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Postharvest Biology and Technology 61 (2011) 137–144

Contents lists available at ScienceDirect

Postharvest Biology and Technology

journa l homepage: www.e lsev ier .com/ locate /postharvbio

hanges in the microstructure and location of some bioactive compounds inersimmons treated by high hydrostatic pressure

.L. Vázquez-Gutiérrez, A. Quiles ∗, I. Hernando, I. Pérez-Munuerarupo de Microestructura y Química de Alimentos, Departamento de Tecnología de Alimentos, Universitat Politècnica de València Camino de Vera s/n, 46022 Valencia, Spain

r t i c l e i n f o

rticle history:eceived 28 September 2010ccepted 21 March 2011

eywords:ersimmonigh hydrostatic pressureicrostructure

ryo-SEMight microscopyonfocal microscopy

a b s t r a c t

Persimmon fruit are an important source of phenolic compounds, dietary fibre, and carotenoids. How-ever, the location of these elements in the tissue is directly connected with processing techniques used.The aim of this work was to study the effect of high hydrostatic pressure (HHP) on the microstruc-ture of persimmon fruit cv. ‘Rojo Brillante’ during different ripening stages and the relationship of thistreatment with changes in the location of some bioactive compounds. Samples from persimmon fruitcv. ‘Rojo Brillante’ from two ripening stages, with and without deastringency treatment (95–98% CO2),were treated by HHP at 200 and 400 MPa during 1, 3, and 6 min. A microstructural study using cryo-scanning electron microscopy, light microscopy, and confocal laser scanning microscopy (Cryo-SEM, LM,and CLSM) was carried out. Total soluble solid (TSS) content and some textural properties were alsoanalyzed. The microstructural study showed that application of HHP caused cell wall disruption and

extural properties intracellular component dispersion throughout the tissue, together with some nutritionally interest-ing compounds (tannins, fibres, and carotenoids). TSS content diminished in astringent samples whenHHP was applied. This was attributed to precipitation of soluble tannins, which are responsible for fruitastringency. Therefore, it might be possible to omit a deastringency treatment with CO2 before HHP treat-ments. HHP treatments caused an overall decrease in firmness for both ripening stages and an increasein cohesiveness (TPA analysis) in the more advanced stage.

© 2011 Elsevier B.V. All rights reserved.

. Introduction

The beneficial effect of bioactive compounds and other nutri-nts in food, both vegetable and animal, depends on appropriatemounts of the compound reaching certain areas of the body inn active chemical form. During food processing, the reaction ofnzymes with their corresponding substrates and the extractionf nutrients and bioactive compounds from organelles and otherellular compartments is favoured. This leads to changes that alterensory (colour, texture, aroma) and nutritional qualities. Accord-ngly, the bioavailability of nutrients is affected by changes in the

icrostructure caused by processing (Parada and Aguilera, 2007).Food processing using high hydrostatic pressure (HHP) involves

he application of pressure on the product at a rate of between0 and 1000 MPa. The main objective of HHP food processing iso obtain safe and wholesome food, while maintaining the sen-

ory qualities. Most studies regarding the potential and limitationsf HHP food processing have focused on microbial and enzymaticnactivation. The effect of this technology on nutritional com-

∗ Corresponding author. Tel.: +34 963879830; fax: +34 963877369.E-mail address: mquichu@tal.upv.es (A. Quiles).

925-5214/$ – see front matter © 2011 Elsevier B.V. All rights reserved.oi:10.1016/j.postharvbio.2011.03.008

pounds, vitamins, and bioactive compounds in foods has beenless studied (Sancho et al., 1999; Patterson, 2000; Cano and DeAncos, 2005; Oey et al., 2008). Improvements in equipment designand research have enabled the development of a wide range ofcommercial fruit and vegetable produce treated by HHP (orangejuice, apple juice, guacamole, fruit jam, mashed tomato, and onionslices) (Torres and Velazquez, 2005; Rastogi et al., 2007). However,there are other fruit and vegetables, such as persimmon, wherethe implementation of HHP is still at an experimental stage and ithas only been tested on purees (De Ancos et al., 2000). Therefore itwould be interesting to evaluate the effects of HHP treatments onfresh-cut persimmon.

