2012 - DeBON - Storage Stability of Prebiotic Fermented Milk Obtained From Permeate Resulting of the...

Accepted Manuscript

Storage stability of prebiotic fermented milk obtained from permeate resulting of themicrofiltration process

Janaina Debon, Elane Schwinden Prudêncio, José Carlos Cunha Petrus, CarliseBeddin Fritzen-Freire, Carmen M.O. Müller, Renata D. de M. Castanho Amboni,Cleide Rosana Werneck Vieira

PII: S0023-6438(11)00429-4

DOI: 10.1016/j.lwt.2011.12.029

Reference: YFSTL 2941

To appear in: LWT - Food Science and Technology

Received Date: 25 July 2011

Revised Date: 22 December 2011

Accepted Date: 28 December 2011

Please cite this article as: Debon, J., Prudêncio, E.S., Cunha Petrus, C., Fritzen-Freire, C.B., Müller,C.M., de M. Castanho Amboni, R.D., Werneck Vieira, C.R., Storage stability of prebiotic fermented milkobtained from permeate resulting of the microfiltration process, LWT - Food Science and Technology(2012), doi: 10.1016/j.lwt.2011.12.029

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http://dx.doi.org/10.1016/j.lwt.2011.12.029

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Storage stability of prebiotic fermented milk obtained from permeate resulting of 1

the microfiltration process 2

3

Janaina Debona, Elane Schwinden Prudênciob*, José Carlos Cunha Petrusa, Carlise 4

Beddin Fritzen-Freireb, Carmen M. O. Müllera, Renata D. de M. Castanho Ambonib & 5

Cleide Rosana Werneck Vieirab 6

a Departament of Chemistry and Food Engineering, Technology Center, Federal 7

University of Santa Catarina, Trindade, 88040-970, Florianópolis, SC, Brazil. 8

b Department of Food Science and Technology, Center of Agricultural Science, Federal 9

University of Santa Catarina, Rod. Admar Gonzaga, 1346, Itacorubi, 88034-001, 10

Florianópolis, SC, Brazil 11

* Corresponding author. Tel: (55) 48 37215366: Fax: (55) (48) 37219943 12

E-mail: [email protected] 13

14

Abstract 15

The permeate, obtained from the best microfiltration process, was employed in the 16

preparation of fermented milks, without inulin (control) and with 5 g 100g-1 inulin 17

(prebiotic), stored at 5 ± 1 °C for 28 days. It could be verified that the storage period 18

and addition of inulin increased the total solids and carbohydrate contents, the caloric 19

value and the acidity, and decreased the pH. The addition of inulin resulted in a 20

fermented product with a lower syneresis index, and greater firmness and cohesiveness. 21

The inulin employed resulted in a product with a greater tendency toward a greenish 22

coloration. 23

Keywords: Microfiltration; permeate; fermented milk; prebiotic; inulin. 24

25

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1. Introduction 26

27

Currently, separation processes which employ membranes, such as 28

microfiltration (MF), are widely used and mainly by the dairy industry. Among the 29

various applications of MF, is the removal of bacteria (Pafylias, Cheryan, Mehaia, & 30

Saglam, 1996; Lawrence, Kentish, O’Connor, Barber, & Stevens, 2008). Microfiltration 31

produces a permeate, which is the liquid that passes through the membrane (Saboya & 32

Maubois, 2000) and can be used to prepare dairy products that have distinct properties 33

(Lawrence et al., 2008) and that are thus well accepted by the market (Saboya & 34

Mabouis, 2000). However, the composition and the quality of the permeate depend on 35

factors such as the operational parameters employed during the process, as well the pore 36

size of membrane. Pafylias et al. (1996) reported that larger pore of microfiltration 37

membranes of about 1.4 µm pore size, can achieve the right balance between rejection 38

of other milk components, such as the protein, lactose and ash component. 39

In comparison with the conventional thermal treatments of milk, MF has the 40

advantages of requiring lower temperatures, and thus maintaining the structure of 41

casein, which results in a raw material with fewer functional and nutritional 42

modifications (Lawrence et al., 2008). Dairy products, such as cheese and fermented 43

milk, have been successfully prepared by using microfiltration (Saboya & Maubois, 44

2000). However, few studies, as that by Debon, Prudêncio, and Petrus (2010), have used 45

a functional ingredient, such as inulin, to obtain previously microfiltered fermented 46

milk. 47

Inulin, which is a fiber that can be extracted from chicory root, for example, and 48

is classified as a prebiotic food ingredient, offers both nutritional and technological 49

benefits when added to the dairy products (Gibson & Fuller, 2000). Studies carried out 50

