The Study On Chemical Compositions Of Houttuynia Resources In Sichuan And Chongqing

HERBA HOUTTYNIAE,is a kind of traditional Chinese medicine and also a kind of delicious vegetable.According to Chinese Pharmacopoeia(2005),the fresh aerial and underground parts and the dry aerial parts during flowering period could be used as Chinese medicine.In the meantime,the fresh young stems and leaves are also delicious vegetable in its main spreading regions.HERBA HOUTTYNIAE came from Houttuynia cordata Thunb., which belongs to genus Houttuynia of Saururaceae.H.cordata is the only species in Houttuynia Thunb.according to Florae Reipublicae Popularis Sinicae and Flora Sichuanica. Zhu et al.(2001)classified a new species H,emeiensis Z.Y.Zhu et S.L.Zhang in the genus Houttuynia collected at Emei mountain in Sichuan province.It was called commonly as white Yuxingcao,which was also used as vegetable and HERBA HOUTTYNIAE in Emei and Leshan regions.Houttuynia is widely spread in Sichuan and Chongqing regions of China.There existed plentiful genetic variations among genus Houttuynia germplasm resources.The chemical components of genus Houttuynia is the main reason why people utilized it as a kind of medicinal plants or vegetables.The diversities among Houttuynia germplasm resources would influence on these chemical components.Evaluation correctly and effectively on the variations among the chemical components of Houttuynia resources will help to the exploitation and ulilization of Houttuynia resources.In this paper,the compositions and their contents of the essential oils, flavones,vitamins and the rudimental contents of some heavy metals and pesticides of Houttuynia accessions in Sichuan and Chongqing regions were analyzed.The main results were as follows:1.The essential oil(EO)compositions of 18 fresh Houttuynia accessions from Sichuan and Chongqing in China were analyzed by using GC/MS method.In total,21 components could be identified from the essential oils(EO)of the aerial parts of Houttuynia.High level of chemical polymorphism in the EOs of the aerial parts of Houttuynia was observed.On the basis of GC/MS data of the EOs of the aerial parts,two chemotypes could be identified for the frist time within the investigated Houttuynia accessions as follows:chemotypes decanal(D) and myrcene(M).The results of cluster analysis(CA)and principal component analysis(PCA) showed that the chemotypes of Houttuynia were correlated with the chromosome numbers. All the accessions classified into chemotype D have the chromosome number less than 80, while chemotype M had the chromosome number equal to or more than 80.The chemotype D showed lower amounts of monoterpenes and sequiterpenes,on average of 5.72%and 0.16%, respectively.The other non-terpenic straight chain components made up 76.94%to 94.71%of the whole EO components.In general,the chemotype M showed higher amounts of monoterpenes(average 36.16%)and sequiterpenes(1.38%),while the concentration of other straight chain components rang was from 37.88%to 64.89%.There was no significant relationship between the chemotypes and their geographical distributions.Totally,16 compounds were characterized from the essential oils(EO)of the underground parts of Houttuynia.There was no significant chemical difference between the EO of the underground parts of the different Houttuynia accessions.In this study,no significant chemical component difference was detected between the H.cordata and H.emeiensis accessions.2.The relationships between the genetic diversity among Houttuynia germplasm resources and the essential oil chemotypes were analyzed by using RAPD,RAMP,ISSR and PCR-RFLP DNA markers technology.The results showed that the ISSR marker was more effective to evaluate the relationships than RAPD,RAMP and PCR-RFLP markers on the evualation of the relationships between the genetic diversity and the essential oil chemotypes.The cluster results of genetic distances(GDs)based on ISSR marker were almost coincident with the cluster results of euclidean distances based on the essential oils chemical components.However,the results of matrix correlation showed,there were insignificant correlation between EDs based on the essential oils chemical components and GDs based on any kind of DNA markers.3.Through analyzing the UV spectrum of the essential oil of Houttuynia,it was found that theλmax of the essential oil of Houttuynia were at 252 nm and 267 nm in acetic ether solvent. Twenty-three samples could be divided into 5 types,which could not be coincident with the essential oil chemotypes.The differences of the absorbency of these samples at 252 nm and 267 nm were remarkable.Twenty-three samples counld be divided into 3 types based on the results of absorbency.All of the correlations between the contents of terpenes of EOs and the absorbencies of EOs at the two wavelengths were positive,and most of them reached 1% remarkable levlel.But all the correlations between the contents of unsaturated alkane of EOs and the absorbencies were negative,and all of them reached 5%or 1%remarkable levlel.4.The abilities to restrain the growths of Fusarium graminearum,Colletotrichum coccodes, Botrytis cinerea,Escherichia coli and Staphylococcus aureus were compared by using the different concentrations of D chemotype and M type essential oils.The results showed that the difference of the