In recent years, the cultivation of persimmon or kaki (Diospy-ros kaki L.) cv. ‘Rojo Brillante’ has expanded greatly and the fruitis now considered an important alternative to other crops. Thepersimmon is a good source of phenolic compounds, mainly con-sisting of condensed tannins (group B proanthocyanidins). Thesecompounds have the ability to form stable complexes with metalsand proteins and are responsible for the astringency of persimmon

(Santos-Buelga and Scalbert, 2000; Nakatsubo et al., 2002). Theconversion of soluble tannins (in astringent kaki), into insolubletannins occurs during ripening or treatments such as the appli-cation of modified atmospheres with ethanol or CO2 (Arnal and

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38 J.L. Vázquez-Gutiérrez et al. / Postharves

el Río, 2003). Persimmons also contain high levels of phenoliccids, such as ferulic or p-coumaric, and contain twice as muchietary fibre as apples (Gorinstein et al., 2001). Persimmons are alsooted for their high level of carotenoid compounds and these con-ain significant antioxidants (�-carotene, �-cryptoxantina, lutein,eaxanthin, and lycopene). Some also are active with provitamin A�-caroteno and �-cryptoxantina) (De Ancos et al., 2000). Persim-

ons are also high in vitamin C and minerals such as potassiumWright and Kader, 1996).

There are few studies on the microstructure of ‘Rojo Brillante’Salvador et al., 2007, 2008; Pérez-Munuera et al., 2009a,b). ‘Rojorillante’ persimmons belong to the group that is astringent whenarvested, and astringency is removed with CO2. Firmness is thearameter that best describes the quality of this fruit (Salvadort al., 2007), and this is directly related to cell wall polysaccharides,ost notably pectins and hemicelluloses (Yakushiji and Nakatsuka,

007). The aim of this study was to examine the effect of highydrostatic pressure (HHP) on the microstructure of ‘Rojo Brillante’ersimmons at two ripening stages and changes in the location ofome bioactive compounds. This will help to a better understand-ng of the commercial viability of fresh-cut persimmons treated byHP.

. Materials and methods

.1. Sample preparation

Persimmon fruit (Diospyros kaki cv. Rojo Brillante) were har-ested at two different ripening stages of commercial maturityn Carlet and Alzira (Spain) at the beginning of November andecember of 2009, respectively. The maturity index used is a visualbservation of the external colour of the fruit (Salvador et al., 2007)nd six maturity stages are accordingly defined, ranging from Iyellow-green) to VI (orange-red). Stages III (RS1) and V (RS2) ofhis scale were studied in this work. For each ripening state, half ofhe batch was treated to remove astringency in closed containersith a 95% CO2 atmosphere for 20 h at 18 ◦C (non-astringent sam-les). After the treatment the fruit were stored at 4 ◦C, together withhe fruit that was not treated for astringency (astringent samples).

Cubes (15 mm) were taken from the equatorial area and placedn 110 mm × 220 mm plastic bags (Doypack type, Amcor, Spain).ach bag contained approximately 80 g of sample, that is to say,etween 20 and 22 cubes. The bags were gently pressed to partiallyxtract the air without damaging the cubes and then they wereeat-sealed. The bags were placed inside a hydrostatic pressurenit with a 2350 mL capacity and water was used as the pressureedium (GEC Alsthom ACB 900 HP, type ACIP 665, Nantes, France).

he pressures employed in the treatments were 200 and 400 MPa,or 1, 3, and 6 min, respectively, at 25 ◦C. Microstructure, total solu-le solids (TSS) and textural properties were analyzed within 24 hfter the treatment.

