VOL. 12, NO. 2, JANUARY 2017 ISSN 1819-6608 ARPN Journal of Engineering and Applied Sciences © 2006-2017 Asian Research Publishing Network (ARPN). All rights reserved. www.arpnjournals.com 626 IDENTIFICATION AND QUANTIFICATION OF ANTHOCYANINS IN MUSCADINE GRAPES BY HPLC AND HPLC-MS Eduardo Pastrana-Bonilla 1 , Casimir C. Akoh 2 and Nestor Enrique Cerquera 1 1 Universidad Surcolombiana, Avenida Pastrana-Carrera 1, Neiva, Huila, Colombia 2 Universityof Georgia, Athens, USA E-Mail: pastrana@usco.edu.com ABSTRACT Total anthocyanin content and individual anthocyanin profile of ten cultivars of muscadine grapes were assessed. Total anthocyanin content was determined by a pH differential method. Individual anthocyanins were analyzed by HPLC and their identity confirmed by HPLC-MS. The total anthocyanin content and the sum of the individual anthocyanins had a high correlation (R = 0.98). The average anthocyanin content of muscadine grapes was lower than published values for red European and other American red grapes. However, the purple muscadine grapes have anthocyanins levels that may be considered important from the nutraceutical point of view. Keywords: anthocyanins, food analysis, muscadine grapes, polyphenols, vitisrotundifolia. 1. INTRODUCTION Muscadine grapes (VitisrotundifoliaMichx.) are indigenous to the southeastern United States. Muscadines are vigorous vines that may grow up to 30 meters in the wild. They differ botanically from other grapes and are placed in a separate sub-genus, Muscadinia. Muscadine fruits are round, 2.5 to 3.5 cm in diameter with thick, tough skin and may have up to 5 seeds (Pastrana-Bonilla, Akoh, Sellappan, and Krewer, 2003). Anthocyanins are water soluble, glycosy1ated derivatives based on the cyanidinaglycon(Chandra, Rana, and Li, 2001; J.P. Goiffon, M. Brun, and M.J. Bourrier, 1991) and are part of the flavonoid family. These pigments are responsible for the red, purple and blue colors of most fruits and flowers (da Costa, Nelson, Margolis, and Derek, 1998). Anthocyanins have the potential to be used as natural food colorants (Wu et al., 2014). However, enzymes, pH, temperature, and oxygen affect their color quality (Chandra et al., 2001; Wang, Tong, Chen, and Gangemi, 2010). Co-pigmentation with other phenolic compounds may increase their stability (Darias-Martı ́ n, Carrillo, Dı ́ az, and Boulton, 2001). Anthocyanins are part of the human diet and they occur in many fruits and vegetables (Sellappan, Akoh, and Krewer, 2002). The average daily intake of anthocyanins in the United States was estimated at 215 mg during the summer and 180 mg during the winter (Clifford, 2000). The beneficial antioxidant activity and therefore, positive health effects of anthocyanins are a significant added value for their use as food colorants (Wu et al., 2014). The antioxidant function of anthocyanins seems to be related to their hydrogen donation capacity, metal chelation and protein binding (SatueGracia, Heinonen, and Frankel, 1997). Anthocyanins have been found to be powerful antioxidants in comparison to other common antioxidants like butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT) and a-tocopherol (Espín, Soler-Rivas, Wichers, and García-Viguera, 2000; Narayan, Akhilender Naidu, Ravishankar, Srinivas, and Venkataraman, 1999). Anthocyanin-rich fruit extracts have been used in traditional medicine as anti-inflammatory agents (Wang etal., 2010), for the treatment and prevention of vascular diseases due to cholesterol-induced atherosclerosis, and as anti-carcinogenic agents (Lauro and Francis, 2000; Xu et al., 2015). Anthocyanins have also been reported to have antiulcer activity and to provide protection against UV radiation (Mazza and Miniati, 1993). Possible mechanisms for the anti-inflammatory activity of anthocyanins include inhibition of arachidonic acid metabolism and the prostaglandin synthase cyclooxygenase activity (Aruoma and Cuppett, 1997). Anthocyanins in red wine may have antiatherogenic effects in conjunction with other polyphenols found in the wine (Frankel, German, Kinsella, Parks, and Kanner, 1993). Kamei et al. (1995)Studied the in vitro anticarcinogenic effect of anthocyanins on tumor cells. Glycosides of the aglyconscyanidin and delphinidin have been found to be the most abundant anthocyanins in plants (Meiers et al., 2001). Cyanidin lowered serum thiobarbituric acid-reactive substance (TBARS) concentration and increased the oxidation resistance of the serum to lipid peroxidation in rats (Tsuda, Horio, Kitoh, and Osawa, 1999). Delphinidin has been reported to inhibit the growth of human tumor cell line by shutting off the epidermal growth-factor receptor downstream signaling cascade (Tsuda et al., 1999; Zheng et al., 2011). Mazza and Miniati and Talcott and Lee (S. T. Talcott and J. H. Lee, 2002) reported the presence of anthocyanidindiglucosides as the anthocyanins present in muscadine grapes. The objective of this paper was to identify and quantify the anthocyanins present in 10 cultivars of muscadine grapes grown in South Georgia, USA. 2. MATERIALS AND METHODS Chemicals: Standards (with more than 97% purity) of malvidin-3-O-β-glucopyranoside (mv-3-gl), delphinidin-3-O-β-glucopyranoside (dp-3-glc) petunidin- 3-O-β-glucopyranoside(pt-3-glc), peonidin-3-O-β- glucopyranoside (pn-3-glc), and cyanidin-3-3-O-β- glucopyranoside (cy-3-glc) were purchased from Polyphenols Laboratories AS (Sandnes, Norway).