restraining abilities of the two chemotypes was obvious.The restraining abilities of D chemotype essential oil were stronger than that of M chemotype essential oil, especially to restrain the growths of the epiphytes.And the same chemotype EO had different abilities to restraine the different microbial species.5.The essential oil constituents of the fresh aerial parts of two lines were analyzed by GC/MS.The results of variance analysis showed that the essential oil of the two new lines w01-100 and w01-94 did not change significantly during daytime.The contents of the main components of the essential oil of w01-94 were more stable.Lines w01-100 and w01-94 can be harvested during the whole daytime.There were significant correlations between the content changes of some main components of the essential oil of the fresh aerial parts of Houttuynia,but the correlations between the two same components might be different in different materials.6.The flavone components in the residuary water solution after essential oil distillation were analyzed by using UV-VIS spectrum and TLC.The results of displaying colors identifications and UV-VIS spectrum showed that there were abundance flavone components in the residuary water solution,mainly included flavone,flavonol,isoflavone, dihydroflavone and dihydroflavonol.The results of TLC showed that most of these flavone components in the residuary water solution were glycoside components,containing rutin and hyperin.The average content of total flavone of 14 Houttuynia accessions was 1.93 mg/g, the content of w01-16 was the highest,reaching to 3.09 mg/g.The difference of the content of total flavone among the 14 Houttuynia accessions was distinct.There were insignificant correlationships between the types of total flavone components,contents of total flavone and the chromosome numbers of Houttuynia accessions.The recycled values of the residuary water solution were obvious.7.The difference of quercetin content of different Houttuynia accessions was determined by using HPLC on Hypersil ODS C18(250mm×4.0mm 5μm)at 30℃with acetonitrile:0.4% phosphoric acid(45:55),pH=3 as the mobile phase.Detection wavelength was at 370nm. The flow rate was 1.0mL/min.The calibration curve of quercetin was 22.43ng～2.24ug(r= 0.999 7).The quercetin content of the accession w01-120 was the highest,arriving at 4.02mg/g.8.The vitamin constituents and content of fresh young Houttuynia accessions were determined by using HPLC methods.The fat-soluble vitamins VA,δ-VE,VD₂,VD₃,α-VE and VK₁ were determined by Hyperclone BDS C₁₈column(150mm×4.6mm 5u)at 25℃with methanol:ethyl acetate(95:5)as the mobile phase at 275nm;the water-soluble vitamins VB₁, VB₂,VB₆,Nicotinic acid and V_C were determined by Luna C₁₈(2)100(?)column(200 mm×4.6mm 5u)at 30℃with 0.005mol/L hexane sulfonic sodium(pH 3.2):methanol(70:30) as the mobile phase.The flow rate was at 1.0 mL/min.Detection wavelength was set at 254nm to determine the content of VB₁,VB₂,VB₆,Nicotinic acid and V_C of Hauttuynia.The mainly vitamin constituents wereα-VE and VC,less of VD₃,VK₁ and Nicotinic acid in Houttuynia,others 6 vitamins not detected in this research.The vitamin constituents and contents of the aerial parts were more abundant than that of underground parts in Houttuynia. The difference of the vitamin constituents among Houttuynia resource were little,but were notable in the content of their vitamins.The nutrition value of the aerial parts was higher than that of underground parts.9.The remains of As and Hg in the young aerial and underground parts of 10 Houttuynia accessions and the soil cultivated these Houttuynia accessions were determined simultaneously by using AFS.The results showed that the content of As was 11.557mg/kg, Hg was 0.387mg/kg in soil,the mean content of As was 0.161mg/kg and Hg was 0.004mg/kg in the young aerial parts.And in the underground parts,the mean content of As was 0.130mg/kg and Hg was 0.0025mg/kg.The diference of the remains As and Hg among the Houttuynia accessions was insignificant.According to GB4810-94 and GB2762-94,the remains of heavy metals As and Hg were not beyond the standard.10.The remains of 666 and DDT in the young aerial and underground parts of 10 Houttuynia accessions and the soil were determined simultaneously by using GC.The results showed that the remains ofδ-666 in soil was 0.17ng/kg,but the trace remains of the young aerial and underground parts in several accessions were detected.According to GB/T14551-2003,the remains of pesticides 666 and DDT were not beyond the standard.To sum up,the differences of the self-metablism chemical components,such as the essential oils,flavones and vitamins were significant among the Houttuynia germplasm resources.And the variations on the rudimental contents of some heavy metals and pesticides were insignificant.The chemotyoes of EOs of fresh aerial parts of Houttuynia were closely related to the diversity of the chromosome numbers,while other chemical components were not related to the diversity of the chromosome numbers,obviously.No significant chemical component difference was detected between the H.cordata and H.emeiensis accessions.The different ehemotypes of EOs could cause the different effects on restraining the microbial species;and it was important to the drugs production and utlization,which used EOs of Houttuynia as the raw materials.