PROTEOLYTIC POTENTIAL OF PSEUDOMONAS FLUORESCENS ISOLATED FROM REFRIGERATED RAW MILK
DOI:
https://doi.org/10.21206/rbas.v4i2.254Resumo
The growth rate and the proteolytic activity of Pseudomonas fluorescens strains 07A and 041, isolated from cow’s milk, were evaluated at 2, 4, 7 and 10ºC. P. fluorescens promoted protein degradation during storage of milk samples as observed by Proteolytic activity measurement, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and heat stability of milk. Casein hydrolysis resulted in loss of thermal stability of milk and in formation of fragments of low and medium molecular mass. Temperatures up to 10°C did not
guarantee raw milk quality when contamination by P. fluorescens was equal or higher than 106 cfu/mL.
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Referências
ADAMS, D.M.; BARACH, J.T.; SPECK, M.L. Effect of psychrotrophic bacteria from raw milk on milk proteins and stability of milk proteins to
ultrahigh temperature treatment. Journal of Dairy Science, v.59, n.5, p.823-827, 1976.
BAGLINIÈRE, F.; TANGUY, G.; JARDIN, J. et al. Quantitative and qualitative variability of the caseinolytic potential of different strains of
Pseudomonas fluorescens: implications for the stability of casein micelles of UHT milks during their storage. Food Chemistry, v.135, n.4, p.2593-2603, 2012.
BRASIL. Regulamento Técnico de Produção, Identidade e Qualidade do Leite tipo A, Regulamento Técnico de Identidade e Qualidade de Leite Cru Refrigerado, Regulamento Técnico de Identidade e Qualidade de Leite Pasteurizado e Regulamento Técnico da Coleta de Leite Cru Refrigerado e seu Transporte a Granel. Instrução Normativa nº 62, de 29 de de dezembro de 2011.
Diário Oficial [da] República Federativa do Brasil, Brasília, DF: Ministério da Agricultura, Pecuária e Abastecimento: p.6-37, 2011.
BRAUN, P.; SUTHERLAND, J.P. Predictive modelling of growth and enzyme production and activity by a cocktail of Pseudomonas spp.,
Shewanella putrefaciens and Acinetobacter. International Journal of Food
Microbiology, v.86, n.3, p.271-282, 2003.
BROCK, T.D. Biology of microorganisms. 7th Ed. New Jersey: Prentice Hall, 1994, 909p.
CELESTINO, E.L.; IYER, M.; ROGINSKI, H. Reconstituted UHT-treated milk: Effects of raw milk, powder quality and storage conditions of
UHT milk on its physico-chemical attributes and flavour. International Dairy Journal, v.7, n.2-3, p.129-140, 1997.
CHEN, L.; DANIEL, R.M.; COOLBEAR, T. Detection and impact of protease and lipase activities in milk and milk powders. International Dairy Journal, v.13, n.4, p.255-275, 2003.
COSTA, L.M.; GÓMEZ, F.S.; MOLINA, L.H.C. et al. Purificación y caracterización de proteasas de Pseudomonas fluorescens y sus efectos sobre las proteínas de la leche. Archivos Latinoamericanos de Nutrición, v.52, n.2, p.1-13, 2002.
CRUDDEN, A.; FOX, P.F.; KELLY, A.L. Factors affecting the hydrolytic action of plasmin in milk. International Dairy Journal, v.15, n.4,
p.305-313, 2005.
DECIMO, M.; MORANDI, S.; SALVETTI, T. et al. Characterization of gram-negative psychrotrophic bacteria isolated from Italian bulk tank milk. Journal of Food Science, v.79, n.10, p.M2081-M2090, 2014.
DE JONGHE, V.; COOREVITS, A.; VAN HOORDE, K. et al. Influence of storage conditions on the growth of Pseudomonas species in refrigerated
raw milk. Applied and Environmental Microbiology, v.77, n.2, p.460-470, 2011.
DOGAN, B.; BOOR, K.J. Genetic diversity and spoilage potentials among Pseudomonas spp. isolated from fluid milk products and dairy
processing plants. Applied and Environmental Microbiology, v.69, n.1,
p.130-138, 2003.
ENEROTH, A.; AHRNÉ, S.; MOLIN, G. Contamination routes of Gram-negative spoilage bacteria in the production of pasteurized milk,
evaluated by randomly amplified polymorphic DNA (RAPD). International Dairy Journal, v.10, n.5-6, p.325-331, 2000.
FAIRBAIRN, D.J.; LAW, B.A. Proteinase of psychrotrophic bacteria: their production, properties, effects and control. Journal of Dairy Research, v.53, n.1, p.139-77, 1986.
