Effect of whey nanofiltration process combined with diafiltration on the rheological and physicochemical properties of ricotta cheese

Food Research International 56 (2014) 92–99

Contents lists available at ScienceDirect

Food Research International journal homepage: www.elsevier.com/locate/foodres

Effect of whey nanofiltration process combined with diafiltration on the rheological and physicochemical properties of ricotta cheese Elane Schwinden Prudêncio a,⁎, Carmen M.O. Müller a, Carlise B. Fritzen-Freire b, Renata D.M. Castanho Amboni a, José C. Cunha Petrus c a b c

Departamento de Ciência e Tecnologia de Alimentos, Universidade Federal de Santa Catarina, Rod Admar Gonzaga 1346, Itacorubi, 88034-001 Florianópolis, SC, Brazil Instituto Federal de Santa Catarina 89820-000, Xanxerê, SC, Brazil Departamento de Engenharia Química e Alimentos, Universidade Federal de Santa Catarina, Campus Universitário, Trindade, 88040-900 Florianópolis, SC, Brazil

a r t i c l e

i n f o

Article history: Received 4 September 2013 Accepted 10 December 2013 Keywords: Nanofiltration Diafiltration Whey Ricotta cheese Rheological properties

a b s t r a c t Whey after thermocalcic precipitation (TP) was submitted to nanofiltration (NF) and to nanofiltration followed by diafiltration (NF/DF). Because of their high protein content, the NF and NF/DF retentates from volume reduction factor (VRF) equal to 2.0 were chosen to produce ricotta cheese 1 and ricotta cheese 2, respectively. The ricotta cheeses were evaluated for their physicochemical, color, rheological and microstructural properties. No differences were noted between the protein contents and total solids of the ricotta cheeses produced through NF and NF/DF retentates. However, considering the low luminosity values, it was possible to note that the color parameters of ricotta 1 were more affected by manufacturing. The same occurred for the rheological properties, indicating that ricotta 1 showed greater deformability before cheese rupture, which was also confirmed through its greater hardness. All the ricotta cheeses were characterized by a tendency to be more viscous than elastic, which was confirmed by their microstructural characteristics. Therefore, the ricotta cheeses obtained from NF and NF/DF retentates can be considered a good alternative for utilization of whey. © 2013 Elsevier Ltd. All rights reserved.

1. Introduction Whey is watery liquid that remains after the casein curd that separates from the milk upon coagulation of the casein proteins in cheese production (Smithers, 2008). Considering whey as a valuable byproduct, it seems to be a very promising route in the development of membrane technology. Developments in nanofiltration (NF) technology have created the opportunity for an entirely new approach to utilization of whey (Román, Wang, Csanádic, Hodúr, & Vatai, 2009). The NF and diafiltration (DF) processes are applied for whey processing in order to increase protein content (Baldasso, Barros, & Tessaro, 2011; Butylina, Luque, & Nyström, 2006; Wang, Yang, Xing, & Xu, 2008). DF is also used for protein purification because it is able to eliminate problems related to high concentration in the concentrate/retentate and thus generate high purification (Ebersold & Zydney, 2004; Paulen, Foley, Fikar, Kovács, & Czermak, 2011). However, the lack of uniformity in its composition and the presence of fat in whey retentate may restrict its utilization (Morr & Ha, 1993). According to Pereira, Diaz, and Cobos (2002), it is recommended that whey be submitted to thermocalcic precipitation (TP) before the membrane process aiming to remove lipoprotein and, therefore, avoid membrane fouling. This pre-treatment is based on the formation of insoluble lipid-calcium phosphate aggregates at moderate temperatures and neutral pH. The associated proteins ⁎ Corresponding author. Tel.: +55 48 37215395; fax: +55 48 37219943. E-mail address: [email protected] (E.S. Prudêncio). 0963-9969/$ – see front matter © 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.foodres.2013.12.017

precipitate with the lipid-calcium phosphate aggregates (Baumy, Gestin, Fauquant, Boyaval, & Maubois, 1990). NF can be defined as a pressure driven membrane process for the separation and concentration of substances having a molar mass between 100 and 1000 Da (g mol−1) (Van Der Bruggen, Mänttäri, & Nyström, 2008). However, there is no exact and uniform definition for the term DF. As suggested by Fikar, Kovacs, and Czermak (2010), DF is a membrane-assisted process that is designed to achieve the twinobjectives of concentrating and purifying a multi-solute system according to a specific wash-water utilization strategy. DF is considered a work of art in membrane separation technology, where the water is added into the batch tank until the desired concentration of the retentate is reached (Lipnizki, Boelsmand, & Madsen, 2002). Unlike the conventional methods of whey concentration, such as thermal evaporation, NF and DF represent very attractive alternative methods because they employ temperatures that do not involve phase change, which make the whey concentration process more economical. Román et al. (2009) reported that the whey cannot be stored in liquid state for a long time, so the NF and NF combined with DF would open doors to reduce production costs. It is noteworthy that heat treatment can change the characteristics of whey components in the concentrate, mainly the proteins, which are thermolabile and can decrease their nutritional and functional properties, such as the change in their physical properties when used in food production. Moreover, the other disadvantage of the conventional method is its high energy consumption (Baldasso et al., 2011).

