Effect of exopolysaccharide-producing strains of Streptococcus thermophilus on technological attributes of fat-free lassi

This article was downloaded by: [Mariano Garcia-Garibay]On: 30 April 2014, At: 13:22Publisher: Taylor & FrancisInforma Ltd Registered in England and Wales Registered Number: 1072954 Registeredoffice: Mortimer House, 37-41 Mortimer Street, London W1T 3JH, UK

International Journal of Food PropertiesPublication details, including instructions for authors andsubscription information:http://www.tandfonline.com/loi/ljfp20

Effect of an Exopolysaccharide-Producing Strain of StreptococcusThermophilus on the Yield and Textureof Mexican Manchego-Type CheeseDiana Lluis-Arroyoa, Angélica Flores-Nájeraa, Alma Cruz-Guerreroa,Francisco Gallardo-Escamillaa, Consuelo Lobato-Callerosb, JudithJiménez-Guzmána & Mariano García-Garibaya

a Departamento de Biotecnología, Universidad AutónomaMetropolitana, Col. Vicentina, México D.F., Méxicob Departamento de Preparatoria Agrícola, Universidad AutónomaChapingo, Texcoco, MexicoAccepted author version posted online: 26 Jun 2013.Publishedonline: 24 Apr 2014.

To cite this article: Diana Lluis-Arroyo, Angélica Flores-Nájera, Alma Cruz-Guerrero, FranciscoGallardo-Escamilla, Consuelo Lobato-Calleros, Judith Jiménez-Guzmán & Mariano García-Garibay(2014) Effect of an Exopolysaccharide-Producing Strain of Streptococcus Thermophilus on the Yieldand Texture of Mexican Manchego-Type Cheese, International Journal of Food Properties, 17:8,1680-1693, DOI: 10.1080/10942912.2011.599091

To link to this article: http://dx.doi.org/10.1080/10942912.2011.599091

PLEASE SCROLL DOWN FOR ARTICLE

Taylor & Francis makes every effort to ensure the accuracy of all the information (the“Content”) contained in the publications on our platform. However, Taylor & Francis,our agents, and our licensors make no representations or warranties whatsoever as tothe accuracy, completeness, or suitability for any purpose of the Content. Any opinionsand views expressed in this publication are the opinions and views of the authors,and are not the views of or endorsed by Taylor & Francis. The accuracy of the Contentshould not be relied upon and should be independently verified with primary sourcesof information. Taylor and Francis shall not be liable for any losses, actions, claims,proceedings, demands, costs, expenses, damages, and other liabilities whatsoever orhowsoever caused arising directly or indirectly in connection with, in relation to or arisingout of the use of the Content.

This article may be used for research, teaching, and private study purposes. Anysubstantial or systematic reproduction, redistribution, reselling, loan, sub-licensing,

systematic supply, or distribution in any form to anyone is expressly forbidden. Terms &Conditions of access and use can be found at http://www.tandfonline.com/page/terms-and-conditions

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

International Journal of Food Properties, 17:1680–1693, 2014Copyright © Taylor & Francis Group, LLCISSN: 1094-2912 print / 1532-2386 onlineDOI: 10.1080/10942912.2011.599091

EFFECT OF AN EXOPOLYSACCHARIDE-PRODUCINGSTRAIN OF STREPTOCOCCUS THERMOPHILUS ON THEYIELD AND TEXTURE OF MEXICAN MANCHEGO-TYPECHEESE

Diana Lluis-Arroyo1, Angélica Flores-Nájera1, AlmaCruz-Guerrero1, Francisco Gallardo-Escamilla †1 , ConsueloLobato-Calleros2, Judith Jiménez-Guzmán1, and MarianoGarcía-Garibay1

1Departamento de Biotecnología, Universidad Autónoma Metropolitana, Col.Vicentina, México D.F., México2Departamento de Preparatoria Agrícola, Universidad Autónoma Chapingo,Texcoco, Mexico

An exopolysaccharide-producing strain of Streptococcus thermophilus was evaluatedin the production of Mexican manchego-type cheese. This ropy strain improved waterand fat retention, and significantly increased cheese yield. Furthermore, the ropy straincheese retained more moisture than control cheese during ripening, suggesting thatexopolysaccharide strongly bound water within the protein matrix of the cheese. Scanningelectron microscopy confirmed that exopolysaccharide bound to the protein matrix of thecheese, producing a dense network that helped to increase water and fat retention andleading to a more open structure of the cheese that gave a softer product, as confirmedby instrumental texture profile analysis and sensory evaluation. Comparison of scanningelectron microscopy micrographs of the different sections of the cheese showed higher con-centration of exopolysaccharide in the centre than in the outer sections, indicating thatexopolysaccharide production continued during ripening and that the environment at thecentre of the cheese (moisture and/or oxygen concentration) favoured exopolysaccharideproduction. Instrumental texture profile analysis also demonstrated that the ropy straincheese was more cohesive and less elastic than the control; in contrast, exopolysaccharidedid not affect chewiness. The changes in texture could be correlated to composition: hard-ness increased as water and fat decreased, while springiness decreased with increasing fat.The interactions of exopolysaccharide with the cheese protein matrix had an affect on theincrease in cohesiveness of the ropy strain cheese.

