β-lactoglobulin and α-lactalbumin Hydrolysates as Sources of Antibacterial Peptides

Authors
M. Sedaghati 1
H. Ezzatpanah 2
M. Mashhadi Akbar Boojar 3
M. Tajabadi Ebrahimi 4
1 Department of Food Science and Technology, Faculty of Agriculture and Natural Resources, Science and Research Branch, Islamic Azad University, Tehran, Iran
2 Department of Food Science and Technology, Faculty of Agriculture and Natural Resources, Science and Research Branch, Islamic Azad University, Tehran 14515.775, Iran.
3 Department of Biochemistry, University of Kharazmi, Tehran, Iran
4 Department of Science, Faculty of Biology, Islamic Azad University, Tehran Central Branch, Iran
Abstract
The presence of antibacterial activity in bovine β-lactoglobulin and in α-lactalbumin hydrolysates was investigated. The Plasmin-Digest of β-lactoglobulin (PDβ) and of α-lactalbumin (PDα) were fractionated, using reversed phase high performance liquid chromatography. The antibacterial activity of β-lactoglobulin, α-lactalbumin, nisin, plasmin, PDβ and PDα were in vitro tested against pathogenic (Escherichia coli and Staphylococcus aureus) and probiotic (Lactobacillus casei and Lactobacillus acidophilus) bacteria. Although α-lactalbumin, β-lactoglobulin and plasmin exhibited no antibacterial activity, PDβ, PDα and nisin revealed antibacterial activity against the bacteria tested. The Minimum Inhibitory Concentration (MIC) of these compounds was determined for the bacteria cultures. Similar to nisin, the MIC of PDβ and of PDα against Gram-positive bacteria was recorded as considerably lower than the MICs against Gram-negative bacteria. The study also evaluated the effect of PDβ, PDα and nisin on the growth curves and on the plate count confirmations of the target bacteria. The results revealed that nisin, PDβ and PDα have inhibitory effects on the lag phase, maximum OD620 and on plate count confirmation of the bacteria tested. The maximum inhibitory effect of these compounds was created during the log phase. Their inhibitory effects depended upon their concentrations, higher concentration causing stronger antibacterial activity. The PDβ and PDα proved more active against Gram-negative bacteria than did nisin, but nisin revealed substantial inhibitory activity against Gram-positive bacteria.

