J. Agric. Food Chem. 2004, 52, 523−530
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Effect of Ripeness and Postharvest Storage on the Phenolic Profiles of Cherries (Prunus avium L.) BERTA GONC¸ ALVES,*,† ANNE-KATRINE LANDBO,‡ DAVID KNUDSEN,‡ ANA P. SILVA,§ JOSEÄ MOUTINHO-PEREIRA,† EDUARDO ROSA,§ AND ANNE S. MEYER‡ Department of Biological and Environmental Engineering, CETAV, and Department of Plant Science and Agricultural Engineering, CECEA, University of Tra´s-os-Montes e Alto Douro, Apartado 1013, 5001-911 Vila Real, Portugal, and Food Biotechnology and Engineering Group, BioCentrum-DTU, Building 221, Technical University of Denmark, 2800 Lyngby, Denmark
The phenolic compounds hydroxycinnamates, anthocyanins, flavonols, and flavan-3-ols of sweet cherry cultivars Burlat, Saco, Summit, and Van harvested in 2001 and 2002 were quantified by HPLC-DAD. Phenolics were analyzed at partially ripe and ripe stages and during storage at 15 ( 5 °C (room temperature) and 1-2 °C (cool temperature). Neochlorogenic and p-coumaroylquinic acids were the main hydroxycinnamic acid derivatives, but chlorogenic acid was also identified in all cultivars. The 3-glucoside and 3-rutinoside of cyanidin were the major anthocyanins. Peonidin and pelargonidin 3-rutinosides were the minor anthocyanins, and peonidin 3-glucoside was also present in cvs. Burlat and Van. Epicatechin was the main monomeric flavan-3-ol with catechin present in smaller amounts in all cultivars. The flavonol rutin was also detected. Cultivar Saco contained the highest amounts of phenolics [227 mg/100 g of fresh weight (fw)] and cv. Van the lowest (124 mg/100 g of fw). Phenolic acid contents generally decreased with storage at 1-2 °C and increased with storage at 15 ( 5 °C. Anthocyanin levels increased at both storage temperatures. In cv. Van the anthocyanins increased up to 5-fold during storage at 15 ( 5 °C (from 47 to 230 mg/100 g of fw). Flavonol and flavan-3-ol contents remained quite constant. For all cultivars the levels of phenolic acids were higher in 2001 and the anthocyanin levels were higher in 2002, which suggest a significant influence of climatic conditions on these compounds. KEYWORDS: Cherry; Prunus avium; ripeness; storage; HPLC-DAD; phenolics; anthocyanins; flavan-3ols; flavonols; hydroxycinnamates.
INTRODUCTION
High fruit and vegetable consumption is associated with a reduced risk of major chronic diseases such as cancer, atherosclerosis, and cardiovascular disease (1-4). Phenolic compounds have been widely identified in fruits and vegetables and partly related to those protective effects (5-7). The phenolic composition of plant foods is dependent on plant genetic information (8) and environmental factors during growth and postharvest. Cherries are considered to be a major source of phenolic compounds, which are also responsible for their color and taste and presumably also their antioxidant properties. Cherry phenols include flavonoids such as anthocyanins, flavan3-ols, and flavonols in addition to the nonflavonoid compounds hydroxycinnamic acids and hydroxybenzoic acids (9, 10). Fruit tissues are able to synthesize phenolic compounds, and changes in this content can be induced by biotic and abiotic * Corresponding author (telephone +351 259 350740; e-mail bertag@ utad.pt). † Department of Biological and Environmental Engineering, CETAV. ‡ Food Biotechnology and Engineering Group, BioCentrum-DTU. § Department of Plant Science and Agricultural Engineering, CECEA.
stress conditions (8, 11). Water availability and soil composition (mineral and organic nutrients) have a marked effect on the phenolic content of plants (12) and on the ability of plant products to suffer browning and other phenolic-related physiological disorders that appear during the maturity stage and postharvest (13). Storage at low temperatures might have positive or negative effects on phenolics and in turn on fruit quality, depending on the commodity and the storage temperature (13, 14). Cherries are nonclimacteric fruits that are usually picked at peak maturity for optimal taste and appearance. However, in Portugal, and in other countries that produce sweet cherries for fresh consumption, the cherries are often stored for up to 3-4 weeks at cold temperatures to increase the seasonal supply. To our knowledge, information about the changes in the phenolic content of cherries during maturity and postharvest storage is limited; however, this information is relevant to the understanding of the parameters that affect fruit color and stability and the potential health protective effects of the phenolics. The aims of this study were to (1) identify and quantify individual phenolic compounds in sweet cherries at partially ripe and ripe
10.1021/jf030595s CCC: $27.50 © 2004 American Chemical Society Published on Web 01/15/2004
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J. Agric. Food Chem., Vol. 52, No. 3, 2004
Table 1. Quality Indices of Cherries at Two Ripeness Stages (Partially Ripe and Ripe)a cv. Burlat Saco Summit Van