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by Gibson, Beatty, Wang, and Cummings (1995) showed that regular ingestion of inulin 51

is beneficial to the microbiota of the human intestine. From a technological point of 52

view, inulin can be used as a substitute for fat in milk products (Paseephol, Small, & 53

Sherkat, 2008; Debon et al., 2010) since it forms microcrystals in the presence of milk, 54

resulting in products with a fine and creamy texture (Kaur & Gupta, 2002). 55

The objective of this study was to use MF to replace conventional thermal 56

treatments of milk to obtain a permeate that is microbiologically and physico-57

chemically acceptable for the production of fermented milks with or without the 58

addition of inulin. The products were evaluated for their physico-chemical properties, 59

syneresis index, instrumental texture profile analysis, and color parameters during 60

storage for 28 days at 5 ± 1 °C. 61

62

2. Materials and methods 63

64

2.1. Materials 65

66

Raw skimmed milk, thermophilic milk culture (YC-X11 Yo Flex®, Chr. 67

Hansen) composed of Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus 68

salivarius ssp. Thermophilus, saccharose, and inulin (HP-Gel-Beneo®, Orafti, Oreye, 69

Belgium) were used in the production of fermented milk. All the reagents used were of 70

analytical grade (P.A.). 71

72

2.2. Microfiltration process 73

74

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The raw skimmed milk was microfiltered in a pilot unit using an organic 75

poly(imide) membrane (MF-1, PAM Membranas Seletivas, Rio de Janeiro, Brazil), 76

hollow fiber type, with an average pore size of 1.4 µm and 0.4 m2 of useful filtering 77

area. In this stage, five experiments were carried out using the following transmembrane 78

pressures and tangential velocities: 100 kPa and 0.8 m s-1, 300 kPa and 0.8 m s-1, 100 79

kPa and 1.4 m s-1, 300 kPa and 1.4 m s-1, and 200 kPa and 1.2 m s-1. The best 80

microfiltration process for the raw skimmed milk, i.e., with higher permeate flux (J) (L 81

h-1 m-2), volumetric reduction factor (VRF) and protein content (g 100g-1) VRF, was the 82

one selected for the production of the fermented milks. The temperature used in the 83

experiments was 45 ± 1 °C. All the experiments were carried out in duplicate. 84

During microfiltration, the J was calculated every five minutes according to Eq. 85

1. 86

87

(L h-1 m-2) (1) 88

89

where Vp is the permeate volume collected during the time interval t (h) and Ap (m2) is 90

the membrane surface area of permeation. 91

The volumetric reduction factor (VRF) was determined as follows (Eq. 2): 92

93

(2) 94

95

2.3. Elaboration of fermented milks 96

97

The fermented milks, which were manufactured from the permeate of the best 98

microfiltration process, were produced by using a methodology adapted from the one 99

tA

VJ

P

P

.=

( )( ) ( )Lfinalvolumeretentate

LvolumemilkinitialVRF=

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developed by Almeida, Bonassi, and Roça (2001). The permeate was used to produce 100

two fermented milks, denominated as control (without inulin) and prebiotic (with 5 g 101

100g-1 of inulin). The permeate samples with and without inulin, at 42 ± 1 °C, were both 102

added with 8 g 100g-1 saccharose and the thermophilic milk culture for the fermentation 103

stage. The fermentation was stopped when pH reached between 4.5 and 4.7, and the 104

fermented milks were cooled at 10 ± 1 °C, being gently stirred, and then conditioned in 105

plastic flasks and stored at 5 ± 1 °C. The physico-chemical composition, syneresis 106

index, instrumental texture profile analysis (TPA), and color parameters were 107

determined every 7 days for 28 days of storage at 5 ± 1 °C. All these evaluations were 108

carried out in triplicate. 109

110

2.4. Microbiological analysis 111

112

Samples of the raw skimmed milk and of the permeate were submitted to 113

mesophilic and psychrophilic bacteria counts, using the values of CFU (Colony 114

Forming Units) mL-1, following the methodology of APHA (2001). The mesophilic and 115

psychrophilic bacteria counts were carried out on the samples stored at 32 °C for 48 h 116

and at 7 ºC for 10 days, respectively. 117

118

2.5. Physico-chemical analysis 119

120

The raw skimmed milk, the permeate, and the fermented milks (control and 121

prebiotic) were analyzed for content of total solids (g 100g-1), through the drying of the 122

samples until reaching constant weight (method 925.23), total proteins (g 100g-1) by the 123

Kjeldahl method (N x 6.38) (method 991.20) and ash (g 100g-1) through a gravimetric 124

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method (method 945.46). The lipid content (g 100g-1) of the raw skimmed milk and of 125

the permeate were determined through the Gerber method, while the lipid content of the 126

fermented milks were determined according to the Monjonnier method (method 989.05) 127

