| Objective:The aim of this study was to investigate the extraction and purification, quality standard and antianxiety activity of iridoids effective fraction of roots and rhizomes of Valeriana jatamansi Jones (IEFV).Methods:The extraction process of IEFV was worked according to preliminary experiments. The purification of IEFV was studied through macroporous resin, the dynamic absorption-desorption was used to find the optimum resin, and the purification process was optimized with purity guided through single factor experiment, such as sample capacity, elution concentration, elution volume, elution flow rate. IEFV was obtained by pilot test. IEFV content was analyzed by TLC, UV and HPLC methods. Further, study on quality standard of IEFV.Rats and mice were used to evaluate the antianxiety activity by the elevated plus maze (EPM), light-dark box (LDB), Vogel’s drinking conflict (VDC) and open field drinking (OFD) tests. After OFD test, the brain and blood of rats was collected immediately for the assays of GABA and5-hydroxytryptamine (5-HT) by ELISA.Results:The optimum extraction process was:crude powder of Valeriana jatamansi Jones,70%ethanol soaking for2times,8vol (w/v) for soaking24h, then6vol (w/v) for soaking12h, vacuum distillation. The D101macroporous resin was found with the best separating efficiency which dynamic absorption capacity and desorption ratio could reach72mg/g and83.3%, respectively. The optimum purification process was as follows:uniformly dispersed sample, sample capacity-72mg/g, adsorption flow rate-1BV/h, elution flow rate-2BV/h, ordinal gradient elution with water of6BV,60%ethanol of4BV and95%ethanol of4BV, collecting95%ethanol eluted fraction and vacuum distillation, vacuum drying at40℃. The spots of TLC were round and well separated. The purity of IEFV through pilot production was71.5%, chlorovaltrate content was8.91mg/g and Valjatrate B was7.00mg/g. Drafting IEFV quality standard according to the style of writing of the "Chinese Pharmacopoeia" of 2010edition (vol. I).In the four tests, the animals of model groups (Vehicle:0.5%CMC-Na) did produce anxiety disaster, the anxious behaviors of animals were obviously improved (P>0.05, P<0.05or P<0.01) in control groups (Estazolam:1.5mg/kg) and IEFV groups (IEFV:6,9and12mg/kg). After OFD test, GABA and5-HT content in rats of model groups were significantly reduced (all P<0.01) compared with normal group (Vehicle:0.5%CMC-Na), while that of control groups and IEFV groups were significantly increased (all P<0.01) compared with model groups, respectively.Conclusion:In the optimum extraction-purification process, the yield of IEFV was2.1%(raw herb weight as a benchmark) and the purity was up to71.5%(chlorovaltrate as a standard). It was feasible and satisfactory that the process could be used for preparation of IEFV. The quality standard was preliminarily established. IEFV content was not less than68%compared with chlorovaltrate as the control. Chlorovaltrate and Valjatrate B contents in IEFV were not less than8.0mg/g and6.5mg/g, respectively. It could be provided a reference about pharmaceutical research for the development and utilization.The antianxiety activity of IEFV can be confirmed by the four antianxiety tests of EPM, LDB, VDC and OF, but it was not show a dose-effect relationship, and the best results was middle dose group. The mechanism maybe is associated with increasing the GABA and5-HT content. |
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