International Journal of Food Microbiology 136 (2009) 26–31

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International Journal of Food Microbiology

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Effect of high pressure homogenization on Saccharomyces cerevisiae inactivation andphysico-chemical features in apricot and carrot juices

Francesca Patrignani a, Lucia Vannini a, Sylvain Leroy Sado Kamdem a,b,Rosalba Lanciotti a,⁎, M. Elisabetta Guerzoni a

a Dipartimento di Scienze degli Alimenti, University of Bologna, Viale G. Fanin 46, 40127 Bologna, Italyb Laboratoire de Microbiologie, Department of Biochemistry, University of Yaounde I, P.O. Box 812 Yaounde, Cameroon

⁎ Corresponding author. Campus Scienze degli AlimeCesena, Italy. Tel.: +39 0547 636132.

E-mail address: rosalba.lanciotti@unibo.it (R. Lancio

0168-1605/$ – see front matter © 2009 Elsevier B.V. Adoi:10.1016/j.ijfoodmicro.2009.09.021

a b s t r a c t

a r t i c l e i n f o

Article history:Received 15 April 2009Received in revised form 3 September 2009Accepted 18 September 2009

Keywords:Fruit juicesHigh pressure homogenizationSaccharomyces cerevisiaeViscosity

This experimental work aimed to evaluate the potential of high pressure homogenization (HPH) to inactivateSaccharomyces cerevisiae 635 inoculated both in apricot and carrot juices. The sensitivity of the yeast wasevaluated both in relation to its initial inoculum level (about 3.0 and 6.0 Log10 cfu/ml) and number of passesapplied at 100 MPa. Moreover, the effects of repeated pressure treatments at 100 MPa were evaluated for pH,water activity and viscosity of carrot and apricot juices.Data obtained showed that repeated high pressure homogenization passes at 100 MPa allowed a significantinactivation of the spoilage yeast inoculated in both juices. However, as expected, the inactivation of theconsidered strain was greatly affected by the food matrix. In fact, a higher viability loss of S. cerevisiae 635was observed in carrot juice than in apricot juice. Concerning the recovery, data obtained showed that thedecrease of the inoculum level to 3.0 Log10 cfu/ml prevented (at least for 6 days) cell proliferation in thesamples of apricot and carrot juices treated with more than four and seven passes, respectively. Also therefrigeration of the treated samples prevented cell recovery and, in some cases, induced a further decrease incell viability also in the samples inoculated with 6.0 Log10 cfu/ml allowing a further increase in the juiceshelf-life.An interesting and promising result was the enhanced viscosity of apricot juices treated up to five passes at100 MPa.

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© 2009 Elsevier B.V. All rights reserved.

1. Introduction

Fruit and vegetable juices are popular drinks consumed by peopleof all ages for their sensory and nutritional qualities. In fact, they areperceived as “healthy” foods due to their low content of sodium,minimal cholesterol, fat and richness in vitamin C, polyphenols andflavonoids contributing to good antioxidant properties (Kumar et al.,2009). Spoilage of fruit and vegetable juices is primarily due to theproliferation of its natural acid tolerant and osmophilic microflora(Tahiri et al., 2006). The microflora is represented by yeasts,responsible for the fermented taste and carbon dioxide production,lactic acid bacteria, which can produce a buttermilk off-flavour, andmoulds which contribute to the spoilage of juices by their surfacegrowth (Tournas et al., 2006). In addition, acid foods such as fruitjuices have always been considered as safe (due to the low pH).However, recent outbreaks of food-borne diseases, which have beenattributed to unpasteurized juices contaminated with pathogens suchas Salmonella spp. and Escherichia coli O157:H7, have demonstrated