(AOAC, 2005). The value for total carbohydrate was obtained by difference. The 128

caloric value for the fermented milks were calculated from the conversion factors, 129

which were 4.0 kcal g-1 for proteins, 4.0 kcal g-1 for carbohydrates and 9.0 kcal g-1 for 130

lipids. The acidity (% lactic acid) of the fermented milks was determined according to 131

the methodology described by IAL (2005). The measurements of pH were carried out 132

with a pH meter (MP220, Metler-Toledo, Greinfensee, Switzerland). 133

134

2.6. Percent recovered of the permeate components (% Rc) 135

136

The percent recovered of the permeate components (% Rc), which indicates how 137

much of a component passes through the membrane in a filtration process, was 138

calculated only for the permeate obtained with the selected variables of the MF process, 139

through Eq. 3. 140

141

(3) 142

143

2.7. Syneresis index 144

145

The syneresis indexes of the two fermented milk samples (control and prebiotic) 146

were determined according to the method proposed by Farnsworth, Li, Hendricks, and 147

Guo (2006), with modifications. The samples (15 g) were centrifuged at 350 g in a 148

refrigerated centrifuge (5 ± 1 °C) (Jaetzki K24, Jena, Germany) for 10 min. The 149

100% xmilktheincomponentaofkg

permeatetheincomponentaofkgRc

=

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supernatant was collected and weighed, and the syneresis index was calculated through 150

Eq. 4. 151

152

(4) 153

154

2.8. Instrumental texture profile analysis (TPA) 155

156

The textural properties of the fermented milks were measured using a 157

texturometer model TA-XT2 (Stable Micro System, Texture Expert, Surrey, UK), 158

operating with the Texture Expert software program. The double compression analysis 159

was carried out using a 25 mm-diameter acrylic probe (P25/L). The analysis was 160

performed in a 50 mL aluminum capsule with the samples at a temperature of 5 ± 1 °C. 161

The test velocity, the time, and the distance were equal at 2.0 mm.s-1; 5.0 s, and 5.0 mm, 162

respectively. From the TPA curve, the texture parameters obtained were firmness, 163

gumminess, and adhesiveness. Firmness is defined by peak force during the first 164

compression cycle, whereas gumminess is the product of the firmness and the 165

cohesiveness, and adhesiveness is the negative area under the curve obtained between 166

the cycles. 167

168

2.9. Color measurements 169

170

The measurements of the color parameters of the fermented milks were carried 171

out with a previously calibrated colorimeter Minolta Chroma Meter CR-400 (Minolta®, 172

Japan), adjusted to operate with a D65 illuminant and an observation angle of 10°. 173

Luminosity (L*), red color intensity (a*) and yellow color intensity (b*) were measured. 174

( ) ( ) 100(g)tSupernatan

% xgmilkFermented

Syneresis =

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175

2.10. Statistical Analysis 176

177

The data were expressed as means and standard deviation. One-way analyses of 178

variance (ANOVA) and Tuckey’s studentized range (5 g 100g-1 significance) were carried 179

out to test any significant differences between the results. The data were obtained using the 180

software STATISTICA version 6.0 (2001) (StatSoft Inc., Tulsa, OK, USA). 181

182

3. Results and discussion 183

184

3.1. Microfiltration 185

186

There were small variations in the behavior of the permeate flux (J) as a function of 187

time for all of the experiments, as shown in Fig. 1. A similar behavior was observed by 188

Beolchini, Veglio, and Barba (2004) and Akbache et al. (2009) during the first 30 189

minutes of microfiltration of skimmed sheep’s milk (with 0.1 g 100g-1 of lipids) using 190

tubular ceramic membrane, and in the ultrafiltration of whey, using a hollow fiber 191

membrane. According to Nóbrega, Borges, and Habert (2005), in the tangential flow 192

there usually is an initial drop in the J values, which later stabilize during the process. In 193

this present study, a positive variation in the J was noted, mainly according as the 194

increase in the transmembrane pressure. It was also possible observed that the J of 195

experiment 4 (300 kPa, 1.4 m s-1) was lower than experiment 2 (300 kPa, 0.8 m s-1). 196

According to Mourouzidis-Mourozidis and Karabelas (2006) this effect is more 197

pronounced at higher filtration pressure (300 kPa). These authors observed that 198

aggregates of measured diameter less than the mean pore size, can infiltrate the internal 199

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membrane structure, possibly blocking some passages, moreover we should also take 200

into consideration the aggregates formed. Thus, during filtration, the deposition of 201

casein micelles, the size of which is close to 200 nm (much higher than the size of other 202

skimmed milk constituents, smaller than 4 nm), could lead to the formation of a porous 203

layer becoming a host network for small components (Rabiller-Baudry, Gesan-Guiziou, 204