FRANK, J.F.; CHRISTEN, G.L.; BULLERMAN, L.B. Tests for groups of microorganisms. In: MARSHALL, R.T. (Ed.). Standard methods
for the examination of dairy products. New York: APHA, p.271-286, 1992.
HACHANA, Y.; KRAIEM, K.; PAAPE, M.J. Effect of plasmin, milk somatic cells and psychrotrophic bacteria on casein fractions of ultra high temperature treated milk. Food Science and Technology, v.16, n.1, p.79-
86, 2010.
HORNE, D.S. Casein micelle structure: Models and muddles. Current Opinion in Colloid & amp; Interface Science, v.11, n.2-3, p.148-153, 2006.
HULL, M.E. Studies on milk proteins. II Colorimetric determination of the partial hydrolysis of the proteins in milk. Journal of Dairy Science, v.30, p.881-894, 1947.
KUMARESAN, G.; ANNALVILLI, R.; SIVAKUMAR, K. Psychrotrophic spoilage of raw milk at different temperatures of storage. Journal of Applied Sciences Research, v.3, n.11, p.1383-1387, 2007.
LAEMMLI, U.K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, v.227, n.5259, p.680-685, 1970.
LARSON, N.K.; ISMAIL, B., NIELSEN, S.S. et al. Activity of Bacillus polymyxa protease on components of the plasmin system in milk.
International Dairy Journal, v.16, n.6, p.586-592, 2006.
MARCHAND, S.; HEYLEN, K.; MESSENS, W. et al. Seasonal influence on heat-resistant proteolytic capacity of Pseudomonas lundensis and Pseudomonas fragi, predominant milk spoilers isolated from Belgian raw milk samples. Environmental Microbiology, v.11, n.2, p.467-482, 2009a.
MARCHAND, S.; VANDRIESCHE, G.; COOREVITS, A. et al. Heterogeneity of heatresistant proteases from milk Pseudomonas species. International Journal of Food Microbiology, v.133, n.1-2, p.68-77, 2009b.
MARTINS, M.L.; ARAÚJO, E.F.; MANTOVANI, H.C. et al. Detection of the apr gene in proteolytic psychrotrophic bacteria isolated from refrigerated raw milk. International Journal of Food Microbiology, v.102, n.2, p.203-
211, 2005.
MARTINS, M.L.; PINTO, C.L.O.; ROCHA, R.B. et al. Genetic diversity of Gram-negative, proteolytic, psychrotrophic bacteria isolated from
refrigerated raw milk. International Journal of Food Microbiology, v.111, n.2, p.144-148, 2006.
MORTON, R.D. Aerobic plate count. In:DOWNES, F.P. and ITO, K. (Ed.).
Compendium of Methods for the Microbiological Examination of
Foods. 4th Ed. Washington: American Public Health Association, p.63-67, 2001.
MUNSCH-ALATOSSAVA, P.; ALATOSSAVA, T. Phenotypic characterization of raw milkassociated psychrotrophic bacteria. Microbiological Research, v.161, n.4, p.334-346, 2006.
NERO, L.A.; VIÇOSA, G.N.; PEREIRA, F.E.V. Qualidade Microbiológica do leite determinada por características de produção. Ciência e Tecnologia de Alimentos, v.29, n.2, p.386-390, 2009.
NICODÈME, M.; GRILL, J.-P.; HUMBERT, G. et al. Extracellular protease activity of different Pseudomonas strains: dependence of proteolytic
activity on culture conditions. Journal of Applied Microbiology, v.99, p.641-648, 2005.
NIELSEN, S.S. Plasmin system and microbial proteases in milk: characteristics, roles and relationship. Journal of Agricultural and
Food Chemistry, v.50, n.22, p.6628-6624, 2002.
PINTO, C.L.O.; MARTINS, M.L.; VANETTI, M.C.D. Qualidade microbiológica de leite cru refrigerado e isolamento de bactérias
psicrotróficas proteolíticas. Ciência e Tecnologia de Alimentos, v.26, n.3, p.645-651, 2006.
RAATS, D.; OFFEK, M.; MINZ, D. et al. Molecular analysis of bacterial communities in raw cow milk and the impact of refrigeration on its structure and dynamics. Food Microbiology, v.28, n.3, p.465-471, 2011.
R CORE TEAM. R: A language and environment for statistical computing. 2013. In: http://www.Rproject.org/ (accessed on January 4th 2015).
SØRHAUG, T.; STEPANIAK, L. Psychrotrophs and their enzymes in milk and dairy products: Quality aspects. Trends in Food Science & Technology, v.8, n.2, p.35-41, 1997.
