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Relationship between somatic cell counts and the properties of yoghurt made from ewes’ milk
ARTICLE IN PRESS
International Dairy Journal 16 (2006) 262–267 www.elsevier.com/locate/idairyj
Relationship between somatic cell counts and the properties of yoghurt made from ewes’ milk A.M. Vivar-Quintanaa,, E. Beneitez De La Mano, I. Revillaa a
Area of food Technology Area, Universidad de Salamanca, E.P.S. de Zamora, Campus Viriato. Avda Requejo, 33. 49022 Zamora, Spain b ASOVINO Cooperative Association, Avda Tres cruces, 35. 49008 Zamora, Spain Received 12 February 2004; accepted 22 March 2005
Abstract Mastitis is one of the most serious diseases that can affect ewes. Owing to the infection of the mammary gland, its synthetic functions decrease and this is accompanied by a deterioration of the physiological barrier to the blood. This may affect the composition of milk and hence the products derived from it. In the present work, yoghurts were made from ewes’ milk with three different somatic cell count (SCC), and the composition of the starting milk and the characteristics of the final product were analysed. High SCC affected the pH of the milk and its lactose content, and changes pH and acidity during the yoghurt fermentation process. The yoghurts made with high counts had an unsuitable texture and sensory analyses revealed that they were rejected by consumers. r 2005 Elsevier Ltd. All rights reserved. Keywords: Somatic cell count; Yoghurt; Ewes’ milk
1. Introduction Currently, yoghurt is an important milk product in the western world. In simple terms, it can be described as milk fermented by the growth of two thermophilic lactic bacteria: Streptococcus thermophilus and Lactobacillus delbrueckii subspecies bulgaricus (Hartley, Duong, David, & Fazel, 1989). Ewes’ milk is widely used in different regions of the word for the production of yoghurt. Sheep milk is especially suitable for yoghurt production because of its high content in proteins and solids (Haenlein, 1998). In general, the overall properties of yoghurt—such as its level of acidity, free fatty acid content, the production of aromatic compounds, as well as its sensory profile and nutritional value—are important characteristics of the product. These aspects are affected by the chemical
Corresponding author. Tel.: +34 980545000; fax: +34 980545002.
E-mail address: [email protected] (A.M. Vivar-Quintana). 0958-6946/$ - see front matter r 2005 Elsevier Ltd. All rights reserved. doi:10.1016/j.idairyj.2005.03.006
composition of the milk base, the processing conditions employed, the flavour added, and the activity of the starter culture during the incubation period (Bonczar, Wszolek, & Siuta, 2002). All milk contains some level of somatic cells (SC). Neutrophils comprise the major cell type in milk from uninfected sheep (Paape, Poutrel, Contreras, Marco, & Capucol, 2001). When there is bacterial infection, tissue damage, or other inflammation processes affecting the mammary tissue the number of SCs in milk increases dramatically (Jaeggi et al., 2003). This increase in SC count results from a transfer of white blood cells from the blood into the milk (Pirisi et al., 2000). In addition, the relative proportions of different types of SCs present in milk change significantly. Mastitis induces changes in milk that influence dairy product quality (Barbano, Verdi, & Rasmussen, 1987). In the dairy industries of many countries, somatic cell counts (SCCs) are widely used as monitors of udder health and milk quality. Maximum levels in milk are held as standards by health officials to ensure quality
ARTICLE IN PRESS A.M. Vivar-Quintana et al. / International Dairy Journal 16 (2006) 262–267
milk supplies from dairy farmers to the consumers of milk and dairy products (Harmon, 1994). The criteria for hygienic and bacteriological quality in sheep milk are outlined in the European Community (CE) directives 92/46 and 94/71, which regulate the various aspects of the production of milk of different species of mammals. The level of SCs in ewes’ milk fluctuates considerably. In particular, it is high in colostrum, although it is also affected by many other factors such as age, animal husbandry practices used, climate, and the health status of the animals’ udders (Duling, Paape, Schultze, & Weinland, 1983; Fruganti, Ranicci, Tesei, & Valente, 1985). In the case of sheep milk, the limits of SC counts have not yet been definitely established, although it has been suggested that a threshold level for subclinical mastitis in sheep should be close to 1,500 000 cell mL1 (Boyazoglu & Morand-Fehr, 2001), much higher than that set for cows (o500,000 cell mL1). The effect of SCC in sheep milk on yoghurt has not been studied. The few studies addressing the effects of SCC in sheep milk on its suitability for cheese manufacture have been inconclusive. This is partly because of the incidence of sub-clinical and clinical infections in ovine udders is less than in bovine udders (Boyazoglu & Morand-Fehr, 2001). The aim of the present work was to establish the effect of SC counts on the rheological and sensory properties of yoghurt made from ewes’ milk.
2. Materials and methods 2.1. Raw material Milk (morning milking) was collected three times during the winter period in January 2002, from ewes which were a cross-breed of the Assaf with the Churra and Castellana breeds. Milk with three SC counts was collected: o500,000, 1,000,000–1,500,000, and 43,000,000 cells mL1. The milk was from herds on three different farms with identical husbandry practices and feeding regimes.