Keywords: Streptococcus thermophilus, Exopolysaccharide-producing lactic acid bacte-ria, Mexican manchego-type cheese, Texture profile analysis, Ropy culture.

Received 25 February 2011; accepted 8 June 2011.†(1955-2010).Address correspondence to Mariano García-Garibay, Departamento de Biotecnología, Universidad

Autónoma Metropolitana, Iztapalapa Av. San Rafael Atlixco 186, Col. Vicentina, México D.F. 09340, México.E-mail: [email protected]

1680

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

YIELD AND TEXTURE OF ROPY MANCHEGO-TYPE CHEESE 1681

INTRODUCTION

Exopolysaccharide (EPS)-producing lactic acid bacteria (LAB) have been used in theproduction of some dairy products. Most studies that have used this kind of bacteria havebeen performed in yoghurt, where it has been shown to increase viscosity or decrease prod-uct syneresis.[1] These characteristics could be useful in generating other dairy products,such as cheese. Among the studies involving cheese, it has been reported that the use of acapsule-forming strain produced microstructural changes in the curd of mozzarella cheese,eliciting a more open casein network with larger pore sizes and softer texture than con-trol cheese.[2] Other investigations have reported that the use of a capsule-forming strainof Streptococcus thermophilus in mozzarella cheese production increases its moisture con-tent, hence improving its melting properties.[3–5] Besides mozzarella, other types of cheeseusing EPS-producing cultures include feta,[6] in which non-fat cheese made with a non-capsule-forming culture have a more compact structure compared to that made with acapsule-forming culture; karish, an Egyptian acid-coagulated cheese;[7] half-fat cheddar;[8]

and panela, a fresh Mexican cheese.[9] In all cases, textural modifications were observed.However, to the best of our knowledge, very little work has been undertaken with full fatcheese, where the fat content determines characteristics, such as flavour and texture. Theinfluence of EPS on fat retention, yield improvement and textural properties has not beenwidely assessed.

Most types of Mexican cheese are fresh, similar to most fresh white Latin Americancheeses. However, about 20% of the Mexican cheese market is composed of ripened cheese,the main representatives being the manchego-type and Chihuahua. These are very sim-ilar: slightly ripened with a semi-hard texture and usually made with a combination ofmesophilic and thermophilic cultures. The yield achieved for slightly ripened cheese islower than that of fresh types of cheese, which makes them less competitive in the Mexicanmarket. Thus, anything that could help increase yield without diminishing its taste and qual-ity would increase the competitiveness of these products. The aim of the current study wasto evaluate the use of Streptococcus thermophilus SY-102, an EPS-producing strain, on theyield and fat retention and some physical attributes of Mexican manchego-type cheese.

MATERIALS AND METHODS

Milk

Fluid pasteurised and homogenised whole cow’s milk containing 30 g·L−1 fat,31 g·L−1 protein, and a density of 1.032 g·mL−1 (Santa Clara, Productos Lácteos S.A.de C.V., Mexico City, Mexico) was used to produce cheese.

Starter Cultures

The ropy strain cheese was developed with a commercial mixed culture ofLactococcus lactis subsp. lactis and Lactococcus lactis subsp. cremoris (Rhône-Poulencde Mexico, Mexico City, Mexico) plus the ropy strain Streptococcus thermophilus SY-102(Rhodia-Texel Ltd., London, UK). The milk for cheese production was inoculated with 2%of the mesophilic cultures plus 0.5% of the S. thermophilus culture. The control cheese wasproduced with 2% of the mesophilic cultures, but the EPS-producing S. thermophilus strainwas replaced with the non-ropy strain Streptococcus thermophilus TA050 (Rhône-Poulencde Mexico). Starter cultures were prepared by incubating at either 37◦C (mesophilic) or

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

1682 LLUIS-ARROYO ET AL.

42◦C (S. thermophilus) for 18 h in skimmed milk with 10% total solids (Svelty, Nestlé,Mexico City, Mexico), sterilised at 121◦C for 5 min.

Cheese Making

The cheese was prepared in triplicate batches in bench cheese vats with 3.5 L of milkand 0.2 mL·L−1 of 0.3% calcium chloride solution (w/w). The milk was incubated at 37◦Cusing 2% of the corresponding starter mixture culture with constant measurements (every30 min) of viscosity, acidity, and pH until pH reached a value of 6.0. The milk was thenclotted using microbial rennet (3.5 mL, strength 1:10000; Cuamex, Mexico City, Mexico)and incubated for approximately 30 min. After coagulation, the curd was cut into cubesof approximately 1 cm per side; the temperature was slowly increased (1◦C min−1) up to45◦C ± 2◦C, while the curd was gently stirred. Once the temperature was reached, mostof the whey (approximately 80% of volume) was drained and the curd was kept in the vatflooded as small blocks in the remaining whey, turning the pieces periodically until thelactic acid in the whey reached 0.20 to 0.22%. The cheese was moulded into 1 kg roundmoulds and pressed for 24 h at a constant pressure of 1 kg·cm−2. After pressing, the cheeseswere submerged in 10% NaCl brine and allowed to salt for 24 h at 10◦C. The cheese wasripened in a chamber (10 to 12◦C, 80 to 90% relative humidity) for 2 weeks, turning thecheeses every third day and cleaning the surface with a 0.2% solution of potassium sorbate.