E.S. Prudêncio et al. / Food Research International 56 (2014) 92–99

Román et al. (2009) reported that normally whey retentate obtained with membrane technologies is utilized by the sweets and by the pharmaceutical industries, as well as in the production of soft drinks. However, the use of whey retentate resulting from NF and/or DF, in cheese manufacture such as ricotta cheese, is practically inexistent. Ricotta cheese has traditionally been prepared by heating whey and acidifying the hot liquid with lactic acid to coagulate whey proteins (Di Pierro, Sorrentino, Mariniello, Giosafatto, & Porta, 2011; Modler & Emmons, 2001; Pizzillo, Claps, Cifuni, Fedele, & Rubino, 2005). The coagulated curd mass floats to the surface and is scooped off and placed in perforated trays for drainage (Modler & Emmons, 2001). Rheological measurements, as uniaxial compression and stress relaxation, have been successfully used in order to describe the changes that are typical of several cheeses (Del Nobile et al., 2007). When the technology involved in the production of a cheese is modified, its physical, chemical and rheological properties can change. Moreover, the development of technologies capable of solving the problem of whey and its utilization as a good source of protein can bring economic and environmental advantages. In this context, the aim of this present work was to evaluate the effect of NF and NF followed by DF to obtain two types of retentates (NF and NF/DF retentates). Each of the retentates was used to produce the ricotta cheeses, which were evaluated for their physicochemical, color, rheological and microstructural properties. 2. Materials and methods 2.1. Materials Commercial pasteurized milk (3 g fat 100 mL−1), lactic acid foodgrade solution (Purac Sínteses, Rio de Janeiro, Brazil), calcium chloride (Vetec, Rio de Janeiro, Brazil), and commercial rennet with a chymosin produced by Aspergillus niger var. awamori (with a strength of 1/3000, Ha La®, Chr. Hansen, Valinhos, Brazil) were used. All the chemicals used were of analytical grade. 2.2. Manufacture of cheese whey The whey was obtained from Minas Frescal cheese according to the procedures suggested by Souza and Saad (2009), with modifications. The Minas Frescal cheese was produced in a 150 L vat from pasteurized milk heated to 37 ± 1 °C, with addition of lactic acid (0.25 mL L−1 of an 85% lactic acid solution). Calcium chloride (0.4 mL L−1 of a 40% calcium chloride solution) and commercial rennet at a ratio of 1:3000 (0.9 mL L−1 of milk) were added to the cheese-milk followed by incubation at 37 ± 1 °C for 40 min. The resulting gel was gently cut into cubes, allowed to drain, and placed in cylindrical perforated containers, each with capacity for 500 g. One part of the whey resulting from this procedure was submitted to thermocalcic precipitation (TP) in order to remove the lipoprotein fraction, as proposed by Fauquant, Vieco, Brule, and Maubois (1985). These authors reported that this fraction in whey favors rapid membrane fouling with substantial decrease in its performance. 2.3. Nanofiltration (NF) and diafiltration (DF) procedures Approximately 15 L of whey after TP were used as the feed for the nanofiltration (NF) process and 15 L for the nanofiltration followed by diafiltration (NF/DF). Both filtration processes were performed using a tangential filtration system on a pilot scale equipped with a spiral membrane module (Osmonics membranes, HL2521TF, Minnetonka, United States), with an approximate molecular weight cut-off of 150 and 300 Da, 0.6 m2 of filtration area and 98% rejection of MgSO4 in a test performed using a spiral module at 25 °C and 690 kPa. Previous experiments were performed to determine the optimal parameters for the NF and NF/DF processes. The operating parameters controlled during NF and NF/DF were at a temperature of 24 ± 1 °C, which is suitable to

93

preserve the properties of whey (Ruffin, Schmit, Lafitte, Dollat, & Chambin, 2013) and a pressure of 300 kPa. During these filtration processes (NF and DF), the permeate flux (J) was measured every 10 min and calculated according to Eq. (1): J¼

V P  −1 −2  Lh m t  AP

ð1Þ

where VP(L) is the volume of permeate collected during the time interval t(h) and AP(m2) is the membrane surface area of permeation. The volume reduction factor (VRF) was calculated as the ratio between the initial feed volume (L) and the volume of the remaining retentate (L) after the considered operation time. The retention index, which shows the relationship between the amounts of the compound of interest in the permeate solutions and in the retentate solutions and also shows the ability of the membrane to retain this compound within the experimental conditions, was calculated according to Eq. (2). R¼

  C 1− P CR

ð2Þ

where CP (g 100 g−1) is the concentration of the protein content in the permeate and CR (g 100 g−1) is the concentration of the protein content in the retentate. In the DF process, deionized water (volume equal to the volume of the permeate obtained in the final stage of NF) was added to the NF retentate, followed by concentration up to the VRF chosen for the NF process, resulting in the NF/DF retentate. After each processing, the pilot unit and the membrane were cleaned and hygienized, according to the manufacturer's instructions. All the filtration experiments were performed in triplicate. 2.4. Manufacture of ricotta cheeses The whey, the NF retentate and the NF/DF retentate were used to produce three batches in triplicate of each type of the ricotta cheeses denominated of control, ricotta 1 and ricotta 2, respectively, according to the procedures suggested by Pizzillo et al. (2005), with modifications. In the manufacture of the ricotta cheeses, the whey, the NF retentate and NF/DF retentate were heated up to 85–90 °C and then coagulated with 100 mL of lactic acid solution, obtained from dilution of 85 mL of lactic acid in 100 mL of water. The precipitation occurred rapidly at this temperature, and after about 2 min the precipitate was stirred gently, drained, removed with a skimmer and placed in perforated circular containers and kept overnight under refrigeration (5 ± 1 °C) for complete draining. After that, the ricotta cheeses were vacuum sealed in plastic bags Cryovac® (BN, São Paulo, Brazil) and storage also at 5 ± 1 °C. 2.5. Physicochemical properties The samples of whey, retentates, permeates and cheese were analyzed for content of total solids (g 100 g− 1) by drying to constant weight at 105 °C, as described by the Analytical Norms of the Adolfo Lutz Institute (IAL, 2005), and for proteins (g 100 g−1), by the Kjeldahl method (N × 6.38) (AOAC, 2005). All analyses were performed in triplicate. 2.6. Color measurements The color measurements of the cheese samples were carried out with a previously calibrated Minolta Chroma Meter CR-400 colorimeter (Konica Minolta, Osaka, Japan) adjusted to operate with D65 illuminant and observation angle of 10°. The CIElab color scale was used to measure the L*, a* and b* parameters. The L* parameters indicate luminosity

