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Food Chemistry 142 (2014) 392399

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Food Chemistry
journal homepage: www.elsevier.com/locate/foodchem

Monitoring of phytochemicals in fresh and fresh-cut vegetables: A comparison


Mara Isabel Alarcn-Flores, Roberto Romero-Gonzlez, Jos Luis Martnez Vidal, Francisco Javier Egea Gonzlez, Antonia Garrido Frenich
Group Analytical Chemistry of Contaminants, Department of Chemistry and Physics, Research Centre for Agricultural and Food Biotechnology (BITAL), University of Almera, Agrifood Campus of International Excellence, ceiA3, E-04120 Almera, Spain

a r t i c l e

i n f o

a b s t r a c t
Bearing in mind that fresh-cut market is currently the fastest growing subsector in the food industry, a comparison of the levels of phytochemicals in fresh and fresh-cut vegetables has been carried out. Thus, several families of phytochemicals, such as phenolic acids, isoavones, avones, avonols and glucosinolates were determined in fresh and fresh-cut samples including tomato, carrot, grape, eggplant and broccoli. Both type of products have potential and similar benecial properties, regarding its content as phytochemicals, except tomato, which should be consumed as fresh. Other factors such as commercial presentation (sliced, grated, diced) and storage conditions (temperature and light) were evaluated observing that in eggplant, the content of phenolic acids is statistically different depending on the presentation. On the other hand, the content of phytochemicals was higher when fresh-cut carrots were stored at 4 C regardless of the presence or absence of light. Multivariate analysis, based on cluster analysis was used as a rst approach to distinguish between fresh and fresh-cut samples, obtaining good results except for eggplant and carrot. 2013 Elsevier Ltd. All rights reserved.

Article history: Received 9 January 2013 Received in revised form 23 May 2013 Accepted 16 July 2013 Available online 24 July 2013 Keywords: Fresh-cut vegetables Fresh vegetables Phytochemicals Storage conditions Food composition UHPLCMSMS

1. Introduction It is well known that fruit and vegetables are important components of a healthy diet, and their daily consumption could help to prevent major diseases, such as cardiovascular diseases (Bhupathiraju & Tucker, 2011; Ness et al., 2005) and certain cancers (Soerjomataram et al., 2010; Wicki & Hagmann, 2011). These benecial effects of fruits and vegetables have been attributed to non-essential food constituents, which are known as phytochemicals or bioactive compounds, that possess a relevant bioactivity when they are frequently consumed as a part of a regular diet (Mudgal, Madaan, Mudgal, & Mishra, 2010). In general, these compounds could possess antioxidant capacity (AOC) (Kim, PadillaZakour, & Grifths, 2004), antiinammatory (Gonzlez-Gallego, Garca-Mediavilla, Snchez-Campos, & Tun, 2010; Vincent, Bourguignon, & Taylor, 2010), lipid prole modication (Perez-Vizcaino & Duarte, 2010; Wang, Melnyk, Tsao, & Marcone, 2011) and antitumor effects (Collins, 2005; Pietta, Minoggio, & Bramati, 2003; Stan, Kar, Stoner, & Singh, 2008). Nowadays, there is an increased interest in health and consumers are concerned with the role of food for maintaining and improving human well-being. In this sense, the practical
Corresponding author. Tel.: +34 950015985; fax: +34 950015008.
E-mail address: agarrido@ual.es (A. Garrido Frenich). 0308-8146/$ - see front matter 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.foodchem.2013.07.065

advantages and convenience that the fresh-cut products provide to the consumers makes this market currently the fastest growing subsector in the food industry, and it still has a high potential of growth worldwide (Ragaert, Verbeke, Devlieghere, & Debevere, 2004). Currently, minimal processing is applied to fruit and vegetables, involving at industrial scale initial rinsing, peeling, slicing, washing, packaging and storage (Laurila & Ahvenainen, 2000). Therefore, it is important to investigate if this minimal processing may result in loss of important phytochemicals. There are several studies that evaluate the effects of minimally processing on phenolic acids, vitamin C and carotenoids in fresh-cut carrot (Alasalvar, Al-Farsi, Quantick, Shahidi, & Wiktorowicz, 2005; Simes, Allende, Tudela, Puschmann, & Gil, 2011; Simes, Tudela, Allende, Puschmann, & Gil, 2009) or lettuce (Martnez-Snchez et al., 2012; Selma et al., 2012), but there are few studies regarding avonoids (Gil, Ferreres, & Toms-Barbern, 1998; Selma et al., 2012) or glucosinolates (Jones, Faragher, & Winkler, 2006) in other fresh-cut vegetables or fruits such as tomato (Odriozola-Serrano, Soliva-Fortuny, & Martn-Belloso, 2008), eggplant (Mishra, Gautam, & Sharma, 2012), grape (Costa et al., 2011) and broccoli (Jones et al., 2006). In general, it was observed that the content of phenolic acids increased in fresh-cut products. This fact could be explained considering that when a sharp blade was used for cutting, it produced a lower release of phenolic acids and lowered the polyphenol

