| Two-dimensional liquid chromatography has been widely used in the analysis and separation of natural products due to its good separation capability.NPLC x RPLC has the best orthogonality but always has solvent incompatibility issues so that brought many difficulties for system development.However,when we studied in different organic solvents,an unexpected thing was discovered that n-hexane can be used as a special normal phase chromatography solvent in NPLC × RPLC two-dimensional liquid chromatography,and does not disadvantageously influent the separation of reversed-phase chromatography.Dissolving the sample with it can even obtain better separation effect than methanol and other solvents,so the traditional theory about the solvent effect of organic solvents remains to be confirmed.In order to prove the correctness of the discovery,we used different polarity model compounds to investigate the influence of different solvent systems on the separation performance of reversed-phase chromatography.We discovered the specificity of n-hexane and investigated on this basis a series of parameters including temperature,solvent injection volume on this basis.Finally,we confirmed that reversed-phase chromatography can tolerate an amount of n-hexane to obtain better separation effect than methanol when dissolved samples.A new two-dimensional off-line NPLC x RPLC preparation system was established.And this system was applied to the separation of the components in the clone 4-1.A total of 20 compounds were isolated from the extract,which simplified the two-dimensional systerm.This paper first proposes that n-hexane as a special normal phase chromatography solvent does not influent the reversed-phase chromatography system in a certain range,and provides a theoretical basis for the development of subsequent reversed-phase reversed-phase chromatography systems.A new NPLC x RPLC preparative chromatography developed for the separation analysis of natural products. |
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