Application Research For Separation And Determination Of Uncaria And Gelsemium Alkaloids By HPCE

Objective:In the first chapter of this work,hydroxypropyl-?-cyclodextrin(HP-?-CD)and tetrabutylammonium-L-glutamic acid(TBA-L-Glu)were used as additives in the separation system to research the influence of additives on the separation and determination of corynoxine(C)and corynoxine B(CB).A sensitive,fast and effective method that field-amplified sample stacking(FASS)in capillary electrophoresis(CE)has been established and applied to determination of C and CB in the stems with hooks of uncaria rhynchophylla and its formulations.The second chapter of this paper studied the effects of cyclodextrin-modified micellar electrokinetic capillary chromatography(CD-MEKC)on the separation of four gelsemium alkaloids(gelsemine,humantenine,koumine,gelsenicine).A comprehensive,rapid and practical CD-MEKC method was developed and applied to determination of four gelsemium alkaloids in Gelsemium elegans Benth and its formulations.Method:The experiments were performed by using a P/ACE MDQ system with P/ACE DAD.Uncoated fused-silica capillary with 40.2 cm×75?m id(30 cm to the detector)was used.(1)The effects of separation and stacking efficiency were investigated in detail.The main parameters include concentration and pH of buffer,concentration of additives(HP-?-CD and TBA-L-Glu),injection time of water and sample and separation voltage.Electrokinetic injection was chosen to introduce sample from inlet at 10 kV for 50 s after a water plug(0.5 psi,4 s)was injected to permit FASS.The running buffer(pH 6.1)was composed of 40 mM sodium dihydrogen phosphate solution,130 mM HP-?-CD and 10mM TBA-L-Glu and the separation voltage was 20 kV.(2)The parameters,including buffer concentration and pH,sodium cholate(SC)and dimethyl-?-cyclodextrin(DM-?-CD)concentration,separation voltage were optimized.The optimal BGE composition was 40 mM sodium dihydrogen phosphate solution(pH 6.3)containing 12.5mM SC and 120 mM DM-?-CD;running voltage was 15 kV,temperature was 25°C,injection samples was 0.5 psi for 4 s.Results:Under the optimum conditions,calibration curves were in a good linear relationship within the range of 62.5~5.00×10~3 ng/mL both for C and CB,analytes were successfully enriched and separated within 12 min,the resolution was 2.5 and the sensitivity was improved approximately by 700~900 folds;the intraday RSDs<2.7%and interday RSD<3.7%;the recoveries of samples were ranged from 98.1%to 101.3%?Under this condition,a good linearity was obtained with correlation coefficients from 0.9971 to 0.9995;four gelsemium alkaloids were baseline separated within 11 min,the resolutions were from 1.8 to 3.4;the intraday RSDs<2.9%and interday RSD<4.1%;the recoveries of the samples varied from 96.5 to 101.6%.Conclusion:(1)The experimental results of this study demonstrated that the separation and enrichment of C and CB by FASS-CE was effective.Therefore,this study has developed a simple method,which is promising for the QC of the C and CB in uncaria rhynchophylla.(2)The experimental results of this study demonstrated that the separation and determination of four gelsemine alkaloids by CD-MEKC was possible.Finally,this method was successfully applied to determination of four gelsemium alkaloids in Gelsemium elegans Benth and its formulations.