that unpasteurized juices can be a vehicle for food-borne illnesses(Briñez et al., 2006b). Despite its negative effects on productnutritional and organoleptic values (Vachon et al., 2002; Pathanibulet al., 2009), heat treatment is the most widely used method forpreservation of fruit and vegetable juices due to its effectiveness inmicrobial inactivation (Tribst et al., 2008). Consumer demand forfresh unpasteurized fruit juices has notably increased worldwide inrecent years; consequently, many producers are reluctant to pasteur-ize their products because of the fore mentioned side effects(McDonald et al., 2000). The growing demand for fresh foods ofhigh microbiological quality has generated an increased interest innon-thermal processing technologies for the inactivation of themicroorganisms with negligible changes in the organoleptic andnutritional value of the raw materials (Diels et al., 2005). The non-thermal technologies proposed for preservation of fruit juices includeUV light (Koutchma et al., 2004), pulsed electric fields (Evrendileket al., 1999), high hydrostatic pressure (Cano et al., 1997; Houskaet al., 2006) and high pressure homogenization (HPH) (Lacroix et al.,2005; Diels and Michiels, 2006; Briñez et al., 2006a,b; Tahiri et al.,2006; Kumar et al., 2009; Pathanibul et al., 2009).

High pressure homogenization is considered to be one of the mostpromising of these new food processing technologies because of the

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recent improvements in high-pressure homogenizers and the accep-tance by consumers of pressure processed foods (Kheadr et al., 2002;Briñez et al., 2006a,b). HPH treatments have been recently used forthe stabilization of liquid food mixtures and the recovery of cellproteins and other biochemicals from microorganisms after celldisintegration. Several papers report the effectiveness of HPH indeactivating pathogenic and spoilage microorganisms in modelsystems and real foods (Lanciotti et al., 1994; Guerzoni et al., 2002;Diels et al., 2003; Briñez et al., 2006a,b; Taylor et al., 2007; Pathanibulet al., 2009), thus stimulating investigations on the application of HPHto improve food safety and shelf-life. Spatial pressure and velocitygradients, turbulence, impingement, cavitation and viscous shearhave been identified as the primary mechanisms of microbial celldisruption and food constituent modification during HPH treatments(Middelberg, 1995; Kleinig and Middelberg, 1998). The HPH treat-ment is reported also to act on food constituents, especially proteins,fat and polysaccharides with consequent modification of theirfunctional properties and susceptibility to enzymatic attack as wellas of food microstructure and rheological properties (Sandra andDalgleish, 2005). Although HPH has a good potential in several fields(i.e. pharmaceutical, biotechnological, chemical) in food industry ithas been studied and applied mainly for the dairy sector (Lanciottiet al., 2007; Vannini et al., 2008). On the other hand relatively fewinvestigations concerning fruit and vegetable juices stabilization areavailable. However, some studies showed the ability of HPH toincrease the safety and shelf-life of orange, carrot and apple juices. Inparticular, Tahiri et al. (2006) showed the great potential of HPHfor the inactivation of E. coli O157:H7 ATCC 35150, Lactobacillusplantarum ATCC 14917, Leuconostoc mesenteroides ATCC 23386 as wellas wild strains of Saccharomyces cerevisiae and Penicillium ssp. inoc-ulated in orange juices. Also Briñez et al. (2006b) reported cellreductions of E. coli O58:H21 ATCC 10536 and E. coli O157:H7 CCUG44857 inoculated into orange juice by HPH treatment. However,significant differences in lethality values between the two strainswere observed. The application of pressure levels ranging between100 and 200 MPa in combination with chitosan allowed the reductionof up to 5 Log10 cfu/ml of E. coli without thermal damages to the rawmaterials. However, pressure levels higher than 200 MPa gave rise toheat damages to juices. In addition HPH used alone or in combinationwith a pre-warming (50 °C for 10 min) showed good potential, tostabilize the opalescence of orange juices (Lacroix et al., 2005).

In this perspective, this study evaluated the effect of HPH at100 MPa on the survival of the yeast S. cerevisiae in apricot and carrotjuices after 1–8 successive passes through the homogenizer. Growthof surviving yeasts was measured during storage of treated juices at25°C and 4°C. In addition, the effects of the adopted HPH treatmentson the pH, water activity and viscosity of apricot and carrot juiceswere studied.

2. Materials and methods

2.1. Fruit juices

Commercial carrot and apricot juices were purchased from a localsupermarket. The samples were stored at refrigerated conditions(4 °C) before their use and warmed to room temperature beforeinoculation.