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oxidase (PPO) activity compared to when a knife was used. This cutting process is known to be a key player in the browning process of various raw and cut fruit and vegetables (Mayer, 2006; Mishra et al., 2012). Furthermore, the abiotic stress (Cisneros-Zevallos, 2003), such as altered O2 and CO2 levels in controlled and modied atmospheres packaging (MAP), or C2H4 gassing for ripening and degreening, could also affect phytochemical accumulation. For example, in the phenylpropanoid pathway, the phenylalanine ammonialyase (PAL) generates an increase in phenolic compounds in certain fresh-cut products (Kenny & OBeirne, 2010). Moreover, in some cases, natural additives, such as ascorbic acid or citric acid (Robles-Snchez, Rojas-Grab, Odriozola-Serrano, Gonzlez-Aguilar, & Martn-Belloso, 2009; Son, Moon, & Lee, 2001) were added to enhance the shelf life of the fresh-cut product, and therefore it is possible that the phytochemicals content remained constant or even increased. In contrast to fresh products, the shelf life of fresh-cut products usually extend upto 16 days at 4 C, 12 days at 10 C and 5 days at 26 C storage temperatures, although this depends on the type of matrix (Mishra et al., 2012). In relation to the evolution of phytochemical in fresh-cut products, some studies reported that total phenolic indices signicantly decreased after 5 days of storage at 24 C in apple (Roossle, Wijngaard, Gormley, Butler, & Brunton, 2010), although some studies reported by Simes et al. (2011), indicate that the phenolic compounds, mainly chlorogenic acid, increased twofold in baby carrots stored under MAP, where the concentration of the gases was used as an important parameter. Furthermore, Odriozola-Serrano et al. (2008), reported that a minimal processing maintains the main antioxidant compounds of sliced tomatoes for 21 days at 4 C, preserving their initial nutritional value. Finally, Cisneros-Zevallos (2003) reported that the total phenolic content and the antioxidant capacity of the tissue increased in sliced orange carrots and purple potatoes stored during 2 days at 20 C. Considering that current approaches have been based on the evaluation of a few family of compounds in some matrices, the purpose of this work was the comparison of the content of several families of phytochemicals (phenolic acids, avonols, avones, glucosinolates and isoavones) in fresh and fresh-cut products stored under MAP, such as tomato, eggplant, grape, carrot and broccoli, in order to get a comprehensive view of the presence of phytochemicals in fresh and fresh-cut products. Other variables such as type of cut and shelf life were also evaluated.