263
2.3. Manufacture of yoghurts A total of 270 yoghurts were manufactured. Ninety yoghurts were made for each of the SCC (milk was collected three times for each SCC, and each time 30 yoghurts were made). Milks were standardised (20% total solids) by the addition of skim milk powder. The standardised milks were pasteurised at 65 1C for 30 min and cooled to 37 1C. The pasteurised milk was inoculated with a commercial yoghurt culture comprised of Lactobacillus delbrueckii subsp bulgaricus and Streptococcus thermophilus (MY087, Laboratorios Arroyo, Santander, Spain). Aliquots of inoculated milk (125 mL) were poured into plastic containers, which were covered with lids and incubated at 45 1C until the pH had decreased to about 4.5. The containers were then transferred to forced-air storage at 4 1C to cool the product and to terminate acid development. Some of the samples were analysed after 72 h, while the remaining samples were stored cool (4 1C) for 28 days before examination. 2.4. Chemical analyses of yoghurt Analyses were performed after mechanical stirring (magnetic stirrer, 10 min) of the yoghurts at 20 1C. The yoghurts were analysed for pH (pH-meter, Crison Basic 20), titratable acidity using the Dornic method (after mixing 10 g of yoghurt with an equal mass of distilled water (Katsiari, Voutsinas, & Kondyli, 2002), and total solids (IDF, 1991)). All analyses were carried out on nine yoghurts for each of the SCCs studied. 2.5. Syneresis of yoghurt Syneresis was defined as the volume of serum that was not retained within the structure on centrifugation. Tubes containing 30 g of yoghurt were centrifuged for 10 min at 6 1C at 680 g (Gonzalez Andrada, Romero Estevez, & Jime´nez-Pe´rez, 1994). The amount of serum released from the coagulum was measured in a calibrated measuring flask (Harwalkar &Kalab, 1983). 2.6. Consistency of the yoghurt
2.2. Chemical analyses of the milk Milks were analysed for fat (Gerber method, British Standards Institution (BSI), 1995), pH (pH-meter, Crison Basic 20), titratable acidity (Dornic method, Tamine & Robinson, 1999), lactose (IDF, 1974), protein (IDF, 1986), total solids (IDF, 1987). On the same day as they were collected, milk samples were submitted for Fossomatic SCC analyses at the certified laboratory of the Junta de Castilla y Leo´n (Spain) (LILCYL; Palencia, Spain). All analyses were carried out in triplicate.
The consistency (firmness) of the unbroken yoghurt coagulum was measured in samples (4 1C) using a Universal TA-XT2i Texture Analyser. The operating conditions were as follows. A 50 mm diameter probe was inserted into each product to a depth of 6 mm at a speed of 1 mm s1. The diameter of yoghurt cup 70 cm, the load cell was 25 Kg, and the temperature of samples during measurement 4 1C. Eighteen yoghurts was analysed for each SCC studied. The resulting force–time curve were analysed using texture profile analysis (Bourne, 1978). Hardness and
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adhesiveness were evaluated. Hardeness is defined as the maximun peak force during the first compression cycle and adhesiveness is defined as the negative force area for the first bite and represents the work required to overcome the attractive forces between the surface of yoghurt and the surface of probe with wich the yoghurt comes into contact. 2.7. Sensory grading of the product Sensory analyses were performed on 72 h-old yoghurt samples. The products were assessed by six judges using a grading scale of 1–9. The expert panel comprised nonsmokers who were very familiar with dairy products. The judges were instructed to classify the samples according to their overall acceptability, and reasons for rejection or acceptance were requested. 2.8. Statistical analysis Three replicate trials were undertaken on three separate occasions during January 2002. In each trial, three different yoghurts (treatement) were made from milks with different SC counts (o500,000; 1,000,000– 1,500,000; 43,000,000 cells mL1. The data on the chemical analyses and textural attributes were analysed by one-way analysis of variance (ANOVA). The statistical significance of each factor considered was calculated at the po0:05 level using the F-test. The LSD Fisher-test was employed to test for statistically significant differences between samples. All statistical analyses were carried out using the Statgraphic 2.0 program (Statistical Graphics Corp. Rockville, USA).
3. Results and discussion 3.1. Milk composition The analyses of the three milks used to manufacture the yoghurts are shown in Table 1. These compositional data are similar to those reported by Alichanidis and Polychroniadon (1995) and Haenlein (1995). The results show that SCC had a significant effect (po0:05) on the pH of milk, which increased with SCC. Acidity (mL 0.1 N NaOH mL1) was slightly lower for the milk with high SCC, although no statistically significant differences were observed between the values for the milks with different SCC. Similarly, Muelas, Molina, Diaz, and Peris (1996) found that the production of lactic acid during fermentation was low in mastitic milk, resulting in yoghurts with low levels of lactic acid and high pH. However, owing to the buffering effect of milk this did not lead to significantly lower acidity values in the current study.
Table 1 Compositional parameters of fresh ewes’ milk used in the manufacture of yoghurta Somatic cell count of milk ( 1000 mL1) o500
1000–1500 a
pH 6.670.08 Acidityb 23.371.09a Total solids (%, w/w) 18.971.98a Lactose (%, w/w) 4.970.09a Fat (%, w/w) 7.571.08a Protein (%, w/w) 6.470.27a
43000 b
6.770.03 23.371.26a 19.170.87a 4.370.07b 7.670.92a 5.970.63a
6.870.02c 22.271.09a 17.870.19a 4.170.26c 7.470.69a 5.970.67a
a
Presented values are the means7standard deviation of three replicate trials. Values in the same row with different superscripted letters differ significantly (po0:05). b mL 0.1 N NaOH mL1 milk.
Lactose concentration also decreased significantly (po0.05) with the SCC. As a result of the damage occurring to epithelial tissue in mastitis, the synthesis capacity the mammary gland decreases and this is reflected in a reduction in the lactose content of milk (Pirisi, Piredda, Podda, & Piutus, 1996). This decrease in lactose levels could also be due to the fact that the blood flow to the udder is reduced when SCC increases and, as a result, the mammary gland has a lower amount of glucose (the main precursor in lactose synthesis) available (Martı´ de Olives & Molina Pons, 1998). SCC did not significantly affect the fat content of the milk. The effect of ovine mastitis on fatty material is controversial and some authors have supported the generalised notion that mastitis gives rise to a slight reduction in the percentage of fat in the milk (Jaeggi et al., 2003) while others hold the opposite view (Pirisi et al., 1996; Dı´ az, Muelas, Segura, Peris, & Molina, 1996). The increase in fat level reported in some studies may be due to the fact that the drop in milk production is much more pronounced than the reduction in the level of fat synthesised (Martı´ de Olives & Molina Pons, 1998). SCC did not significantly influence protein content. However, numerically higher protein values were observed for the milk with low SCC and slightly lower levels were observed in milk with high SCC. During mastitis, the proteins synthesised by the udder decrease, whereas proteins coming from the bloodstream increase. These increases and decreases may compensate each other, and hence the total protein remains unaffected (Pirisi et al., 2000). The dry matter content of milk was not affected statistically by SCC. 3.2. Yoghurt fermentation The changes in milk pH and acidity during incubation of the yoghurt are shown in Fig. 1. The rapid increase in
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6.5
Table 2 Properties of 72 h-old yoghurt made from standardised ewes’ milk with different somatic cell countsa
6.0
Yoghurt properties
pH
7.0
1000–1500
43000
Physical and chemical data 4.270.07a 4.370.05a pH 4.370.08a Acidityb 139.072.27a 138.173.56a 134.072.4a Total solids (%, w/w) 21.771.22a 21.270.97a 21.370.39a Syneresis (mL) 1.071.13a 2.870.98a 7.270.07b
4.5 4.0 0
30
(a)
60 90 120 150 Incubation Time (min)
180
210
120 Acidity (mL 0.1N NaOH mL-1)
Somatic cell of milk ( 1000 mL1) o500
5.5 5.0
Rheological Hardness (N) Adhesiveness (N s) Sensory gradingc Sensory evaluation (score)
100
4.2972.12a 4.4172.05a 3.6672.08b 4.1272.81a 4.1572.82a 5.7072.61b 7.770.51a
7.270.98a
4.270.41b
a
Presented values are the means7standard deviation of three replicate trials. Values in the same row with different superscripted letters differ significantly (po0:05). b mL 0.1 N NaOH mL1 milk. c Details of test are given in the text.