Physico-Chemical Analyses

Milk apparent viscosity was analysed by means of a Brookfield LV-DV III viscome-ter (Brookfield Engineering Laboratories Inc., Stoughton, MA, USA). Readings were takenwith LV4 or ULA spindles at speeds of 40 and 60 rpm, respectively, every 30 min up to180 min. Milk and whey acidity were determined by titration with 0.1 mol·L−1 NaOH(Sigma Chemicals, Mexico City, Mexico), using phenolphthalein as an indicator. ThepH was determined with a pH-meter Conductronic pH 20 (Conductronic, Mexico City,Mexico). The moisture content of cheese was analysed using 1 g of sample dried at 100◦Cto constant weight; total solids were calculated by difference of total weight and moisture.Fat content was determined using the Gerber method. Thereafter, fat in dry weight basesand total solids were calculated.[9] Total protein in the cheese was determined by extractionwith an 8 mol·L−1 urea solution at pH 8.0 as reported by Yong and Young.[10]

Cheese yield, reported as percentage, was calculated by the ratio between the finalweight of the cheese at different stages of production and the weight of the milk used forits production.

Scanning Electron Microscopy

The samples were taken with a cheese probe after 15 days of ripening from threedifferent sections of the cheese (centre, middle, and rind). They were cut into pieces ofapproximately 3 × 3 × 1 mm, fixed overnight at 4◦C in a 10 mL·L−1 glutaraldehydesolution (Electron Microscopy Science, Washington, DC, USA) with 5 mg·L−1 ofruthenium red to stain the EPS, followed by post fixation in 20 g·L−1 osmium tetroxide(Electron Microscopy Science). Dehydration was carried out at room temperature by anethanol (J.T. Baker, Xalostoc, Mexico) dehydration series (10 min each in 25, 50, 75, and99% ethanol solution, and finally three 10-min changes in 100% ethanol). Final dehydration

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

YIELD AND TEXTURE OF ROPY MANCHEGO-TYPE CHEESE 1683

was performed in a Samadri 780-B critical point dryer (Tousimis Research Corporation,Santa Clara, CA, USA) using CO2 as a transition medium. The specimens were mountedon aluminium scanning electron microscopy (SEM) stubs with carbon paint and coatedwith a layer of gold with a Sputter Coater SCD 050 (Bal-Tec, Balzers, Lietchtenstein).Specimens were viewed on a Zeiss DSM 940 digital scanning microscope (Oberkochen,Germany) at either 10 or 20 kV.

In spite of certain shrinking that could originate from sample treatment for SEM,the technique used to dehydrate the samples consisted of gradually replacing water withethanol followed by the replacement of ethanol with CO2. After that, CO2 was quicklysublimated to leave a dry sample. This technique allows the preservation of the originalstructure of the sample as much as possible. Thus, we can assume that even though themicrostructure of the cheese might have changed during the process, the structure observedin the micrographs is close to the original.

Instrumental Texture Profile Analysis (TPA)

Texture profile analysis was carried out in a TA.XT2i texture analyser (Stable MicroSystems, Texture Technologies Corp., White Plains, NY, USA). Cylindrical samples wereobtained from the centre of 15-day ripened cheese pieces using a 0.5-inch borer. Theirheight was adjusted to 1.1 cm with a blade. Samples were placed into Petri dishes and keptat 20◦C for 1 h; afterwards, they were compressed 60% in two cycles using the P50 probeand a crosshead speed of 120 mm·min−1; five samples of every cheese were analysed. Thevalues of the textural characteristics, hardness, springiness, cohesiveness, and chewinesswere calculated from the resulting curves by means of the equipment software (TextureExpert 1.20, Stable Micro Systems Ltd.).[11]

Sensory Evaluation

A selected and trained panel of ten members was used for the sensorial textureevaluation. Selection of the panel was done using duo-trio and triangle tests to differen-tiate commercial cheeses based on textural attributes. The selected members of the panelwere trained using duo-trio, triangle tests, and ranking multi-sample tests through severalsessions. Additional sessions were done to fully define textural attribute descriptors andscales knowledge. Training sessions were done with five commercial cheeses for everyattribute. To evaluate the textural attributes, the method of scaling was used by meansof a 10-cm unstructured scale with references at the ends and the evaluated attributeswere: hardness, springiness, cohesiveness, and chewiness. Cheese produced with eitherropy or non-ropy starter cultures were evaluated by the quantitative description analysistechnique. Samples were prepared by cutting cubical portions of approximately 1 cm perside, kept at room temperature for 1 h before tasting, and coded using random three-digitnumbers.

