Separation And Purification Of Dicaffeoylquinic Acid From Mugwort And The Microbial Transformation Of Scutellarin

Artemisia lavandulaefolia is a medicinal plant with rich resources.On the basis of phenolic acids which were identified in A.lavandulaefolia,the work was carried out to study the extraction and purification process of dicaffeoylquinic acids,and the transformation of scutellarin.(1)This work was the first time to analyze caffeoylquinic acids detailedly in A.lavandulaefolia.A method using Liquid Chromatography and Negtive Electrospary Ionization Tandem Mass Spectrometry method was established for the identification of caffeoylquinic acids in the extract of A.lavandulaefolia.Eighteen caffeoylquinic acids were successfully identified.These caffeoylquinic acids comprised three monocaffeoylquinic acids,six dicaffeoylquinic acids,one tricaffeoylquinic acid,three feruloylquinic acids,three caffeoylferuloylquinic acids,one diferuloylquinic acid and one dimethoxycinnamoylquinic acid.Especially,the contents of monocaffeoylquinic acids and dicaffeoylquinic acids were higher than others.Rutin and acutellarin were also found in the extract.(2)This work found that the pretreatment to A.lavandulaefolia by petroleum ether will not have a significant impact on the dicaffeoylquinic acids,and essential oil was got.According to the principle of Box-Behnken experimental design,the three-level-four-factor experiment was designed.Based on the single-factor-test,response surface methodology was used to optimize the extraction variables,including ethanol concentration,extraction temperature,extraction time and solid-liquid ratio,for the achievement of high yield of diCQAs.The order of factors influencing the yield of diCQAs was as follows:ethanol concentration>extraction temperature>solid-liquid ratio>extraction time.The optimum conditions for extraction were:ethanol concentration 72%,extraction temperature 53?,extraction time 102 min,solid-liquid ratio 1:54(g/mL),and the yield was 3.30%while the extraction rate was 91%.(3)Macroporous resin CN205 was appropriate for the separation of dicaffeoylquinic acids in A.lavandulaefolia and the purity of dicaffeoylquinic acid was 48.4%.After the purification of macroporous resin,a column packed with sephadex was used to perform the secondary purification.The purity of dicaffeoylquinic acids was 82.3%.The three dicaffeoylquinic acid monomers were separated by preparative chromatography and the purity was 85%.The three dicaffeoylquinic acid monomers were 1,5-diCQA,3,5-diCQA,4,5-diCQA and the content of three diCQAs were 96.6 mg,107.4 mg and 61.4 mg respectively while the raw material was 15 g.(4)The content of flavonoid in A.lavandulaefolia was 1.33%and the content of scutellarin was 0.59 mg/g.The transformation of scutellarin was carried out by enzyme of Neurospora.The transformation product was identified to be scutellarein by LC/MS.Scutellarin was transformed effectively by the enzyme under the following conditions:temperature 55?,pH5.0,scutellarin concentration 2.0 mg/mL and time 90 min.The transformation rate was 72.3%.(5)Macroporous resin CN206 was used to isolate scutellarein.30%ethanol and 50%ethanol were used as eluent.Scutellarein mainly was retained in 50%ethanol fraction and the purity was 90%after purification.