2.2. Strain and experimental procedures

S. cerevisiae 635, belonging to the collection of the Department ofFood Science, University of Bologna, was employed in this experi-mental work. The strain was cultured in Sabouraud medium (Oxoid,Basingstoke, UK) at 25 °C for 48 h and, in a first set of trials, inoculatedin carrot juices at levels of 3 and 6 Log10 cfu/ml. Before the inoculation,the cells were harvested by centrifugation (4000 rpm×6 min) and

washed with NaCl 0.9% solution to remove spent growth media. Thesame procedures and inoculum levels were adopted for apricot juices.

In a second set of trials, S. cerevisiae 635 was inoculated, adoptingthe procedure previously described, in refrigerated carrot and apricotjuices at a level of about 4.0 Log10 cfu/ml.

2.3. High pressure homogenization treatment

The inoculated fruit juices were subjected to repeated high pressurehomogenization (HPH) treatments at 100 MPa ranging from 1 to8 passes. A continuous high pressure homogenizer PANDA (Niro Soavi,Parma, Italy) was used for all the homogenizing treatments. Themachine was supplied with a homogenizing PS type valve with a flowrate of 10 l/h; the valve assembly includes a ball type impact headmadeof ceramics, a stainless steel large inner diameter impact ring and atungsten carbide passage head. The inlet temperature of fruit juicesinoculated at levels of 3 and 6 Log10 cfu/ml was 15 °C and the increaserate of temperature was about 2.5 °C/10 MPa. After each pass at100 MPa, the fruit juices were cooled by using a thermal exchanger(Niro Soavi, Parma, Italy). The maximum temperature reached by thesamples did not exceed 40 °C. As controls, untreated and 0.1 MPatreated inoculated juices were used. The samples, collected after eachpass at 100 MPa and the controls were stored at 25 °C. Differently, theinlet temperature of samples inoculated at a level of 4.0 Log10 cfu/mlwas 4 °C and the outlet temperature did not exceeded 30 °C. The treatedsamples were stored at 4 °C for 216 h.

2.4. Microbiological and physico-chemical analyses

The microbiological analyses were performed immediately afterhomogenization treatments and after 48, 96 and 144 h of storage at25 °C. Appropriate serial dilutions of untreated and treated sampleswere performed in sterile 0.9% (w/v) NaCl solution. In all cases, 0.1 mlvolumes from the selected dilutions were surface plated ontoSabouraud Agar (Oxoid, Basingstoke, UK). The plates were incubatedat 25 °C for 48 h. Treatment efficiencywas expressed as absolute valueof |Log10 (N/N0)|, where N was the survivors after pressure treatmentand N0 was the initial inoculated population into the sample prior toeach HPH treatment.

The pH and water activity (aw) were measured immediatelyafter treatment by using a pH-meter Basic 20 (Crison Instruments,Barcelona, Spain) and an Aqualab CX3-TE (Labo-Scientifica, Parma, Italy),respectively.

The viscosity was measured immediately after pressure treat-ments by using a falling ball viscometer (Thermo Electron, Karlsruhe,Germany). The viscosity (mPa/s) was calculated according to thefollowing equation:

η = K⁎ðρ1−ρ2Þ⁎t ð1Þ

where η=sample viscosity;K=constant in relation to the used sphere;ρ1=sphere density; ρ2=sample density (mass/volume); t=timenecessary to the sphere to run 10 cm (which is the value recordedwith the viscometer).

All the measurements were carried out at 20 °C.Microbiological and chemico-physical analyses are the mean of

three repetitions of the same experiment. The data and the graphswere analyzed and drown by means of Statitistica 6.0 for Windows(StatSoft. Inc.).

3. Results

In order to study the HPH inactivation dynamics of S. cerevisiae635 inoculated in apricot and carrot juices, repeated passes at100 MPa and two inoculation levels were used. The carrot and apricot

Fig. 2. Growth of Saccharomyces cerevisiae 635 inoculated (3 Log10 cfu/ml) in apricotjuice at 25 °C after successive passes through a high-pressure homogenizer at 100 MPa.Number of passes: control (♦), 1 (■), 2 (▲), 3 (□), 4 (●), 5 (○), 6 (♢), 7 (Δ), and 8 (⁎).The inlet temperature of the samples during HPH treatment was 15 °C. The data are themean of three repetitions. Variability coefficient ranged between 5% and 7%.