Keywords

1. Bellamy, W., Takase, M., Yamauchi, K., Wakabayashi, H., Kawase, K. and Tomita, M. 1992. Identification of the Bactericidal Domain of Lactoferrin. Biochim. Biophys. Acta., 1121: 130–136.
2. Benkerroum, N. 2010. Antimicrobial Peptides Generated from Milk Proteins a Survey and Prospects for Application in the Food Industry. Int. J. Dairy. Technol., 63: 320-338.
3. Boziaris, I. S. and Adams, M. R. 1999. Effect of Chelators and Nisin Produced In situ on Inhibition and Inactivation of Gram Negatives. Int. J. Food. Microbiol., 53:105-113.
4. Brew, K. and Grobler, J. A. 1992. α-Lactalbumin. In: ״ Advanced dairy chemistry: Proteins״ , (Ed.): Fox, P. F.. Elsevier Applied Science Publishers, London, PP. 191-229.
5. Chaneton, L., Pérez Sáez, J. M. and Bussmann, L. M. 2011. Antimicrobial Activity of Bovine β-Lactoglobulin against Mastitis-causing Bacteria. J. Dairy. Sci., 94:138–145.
6. Chatterton, D. E. W., Smithers, G., Roupas, P. and Brodkorb, A. 2006. Bioactivity of β-lactoglobulin and α-lactalbumin Technological Implications for Processing. Int. Dairy. J., 16: 1229–1240.
7. Chen, J. H. and Ledford, R. A. 1971. Purification and Characterization of Milk Protease. J. Dairy. Sci., 54:763.
8. Dalasgaard, T. K., Heegaard, C. W. and Larsen, L. B. 2008. Plasmin Digestion of Photooxidized Milk Proteins. J. Dairy. Sci., 91: 2175- 2183.
9. El-Zahar, K., Sitohy, M., Choiset, Y., Metro, F., Haertle, T. and Chobert, J. M. 2004. Antimicrobial Activity of Ovine Whey Protein and Their Peptic Hydrolysates. Milchwissenschaft., 59: 653–656.
10. Farouk, A., 1982. Antibacterial Activity of Proteolytic Enzymes. Int. J. Pharm., 12: 295-298.
11. Fitzgerald, R. J., Murray, B. A. and Walsh, D. J. 2004. Hypotensive Peptides from Milk Proteins. J. Nutri., 134: 980S–988S.
12. Floris, R., Recio, I., Berkhout, B. and Visser, S. 2003. Antibacterial and Antiviral Effects of Milk Proteins and Derivatives Thereo. Curt. Pharm. Design., 9: 1257-1275.
13. Fox, P. F., 1991. Proteinase. In: ״Food Enzymology״, (Ed.): Fox, P. F.. Elsevier Applied Science, New York, PP. 79-88.
14. Ganzle, M. G., Hertel, C. and Hammes, W. P. 1999. Resistance of Escherichia coli and Salmonella against Nisin and Curvacin A. Int. J. Food. Microbiol., 48: 37–50.
15. Guo, M. R., Fox, P. F., Flynn, A. and Kindstedt, P. S. 1995. Susceptibility of Beta-lactoglobulin and Sodium Caseinate to Proteolysis by Pepsin and Trypsin. Food. Chem., 78: 2336-2244.
16. Hancock, R. E. W. and Lehrer, R. 1998. Cationic Peptides: A New Source of Antibiotics. Trends. Biotechnol., 16: 82-88.
17. Heike, B. and Sahl, H. G. 2000. New Insights in to the Mechanism of Action of Lantibiotics-diverse Biological Effects by Binding to the Same Molecular Target. J. Antimicrob. Chemother., 46: 1-6.
18. Hernández-Ledesma, B., Recio, I. and Amigo, L. 2008. Beta-lactoglobulin as Source of Peptides. Amino. Acids., 35: 257-65.
19. Kordel, M., Schuller, F. and Sahl, H. G. 1989. Interaction of the Pore Forming-peptide Antibiotics Pep 5, Nisin and Subtilin with Non-energized Liposomes. FEBS Lett., 244: 99-102.
20. Korhonen, H. and Pihlanto-Leppälä, A. 2004. Milk-derived Bioactive Peptides: Formation and Prospects for Health Promotion. In: ״Handbook of Functional Dairy Products״, (Eds.): Shortt, C. and Brien, J. O.. CRC Press, Boca Raton, FL, PP. 109 – 124.
21. Lopez-Exposito, I. and Recio, I. 2006. Antibacterial Activity of Peptides and Folding Variants from Milk Proteins. Int. Dairy. J., 16: 1294–1305.
22. McCann, K. B., Shiell, B. J., Michalski, W. P., Lee, A., Wan, J., Roginski, H. and Coventry, M. J. 2006. Isolation and Ccharacterization of a Novel Antibacterial Peptide from Bovine αS1-casein. Int. Dairy. J., 16: 316–323.
23. Mortazavian, A. M. and Sohrabvandi, S. 2006. Probiotic Products. In: ״Probiotics and Food Probiotic Products Based on Dairy Probiotic Products״, (Ed.): Mortazavian, A. M.. Eta Publication, Tehran, PP. 330-372
24. Pakkanen, R. and Aalto, J. 1997. Growth Factors and Antimicrobial Factors of Bovine Colostrum. Int. Dairy. J., 7: 285–297.
25. Park, Y. W. 2009. Overview of Bioactive Components in Milk and Dairy Products. In: ״Bioactive Components in Milk and Dairy Products״, (Ed.): Park, Y. W.. Wiley-Blackwell, Ames, Iowa, USA, PP. 3-5.
26. Pellegrini, A., Thomas, U., Bramaz, N., Hunziker, P. and von Fellenberg, R. 1999. Isolation and Identification of Three Bactericidal Domains in the Bovine α-lactalbumin Molecule. Biochim. Biophys. Acta., 1426: 439–448.
27. Pellegrini, A., Dettling, C., Thomas, U. and Hunziker, P. 2001. Isolation and Characterization of Four Bactericidal Domains in the β-lactoglobulin. Biochim. Biophys. Acta., 1526: 131–140.
28. Pihlanto-Leppala, A., Marnila, P., Hubert, L., Rokka, T., Korhonen, H. J. and Karp, M. 1999. The Effect of α-lactalbumin and β-lactoglobulin Hydrolysates on the Metabolic Activity of Escherichia coli JM103. J. Appl. Microbiol., 87: 540–545.
29. Schmidt, D. G. and Poll, J. K. 1991. Enzymatic Hydrolysis of Whey Proteins. Hydrolysis of α-lactalbumin and β-lactoglobulin in Buffer Solutions by Proteolytic Enzymes. Neth. Milk. Dairy. J., 45: 225–240.
30. Sternhagen, L. G. and Allen, J. C. 2001. Growth Rates of a Human Colon Adenocarcinoma Cell Line Are Regulated by the Milk Protein α-lactalbumin. Adv. Exp. Med. Biol., 501: 115–120.
31. Thompson, A., Boland, J. M. and Singh, H. 2009. Milk Proteins: From Expression to Food. Amsterdam, Elsevier Academic Press, New York, PP. 25-32.
32. Vongsawasdi, P., Nopharatana, M., Supanivatin, P. and Matthana, P. 2012. Effect of Nisin on the Survival of Staphylococcus aureus Inoculated in Fish Balls. Asian. J. Food. Agro. Ind., 5: 52-60.
Journal of Agricultural Science and Technology
Volume 16, Issue 7
November and December 2014
Pages 1587-1600