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Animal Science Journal (2014) ••, ••–••

doi: 10.1111/asj.12323

ORIGINAL ARTICLE Influence of race and crossbreeding on casein micelles size Denise R. FREITAS,1 Leorges M. FONSECA,2 Fernando N. SOUZA,1 Cristiane V. G. LADEIRA,1 Soraia A. DINIZ,1 João Paulo A. HADDAD,1 Diêgo S. FERREIRA,3 Marcelo M. SANTORO4* and Mônica M. O. P. CERQUEIRA2 1

Department of Veterinary Preventive Medicine, 2Technology and Inspection of Animal Products, 3Pharmaceutical Products and 4Biochemistry and Immunology, Federal University of Minas Gerais, Brazil, Belo Horizonte, Brazil

ABSTRACT Casein (CN) micelles are colloidal aggregates of protein dispersed in milk, the importance of which in the dairy industry is related to functionality and yield in dairy products. The objective of this work was to investigate the correlation of milk CN micelles diameter from Holstein and Zebu crossbreds with milk composition (protein, fat, lactose, total and nonfat solids and milk urea nitrogen), somatic cell count (SCC), age, lactation stage and production. Average casein micelles diameters of milk samples obtained from 200 cows were measured using photon correlation spectroscopy and multiple regression analysis was used to find relationship between variables. CN micelle diameter, SCC and nonfat solids were different between animals with different Holstein crossbreed ratios, which suggests influence of genetic factors, mammary gland health and milk composition. Overall, results indicate the potential use of CN micelle diameter as a tool to select animals to produce milk more suitable to cheese production.

Key words: milk urea nitrogen, nonfat solids, protein, somatic cell count.

INTRODUCTION The amount of milk processed as derivatives in Brazil has been increasing during the last years (United States Department of Agriculture 2013). For a sustainable production, milk quality and composition are essential conditions to obtain high-quality and profitable products (Lucey & Singh 1997). Casein is among the important components of milk, with four major types, αs1, αs2, β and κ, and together with calcium phosphate, 1000s of casein molecules form aggregates of micelles with an average diameter of 150 to 200 nm (de Kruif 1998). The micelle structure influences milk stability to heating, freezing and drying, gelling properties (de Kruif 1998), and functionality of products, such as cheese and yoghurt (Dalgleish & Corredig 2012). Previous reports indicate that genetic, feed management and milk composition can affect casein micelle size (Devold et al. 2000; de Kruif & Huppertz 2012). Therefore, animal selection toward production of milk with smaller casein micelles might improve milk renneting properties and consequently cheese production. Smaller micelles are more compact and have higher proportion of κ-casein (Devold et al. 2000) resulting in a firmer gel during initial steps of cheese production (Ford & Grandison 1986). © 2014 Japanese Society of Animal Science

Crossbreeding of Holstein with Zebu races is widely used for dairy cows in Brazil, especially to obtain animals with important economic implications. The Gyr is one of the principal Zebu breeds used for dairy production. Crossbreeding of Holstein and Gyr have a significant heterosis effect for characteristics of economic impact and adaptability to tropical climate (Madalena 2002). However, to the best of our knowledge, no evaluation of crossbreeding influence over casein micelles has been reported. Therefore, the objective of this work was to investigate the average casein micelles size from different Holstein × Zebu crossbreeding ratios, and to correlate it with milk production and composition, somatic cell count (SCC), animal age, and lactation stage. Correspondence: Denise Ribeiro de Freitas, Department of Veterinary Preventive Medicine, Escola de Veterinária, Universidade Federal de Minas Gerais, Av. Presidente Antônio Carlos, 6627, Belo Horizonte 30123-970, Minas Gerais, Brazil. (Email: [email protected]); Mônica Maria Oliveira Pinho Cerqueira, Department of Technology and Inspection of Animal Products, Federal University of Minas Gerais, Brazil. (Email: [email protected]) *In Memoriam. Received 2 April 2014; accepted for publication 4 August 2014.