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juices had the following composition, respectively: protein, 0.7% and0.2%; carbohydrates, 6.0% and 14.0%.

Data relative to the cell load reductions of S. cerevisiae 635 inapricot juice, immediately after HPH, in relation to the severity of HPHtreatment and inoculum level are shown in Fig. 1. In apricot juiceinoculated with 6.0 Log10 cfu/ml a significant viability decrease(2.2 Log10 cfu/ml) was obtained only with four repeated passes at100 MPa. A further increase of the number of pushes at 100 MPa did notsignificantly increase the effectiveness of HPH treatment. In the samesamples inoculated at a level of 3.0 Log10 cfu/ml, three passes at100 MPa reduced cell viability under the detection limit. However, afterthe first pass at 100 MPa, the inactivation rate remained unchangedregardless of the inoculum level and the number of passes (data notshown). All the samples inoculated at the highest level spoiled duringthe storage at 25 °C regardless of the passes applied. In particular, themost severely treated samples reached a yeast level higher than6.0 Log10 cfu/ml after 96 h of storage at 25 °C (data not shown).According to Thomas et al. (2004), this cell load can be regarded as thethreshold for yeast spoilage. In the samples inoculated at a level of3 Log10 cfu/ml and subjected to four or more repeated passes at100 MPa, S. cerevisiae cell load remained under the detection limit atleast up to 144 h of storage at 25 °C (Fig. 2).

Concerning carrot juice, when S. cerevisiae was inoculated at alevel of about 6.0 Log10 cfu/ml, eight repeated passes at 100 MPawereunable to completely inactivate the inoculated cells (Fig. 3) and, after96 h of storage at 25 °C, cell loads exceeding 7.0 Log10 cfu/ml weredetected in all the samples (data not shown). Differently, with aninoculum level of 3.0 Log10 cfu/ml, three repeated passes at 100 MPawere able to decrease S. cerevisiae 635 cell viability under thedetection limit (Fig. 3). However, after 144 h of storage at 25 °C onlythe samples subjected to five up to eight repeated HPH treatments at100 MPa showed cell loads lower than 5 Log10 cfu/ml (Fig. 4). Aspreviously observed in apricot, also in carrot juice the survival ratio ofS. cerevisiae 635 as a function of the number of passes at 100 MPa wasnot influenced by the inoculum level (Fig. 5). The comparison of theinactivation levels in carrot and apricot juices clearly indicates ahigher sensitivity of S. cerevisiae strain 635 when inoculated in carrotjuice, even though an earlier recovery was observed in carrot samples.

One of the principal aims of using HPH at industrial level is theextension of shelf-life and the improvement of safety of refrig-erated fluids. Therefore, a second trial in which the inlet and outlettemperatures were 4 °C and not-exceeding 15 °C respectively, was

Fig. 1. Cell load reductions (Log10 cfu/ml) of Saccharomyces cerevisiae 635 inoculated inapricot juice (3 and 6 Log10 cfu/ml) after successive passes through a high-pressurehomogenizer at 100 MPa. The inlet temperature of the samples during HPH treatmentwas 15 °C.

performed with both the juices inoculated with S. cerevisiae 635. Thesamples were stored at 4 °C and the re-growth of the inoculated yeastwas monitored. The results obtained evidenced a decrease in cellviability of S. cerevisiae strain 635 in the apricot samples treated at100 MPa for 1–5cycles over storage (Table 1). In the most severelytreated samples S. cerevisiaewas under the detection limit immediatelyafter the treatment. However, after 96 h cell loads ranging between 3.3and 4.2 Log10 cfu/ml were observed in the samples treated at 100 MPafor 5–8cycles. However, no further cell load increasewas observed overthe refrigerated storage suggesting that a satisfactory extension of thejuices shelf-life occurred. The same behaviour during refrigeratedstorage was observed for S. cerevisiae in carrot juice samples treated at100 MPa for seven and eight passes (Table 1).

