Identification And Evaluation Of Paris Polyphylla Var. Yunnanensis Using Spectroscopic And Chromatographic Techniques

Paris polyphylla var.yunnanensis(PPY),as the material of Chinese patent medicine,“Ji Desheng Antivenom Drug”,“Gongxuening Capsule”,“Yunnan Baiyao”,which has been used to clear heat and remove toxin,disperse swelling and relieve pain,cool the liver to settle convulsion.With the development of demand of industrialization,recently,the number of PPY has decreased and even at the edge of extinction.It is necessary to establish a method to change the situation.The secondary metabolite of medicinal plant varied significantly due to growing condition,individual,harvesting time and processing condition.It is therefore to develop a robust,reliable and fast strategy to discriminate geographical origin,growing years in PPY and its relative species,which can provide a screening method for the best regions,alternative species and harvesting time for PPY.Up to now,high performance liquid chromatography determine polyphyllin I,II,VI and VII for PPY with long time and a lot of organic solvent.Infrared and ultraviolet spectroscopy has been used for evaluation of PPY comprehensively,while they fail to quantify the index components.The aim of this study is to evaluate PPY and its relative qualitatively and quantitatively in different regions,growing years and species using ultra high performance liquid chromatography(UHPLC),ultraviolet(UV)and Fourier infrared(FTIR)spectroscopy combined with chemometrics,which can provide a foundation for screening good germplasm resource of PPY.The investigation could divide into five parts.Part 1,UHPLC coupled to chemometrics discriminate six species of Paris to illuminate the relationship between PPY and its relative in the chemical components.Part 2,UHPLC and UV spectroscopy evaluate PPY collected from three regions qualitatively and quantitatively to discuss the chemical difference of PPY in different geographical origin.Part 3,distinguishing different growing years of PPY is to discriminate the chemical constituents of different harvesting times.Part 4,it is to establish a method for quantitation of total saponins of different species for Paris using FTIR combined with chemometrics.Part 5,FTIR coupled to chemometrics is to determine total flavonoids of PPY in different regions.Part 6,a review,on the research of chemical components and pharmacological activity of Paris.The investigation is:(1)to develop an qualitative and quantitative method for different species of Paris using UHPLC fingerprint,the results indicate that the content of different species varied obviously,and P.axialis(PA)and PPY is higher than others.PPY has the apparent content ratio(equal to polyphyllin II against polyphyllin I)with 46.61%-122.44%,while the moderate was for PA(8.61%-43.08%).UHPLC combined with principal component analysis and partial least square-discriminant analysis(PLSDA)can discriminate different species of Paris,and similarity analysis can evaluate PPY and PA.In addition,PA had better performance than PPY in the content,content ratio,clustering effect.It is therefore necessary that PA can be an alternative source of PPY for the further investigation and development.(2)to evaluate PPY of different geographical origin in Yunnan Province qualitatively and quantitatively,the results indicate that the average content of western and southern region is higher than that of central district.For the most samples,there is an obvious trend that the content of polyphyllin I is highest and that of polyphyllin VI was lowest,and polyphyllin II is higher than polyphyllin VII.The analysis found that PLS-DA for UV spectroscopy could separate the samples from three regions,was more appropriate than UHPLC.Retention times during 20-30.75 min of UHPLC,and absorption at 200-300 nm of the UV spectrum were identified as significant peaks for distinguishing PPY from different regions.(3)to discriminate PPY of different growing years using UHPLC and FTIR spectroscopy,the results indicate that the content of total saponins(including polyphyllin I,II,VI and VII)in eight years is higher than others.The highest percentage of polyphyllin I is found at 12 years,and polyphyllin II,polyphyllin VI and polyphyllin VII are distributed at 5,8 and 4 years.The PLS-DA of FTIR spectra has a better performance than that of UHPLC for discrimination of PPY from different harvesting times.(4)to determinate total saponins of different species of Paris using FTIR and chemometrics,the results indicate that a combination pretreatment of standard normal variate(SNV),second derivative(SD)and orthogonal signal correction(OSC)has the best performance with the highest residual predictive deviation(4.8644 in the PLSR and 2.9261 in the SVMR).SD always has the better performance than first derivative(FD).Compared with full wavenumber,selected wavenumber has the better performance in the SVMR for the most of samples,while in the PLSR it has the various results.(5)to determinate total flavonoids of different regions for PPY using UV and FTIR spectroscopy,the results indicate the content is highest in Yuxi and lowest in Xishuangbanna.Geographical origins have not so much correlation with content of total flavonoids.A combination of pretreatment include SNV,OSC,FD and SavitzkyGolay(7)has the best performance in the determination of total flavonoids.