Statistical Analyses

All results were analysed by analysis of variance (ANOVA) using the StatisticalAnalysis Software (Statistica 5.0, Stat Soft, Tulsa, OK, USA), with P < 0.05 used as thethreshold for statistical significance.

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

1684 LLUIS-ARROYO ET AL.

RESULTS

Milk Fermentation

Results obtained for pH, acidity, and apparent viscosity during milk fermentationwith either control or the ropy strain are shown in Fig. 1. There were no significantdifferences in pH (P = 0.0928) and acidity (P = 0.5835) in milk fermented with eitherculture during the whole fermentation process. Figure 1 illustrates that the apparent viscos-ity of milk fermented with either the control or the ropy strain remained constant and equalfor both cultures during the first hour (P = 1.000 for time and P = 0.98 between cultures),and increased slightly (P = 0.0007 for time), but still with no difference between the con-trol and the ropy culture (P = 0.98), up to 90 min of fermentation. When the pH reacheda value below 6.2 (after 120 min of fermentation), the apparent viscosity of the ropy strainculture increased significantly (P < 0.02) with respect to the control, despite the fact thatpH diminished equally for both cultures.

Yield and Chemical Composition of the Cheese

The yield of manchego-type cheese was significantly higher (P = 0.0003) for the ropystrain cheese than for the control. Table 1 shows the cheese yield before and after saltingand in the final product. The increase in yield was statistically significant (P < 0.001) atthe three stages of production, demonstrating the effect of EPS. Composition analysis ofthe cheese is shown in Table 2, where it can be observed that water and fat contents arehigher in the ropy strain cheese with a slight decrease in total protein content. Figure 2shows the loss of weight of cheese during ripening, demonstrating a greater loss of waterin the control cheese.

Figure 1 Apparent viscosity (η), pH, and acidity changes during milk fermentation. �, pH control; �, pH ropy;�, η control; �, η ropy; ©, acidity control; �, acidity ropy.

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

YIELD AND TEXTURE OF ROPY MANCHEGO-TYPE CHEESE 1685

Table 1 Cheese yield.

Before salting After salting After ripening

Control cheese 11.16 ± 0.19a 11.67 ± 0.07c 9.43 ± 0.04d

Ropy-strain cheese 12.37 ± 0.28b 12.53 ± 0.28b 10.45 ± 0.06e

% enhancement of yield 10.8 7.4 10.9

Different superscript letters indicate significantly different values (P < 0.001).

Table 2 Chemical composition of the cheese (g·100 g−1).

TS Moisture FatTotal

protein Fat (dwb)Total protein

(dwb)

Control cheese 59.7 ± 0.5a 40.3 ± 0.5a 30 ± 0.0a 21.0 ± 1.2a 50.3 ± 1.0a 35.3 ± 0.6a

Ropy straincheese

55.3 ± 0.2b 44.7 ± 0.2b 34 ± 0.0b 19.0 ± 1.0a 61.5 ± 1.0b 34.3 ± 0.1b

TS: total solids; NFS: non-fat solids; dwb: dry weight basis.Different superscript letters in the same column indicate significantly different values (P <

0.005).

Figure 2 Weight variation in cheese during ripening. �, control; �, ropy.

Microstructure

Figures 3 and 4 show the microstructure by SEM of all the sections of both controland ropy strain cheese. Microscopic studies of the control cheese demonstrated no EPSproduction (Fig. 3) and a three-dimensional protein matrix formed by casein micelles towhich microorganisms adhered. SEM showed that in the ropy strain cheese (Fig. 4), theprotein network had more and larger open cavities than the control (Fig. 3). The polymerin the ropy strain cheese could be seen as part of the network binding to the protein matrixand to the fat globules; moreover, the number of visible fat globules was higher than in thecontrol cheese.

As manchego-type cheese is a pressed cheese, a different microstructure wasexpected across the different sections of the cheese. Due to this, for the SEM, three different

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

1686 LLUIS-ARROYO ET AL.

Figure 3 Microstructure (×5000) of three different sections of control Mexican manchego-type cheese: (A) rind,(B) medium, and (C) centre. The scale bar indicates 5 µm.

Figure 4 Microstructure (×5000) of three different sections of ropy Mexican manchego-type cheese: (A) rind,(B) medium, and (C) centre. The scale bar indicates 5 µm.

Table 3 Textural characteristics of ropy and control cheese.

Hardness (N) Cohesiveness Springiness (mm) Chewiness (Nmm)

Control cheese 8.67 ± 0.26a 0.15 ± 0.02a 6.90 ± 0.42a 8.96 ± 1.21a

Ropy strain cheese 6.73 ± 0.47b 0.18 ± 0.01b 6.24 ± 0.38b 7.66 ± 1.14a

Different superscript letters in the same column indicate significantly different values (P <

0.05).

sections of the cheese were observed: the centre, the middle, and the rind. The cheesemicrostructure was different in the different sections studied: the cavities in the proteinmatrix were wider in the centre than in the outer sections of the cheese, which is inagreement with the fact that the centre retained more water than the rind. Furthermore,comparison of the three sections showed that there was a higher concentration of EPS inthe centre than in the rind (Figs. 4a, 4b, and 4c).