2 D. R. FREITAS et al.

MATERIALS AND METHODS Milk samples and data from crossbred cows (n = 200) of five dairy farms (Minas Gerais State, Brazil) were obtained during the months of March to April, 2013, with the following Holstein × Gyr ratios: 1/2 (n = 41); 9/16 (n = 21); 5/8 (31); 3/4 (29); 7/8 (51); and 15/16 (27). Composition (fat, protein, lactose, nonfat solids and milk urea nitrogen (MUN)) was analyzed by Fourier Transform Infrared spectroscopy (FTIR) (International Dairy Federation 2000), and SCC by flow cytometer (International Dairy Federation 1995), using a CombiscopeTM FTIR 400 equipment (Advanced/Delta Instruments®, Drachten, Netherlands). SCC was transformed in somatic cell score (SCS) using log transformation (SCS = Log2 (SCC/100 000) + 3) (Dabdoub & Shook 1984). Casein micelles diameter was estimated using photon correlation spectroscopy (PCS) (Devold et al. 2000) after fat layer removal (centrifugation at 2000 × g/30 min, and 5°C). Analysis was done using a Zetasizer 3000HS (Malvern Instruments Ltd, Malvern, UK) equipped with a helium-neon laser (632.8 nm), and set for unimodal analysis, and light scattering at 25°C at a 90° angle. Defatted milk was diluted (1:1000) in simulated milk ultrafiltrate (SMUF) (Jenness & Koops 1962) to reach the optimum counting condition specified for the equipment. The solution was previously filtered in a 0.22 μm filter (Millex-HV 0.45 μm, polyvinyl difluoride, 33 mm, non-sterile, Millipore Ltd, Hertfordshire, UK). Variables with P < 0.20 in a simple regression model were selected for analysis in a multiple regression model. Final multiple regression model was composed by variables with P < 0.05 (STATA v. 12; Stata Corp., College Station, TX, USA). The variables under examination were first analyzed individually to verify the significance and then in combination to assess the effect of the single variables on all others. In the first stage of the analysis, unconditional logistical model for each variable related to the casein micelles size, with P-values lower than 0.20, were considered as a select variable and passed to the next stage of the analysis. In the final model, variables selected in the first stage were used to develop a multivariate logistical model in which variables with P ≤ 0.05 were retained in the final model.

RESULTS AND DISCUSSION Average diameter of casein micelles was 170.22 ± 21.18 nm (121.8–235.6 nm). Regression model was adjusted according to the following exploratory variables (P < 0.20): crossbreeding ratio, animal age, days in lactation, milk production, SCS, composition (fat, protein, lactose, nonfat solids and MUN, Table 1). Only three variables were statistically significant (P ≤ 0.05) to remain in the final multivariate model to estimate milk casein micelles diameter (crossbreeding ratio, SCS and nonfat solids, Table 2). Age, milk production, fat, protein and MUN were associated with average casein micelle diameter (P ≤ 0.05) only through univariate regression (Table 1). Milk casein micelles presented a wide size variation, between 121.8 and 235.6 nm (Fig. 1), but similar to sizes previously reported for European races (Horne & Dalgleish 1985; de Kruif 1998; Devold et al. 2000). The positive coefficient for crossbreeding ratio in multiple regression indicates that the average diameter © 2014 Japanese Society of Animal Science

Table 1 Variables eligible for entry in the final model by logistic regression model (P ≤ 0.20)

Variables

Odds ratio P-value

95% CI

Overall P-value ≤ 0.001 1/2 Holstein × Gyr −7.96 9/16 Holstein × Gyr −0.72 5/8 Holstein × Gyr 21.02 3/4 Holstein × Gyr 11.40 7/8 Holstein × Gyr 13.92 15/16 Holstein × Gyr 20.84 Age −1.35 Milk production 0.73 Somatic cell score −1.40 Fat −3.97 Protein −14.99 Nonfat solids −7.04 Milk urea nitrogen 0.91

< 0.01 18.45, 2.53 < 0.01 −40.32, 38.87 < 0.01 11.71, 30.33 < 0.01 1.91, 20.90 < 0.01 5.71, 22.12 < 0.01 11.03, 30.65 0.024 −2.53, −0.18 0.001 0.31, 1.15 0.002 −2.27, −0.53 0.010 −7.01, −0.94 0.002 −24.59, −5.40 0.024 −13.15, −0.92 0.005 0.27, 1.54

