title: Preparation of a super-long two column chromatography system and its application in separating glycosylated puerarin.
abstract: Separation of Puerarin-7-O-glucoside from its precursor, puerarin, using a common chromatography column packed with AB-8 macroporous resin was unsuccessful. Therefore, in this study a 8 m super-long flexible reinforced PVC column was externally added to the common column in order to improve the chromatography efficiency by increasing the number of theoretical plates. Both the PVC and common columns were separately packed with AB-8 macroporous resin slurry. The packed PVC column was coiled after washing and stored until use. The microbial transformation mixture with puerarin-7-O-glucoside and puerarin (250 mL) was loaded onto the common column, followed by washing with 2000 mL H(2)O. After attaching the coiled external PVC column to the common column, a linear gradient of 10-30% ethanol was applied to elute the target compound. Two peaks appeared: peak I contained puerarin-7-O-glucoside at 97.9% purity and 88.1% recovery rate, and peak II was puerarin at 98.7% purity and 87.0% recovery rate. The use of the coiled external flexible reinforced PVC column avoided spatial restriction for long columns, which made it much more convenient for column packing and chromatography operations. Furthermore, this method eliminated the resin blockage problem caused by stationary water pressure in a rigid vertical long column. Using an external super-long column, the PVC tube was connected with the common column only during elution, which avoided delay in time period during sample loading and column washes associated with the use of long external columns.
authors: Jiangsu Engineering and Technology Research Center for Industrialization of Microbial Resources, Jiangsu Key Laboratory for Biodiversity and Biotechnology, College of Life Science, Nanjing Normal University, Nanjing, PR China.
authors: Shouchuang
authors: S
authors: Zhu
authors: Haidong
authors: H
authors: Xu
authors: Cigang
authors: C
authors: Yu
authors: Juanfang
authors: J
authors: Ding
authors: Ting
authors: T
authors: Chen
authors: Jierong
authors: J
authors: Jiang
authors: Yijun
authors: Y
authors: Dai
authors: Guiyou
authors: G
authors: Liu
authors: Guodong
authors: G
authors: Huang
authors: Nan
authors: N
authors: Chai
authors: Xuhua
authors: X
authors: Jiang
authors: Sheng
authors: S
authors: Yuan
mesh_terms: D001711:Biotransformation; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation*; D006031:Glycosylation; D007529:Isoflavones / Q000302:isolation & purification* / Q000493:pharmacokinetics; D013056:Spectrophotometry, Ultraviolet
chemical_list: D007529:Isoflavones; C033607:puerarin