Isolation、Identification Of Active Saponins From Schima Superba Resisting Magnaporthe Oryzae

This article studied how to separate the active saponins resisting magnaportheoryzae from Schima superba leaf complex saponins samples. The similarity in bothpolarity and structure of various schima superba saponins posed great difficulties tothe separation. By investigating and analyzing the influence of HPLC columnpacking,flow matching ratio and velocity on the separation of active Schima superbasaponins, compositing the plate theory, rate theory and linear amplification principlereasoning them, we successfully explored the liquid chromatographic behavior of theactive saponins resisting magnaporthe oryzae from Schima superba and efficientpreparation of Schima saponin monomers efficient means of technology. And wemanaged to acquire seven schima superba saponins monomers within a short periodof time. Furthermore, experiments on resistance to magnaporthe oryzae activity wereconducted, locking three highly active monomers. The main study results are asfollow:1. The best liquid chromatographic analysis conditions of schima superbasaponins.We studied the influence of chromatographic packing, flow velocity, flow ratioand sample amount on the effect of schima superba saponins separation. Theconditions of schima superba saponins separation were determined: C18packingwith the aperture of120, specific surface area about300m2/g, the velocity of0.878ml/min, specific gradient elution procedures. we received a relativelysatisfactory chromatographic analysis map and deduced the rate formula of schimasuperba saponins:H=-0.0235195+0.1155/u+0.0149675u.we developed a systematicprocess of mobile phase elution program optimization, through which we arrangedexperiments and acquired the best mobile phase program optimization. We studiedthe relation between the sample quantity and the degree of separation and drew theconclusion that the sample quantity should be less than0.15mg. Thus we obtainedthe best liquid chromatographic analysis conditions of schima superba saponins. Andthen we developed a complete set of liquid chromatographic analysis methods ofschima superba saponins and provided technological ground for future study.2. Successfully obtained monomers by amplifying to half preparation liquid phase.We got the half preparation conditions of velocity of3.78mL/min, sample quantityof0.7mg, specific elution program through the linear equation. By separating theschima superba saponins samples M2IGR5R2、M2IGR5R3, we acquired sevenschima superba saponins monomers. First achieved Schima saponin monomerssemi-preparative liquid separation from the system, has been exceeded the technicalbottleneck of in the current phase of such substances can not be separated on themeans of preparing.3. The determination of resisting magnaporthe oryzae activityWe determined the resistance to magnaporthe oryzae activity between the twoschima superba saponins total grade sample and each monomer. The three out ofseven monomers possess strong resistance to magnaporthe oryzae activity and are generally higher than the schima superba saponins total samples. EC₅₀ ofM2IGR5R2is9.1313μg/mL，EC₉₀ is17.6899μg/mL；EC₅₀ of M2IGR5R3is8.7662μg/mL，EC₉₀ is17.5825μg/mL；EC₅₀ of R2-3is5.9123μg/mL，EC₉₀ is21.6912μg/mL；EC₅₀ of R2-4is5.2064μg/mL，EC₉₀ is16.9781μg/mL. We lockedthree high activity schima superba saponins monomers that are resistant tomagnaporthe oryzae and provided ground for further study.