| Qianghuo is the root and rhizome of the Umbelliferae plant Notopterygium incisum Ting ex H.T.Chang(NI)or N.franchetii H.de Boiss.(NF),mainly grows in regions of the high altitude such as Sichuan,Qinghai,Shanxi and Gansu,etc.It has been used to disperse cold,prevent painful obstructions from wind,damp and warm pain.Pharmacological research shows it has various therapeutic effects such as analgesic,anti-inflammatory,anti-cancer and antioxidants effects.Besides,it also has many new clinical efficacies such as anti-coagulation,anti-oxidation,anti-Alzheimer’s disease,etc.Qianghuo has a long history of medical use in China,and it also has systematic research on its chemistry and pharmacology since 1980s.Recently,there has been interested in difference between NI and NF.We choosed market popular varieties NI to study on its chemical ingredients and quality control based on summary of a large number of literatures with the five principles of Q-markers screening.First,ingredients of Qianghuo were classified and sorted out according to the principles of specificity,and the biosynthesis pathways were analyzed to select the specific components of Qianghuo.Then according to the principles of effectiveness,the active components that can represent both the medicinal properties and the medicinal effects were selected.According to their content in Qianghuo(Measurability),the selected compounds were carried on for fingerprint and quantitative analysis.More than 15 batches of NI were collected to get the common peaks in fingerprint.20 kg of the roots and rhizomes of NI was extracted and isolated based on the common peaks.The structures were determined by comprehensive using of LC-MS,1D-NMR,COSY,HSQC,HMBC and other spectroscopy methods.The isolated compounds can be used to identify the common peaks in fingerprint,as well as for quantitative analysis.According to literature summary and the "five principles" of Q-marker,we found that notopterol,which has good specificity and activity related to the traditional functions,is the best candidate for the Q-marker of Qianghuo.Falcarindiol,nodakenetin,isoimperatorin and phenethyl ferulate with strong activities but not enough specificity can be used as second candidates.Volatile oil with strong pharmacological activity is the material basis of Qianghuo’s "medicinal properties",it should be used as an important reference for Qianghuo Q-marker.Sixteen compounds were isolated from the roots and rhizomes of NI and identified as:nodakenetin(1),isoimperatorin(2),pregnenolone(3),trans-ferulic acid(4),bergapten(5),p-hydroxyphenethyl anisate(6),dimethoxylapigenin(7),6-methoxy-7-geranyloxycourmarin(8),phenethyl ferulic acid(9),notopterol(10),falcarindiol(11),ostruthine(12),ostruthene(13),8-bergapten(14),ostruthine(15),daidzein(16).13 is a new compound of coumarin;7,14,16 are isolated from NI for the first time;1,2,3,6,9,10 and 11 have higher content in NI,are the main component of NI;1,2,4,5,9,10 and 12 were used in fingerprint;2,9,10 and 11 were used in quantitative analysis.An efficient and convenient HPTLC method avoiding the use of controlled reagents such as toluene,chloroform etc.in the Pharmacopoeia for Qianghuo identification was established.The fingerprint of Qianghuo base on HPLC-DAD was established.14 common peaks were selected and 7 of them(nodakenetin,isoimperatorin,trans-ferulic acid,bergapten,phenethyl ferulic acid,notopterol,Ostruthine)were identified by comparing with those of standard substances.The similarity of 18 batches NI was between 0.887 and 0.992.Both HPTLC and fingerprint can effectively distinguish N.incisum,N.franchetii and other confusable varieties.For the multi-components quantitative analysis in NI,an HPLC method was established to detect quantity of phenethyl ferulate,notopterol,isoimperatorin and falcarindiol.The four peaks were separated well and had a good linear relationship(r>0.9998)under the condition.The average recovery rates were 95.3%,99.5%,98.5%and 98.5%,respectively.The results show that the content of active components of Qianghuo varies greatly among different batches.In the future,clustering analysis could be considered base on different origins,different growth years and commodity specifications of Qianghuo to establish the classification standard and detection method of Qianghuo.This study provides theoretical guidance for comprehensively improving the quality control level of Qianghuo. |