94

E.S. Prudêncio et al. / Food Research International 56 (2014) 92–99

while, the a* axis shows the variation from red (+a*) to green (−a*) and the b* axis is the variation from yellow (+b*) to blue (− b*). The measurements were performed ten times for each sample. 2.7. Rheological properties The uniaxial compression and stress relaxation of cheese samples were carried out using the TA.XT plus texture analyzer (Stable Micro Systems Ltd., texture exponent for Windows software, Surrey, United Kingdom) fitted with a 50 kg load cell and a 25 mm diameter aluminum probe. Five samples were prepared by removing cylindrical pieces from different points of each cheese (19 mm diameter, 20 mm height) which were then kept in refrigeration (5 ± 1 °C) until testing, without addition of any lubricants. 2.7.1. Uniaxial compression The cylindrical cheese samples were compressed to 50% of their height at a cross-head speed of 1 mm s−1. The stress (σ) was calculated through Eq. (3), as proposed by Calzada and Peleg (1978): σ ðt Þ ¼

F ðt Þ

ð3Þ

Að t Þ

where σ(t) is the stress at time (t); F(t) is the force at time (t); A(t) is the area at time (t). The strain (ε) was also calculated as proposed by Calzada and Peleg (1978), as shown in Eq. (4): ε ¼ ln

H0 H 0 −ΔH

ð4Þ

where ε is the strain; H0 is the original height; ΔH is the change in height. The fracture stress (σf) and the fracture strain (εf) were derived in relation to the fracture point, which is defined as the local of maximum stress/strain of the curve (Wium & Qvist, 1997). 2.7.2. Stress relaxation The cheese samples were subjected to 10% compression for 60 s at a cross-head speed of 1 mm s−1. The experimental results for the stress relaxation were normalized and analyzed through the empirical model proposed by Peleg (Laurindo & Peleg, 2007; Müller, Laurindo, & Yamashita, 2009; Peleg, 1980), shown in Eq. (5): F ðt Þ F0

¼ 1−

c1  t c2 þ t

ð5Þ

2.8. Scanning electron microscopy (SEM) The cheese samples were prepared for scanning electron microscopy (SEM) by a method adapted from that proposed by Lobato-Calleros, Ramirez-Santiago, Osorio-Santiago, and Vernon-Carter (2002). Cylindrical samples of 0.5 cm diameter by 0.5 cm height were fixed in 2% buffered glutaraldehyde (0.1 M phosphate buffer, pH 7.2) for 6 h. The samples were dehydrated in increasing concentrations of aqueous ethanol solutions (50%, 60%, 70%, 80%, 90% and 100%, 30 min in each) and placed in acetone for 1 h. After that, the samples were dried in a liophilizator Terroni (LD 3000, São Carlos, Brazil) and each sample was then fractured perpendicular to its long axis, mounted on stubs with fractured face upwards, and coated with a thin layer of gold using a Leica EM SCD 500 sputter coater (Wetzlar, Germany). A scanning electron microscope (Jeol JSM 6390 LV, Tokyo, Japan) was used at 7 kV to view each sample at a magnification of 100× (200 μm). 2.9. Statistical analysis The data were expressed as means and standard deviation. One-way analyses of variance (ANOVA) and Tukey's studentized range (5% significance) were carried out to test for any significant differences between the results. The data were obtained using the software STATISTICA version 7.0 (StatSoft Inc., Tulsa, OK, USA). 3. Results and discussion 3.1. Nanofiltration (NF) and diafiltration (DF) procedures The initial permeate flux (J) values for NF and for diafiltration (DF) after NF process (Fig. 1) of whey were similar to and/or lower than those obtained by Yorgun, Balcioglu, and Saygin (2008) (~24 L h−1 m−2) and Suárez, Lobo, Alvarez, Riera, and Álvarez (2009) (~30 L h−1 m−2), respectively. As noted by Baldasso et al. (2011), the J value in the DF steps was below the initial flux. According to Suárez et al. (2009), the lower values of J are probably due to the presence of proteins, which can form a gel layer on the membrane surface. As reported by Pan, Song, Wang, and Cao (2011), an initial drop in the J values was observed in NF and DF after NF process, with a severe flux decline, which later stabilizes during the process. In accordance with the results obtained by Yorgun et al. (2008), it was also verified that the membrane is quickly fouled in the first 20 min. Several factors can cause flux decline, such as the increase in the osmotic pressure of the feed solution (Suárez et al., 2009), concentration polarization phenomena (Pan et al., 2011; Rinaldoni, Tarazaga, Campderrós, & Padilla, 24

RRELAX ¼

F ð60 sÞ  100ð% Þ F0

where F0 is the force at t = 0 and F(60 s) is the force at t = 60 s.

22

JNF = 21.38e -3.24t

20

JDF = 17.77e -4.16t

18

J(L.h-1.m-2 )

where F(t) is the force at time (t); F0 is the initial force. The parameters c1 and c2 were estimated by non-linear regression using STATISTICA version 7.0 (StatSoft Inc., Tulsa, OK, USA). In this model, 1 − c1 and c1/c2 provide information about the viscoelastic characteristics of the samples. The value of 1 − c1 can be seen as a “degree of solidity”, while the ratio c1/c2 represents the initial rate of the stress decay. The “degree of solidity” is associated with the global behavior of the material, thus all the experimental data were considered. In order to obtain a precise estimation of the initial decay rate, the data were adjusted using the first one hundred experimental points from the relaxation curve, without dimension through the ratio F(t)/F0 versus time as suggested by Müller et al. (2009). The relaxation ratio at 60 s (RRELAX) was determined through Eq. (6), as proposed by Peleg (1979):

16 14 12 10 8 6 4 2 0

0

1

2

Time (h) ð6Þ Fig. 1. Permeate flux (J) of whey after thermocalcic precipitation submitted to (●) nanofiltration (NF) and to (□) diafiltration (DF) after NF process, up to volume reduction factor equal to 2.