Roldan-Calbo, Beaulieu, & Michel, 2005). Mourouzidis-Mourozidis and Karabelas 205

(2006) related that aggregates higher than 1 µm can effectively block the pores of 206

membrane. It is reasonable to assume that protein aggregates can be firmly “packed” 207

inside membrane pores, also contributing to the decrease of protein content in the 208

permeate when compared with raw skimmed milk. Hwang, Hsu and Tung (2006) cited 209

that an increase in filtration pressure led to a decrease in cake porosity, but to an 210

increase in the mass and average filtration resistance of the cake. However, the mass 211

and porosity of the cake decreased, but the average filtration resistance increased with 212

the increase of the velocity, as observed in present work. 213

The results for the parameters of the five experiments employed in the MF are 214

shown in Table 1. The best microfiltration process for the raw skimmed milk was that 215

of Experiment 2, where a transmembrane pressure of 300 kPa and tangential velocity of 216

0.8 m s-1 were employed and whose VRF, J, and protein content were higher. According 217

to Nóbrega et al. (2005), separation processes with membranes that use a pressure 218

gradient as the driving force will result, within a certain range, in a J directly 219

proportional to the pressure gradient employed. Similar VRF values were detected by 220

Ozer, Robinson, Grandison, and Bell (1998) for cow’s milk. The J value of best the 221

microfiltration was higher than that obtained by Krstic, Teric, Caric, and Milanovic 222

(2002) (35 L h-1 m-2), who employed a 0.1 µm membrane and pressure of 100 kPa. The 223

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difference in these values may be related to the average membrane pore diameter and 224

the pressure employed. 225

226

3.2. Microbiological analysis 227

228

The raw milk and the permeate showed mesophilic counts of 4 log CFU mL-1 229

and 3 log CFU mL-1, respectively. The psychrophilic count for the raw milk was 3 log 230

CFU mL-1, whereas for the permeate it was 2 log CFU mL-1. Bansal, Al-Ali, Mercadé-231

Prieto, and Chen (2006) and Lawrence et al. (2008) verified that MF can be used in the 232

separation of microorganisms, and thus reduce the nutrient damage caused by high 233

temperature. According to Tomasula et al. (2011), in practical the use of MF has the 234

potential to serve as a nonthermal intervention to high-temperature/short-time 235

pasteurization to ensure the safety and quality of milk, because it showed efficient 236

removal of the native microorganisms, as a transmission of the casein proteins to 237

permeate. Comparing MF and pasteurization, Walkling-Ribeiro, Rodríguez-Gonzalez, 238

Jayaram and Griffiths (2011) observed a similar reduction of the native microorganism 239

in milk. Pafylias et al. (1996) and Saboya and Maubois (2000) stated that the bacterial 240

decrease in the MF of milk is from 4 to 5 log CFU mL-1, such decrease could be even 241

lower, according to the findings of Trouvé et al. (1991), because of a lower bacterial 242

contamination initially detected in the raw milk. 243

The bacterial counts (mesophilic and psychrophilic) in the permeate show that it 244

can be employed in the production of a prebiotic fermented milk, since Angelidis et al. 245

(2006) noted that a count above 6 log CFU mL-1 is indicative of utilization of low-246

quality raw material. 247

248

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3.3. Physico-chemical analysis 249

250

The MF of the milk resulted in a permeate with lower contents of total solids 251

(TS) (p < 0.05), proteins, and lipids (Table 2), when compared with the raw skimmed 252

milk. Thus, the reduction in the TS content could be related with the reduction in the 253

lipid and protein contents. 254

The result obtained for the % Rc of the total proteins, approximately 96 %, is in 255

agreement with that of the study carried out by Mourouzidis-Mourouzis and Karabelas 256

(2006) on whey. Such authors used a tubular ceramic MF membrane, with a pore 257

diameter of 0.8 µm, where the percent permeability of the proteins was greater than 96 258

%. These observations are also in agreement with those in previous studies carried out 259

by Guell and Davis (1996) in the tangential MF process, where the retention of proteins 260

was below 15 %. In this present study, the % Rc obtained at the end of the MF process 261

showed that 73 % of the lipids initially contained in the raw skimmed milk were 262

transferred to the permeate. 263

Through the physico-chemical analysis of the permeate it is possible to note that 264

the pH and acidity values (Table 2) were not affected by the MF. The physico-chemical 265

composition of the permeate was similar to that of the skimmed milk used by Sivieri 266

and Oliveira (2002) in the preparation of lactic beverage with fat substitutes, where the 267

pH value was 6.65 and the acidity was 0.197 % lactic acid, whereas the lipid and TS 268

contents were 0.40 g 100g-1 and 8.56 g 100g-1, respectively. The pH and acidity values 269

were also in agreement with those obtained for the pasteurized milks used by Sivieri 270

and Oliveira (2002) (pH = 6.68 and acidity = 0.193 % lactic acid) and by Cunha, 271