Stock standard solutions of individual compounds (with concentrations between 200 and 300 mg/l) were prepared by exact weighing of the powder and dissolved in 10 ml of HPLC grade methanol or in a mixture of methanol:water (50:50, v/v). Then they were stored at 20 C in dark bottles. A multicompound working standard solution at a concentration of 5 mg/l of each compound was prepared by appropriate dilutions of the stock solutions with methanol and stored in screw-capped glass tubes at 20 C. The solutions were prepared each 6 months. Ultrapure water was obtained from a Milli-Q Gradient water system (Millipore, Bedford, MA, USA). Ammonium acetate was purchased from Panreac (Barcelona, Spain). Formic acid (purity >98%), HPLC-grade methanol was provided by Sigma (Madrid, Spain). Millex-GN nylon lters of 0.20-lm were provided by Millipore (Millipore, Carrightwohill, Ireland). 2.2. Apparatus and software Chromatographic analyses were carried out using an Agilent series 1290 RRLC instrument (Agilent, Santa Clara, CA, USA) equipped with a high-performance autosampler (G4226A), a binary pump (G4220A), a column compartment thermostat (G1316C) and an autosampler thermostat (G1330B). The system was coupled to an Agilent triple quadrupole mass spectrometer (6460A) with a Jet Stream ESI ion source (G1958-65138). For the chromatographic separation of the extracts, a Zorbax Eclipse Plus C18 column (100 mm 2.1 mm, 1.8 lm particle size) from Agilent was used. The injection volume was 5 ll and column temperature was set at 30 C. Chromatographic separation was carried out using a gradient elution with methanol as eluent A, and an aqueous solution of ammonium acetate (30 mM), adjusted to pH 5 with formic acid, was used as eluent B. The elution started at 5% of eluent A for 1.5 min, and then it was increased to 30% in 2.5 min. After that, it was increased to 100% at 4 min. This composition was kept constant during 2 min, before being returned to the initial conditions after 0.5 min, keeping this composition during 1.5 min prior to the next analysis, obtaining a total run time of 12 min. The ow rate was set at 0.2 ml/min. The Jet Stream ion source parameters were: drying gas temperature and sheath gas temperature at 325 C and 400 C respectively; drying gas ow and sheath gas ow at 7 and 12 ml/min respectively; nebulizer pressure at 40 psi; capillary voltage was set at 4000 and 3500 V in positive and negative acquisition respectively. An Agilent Mass Hunter Quantitative analysis (Agilent Technologies, Inc.) was used for data acquisition and quantication of samples. Statistical analysis, analysis of variance and cluster analysis were carried out with JMP v9 (Cary, NC, USA). Lyophiliser Alpha from Martin Christ (Osterode, Germany) was also used; an analytical balance AB204-S from Mettler Toledo (Greifensee, Switzerland), a Reax-2 rotary agitator from Heidolph (Schwabach, Germany), and vacuum pump from Vacuubrand (Wertheim, Germany) were also utilised. 2.3. Extraction procedure Samples were homogenised and they were transferred to a Petri dish, and weighed and cooled to 18 C. Then, all samples were processed according to the following procedure: 150 mg of lyophilized sample was weighed in a 15 ml polypropylene centrifuge tube and 3 ml of a mixture of methanol:water (80:20, v/v) was added. The mixture was agitated for 30 min with a rotary shaker. After that, the extract was ltered and 100 ll were transferred into a vial containing 400 ll of a mobile phase (50:50 v/v of eluent A and B), and 5 ll were injected into the chromatographic system

2. Materials and methods 2.1. Chemicals and reagents Commercial phenolic compound standards such as progoitrin, gluconasturtin and glucoraphanin were supplied by PhytoLab GmbH & Co (Vestenbergsgreuth, Germany). Glucotropaeolin, glucoerucin and glucoiberin were purchased from Scharlab (Barcelona, Spain). Other standards as genistein, apigenin, quercetin, quercetin-3-O-glucoside, gallic acid, sulforaphane, ferulic acid, baicalein, gallic acid and caffeic acid were purchased from SigmaAldrich (Steinheim, Germany). Other standards as daidzein, glycitein, luteolin-4-O-glucoside, luteolin-7-O-glucoside, apigenin-7-O-neohesperoside, kaempferol, kaempferol-3-O-glucoside, kaempferol-3-O-rutinoside, luteolin, luteolin-6-C-glucoside, luteolin-8-C-glucoside, apigenin-7-O-glucoside, apigenin-6-C-glucoside, apigenin-8-C-glucoside, quercetin-3-O-ramnoside, quercetin-3-Ogalactoside, quercetin-3-O-rutinoside, quercetin-3-O-ramnoside, quercetin-3-O-galactoside, isorhamnetin, isorhamnetin-3-O-rutinoside, isorhamnetin-3-O-glucoside, apigenin-7-O-rutinoside and tamarixetin were purchased from Extrasynthese (Genay, France).

