Ningxia, Delingha Qinghai province, Gaoquan Xinjiang and Zhongqu Mingqin Gansu province have better antioxidant activities. Partial leasts squares regression (PLSR) model was applied to analyze the correlations between the antioxidant activity and the polyphenols.The results showed that the contents had the largest effects on the antioxidant activities of Lycium ruthenicum Murr., the contents of petunidin-3-O-rutinoside (cis-p-coumaroyl)-5-O-glucoside, the contents of total anthocyanins and the contents of total flavones, subsequently, the Weighting coefficients of regression were 2.243,1.418, 1.392 and 1.298, subsequently, All of the polyphenols indexes had positive correlations with all of the antioxidant activity indexes, excluding the peak 31 and peak 10, which are Naringenin-3-O-rutinose-7-O-hexoside and petunidin-3-O-rutinoside (cis-p-coumaroyl)-5-O-glucoside, identified by HPLC-MS tentertively.