E.S. Prudêncio et al. / Food Research International 56 (2014) 92–99

2009), adsorption, pore plugging and gel-layer formation due to presence of protein (Rinaldoni et al., 2009). The data in Fig. 1 also confirm what Atra, Vatai, Bekassy-Molnar, and Balint (2005) had already noted, i.e., that probably there was a deposit of the largest and more dense layer, which reduced the permeate flux until reaching a static condition. In the present experiment, it was also noted that the J of whey during the DF processes was lower than the fluxes of whey permeate during the NF. It is noteworthy that the DF is a method that not only enhances the purity of the retained stream (Fikar et al., 2010; Yin, Yang, Zhong, & Xing, 2011), but also can eliminate the impurities from the solution (Paulen et al., 2011). Thus, the process of whey nanofiltration and diafiltration (NF/DF) may be able to generate a purer retentate, principally because of the type of protein present, which in turn contributes to the increase in membrane fouling, resulting in lower J values. The same was observed by Suárez et al. (2009) for the experiments performed with whey, which may represent a significant source of membrane fouling, showing an additional resistance to J. MourouzidisMourouzis and Karabelas (2006) reported that aggregates higher than 1 μm, as protein aggregates, can become firmly “packed” inside membrane pores. The volumetric reduction factor (VRF) reached a maximum of 2.0. Similar value of VRF in whey was obtained by Cuartas-Uribe et al. (2009), while slightly higher values (VRF = 2.5) were obtained by Román et al. (2009). Cuartas-Uribe et al. (2009) reported that NF avoids protein adsorption onto the NF membrane surface, which could produce membrane fouling, and thus also resulting in low VRF values. However, a number of factors may affect the VRF, such as distribution of pore size, surface morphology, and adhesion onto membrane (Tsuru, Sudoh, Yoshioka, & Asaeda, 2001) and composition of the raw material (AlMalack & Anderson, 1997). During NF, for both VRFs (1.5 and 2.0), the retention index (R) of protein was equal to 0.9. Both values were higher than that reported by Sarkar, Ghosh, Dutta, Sen, and Bhattacharjee (2009), who obtained an R value of around 0.4, when applying the same pressure that the used in this present experiment. These authors presume that conformational changes of different proteins may have occurred. In fact, a low VRF and a high R of protein is expected because, according to Baldasso et al. (2011), the process becomes less capable of delivering the desired product specification after long hours of operation when long-term fouling is more significant. It is noteworthy that the high value of R could be because of the thermocalcic precipitation (TP) carried out on the whey before the NF and NF/DF processes. The presence of lipoproteins in whey causes an increase of fouling with substantial decrease in membrane performance (Maubois, Pierre, Fauquant, & Piot, 1987). Furthermore, Wang and Tang (2011) observed that NF membranes have low permeability of organic compounds, as noted in this present work by the high R protein value.

95

Table 1 Results of physicochemical composition of whey after thermocalcic precipitation (TP), retentates and permeates from nanofiltration (NF) and retentate from nanofiltration followed by diafiltration (NF/DF). Samples

Total solids (g 100 g−1)

Protein (g 100 g−1)

Whey after TP

c

5.851 ± 0.031

0.721 ± 0.010c

NF retentate VRF = 1.5 VRF = 2.0

7.920 ± 0.041b 8.702 ± 0.020a

0.950 ± 0.002b 1.050 ± 0.010a

NF permeate VRF = 1.5 VRF = 2.0

0.921 ± 0.001e 1.372 ± 0.001d

0.110 ± 0.001d 0.120 ± 0.010d

NF/DF retentate VRF = 2.0

5.850 ± 0.022c

0.720 ± 0.021c

VRF: volume reduction factor. Results expressed as mean ± standard deviation, among three batches realized in triplicate for each type of the ricotta cheese, with three repetitions for total solids and for protein contents. a–e Within a column, different superscript lowercase letters denote significant differences (P b 0.05) among the samples.

the retention of organic compounds, as observed for the protein contents of NF retentates and NF/DF retentates. Due to its higher protein content (P b 0.05), the NF retentate from VRF equal to 2.0 was chosen to produce ricotta 1, and also to obtain the NF/DF retentate and, therefore, ricotta 2. However, no differences (P N 0.05) were noted between the protein content and total solids of both ricotta 1 and ricotta 2 (Table 2). Moreover, these contents were also slightly smaller (P b 0.05) than in the ricotta control. The decrease of protein content and total solids of ricotta 1 and ricotta 2, when compared with their respective NF and NF/DF retentates, may be due to TP of whey. According to Rosenberg (1995) one of the main problems related to whey protein is the variations in their functionality caused by differences in whey composition, due to whey-processing conditions such as the removal of lipoprotein. Khalloufi, Alexander, Goff, and Corredig (2008) reported that the protein/lipid ratio is a factor that can directly affect the stability of protein and is responsible for interactions between both compounds, consequently resulting in a cheese with lower protein content and, therefore, with lower total solids. The lower protein content in ricotta 1 and ricotta 2 may also be attributed to denaturation accelerated by the action of the fluid shear observed during the NF and DF processes. According to Agrawal, Bund, and Pandi (2008), it is likely that the denaturation may be accelerated by the action of the fluid shear as some proteins show sensitivity to this condition. The effects of agitation rate on the protein as well as the particle size distribution are responsible for separation of proteins (Agrawal et al., 2008), and thus the decrease of its content in the samples of ricotta 1 and ricotta 2.

3.2. Physicochemical properties 3.3. Color measurements The NF retentates (VRF = 1.5 and 2.0) showed higher (P b 0.05) contents of total solids and protein than the NF permeates and whey after TP (Table 1), while the contents of NF/DF retentate (VRF = 2.0) were similar to those of whey after TP (P N 0.05). The high retention of compounds, such as the total solids and the proteins found in the NF retentates, could be due to diffusive and convective transports. Bracken et al. (2006) noted that convection is the dominant mechanism involved in solute transport during nanofiltration. However, these authors also affirm that the contribution of diffusion increases when the convective transport is hindered due to electrostatic repulsion or steric hindrance. Already, Bellona, Drewesa, Xua, and Amy (2004) reported that the retention of organic compounds represents a complex interaction of steric hindrance, electrostatic repulsion, effects of raw material on the membrane and membrane properties. In addition, the composition of the feed material and the membrane fouling may affect