Instrumental Texture Profile Analysis (TPA)

Figure 5 representative TPA curves obtained for both kinds of cheeses. Fromthese force-time graphs, the following parameters were evaluated: hardness, cohesive-ness, springiness, and chewiness, which were calculated according to Bourne.[11] Table 3shows the mean values of these TPA parameters and their standard deviation. Hardness andspringiness were significantly lower (P < 0.0001, P = 0.0299, respectively) in the ropystrain cheese, while cohesiveness was significantly higher (P = 0.0321) and chewiness wasnot significantly different (P = 0.1184) between the two kinds of cheese.

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

YIELD AND TEXTURE OF ROPY MANCHEGO-TYPE CHEESE 1687

Figure 5. TPA curves obtained for the two kinds of cheese. - - - Control, — ropy.

Sensory Evaluation

The same textural attributes that were evaluated by TPA were also assessed by theirsensorial appraisal by a trained panel. Results are shown in Fig. 6 and in this case hardnessand cohesiveness were significantly lower (P < 0.03) in the ropy strain cheese than in thecontrol, while springiness was significantly higher (P = 0.0299). Chewiness, on the otherhand, was not perceived as significantly different by the panellists (P = 0.3629).

DISCUSSION

Milk Fermentation

The decrease in pH together with the increase in acidity observed for both cultures(Fig. 1) indicate that the starter cultures developed as expected. When analysing the appar-ent viscosity obtained with both cultures (Fig. 1), it was observed that it remained almostthe same until a pH of 6.2 was reached. After that point, the apparent viscosity of the ropystrain culture increased significantly despite the fact that pH diminished very similarly forboth cultures. The measurement of apparent viscosity (Fig. 1) was used as an indirect indi-cator of EPS production. The changes in casein conformation due to acidification modifythe apparent viscosity of acidified milk. Nevertheless, it is generally agreed that even thoughthere is no straight relationship between an increase in viscosity and the concentration ofEPS, when EPS is produced the viscosity of the medium increases.[1,12–14] Therefore, thehigher increase in apparent viscosity of milk fermented with the ropy culture indicates thatEPS was produced.

Yield and Chemical Composition of the Cheese

The yield of manchego-type cheese at the three stages of production was higherwhen the ropy strain was used (P < 0.001), with an increase of 10.9% of the final

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

1688 LLUIS-ARROYO ET AL.

Figure 6 Comparison of the sensory attributes of ropy and control Mexican manchego-type cheese. �, control;�, ropy.

product (P = 0.0003) (Table 1). Before salting, an increase of 10.8% was observed(P = 0.0002), reflecting the retention, despite the pressing procedure, of water and fatbecause of their interactions with the EPS. Since salting was done by submerging inbrine, some water could be absorbed by the cheese to increase its weight, as seen inthe control cheese, which showed a significant increase (P = 0.0468) of 4.54%. Theropy strain cheese did not show a significant increase in weight (P = 0.8765), mostlikely because after pressing, EPS was already holding as much water as it could and itcould not absorb any more. During ripening, loss in weight occurs due to the evapora-tion of water. As shown in Fig. 2, the ropy strain cheese lost significantly less water thanthe control cheese (16.65% for the ropy and 19.16% for the control, P = 0.0003) as aconsequence of the ability of EPS to retain moisture efficiently. An increment in yieldbetween 13.7 and 16.8% in Mexican panela cheese has been reported when an EPS-producing culture is used.[9] However, panela is a fresh cheese with high moisture content,while manchego-type is a pressed and ripened cheese. Therefore, a lower increase inthe yield was expected, but the increase in the yield was statistically significant (P =0.0003), demonstrating the effect of EPS on cheese yield. Observing the composition ofthe two types of cheese (Table 2), yield improvement could be related to higher mois-ture and fat retention in the ropy culture cheese than in the control. Similar conclusionswere reported by Rynne et al.[8] in half-fat cheddar cheese made with a ropy culture;they obtained higher moisture and milk fat content, and an increase in the actual cheeseyield.

Total protein content (Table 2) decreased slightly in the ropy strain cheese.As reported elsewhere, the ropy behaviour of EPS-producing strains might be due to acomplex relationship between the exopolysaccharides and milk proteins beyond the soleproduction of the EPS, which has been explained partially by texture profile analysis

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

YIELD AND TEXTURE OF ROPY MANCHEGO-TYPE CHEESE 1689

by Dominguez-Soberanes et al.,[15] who determined that the higher the protein content,the more EPS-protein interactions occur in milk, leading to enhanced texture propertiesof the product. However, despite protein concentration, EPS-protein interactions con-tributed to the textural characteristics of the product, which was observed as an increasein viscosity during milk fermentation due to the matrix formed by caseins and EPS,as suggested by Hess et al.[16] and Dominguez-Soberanes et al.[15] This matrix couldretain more fat and water as demonstrated in the chemical analysis of the cheese(Table 2).