Burlat Saco Summit Van
a
stage
wt (g)
skin color a* value
partially ripe ripe partially ripe ripe partially ripe ripe partially ripe ripe
6.2 ± 0.6 d 7.4 ± 0.9 e 4.3 ± 0.4 a 5.0 ± 0.6 b 6.6 ± 0.8 d 9.0 ± 1.2 f 5.6 ± 0.6 c 7.4 ± 0.7 e
Year 2001 36.4 ± 6.8 bc 36.2 ± 5.9 bc 42.0 ± 2.1 bc 35.8 ± 10.3 e 38.5 ± 3.0 ab 34.2 ± 10.2 cd 40.9 ± 2.3 a 32.8 ± 11.3 de
partially ripe ripe partially ripe ripe partially ripe ripe partially ripe ripe
4.2 ± 0.6 a 7.2 ± 0.8 c 4.3 ± 0.8 a 5.2 ± 0.8 b 5.3 ± 0.7 b 7.0 ± 0.8 c 5.5 ± 0.6 b 7.0 ± 1.2 c
Year 2002 43.2 ± 4.8 f 18.5 ± 6.5 a 36.3 ± 7.8 e 26.6 ± 6.6 c 41.0 ± 4.7 f 33.5 ± 4.6 d 23.4 ± 12.9 b 21.5 ± 5.4 b
soluble solids (°Brix)
titratable acidity (meq/L)
pH
10.4 ± 0.1 a 13.4 ± 0.1 c 14.7 ± 0.2 d 15.1 ± 0.1 e 12.5 ± 0.0 b 18.6 ± 0.1 g 15.2 ± 0.2 e 16.5 ± 0.1 f
79.3 ± 7.6 b 70.3 ± 13.9 ab 89.3 ± 1.2 c 68.7 ± 1.2 a 112.3 ± 0.6 e 99.3 ± 1.2 d 133.7 ± 1.5 f 143.3 ± 2.1 f
3.70 ± 0.02 b 3.74 ± 0.01 c 4.21 ± 0.02 f 4.27 ± 0.01 g 3.82 ± 0.02 d 4.16 ± 0.00 e 3.64 ± 0.01 a 3.66 ± 0.01 a
11.9 ± 1.4 a 16.3 ± 2.1 cd 15.8 ± 1.1 c 17.6 ± 0.9 e 13.6 ± 1.4 b 16.7 ± 2.1 cde 17.3 ± 1.3 de 19.2 ± 2.0 f
58.7 ± 1.6 b 67.2 ± 1.7 c 76.2 ± 0.6 d 81.4 ± 1.8 ef 74.3 ± 2.3 d 79.9 ± 0.3 e 48.7 ± 2.2 a 83.6 ± 1.4 f
3.92 ± 0.01 d 3.91 ± 0.04 cd 3.95 ± 0.03 d 3.84 ± 0.02 ab 3.87 ± 0.02 bc 3.87 ± 0.02 bc 4.16 ± 0.04 e 3.80 ± 0.02 a
Means of each year followed by the same letter are not significantly different at P < 0.05 (Duncan’s test). Mean values ± SD (n ) 20) for the year 2001 and 2002.
stages, (2) evaluate the effects of postharvest storage on the phenolic composition of sweet cherries, and (3) assess the variations caused by natural harvest fluctuations in different harvest years. MATERIALS AND METHODS Sample Preparation. Sweet cherries from the cultivars Burlat, Saco, Summit, and Van, grown in Vila Real, Portugal, were randomly harvested by hand in 2001 and 2002, at two different stages of ripeness: partially ripe and ripe. Skin color is the main criterion used for indicating maturity for cherry picking. For each cultivar the maturity was assessed for 20 fruits by the following indices: weight, skin color (by a Minolta colorimeter), soluble solids content (°Brix by a refractometer), titratable acidity (by an automatic titration system), and pH (by a pH-meter). The ranges of these indices for each cultivar are shown in Table 1. The phenolic analyses were made on days 0, 5, 10, 15, 20, 25, and 30 in the fruits under storage at cold treatment [1-2 °C and 90% relative humidity (RH)] and on days 0, 3, and 6 for fruits subjected to room temperature treatment (15 ( 5 °C). Cherries were cut in half (the stone was taken off), and the cherry halves were frozen in liquid nitrogen, crushed, and freeze-dried prior to analysis. Extraction of Phenolic Compounds. Pitted and freeze-dried cherry samples (0.5 g) were mixed in 5 mL of 60% MeOH, flushed with N2, and extracted during shaking using a thermostated (25 ( 1 °C) water bath, at 200 rpm for 10 min. The suspension was initially filtered under vacuum through one layer of Whatman no. 1 filter paper. The extract was then filtered through a 0.45 µm hydrophilic Durapore filter (Millipore Corp., Bedford, MA), flushed with N2, and the filtrate was injected into the HPLC, after a period not exceeding 24 h, for the separation and quantification of phenolic compounds. The cherry samples were submitted to a second and a third extraction and analyzed by HPLC, and the total content was determined as the sum of the three values. HPLC-DAD Analyses. Samples of 10 µL of extracts were analyzed using an HPLC system equipped with a diode array detector (DAD) (Hewlett-Packard 1100 system, Waldbronn, Germany) operated by HP ChemStation software with a Nova-Pak C18 column (3.9 × 150 mm, Waters) at 40 °C. The mobile phase was made of three solvents delivered in a gradient system at a flow rate of 0.5 mL/min essentially as described by Lamuela-Ravento´s and Waterhouse (15). Identification and Quantification of Phenolic Compounds. The phenolic compounds in cherry extracts were identified by their spectral and retention time characteristics, recorded with a diode array detector,
and, wherever possible, by spectral chromatographic comparisons with authentic markers (16). The quantities of the different phenolic compounds were assessed from peak areas and calculated as equivalents of seven representative standard compounds (from standard, linear regression curves of authentic standards) as follows: at 280 nm (flavan-3-ols), catechin and epicatechin, respectively; at 316 nm (hydroxycinnamates), neochlorogenic acid, chlorogenic acid, and other hydroxycinnamaic acids as chlorogenic acid equivalents and p-coumaroylquinic acid as p-coumaric acid equivalents; at 365 nm (flavonols), quercetin glucosides as rutin equivalents; at 520 nm (anthocyanins), cyanidin 3-glucoside and cyanidin 3-rutinoside, respectively, and other anthocyanins as cyanidin 3-rutinoside. Coefficients of variation on the HPLC quantifications were