Table 2 Final model (P ≤ 0.05) for variables associated with casein micelles diameter using multivariate analysis (R2 = 0.132; adjusted R2 = 0.119) (n = 200)

Variables

Odds ratio (95% CI)

P-value

Crossbreeding ratio Somatic cell score Nonfat solids

0.270 (0.10, 0.43) −1.396 (−2.23, 0.55) −6.549 (−12.56, 10.53)

0.001 0.001 0.033

of casein micelles is positively correlated with the increasing Holstein/Gyr ratio. Therefore, crossbreeding ratio is influential on milk casein micelle structure. Aspects such as casein genotypes may help to understand the factors involved in Holstein crossbreeding effects on milk micelles, since there are polymorphisms of κ casein in the cattle of crossbred Holstein (Molee et al. 2011), and κ casein genotype is correlated with micelle size. Micelles containing κ casein AB are smaller than those with AA and AE polymorphs (Devold et al. 2000). No work about the effect of Holstein × Zebu crossbreed ratios on the casein micelle structure and composition has been reported. Hence, investigation on markers for animal selection with desirable characteristics for the dairy industry is a potential field, especially regarding their economic implications. The negative sign of SCS and nonfat solids indicates inverse relationship between these variables and average casein micelle diameter. SCS influence may be related to the increase in proteolitic activity of milk indigenous and endogenous enzymes associated with leucocytes from blood (Roux et al. 2003). Plasmin is the most common indigenous enzyme found in milk with high SCC (Donnelly & Barry 1983), followed by catepsin D. Other indigenous enzymes include catepsins G and B, elastase and collagenase (Owen & Campbell 1999). These enzymes, if present in high concentration due to elevated SCC, will hydrolyze caseins, with plasmin affinity for αs1, αs2 and β caseins Animal Science Journal (2014) ••, ••–••

CASEIN MICELLES SIZE IN ZUBE CATTLE

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Figure 1 Micelle size casein distribution according to percentage of samples.

(Bastian & Brown 1996), while catepsin G, a serine protease, hydrolyzes only αs1 and β caseins (Considine et al. 2002). Catepsin D is an aspartic protease, which hydrolyzes αs1, αs2, β and κ caseins. Casein hydrolysis is correlated with high SCC (Senyk et al. 1985; Verdi et al. 1987), and casein micelles diameter might be reduced in milk with SCC higher than 200 000 cells/mL (Moslehishad & Ezzatpanah 2010), maybe due to the structural changes induced by the casein hydrolysis. The negative correlation of casein micelles size with nonfat solids content indicate that milk composition is a factor to be considered. Although the multiple linear regression did not indicate which component was individually influential, it is possible to infer using the univariate regression model (Table 1) that average micelle diameter is associated with protein (P = 0.002), fat (P = 0.01) and MUN (P = 0.005). The possible explanation is that these variables interact with others in the model. Glantz et al. (2010) did not observe correlation between protein and average diameter of casein micelles. Overall, results indicate the potential use of casein micelle diameter as a tool to select animals to produce milk more suitable to cheese production. However, more studies are necessary to investigate crossbreeding influence in this index, and technological impacts thereof.

Conclusion Smaller casein micelles and higher SCC were found in milk produced by cows with lower Holstein/Gyr ratio, Animal Science Journal (2014) ••, ••–••

which indicates genetic, mammary gland health and milk composition influence.

ACKNOWLEDGMENTS The authors are grateful for the financial support from the Fundação de Amparo à Pesquisa do estado de Minas Gerais (FAPEMIG).