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(Alarcn Flores, Romero-Gonzlez, Garrido Frenich, & Martnez Vidal, 2013). 2.4. Samples Fresh tomato, broccoli, carrot, grape and eggplant samples were obtained from different supermarkets located in the province of Almera (southeast of Spain). Fresh-cut products were obtained from Guzmn Gastronoma (Barcelona, Spain). In both cases, six samples of each matrix were analysed. However for fresh-cut carrot and fresh-cut eggplant, twelve samples per matrix were analysed considering that two types of cuts were evaluated for each matrix (grated and sliced for carrots, and diced and sliced for eggplant). Samples were chopped and homogenised at room temperature. After that, they were kept at 18 C until lyophilisation. 3. Results and discussion 3.1. Comparison of the content of phytochemicals in fresh and freshcut matrices First, the content of phytochemicals was determined in the fresh and fresh-cut matrices included in this survey, using a method developed previously by our research group (Alarcn Flores et al., 2013). Tables 1 and 2 show the obtained results, expressed as mg of phytochemical per kg of dry weight (DW). It can be observed that the highest concentration of phytochemicals was detected in broccoli (4517.4 and 3369.7 mg/kg DW in fresh and fresh-cut products respectively, whereas tomato (174.5 and 96.8 mg/kg DW), carrot (365.8 and 261.9 mg/kg DW) and grape (266.3 and 200.7 mg/kg DW), had the lowest concentrations; nally, the level of phytochemicals in eggplant (782.6 and 1520.8 mg/ kg DW) can be considered as an intermediate, regardless the type of matrix (fresh or fresh-cut). Fig. 1 shows the chromatograms of

quercetin-3-O-rutinoside at 26 mg/kg in fresh-cut tomato sample, sulforaphane at 15 mg/kg in fresh broccoli sample, quercetin-Oderivate at 172 mg/kg in fresh grape samples, chlorogenic acid at 694 mg/kg in fresh eggplant samples and ferulic acid at 2 mg/kg in fresh-cut carrot samples. Considering the different families, it can be observed that phenolic acids are the most abundant compounds in tomato, although a signicant difference between their levels in fresh (102.3 mg/kg) and fresh-cut tomato (54.3 mg/kg), were obtained when ANOVA was carried out (p = 0.02), observing that the highest levels were obtained in fresh tomato (Fig. 2). For the rest of families of phytochemicals detected in tomato, avones and avonols, there was no signicant difference between fresh and fresh-cut products, although for avonols, the p-value obtained was lower than 0.1. As indicated before, broccoli contains the highest levels of phytochemicals due to the presence of glucosinolates (Fig. 2), and the total content of these compounds in fresh and fresh-cut samples ranges from 4517.4 mg/kg to 3369.7 mg/kg, respectively. This decrease can be attributed to the diminution of glucoraphanin concentration in fresh-cut samples, although this difference was not signicant. However, it was observed that the level of avonols in fresh-cut broccoli was signicantly lower (p = 0.01) compared with fresh samples. In relation to grapes, avonols were the compounds detected at highest concentration, which remains quite stable from fresh to fresh-cut, except for quercetin-3-O-derivate that showed a nonsignicant decrease. As previously, the content of avonols was lower in fresh-cut products (150 mg/kg) than in fresh (230 mg/ kg), although in this case, this difference was not signicant (p = 0.32). Phenolic acids were the predominant phytochemicals detected at higher concentration in carrots, The highest concentration was quantied in fresh samples (340 mg/kg), whereas the concentration in fresh-cut samples was 237 mg/kg. Lower concentrations were detected for avones and avonols. In this matrix, there

Table 1 Content of phytochemicals (as mg/kg of DW) in tomato, broccoli and grape. Tomato Fresha Apigenin Baicalein Luteolin Luteolin C-glucoside Luteolin O-glucoside Caffeic acid Chlorogenic acid Ferulic acid Gallic acid Glycitein Glucoerucin Glucoiberin Gluconasturtin Glucoraphanin Sinigrin Sulforaphane Isorhamnetin Isorhamnetin 3-O-glucoside Kaempferol Kaempferol 3-O-glucoside Kaempferol 3-O-rutinoside Quercetin Quercetin O-derivate Quercetin 3-O-rutinoside Quercitrin Tamarixetin Total content
a

Broccoli Fresh-cuta 1.6 (1.1) 11.9 (3.7) 4.8 (1.4) 2.2 (1.5) 0.7 (0.4) 16.5 (4.7) 25.4 (22.5) 12.4 (2.6) Fresha 3.3 4.0 5.9 2.3 1.0 (0.6) (2.6) (1.9) (0.3) (0.8) Fresh-cuta 1.9 (1.3) 13.6 (4.7) 3.1 (1.0) 3.5 (4.7) 1.5 (0.5) 28.5 (8.0) 1.3 (0.8)