Castro, Barreto, Benedet, and Prudêncio (2008) (pH = 6.69 and acidity = 0.173 % lactic 272

acid) in the preparation of fermented milks. 273

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Table 3 shows the results of the physico-chemical composition for the two 274

fermented milks (control and prebiotic) during the 28 days of storage. As expected, the 275

addition of inulin increased the TS content, reflecting also in an increase in the total 276

carbohydrates and, consequently, in the caloric value (p < 0.05). During storage the 277

control and prebiotic fermented milk showed an increase (p < 0.05) in the TS content 278

due to moisture loss, which can be attributed to the form of storage employed, i.e., the 279

use of plastic packaging without a perfect sealing system. For the prebiotic fermented 280

milk, the TS values were similar to those found by Penna, Sivieri, and Oliveira (2001) 281

(19.01 g 100g-1 to 21.71 g 100g-1) for commercial lactic beverage. 282

The storage period did not influence on (p > 0.05) the protein or lipid contents, 283

neither in the control nor in the prebiotic fermented milk. When the protein and lipid 284

contents of the control and prebiotic fermented milks are compared, it is possible to note 285

(Table 3) that there were no differences (p < 0.05) on days 1 and 28 of storage. 286

The storage period and the addition of inulin did not influence on the ash 287

contents of the fermented milks evaluated (p > 0.05). Moreover, the addition of a 288

prebiotic did not change the ash content in a study carried out by Thamer and Penna 289

(2006) (0.61 g 100g-1), Castro, Cunha, Barreto, Amboni, and Prudêncio (2008) (0.66 g 290

100g-1) and Cunha et al. (2008) (0.65 g 100g-1) in lactic beverages. 291

The fermented milks prepared in this present study showed a decrease (p < 0.05) 292

in the pH values on days 1 and 21 for the control and on days 1 and 28 for the prebiotic 293

(Fig. 2). When evaluated on the same day of storage, the pH values for the fermented 294

milks were not different (p > 0.05). A similar behavior was noted by Fuchs, Tanamati, 295

Antonioli, Gasparello, and Doneda (2006) in yogurt containing 5 g 100g-1 oligofructose 296

and 1 g 100g-1 inulin, where the pH values remained around 4.33 and 4.20 on days 1 297

and 28 of storage, respectively. 298

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For the control fermented milk, the difference in acidity (p < 0.05) was verified 299

from day 7 on, while for the prebiotic fermented milk the increase (p < 0.05) was 300

verified only from day 21 on. However, the acidity remained constant (p > 0.05) until 301

day 28 of storage (Fig. 3). According to Kailasapathy (2006), the decrease in the pH 302

values can occur due to the post-acidification, which is detected in fermented milks 303

stored at temperatures between 0 °C and 5 °C. 304

The acidity values were higher (p < 0.05) for the prebiotic fermented milk when 305

compared with the control, on days 1 and 7 of storage. However, these values were 306

higher than those found by Fuchs et al. (2006) (1.76 %) for probiotic yogurts 307

supplemented with prebiotics. 308

309

3.4. Syneresis index 310

311

The syneresis index of fermented milks (control and prebiotic) increased (p < 312

0.05) on day 7, however, it remained constant (p > 0.05) until day 28 of storage (Fig. 4). 313

As expected, the use of inulin in the preparation of the prebiotic fermented milk 314

contributed to the obtainment of a product with a lower syneresis index (p < 0.05). This 315

decrease may be related to TS present in the prebiotic fermented milk because, 316

according to Lucey (2001), a greater content of TS leads to a lower syneresis index. 317

Neven (2001) and Kaur and Gupta (2002) stated that the main use of inulin in fermented 318

milks can be as a substitute for fat. 319

However, Gauche, Tomazi, Barreto, Ogliari, and Bordignon-Luiz (2009) noted a 320

lower syneresis (22.93 %) index for yogurts, prepared from pasteurized milk, than those 321

verified for the fermented milks produced in this present study. It is important to note 322

that this difference may be associated with the thermal treatment initially applied to the 323

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milk. These fermented milks (control and prebiotic) were produced from the permeate 324

without undergoing any thermal treatment. According to Varnan and Sutherland (1994) 325

thermal treatments, as the pasteurization employed by Gauche et al. (2009), result in the 326

association of whey proteins with caseins, thus generating a more stable gel with a 327

decrease in syneresis. 328

329

3.5. Instrumental texture profile analysis (TPA) 330

331

No alterations in the firmness, gumminess, or adhesiveness were observed in the 332

control and prebiotic fermented milks during the storage period (Table 4). Also, it was 333

possible to verify that the use of inulin contributed to the obtainment of a prebiotic 334

fermented milk with greater firmness (p < 0.05) than that of the control product. Castro 335

et al. (2008) also verified that the addition of oligofructose at the proportion of 336

