Anthocyanin profiles in south Patagonian wild berries by HPLC-DAD-ESI-MS/MS

Abstract

Chilean Patagonia is one of the most beautiful natural scenarios, with a great diversity of habitats and vascular plants, including different berry fruits. There is no scientific information about the polyphenol profiles of most of these fruits, as well as about their potential use as functional food.

Samples of 10 different berry species were collected in the Magallanes region in the extreme South of Chile. Their anthocyanin profiles were studied using liquid chromatography with photodiode array and mass spectrometry detection on the basis of retention times, UV and MS/MS spectra. In addition, total anthocyanin concentrations obtained by HPLC-DAD, antioxidant capacity estimated by TEAC (as trolox equivalent), and ascorbic acid contents were determined in these fruits. The studied species differed in their berry anthocyanin profiles and concentrations. In the berries of Berberis genus (Berberis microphylla, Berberis empetrifolia and Berberis ilicifolia), a predominance of delphinidin, petunidin and malvidin 3-glucoside was observed; they are the fruits with the highest total anthocyanin concentration (22.91–35.99, 16.11–21.40 and 13.70 μmol/g, respectively). In Ribes magellanicum and Ribes cucullatum, the 3-glucoside and 3-rutinoside derivates of cyanidins predominated, especially for R. cucullatum, showing intermediate total anthocyanin concentrations. The other studied berries (Gaultheria mucronata, Gaultheria antarctica, Rubus geoides, Myrteola nummularia and Fuchsia magellanica) presented as main anthocyanidin cyanidin and/or delphinidin derivates, with lower total concentrations in comparison with the other studied species. Antioxidant activity and especially the ascorbic acid concentration observed in these fruits were higher than those described for other widely consumed berries, reaching levels up to 75.1 μmol/g and 198.8 mg/100 g, respectively. These findings suggest that some of the berry species from Patagonia have an interesting potential to be used as functional food.

Introduction

Fruits and vegetables contain several health-promoting components, including dietary fiber and high concentrations of phenolic acids, flavonoids, vitamins and minerals. Berry fruit consumption has become important in population health promotion, mainly due to their phenolic compounds, which have been associated with protection against different pathologies, e.g., several types of human cancer and because of their anti-inflammatory, gastroprotective, antimicrobial and other biological activities (Basu et al., 2010, Paredes-López et al., 2010). One of the most important groups of phytochemical compounds in berries is represented by anthocyanins, a major cluster of water-soluble pigments from the flavonoid group. Anthocyanins exist abundantly in the plant kingdom and confer the blue, red, violet and purple colors to the fruits and vegetables, including berries (Clifford, 2000). Anthocyanins in fruits exist primarily as glycosides or acylglycosides of their corresponding aglycone, which are known as anthocyanidins. Recent studies provide convincing evidence that anthocyanins possess potent anticarcinogenic properties. Between them, Huang et al., 2008 showed that a Mulberry anthocyanin-rich extract can inhibit melanoma metastasis and can potentially inhibit gastric cancer (Huang, Shih, Chang, Hung, & Wang, 2008). Other studies of these authors show that Hibiscus anthocyanin extracts mediated the apoptosis of human promyelocytic leukemia cells via the p38/Fas and Bid pathway (Chang, Huang, Hsu, Yang, & Wang, 2006). In the same context, Bishayee et al., (2011) proposed that an anthocyanin-rich extract from blackcurrant exerts a striking chemopreventive effect against experimentally induced in vivo hepatocarcinogenesis in rats (Bishayee et al., 2011). These and other studies show the importance of berry anthocyanins in promoting human health.

Chilean Patagonia is one of the most beautiful natural scenarios, with a great diversity of habitats and vascular plants, including different wild edible berry fruits from Berberidaceae, Grossulariaceae, Ericaceae, Rosaceae, Myrtaceae, Onagraceae and other families (Vidal, 2007). Human populations consume these plants since pre-hispanic times, being known by ancestral cultures as medicinal plants. In folk medicine, the laxative and antipyretic effects of infusions of some Chilean native wild species of the Berberidaceae family (Donoso & Ramirez, 2005) as well as the diuretic and also antipyretic effects of Fuchsia magellanica from the Onagraceae family (Hoffmann, 2005) are known. In a very recent publication, the inhibition of cancer cell proliferation of Myrteola nummularia has been described (Flis et al., 2012). However, there is a lack of scientific information about anthocyanin profiles of most of these edible berries and their concentrations. Only one of them, “calafate” (Berberis microphylla), has been previously characterized by high performance liquid chromatography with photodiode array and mass spectrometry detection (HPLC-DAD-ESI-MS/MS), showing an interesting anthocyanin composition and reaching high anthocyanin contents (Ruiz et al., 2010). Predominant anthocyanins in calafate correspond to 3-glucoside derivatives of delphinidin, petunidin and malvidin; 18 anthocyanins have been identified in total. The higher antioxidant capacity shown by calafate compared with other berries such as “murtilla” (Ugni molinae) and blueberries has been attributed to their high anthocyanin content (Ruiz et al., 2010). In a recent publication, the calafate fruit has been classified as the fruit with the highest antioxidant capacity measured by the oxygen radical absorbance capacity (ORAC) assay (Speisky, López Alarcón, Gómez, Fuentes, & Sandoval Acuña, 2012).

Other interesting wild species of the Berberidaceae family from Chilean Patagonia are “calafatillo” (Berberis empetrifolia) and “michay” (Berberis ilicifolia), both species being widely distributed along the south half of Chile. The first is a dwarf shrub with linear leaves that give sub-globular dark blue berries of 4 to 10 mm, whereas “michay” is a large and well-branched shrub with larger dark blue berries (Marticorena & Rodriguez, 2003). From the Grossulariaceae family, it is possible to find Ribes magellanicum, known as wildcurrant (“parra silvestre” or “parrilla” in Spanish) and Ribes cucullatum (called “parrillita”). Both shrubs produce light purple globular berries (Hoffmann, 2005, Vidal, 2007). From the Ericaceae family, 2 different globular berries with a dry calyx at its base grow in this zone. They are Gaultheria mucronata, called “prickly heath” or “chaura” and Gaultheria antarctica, locally known as “mutilla,” both widely distributed in the south Patagonia. From the Rosaceae family, Rubus geoides is a wild raspberry known as rainberry (“frambuesa silvestre,” “frutilla silvestre” or “frutilla de Magallanes” in Spanish). This plant is a small perennial and creeping herb that produces a reddish polydrupe of excellent taste (Vidal, 2007). M. nummularia is a Myrtaceae that produces a pink-red small fruit known as “sarapito” or “daudapo” by the local population with remarkable antioxidant activity (Arancibia-Avila et al., 2012), whereas F. magellanica, which is usually used as an ornamental plant, produces an elongated pink fruit known as “chilco.” The last 2 species are reported to occur between Coquimbo and Tierra del Fuego, covering almost the whole territory of Chile, except the desert in the North of the country. As far as we know, there are no scientific publications concerning the anthocyanin profiles or concentration levels of these compounds for the aforementioned wild edible fruits.

In this study, the anthocyanin profiles in the main, wild, edible berry fruits collected in the Magallanes region, in south Chilean Patagonia, have been determined by HPLC-DAD-ESI-MS/MS, together with the individual and total concentration of these anthocyanins. In addition, the results have been discussed with the objective to find some connection between the measured antioxidant capacity and the concentration of the detected anthocyanins and a main antioxidant such as ascorbic acid.