Article pubs.acs.org/JAFC
Origin of the Pinking Phenomenon of White Wines Jenny Andrea-Silva,†,‡ Fernanda Cosme,‡ Luís Filipe Ribeiro,‡ Ana S. P. Moreira,§ Aureliano C. Malheiro,# Manuel A. Coimbra,§ M. Rosário M. Domingues,§ and Fernando M. Nunes*,† †
CQ-VR, Chemistry Research Centre, Chemistry Department, Universidade de Trás-os-Montes e Alto Douro, 5001-801 Vila Real, Portugal ‡ Institute for Biotechnology and Bioengineering, Centre of Genomics and Biotechnology, (IBB/CGB-UTAD), Edifício de Enologia, Universidade de Trás-os-Montes e Alto Douro, Apartado 1013, 5001-801 Vila Real, Portugal § Mass Spectrometry Centre, QOPNA, Chemistry Department, Universidade de Aveiro, 3810-193 Aveiro, Portugal # Centre for the Research and Technology of Agro-Environmental and Biological Sciences (CITAB), Universidade de Trás-os-Montes e Alto Douro, 5001-801 Vila Real, Portugal S Supporting Information *
ABSTRACT: Pinking is the terminology used for the salmon-red blush color that may appear in white wines produced exclusively from white grape varieties. The isolation of pinking compounds and their analysis by RP-HPLC-DAD and ESI-MSn showed that the origin of the pinking phenomenon in white wines from Vitis vinifera L. of Sı ́ria grape variety are the anthocyanins, mainly malvidin-3-O-glucoside. The analysis showed that the anthocyanins were located both in the pulp and in the skin. Wine pinking severity was negatively related with the increase of the average temperature of the first 10 days of October, the final period of grape maturation. The minimum amount of anthocyanins needed for the pink color visualization in wine was 0.3 mg/L. The appearance of pinking in white wines after bottling is due to the lowering of free sulfur dioxide, which leads to an increase of the relative amount of the anthocyanins red flavylium form and their polymerization, resulting in the formation of colored compounds resistant to pH changes and sulfur dioxide bleaching. KEYWORDS: Vitis vinifera, white grapes, Sı ́ria variety, white wines, pinking, anthocyanins
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materials.2 Also, it was reported that a 2-S-glutathionyl-caftaric acid derivative may be the pink chromophore.4 There are various preventive or curative enological treatments for the pinking problem, including adding polyvinylpolypyrrolidone (PVPP) or PVPP associated with bentonite or increasing the redox potential using ascorbic acid in the prebottling stage.10 Nevertheless, the use of these treatments increases wine production costs and can change wine sensory properties.13 Also, the use of high levels of ascorbic acid has been shown to increase the browning potential of white wines.14 In white wine production, hyperoxidation of juice is not recommended, as it alters significantly the white wine flavor, decreasing the fruity aroma intensity and consequently the wine quality.4,8 The main purpose of this work was to get a deeper insight into the pinking phenomenon, its origin, and technological factors influencing its appearance in white wines. For this, Sı ́ria, a white grape variety, was used. Knowledge of the compounds responsible for this color alteration would increase the treatment efficiency of those wines by allowing the choice of the most correct treatment to optimize their removal.
INTRODUCTION Pinking is a term that describes the appearance of a salmon-red blush in white bottled wines produced exclusively from white grape varieties. It is perceived as an undesirable phenomenon by both wine consumers and the industry.1,2 Although with seasonal and regional variations, pinking has been observed worldwide, with predominance in white wines produced from Vitis vinifera L. grape varieties such as Chardonnay, Chenin blanc, Crouchen, Muscat Gordo blanco, Palomino, Riesling, Sauvignon blanc, Semillon, Sultana, and Thompson Seedless.1,3 Pinking is mainly observed when white wines are produced under reducing conditions.1−4 The pinking phenomenon is usually observed after bottling and storage of white wines or after alcoholic fermentation, although sometimes it occurs as soon as the grape must is extracted.5−7 Susceptibility of reductively produced wines to pinking may be effectively overcome by maintaining adequate levels of free sulfur dioxide, especially when ascorbic acid is used.8,9 Pinking is thought to be due to oxidative changes in white wines when exposed to oxygen,1,7 although the compounds responsible for the pinking phenomena are unknown.10 In addition, inhibition of polyphenol oxidase activity has been linked to pinking potential in white wines,11 and added hydrogen peroxide can induce it.1 It has been proposed that white wine pinking can be a result of the rapid conversion of accumulated flavenes to the corresponding red flavylium salts formed from the hydrolysis of leucoanthocyanins.12 However, this coloration may be caused by at least 10 different compounds and polymeric © 2014 American Chemical Society
Received: Revised: Accepted: Published: 5651
February 18, 2014 May 18, 2014 May 24, 2014 May 25, 2014 dx.doi.org/10.1021/jf500825h | J. Agric. Food Chem. 2014, 62, 5651−5659
Journal of Agricultural and Food Chemistry
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Article