80 60 40 20 0 0
(b)
265
30
60 90 120 150 Incubation Time (min)
180
210
Fig. 1. Effect of somatic cell count in ewes’ milk on time-related changes in pH (a) and titratable acidity (b) on incubation with starter culture during yoghurt manufacture. Somatic cell count: cells mL1: o500,000 (’); 1,000,000–1,500,000 (m); 43,000,000 (J).
acid production is due to the logarithmic phase of starter growth, which tapers off as the level of acidity becomes limiting (Shaker, Jumah, & Abu-Jdayil, 2000). The change in pH during the incubation of the milks was similar in the milks with low and medium counts, and about 210 min was required to reach a pH of 4.5, after which fermentation ceased. The pH decreased much faster in the milk with the highest SCC, fermentation ceasing at 150 min, despite the initial pH of the milk being higher. The fermentation of milk with high SCC may be inhibited owing to the inhibition of the starter cultures as a result of the antimicrobial action of SCs (Heras, Dominguez, & Fernandez-Garayzabal, 1999). However, our results would suggest the opposite. The changes in pH and acidity in milk with high SCC were different from those of the milk with low SCC. 3.3. Properties of 72 h old yoghurt The SCC of the milk did not significantly influence the pH or acidity of the yoghurt at 72 h. However, the SCC significantly affected the levels of syneresis. The milk with the highest SCC had the highest level of syneresis, pointing to the poor consistency of the coagulum of this milk and its inability to retain serum. The milk from
animals with mastitis had a high level of serum proteins, and low level of casein (Pirisi et al., 1996; Martı´ de Olives & Molina Pons, 1998). For this reason, milk with high SCC would give rise to soft coagula with a reduced ability to retain serum (Tamine & Robinson, 1999). The rheological characteristics of the yoghurts are shown in Table 2. Yoghurts manufactured from milk with low- or medium-SCC did not statistically differ from each, but differed significantly from those manufactured from milk with a SCC43,000,000 cells mL1. Yoghurt from high-SCC milk was softer and more adhesive than yoghurt from milk with lower SCC. According to Tamine and Robinson (1999), one of the factors that can affect the firmness and adhesiveness of yoghurt is the amount of lactose present in the milk. Milk with SCC43,000,000 cells mL1 had the lowest lactose content, which may have affected the texture of the yoghurt. Similarly, the results of Urech, Puhan, and Scha¨llibaum (1999) showed that milk with high SCC had a relatively low level of casein, and a high level of serum proteins. The SCC of the milk significantly influenced the grading score of yoghurts (Table 2). The high-SCC milk gave yoghurt which received the lowest grading score. The yoghurts manufactured from milk with high SCC were described by the tasters as having a piquant and bitter taste. Owing to the damage caused to the mammary tissue by mastitis, blood plasma leaks into the milk (Barbano et al., 1987) and such plasma contains many enzymes, such as proteases and lipases. These enzymes hydrolyse protein and fat, causing losses in yield (Barbano et al., 1987) and, possibly, result in the development of undesirable flavours and textures in the dairy products manufactured from this milk.
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266
4.9 4.7
pH
4.5 4.3 4.1 3.9 3.7 3.5 3
6
9
(a)
12 15 18 21 Storage time (days)
24
28
Acidity (mL 0.1N NaOH mL-1)
180
Increasing the SCC of the milk from o500,000 to 1,000,000–1,500,000 cell mL1 did not significantly influence the texture, syneresis or grading quality of the resultant yoghurt. However, SCCs43,000,000 cell mL1 resulted in yoghurt that had a very soft consistency and a reduced ability to retain serum, and that was characterised by flavours described as piquant and bitter. SCC resulted in a marked non-significant decreased in milk protein level, a significant decrease in yoghurt hardness, and significant increases in the adhesiveness and syneresis of yoghurt. From these results, it seems that the SCC in ewes’ milk destined for the manufacture of yoghurts should not exceed 1,500,000 cell mL1.
160 140 120
Acknowledgements
100 80
Thanks are due to the ASOVINO Cooperative Association for funding this project.
60 40 20 0 3
(b)
6
9
15 12 18 21 Storage time (days)
24
28
Fig. 2. Effect of somatic cell count in ewes’ milk on changes in pH (a) and titratable acidity (b) on the resultant yoghurt. Somatic cell count: cells mL1: o500,000 (’); 1,000,000–1,500,000 (m); 43,000,000 (J).
3.4. Changes in yoghurt during storage The pH and titratable acidity values of the yoghurt stored at 4 1C for 28 days are shown in Fig. 2. The pH values of all the yoghurts decreased and acidity values increased with storage time. This can be attributed to acid production during cold storage (post-acidification) as a result of the conversion of lactose to lactic acid by the bacterial cultures (Katsiari et al., 2002). There were no significant (po0:05) differences in either pH or acidity between the yoghurts from the milk with low- or medium-SCCs at storage times 415 days. However, in the case of milk with high SCC, the pH values at storage time 415 days were significantly lower than those of the others milks. The SCC in the milk did not significantly influence the acidity of the yoghurts during storage.