.2. Microstructural analysis

.2.1. Cryo-scanning electron microscopyA JSM-5410 SEM microscope (JEOL, Tokyo, Japan) was used with

Cryo CT-1500C unit (Oxford Instruments, Witney, UK) for theryo-SEM observation. Samples (1 mm thick pieces from the per-immon cubes) were placed in the holder, frozen in liquid nitrogenlush (T ≤ −210 ◦C). The frozen samples were then transferred to

he Cryo unit, fractured, etched for 15 min (−90 ◦C) and gold-coated2 mbar and 2 mA). Samples were then transferred to the micro-cope and examined at 10 kV, −130 ◦C and at a working distance of5 mm.

gy and Technology 61 (2011) 137–144

2.2.2. Light microscopyFor light microscopy (LM) observation, cryostat sections

(10 �m) were obtained from the persimmon cubes (CM1950, LeicaBiosystems, Nussloch, Germany).

Persimmon cubes were frozen at −60 ◦C in an ultra low tem-perature freezer (Dairei Europe, Denmark) for 24 h, transferred tothe cryostat and allowed to equilibrate with the warmer temper-ature of the cryostat (−25 ◦C) for 15 min. The frozen samples wereplaced in the holder and sections were cut with a disposable bladeusing an anti-roll plate. Sections were picked up with an artistbrush and transferred to coated glass slides, which had been previ-ously put inside the cryo chamber for improved adherence of tissuesections.

Some sections were stained with a 0.2% aqueous solution of tolu-idine blue. Other sections were stained with 10% vanillin-HCl (1:1,v/v) for the identification of tannins (Valette et al., 1998). Stainedsamples were examined under a Nikon Eclipse E800 light micro-scope (Nikon, Tokyo, Japan). Some non-stained cryostat sectionswere also studied.

2.2.3. Confocal laser scanning microscopyMicrostructural analysis was also performed using confocal

laser scanning microscopy (CLSM). This was achieved with aNikon confocal microscope C1 unit fitted on a Nikon Eclipse E800microscope (Nikon, Tokyo, Japan). An Ar laser line (488 nm) wasemployed as a light source and the autofluorescence of the samplewas observed through a 515/530 band-pass emission filter.

2.3. Total soluble solids and textural properties

2.3.1. Total soluble solidsTotal soluble solids of the fruit juice were determined using a

hand-held refractometer (Atago ATC-1E) and expressed as %. Juicewas prepared in triplicate for each treatment and 60 g of persim-mon cubes were blended for each juice.

2.3.2. Textural propertiesFlesh firmness and cohesiveness were determined at room tem-

perature with a TA.XTplus Texture Analyzer (Stable Micro Systems).Flesh firmness was expressed as the maximum breaking force innewtons (N) using a 4 mm diameter flat-tipped cylindrical probeat 1 mm/s test speed. A texture profile analysis was performed todetermine cohesiveness. The samples were axially compressed intwo consecutive cycles at 1 mm/s test speed and 75% compres-sion, 3 s apart, with a 50 mm diameter flat plunger. Cohesivenesswas calculated as the ratio of the area under the second curve tothe area under the first curve (dimensionless). Firmness and cohe-siveness values were an average of the measurements from eightcubes.

2.4. Statistical analysis

Results presented in graphs were reported as mean ± standarddeviation. Data was subjected to variance analysis (one-wayANOVA), using Least Significant Differences (LSD) multiple com-parison test with a 95% confidence interval for differences betweenmeans (Statgraphics Plus 5.1, Manugistics, Inc., Rockville, MA,USA). One-way ANOVA and LSD tests were performed between

the control and the different HHP treatments for both astringentand non-astringent samples. One-way ANOVA and LSD tests werethen performed to show differences between astringent and non-astringent samples with the same HHP treatment.