Technologies, Scotland). Conditions of HPLC analysis were as follows: solvent A was a mixture of 95:5 water/formic acid (v/v), and solvent B was methanol. A linear gradient analysis for a total run time of 80 min was used as follows: starting from 5% solvent B during 2 min, increase to 80% solvent B over 68 min and then isocratic for 8 min, decrease to 5% solvent B over 2 min, and finally isocratic for 5 min. The sample volume injected was 100 μL, the flow rate was 1.0 mL/min, and the column temperature was maintained at 35 °C during the run. The eluent was continuously monitored from 250 to 600 nm with a photodiode array detector (PDA-100, Dionex). All analyses were performed in duplicate. Anthocyanin identifications were made by injection of pure standards in the case of cyanidin-3-O-glucoside, delphinidin-3-O-glucoside, and malvidin-3-O-glucoside and by comparison of their retention times and UV−vis spectra. Other anthocyanins were identified by comparison of the elution order and UV−vis spectra reported in the literature. Additionally, all anthocyanin-3-O-glucosides were confirmed by tandem mass spectrometry. Determination of Total Monomeric Anthocyanins. Total monomeric anthocyanins were determined using the pH differential method AOAC Official Method 2005.02 with a minor modification.16 Due to the low concentration of anthocyanins present in pinking white wines, for the anthocyanin determination by the pH differential method, the absorbance was measured at 520 and 700 nm using a 10 cm path length glass cell, and total monomeric anthocyanins were expressed as milligrams of malvidin-3-O-glucoside per liter (molar extinction coefficient of 28000 L/cm/mol and molecular weight of 493.4 g/mol). All analyses were performed in duplicate. Electrospray Ionization Mass Spectrometry (ESI-MS). ESI-MS and tandem ESI-MS (ESI-MSn) spectra of SPE-purified and concentrated must and wines were carried out on an LXQ linear ion trap mass spectrometer (Thermo Fisher Scientific Inc., Waltham, MA, USA). Typical operating conditions were as follows: electrospray voltage was 5 kV, capillary temperature was 275 °C, capillary voltage was 1 V, and tube lens voltage was 40 V. Samples were introduced at a flow rate of 8 μL/min into the ESI source. Nitrogen was used as nebulizing and drying gas. In the ESI-MSn experiments, the collision energy used was set between 18 and 31 (arbitrary units). Data acquisitions were carried out on an Xcalibur data system. For all ESIMS analyses, samples dissolved in water were diluted in methanol/ formic acid (99:1, v/v), and the spectra were acquired in the positive mode, scanning the mass range from m/z 100 to 1500. All analyses were performed in duplicate. Tissue Fractionation and Anthocyanin Content. For the analysis of anthocyanin distribution in Sı ́ria white grape variety, the skins were removed from the frozen grapes (hypodermal tissue samples) and the seeds were removed from the flesh (mesocarp tissue samples).17 The hypodermal and mesocarp samples were homogenized in a mortar and pestle at 4 °C in the presence of 0.1% HC1 (v/v) in methanol containing 50 ppm of SO2. The homogenate was centrifuged at 3400g for 6 min, and the supernatant was concentrated under vacuum. The residue was suspended in 2 mL of a water/ methanol solution (1:1 v/v) and analyzed for anthocyanin by HPLCDAD. All analyses were performed in duplicate. Climate Data. The climate data corresponds to the period from 2005 to 2011 of the vineyards located at Figueira de Castelo Rodrigo (40° 52′ N, 6° 54′ W, 635 m). The climate is of the Mediterranean type (warm temperate climate with dry and warm summer) characterized by a sinusoidal pattern of precipitation with dry summers and wet winters. In the experimental area, long-term annual precipitation is about 590 mm, with roughly a third falling from April to September. Corresponding annual average minimum and maximum temperatures are 6.4 and 18.3 °C.18 Weather variables kindly supplied by the Portuguese state agricultural services (Direçaõ Regional de Agricultura e Pescas do Centro) included daily records of maximum, minimum, and average temperatures and total precipitation, from which 10 day averages were computed. This set of data was recorded from a standard automatic weather station close to the experimental site. Additionally, thermal
MATERIAL AND METHODS
Chemicals. Oenin chloride, kuromanin chloride, and delphinidin 3-O-β-D-glucoside chloride were purchased from Sigma (USA). Grape and Wine Samples. The white grapes and white wines from V. vinifera L., Sı ́ria variety, used in this study were from the vintages 2010−2012, harvested at the designation of origin Beira Interior, subregion Castelo Rodrigo, in Portugal, and provided by the Producers Union “Adega Cooperativa de Figueira de Castelo Rodrigo” (ACFCR). This region is characterized by an altitude ranging between 400 and 700 m. The soils are mostly granite or schist. In this subregion the principal white grape varieties are Sı ́ria and Malvasia Fina. The white wines from this region have medium acidity (4−5 g/L tartaric acid), pH 3.2−3.4, light color, medium sensitivity to oxidation, intense aroma, and balanced notes of unripe tropical and citrus fruits. Winemaking was carried at the ACFCR, in Portugal. Grapes were manually harvested at optimum technological maturity (18−20 °Brix) and transported to the winery. They were immediately destemmed and crushed, and the grape juice was treated with sulfur dioxide (60 mg/L) to avoid must oxidation. Then, crushed grapes were pressed at a maximum pressure of 2 bar in a pneumatic press machine. Grape juice was clarified by decantation in a stainless steel tank during 24 h at 8 °C. The alcoholic fermentation was started using active dry yeasts, carried out at controlled temperature (14−15 °C), and finished when residual sugars were