4. Conclusions As the SCC of ewes’ milk increased, the pH increased and the lactose concentration decreased. The titratable acidity and the contents of fat, protein, and total solids were not significantly influenced by the SCC of the milk.
References Alichanidis, E., & Polychroniadon, A. (1995). Special features of dairy products from ewe and goat milk from the physiochemical and organoleptic point of view. IDF Greek National Committee of IDF CIRVAL Seminar, Creta, Greece, October 21– 29, 1995. (pp. 122–127). Barbano, D.M., Verdi, R.J., & Rasmussen, R. (1987). Influence of milk somatic cell count on cheese manufacturing and cheese yield. 24th Annual Marschall invitational Italian Cheese Seminar, Wisconsin, USA, September 16– 17, 1987. (pp.12–16). Bonczar, G., Wszolek, M., & Siuta, A. (2002). The effects of certain factors on the properties of yoghurt made from ewe’s milk. Food Chemistry, 79, 85–91. Bourne, M. C. (1978). Texture profile analysis. Food Technology, 32, 62.66. Boyazoglu, J., & Morand-Fehr, P. (2001). Mediterranean dairy sheep and goat products and their quality. A critical review. Small Rumiant Research, 40, 1–11. British Standards Institution (BSI). (1995). Gerber method for the determination of fat in milk and milk products. British Standard 696. London: British Standards Institution. Dı´ az, J. R., Muelas, R., Segura, C., Peris, C., & Molina, P. (1996). Effect of mastitis on milk composition in Manchega ewes: Preliminary results. In Somatic cell and milk of small ruminants (pp. 240–246). Wageningen, The Netherlands: Pers Wageningen. Duling, A. M., Paape, M. J., Schultze, W. D., & Weinland, B. T. (1983). Effect of parity, stage of lactation and intramammary infection on concentration of somatic cells and cytoplasmic particles in goat milk. Journal of Dairy Science, 66, 2426–2433. Fruganti, G., Ranicci, S., Tesei, B., & Valente, C. (1985). Valutazione dello stato sanitario della mamella di pecora durante un intero ciclo di lattazione. Clinica Veterinaria, 108, 286–288. Gonzalez Andrada, J. I., Romero Estevez, C., & Jime´nez-Pe´rez, S. (1994). Control de calidad en la fabricacio´n de yoghurt. Alimentacio´n, Equipos y Tecnologı´a, 7, 77–81. Haenlein, G. F. W. (1995). Topics for success with dairy goats. In Dairy Extension Bulletin (pp. 105–112). Delaware, USA: University of Delaware.
ARTICLE IN PRESS A.M. Vivar-Quintana et al. / International Dairy Journal 16 (2006) 262–267 Haenlein, G. F. W. (1998). The value of goats and sheep to sustain mountain farmers. International Journal of Animal Science, 13, 187–194. Harmon, R. J. (1994). Physiology of mastitis and factors affecting somatic cell counts. Journal of Dairy Science, 77, 2103–2112. Hartley, D., Duong, C., David, P., & Fazel, A. (1989). Symbiosis of yogurt microorganisms. In Les laits fermente´s (pp. 139–145). Montrouge, France: John Libbery Eurotext. Harwalkar, V. R., & Kalab, M. (1983). Susceptibility of yoghurt to syneresis. Comparison of centrifugation and drainage methods. Milchwissenschaft, 38, 517–522. Heras, A., Dominguez, L., & Ferna´ndez-Garayzabal, J. F. (1999). Prevalence and aetiology of subclinical mastitis in dairy ewes of the Madrid Region. Small Ruminant Research, 32, 21–29. IDF. (1974). Determination of the lactose content of milk. IDF Standard 28 A. Brussels, Belgium: International Dairy Federation. IDF. (1986). Milk. Determination of nitrogen content (Kjeldahl method) and calculation of crude protein content. IDF Standard 20 A. Brussels, Belgium: International Dairy Federation. IDF. (1987). Milk, cream and evaporated milk. Determination of total solids content. IDF Standard 21B. Brussels, Belgium: International Dairy Federation. IDF. (1991). Yoghurt. Determination of total solids content. IDF Standard 151. Brussels, Belgium: International Diary Federation. Jaeggi, J. J., Govindasamy-Lucey, S., Berger, Y. M., Johnson, M. E., McKusick, B. C., Thomas, D. L., & Wendorff, W. L. (2003). Hard ewe0 s milk cheese manufactured from milk of three different groups of somatic cell counts. Journal of Dairy Science, 86, 3082–3089.
267
Katsiari, M. C., Voutsinas, L. P., & Kondyli, E. (2002). Manufacture of yoghurt from stored frozen sheep’s milk. Food Chemistry, 77, 413–420. Martı´ de Olives, A., & Molina Pons, P. (1998). Mamitis y calidad de leche de oveja. Ovis, 59, 11–25. Muelas, R., Molina, P., Diaz, J. R., & Peris, C. (1996). Relationship between somatic cell count, acidity and California Mastitis test in Manchega ewe milk. In Somatic cells and milk of small Ruminants (pp. 213–219). Wageningen. The Netherlands: Pers Wagening. Paape, M. J., Poutrel, B., Contreras, A., Marco, J. C., & Capucol, A. V. (2001). Milk somatic cells and lactation in Small Ruminants. Journal of Dairy Science, 84, 236–244. Pirisi, A., Pireda, G., Corona, M., Pes, M., Pinutus, S., & Ledda, A. (2000). Efecto de las ce´lulas soma´ticas sobre la composicio´n de la leche de oveja y las caracterı´ sticas del producto elaborado. Ovis, 66, 49–58. Pirisi, A., Piredda, G., Podda, F., & Piutus, S. (1996). Effect of somatic cell count on sheep milk composition and cheese making properties. In Somatic cell and milk of small ruminants (pp. 187–192). Wageningen, The Netherlands: Pers Wageningen. Shaker, R. R., Jumah, R. Y., & Abu-Jdayil, B. (2000). Rheological properties of plain yoghurt during coagulation process: Impact of fat content and preheat treatment of milk. Journal of Food Engineering, 44, 175–180. Tamine, A. Y., & Robinson, R. K. (1999). Yoghurt science and technology. Cambridge, UK: Woodhead Publishing Limited. Urech, E., Puhan, Z., & Scha¨llibaum (1999). Changes in milk protein fraction as affected by subclinical mastitis. Journal Dairy Science, 82, 2402–2411.