5 g 100g-1 in fermented lactic beverages increased the firmness of the product. 337

Inulin has also contributed to prebiotic fermented milks with higher values (p < 338

0.05) for adhesiveness on days 1 and 7 of storage. El-Nagar, Clowes, Tudoricã, and 339

Kuri (2002) noted an increase in adhesiveness when inulin was used in frozen yogurt 340

and these authors attributed this increase to a higher gel viscosity resulting from this 341

process. This statement is in agreement with that of Neven (2001), who defined inulin 342

as a substitute for fat in milk derivatives, contributing to the increase in adhesiveness. 343

344

3.6. Color measurements 345

346

Table 5 shows the parameters L*, a*, and b* for the fermented milks (control 347

and prebiotic) during storage period. On the same day of storage, it was possible to note 348

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that for the fermented milks only the parameter a* differed (p < 0.05), whereas the 349

parameters L* and b* remained practically unaltered (p > 0.05). The values obtained for 350

the parameter a* indicate that the control fermented milk tends towards a slightly more 351

greenish color than the prebiotic fermented milk, i.e., with the addition of inulin, a 352

reducing sugar. The decrease in the greenish color and the increase in the reddish color 353

may be attributed to the inulin employed associated with the lower moisture content and 354

fermentation temperature. According to Dattatreya and Rankin (2006), factors as the 355

reducing carbohydrates content and the temperature may contribute to maillard reaction, 356

resulting in change in color of dairy products. The results obtained in this present study 357

are in accordance with those reported by Castro et al. (2008), where the 358

supplementation of fermented milk with a prebiotic of white color did not affect the 359

parameters L* and b*. 360

Moreover, the storage of the fermented milks for 28 days did not influence (p > 361

0.05) on the parameters L* and a*. However, after 14 days of storage it was possible to 362

verify an increase (p < 0.05) in the yellow coloration (b* values) of the prebiotic 363

fermented milk. This occurrence may be related to the addition of the prebiotic, possibly 364

attributable for the maillard reaction. According to Dattatreya and Rankin (2006), the 365

increase in the parameter b* is related to the intermediate phase of the maillard reaction, 366

where a greater production of yellow compounds occurs, thus confirming the instability 367

of the b* values obtained. In relation to the parameters L* and a* during storage, a 368

similar behavior was observed by Dello Staffolo, Bertola, Martino and Bevilacqua 369

(2004) in yogurts added with inulin and stored for 21 days. Therefore, the use of 370

microfiltration as a replacement for the thermal treatment of pasteurization in the 371

preparation of the fermented milks (control and prebiotic) may be attributable for the 372

lower values obtained to parameters a* and b*. 373

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374

4. Conclusion 375

376

The best microfiltration process was when a pressure of 300 kPa and a velocity 377

of 0.8 m s-1 were used, resulting in an average permeate flux of 41.27 L h-1 m-2, a 378

volumetric reduction factor of 4.24, and a protein content of 2.59 g 100g-1. The 379

permeate showed a reduction in the mesophilic and psychrophilic bacterial counts and 380

in total solids, protein, and lipid contents when compared with the raw skimmed milk. 381

Neither the storage period (28 days) nor addition of inulin affected the lipid, protein, or 382

ash contents of the prebiotic fermented milk. As happened with the control fermented 383

milk, the storage period of 28 days also led to an increase in the total solids, 384

carbohydrate content, caloric value, and of acidity, which consequently decreased the 385

pH of the prebiotic fermented milk. 386

The addition of inulin resulted in a fermented milk with a lower syneresis index 387

and greater firmness and cohesiveness. There were no modifications in firmness, 388

gumminess, or cohesiveness during the 28 days of storage. The addition of inulin 389

caused the fermented milk to show a lower tendency towards a green coloration, 390

whereas the storage period of the prebiotic fermented milk showed a greater tendency 391

towards a yellow coloration. 392

Finally, the results showed that microfiltration can be considered as a good 393

alternative to obtain a permeate from microfiltration with appropriate characteristics to 394

be employed in obtaining of prebiotic fermented milk with storage stability. 395

396

Acknowledgements 397

The authors wish to thank to Beneo® Orafti and Victoria Alimentos Ltda. 398

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- Storage stability of prebiotic fermented milk obtained from permeate.

- Evaluation of the permeate obtained from the microfiltration process.

- Preparation of prebiotic fermented milk obtained from permeate.

- Evaluation of fermented milks without and with the addition of inulin.

- The use of microfiltration to replace conventional thermal milk treatments.