J.L. Vázquez-Gutiérrez et al. / Postharvest Biology and Technology 61 (2011) 137–144 139

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ig. 1. Cryo-SEM. Astringent persimmons at the RS1 ripening stage. Control (A andntercellular space; FIS: flooded intercellular space; SM: soluble material; TC: tanni

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. Results and discussion

.1. Microstructural study

The parenchyma of astringent ‘Rojo Brillante’ persimmons atS1 was quite compact, with small air-filled intercellular spaces.owever, some of the spaces were filled with soluble material from

he cells due to degradation during the maturation process (Fig. 1A).he parenchymatic cells observed in cross section were rounded,00–130 �m in diameter. The vacuole, full of soluble material, cov-red totally the interior of the cell, so that the tonoplast could note observed (Fig. 1D). Soluble solids were observed in the sampless a network; this is the typical eutectic artefact generated dur-ng the process of water sublimation while samples are preparedy Cryo-SEM. After HHP treatment at 200 MPa/1 min, a similar butore compact structure was maintained (Fig. 1B). Due to the degra-

ation of the tissue caused by pressure, cells were deformed andhigher number of intercellular spaces were filled with solubleaterial from the inside of the cells. When pressure was applied,

ntracellular liquid spread throughout the tissue. The tonoplast orlasmalemma could now be observed as a consequence of the com-ression during the HHP treatment. Moreover, some cell walls wereegraded. Some cells had a compact material inside their vacuolesFig. 1E). This structure corresponded to insoluble material, sincehe typical eutectic artefact could not be observed in these areas,nd it has been associated with the precipitation of soluble tan-ins (Salvador et al., 2007, 2008). This new structural appearance isnly observed in what some authors call tannic cells (Gottreich andlumenfeld, 1991; Yonemori et al., 1997). Some authors (Cheftel,992) have already shown that pressure produces a decrease inhe total molar volume associated with some chemical reactions.herefore, pressure could encourage the precipitation and poly-erization of tannins. At 400 MPa/6 min, cells collapsed, cell wallsere damaged and cells became hardly distinguishable (Fig. 1C).

n these samples, some small cavities occurred due to the expan-ion and compression produced by HHP treatment (Préstamo andrroyo, 1998). The intercellular spaces were completely filled with

iquid and insoluble material was also observed outside the cells inhese samples (Fig. 1F).

Non-astringent samples at RS1 showed, after all the treatments

Fig. 2), a similar structure to the corresponding astringent samplesFig. 1). However, the tissue was more sensitive to the application ofressure because the previous treatment with CO2 slightly affectshe cellular structure (Salvador et al., 2007). Non-astringent control

HHP treated samples at 200 MPa, 1 min (B and E) and 400 MPa, 6 min (C and F). IS:D: detached tonoplast/plasmalemma; CV: cavity; IM: insoluble material; CW: cell

samples (Fig. 2A and D) had a higher proportion of intercellularspaces filled with liquid. Moreover, the tannic cells were filled withinsoluble material (polymerized tannins) because of the applieddeastringency treatment. Probably, the HHP treatment producedpolymerization of some of the tannins that remained soluble afterthe deastringency treatment, not only inside but also outside thecells (Fig. 2E and F).

Tissue from the control samples of persimmon harvested atRS2 showed structural changes related to the maturation process(Fig. 3), when compared with tissue from control samples harvestedat RS1 (Figs. 1 and 2). The plasmalemma was highly degraded andseparated from the cell wall in many cells (Fig. 3A) and there wasan increase in cell-cell separation, above all in non-astringent sam-ples. Moreover, the majority of intercellular spaces were filled withsoluble material. Non-astringent samples showed the typical tan-nic cells from the treatment with CO2 (Fig. 3D). The applicationof pressure (Fig. 3B, C, E and F) generated changes equivalent tothose described for samples at RS1. At 400 MPa/6 min, degrada-tion of the tissue was much greater and most cells were no longerdistinguishable (Fig. 3C and F).

Tannins were localized before and after HHP treatment usinglight microscopy. In the control astringent samples at RS1, with-out HHP treatment, cell walls were stained in purple (Fig. 4A)and soluble tannins stained in red with vanillin-HCl (Fig. 4B).These tannins, which were originally inside tannic cells, were dis-persed throughout the tissue while the cryostat sections were beingobtained. The effect of pressure on cell wall fracture was noticeablewhen low pressure was applied (Fig. 4C) for short time periods(200 MPa/1 min). Moreover, because of the pressure applied, theprecipitated tannins were found inside tannic cells and in inter-cellular spaces stained in dark blue with toluidine blue (Fig. 4C).Tannins dispersed from the inside of cells and precipitated in theintercellular spaces were more clearly observed when sampleswere stained with vanillin-HCl (Fig. 4D). When higher pressure wasapplied, the tissue was more affected and the dispersion of tan-nins was greater (Fig. 4E and F). This could be related to a higheraccessibility of these compounds in HHP-treated samples.