International Dairy Journal 16 (2006) 262–267 www.elsevier.com/locate/idairyj
Relationship between somatic cell counts and the properties of yoghurt made from ewes’ milk A.M. Vivar-Quintanaa,, E. Beneitez De La Mano, I. Revillaa a
Area of food Technology Area, Universidad de Salamanca, E.P.S. de Zamora, Campus Viriato. Avda Requejo, 33. 49022 Zamora, Spain b ASOVINO Cooperative Association, Avda Tres cruces, 35. 49008 Zamora, Spain Received 12 February 2004; accepted 22 March 2005
Abstract Mastitis is one of the most serious diseases that can affect ewes. Owing to the infection of the mammary gland, its synthetic functions decrease and this is accompanied by a deterioration of the physiological barrier to the blood. This may affect the composition of milk and hence the products derived from it. In the present work, yoghurts were made from ewes’ milk with three different somatic cell count (SCC), and the composition of the starting milk and the characteristics of the final product were analysed. High SCC affected the pH of the milk and its lactose content, and changes pH and acidity during the yoghurt fermentation process. The yoghurts made with high counts had an unsuitable texture and sensory analyses revealed that they were rejected by consumers. r 2005 Elsevier Ltd. All rights reserved. Keywords: Somatic cell count; Yoghurt; Ewes’ milk
1. Introduction Currently, yoghurt is an important milk product in the western world. In simple terms, it can be described as milk fermented by the growth of two thermophilic lactic bacteria: Streptococcus thermophilus and Lactobacillus delbrueckii subspecies bulgaricus (Hartley, Duong, David, & Fazel, 1989). Ewes’ milk is widely used in different regions of the word for the production of yoghurt. Sheep milk is especially suitable for yoghurt production because of its high content in proteins and solids (Haenlein, 1998). In general, the overall properties of yoghurt—such as its level of acidity, free fatty acid content, the production of aromatic compounds, as well as its sensory profile and nutritional value—are important characteristics of the product. These aspects are affected by the chemical
Corresponding author. Tel.: +34 980545000; fax: +34 980545002.
E-mail address: [email protected] (A.M. Vivar-Quintana). 0958-6946/$ - see front matter r 2005 Elsevier Ltd. All rights reserved. doi:10.1016/j.idairyj.2005.03.006
composition of the milk base, the processing conditions employed, the flavour added, and the activity of the starter culture during the incubation period (Bonczar, Wszolek, & Siuta, 2002). All milk contains some level of somatic cells (SC). Neutrophils comprise the major cell type in milk from uninfected sheep (Paape, Poutrel, Contreras, Marco, & Capucol, 2001). When there is bacterial infection, tissue damage, or other inflammation processes affecting the mammary tissue the number of SCs in milk increases dramatically (Jaeggi et al., 2003). This increase in SC count results from a transfer of white blood cells from the blood into the milk (Pirisi et al., 2000). In addition, the relative proportions of different types of SCs present in milk change significantly. Mastitis induces changes in milk that influence dairy product quality (Barbano, Verdi, & Rasmussen, 1987). In the dairy industries of many countries, somatic cell counts (SCCs) are widely used as monitors of udder health and milk quality. Maximum levels in milk are held as standards by health officials to ensure quality
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milk supplies from dairy farmers to the consumers of milk and dairy products (Harmon, 1994). The criteria for hygienic and bacteriological quality in sheep milk are outlined in the European Community (CE) directives 92/46 and 94/71, which regulate the various aspects of the production of milk of different species of mammals. The level of SCs in ewes’ milk fluctuates considerably. In particular, it is high in colostrum, although it is also affected by many other factors such as age, animal husbandry practices used, climate, and the health status of the animals’ udders (Duling, Paape, Schultze, & Weinland, 1983; Fruganti, Ranicci, Tesei, & Valente, 1985). In the case of sheep milk, the limits of SC counts have not yet been definitely established, although it has been suggested that a threshold level for subclinical mastitis in sheep should be close to 1,500 000 cell mL1 (Boyazoglu & Morand-Fehr, 2001), much higher than that set for cows (o500,000 cell mL1). The effect of SCC in sheep milk on yoghurt has not been studied. The few studies addressing the effects of SCC in sheep milk on its suitability for cheese manufacture have been inconclusive. This is partly because of the incidence of sub-clinical and clinical infections in ovine udders is less than in bovine udders (Boyazoglu & Morand-Fehr, 2001). The aim of the present work was to establish the effect of SC counts on the rheological and sensory properties of yoghurt made from ewes’ milk.
2. Materials and methods 2.1. Raw material Milk (morning milking) was collected three times during the winter period in January 2002, from ewes which were a cross-breed of the Assaf with the Churra and Castellana breeds. Milk with three SC counts was collected: o500,000, 1,000,000–1,500,000, and 43,000,000 cells mL1. The milk was from herds on three different farms with identical husbandry practices and feeding regimes.
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2.3. Manufacture of yoghurts A total of 270 yoghurts were manufactured. Ninety yoghurts were made for each of the SCC (milk was collected three times for each SCC, and each time 30 yoghurts were made). Milks were standardised (20% total solids) by the addition of skim milk powder. The standardised milks were pasteurised at 65 1C for 30 min and cooled to 37 1C. The pasteurised milk was inoculated with a commercial yoghurt culture comprised of Lactobacillus delbrueckii subsp bulgaricus and Streptococcus thermophilus (MY087, Laboratorios Arroyo, Santander, Spain). Aliquots of inoculated milk (125 mL) were poured into plastic containers, which were covered with lids and incubated at 45 1C until the pH had decreased to about 4.5. The containers were then transferred to forced-air storage at 4 1C to cool the product and to terminate acid development. Some of the samples were analysed after 72 h, while the remaining samples were stored cool (4 1C) for 28 days before examination. 2.4. Chemical analyses of yoghurt Analyses were performed after mechanical stirring (magnetic stirrer, 10 min) of the yoghurts at 20 1C. The yoghurts were analysed for pH (pH-meter, Crison Basic 20), titratable acidity using the Dornic method (after mixing 10 g of yoghurt with an equal mass of distilled water (Katsiari, Voutsinas, & Kondyli, 2002), and total solids (IDF, 1991)). All analyses were carried out on nine yoghurts for each of the SCCs studied. 2.5. Syneresis of yoghurt Syneresis was defined as the volume of serum that was not retained within the structure on centrifugation. Tubes containing 30 g of yoghurt were centrifuged for 10 min at 6 1C at 680 g (Gonzalez Andrada, Romero Estevez, & Jime´nez-Pe´rez, 1994). The amount of serum released from the coagulum was measured in a calibrated measuring flask (Harwalkar &Kalab, 1983). 2.6. Consistency of the yoghurt
2.2. Chemical analyses of the milk Milks were analysed for fat (Gerber method, British Standards Institution (BSI), 1995), pH (pH-meter, Crison Basic 20), titratable acidity (Dornic method, Tamine & Robinson, 1999), lactose (IDF, 1974), protein (IDF, 1986), total solids (IDF, 1987). On the same day as they were collected, milk samples were submitted for Fossomatic SCC analyses at the certified laboratory of the Junta de Castilla y Leo´n (Spain) (LILCYL; Palencia, Spain). All analyses were carried out in triplicate.