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Table 1 521

Results of permeate flux (J), volumetric reduction factor (VRF) and protein content of 522

the permeate during the microfiltration process with different pressure and tangential 523

velocity 524

EXPERIMENT J (L h-1 m-2) VRF PROTEIN CONTENT (g 100g-1)

1 16.79 ± 0.01 2.26 ± 0.01 1.68 ± 0.01

2 41.27 ± 0.01 4.24 ± 0.01 2.59 ± 0.01

3 15.93 ± 0.01 2.55 ± 0.01 1.67 ± 0.01

4 29.41 ± 0.01 2.58 ± 0.01 2.46 ± 0.00

5 24.72 ± 0.02 3.41 ± 0.01 2.10 ± 0.00

Results expressed as mean ± standard deviation (n = 2) 525

Experiment 1: Pressure = 1 bar and tangential velocity = 0.8 m s-1, during 30 minutes ; experiment 2 = 3 526

bar and 0.8 m s-1, during 25 minutes; experiment 3: 1 bar and 1.4 m s-1, during 25 minutes; experiment 4: 527

3 bar and 1.4 m s-1, during 25 minutes; experiment 5: 2 bar and 1.2 m s-1, during 30 minutes 528

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Table 2 541

Physico-chemical composition of the raw skimmed milk and of the permeate obtained 542

of microfiltration process with pressure of 3 bar and tangential velocity of 0.8 m s-1 543

RAW SKIMMED MILK PERMEATE

Proteins (g 100g-1) 3.27a ± 0.01 3.13b ± 0.01

Lipids (g 100g-1) 0.55a ± 0.07 0.40b ± 0.01

Total solids (TS) (g 100g-1) 8.80a ± 0.02 8.41b ± 0.03

Ash (g 100g-1) 0.74a ± 0.01 0.72a ± 0.01

Carbohydrates (g 100g-1) 4.23a ± 0.04 4.15a ± 0.01

pH 6.66a ± 0.00 6.67a ± 0.00

Acidity (% lactic acid) 0.195a ± 0.007 0.185a ± 0.007


a,b Within a line, different superscript lowercase letters denote significant differences (p < 0.05) between 545

the samples (raw skimmed milk and permeate) 546

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Table 3 559 Composition of the control (without inulin) and prebiotic fermented milk (with inulin) during 28 days of storage at 5 ± 1 °C 560

FERMENTED

MILK

DAY

TOTAL SOLIDS

(g 100g-1)

PROTEINS

(g 100g-1)

LIPIDS

(g 100g-1)

ASH

(g 100g-1)

CARBOHYDRATES

(g 100g-1)

CALORIC VALUE

(kcal g-1)

1 14.50Aa ± 0.03 2.78Aa ± 0.02 0.34Aa ± 0.03 0.63Aa ± 0.03 10.75Aa ± 0.04 57.44Aa ± 0.23

7 14.69Ab ± 0.03 2.82Aa ± 0.02 0.35Aa ± 0.02 0.63Aa ± 0.01 10.89Ab ± 0.03 57.99Aa ± 0.18

14 14.71Ab ± 0.04 2.84Aa ± 0.02 0.36Aa ± 0.02 0.62Aa ± 0.04 10.89Ab ± 0.03 58.16Aa ± 0.01

21 15.20Ac ± 0.04 2.84Aa ± 0.03 0.37Aa ± 0.03 0.65Aa ± 0.03 11.34Ac ± 0.02 60.05Ab ± 0.48

CONTROL

28 15.22Ac ± 0.03 2.85Aa ± 0.03 0.37Aa ± 0.02 0.66Aa ± 0.04 11.34Ac ± 0.04 60.09Ab ± 0.18

1 18.39Ba ± 0.08 2.72Aa ± 0.03 0.30Aa ± 0.03 0.66Aa ± 0.03 14.71Ba ± 0.03 72.42Ba ± 0.03

7 18.49Ba ± 0.05 2.72Ba ± 0.03 0.33Aa ± 0.01 0.68Aa ± 0.03 14.76Ba ± 0.04 72.89Ba ± 0.54

14 18.47Ba ± 0.03 2.69Ba ± 0.03 0.33Aa ± 0.04 0.67Aa ± 0.04 14.78Ba ± 0.04 72.85Ba ± 0.03

21 19.01Bb ± 0.06 2.74Aa ± 0.03 0.34Aa ± 0.03 0.67Aa ± 0.02 15.26Bb ± 0.04 75.06Bb ± 0.19

PREBIOTIC

28 19.04Bb ± 0.04 2.75Aa ± 0.01 0.35Aa ± 0.01 0.69Aa ± 0.01 15.25Bb ± 0.03 75.15Bb ± 0.07