Tissue from control non-astringent samples (Fig. 5A and B)showed polymerized tannins inside tannic cells owing to the effectof CO2 (Gottreich and Blumenfeld, 1991; Oshida et al., 1996). When

200 MPa/1 min was applied (Fig. 5C and D), greater cell distor-tion and cell wall damage than in the astringent samples occurred(Fig. 4C). Tannins had been already polymerized inside the cellsbefore the application of HHP. Therefore, they did not spread and

140 J.L. Vázquez-Gutiérrez et al. / Postharvest Biology and Technology 61 (2011) 137–144

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o precipitated tannins were observed outside the cells (Fig. 4D). At00 MPa/1 min, the tissue was completely degraded and the poly-erized tannins from the tannic cells were observed among the

ell wall remains (Fig. 5E and F).The effects of HHP treatment on the structure of persimmons at

S2 were similar to the effects previously described (micrographsot shown). However, the tissue from the control samples was moreegraded due to the maturation process in both astringent and non-stringent samples.

In view of the results obtained, it can be deduced that HHPreatment produces changes in the distribution and aggregationf tannins which could affect their extractability when the fruit hasot undergone CO2 treatment.

Autofluorescent bodies in the persimmon samples were distin-uished using confocal laser scanning microscopy. In a previoustudy regarding the quantification of carotenoids in yeast cells,n et al. (2000) concluded that autofluorescence at wavelengthsver 515 nm using an argon ion laser emitting 488 nm light was

rimarily due to carotenoids. Therefore, the brightest autofluo-escent bodies found in persimmon samples have been identifieds carotenoids, although cell walls were also observed as fluores-ent structures (Fig. 6A and D). When these non-stained sections

ig. 3. Cryo-SEM. Astringent (A–C) and non-astringent (D–F) persimmons at the RS2 ripend 400 MPa, 6 min (C and F). FIS: flooded intercellular space; DP: detached plasmalemm

) and HHP treated samples at 200 MPa, 1 min (B and E) and 400 MPa, 6 min (C and

were observed using light microscopy, carotenoids were found dis-tributed throughout the tissue (Fig. 6B) and they seemed to beassociated with cell walls forming spherical bodies (chromoplasts)surrounded by a membrane (Fig. 6C). These chromoplasts wereapproximately 15 �m in diameter.

As previously described by cryo-SEM, cell walls were degradedafter HHP treatment and this may explain the loss of fluorescentemission, although carotenoids still emit bright green fluorescence(Fig. 6D). In Fig. 6E, it can be observed using light microscopythat pressure favoured carotenoid diffusion from the chromoplasts,whose membrane could have been affected by the treatment. Indetail (Fig. 6F), carotenoids seemed to be composed of globules(approximately 0.75 �m in diam.) associated in chains. Pressurefavours the dispersion of these globules in the tissue. This couldfavour the bioavailability of carotenoids since they would now bemore dispersed.

3.2. Total soluble solids

The TSS content for control samples in both ripening stageswas approximately 17% in astringent samples and 15–16% in non-astringent samples (data not shown). A significant decrease in TSS

ning stage. Control (A and D) and HHP treated samples at 200 MPa, 1 min (B and E)a; TC: tannic cell.

J.L. Vázquez-Gutiérrez et al. / Postharvest Biology and Technology 61 (2011) 137–144 141

Fig. 4. Light microscopy. Astringent persimmons at the RS1 ripening stage. Control (A and B) and HHP treated samples at 200 MPa, 1 min (C and D) and 400 MPa, 1 min (Ea w: brc referr

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nd F). Toluidine blue stain (A, C and E) and vainillin-HCl stain (B, D and F). Red arroell. (For interpretation of the references to color in this figure legend, the reader is

ontent after CO2 treatment was observed for both ripening stages.his could be related to the insolubilization of tannins (Salvadort al., 2007).

