Antifungal Activity against Aspergillus parasiticus of Supernatants from Whey Permeates Fermented with Kefir Grains
Raúl Ricardo Gamba1,2,3, Carolina Ni Colo1, Mariana Correa1,3, Andrea Astoreca3,4, Teresa Alconada3,4, Graciela De Antoni1,2,5, Angela León Peláez1
1Cátedra de Microbiología, Facultad de Ciencias Exactas, Universidad Nacional de La Plata, La Plata, Argentine.
2CIDCA (Centro de Investigación y Desarrollo en Criotecnología de los Alimentos), La Plata, Argentine.
3CONICET (Consejo Nacional de Investigaciones Científicas y Tecnológicas), CCT-La Plata, La Plata, Argentine.
4CINDEFI (Centro de Investigación y Desarrollo en Fermentaciones Industriales), Universidad Nacional de La Plata, La Plata, Argentine.
5CIC-PBA (Comisión de Investigaciones Científicas-Provincia de Buenas Aires), Buenas Aires, Argentine.
DOI: 10.4236/aim.2015.56049   PDF   HTML   XML   3,779 Downloads   5,165 Views   Citations


Aspergillus parasiticus, a common fungal contaminant in food, produces aflatoxin B1, which is classified as human carcinogen. Kefir is an ancient fermented beverage obtained by the fermentation of different substrates with kefir grains. A very important waste produced by the dairy cheese industry is the whey permeate, which nowadays is a strong ambient contaminant. The aim of this work was to assess the effect of whey permeates fermented with kefir grains against A. parasiticus growth, aflatoxin B1 biosynthesis, and the kefir microorganisms protection against the cell damage produced by aflatoxin B1. It was observed that kefir-cell-free-supernatants (CFS) produced fungal inhibition. A fungicidal effect was observed with 65% v/v of CFS in the culture medium (final pH 4.55 and total undissociated lactic and acetic acid concentration 34.08 mM). Under these conditions, aflatoxin production was not detected. Finally, it was found that non-viable kefir microorganisms protected HepG2 cells from the damage produced by aflatoxin B1.

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Gamba, R. , Colo, C. , Correa, M. , Astoreca, A. , Alconada, T. , Antoni, G. and Peláez, A. (2015) Antifungal Activity against Aspergillus parasiticus of Supernatants from Whey Permeates Fermented with Kefir Grains. Advances in Microbiology, 5, 479-492. doi: 10.4236/aim.2015.56049.

Conflicts of Interest

The authors declare no conflicts of interest.