According to Ramos et al. (2013), color is extremely important in food because it directly influences product appeal and consumer acceptability. In the present work it was possible to note that the L*, a* and b* values were lower (P b 0.05) for ricotta 1 and ricotta 2 (Table 2), when compared to ricotta control. Dattatreya and Rankin (2006) attributed the decrease of L* value to the browning due to oxidation, which is undesirable in cheese because it may influence on consumer acceptability. All the ricotta cheese samples showed a low a* value, indicating a tendency to color green. These results occurred probably due to the presence of riboflavin in whey, since it is attributable for whey's slightly green coloration, as stated by Mestdagh, Kerkaert, Cucu, and Meulenaer (2011). Meanwhile, parameter b* showed no differences (P N 0.05) between ricotta 1 and ricotta 2. This occurrence may be related with Maillard reaction, i.e., non-enzymatic browning, principally because

96

E.S. Prudêncio et al. / Food Research International 56 (2014) 92–99

Table 2 Results of physicochemical composition and color parameters (L*, a*, b*) of ricotta cheeses produced by whey (control), NF retentate (ricotta 1) and NF/DF retentate (ricotta 2). Samples

Total solids (g 100 g−1)

Protein (g 100 g−1)

Color parameters L*

Control Ricotta 1 Ricotta 2

a

a

14.92 ± 0.28 13.96 ± 0.53b 14.29 ± 0.10b

33.16 ± 0.21 32.12 ± 0.06b 31.99 ± 0.62b

a* a

b* a

15.42 ± 0.69a 13.92 ± 0.34b 13.62 ± 0.73b

−2.56 ± 0.10 −2.24 ± 0.07b −1.52 ± 0.15c

99.97 ± 0.00 92.59 ± 0.38c 98.43 ± 0.30b

NF retentate: retentate from nanofiltration process up to volume reduction factor (VRF) equal to 2. NF/DF retentate: retentate from nanofiltration followed by diafiltration (NF/DF) up to VRF equal to 2. Results expressed as mean ± standard deviation, among three batches realized in triplicate for each type of the ricotta cheese, with three repetitions for total solids and for protein contents; and ten repetitions for each one of rheological parameters. a–b Within a column, different superscript lowercase letters denote significant differences (P b 0.05) among the samples.

the manufacture of ricotta cheese employs high temperature. According to Dattatreya and Rankin (2006), high parameter b* value is probably due to the high concentration of yellow-colored compounds formed during the intermediate stage of Maillard reaction. These authors also reported that the drop in b*, as observed for ricotta 1 and ricotta 2 in this present work, may be attributed to the following stage of Maillard reaction, wherein brown compounds are formed, and thereby reducing the yellowness of ricotta 1 and ricotta 2. Finally, because of the high luminosity and yellowness values, it was possible to observe that ricotta control was less affected by manufacturing, followed by ricotta 2 and ricotta 1, respectively. 3.4. Rheological properties The rheological data of uniaxial compression, fracture stress (σf) and fracture strain (εf); as well as stress relaxation, “degree of solidity” (1 − c1), stress decay (c1/c2) and relaxation ratio (RRELAX); are shown in Table 3. The three ricotta cheeses manufactured showed differences (P b 0.05) between their σf values. This behavior is in accordance with that observed by Hinrichs (2001), who states that modifications of process technology, as the use of technology membrane, are accompanied by rheological changes in cheeses. Ricotta 1, produced from NF retentate, showed higher σf value (P b 0.05) than the other two cheeses, from ricotta control and ricotta 2, indicating typical behavior of a product with higher hardness, as noted by Fritzen-Freire, Müller, Laurindo, and Prudêncio (2010). This behavior may be because of TP and, therefore, there may have been a removal of lipoprotein from whey. Pereira et al. (2002) emphasized the importance of the potential functional properties, namely, the emulsifying capacity of the extracted lipoprotein. Therefore, the removal of lipoprotein may have contributed to the increase in hardness of ricotta 1. Despite the whey used in the manufacture of ricotta 2 also having been submitted to TP, the σf value was lower (P b 0.05). In this case, the

whey was submitted to two stages of membrane process, i.e., NF followed by DF and probably, according to Almécija, Ibáñez, Guadix, and Guadix (2007), the denaturation of protein and/or a new association between proteins may have occurred because some whey proteins are sensitive to the fluid shear from membrane process. Buffa, Trujillo, Pavia, and Guamis (2001) reported that denaturation and/or association between proteins may disturb the homogeneity of the network and result in a reduction of the hardness, as noted for ricotta 2. Fox, Guinee, Cogan, and McSweeney (2000) verified decrease in the hardness of cheese, through disruption of the protein matrix due to the release of small peptides and free amino acids. According to Buffa et al. (2001), εf describes the deformability of cheese, which may express cohesion properties. Therefore, it was possible to note that ricotta 1 showed the highest εf value (P b 0.05), while no differences (P N 0.05) were noted between ricotta 2 and ricotta control. These results indicate that ricotta 1 showed a greater deformability before cheese rupture. As reported by Ferrandini, López, Castillo, and Laencina (2011), the differences noted in deformability may be related to the chemical structures of the components inside the cheese, resulting from changes in the processing. Foegeding and Davis (2011) reported that processing steps, such as the concentration process, are capable of modifying intermolecular interactions of proteins. Cunha, Viotto, and Viotto (2006) reported that the strong protein intermolecular attractions confirm a higher hardness, which was verified in this present work for the σf value of ricotta 1. Furthermore, Fig. 2 shows the stress–strain curves of the ricotta cheeses, where it is possible to verify that ricotta 2 was the most fragile, followed by the ricotta control and ricotta 1, respectively. The three ricotta cheeses manufactured had no differences (P N 0.05) on the “degree of solidity” (1 − c1). According to the model proposed by Peleg (1980), when t → ∞, F(t)/F0 → 1 − c1. Therefore, if the stress of the specimen fully relaxes, c1 = 1 and 1 − c1 = 0. When 1 − c1 goes to 1 the material behaves as an elastic solid, and when it goes to 0 the 40.0

Table 3 Results of the rheological parameters of uniaxial compression and relaxation of the ricotta cheeses produced by whey (control), NF retentate (ricotta 1) and NF/DF retentate (ricotta 2).

