Journal of Chromatography A, 1077 (2005) 188–194 Separation of eight selected flavan-3-ols on cellulose thin-layer chromatographic plates Irena Vovk a,∗ , Breda Simonovska a , Heikki Vuorela b a Laboratory for Food Chemistry, National Institute of Chemistry, Hajdrihova 19, SI-1001 Ljubljana, Slovenia b Division of Pharmacognosy, Department of Pharmacy, P.O. Box 56, FIN-00014 University of Helsinki, Helsinki, Finland Received 28 July 2003; received in revised form 22 March 2005; accepted 31 March 2005 Abstract The potential of microcristaline cellulose as sorbent in the separation of eight compounds: (+)-catechin (C), (-)-epicatechin (EC), (-)- gallocatechin (GC), (-)-epigallocatechin (EGC), (-)-epicatechin gallate (ECg), (-)-epigallocatechin gallate (EGCg), procyanidin B1 and procyanidin B2 was studied. Cellulose HPTLC plates prewashed in water (not necessary, when water was used as developing solvent) and dried with a hair dryer, bandwise application and development in horizontal developing chamber (sandwich configuration) gave the best results. Detection was performed using vanillin–H 3 PO 4 reagent. Four new developing solvent systems were proposed: water, 1-propanol–water (20:80, v/v), 1-propanol–water–acetic acid (4:2:1, v/v) and 1-propanol–water–acetic acid (20:80:1, v/v), and at least two of them were needed for the differentiation between all eight compounds. Surprisingly, water enabled the separation of epimers C from EC and GC from EGC, as well as the dimers procianidin B1 and B2. Additionally, C, EGC, B1 and B2 were separated from all the other compounds. The best choice for developing solvent is given for each of the studied compounds. The best separation of the five main catechins (EC, GC, EGC, ECg, EGCg) present in green tea extract was achieved using 1-propanol–water–acetic acid (20:80:1, v/v). The chromatograms of oak bark extract developed in solvents with higher water content (1-propanol–water (1:4, v/v) and 1-propanol–water–acetic acid (20:80:1, v/v)) showed less bands than chromatograms developed in solvents with higher organic modifier content (e.g. 1-propanol–water–acetic acid (4:2:1, v/v)). It was proved that such behavior was due to the presence of procyanidins beside the main component catechin. © 2005 Published by Elsevier B.V. Keywords: Flavan-3-ols; Catechins; Procyanidins; Thin-layer chromatography; Cellulose HPTLC plates; Green tea; Oak 1. Introduction Flavan-3-ols (catechins) are polyphenolic compounds that have been shown to exhibit a wide variety of beneficial ef- fects on human health [1–3] when using in appropriate con- centrations. These compounds are present in relatively high concentrations in tea leaves [4,5], oak [6], cocoa beans [7,8], grapes [9,10] and many other plants. They are also the build- ing blocks of oligomeric procyanidins (condensed tannins). The analysis of flavan-3-ols has usually been performed by HPLC using UV, electrochemical or MS detection [10–16] and also by thin-layer chromatography (TLC) ∗ Corresponding author. Tel.: +386 1 4760 341; fax: +386 1 4760 300. E-mail address: irena.vovk@ki.si (I. Vovk). [7,10,16–21]. The resolution of critical pairs in some TLC studies on silica gel [16,19] is relatively poor and does not allow reliable separation, although it is sometimes sufficient for qualitative screening purposes. Recently, some new TLC systems for the separation of (+/-)-catechin and (-)-epicatechin based on different modified silica sorbents [17] and cellulose [18,20,21] as sorbent were published. Additionally, TLC cellulose plates and pure water as developing solvent enabled us to distinguish between the (+)-catechin and (+/-)-catechin [21]. The aim of this study was to investigate the separation of selected flavan-3-ols and dimeric procyanidins (Fig. 1) using cellulose TLC or HPTLC plates. The applicability of the de- veloped method was checked for screening of the extracts of oak bark and green tea. 0021-9673/$ – see front matter © 2005 Published by Elsevier B.V. doi:10.1016/j.chroma.2005.03.096