Analysis Of Chemical Composition Of Dendrobii Officinalis Caulis

Dendrobii officinalis caulis is the stems of Dendrobium officinale Kimura et Migo belonging to orchid family.The fresh stems of D.officinale could be used as a medicine directly or dried and processed into Fengdou,strip or slice in the market.In recent years,with the development of cultivation techniques,various cultivation modes have been developed.However,due to depletion of wild resources and high cost of planting artificially,the stems of many other morphologically similar Dendrobium species were collected and processed.These stems are easily confused with the processed stem of D.officinale,even named as Dendrobii officinalis caulis to use by some illegal business behaviors for higher benefits in China,which may cause highly variable and inconsistent therapeutic effects.In order to identify accurately Dendrobii officinalis caulis and investigate changes of chemical component during cultivation and processing,the newly established thin layer chromatography(TLC)was used to distinguish D.officinale from its same genus species,the quantitative analysis of multi-components with a single marker(QAMS)methods combined with double wavelength HPLC method was established to analyze six flavonoids in Dendrobii officinalis caulis.And chemical composition of Dendrobii officinalis caulis was determined by the 2D-LC/MS method.1.Using violanthin and reference drug of Dendrobii officinalis caulis as the reference standards,the TLC behaviors of the D.officinale and other seven Dendrobium species were compared by using established TLC method.The results show that the identification method has strong stability and specificity,and D.officinale can be effectively distinguished from its related Dendrobium species.2.Established a simple HPLC method for fast and accurate determination of six flavonoids in D.officinale with a single reference standard strategy by calibrating the standard curve with internal standard and relative correction factor(RCF),and combined with double wavelength detection.The easily available compound rutin was selected as the internal reference compound for calculating the RCFs of other four flavonoids,including schaftoside,isoschaftoside,vicenin-2 and glucosylvitexin and calculating their contents at 340 nm,while naringenin was detected at 290 nm to determine its content.The established method indicated that there was no statistically significant difference compared with the traditional external standard method.The present results showed that a significant variation in the average total amount of sixflavonoids between D.officinale cultivated under greenhouse and imitative natural condition,and among D.officinale collected from different regions of China.The average amount of six flavonoids in D.officinale cultivated under imitative natural condition was significantly higher than that of D.officinale cultivated in greenhouse.And the total content of the six flavonoids in fresh slice was significantly higher than those of Fengdou and dried strip.The content of rutin decreased,while the content of naringenin increased with increasing the dried temperature of D.officinale.PCA analysis of the six flavonoids showed that the three principal components with a cumulative contribution rate of 87.43%,the first principal component was highly positively correlated with vicenin-2,schaftoside,and isoschaftoside,and the second principal component was positively correlated with naringenin.The third principal component was positively correlated with rutin.Pearson correlation analysis showed that naringenin was significantly positively correlated with vitaminin-2,schaftoside,isoschaftoside and glucosylvitexin.Isoschaftoside was significantly positively correlated with vicenin-2 and schaftoside,and schaftoside was significantly positively correlated with vicenin-2,while glucosylvitexin was significantly negatively correlated with isoschaftoside.The Pearson correlation analysis showed that the five flavonoids were synergistic active ingredients in D.officinale.3.Established the method of full two-dimensional liquid chromatography to isolate the chemical compounds of Dendrobii officinalis caulis,and combined the high-resolution mass spectrum to characterize qualitatively its chemical compound.Finally,61 components were identified according to their composition of molecular weights together with multi-stage mass fragmentation,retention time and relative abundance in comparison with those found in reference compounds and literature by the offline full two-dimensional liquid chromatography with high resolution mass spectrometry,including 8 flavonoid O-glycosides,13 flavonoid C-glycosides,5flavonoid aglycones,10 bibenzyl compounds and 25 other compounds,which are alkaloid,phenanthrene and sesquiterpene,coumarin,anthraquinone,organic acid,sugar and other types of compound.