Control Fracture stress (σf) (kPa) Fracture strain (εf) (−) Degree of solidity (1 − c1) (−) Initial stress decay rate (c1/c2) (s−1) Relaxation rate (RRATE) (%)

21.57 0.56 0.35 1.02 35

± ± ± ± ±

Ricotta 1 0.39b 0.09b 0.03a 0.09a 2a

38.97 0.69 0.37 0.64 37

± ± ± ± ±

Ricotta 2 2.32a 0.01a 0.04a 0.10b 4a

7.33 0.46 0.34 0.43 34

± ± ± ± ±

0.24c 0.04b 0.03a 0.33b 3a

NF retentate: retentate from nanofiltration process up to volume reduction factor (VRF) equal to 2. NF/DF retentate: retentate from nanofiltration followed by diafiltration (NF/DF) up to VRF equal to 2. Results expressed as mean ± standard deviation, among three batches realized in triplicate for each type of the ricotta cheese, with three repetitions for each one of the rheological parameters. a–c Within a row, different superscript lowercase letters denote significant differences (P b 0.05) among the samples.

30.0

Stress (kPa)

Samples

Ricotta 1

Control

20.0

10.0

0.0 0.0

Ricotta 2

0.2

0.4

0.6

0.8

Strain (-) Fig. 2. Stress–strain curves of the ricotta cheeses produced by whey (control); by NF retentate (volume reduction factor — VRF equal to 2) (ricotta 1); and by NF/DF retentate (VRF equal to 2) (ricotta 2).

E.S. Prudêncio et al. / Food Research International 56 (2014) 92–99

97

material behavior is controlled by its viscous component (Müller et al., 2009). Therefore, it was possible to verify from the data obtained in this present study that all the ricotta cheeses produced were characterized by a tendency to be more viscous than elastic. A similar behavior was observed by Fritzen-Freire et al. (2010) with Minas Frescal cheese. Fig. 3 shows the typical stress relaxation curves of the ricotta cheeses. It was possible to note that the force decay rate with time to a residual value (60 s) also confirmed that all the ricotta cheeses showed a viscoelastic behavior. This result is in accordance with Messens, Van Dewalle, Arevalo, Dewettinck, and Huyghebaert (2000), who state that, like most solid foods, cheese is viscoelastic in nature, meaning that it has both solid (elastic) and fluid (viscous) behavior. Fig. 3, also shows that ricotta 1 showed a F0 (t = 0) higher than ricotta control and ricotta 2, which is consistent with the higher σf value shown in the uniaxial compression. An accurate estimation of c2 is important to determinate the rate of stress decay (c1/c2). The value of c1/c2 for ricotta control was higher (P b 0.05) than ricotta 1 and ricotta 2. It was also possible to confirm that the use of retentate from whey after TP submitted to NF and NF/DF, respectively, contributed to the production of ricotta cheeses (1 and 2) with low mechanical resistance and, therefore, with less elasticity. The values of the relaxation rate (RRATE) for the three ricotta cheeses showed no differences (P N 0.05), indicating that they are consistent with the “degree of solidity” (1 − c1).

3.5. Microstructural properties Fig. 4(a, b, and c) shows the micrographs of the ricotta cheeses (control, ricotta 1 and ricotta 2). Differences in microstructure were noted between the ricotta cheeses, where the matrix of ricotta 1 showed a more compacted structure. Thus, the more compact matrix seen in Fig. 4b could explain and confirm the higher hardness and deformability noted in the rheological analysis of ricotta 1. According to Fachin and Viotto (2005) disturbing the homogeneity of the network of protein may change the characteristics of cheeses. Yorgun et al. (2008) also confirm that when different technologies of manufacture are used, the proteins in the curd cheese whey are de-solubilized, losing their structure and modifying their properties. Therefore, the uniaxial compression and stress relaxation tests showed textural differences between the cheeses, which probably derived from the microstructural characteristics. The close relationship between the microstructure and the rheology of cheeses is often highlighted and it was also noted by Fritzen-Freire et al. (2010) and Madadlou, Asl, Mousavi, and Farmani (2007).

2.6 2.4 2.2 2.0

Force (N)

1.8 1.6 1.4 1.2

Ricotta 1

Fig. 4. SEM micrographs of the ricotta cheeses produced by: (a) whey (control); (b) NF retentate up to volume reduction factor (VRF) equal to 2 (ricotta 1); and (c) NF/DF retentate up to VRF equal to 2 (ricotta 2).

1.0 0.8 0.6 0.4

Control

0.2 0.0

4. Conclusions

Ricotta 2

0

10

20

30

40

50

60

Time (s) Fig. 3. Experimental stress relaxation curves of the ricotta cheeses produced by whey (control); by NF retentate (volume reduction factor —VRF equal to 2) (ricotta 1); and by NF/DF retentate (VRF equal to 2) (ricotta 2).

Despite detecting the differences between the compositions of the retentates, no differences were noted between the protein and the total solids of the ricotta cheeses from nanofiltration (NF) and nanofiltration followed by diafiltration (NF/DF) processes. Because of its high luminosity and yellowness values, it was possible to note that