The consistency (firmness) of the unbroken yoghurt coagulum was measured in samples (4 1C) using a Universal TA-XT2i Texture Analyser. The operating conditions were as follows. A 50 mm diameter probe was inserted into each product to a depth of 6 mm at a speed of 1 mm s1. The diameter of yoghurt cup 70 cm, the load cell was 25 Kg, and the temperature of samples during measurement 4 1C. Eighteen yoghurts was analysed for each SCC studied. The resulting force–time curve were analysed using texture profile analysis (Bourne, 1978). Hardness and
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adhesiveness were evaluated. Hardeness is defined as the maximun peak force during the first compression cycle and adhesiveness is defined as the negative force area for the first bite and represents the work required to overcome the attractive forces between the surface of yoghurt and the surface of probe with wich the yoghurt comes into contact. 2.7. Sensory grading of the product Sensory analyses were performed on 72 h-old yoghurt samples. The products were assessed by six judges using a grading scale of 1–9. The expert panel comprised nonsmokers who were very familiar with dairy products. The judges were instructed to classify the samples according to their overall acceptability, and reasons for rejection or acceptance were requested. 2.8. Statistical analysis Three replicate trials were undertaken on three separate occasions during January 2002. In each trial, three different yoghurts (treatement) were made from milks with different SC counts (o500,000; 1,000,000– 1,500,000; 43,000,000 cells mL1. The data on the chemical analyses and textural attributes were analysed by one-way analysis of variance (ANOVA). The statistical significance of each factor considered was calculated at the po0:05 level using the F-test. The LSD Fisher-test was employed to test for statistically significant differences between samples. All statistical analyses were carried out using the Statgraphic 2.0 program (Statistical Graphics Corp. Rockville, USA).
3. Results and discussion 3.1. Milk composition The analyses of the three milks used to manufacture the yoghurts are shown in Table 1. These compositional data are similar to those reported by Alichanidis and Polychroniadon (1995) and Haenlein (1995). The results show that SCC had a significant effect (po0:05) on the pH of milk, which increased with SCC. Acidity (mL 0.1 N NaOH mL1) was slightly lower for the milk with high SCC, although no statistically significant differences were observed between the values for the milks with different SCC. Similarly, Muelas, Molina, Diaz, and Peris (1996) found that the production of lactic acid during fermentation was low in mastitic milk, resulting in yoghurts with low levels of lactic acid and high pH. However, owing to the buffering effect of milk this did not lead to significantly lower acidity values in the current study.
Table 1 Compositional parameters of fresh ewes’ milk used in the manufacture of yoghurta Somatic cell count of milk ( 1000 mL1) o500
1000–1500 a
pH 6.670.08 Acidityb 23.371.09a Total solids (%, w/w) 18.971.98a Lactose (%, w/w) 4.970.09a Fat (%, w/w) 7.571.08a Protein (%, w/w) 6.470.27a
43000 b
6.770.03 23.371.26a 19.170.87a 4.370.07b 7.670.92a 5.970.63a
6.870.02c 22.271.09a 17.870.19a 4.170.26c 7.470.69a 5.970.67a
a
Presented values are the means7standard deviation of three replicate trials. Values in the same row with different superscripted letters differ significantly (po0:05). b mL 0.1 N NaOH mL1 milk.
Lactose concentration also decreased significantly (po0.05) with the SCC. As a result of the damage occurring to epithelial tissue in mastitis, the synthesis capacity the mammary gland decreases and this is reflected in a reduction in the lactose content of milk (Pirisi, Piredda, Podda, & Piutus, 1996). This decrease in lactose levels could also be due to the fact that the blood flow to the udder is reduced when SCC increases and, as a result, the mammary gland has a lower amount of glucose (the main precursor in lactose synthesis) available (Martı´ de Olives & Molina Pons, 1998). SCC did not significantly affect the fat content of the milk. The effect of ovine mastitis on fatty material is controversial and some authors have supported the generalised notion that mastitis gives rise to a slight reduction in the percentage of fat in the milk (Jaeggi et al., 2003) while others hold the opposite view (Pirisi et al., 1996; Dı´ az, Muelas, Segura, Peris, & Molina, 1996). The increase in fat level reported in some studies may be due to the fact that the drop in milk production is much more pronounced than the reduction in the level of fat synthesised (Martı´ de Olives & Molina Pons, 1998). SCC did not significantly influence protein content. However, numerically higher protein values were observed for the milk with low SCC and slightly lower levels were observed in milk with high SCC. During mastitis, the proteins synthesised by the udder decrease, whereas proteins coming from the bloodstream increase. These increases and decreases may compensate each other, and hence the total protein remains unaffected (Pirisi et al., 2000). The dry matter content of milk was not affected statistically by SCC. 3.2. Yoghurt fermentation The changes in milk pH and acidity during incubation of the yoghurt are shown in Fig. 1. The rapid increase in
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6.5
Table 2 Properties of 72 h-old yoghurt made from standardised ewes’ milk with different somatic cell countsa
6.0
Yoghurt properties
pH
7.0
1000–1500
43000
Physical and chemical data 4.270.07a 4.370.05a pH 4.370.08a Acidityb 139.072.27a 138.173.56a 134.072.4a Total solids (%, w/w) 21.771.22a 21.270.97a 21.370.39a Syneresis (mL) 1.071.13a 2.870.98a 7.270.07b
4.5 4.0 0
30
(a)
60 90 120 150 Incubation Time (min)
180
210
120 Acidity (mL 0.1N NaOH mL-1)
Somatic cell of milk ( 1000 mL1) o500
5.5 5.0
Rheological Hardness (N) Adhesiveness (N s) Sensory gradingc Sensory evaluation (score)
100
4.2972.12a 4.4172.05a 3.6672.08b 4.1272.81a 4.1572.82a 5.7072.61b 7.770.51a
7.270.98a
4.270.41b
a
Presented values are the means7standard deviation of three replicate trials. Values in the same row with different superscripted letters differ significantly (po0:05). b mL 0.1 N NaOH mL1 milk. c Details of test are given in the text.