To better understand the different inactivation and recoverybehaviours of S. cerevisiae 635 in carrot and apricot juices, the effectsof the different HPH treatments on pH, water activity (aw) andviscosity of the treated samples were studied (Fig. 6a, b and c). TheHPH treatments had a significant effect on the pH and viscosity ofapricot juices (Fig. 6a and c), while no significant influence wasobserved in carrot samples (data not shown). Moreover the pH values

Fig. 3. Cell load reductions (Log10 cfu/ml) of Saccharomyces cerevisiae 635 inoculated incarrot juice (3.3 and 6 Log10 cfu/ml) after successive passes through a high-pressurehomogenizer at 100 MPa. The inlet temperature of the samples during HPH treatmentwas 15 °C.

Fig. 4. Growth of Saccharomyces cerevisiae 635 inoculated (3 Log10 cfu/ml) in carrotjuice at 25 °C after successive passes through a high-pressure homogenizer at 100 MPa.Number of passes: control (♦), 1 (■), 2 (▲), 3 (□), 4 (●), 5 (○), 6 (♢), 7 (Δ), and 8 (⁎).The inlet temperature of the samples during HPH treatment was 15 °C. The data are themean of three repetitions. Variability coefficient ranged between 5% and 7%.

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of apricot juices decreased by increasing the severity of the HPHtreatments, while the viscosity increased. The aw values of both juiceswere not affected by the increase of the number of repeatedtreatments. The application of five passes at 100 MPa resulted in themaximal viscosity increase.

4. Discussion

In recent years the interest on the application of high pressurehomogenization in order to improve the safety (Lanciotti et al., 1994,1996; Vachon et al., 2002; Kheadr et al., 2002), shelf-life (Guerzoniet al., 2002), organoleptic and functional properties (Lanciotti et al.,2004; Patrignani et al., 2007, 2009; Burns et al., 2008) of foods hasincreased. Nevertheless, few data on the effectiveness of HPH againstspoilagemicroorganisms inoculated in fruit juices are available (Tahiriet al., 2006).

The data obtained in the present research showed that repeatedHPH passes at 100 MPa allowed a significant inactivation of thespoilage yeast S. cerevisiae 635 inoculated in apricot and carrot juices.

Fig. 5. Survivors (Log10 N/N0) of Saccharomyces cerevisiae 635 inoculated in carrot juicein relation to the inoculum level and after successive passes through a high-pressurehomogenizer at 100 MPa. The inlet temperature of the samples before HPH treatmentwas 15 °C.

As expected, the inactivation of the considered strain was greatlyaffected by the food matrix. In fact a higher sensitivity of S. cerevisiae635was observed in carrot juice than in apricot juices regardless of theinitial contamination level. In fact it is well known that the suspensionmedium is one of themain factors affecting themicrobial sensitivity toHPH (Diels et al., 2005; Diels and Michiels, 2006). Its effect on theeffectiveness of HPH treatments has been mainly attributed to theinteraction of some constituents of the mediumwith the microorgan-isms (Donsì et al., 2009). The importance of the suspensionmedium isalso outlined by some papers on the application of HPH to improve thesafety and shelf-life of vegetable juices. Tahiri et al. (2006) comparedthe effectiveness of HPH treatment at 200 MPa for one, three and fivepasses on L. plantarum and L. mesenteroides in orange juice andphosphate buffer evidencing that these microorganisms were moreresistant in real system than in buffer. An opposite behaviour wasshown by Penicillium spp., while S. cerevisiae and E. coli resulted to beunaffected by the suspension medium. The increase of apricot juiceviscosity following the repeated HPH treatments is one of the factorsthat may be involved in the cell/system interaction. Cheftel (1995)attributed to the high sugar concentration of mandarin juices theprotective effect against microbial inactivation by high-pressuretreatment. Consequently, the higher sugar content and the increasingviscosities of the treated apricot juices with respect to carrot canexplain the greater effectiveness of HPH treatments against S.cerevisiae 635 in the latter food matrix. However, the treated apricotjuice was more stable over the refrigerated storage at 4 °C. Althoughthe relationship between medium viscosity and microbial growth hasnot been deeply investigated, Stecchini et al. (1998) reported asignificant reduction of Bacillus cereus growth rate with an increasingmediumviscosity. This reduction can be attributed to an indirect effectdue to a limited nutrient diffusion.