Microstructure

A higher water or fat content would change the cheese microstructure. To evaluatethe effect of EPS on cheese microstructure, SEM was used. Microscopic studies of controlcheese (Fig. 3) demonstrated no EPS production and a three-dimensional protein matrixformed by casein micelles to which microorganisms adhered. This is in accordance withother reports.[15,16]

SEM of the ropy strain cheese (Fig. 4) showed that the protein network had moreand larger open cavities than the control. These cavities contained higher amounts of water,possible due to the interactions of EPS with milk components and the protein matrix ofthe cheese, as observed in the SEM micrographs (Fig. 4) that show the interaction betweenEPS and the protein matrix leading to larger hollow spaces than in the control cheese.Data reported in Table 2 confirm the higher retention of water and fat within the cheese,hence increasing the yield. These results agree with those reported by Hassan et al.[6] whoobserved by confocal scanning laser microscopy a more open structure in feta cheese madewith a capsule-forming culture of S. thermophilus. Guzel-Seydim et al.[13] reported thatEPS strongly bound to water within the cheese matrix, retaining it for a longer time thanthe control cheese. This was observed during the ripening of the cheese, in which the ropystrain cheese retained more water than the control (Fig. 2).

The cheese microstructure was different along the different sections studied: the cav-ities in the protein matrix were wider in the centre than in the outer sections of the cheese,since the centre retained more water than the rind.

Comparison of the three sections of the ropy strain cheese (Figs. 4a, 4b, 4c) demon-strated that there was a higher concentration of EPS in the centre than in the rind. It seemsthat the production of EPS continued during ripening and occurred more frequently in thecentre where higher moisture content and an anaerobic environment was found comparedto the outer sections like the rind, where moisture is lower and oxygen concentration higher.Thus, the environment influences the growth of bacteria and/or EPS production.

Instrumental Texture Profile Analysis (TPA)

The higher content of water and fat in the ropy strain cheese (Table 2) contributes toa more open structure (Figs. 3 and 4), leading to a softer cheese that is also more cohesivebut less elastic (Table 3). This could be explained by the fact that higher fat and water con-tents tend to weaken the protein structure, resulting in a softer cheese, as has been reportedby several authors.[17–19] Additionally, according to Chen et al.[20] and Emmons et al.,[21]

higher fat content results in smoother and softer cheese. Furthermore, data reported byBryant et al.[22] regarding cheddar cheese with different fat contents established that themicrostructure of lower-fat cheese had a more compact protein matrix with less open

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

1690 LLUIS-ARROYO ET AL.

spaces that would have otherwise been occupied by fat globules. This is associated witha harder texture, even when the moisture content is high. These authors also showed thatthe open-intricate microstructure of the cheese analysed by SEM was lost, accompaniedby a decrease in fat content. The same effect can be observed in Figs. 3 and 4, where theropy strain cheese shows a more open structure, with milk fat globules observed withinthe protein matrix. All these findings are in accordance with data reported by Lobato-Calleros et al.[23] who observed that in Mexican manchego-type cheese, the reduction offat resulted in a harder cheese due to the compact and denser larger areas of protein matrixuninterrupted by fat globules.

Moreover, it is well established that cheese with higher moisture are less firmthan their lower-moisture counterparts due to the fact that the hydrophobic interactions,important for stabilising casein matrix structure, are weakened by adsorption of water byproteins.[17,24] This could contribute to the softer texture found in the ropy strain cheese.Since EPS increased water and fat contents, these factors could have primarily contributedto a softer cheese when the ropy culture was used. However, beyond the augmented con-tent of water and fat in the ropy strain cheese, EPS and its interaction with proteins andother cheese constituents should influence the texture profile. Cohesiveness is expected todiminish as water and fat contents increase,[22] but on the other hand, interactions betweenEPS and milk components, mainly proteins, could increase cohesiveness in the ropy straincheese, as has been established by Dominguez-Soberanes et al.,[15] who suggested that aninteraction between EPS and caseins increased cohesiveness in fermented milk. In a solidproduct, cohesiveness represents how well its structure withstands a deformation and/ortearing apart; the higher cohesiveness of the ropy strain cheese found in this study suggestsa strong interaction between EPS and caseins of the cheese matrix, in spite of the higher fatand water contents.

The results in springiness showed a less elastic ropy strain cheese than control. Bryantet al.[22] reported that springiness in cheddar cheese increased with decreasing fat con-tent, consistent with Emmons et al.[21] whom established that reduced fat cheese tends tobe more elastic. Based only on fat content, our results would be in agreement with thosereported by Emmons et al.[21] and Bryant et al.[22] However, the effect of moisture contenton springiness is not clear, and both increases and decreases in springiness with moisturecontent have been reported.[24]

It is clear that the relationship between cheese composition and its texture profileis very complex, and that other factors, such as process conditions and microbiota, mightalso influence the texture of the product. This makes it hard to establish a straight rela-tionship between the components and the texture profile alone, as has been reported by vanHekken et al.[25] and Tunick et al.[26] for Chihuahua cheese, which is similar to the Mexicanmanchego-type cheese.