80 60 40 20 0 0
(b)
265
30
60 90 120 150 Incubation Time (min)
180
210
Fig. 1. Effect of somatic cell count in ewes’ milk on time-related changes in pH (a) and titratable acidity (b) on incubation with starter culture during yoghurt manufacture. Somatic cell count: cells mL1: o500,000 (’); 1,000,000–1,500,000 (m); 43,000,000 (J).
acid production is due to the logarithmic phase of starter growth, which tapers off as the level of acidity becomes limiting (Shaker, Jumah, & Abu-Jdayil, 2000). The change in pH during the incubation of the milks was similar in the milks with low and medium counts, and about 210 min was required to reach a pH of 4.5, after which fermentation ceased. The pH decreased much faster in the milk with the highest SCC, fermentation ceasing at 150 min, despite the initial pH of the milk being higher. The fermentation of milk with high SCC may be inhibited owing to the inhibition of the starter cultures as a result of the antimicrobial action of SCs (Heras, Dominguez, & Fernandez-Garayzabal, 1999). However, our results would suggest the opposite. The changes in pH and acidity in milk with high SCC were different from those of the milk with low SCC. 3.3. Properties of 72 h old yoghurt The SCC of the milk did not significantly influence the pH or acidity of the yoghurt at 72 h. However, the SCC significantly affected the levels of syneresis. The milk with the highest SCC had the highest level of syneresis, pointing to the poor consistency of the coagulum of this milk and its inability to retain serum. The milk from
animals with mastitis had a high level of serum proteins, and low level of casein (Pirisi et al., 1996; Martı´ de Olives & Molina Pons, 1998). For this reason, milk with high SCC would give rise to soft coagula with a reduced ability to retain serum (Tamine & Robinson, 1999). The rheological characteristics of the yoghurts are shown in Table 2. Yoghurts manufactured from milk with low- or medium-SCC did not statistically differ from each, but differed significantly from those manufactured from milk with a SCC43,000,000 cells mL1. Yoghurt from high-SCC milk was softer and more adhesive than yoghurt from milk with lower SCC. According to Tamine and Robinson (1999), one of the factors that can affect the firmness and adhesiveness of yoghurt is the amount of lactose present in the milk. Milk with SCC43,000,000 cells mL1 had the lowest lactose content, which may have affected the texture of the yoghurt. Similarly, the results of Urech, Puhan, and Scha¨llibaum (1999) showed that milk with high SCC had a relatively low level of casein, and a high level of serum proteins. The SCC of the milk significantly influenced the grading score of yoghurts (Table 2). The high-SCC milk gave yoghurt which received the lowest grading score. The yoghurts manufactured from milk with high SCC were described by the tasters as having a piquant and bitter taste. Owing to the damage caused to the mammary tissue by mastitis, blood plasma leaks into the milk (Barbano et al., 1987) and such plasma contains many enzymes, such as proteases and lipases. These enzymes hydrolyse protein and fat, causing losses in yield (Barbano et al., 1987) and, possibly, result in the development of undesirable flavours and textures in the dairy products manufactured from this milk.
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266
4.9 4.7
pH
4.5 4.3 4.1 3.9 3.7 3.5 3
6
9
(a)
12 15 18 21 Storage time (days)
24
28
Acidity (mL 0.1N NaOH mL-1)
180
Increasing the SCC of the milk from o500,000 to 1,000,000–1,500,000 cell mL1 did not significantly influence the texture, syneresis or grading quality of the resultant yoghurt. However, SCCs43,000,000 cell mL1 resulted in yoghurt that had a very soft consistency and a reduced ability to retain serum, and that was characterised by flavours described as piquant and bitter. SCC resulted in a marked non-significant decreased in milk protein level, a significant decrease in yoghurt hardness, and significant increases in the adhesiveness and syneresis of yoghurt. From these results, it seems that the SCC in ewes’ milk destined for the manufacture of yoghurts should not exceed 1,500,000 cell mL1.
160 140 120
Acknowledgements
100 80
Thanks are due to the ASOVINO Cooperative Association for funding this project.
60 40 20 0 3
(b)
6
9
15 12 18 21 Storage time (days)
24
28
Fig. 2. Effect of somatic cell count in ewes’ milk on changes in pH (a) and titratable acidity (b) on the resultant yoghurt. Somatic cell count: cells mL1: o500,000 (’); 1,000,000–1,500,000 (m); 43,000,000 (J).
3.4. Changes in yoghurt during storage The pH and titratable acidity values of the yoghurt stored at 4 1C for 28 days are shown in Fig. 2. The pH values of all the yoghurts decreased and acidity values increased with storage time. This can be attributed to acid production during cold storage (post-acidification) as a result of the conversion of lactose to lactic acid by the bacterial cultures (Katsiari et al., 2002). There were no significant (po0:05) differences in either pH or acidity between the yoghurts from the milk with low- or medium-SCCs at storage times 415 days. However, in the case of milk with high SCC, the pH values at storage time 415 days were significantly lower than those of the others milks. The SCC in the milk did not significantly influence the acidity of the yoghurts during storage.
4. Conclusions As the SCC of ewes’ milk increased, the pH increased and the lactose concentration decreased. The titratable acidity and the contents of fat, protein, and total solids were not significantly influenced by the SCC of the milk.