Although the initial inoculum level is reported to be an importantfactor affecting the HPH effectiveness, the rates of inactivation of S.cerevisiae 635 in relation to the number of passeswere independent ofthe initial cell concentration. On the other hand, many studies reportthat cell concentration has no discernible influence on cell disruptionefficiency over a wide range of cell concentrations and operatingpressure values (Diels and Michiels, 2006). However, few papers areinconsistent with these findings (Middelberg et al., 1991; Kleiniget al., 1995). Kleinig et al. (1995) explained the decrease in disruptionefficacy with increased cell concentrations by the increased viscosityof the homogenate. Moreover, Donsì et al. (2009) showed that theinitial cell concentration (studied in the range 3–8 Log10 cfu/ml) didnot affect the effectiveness of a single pass treatment, while the lowerthe initial microbial cell load was the lower the inactivation levelachieved for the multi-pass processes at 50 and 150 MPa.

Although theuse ofmultiple passes is considered a promisingway toimprove the lethality levels of HPH at reduced pressures (Donsì et al.,2007, 2009), thedata obtained in thiswork showed that the inactivationof S. cerevisiae 635 was not linearly related to the number of passes at100 MPa. In fact, the inactivation level achieved after each pass at100 MPa was lower than that of the previous one and tended to aplateau. The observed asymptotic value was affected by both the initialinoculum level and the juice type considered. Our experimental data arein disagreement with those of Wuytack et al. (2002), Diels et al. (2005)and Diels and Michiels (2006). These authors supported the hypothesisthat in a multi-pass treatment the effect of each pass is additive;therefore, each homogenization pass causes the same reduction of themicrobial load. On the contrary, our data are in agreementwith those ofDonsì et al. (2007, 2009) on E. coli, Lactobacillus delbrueckii and S.cerevisiae who observed a non-additive trend for multiple passprocesses at a given pressure level. They attributed this trend mainlyto the physiological diversity within a microbial population and to theexistence of resistant cells able to survive the repeated passes at thepressure applied. The HPH treatments used for carrot and apricot juicesinoculated with S. cerevisiae 635 at the highest rate were unable to

Table 1Survival and growth of Saccharomyces cerevisiae 635 in apricot and carrot juices during storage at 4 °C after repeated passes through a high-pressure homogenizer at 100 MPa.a

Number ofpasses at 100 MPa

Carrot juice Apricot juice

0b 96 h 144 h 216 h 0 96 h 144 h 216 h

Controlc 3.8±0.3 5.8±0.2 4.4±0.4 3.9±0.4 3.9±0.3 5.2±0.4 3.9±0.4 4.9±0.31 3.4±0.2 4.6±0.4 4.5±0.3 3.4±0.1 3.6±0.1 5.0±0.2 3.9±0.2 3.2±0.22 2.9±0.2 4.7±0.5 4.0±0.3 3.6±0.2 3.3±0.2 4.0±0.2 3.8±0.1 4.2±0.13 2.5±0.4 4.4±0.4 3.9±0.5 3.0±0.4 2.9±0.3 4.0±0.3 3.7±0.3 3.8±0.54 2.1±0.3 4.2±0.3 3.6±0.3 3.6±0.2 1.0±0.2 4.0±0.1 3.3±0.2 3.4±0.35 1.9±0.2 3.8±0.5 3.5±0.2 3.0±0.3 – 4.2±0.4 3.8±0.4 3.7±0.36 1.1±0.1 1.1±0.1 3.3±0.2 3.6±0.1 – 3.5±0.4 3.6±0.2 3.6±0.47 −d – 3.6±0.1 3.7±0.2 – 3.3±0.2 3.7±0.1 3.3±0.38 – – 3.7±0.4 3.5±0.3 – 3.7±0.3 3.5±0.2 2.9±0.1

a The values are expressed as Log10 cfu/ml. The yeast inoculum level was 4.0 Log10 cfu/ml. The inlet temperature of the samples during HPH treatment was 4 °C.b Immediately after treatment.c Sample treated at 0.1 MPa.d Under the detection limit.