When HHP was applied to fruit at RS1, a significant decrease inSS was observed in astringent samples, falling to values similaro those from non-astringent samples. This could be attributed tohe effect of high pressure on the precipitation of soluble tannins,s observed in the microstructural study (Fig. 1). Non-astringentamples did not show a variation in the TSS content. Therefore, thepplication of 200–400 MPa to astringent ‘Rojo Brillante’ persim-on fruit caused a decrease in TSS, mainly tannins, equivalent to

he decrease caused by the deastringency treatment with CO2.In fruit at RS2, TSS content in the samples treated with HHP

as higher than in the corresponding samples at RS1. This cane attributed to broken walls and cell membranes caused by the

pplication of HHP (see cryo-SEM discussion) which encourageshe solubilization of certain substances whose concentration in theruit increases as maturation progresses (sugars, pectic substancesrom cell walls, etc.).

oken cell wall; green arrow: precipitated tannins in intercellular spaces; TC: tanniced to the web version of the article.)

3.3. Textural properties

3.3.1. FirmnessFirmness is an important quality parameter in persimmons. In

this study, persimmons were considered fit for consumption whenfirmness values were above 2.5 N when measured with 4 mm diam-eter cylinder (Arnal and Del Río, 2004; Salvador et al., 2004). Thesamples at RS1 had a firmness of around 10.8 N, while the valuesat RS2 were between 3 and 4 N (Fig. 7). These values are consis-tent with those in previous studies (Salvador et al., 2004, 2005,2007; Besada et al., 2008; Igual et al., 2008). Softening of persim-mon fruit during ripening is associated with the solubilisation andhydrolysis of pectic substances in the cell wall due to the activity ofenzymes such as pectinmethylesterase (PME) and �-galactosidase(Nakamura et al., 2003; Ciardiello et al., 2004).

HHP treatment resulted in diminished firmness at both ripen-ing stages. When 200 MPa was applied for short periods (1 and3 min) to the astringent persimmon samples at RS1, the result wasa significantly lower value for firmness when compared to those

142 J.L. Vázquez-Gutiérrez et al. / Postharvest Biology and Technology 61 (2011) 137–144

Fig. 5. Light microscopy. Non-astringent persimmons at the RS1 ripening stage. Control (A and B) and HHP treated samples at 200 MPa, 1 min (C and D) and 400 MPa, 1 min( nnic cr

tethhtsntdnaetlbH(d

E and F). Toluidine blue stain (A, C and E) and vainillin-HCl stain (B, D and F). TC: taeferred to the web version of the article.)

reated with CO2. According to Basak and Ramaswamy (1998), twoffects can contribute to firmness value after HHP treatment. Onhe one hand, a tissue with completely flooded interstices hasigher resistance than that with air-filled spaces. On the otherand, the enzyme pectin methyl esterase (PME), which is boundo the cell wall in the intact cell, is liberated and contacts its sub-trate, the highly methylated pectin. This causes de-esterificationot only during the HHP application but also after the release ofhe pressure. The de-esterified pectin can form a gel-network withivalent ions such as Ca2+ and Mg2+, contributing to the firm-ess value. Persimmon contains approx. 6.5 and 9 mg of calciumnd magnesium for every 100 g of fresh fruit, respectively (Parkt al., 2006), and this may contribute to strengthening the struc-ure. Since a greater number of intercellular spaces are filled withiquid from the interior of cells in the non-astringent samples,

oth effects are expected to be more relevant in these samples.owever, when the HHP treatment conditions are more severe

200 MPa/6 min or 400 MPa/3 and 6 min), samples may be seriouslyisrupted leading to a permanent damage to the texture; Basak and

ell. (For interpretation of the references to color in this figure legend, the reader is

Ramaswamy (1998) observed this fact when treating apple with200 MPa and other vegetables such as carrots and red peppers with400 MPa.