Grape Fresha 3.2 4.3 9.3 4.1 1.5 (0.2) (4.1) (2.0) (3.7) (0.2) Fresh-cuta 6.1 (4.9) 28.2 (16.7) 11.4 (4.4) 0.9 (1.6) 2.0 (1.3)

2.3 (1.5) 15.1 (2.9) 7.0 (3.1) 2.3 (1.5) 1.9 (1.5) 22.2 (25.4) 51.6 (16.0) 28.2 (18.2)

36.4 (10.3) 1.9 (0.9)

0.8 (0.9) 2.6 (1.8) 33.4 (15.6) 629.9 (36) 33.2 (4.7) 3737.8 (27) 2.3 (1.0) 13.1 (2.7) 6.4 (1.4) 3.9 (0.8) 40.5 (10.5) 734.5 (52.0) 38.7 (7.7) 2473.8 (100.0) 1.7 (1.2) 20.4 (6.0) 3.3 (0.8) 3.2 (2.0) 14.7 (13.7) 4.9 (2.5) 34.8 (16.6) 3.7 (1.2) 13.4 (11.5) 2.6 (1.1) 0.25 (0.5) 5.6 (3.2) 165.3 (128.0) 10.3 (4.2) 2.7 (1.2) 15.7 (6.7) 257.4 (153.5)

3.4 (3.5)

4.2 (3.0)

4.2 (2.0)

9.5 (2.2) 7.8 (5.1)

10.0 (2.6) 7.6 (8.6) 10.3 (7.5) 7.2 (1.8) 85.4 (59.0) 11.7 (11.5) 3.3 (1.9) 13.7 (7.4) 201.2 (66.9)

174.5 (42.2)

96.8 (31.5)

4517.4 (20.7)

3369.7 (76.3)

Mean value. Standard deviation is given in parenthesis (n = 6).

M.I. Alarcn-Flores et al. / Food Chemistry 142 (2014) 392399 Table 2 Content of phytochemicals (as mg/kg of DW) in carrot and eggplant. Carrot Fresh
a

395

Eggplant Fresh-cut Grated


a

Fresha Sliced 4.1 (1.3) 12.4 (1.8) 4.9 (1.7) 2.0 (2.3) 2.6 (2.5) 2.0 (1.6) 180.4 (77.5) 1.1 (1.2) 2.6 (0.8) 5.0 (5.0) 6.2 (1.1) 3.3 (0.9) 2.5 (0.8) 2.9 (2.5) 730.1 (848.0) 0.6 (0.4)

Fresh-cuta Diced 4.2 (2.1) 15.2 (4.2) 8.0 (3.6) 2.9 (1.9) 2.1 (1.7) 6.9 (2.0) 1827.2 (613.3) 3.1 (2.9) Sliced 4.3 (2.9) 12.7 (3.3) 6.9 (1.9) 2.3 (1.6) 2.0 (1.4) 6.5 (1.7) 1104.1 (496.0) 1.5 (1.1)

Apigenin Baicalein Luteolin Luteolin C-glucoside Luteolin O-glucoside Caffeic acid Chlorogenic acid Ferulic acid Gallic acid Glycitein Glucoerucin Glucoiberin Gluconasturtin Glucoraphanin Sinigrin Sulforaphane Isorhamnetin Isorhamnetin 3-O-glucoside Kaempferol Kaempferol 3-O-glucoside Kaempferol 3-O-rutinoside Quercetin Quercetin O-derivate Quercetin 3 O-rutinoside Quercitrin Tamarixetin Total content
a

2.0 (0.4) 3.3 (2.8) 5.9 (1.8) 2.2 (0.7) 2.6 (2.1) 1.1 (0.7) 337.8 (155.5) 1.2 (0.5)

3.7 (1.7) 10.4 (3.3) 6.4 (1.9) 1.7 (1.4) 1.7 (1.9) 1.3 (0.7) 270.6 (129.1) 3.7 (5.0)

5.3 (1.2)

5.6 (3.1)

6.5 (1.1)

5.4 (1.5)

4.8 (0.4)

5.3 (1.7)

4.5 (0.5)

2.3 (2.0)

1.0 (1.5)

5.8 (0.4) 13.4 (9.5) 4.8 (2.7)

1.6 (1.4) 3.5 (0.9) 3.7 (2.1)

365.8 (136.6)

307.4 (133.4)

217.0 (78.3)

782.6 (602.9)

1877.9 (617.6)

1150.9 (494.6)

Mean value. Standard deviation is given in parenthesis (n = 6).