Sensory Evaluation

Despite the fact that instrumental TPA is designed to simulate consumer perceptionof the physical properties of foods, human perception of the textural properties of fooddepends on many complex factors that the instrument does not consider on the wholewhile making the measurements. Because of this, it is always important to measure bothinstrumental and sensory perception of the texture of a product.

When comparing the results obtained in the TPA with those obtained by sensoryevaluation (Fig. 6), a direct correlation was found for hardness and chewiness. In both cases,

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

YIELD AND TEXTURE OF ROPY MANCHEGO-TYPE CHEESE 1691

hardness was higher for the control than for the ropy strain cheese, which (as discussedabove) could be due to the higher water and fat contents of the ropy strain cheese. There wasno significant difference in the chewiness of both types of cheese. The other parameters,cohesiveness and springiness, were perceived differently by the panellists compared to theinstrument.

It is clear that the same textural attributes are not necessarily perceived equally bythe human senses and an instrument, even when equipments such as the texture analyserhave been designed to mimic consumer texture perception. This has been pointed out byMarshall[27] and Jack et al.[28] The texturometer can simultaneously quantify a number oftextural properties, although only hardness has been shown to relate well to data obtainedfrom sensory analysis. Fracture patterns obtained by human actions differ markedly fromthose produced by instruments.[17,27]

In summary, the use of the exopolysaccharide-producing strain of S. thermophilusSY-102 in Mexican manchego-type cheese increased moisture and fat retention within thecheese matrix. This was reflected in the greater yield of cheese obtained. Scanning electronmicroscopy showed that EPS bound to the protein matrix of the cheese, microorganisms,and milk fat globules, leading to a more open structure of the cheese and producing anEPS-protein network that helped increase water and fat retention. Furthermore, interactionsbetween EPS and water were strong enough to retain moisture during cheese ripening.

The higher water and fat contents of the ropy strain cheese was reflected in a changein the textural characteristics, hardness being the most significant and also perceived bythe sensory evaluation panellists. The changes in textural characteristics were related tothe changes in cheese composition: hardness increased as water and fat decreased, whilespringiness decreased with increasing fat. The interactions of EPS with the protein matrixof the cheese affected cohesiveness, increasing it in the ropy strain cheese. The compositionof the cheese also influenced its sensory characteristics, giving a softer texture in the ropythan the control cheese. Other textural parameters, such as cohesiveness and elasticity, wereperceived differently by the panellists and the texturometer.

REFERENCES

1. Wacher-Rodarte, C.; Galván, M.V.; Farrés, A.; Gallardo, F.; Marshall, V.M.E.; Garcia-Garibay,M. Yogurt Production from reconstituted skim milk powders using different polymer and non-polymer forming starter cultures. Journal of Dairy Research 1993, 60, 247–254.

2. Hassan, A.; Frank, J.F. Modification of microstructure and texture of rennet curd by using acapsule-forming non-ropy lactic culture. Journal of Dairy Research 1997, 64, 115–121.

3. Perry, D.B.; McMahon, D.J.; Oberg, C.J. Effect of exopolysaccharide-producing cultureson moisture retention in low fat mozzarella cheese. Journal of Dairy Science 1997, 80,799–805.

4. Low, D.; Ahlgren, J.A.; Horne, D.; McMahon, D.J.; Oberg, C.J.; Broadbent, J.R. Role ofStreptococcus thermophiles MR-1C capsular exopolysaccharide in cheese moisture retention.Applied and Environmental Microbiology 1998, 64, 2147–2151.

5. Petersen, B.L.; Dave, R.I.; McMahon, D.J.; Oberg, C.J.; Broadbent, J.R. Influence of capsularand ropy exopolysaccharide-producing Streptococcus thermophilus on mozzarella cheese andcheese whey. Journal of Dairy Science 2000, 83, 1952–1956.

6. Hassan, A.N.; Frank, J.F.; Corredig, M. Microstructure of feta cheese made using different cul-tures as determined by confocal scanning laser microscopy. Journal of Food Science 2002, 67,2750–2753.

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

1692 LLUIS-ARROYO ET AL.

7. Hassan, A.N.; Corredig, M.; Frank, J.F.; Elsoda, M. Microstructure and rheology of anacid-coagulated cheese (Karish) made with an exopolysaccharide-producing Streptococcusthermophilus strain and its exopolysaccharide non-producing genetic variant. Journal of DairyResearch 2004, 71, 116–120.

8. Rynne, N.M.; Beresford, T.P.; Kelly, A.L.; Tunick, M.H.; Malin, E.L.; Guinee, T.P. Effect ofexopolysaccharide-producing adjunct starter cultures on the manufacture, composition and yieldof half-fat cheddar cheese. Australian Journal of Dairy Technology 2007, 62, 12–18.

