Optimization and Application of FL-HPLC for Folates Analysis in 20 Species of Mediterranean Wild Vegetables P. Morales & V. Fernández-Ruiz & M. C. Sánchez-Mata & M. Cámara & J. Tardío Received: 25 February 2014 /Accepted: 23 April 2014 # Springer Science+Business Media New York 2014 Abstract Folic acid and folates (vitamin B 9 ) are essential for human metabolism. Their nutritional importance mainly lies in the prevention of neural tube formation defects, and other degenerative diseases. The determination of folates in food matrices is a very complex task due to the large number of chemical compounds included under the definition of folates. In the present study, an HPLC-fluorescence detection (HPLC- FL) methodology was optimized and validated to characterize total folates in vegetables. Its application to Mediterranean wild vegetables, provide new data on folates content on these species, scarcely studied so far. The highest folates contents were found in the leaves of Rumex pulcher L., Beta maritima L. and Anchusa azurea Mill. (506, 302 and 278 μg/100 g, respectively), as well as in the tender leaves and stems of Foeniculum vulgare Mill. and Silene vulgaris (Moench.) Garcke (272 and 268 μg/100 g, respectively). These data show that the studied wild vegetables are mostly a very good source of folates, compared to other folate-rich plants, such as spinach, parsley or chard. These under-utilized plants deserve attention for their potential value in current diets, as they are a good alternative for the diversification of our vegetable intake and represent an important added value as rich sources of vitamin B 9 . Keywords Folates . HPLC-FL . Wild vegetables Introduction Folates are a complex group of water-soluble compounds, known as vitamin B 9 . Folic acid (pteroil-glutamic acid) con- sists in a pteridine ring, linked by a methylene bridge to a residue of p-amino benzoic acid, which in turn is linked by amide bond to a glutamic acid residue (Fig. 1). More than 150 folates-derivatives exist (Scott 1999), depending on the sub- stituents linked to this basic structure (5-methylfolate; 5- formylfolate, 5,10-methylenfolate), the oxidation status (mono-, di- or tetrahydrofolic acid), or the number of glutamate residues linked by peptide bond to the pteroil structure. As folates are synthesised only by microorganisms and plants, humans depend on a variety of dietary sources for this vitamin intake, including liver, fresh dark leafy vegetables, legumes, wheat germ and yeast (Lucock 2000). In food, folates are naturally presented as polyglutamates (PteGlu n ), mainly as mono-, penta- and hexaglutamates (Scott 1999). Folates are necessary for one-carbon transfer reaction in a wide number of metabolic pathways in the human body, acting in purine and pyrimidine metabolism, and thus in DNA synthesis (Kim 1999). It has shown preventive effects against different tumor types (Gerber 2001), and some degen- erative diseases such as Alzheimer's, but its main nutritional importance lies in embryonic development due to its activity in the methylation cycle during tissues formation, being this compound closely related to the development of neural tube formation defects (Montgomery et al. 2003). Thus, low ma- ternal folate levels are associated with an increased risk of abnormal development of the newborn (Steegers 1995; Wright et al. 2010). Other important functions of folates are related to the metabolism of homocysteine, as it is known that low folate levels are correlated with hyperhomocysteine plas- ma levels, which are considered an independent risk factor for atherosclerotic and atherothrombotic damage through the P. Morales : V. Fernández-Ruiz : M. C. Sánchez-Mata (*) : M. Cámara Departamento Nutrición y Bromatología II. Bromatología, Facultad de Farmacia, Universidad Complutense de Madrid (UCM), Pza Ramón y Cajal, s/n., 28040 Madrid, Spain e-mail: cortesm@farm.ucm.es J. Tardío Instituto Madrileño de Investigación y Desarrollo Rural, Agrario y Alimentario (IMIDRA), Finca “El Encín”, Apdo. 127, 28800 Alcalá de Henares, Spain Food Anal. Methods DOI 10.1007/s12161-014-9887-6