Fig. 1. UHPLCMS/MS chromatograms for different compounds in different matrices: (a) quercetin-3-O-rutinoside at 26 mg/kg in fresh-cut tomato sample; (b) sulforaphane at 15 mg/kg in fresh broccoli sample; (c) quercetin-O-derivate at 172 mg/kg in fresh grape samples; (d) chlorogenic acid at 694 mg/kg in fresh eggplant samples, and (e) ferulic acid at 2 mg/kg in fresh-cut carrot samples.

was no signicant difference between the content of the different families of phytochemicals in fresh and fresh-cut products. Finally, phenolic acids were also the most abundant phytochemicals in eggplant, basically due to the high concentration of chlorogenic acid. In this case, it was observed that the content of

phenolic acid was lower in fresh samples than in fresh-cut samples. This can be explained considering that fresh-cut eggplant contains ascorbic acid as a natural additive, and PPO activity might explain the signicant increase in fresh-cut eggplant; this last effect has been reported elsewhere in apples (Chung & Moon,

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Fig. 2. Content of the families of phytochemicals evaluated in: (a) tomato; (b) broccoli; (c) grape; (d) eggplant and (e) carrot. p < 0.10. Values are means standard deviation (n = 6).

Signicantly different (p < 0.05).

2009) and mango fruits (Robles-Snchez et al., 2009). On the other hand, it was also observed that the content of avonols was significant higher in fresh samples (p = 0.01), indicating that this family of phytochemicals is most sensitive to processing. 3.2. Inuence of the type of presentation in the content of phytochemicals in fresh-cut product In a second step, the inuence of the type of presentation on the concentration of phytochemicals in fresh-cut carrot and eggplant was evaluated. Thus different types of cuts were studied. Sliced and grated carrots were investigated, whereas for eggplant, sliced and grated products were selected, the obtained results for both matrices are shown in Fig. 3. Furthermore, ANOVA study was carried out and it can be noted that there is no statistical difference between the content of phytochemicals between the different types of cut either in carrot or in eggplant, except for the level of phenolic acids in eggplant. For this matrix, it was observed that the higher results were obtained for diced products, and a mean value of 1877.9 mg/kg was obtained for this family, whereas for sliced products, a mean value of

1150.9 mg/kg was achieved (p = 0.02). This difference is mainly due to the content of chlorogenic acid in both types of presentation (Table 2). A similar situation was observed for carrots, where the lowest values of phenolic acids were obtained in sliced products, whereas for grated carrots, the level of these compounds was found to be higher, although the difference was not statistically signicant (p = 0.19). 3.3. Evolution of the content of phytochemicals in fresh-cut carrot In order to check whether the content of phytochemicals remains constant during the whole shelf-life, fresh-cut carrots stored at different conditions were analysed. Thus, freshly-cut packed carrots were stored at 4 C and 25 C during the life-time of the product in the dark (the lyophilisation and extraction procedure were done in the absence of light) and in the presence of the light. Therefore, and just after the expiration date, the content of phytochemicals was determined. The obtained results are shown in Table 3. ANOVA study was carried out to check if there is a statistical difference between the content of phytochemicals in the assayed product. It was observed that the life-time of the product

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Fig. 3. Effect of the type of cut in the content of phytochemicals in fresh-cut: (a) carrot and (b) eggplant. deviation (n = 6).

Signicantly different (p < 0.05). Values are means standard

Table 3 Content of phytochemicals (mg/kg of DW) in fresh-cut carrots stored at different conditions.a Family Compound Fresh-cut product L4 Flavones Apigenin Baicalein Luteolin Luteolin C glucoside Luteolin O glucoside Caffeic acid Chlorogenic acid Ferulic acid Isorhamnetin Quercetin 1.9 10.4 6.3 2.4 0.5 0.9 72.7 1.0 6.0 3.0 105.1 D4 3.2 16.7 22.2 1.2 0.2 0.6 69.7 2.2 18.6 7.0 141.6 Expired product L4 1.3 9.6 6.0 0.5 1.4 1.3 246.0 1.0 5.4 4.3 276.8 L25 4.1 10.4 3.2 0.5 1.4 0.1 94.3 0.5 3.6 5.4 123.4 D4 2.0 9.9 7.1 4.9 4.6 4.7 204.6 1.5 9.0 5.4 253.8 D25 1.8 26.8 6.8 3.9 4.1 0.2 104.6 1.1 12.5 7.3 169.0