TEH, K.H.; FLINT, S.; Palmer, J. et al. Proteolysis produced within biofilms of bacterial isolates from raw milk tankers. International
Journal of Food Microbiology, v.157, n.1, p.28-34, 2012.
ultrahigh temperature treatment. Journal of Dairy Science, v.59, n.5, p.823-827, 1976.
BAGLINIÈRE, F.; TANGUY, G.; JARDIN, J. et al. Quantitative and qualitative variability of the caseinolytic potential of different strains of
Pseudomonas fluorescens: implications for the stability of casein micelles of UHT milks during their storage. Food Chemistry, v.135, n.4, p.2593-2603, 2012.
BRASIL. Regulamento Técnico de Produção, Identidade e Qualidade do Leite tipo A, Regulamento Técnico de Identidade e Qualidade de Leite Cru Refrigerado, Regulamento Técnico de Identidade e Qualidade de Leite Pasteurizado e Regulamento Técnico da Coleta de Leite Cru Refrigerado e seu Transporte a Granel. Instrução Normativa nº 62, de 29 de de dezembro de 2011.
Diário Oficial [da] República Federativa do Brasil, Brasília, DF: Ministério da Agricultura, Pecuária e Abastecimento: p.6-37, 2011.
BRAUN, P.; SUTHERLAND, J.P. Predictive modelling of growth and enzyme production and activity by a cocktail of Pseudomonas spp.,
Shewanella putrefaciens and Acinetobacter. International Journal of Food
Microbiology, v.86, n.3, p.271-282, 2003.
BROCK, T.D. Biology of microorganisms. 7th Ed. New Jersey: Prentice Hall, 1994, 909p.
CELESTINO, E.L.; IYER, M.; ROGINSKI, H. Reconstituted UHT-treated milk: Effects of raw milk, powder quality and storage conditions of
UHT milk on its physico-chemical attributes and flavour. International Dairy Journal, v.7, n.2-3, p.129-140, 1997.
CHEN, L.; DANIEL, R.M.; COOLBEAR, T. Detection and impact of protease and lipase activities in milk and milk powders. International Dairy Journal, v.13, n.4, p.255-275, 2003.
COSTA, L.M.; GÓMEZ, F.S.; MOLINA, L.H.C. et al. Purificación y caracterización de proteasas de Pseudomonas fluorescens y sus efectos sobre las proteínas de la leche. Archivos Latinoamericanos de Nutrición, v.52, n.2, p.1-13, 2002.
CRUDDEN, A.; FOX, P.F.; KELLY, A.L. Factors affecting the hydrolytic action of plasmin in milk. International Dairy Journal, v.15, n.4,
p.305-313, 2005.
DECIMO, M.; MORANDI, S.; SALVETTI, T. et al. Characterization of gram-negative psychrotrophic bacteria isolated from Italian bulk tank milk. Journal of Food Science, v.79, n.10, p.M2081-M2090, 2014.
DE JONGHE, V.; COOREVITS, A.; VAN HOORDE, K. et al. Influence of storage conditions on the growth of Pseudomonas species in refrigerated
raw milk. Applied and Environmental Microbiology, v.77, n.2, p.460-470, 2011.
DOGAN, B.; BOOR, K.J. Genetic diversity and spoilage potentials among Pseudomonas spp. isolated from fluid milk products and dairy
processing plants. Applied and Environmental Microbiology, v.69, n.1,
p.130-138, 2003.
ENEROTH, A.; AHRNÉ, S.; MOLIN, G. Contamination routes of Gram-negative spoilage bacteria in the production of pasteurized milk,
evaluated by randomly amplified polymorphic DNA (RAPD). International Dairy Journal, v.10, n.5-6, p.325-331, 2000.
FAIRBAIRN, D.J.; LAW, B.A. Proteinase of psychrotrophic bacteria: their production, properties, effects and control. Journal of Dairy Research, v.53, n.1, p.139-77, 1986.
FRANK, J.F.; CHRISTEN, G.L.; BULLERMAN, L.B. Tests for groups of microorganisms. In: MARSHALL, R.T. (Ed.). Standard methods
for the examination of dairy products. New York: APHA, p.271-286, 1992.
HACHANA, Y.; KRAIEM, K.; PAAPE, M.J. Effect of plasmin, milk somatic cells and psychrotrophic bacteria on casein fractions of ultra high temperature treated milk. Food Science and Technology, v.16, n.1, p.79-
86, 2010.
HORNE, D.S. Casein micelle structure: Models and muddles. Current Opinion in Colloid & amp; Interface Science, v.11, n.2-3, p.148-153, 2006.
