Available online at www.sciencedirect.com Talanta 74 (2008) 1218–1223 Pressurized solvent extraction and monolithic column-HPLC/DAD analysis of anthocyanins in red cabbage Panagiotis Arapitsas a,b,∗ , Charlotta Turner a a Uppsala University, Department of Physical and Analytical Chemistry, Analytical Chemistry, P.O. Box 599, 751 24 Uppsala, Sweden b University of Florence, Department of Pharmaceutical Sciences, Via Ugo Schiff 6, 50019 Sesto F.no, Florence, Italy Received 20 May 2007; received in revised form 10 August 2007; accepted 24 August 2007 Available online 1 September 2007 Abstract The aim of this work was to develop a fast method for extraction and analysis of anthocyanins in red cabbage. Pressurized hot water containing 5% of ethanol was used as an extremely efficient extraction solvent. HPLC/DAD with a monolithic column was used to accomplish a fast analysis—24 anthocyanin peaks within 18 min. Statistical design was used to optimize the studied extraction parameters: temperature (80–120 ◦ C); sample amount (1–3 g); extraction time (6–11 min); concentration of formic acid in the extraction solvent (0–5 vol.%). The best extraction conditions for a majority of the anthocyanin peaks were 2.5 g of sample, 99 ◦ C (at 50 bar), 7 min of extraction and a solvent composition of water/ethanol/formic acid (94/5/1, v/v/v). © 2007 Elsevier B.V. All rights reserved. Keywords: Anthocyanins; Red cabbage; Monolithic column; Accelerated solvent extraction; Polyphenols 1. Introduction Red cabbage (Brassica oleracea L. var. capitata f. rubra) belongs to the family of Brassicaceae, and is a native vegetable of the Mediterranean region and south-western Europe that now grows in regions all over the world. Studies concerning the influ- ence of Brassica species in human health have verified their capacity to prevent cardiovascular diseases as well as their activ- ity against some types of cancer [1–5]. Among the substances that seem to be responsible for those properties are polyphenols [2,3,5], and red cabbage is a rich source of phenolic compounds, with the anthocyanins being the most abundant class [5–8]. Anthocyanins are a group of plant pigments that are widely dis- tributed in nature, among flowers, fruits and vegetables, and are responsible for their bright colors such as orange, red and blue [6,9,10]. They play an important role in the plant physiology and are valuable for food industry as well as in human health [2,5–7,9,10]. Red cabbage is a promising source of anthocyanins ∗ Corresponding author at: University of Florence, Department of Pharma- ceutical Sciences, Via Ugo Schiff 6, 50019 Sesto Fiorentino, Florence, Italy. Tel.: +39 055 4573677; fax: +39 055 4573667. E-mail address: panagiotis.arapitsas@unifi.it (P. Arapitsas). for the coloration of food since they are unique in being colored over a very broad pH-range, from red to blue and green, com- pared to other red vegetables [6,9]. Therefore, anthocyanins are not limited to acidic foodstuffs but can be extended to neutral- pH products, for which they may provide a natural alternative to synthetic blue colorants [9]. Regarding the chemical structure of red cabbage anthocyanins, only cyanidin has been identified as aglycon, glycosylated and acylated with various aromatic and aliphatic groups at positions 3 and 5 [5,6,8,9] (Fig. 1). The analysis of anthocyanins is a difficult task, due to their low stability in high temperature, light and pH above 3.5 [6,7,9,11–17]. Hence, food and pharmaceutical industries request the development of fast and efficient analysis meth- ods for these unstable compounds. Traditionally, the techniques employed in the extraction of anthocyanins from semi-solid or viscous samples involve the use of solvent extraction followed by liquid–liquid extraction or solid-phase extraction [5–9,11–15]. These methods are time consuming and use large amounts of organic solvent with negative consequences in the environment. Pressurized fluid extraction (PFE) is an alternative and innova- tive technique that combines elevated temperature and pressures with liquid solvents to achieve fast and efficient extraction [18–25], that has already been reported in the delicate extrac- tion of anthocyanins from fruits and vegetables [15,17], and has 0039-9140/$ – see front matter © 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.talanta.2007.08.029