Journal of Chromatography B, 871 (2008) 1–6 Contents lists available at ScienceDirect Journal of Chromatography B journal homepage: www.elsevier.com/locate/chromb One-step rapid determination and purification of puerarin from Radix puerariae by n-octylamine-modified poly(methacrylate-co-ethylene dimethacrylate) monolith Yong-Qin Lv a , Tian-Wei Tan a,∗ , Man-Yi Wang a , Jan-Christer Janson b a College of Life Science and Technology, Beijing University of Chemical Technology, Beisanhuan East Road 15#, Beijing 100029, China b Department of Surface Biotechnology, Uppsala Biomedical Centre, Uppsala University, P.O. Box 577, Uppsala SE-75123, Sweden article info Article history: Received 4 March 2008 Accepted 4 June 2008 Available online 11 June 2008 Keywords: Monolith Chromatography Radix puerariae Puerarin Isoflavonoids abstract n-Octylamine-modified poly(methacrylate-co-ethylene dimethacrylate) monoliths were prepared for rapid screening, determination and one-step purification of puerarin from Radix puerariae (a crude extract of the root of Pueraria lobata). The modified monolith showed a specific surface area of 17.8m 2 g -1 , an average pore size of 0.76 m and a total porosity of 60.8%. Fast separation of R. puerariae crude extract was achieved within 5 min at a flow velocity of 722 cm h -1 resulting in a puerarin purity of 97%, with a recov- ery of 85%. This demonstrates the potential of n-octylamine-modified poly(methacrylate-co-ethylene dimethacrylate) monolith for the rapid analysis and separation of isoflavonoids. Preparative scale sample loading (12 mg in 2 mL) resulted in a purity of 95%, and a recovery of about 69%. HPLC, FTIR, MS and 1 H NMR spectroscopy were used for the characterization and quantification of puerarin in isolated fraction. © 2008 Elsevier B.V. All rights reserved. 1. Introduction Traditional Chinese Medicines (TCMs) have been extensively used to prevent and cure many diseases in China, thanks to low toxicity and rare complications. The roots of Pueraria lobata (Wild.) Ohwi and Pueraria thomsonii Benth (Fabaceae family), two com- monly used Chinese herbal medicines, were officially included in the Chinese Pharmacopoeia (CP) until the year 2000 edition under the same name ‘Gegen’ (Radix puerariae, RP) [1]. Isoflavonoids, such as puerarin, daidzin, genistin, genistein and daidzein have been responsible for the broad therapeutic effects of RP con- nected to its estrogenic activity (phytoestrogens) [2–4]. Puerarin (7,4 ′ -dihydroxyisoflavone-8-glucopyranoside) with a concentra- tion of approximate 4% in RP has many pharmacological effects, such as anti-hypertension, anti-arteriosclerosis, dilating coronary arteries, decreasing myocardial oxygen consumption and improv- ing microcirculation in both animals and humans suffering from cardiovascular disease [5]. However, most of the TCMs have com- plicated and structurally related components. Therefore, a rapid, simple and effective method is required for the separation and purification of these substances. ∗ Corresponding author. Tel.: +86 10 64416691; fax: +86 10 64715443. E-mail addresses: yongqinlv.student@sina.com (Y.-Q. Lv), twtan@mail.buct.edu.cn, 2007080138@grad.buct.edu.cn (T.-W. Tan), jan-christer.janson@ytbioteknik.uu.se (J.-C. Janson). -Cyclodextrin (-CD) coupled to different media [2,6–9], high- speed counter-current chromatography [10] and extraction [11] were shown useful for the separation and purification of puerarin. Micellar electrokinetic capillary chromatography with electro- chemical detection [12], non-aqueous capillary electrophoresis [13] and HPLC [14–18] were also used for the separation of puerarin, however, primarily for analytical purposes and not for prepar- ative chromatography due to their very low binding capacities. Monolithic stationary phases, directly cast in tubes by radi- cal polymerization were developed by several groups [19–21]. They have high permeability and good mass transfer, leading to improved resolution at high linear flow rates. Monoliths based on poly(methacrylate-co-ethylene dimethacrylate) were usually mod- ified to ion-exchangers [22,23], reversed-phase chromatography media [24,25] and affinity chromatography media [26]. However, since the components of R. puerariae extracts contain aromatic rings and phenolic groups, it is reasonable to assume that hydrophobic interaction and hydrogen bonds may contribute to the separation process. In this paper, monoliths based on n-octylamine-modified poly(methacrylate-co-ethylene dimethacrylate) were shown to be efficient for rapid separation and purification of puerarin from crude extracts of R. puerariae in one step. This work suggests a new approach for the rapid separation of puerarin and intro- duces a new application of monolithic columns for fast separation of natural products. The dynamic binding capacity, adsorption characteristics and fast separation of puerarin were investigated. Compared with published literatures, n-octylamine-modified 1570-0232/$ – see front matter © 2008 Elsevier B.V. All rights reserved. doi:10.1016/j.jchromb.2008.06.010