HULL, M.E. Studies on milk proteins. II Colorimetric determination of the partial hydrolysis of the proteins in milk. Journal of Dairy Science, v.30, p.881-894, 1947.
KUMARESAN, G.; ANNALVILLI, R.; SIVAKUMAR, K. Psychrotrophic spoilage of raw milk at different temperatures of storage. Journal of Applied Sciences Research, v.3, n.11, p.1383-1387, 2007.
LAEMMLI, U.K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature, v.227, n.5259, p.680-685, 1970.
LARSON, N.K.; ISMAIL, B., NIELSEN, S.S. et al. Activity of Bacillus polymyxa protease on components of the plasmin system in milk.
International Dairy Journal, v.16, n.6, p.586-592, 2006.
MARCHAND, S.; HEYLEN, K.; MESSENS, W. et al. Seasonal influence on heat-resistant proteolytic capacity of Pseudomonas lundensis and Pseudomonas fragi, predominant milk spoilers isolated from Belgian raw milk samples. Environmental Microbiology, v.11, n.2, p.467-482, 2009a.
MARCHAND, S.; VANDRIESCHE, G.; COOREVITS, A. et al. Heterogeneity of heatresistant proteases from milk Pseudomonas species. International Journal of Food Microbiology, v.133, n.1-2, p.68-77, 2009b.
MARTINS, M.L.; ARAÚJO, E.F.; MANTOVANI, H.C. et al. Detection of the apr gene in proteolytic psychrotrophic bacteria isolated from refrigerated raw milk. International Journal of Food Microbiology, v.102, n.2, p.203-
211, 2005.
MARTINS, M.L.; PINTO, C.L.O.; ROCHA, R.B. et al. Genetic diversity of Gram-negative, proteolytic, psychrotrophic bacteria isolated from
refrigerated raw milk. International Journal of Food Microbiology, v.111, n.2, p.144-148, 2006.
MORTON, R.D. Aerobic plate count. In:DOWNES, F.P. and ITO, K. (Ed.).
Compendium of Methods for the Microbiological Examination of
Foods. 4th Ed. Washington: American Public Health Association, p.63-67, 2001.
MUNSCH-ALATOSSAVA, P.; ALATOSSAVA, T. Phenotypic characterization of raw milkassociated psychrotrophic bacteria. Microbiological Research, v.161, n.4, p.334-346, 2006.
NERO, L.A.; VIÇOSA, G.N.; PEREIRA, F.E.V. Qualidade Microbiológica do leite determinada por características de produção. Ciência e Tecnologia de Alimentos, v.29, n.2, p.386-390, 2009.
NICODÈME, M.; GRILL, J.-P.; HUMBERT, G. et al. Extracellular protease activity of different Pseudomonas strains: dependence of proteolytic
activity on culture conditions. Journal of Applied Microbiology, v.99, p.641-648, 2005.
NIELSEN, S.S. Plasmin system and microbial proteases in milk: characteristics, roles and relationship. Journal of Agricultural and
Food Chemistry, v.50, n.22, p.6628-6624, 2002.
PINTO, C.L.O.; MARTINS, M.L.; VANETTI, M.C.D. Qualidade microbiológica de leite cru refrigerado e isolamento de bactérias
psicrotróficas proteolíticas. Ciência e Tecnologia de Alimentos, v.26, n.3, p.645-651, 2006.
RAATS, D.; OFFEK, M.; MINZ, D. et al. Molecular analysis of bacterial communities in raw cow milk and the impact of refrigeration on its structure and dynamics. Food Microbiology, v.28, n.3, p.465-471, 2011.
R CORE TEAM. R: A language and environment for statistical computing. 2013. In: http://www.Rproject.org/ (accessed on January 4th 2015).
SØRHAUG, T.; STEPANIAK, L. Psychrotrophs and their enzymes in milk and dairy products: Quality aspects. Trends in Food Science & Technology, v.8, n.2, p.35-41, 1997.
TEH, K.H.; FLINT, S.; Palmer, J. et al. Proteolysis produced within biofilms of bacterial isolates from raw milk tankers. International
Journal of Food Microbiology, v.157, n.1, p.28-34, 2012.
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2014-12-30
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Pinto1, C. L. O., Machado, S. G., Cardoso, R. R., Alves, R. M., & Vanetti, M. C. D. (2014). PROTEOLYTIC POTENTIAL OF PSEUDOMONAS FLUORESCENS ISOLATED FROM REFRIGERATED RAW MILK. Revista Brasileira De Agropecuária Sustentável, 4(2). https://doi.org/10.21206/rbas.v4i2.254
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