98

E.S. Prudêncio et al. / Food Research International 56 (2014) 92–99

ricotta control (not submitted to membrane processes) was less affected, followed by ricotta from NF/DF retentate and from NF retentate, respectively. The membrane processes employed influenced on the rheological and microstructural properties of the ricotta cheeses, making the ricotta from NF retentate more elastic, firmer, and more compact. However, all the ricotta cheeses investigated in this present work showed a higher tendency to viscosity than to elasticity. Finally, NF and NF/DF processes and the use of their retentates in the manufacture of ricotta cheeses offer an opportunity for an entirely new approach regarding the utilization of whey. Acknowledgments The authors gratefully thank the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Brazil, for its financial support; and also to the Universidade Federal de Santa Catarina (UFSC), especially to research supported by LCME-UFSC, such as scanning electron microscopy (SEM). References Agrawal, S. G., Bund, R. K., & Pandi, A.B. (2008). Effect of agitation on heat-induced deproteination process of buffalo milk whey. Journal of Food Engineering, 87, 398–404. Al-Malack, M. H., & Anderson, G. K. (1997). Crossflow microfiltration with dynamic membranes. Water Research, 31, 1969–1979. Almécija, M. C., Ibáñez, R., Guadix, A., & Guadix, E. M. (2007). Effect of pH on the fractionation of whey proteins with a ceramic ultrafiltration membrane. Journal of Membrane Science, 288, 28–35. Association of Official Analytical Chemists (AOAC) (2005). Official methods of analysis of the association analytical chemists (18th ed.). Maryland, DC: AOAC. Atra, R., Vatai, G., Bekassy-Molnar, E., & Balint, A. (2005). Investigation of ultra- and nanofiltration for utilization of whey protein and lactose. Journal of Food Engineering, 67, 325–332. Baldasso, C., Barros, T. C., & Tessaro, I. C. (2011). Concentration and purification of whey proteins by ultrafiltration. Desalination, 278, 381–386. Baumy, J. J., Gestin, L., Fauquant, J., Boyaval, E., & Maubois, J. L. (1990). Technologies de purification des phospholipids du lactoserum. Process, 1047, 29–33. Bellona, C., Drewesa, J. E., Xua, P., & Amy, G. (2004). Factors affecting the rejection of organic solutes during NF/RO treatment—a literature review. Water Research, 38, 2795–2809. Bracken, L., Bettens, B., Boussu, K., Van der Meeren, P., Cocquyt, J., Vermant, J., et al. (2006). Transport mechanisms of dissolved organic compounds in aqueous solution during nanofiltration. Journal of Membrane Science, 279, 311–319. Buffa, M. N., Trujillo, A. J., Pavia, M., & Guamis, B. (2001). Changes in textural, microstructural, and colour characteristics during ripening of cheeses made from raw, pasteurized or high-pressure-treated goats' milk. International Dairy Journal, 11, 927–934. Butylina, S., Luque, S., & Nyström, M. (2006). Fractionation of whey-derived peptides using a combination of ultrafiltration and nanofiltration. Journal of Membrane Science, 280, 418–426. Calzada, J. F., & Peleg, M. (1978). Mechanical interpretation of compressive stress–strain relationships on solids foods. Journal of Food Science, 41, 1087–1092. Cuartas-Uribe, B., Alcaina-Miranda, M. I., Soriano-Costa, E., Mendoza-Roca, J. A., Iborra-Clar, M. I., & Lora-Garcia, J. (2009). A study of the separation of lactose from whey ultrafiltration permeate using nanofiltration. Desalination, 241, 244–255. Cunha, C. R., Viotto, W. H., & Viotto, L. A. (2006). Use of low concentration factor ultrafiltration retentates in reduced fat “Minas Frescal” cheese manufacture: effect on composition, proteolysis, viscoelastic properties and sensory acceptance. International Dairy Journal, 16, 215–224. Dattatreya, A., & Rankin, S. A. (2006). Moderately acidic pH potentiates browning of sweet whey powder. International Dairy Journal, 16, 822–828. Del Nobile, M.A., Chillo, S., Falcone, P.M., Laverse, J., Pati, S., & Baiano, A. (2007). Textural changes of Canestrello Pugliese cheese measured during storage. Journal of Food Engineering, 83, 621–628. Di Pierro, P., Sorrentino, A., Mariniello, L., Giosafatto, C. V. L., & Porta, R. (2011). Chitosan/whey protein film as active coating to extend ricotta cheese shelf-life. LWT - Food Science and Technology, 44, 2324–2327. Ebersold, M. F., & Zydney, A. L. (2004). The effect of membrane properties on the separation of protein charge variants using UF. Journal of Membrane Science, 243, 379–388. Fachin, L., & Viotto, W. H. (2005). Effect of pH and heat treatment of cheese whey on solubility and emulsifying properties of whey protein concentrate produced by ultrafiltration. International Dairy Journal, 15, 325–332. Fauquant, J., Vieco, E., Brule, G., & Maubois, J. -L. (1985). Clarification des lactosérums doux par agrégation thermocalcique de la matière grasse résiduelle. Le Lait, 65, 1–20. Ferrandini, E., López, M. B., Castillo, M., & Laencina, J. (2011). Influence of an artisanal lamb rennet paste on proteolysis and textural properties of Murcia al Vino cheese. Food Chemistry, 124, 583–588. Fikar, M., Kovacs, Z., & Czermak, P. (2010). Dynamic optimization of batch diafiltration processes. Journal of Membrane Science, 355, 168–174.

