Research In Purification And Detection Of L-DOPA Derived From The Seeds Of Stizolobium Cochinchinensis (Lour.)

L-Dopa is clinically main drug for the treatment of Parkinson’s disease which is common.The Stizolobium cochinchinensis（Lour.）is characteristic plant in Guangxi as the resources of food and medicine,whose seeds contain 6%to 9%of the natural L-Dopa,so the process of extracting L-Dopa from Stizolobium cochinchinensis（Lour.）seeds appears largely in the local industry.Further studying its process and analysis methods for the purification will improve the economic output of Stizolobium cochinchinensis（Lour.）seeds in Guangxi.This article simulated the process of extracting L-Dopa from Stizolobium cochinchinensis（Lour.）seeds in the laboratory for preparation,on the basis of optimizing the conditions of refining purification process,to greatly improve the yield and purity of L-Dopa.What’s more,according to the quality standard requirements of United State Pharmacopoeia（USP）,combining with the literature,this study determined the main impurity elements and detection method.In the simulated process of extracting L-Dopa from the Stizolobium cochinchinensis（Lour.）seeds in laboratory,yield of crude products of L-Dopa was（35±5）%,and the content was between 40%to 60%.And the stability of L-Dopa in solution state was good;In the condition of crystallization,L-Dopa at 100℃was not stable;the loss was bigger.Using high performance liquid chromatography（HPLC）method,the study determined the quantitative detection conditions of L-Dopa:chromatographic column was Shimadzu C-18 methyl silane column（250 mm×4.6 mm,5μm）;Mobile phase:methanol:0.1 mol/L acetic acid solution（10:90,v/v）;Uv detector sensitivity:0.05AUFS,detection wavelength of 280 nm;Flow rate:1.0 m L/min;Column temperature:room temperature.At the same time,L-Dopa and Fe Cl₃under acid condition generating complex under ultraviolet had absorption,determining the maximum absorption wavelength of 281 nm of complex.And L-DOPA with Fe Cl₃complex absorbance value and the concentration of L-DOPA making standard curve,it concluded that:the concentration of L-Dopa in 50～1600μg/m L range inside sex good;regression equation:Y=0.0005+0.2242 x,（R²=0.9927）.On the basis of single-factor experiments,the optimal purification condition for maximizing the yield of rough L-Dopa were determined by using a 3-variable,3-level Box-Behnken experimental design combined with response surface methodology analysia as follows:concentration of hydrochloric acid was 0.56 mo L/L;（V/M）was10;（M/M）was 10,resulting in the rough L-Dopa yield of 70.03%.After experimental verification,the actual yield of rough L-Dopa was 67.89%,and relative error was2.83%.The results showed that using the response surface methodology for purification conditions was considerably accurate,and had a certain practical value.Through early detection for refined sample L-Dopa by amino acid analyzer,and the analysis by high performance liquid chromatography（HPLC）method,the refined sample L-Dopa mainly existed L-Tyr and L-Phe,that were the impurity components.And under the ultraviolet detector,HPLC condition was:the chromatographic column:Dikma Spursil C₁₈column（250×4.6 mm,5μm）;Mobile phase:2%acetic acid solution-methanol（90:10）;Flow rate:0.8 m L/min;Detection wavelength of 260nm;Column temperature:room temperature;That were capable of detecting and separating L-Dopa and L-Phe;Under the fluorescence detector,HPLC condition was:the chromatographic column:Dikma Spursil C₁₈column（250×4.6 mm,5μm）;Mobile phase:0.8%formic acid-acetonitrile（98:2）;Sample quantity:10μL;Detection wavelength:Sig=280nm,Ref=314nm;Column temperature:room temperature;That were capable of detecting and separating L-Dopa and L-Tyr.