A,B,C Within a column, different superscript uppercase letters denote significant differences (p < 0.05) among the different fermented milk (without and with inulin) for the 562

same storage period 563

a,b,c Within a column, different superscript lowercase letters denote significant differences (p < 0.05) among the different storage day, for each sample 564

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Table 4 565

Results of the instrumental texture profile (TPA) analysis of the control (without inulin) 566

and prebiotic fermented milk (with inulin) during 28 days of storage at 5 ± 1 °C 567

FERMENTED

MILK DAY FIRMNESS (g) GUMMINESS (g) ADHESIVENESS (g.s)

1 27.13Aa ± 1.71 9.92Aab ± 0.37 - 27.05Aa ± 2.29

7 30.10Ab ± 0.62 10.40Ab ± 0.39 - 26.99Aa ± 1.20

14 24.40Aa ± 0.85 8.92Aa ± 0.69 - 41.87Ab ± 8.26

21 26.50Aa ± 0.69 9.87Aab ± 0.27 - 31.84Aab ± 2.43

CONTROL

28 25.75Aa ± 0.07 9.21Aab ± 0.42 - 38.56Aab ± 8.57

1 36.10Ba ± 1.30 10.83Aa ± 0.47 - 48.52Ba ± 1.77

7 35.20Bba ± 1.84 10.05Aa ± 1.44 - 43.81Bba ± 5.26

14 31.50Bb ± 1.27 10.29Aa ± 0.10 - 45.60Aba ± 6.32

21 34.65Bba ± 1.48 10.29Aa ± 0.51 - 36.24Ab ± 1.61

PREBIOTIC

28 32.87Bba ± 1.23 10.16Aa ± 0.82 - 42.91Aba ± 1.41


A,B,C Within a column, different superscript uppercase letters denote significant differences (p < 0.05) 569

among the different fermented milk (without and with inulin) for the same storage period 570

a,b,c Within a column, different superscript lowercase letters denote significant differences (p < 0.05) 571

among the different storage day, for each sample 572

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Table 5 578

Color parameters L*, a* and b* of the control (without inluin) and prebiotic fermented 579

milk (with inulin) samples during 28 days of storage at 5 ± 1 °C 580

L* a* b*

DAY CONTROL PREBIOTIC CONTROL PREBIOTIC CONTROL PREBIOTIC

1 48.59Aa±1.86 50.46Aa±2.26 -2.44Aa±0.11 -2.12Ba±0.02 6.12Aa±0.14 5.97Aa±0.11

7 50.12Aa±0.51 51.09Aa±1.29 -2.43Aa±0.05 -2.11Ba±0.03 6.20Aa±0.02 6.11Aa±0.02

14 49.07Aa±0.16 49.82Aa±0.48 -2.45Aa±0.01 -2.08Ba±0.03 6.32Aa±0.09 6.24Ab±0.13

21 49.35Aa±1.36 49.51Aa±1.24 -2.50Aa±0.02 -2.10Ba±0.01 6.09Aa±0.06 6.25Bb±0.04

28 50.09Aa±0.92 50.57Aa±0.48 -2.36Aa±0.03 -2.02Ba±0.02 6.29Aa±0.06 6.28Ab±0.04


A,B Within a line, for each color parameter evaluated, different superscript uppercase letters denote 582

significant differences (p < 0.05) among the different fermented milk (without and with inulin) for the 583

same storage period 584

a,b Within a column, different superscript lowercase letters denote significant differences (p < 0.05) 585

among the different storage day, for each sample 586

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Fig. 1. Average permeate flux (J) of milk submitted to microfiltration in different 605

process conditions. P = 100 kPa, v = 0.8 m s-1; P = 100kPa, v = 1.4 m s-1; 606

P = 200 kPa, v = 1.2 m s-1; P = 300 kPa, v = 0.8 m s-1; P = 300 kPa, v = 1.4 m 607

s-1. 608

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Fig. 2. Results of the average(standard deviation) pH during 28 days of storage (5 ± 1 633

°C), of fermented milks (control and prebiotic). Control fermented milk; 634

Prebiotic fermented milk. 635

A,B Different superscript uppercase letters denote significant differences (p < 0.05) among the different 636

storage day, for each sample 637

a,b Different superscript lowercase letters denote significant differences (p < 0.05) among the different 638

fermented milk (without and with inulin) for the same storage period 639

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Fig. 3. Results of the average acidity(standard deviation) (% lactic acid), during 28 days 660

of storage (5 ± 1 °C), of fermented milks (control and prebiotic). 661

Control fermented milk; Prebiotic fermented milk. 662





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Fig. 4. Results of the average ± standard deviation ( ) syneresis index of fermented 687

milks (control and prebiotic), during 28 days of storage at 5 ± 1 °C. Control fermented 688

Milk; Prebiotic fermented milk. 689





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