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prevent the yeast proliferation during the subsequent storage at 25 °Cdue to the survival of the resistant cell fraction also in themost severelytreated samples. The decrease of the inoculum level to 3 Log10 cfu/mlprevented (at least for 6 days) cell proliferation in the samples of apricotand carrot juices treated with more than four and seven passes,respectively. This result can be considered an interesting goal because

Fig. 6. Effects of various passes through a high-pressure homogenizer at 100 MPa on pH(a), water activity (b) and viscosity (c) of apricot juice. The inlet temperature of thesamples before HPH treatment was 15 °C. Control corresponds to unprocessed sample.

the juice shelf-life is calculated on the basis of the time necessary toreach 6.0 Log10 cfu/ml, also in comparison with the commercial asepticpackaged juices, which are stable but have a negligible contamination ofmicroaerophilic fungi. Moreover, the refrigeration of the treatedsamples prevented cell proliferation and in some cases induced a fur-ther decrease in cell viability also in the samples with the highestinoculum resulting in a further shelf-life increase. The observed cellviability decrease during refrigerated storage was probably due to theexistence ofHPH-damaged cells.Also the fact that the cell loadsdetectedimmediately after the treatment were significantly lower than thoserecorded after 3days of refrigerated storage seems to contradict the “allor nothing” mechanism of HPH cell disruption (Diels and Michiels,2006) and support the hypothesis of sub-lethal injuries induced by theprocess (Vannini et al., 2004; Iucci et al., 2007; Donsì et al., 2007, 2009;Taylor et al., 2007). It is likely that HPH-damaged cells of S. cerevisiae635, which were unable to grow immediately after the treatment, re-paired the injuries (counts after 3 days), but were unable to multiplyduring the refrigerated storage.

Froma technological pointof view, the increaseof viscosity in apricotjuice, with the increase of the number of passes at 100 MPa, can beregarded as an important result already observed for other foodmatrices by several authors (Guerzoni et al., 2002; Hayes and Kelly,2003; Lanciotti et al., 2004; Patrignani et al., 2009). The viscositychanges were mainly attributed to the modifications induced by HPHtreatment on proteins conformation as well as to protein aggregationand interactionwith theother constituents of the systems, e.g. fat and/orpolysaccharides. Therefore, it is presumable that the main factorresponsible for the observed viscosity increase in apricot and carrotjuices during HPH treatments is the change of macromolecules such asproteins and polysaccharides, andmainly those involvingpectin (Flouryet al., 2002). It is likely that HPH treatments enhance the activity ofpectin methylesterase (PME — not completely inactivated by thethermal treatment), the exposure of pectin side chains, the interactionbetween protein and pectin molecules as well as the pectin gellingability. Moreover, the use of pasteurized apricot and carrot juices toavoid the interference by naturally occurring microbial population didnot exclude the involvement of PME in the rheological changes inducedby HPH treatment. In fact, the presence of a consistent residual PMEactivity in thermally stabilized industrial preparation of peach, pear,apple, apricot, red grapefruit and pineapple has been reported (Castaldoet al., 1997; Riener et al., 2009). The enhancement of PME activity issupposed to be fundamental for the observed rheological modification.On theother hand theHPH-induced potentiating effect on the activity ofseveral enzymes is documented (Vannini et al., 2004; Iucci et al., 2007).However, also low pH values and high sugar contents of food matricesare fundamental for the thermal gelling. The involvement of pectingelling, favoured also by the temperature raise during HPH treatments,

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is highlighted by the higher viscosity increase in apricot with respect tocarrot juice. The latterwas endowedwith lower concentrations of sugarandwith a higher pHvaluewith respect to apricot juice. It iswell knownthat pectin gelling is favoured by pH decrease and sugar concentrationincrease (Agoub et al., 2009).

Acknowledgement

This work was financed by the European Commission Project“Innovative non thermal processing technologies to improve the qualityand safety of ready-to-eat (RTE) meals— HighQ RTE”, contract no. FP6-FOOD-023140.

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