Phenolic acids

Flavonols

Total content of phytochemicals


a

Sample code: L: storage in light conditions; D: storage in dark conditions; 4: storage at 4 C; 25: storage at 25 C.

as well as the storage temperature has a signicant inuence in the content of total phytochemicals (p values of 0.03 and 0.04 respectively). Thus, it can be observed that the highest content of phytochemicals was obtained just after the expiration date. Furthermore, the storage at a lower temperature (4 C) also provides the highest levels of the assayed compounds. In this study, the presence or absence of the light did not have a signicant inuence on the content of phytochemicals. The reason of the above differences lays in the content of chlorogenic acid, which is the compound with the highest concentration at the end of the storage period kept at 4 C. This could be explained according to the results obtained by Cisneros-Zevallos (2003), who reported and proposed that the use of different abiotic stresses, such as cold storage and stored atmospheric conditions enhances the phytochemicals levels in fresh fruits and vegetables. 3.4. Multivariate analysis: Cluster analysis In order to verify if it is possible to distinguish between fresh and fresh-cut products on the basis of their respective phytochemicals content, a multivariate analysis based on cluster analysis was carried out, considering that it provides useful information related to natural grouping among samples. A hierarchical agglomerative cluster analysis of samples was performed using the concentrations of the phytochemicals detected as variables. The squared Euclidean was used as the similarity measurement and Wards method was selected as the amalgamation rule. Fig. 4 shows the resulting dendogram when the natural grouping of samples was

evaluated. It can be observed that three groups can be distinguished, according to the concentration of the phytochemicals evaluated in this study. Thus, the rst group is formed by tomatoes, carrots and eggplants; although they belong to different families of vegetables, they basically contain phenolic acids as the major phytochemical. Furthermore, it can be observed that tomato differs from the other two matrices, whereas it is difcult to discriminate between carrot and eggplant. Then, the second group is formed by grapes, which basically contains avonols, whereas the third group is composed by broccoli, where a high level of glucosinolates was detected. Furthermore, it can be observed that for some matrices such as tomato, grape and broccoli, it is possible to distinguish between fresh and fresh-cut products according to their phytochemical content, whereas for eggplant and carrot it is more difcult to nd a common pattern between fresh and fresh-cut products, although more data is necessary to get denitive conclusions. Therefore, this study can be considered as a rst approach to distinguish between fresh and fresh-cut products based on their phytochemicals content.

4. Conclusions According to the data obtained in this work, it was found that in both fresh and fresh-cut vegetables, phenolic acids were the most dominant group in tomato, carrot and eggplant, whereas glucosinolates were the most abundant in broccoli. Flavonols were found to

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between fresh and fresh-cut products, and the content was statistically different in broccoli and eggplant. Regarding the presentation of fresh-cut products, it was observed that phenolic acids were the phytochemicals most affected by this factor, and the lowest content was also found in slice presentation, whereas diced or grated products presented a higher concentration of these compounds. Furthermore, it was observed that the content of phytochemicals can increase during the storage of these products at 4 C, leading to an increase in concentration at lower temperatures. Finally, a preliminary classication based on the content of phytochemicals could be used to distinguish between fresh and freshcut products, although more analyses are necessary. In conclusion, fresh and fresh-cut products will have similar benecial properties with respect to their content in phytochemicals, except for tomato, which should be consumed as fresh product. Acknowledgements The authors are grateful to Andalusian Regional Government (Regional Ministry of Innovation, Science and Enterprise) and the Centre for Industrial Technological Development (CDTI) and FEDER for nancial support Project Ref. P11-AGR-7034 and IDI-20110017 respectively. The authors also are grateful to Guzmn Gastronoma for supply of the fresh-cut samples. M.I.A.F. acknowledges her grant (FPU, Ref.: AP 2009-2074) from the Spanish Ministry of Education. RRG is also grateful for personal funding through Ramon y Cajal Program (Spanish Ministry of Economy and CompetitivenessEuropean Social Fund). References
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Fig. 4. Dendogram constructed with Wards method showing the results of cluster analysis. Sample code: First letter B: Broccoli; C: Carrot; E: Eggplant; G: Grape; T: Tomato; Second letter C: Fresh-cut product; F: Fresh product.

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