9. Jiménez-Guzmán, J.; Flores-Nájera, A.; Cruz-Guerrero, A.E.; García-Garibay, M. Use ofan exopolysaccharide-producing strain of Streptococcus thermophilus in the manufacture ofMexican panela cheese. LWT–Food Science and Technology 2009, 42, 1508–1512.

10. Yong, K.J.; Young, W.P. SDS-PAGE of proteins in goat milk cheeses ripened under differentconditions. Journal of Food Science 1996, 61, 490–495.

11. Bourne, M.C. Food Texture and Viscosity: Concept and Measurement; Academic Press: NewYork, 1982.

12. Broadbent, J.R.; McMahon, D.J.; Oberg, C.J.; Welker, D.L. Use of exopolysaccharide-producingcultures to improve the functionality of low fat cheese. International Dairy Journal 2001, 11,433–439.

13. Guzel-Seydim, Z.B.; Sezgin, E.; Seydim, A.C. Influences of exopolysaccharide producingcultures on the quality of plain set type yogurt. Food Control 2005, 16, 205–209.

14. Champagne, C.P.; Barrette, J.; Roy, D.; Rodrigue, N. Fresh cheesemilk formulation fermentedby a combination of freeze-dried citrate-positive cultures and exopolysaccharide-producinglactobacilli with liquid lactococcal starter. Food Research International 2006, 39, 651–659.

15. Dominguez-Soberanes, J.; García-Garibay, M.; Casas-Alencáster, N.B.; Martínez-Padilla, L.P.Instrumental texture of set and stirred fermented milk. Effect of a ropy strain of Lactobacillusdelbrueckii subsp. bulgaricus and an enriched substrate. Journal of Texture Studies 2001, 32,205–217.

16. Hess, S.J.; Roberts, R.F.; Zieglier, G. Rheological properties of non-fat yogurt stabilized usingLactobacillus delbrueckii subsp. bulgaricus producing exopolysaccharide or using commercialstabilizer systems. Journal of Dairy Science 1997, 80, 252–263.

17. Jack, F.R.; Paterson, A. Texture of hard cheeses. Trends in Food Science and Technology 1992,3, 160–164.

18. Tunick, M.H.; Mackey, K.L.; Shieh, J.J.; Smith, P.W.; Cooke, P.; Malin, E.L. Rheology andmicrostructure of low-fat mozzarella cheese. International Dairy Journal 1993, 3, 649–662.

19. Tunick, M.H.; Malin, E.L.; Smith, P.W.; Shieh, J.J.; Sullivan, B.C.; Mackey, K.L.; Holsinger,V.H. Proteolysis and rheology of low-fat and full-fat mozzarella cheeses prepared from homog-enized milk. Journal of Dairy Science 1993, 76, 3621–3628.

20. Chen, A.H.; Larkin, J.W.; Clark, C.J.; Irwin, W.E. Textural analysis of cheese. Journal of DairyScience 1979, 62, 901–907.

21. Emmons, D.B.; Kalab, M.; Larmond, E.; Lowrie, R.J. Milk gel structure X. Texture andmicrostructure in cheddar cheese made from whole milk and homogenized low-fat milk. Journalof Texture Studies 1980, 11, 15–34.

22. Bryant, A.; Ustunol, Z.; Steffe, J. Texture of cheddar cheese as influenced by fat reduction.Journal of Food Science 1995, 60, 1216–1219, 1236.

23. Lobato-Calleros, C.; Robles-Martínez, J.C.; Caballero-Perez, J.F.; Aguirre-Mandujano, E.;Vernon-Carter, E.J. Fat replacers in low-fat Mexican manchego cheese. Journal of TextureStudies 2001, 32, 1–14.

24. Tunick, M.H.; Mackey, K.L.; Smith, P.W.; Holsinger, V.H. Effects of composition and storageon the texture of mozzarella cheese. Netherlands Milk and Dairy Journal 1991, 45, 117–125.

25. Van Hekken, D.L.; Tunick, M.H.; Tomasula, P.M.; Molina-Corral, F.J.; Gardea, A.A. MexicanQueso Chihuahua: Rheology of fresh cheese. International Journal of Dairy Technology 2007,60, 5–12.

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014

YIELD AND TEXTURE OF ROPY MANCHEGO-TYPE CHEESE 1693

26. Tunick, M.H.; Van Hekken, D.L.; Call, J.; Molina-Corral, F.J.; Gardea, A.A. Queso Chihuahua:Effects of seasonality of cheesemilk on rheology. International Journal of Dairy Technology2007, 60, 13–21.

27. Marshall, R.J. Composition, structure, rheological properties, and sensory texture of processedcheese analogues. Journal of the Science of Food and Agriculture 1990, 50, 237–252.

28. Jack, F.R.; Paterson, A.; Piggott, J.R. Relationships between rheology and composition of ched-dar cheeses and texture as perceived by consumers. International Journal of Food Science andTechnology 1993, 28, 293–302.

Dow

nloa

ded

by [

Mar

iano

Gar

cia-

Gar

ibay

] at

13:

22 3

0 A

pril

2014