| Two-dimensional chromatography is one of the most effective method toseparate and analysis the complicated compositions, compared with traditionalseparating technology, it has unparalleled advantages, it can separate signalcomponent from the complicated compositions rapidly and efficiently. However, forthe low amount of purity, the Two-dimensional chromatography can just be adopted toanalyze and identify trace amount material. There are still many shortcomings in theresearch and application of preparation. On the basis of Two-dimensionalchromatography and traditional chromatography, our research group designedmulti-channel two-dimensional chromatography separating equipment to make up theapplication on preparation. The article, based on the multi-channel two-dimensionalchromatography and Pine polyphenols systematically optimize process conditions ofseparation Pine polyphenols by multi-channel two-dimensional chromatography andstudy the active tracking of antioxidants of Pine polyphenols by multi-channeltwo-dimensional chromatography and antioxidant function. The main contents can bestated as follows.According to the principle of multi-channel two-dimensional chromatography,we successfully structure multi-channel two-dimensional chromatography separatingequipment; then adopt two different specifications of chromatographic column to testthe separating effect of multi-channel two-dimensional chromatography withblueberry residue. The results suggest that, compared with the extracting solution ofblueberry residue, the purity of blueberry anthocynin separated by multi-channeltwo-dimensional chromatography have greatly improved; the biggest has increased8.9times. Thus, the multi-channel two-dimensional chromatography is provided withthe separation ability to the bioactive substances of natural plants.With pinecone of Pinus koraiensis Sieb.et Zucc., the best processing parameters,including solid phases, eluent, sample amount, sample concentration, elution rate andcolumn amount, are optimized. The static adsorption-desorption performance is usedto determine that, the X-5macro porous resin with good effect of adsorption anddesorption can be used as solid phases,0.5%formic acid-60%ethanol solution withthe best desorption efficiency can be used as eluent. Next, the dynamic single factortest and response surface methodology test are adopted to get the optimum value ofthe other four factors. The results are respectively260mg,2.0mg/ml,200±20ml/min,And the amount of the hyphenated chromatographic column is4. In theseconditions, the separating effect of polyphenol by multi-channel two-dimensionalchromatography is great; the purity of Pine Polyphenols was developed from initial2.37%to23.357%.MTT test is used to determine the influence of the component which has thestrongest total reducing ability (Ct) and the component which has the most powerful hydroxyl radical scavenging ability (Ch) on mice splenocyte proliferation reaction.Results indicate that the two components have strong ability to stimulate micesplenocyte to proliferate. When the concentration of Ctis5.0ug/ml, the proliferationrate of splenocyte is the highest and can reach15%. The proliferation rate ofsplenocyte can run into10%, when the concentration of Chis20ug/ml.The oxidative-damage model in mice splenocyte is established by H2O2inducedmeasurement. In the oxidation resistant test, the two components both have protectionto cell oxidation-damage: Cthas the best efficiency of protection by high dose(160ug/ml), and Chcan reach the optimal effect of protection by middle dose(40ug/ml). And in the repairing test of oxidation–damage, the two components bothhave restorative effects to cell oxidation-damage by middle dose, but the high dosegroup and the low dose group all have no effect.The two antioxidant componengts are found that they all contain polyphenol,flavonoid and anthocyanin in the component analysis, but the amount of anthocyaninis low. And the polyphenol and flavonoid in Chis less than those in Ct. The proportionof polyphenol, flavonoid and anthocyanin in the two components are250:75:7and785:350:1, respectively.Above all, the structure of the multi-channel two-dimensional chromatography,the process optimization of separating Pine polyphenol and the active tracking ofantioxidants were undertaken to research the multi-channel two-dimensionalchromatography and Pine polyphenols in this article, and we have achieved betterresults. These achievements can be excellent technical and theoretical guide on theseparation of active materials and the application of Pine polyphenols, and they alsolay the foundation for the further study of two-dimensional chromatography andPine polyphenols. |
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