Available online at www.sciencedirect.com Journal of Chromatography A, 1182 (2008) 56–64 Further knowledge on barley (Hordeum vulgare L.) leaves O-glycosyl-C-glycosyl flavones by liquid chromatography-UV diode-array detection-electrospray ionisation mass spectrometry F. Ferreres a,∗ , P.B. Andrade b , P. Valent˜ ao b , A. Gil-Izquierdo a a Research Group on Quality, Safety and Bioactivity of Plant Foods, Department of Food Science and Technology, CEBAS (CSIC), P.O. Box 164, 30100 Campus University Espinardo, Murcia, Spain b REQUIMTE/Servi¸ co de Farmacognosia, Faculdade de Farm´ acia, Universidade do Porto, R. An´ ıbal Cunha, 164, 4050-047 Porto, Portugal Received 17 October 2007; received in revised form 10 December 2007; accepted 17 December 2007 Available online 3 January 2008 Abstract Thirty-seven flavonoids and a hydroxycynnamic acid have been characterized in barley leaves (Hordeum vulgare L.) by liquid chromatography- UV diode-array coupled to ion trap mass spectrometry with electrospray ionisation interface (negative mode). Their structures have been determined by the study of the ion mass fragmentation which characterizes C-glycosyl flavones and O-glycosyl-C-glycosyl flavones, and differentiates di-O-glycosyl flavones from O-diglycosyl-flavones. The majority of them are described for the first time in barley. Saponarin (isovitexin-7-O- glucoside), lutonarin (isoorientin-7-O-glucoside) and isoscoparin-7-O-glucoside derivatives constitute the major part of the detected compounds. Some acylated derivatives are also described, namely, 7-O-[6-acyl]-glucoside and -7-O-[6-acyl]-glc-4 ′ -glucoside of isovitexin, isoorientin and isoscoparin. © 2007 Elsevier B.V. All rights reserved. Keywords: O-Glycosyl-C-glycosyl flavones; Acylated derivatives; Mass spectrometry; Hordeum vulgare 1. Introduction Barley is a plant widely distributed and cultivated with food goals, thus, its economical importance is very marked. By these reasons, it has been studied from different point of views. Among them, its phenolic content (flavonoids and hydroxycinnamic acids) have been analyzed as related to its antioxidant activity. Numerous reports have described the occurrence of C-glycosyl flavones in this plant. However, the difficulties of the C C bond hydrolysis have slowed down the advance on the study of these compounds up to the last decade, when the use of 1 H NMR and 13 C NMR helped in the structure confirmation of these type of compounds [1–5]. The majority of the described compounds are saponarin (isovitexin-7-O-glucoside) and lutonarin (isoorientin- 7-O-glucoside) derivatives (Fig. 1). Acylated derivatives, with hydroxycinnamic acids, of isoorientin and isovitexin-7-O-[6- ∗ Corresponding author. E-mail address: federico@cebas.csic.es (F. Ferreres). acyl]-glucoside and -7-O-[6-acyl]-glc-4 ′ -glucoside have been also reported (Table 1) [1–5]. The use of liquid chromatography-UV diode array detection coupled to ion trap mass spectrometry with electrospray ionisa- tion interface on the screening of polyphenol profiles of plant extracts have input a great advance for the tentative study of these compounds, since the knowledge of their structure can be approximately achieved without previous isolation and clean-up of the sample. Besides, the presence of these compounds can be detected at low concentrations or in traces. In this way, the ions obtained as a result of the preferen- tial MS fragmentation of flavonoid glycosides, as well as their relative abundance, may indicate the type of interglycosidic link- ages, differentiating O-rutinoside (rhamnosyl(1→6)glucoside) from O-neohesperidoside (rhamnosyl(1→2)glucoside) [6,7] and O-gentiobioside (glucosyl(1→6)glucoside) from sophoro- side (glucosyl(1→2)glucoside) [8], among other information. In addition, it is possible to differentiate di-O-glycosides from O-diglycosides of flavonoid derivatives [6–8]. In the mono-C and di-C-glycosyl flavones, the aglycone information, the type 0021-9673/$ – see front matter © 2007 Elsevier B.V. All rights reserved. doi:10.1016/j.chroma.2007.12.070