References Alichanidis, E., & Polychroniadon, A. (1995). Special features of dairy products from ewe and goat milk from the physiochemical and organoleptic point of view. IDF Greek National Committee of IDF CIRVAL Seminar, Creta, Greece, October 21– 29, 1995. (pp. 122–127). Barbano, D.M., Verdi, R.J., & Rasmussen, R. (1987). Influence of milk somatic cell count on cheese manufacturing and cheese yield. 24th Annual Marschall invitational Italian Cheese Seminar, Wisconsin, USA, September 16– 17, 1987. (pp.12–16). Bonczar, G., Wszolek, M., & Siuta, A. (2002). The effects of certain factors on the properties of yoghurt made from ewe’s milk. Food Chemistry, 79, 85–91. Bourne, M. C. (1978). Texture profile analysis. Food Technology, 32, 62.66. Boyazoglu, J., & Morand-Fehr, P. (2001). Mediterranean dairy sheep and goat products and their quality. A critical review. Small Rumiant Research, 40, 1–11. British Standards Institution (BSI). (1995). Gerber method for the determination of fat in milk and milk products. British Standard 696. London: British Standards Institution. Dı´ az, J. R., Muelas, R., Segura, C., Peris, C., & Molina, P. (1996). Effect of mastitis on milk composition in Manchega ewes: Preliminary results. In Somatic cell and milk of small ruminants (pp. 240–246). Wageningen, The Netherlands: Pers Wageningen. Duling, A. M., Paape, M. J., Schultze, W. D., & Weinland, B. T. (1983). Effect of parity, stage of lactation and intramammary infection on concentration of somatic cells and cytoplasmic particles in goat milk. Journal of Dairy Science, 66, 2426–2433. Fruganti, G., Ranicci, S., Tesei, B., & Valente, C. (1985). Valutazione dello stato sanitario della mamella di pecora durante un intero ciclo di lattazione. Clinica Veterinaria, 108, 286–288. Gonzalez Andrada, J. I., Romero Estevez, C., & Jime´nez-Pe´rez, S. (1994). Control de calidad en la fabricacio´n de yoghurt. Alimentacio´n, Equipos y Tecnologı´a, 7, 77–81. Haenlein, G. F. W. (1995). Topics for success with dairy goats. In Dairy Extension Bulletin (pp. 105–112). Delaware, USA: University of Delaware.
ARTICLE IN PRESS A.M. Vivar-Quintana et al. / International Dairy Journal 16 (2006) 262–267 Haenlein, G. F. W. (1998). The value of goats and sheep to sustain mountain farmers. International Journal of Animal Science, 13, 187–194. Harmon, R. J. (1994). Physiology of mastitis and factors affecting somatic cell counts. Journal of Dairy Science, 77, 2103–2112. Hartley, D., Duong, C., David, P., & Fazel, A. (1989). Symbiosis of yogurt microorganisms. In Les laits fermente´s (pp. 139–145). Montrouge, France: John Libbery Eurotext. Harwalkar, V. R., & Kalab, M. (1983). Susceptibility of yoghurt to syneresis. Comparison of centrifugation and drainage methods. Milchwissenschaft, 38, 517–522. Heras, A., Dominguez, L., & Ferna´ndez-Garayzabal, J. F. (1999). Prevalence and aetiology of subclinical mastitis in dairy ewes of the Madrid Region. Small Ruminant Research, 32, 21–29. IDF. (1974). Determination of the lactose content of milk. IDF Standard 28 A. Brussels, Belgium: International Dairy Federation. IDF. (1986). Milk. Determination of nitrogen content (Kjeldahl method) and calculation of crude protein content. IDF Standard 20 A. Brussels, Belgium: International Dairy Federation. IDF. (1987). Milk, cream and evaporated milk. Determination of total solids content. IDF Standard 21B. Brussels, Belgium: International Dairy Federation. IDF. (1991). Yoghurt. Determination of total solids content. IDF Standard 151. Brussels, Belgium: International Diary Federation. Jaeggi, J. J., Govindasamy-Lucey, S., Berger, Y. M., Johnson, M. E., McKusick, B. C., Thomas, D. L., & Wendorff, W. L. (2003). Hard ewe0 s milk cheese manufactured from milk of three different groups of somatic cell counts. Journal of Dairy Science, 86, 3082–3089.
267
Katsiari, M. C., Voutsinas, L. P., & Kondyli, E. (2002). Manufacture of yoghurt from stored frozen sheep’s milk. Food Chemistry, 77, 413–420. Martı´ de Olives, A., & Molina Pons, P. (1998). Mamitis y calidad de leche de oveja. Ovis, 59, 11–25. Muelas, R., Molina, P., Diaz, J. R., & Peris, C. (1996). Relationship between somatic cell count, acidity and California Mastitis test in Manchega ewe milk. In Somatic cells and milk of small Ruminants (pp. 213–219). Wageningen. The Netherlands: Pers Wagening. Paape, M. J., Poutrel, B., Contreras, A., Marco, J. C., & Capucol, A. V. (2001). Milk somatic cells and lactation in Small Ruminants. Journal of Dairy Science, 84, 236–244. Pirisi, A., Pireda, G., Corona, M., Pes, M., Pinutus, S., & Ledda, A. (2000). Efecto de las ce´lulas soma´ticas sobre la composicio´n de la leche de oveja y las caracterı´ sticas del producto elaborado. Ovis, 66, 49–58. Pirisi, A., Piredda, G., Podda, F., & Piutus, S. (1996). Effect of somatic cell count on sheep milk composition and cheese making properties. In Somatic cell and milk of small ruminants (pp. 187–192). Wageningen, The Netherlands: Pers Wageningen. Shaker, R. R., Jumah, R. Y., & Abu-Jdayil, B. (2000). Rheological properties of plain yoghurt during coagulation process: Impact of fat content and preheat treatment of milk. Journal of Food Engineering, 44, 175–180. Tamine, A. Y., & Robinson, R. K. (1999). Yoghurt science and technology. Cambridge, UK: Woodhead Publishing Limited. Urech, E., Puhan, Z., & Scha¨llibaum (1999). Changes in milk protein fraction as affected by subclinical mastitis. Journal Dairy Science, 82, 2402–2411.