Firmness values of samples at RS2 were lower than those of thesamples at RS1, with a decrease in firmness after the HHP treat-ments (Fig. 7B). As discussed in the microstructure section, whenhigh pressure was applied to persimmons at an advanced ripeningstage, damage in the tissue was much greater than for the ear-lier ripening stage. This would be the reason why firmness valueswere below the minimum acceptable levels in this case (Arnal andDel Río, 2004; Salvador et al., 2004). Furthermore, differences infirmness between HHP-treated astringent and non-astringent per-simmons were lower than those at RS1. This could be due to thefact that persimmon tissue at RS2 was already degraded before theHHP treatments due to the maturation process.

3.3.2. CohesivenessThe control samples at RS2 were less cohesive than those of

the samples at RS1. In addition, the astringent samples were sig-

J.L. Vázquez-Gutiérrez et al. / Postharvest Biology and Technology 61 (2011) 137–144 143

Fig. 6. Carotenoids in astringent persimmon parenchyma at the RS1 ripening stage. Control (A–C) and HHP treated samples at 400 MPa, 3 min (D–F). Confocal laser scanningmicroscopy (A and D) and light microscopy (B, C, E and F) images. CW: cell wall; CH: chromoplast; CC: carotenoid-lipid globules forming a chain. (For interpretation of thereferences to color in this figure legend, the reader is referred to the web version of the article.)

Fig. 7. Firmness (N) of samples of astringent and non-astringent persimmons with different HHP treatments: (A) ripening stage RS1; (B) ripening stage RS2. Means from eightreplicates ± standard deviation. Values with the same letter do not have significant differences (P < 0.05). Asterisks mean there are significant differences (P < 0.05) betweenastringent and non-astringent samples for a specific treatment and ripening stage.

Fig. 8. Cohesiveness of astringent and non-astringent persimmon samples with different HHP treatments: (A) ripening stage RS1; (B) ripening stage RS2. Means from eightr differa

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eplicates ± standard deviation. Values with the same letter do not have significantstringent and non-astringent samples for a specific treatment and ripening stage.

ificantly more cohesive than the non-astringent in both ripeningtages (Fig. 8).

When applying HHP to the samples at RS1 (Fig. 8A), there was aecrease in cohesiveness that could be attributed to the rupturef cell walls and membranes, as well as to separation betweenells. Only non-astringent samples treated at 200 MPa for 3 minaintained values similar to untreated samples.

However, at RS2 (Fig. 8B) the astringent samples treated with

HP maintained, or even increased cohesiveness in comparisonith untreated samples; non-astringent samples also experiencedgeneral increase in cohesiveness (which resulted especially

ences (P < 0.05). Asterisks mean there are significant differences (P < 0.05) between

high when 400 MPa was applied). Some authors have associatedincreased cohesiveness in other fruit with the formation of agel-like network between pectin and divalent ions (Basak andRamaswamy, 1998), as previously stated in the firmness section.

4. Conclusions

HHP treatment produced a significant effect on the structure ofpersimmon, affecting the integrity of cell walls and membranes.Much of the soluble tannins spread outside vacuoles, carotenoidsubstances were released from the chromoplasts and cell walls

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44 J.L. Vázquez-Gutiérrez et al. / Postharves

ere degraded. Due to these effects, the distribution of tannins,arotenoids and some fibre fraction changed after HHP treatments.his could affect the extractability of these compounds. The appli-ation of HHP provoked the precipitation of soluble tannins in “Rojorillante” persimmons, which could be related to the loss of astrin-ency. Moreover, the deastringency treatment with CO2 and theipening stage are significant factors in HHP-treated persimmons.

cknowledgements

The authors wish to acknowledge the Spanish Ministry ofcience and Innovation for the financial support (project AGL2008-4 798-C02-02) and the FPU grant given to Jose Luis Vazquezutierrez. The authors also thank to the “Agrupación Nacionale Exportación de Cooperativas Citrícolas” (ANECOOP) and to thepanish Council for Scientific Research (Instituto del Frío-CSIC) forhe supply and processing of the samples, respectively.

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