REFERENCES Bastian ED, Brown RJ. 1996. Plasmin in milk and dairy products: an update. International Dairy Journal 6, 435– 457. Considine T, Geary S, Kelly AL, McSweeney PLH. 2002. Proteolytic specificity of cathepsin G on bovine αs1- and β-caseins. Food Chemistry 76, 59–67. Dabdoub SM, Shook GE. 1984. Phenotypic relations among milk yield, somatic cell count and clinical mastitis. Journal of Dairy Science 67 (Supp. 1), 163–164. Dalgleish DG, Corredig M. 2012. The structure of the casein micelle of milk and its changes during processing. Annual Review of Food Science and Technology 3, 449–467. De Kruif CG. 1998. Supra-aggregates of casein micelles as a prelude to coagulation. Journal of Dairy Science 81, 3019– 3028. De Kruif CG, Huppertz T. 2012. Casein Micelles: size Distribution in Milks from Individual Cows. Journal of Agricultural and Food Chemistry 60, 4649–4655. Devold TG, Brovold MJ, Langsrud T, Vegarud GE. 2000. Size of native and heated casein micelles, content of protein and minerals in milk from Norwegian Red Cattle – effect of milk protein polymorphism and different feeding regimes. International Dairy Journal 10, 313–323. © 2014 Japanese Society of Animal Science

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Donnelly WJ, Barry JG. 1983. Casein compositional studies. III. Changes in Irish milk for manufacturing and role of milk proteinase. Journal of Dairy Research 50, 433– 441. Ford GD, Grandison AS. 1986. Effect of size of casein micelles on coagulation properties of skim milk. Journal of Dairy Research 53, 129–133. Glantz M, Devold TG, Vegarud GE, Lindmark MÅnsson H, StÅlhammar H, Paulsson M. 2010. Importance of casein micelle size and milk composition for milk gelation. Journal of Dairy Science 93, 1444–1451. Horne DS, Dalgleish DG. 1985. A photon correlation spectroscopy study of size distributions of casein micelle suspensions. European Biophysics Journal 11, 249–258. International Dairy Federation. 1995. Milk: Enumeration of Somatic Cells. International Dairy Federation, Brussels. International Dairy Federation. 2000. Whole Milk: Determination of Milkfat, Protein and Lactose Content, Guidance on the Operation of Mid-Infrared Instruments. International Dairy Federation, Brussels. Jenness R, Koops J. 1962. Preparation and properties of a salt solution which simulates milk ultrafiltrate (Amsterdam), Netherlands Milk and Dairy Journal 16, 153–164. Lucey JA, Singh H. 1997. Formation and physical properties of acid milk gels: a review. Food Research International 30, 529–542. Madalena FE. 2002. Animals that Produce Dairy Foods |Bos indicus Breeds and Bos indicus × Bos taurus Crosses. In:

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Fuquay JW (ed.), Encyclopedia of Dairy Sciences, 2nd edn. pp. 576–585. Academic Press, San Diego, CA. Molee A, Boonek L, Rungsakinnin N. 2011. The effect of beta and kappa casein genes on milk yield and milk composition in different percentages of Holstein in crossbred dairy cattle. Anim Science Journal 82, 512–516. Moslehishad M, Ezzatpanah H. 2010. Transmission electron microscopy study of casein micelle in raw milk with different somatic cell count levels. International Journal of Food Properties 13, 546–552. Owen CA, Campbell EJ. 1999. The cell biology of leukocytemediated proteolysis. Journal Leukocyte Biology 65, 137– 150. Roux YL, Laurent F, Moussaoui F. 2003. Polymorphonuclear proteolytic activity and milk composition change. Veterinary Research. 34, 629–645. Senyk GF, Barbano DM, Shipe WF. 1985. Proteolysis in milk associated with increasing somatic cell counts. Journal of Dairy Science 68, 2189–2194. United States Department of Agriculture. 2013. Brazil dairy and products annual. United States Department of Agriculture, Global Agricultural Information Network. Available from URL http://www.thefarmsite.com/reports/ contents/BrazilDairy&Products22Oct2013.pdf Verdi RJ, Barbano DM, Dellavalle ME, Senyk GF. 1987. Variability in true protein, casein, nonprotein nitrogen, and proteolysis in high and low somatic cell milks. Journal of Dairy Science 70, 230–242.

Animal Science Journal (2014) ••, ••–••

Influence of race and crossbreeding on casein micelles size.

Casein (CN) micelles are colloidal aggregates of protein dispersed in milk, the importance of which in the dairy industry is related to functionality ...
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