Foegeding, E. A., & Davis, J. P. (2011). Food protein functionality: A comprehensive approach. Food Hydrocolloids, 25, 1853–1864. Fox, P. F., Guinee, T. P., Cogan, T. M., & McSweeney, P. L. H. (2000). Fundamentals of cheese science. Gaithersburg: Aspen (587 pp.). Fritzen-Freire, C. B., Müller, C. M.O., Laurindo, J. B., & Prudêncio, E. S. (2010). The influence of Bifidobacterium Bb-12 and lactic acid incorporation on the properties of Minas Frescal cheese. Journal of Food Engineering, 96, 621–627. Hinrichs, J. (2001). Incorporation of whey proteins in cheese. International Dairy Journal, 11, 495–503. Instituto Adolfo Lutz (IAL) (2005). Normas Analíticas do Instituto Adolfo Lutz (4th ed.). São Paulo, DC: IAL. Khalloufi, S., Alexander, M., Goff, H. D., & Corredig, M. (2008). Physicochemical properties of whey protein isolate stabilized oil-in-water emulsions when mixed with flaxseed gum at neutral pH. Food Research International, 41, 964–972. Laurindo, J. B., & Peleg, M. (2007). Mechanical measurements in puffed rice cakes. Journal of Texture Studies, 38, 619–634. Lipnizki, F., Boelsmand, J., & Madsen, R. F. (2002). Concepts of industrial-scale diafiltration systems. Desalination, 144, 179–184. Lobato-Calleros, C., Ramirez-Santiago, C., Osorio-Santiago, V. J., & Vernon-Carter, E. J. (2002). Microstructure and texture of Manchego cheese-light products made with canola oil, lipophilic, and hydrophilic emulsifiers. Journal of Texture Studies, 33, 165–182. Madadlou, A., Asl, A. K., Mousavi, M. E., & Farmani, J. (2007). The influence of brine concentration on chemical composition and texture of Iranian white cheese. Journal of Food Engineering, 81, 330–335. Maubois, J. L., Pierre, A., Fauquant, J., & Piot, M. (1987). Industrial fractionation of main whey proteins. IDF Bulletin, 212, 154–159. Messens, W., Van Dewalle, D., Arevalo, J., Dewettinck, K., & Huyghebaert, A. (2000). Rheological properties of high-pressure-treated Gouda cheese. International Dairy Journal, 10, 359–367. Mestdagh, F., Kerkaert, B., Cucu, T., & Meulenaer, B. (2011). Interaction between whey proteins and lipids during light-induced oxidation. Food Chemistry, 126, 1190–1197. Modler, H. W., & Emmons, D. B. (2001). The use of continuous ricotta processing to reduce ingredient cost in ‘further processed’ cheese products. International Dairy Journal, 11, 517–523. Morr, C. V., & Ha, E. Y. W. (1993). Whey protein concentrates and isolates: Processing and functional properties. Critical Reviews in Food Science and Nutrition, 33, 431–476. Mourouzidis-Mourouzis, S. A., & Karabelas, A. J. (2006). Whey protein fouling of microfiltration ceramic membranes—Pressure effects. Journal of Membrane Science, 282, 124–132. Müller, C. M.O., Laurindo, J. B., & Yamashita, F. (2009). Effect of cellulose fibers on the crystallinity and mechanical properties of starch-based films at different relative humidity values. Carbohydrate Polymers, 77, 293–299. Pan, K., Song, Q., Wang, L., & Cao, B. (2011). A study of demineralization of whey by nanofiltration membrane. Desalination, 267, 217–221. Paulen, R., Foley, G., Fikar, M., Kovács, Z., & Czermak, P. (2011). Minimizing the process time for ultrafiltration/diafiltration under gel polarization conditions. Journal of Membrane Science, 380, 148–154. Peleg, M. (1979). Characterization of stress relaxation curves of solid food. Journal of Food Science, 44, 277–280. Peleg, M. (1980). Linearization of relaxation and creep curves of solid biological materials. Journal of Rheology, 24, 451–463. Pereira, C. D., Diaz, O., & Cobos, A. (2002). Valorization of by-products from ovine cheese manufacture: Clarification by thermocalcic precipitation/microfiltration before ultrafiltration. International Dairy Journal, 12, 773–783. Pizzillo, M., Claps, S., Cifuni, G. F., Fedele, V., & Rubino, R. (2005). Effect of goat breed on the sensory, chemical and nutritional characteristics of ricotta cheese. Livestock Production Science, 94, 33–40. Ramos, Ó. L., Reinas, I., Silva, S. I., Fernandes, J., Cerqueira, M.A., Pereira, R. N., et al. (2013). Effect of whey protein purity and glycerol content upon physical properties of edible films manufactured therefrom. Food Hydrocolloids, 30, 110–122. Rinaldoni, A. N., Tarazaga, C. C., Campderrós, E., & Padilla, A. P. (2009). Assessing performance of skim milk ultrafiltration by using technical parameters. Journal of Food Engineering, 92, 226–232. Román, A., Wang, J., Csanádic, J., Hodúr, C., & Vatai, G. (2009). Partial demineralization and concentration of acid whey by nanofiltration combined with diafiltration. Desalination, 241, 288–295. Rosenberg, M. (1995). Current and future applications for membrane processes in the dairy industry. Trends in Food Science & Technology, 61, 12–19. Ruffin, E., Schmit, T., Lafitte, G., Dollat, J. -M., & Chambin, O. (2013). The impact of whey protein preheating on the properties of emulsion gel bead. Food Chemistry, http://dx.doi.org/10.1016/j.foodchem.2013.11.07. Sarkar, P., Ghosh, S., Dutta, S., Sen, D., & Bhattacharjee, C. (2009). Effect of different operating parameters on the recovery of proteins from casein whey using a rotating disc membrane ultrafiltration cell. Desalination, 249, 5–11. Smithers, G. W. (2008). Whey and whey proteins — From ‘gutter-to-gold’. International Dairy Journal, 18, 695–704. Souza, C. H. B., & Saad, S. M. I. (2009). Viability of Lactobacillus acidophilus La-5 added solely or in co-culture with a yoghurt starter culture and implications on physico-chemical and related properties of Minas fresh cheese during storage. LWT - Food Science and Technology, 42, 633–640. Suárez, E., Lobo, A., Alvarez, S., Riera, F. A., & Álvarez, R. (2009). Demineralization of whey and milk ultrafiltration permeate by means of nanofiltration. Desalination, 241, 272–280. Tsuru, T., Sudoh, T., Yoshioka, T., & Asaeda, M. (2001). Nanofiltration in non-aqueous solutions by porous silica–zirconia membranes. Journal of Membrane Science, 185, 253–261.

E.S. Prudêncio et al. / Food Research International 56 (2014) 92–99 Van Der Bruggen, B., Mänttäri, M., & Nyström, M. (2008). Drawbacks of applying nanofiltration and how to avoid them: A review. Separation and Purification Technology, 63, 251–263. Wang, Y. N., & Tang, C. Y. (2011). Protein fouling of nanofiltration, reverse osmosis, and ultrafiltration membranes — The role of hydrodynamic conditions, solution chemistry, and membrane properties. Journal of Membrane Science, 376, 275–282. Wang, L., Yang, G., Xing, W., & Xu, N. (2008). Mathematic model of the yield for diafiltration processes. Separation and Purification Technology, 59, 206–213.

99

Wium, H., & Qvist, K. B. (1997). Rheological properties of UF-feta cheese determined by uniaxial compression and dynamic testing. Journal of Texture Studies, 28, 435–454. Yin, N., Yang, G., Zhong, Z., & Xing, W. (2011). Separation of ammonium salts from coking wastewater with nanofiltration combined with diafiltration. Desalination, 268, 233–237. Yorgun, M. S., Balcioglu, A., & Saygin, O. (2008). Performance comparison of ultrafiltration, nanofiltration and reverse osmosis on whey treatment. Desalination, 229, 204–216.