[1] IARC (International Agency for Research on Cancer) (2002) Some Traditional Herbal Medicines, Some Mycotoxins, Naphthalene and Styrene. IARC Monographs on the Evaluation of Carcinogenic Risks to Humans, 82, 1-556.
[2] Pitt, J. and Hocking, A. (2009) Fungi and Food Spoilage. Blackie Academic and Professional, London.
[3] Wang, J.-S., Kensler, T. and Groopman, J. (1998) Toxicants in Food: Fungal Contaminants. In: Ioannides, C., Ed., Current Toxicology Series: Nutrition and Chemical Toxicity, Wiley, New York, 29-57.
[4] Williams, J., Phillips, T., Jolly, P., Stiles, J., Jolly, C. and Aggarwal, D. (2004) Human Aflatoxicosis in Developing Countries: A Review of Toxicology, Exposure, Potential Health Consequences, and Interventions. American Journal of Clinical Nutrition, 80, 1106-1122.
[5] Kew, M. (2013) Aflatoxins as a Cause of Hepatocellular Carcinoma. Journal of Gastrointestinal and Liver Diseases, 22, 305-310.
[6] Corsetti, A., Gobbetti, M., Rossi, J. and Damiani, P. (1998) Antimould Activity of Sourdough Lactic Acid Bacteria: Identification of a Mixture of Organic Acids Produced by Lactobacillus sanfrancisco CB1. Applied Microbiology and Biotechnology, 50, 253-256.
[7] El-Nezami, H., Kankaanpaa, P., Salminen, S. and Ahokas J. (1998) Ability of Dairy Strains of Lactic Acid Bacteria to Bind a Common Food Carcinogen, Aflatoxin B1. Food and Chemical Toxicolology, 36, 321-326.
[8] Haskard, C., Binnion, C. and Ahokas, J. (2000) Factors Affecting the Sequestration of Aflatoxin by Lactobacillus rhamnosus Strain GG. Chemico-Biological Interactions, 128, 39-49.
[9] Magnusson, J. and Schnurer, J. (2001) Lactobacillus coryniformis Subsp. Coryniformis Strain Si3 Produces a Broad-Spectrum Proteinaceous Antifungal Compound. Applied and Environmental Microbiology, 67, 1-5.
[10] Sjogren, J., Magnusson, J., Broberg, A., Schnürer, J. and Kenne, L. (2003) Antifungal 3-Hydroxy Fatty Acids from Lactobacillus Plantarum MiLAB 14. Applied and Environmental Microbiology, 69, 7554-7557.
[11] Schnurer, J. and Magnusson, J. (2005) Antifungal Lactic Acid Bacteria as Biopreservatives. Trends in Food Science and Technology, 16, 70-78.
[12] Gerez, C., Torres, M., Font de Valdez, G. and Rollán, G. (2013) Control of Spoilage Fungi by Lactic Acid Bacteria. Biological Control, 64, 231-237.
[13] Abbès, S., Ben Salah-Abbès, J., Jebali, R., Ben Younes, R. and Oueslati, R. (2015) Interaction of Aflatoxin B1 and Fumonisin B1 in Mice Causes Immunotoxicity and Oxidative Stress: Possible Protective Role Using Lactic Acid Bacteria. Journal of Immunotoxicology, 1-9.
[14] Cortés-Zavaleta, O., López-Malo, A., Hernández-Mendoza, A. and García, H.S. (2014) Antifungal Activity of Lactobacilli and Its Relationship with 3-Phenyllactic Acid Production. International Journal of Food Microbiology, 173, 30-35.
[15] Miao, J., Guo, H., Ou, Y., Liu, G., Fang, X., Liao, Z., Ke, Ch., Chen, Y., Zhao, L. and Cao, Y. (2014) Purification and Characterization of Bacteriocin F1, a Novel Bacteriocin Produced by Lactobacillus paracasei subsp. Tolerans FX-6 from Tibetan Kefir, a Traditional Fermented Milk from Tibet, China. Food Control, 42, 48-53.
[16] Armando, M., Dogi, C., Pizzolitto, R., Escobar, F., Peirano, M., Salvano, M., Sabini, L., Combina, M., Dalcero, A. and Cavaglieri, L. (2011) Saccharomyces cerevisiae Strains from Animal Environment with Aflatoxin B1 Detoxification Ability and Anti Pathogenic Bacteria Influence. World Mycotoxin Journal, 4, 59-68.
[17] Pizzolitto, R., Armando, M., Combina, M., Cavaglieri, L., Dalcero, A. and Salvano, M. (2012) Evaluation of Saccharomyces cerevisiae Strains as Probiotic Agent with Aflatoxin B1 Adsorption Ability for Use in Poultry Feedstuffs. Journal of Environmental Science and Health, Part B, 47, 933-941.
[18] Yang, Y., Shevchenko, A., Knaust, A., Abuduresule, I., Li, W., Xingjun, H., Changsui, X., Wang, Ch. and Shevchenko, A. (2014) Proteomics Evidence for Kefir Dairy in Early Bronze Age China. Journal of Archaeological Science, 45, 178-186.
[19] Garrote, G., Abraham, A. and De Antoni, G. (2001) Chemical and Microbiological Characterization of Kefir Grains. Journal of Dairy Research, 68, 639-650.
[20] Garrote, G., Abraham, A. and De Antoni, G. (2000) Inhibitory Power of Kefir: The Role of Organic Acids. Journal of Food Protection, 63, 364-369.
[21] Londero, A., Quinta, R., Abraham, A., Sereno, R., De Antoni, G. and Garrote, G. (2011) Inhibitory Activity of Cheese Whey Fermented with Kefir Grains. Journal Food Protection, 74, 94-100.
[22] Londero, A., Iraporda, C., Garrote, G. and Abraham, A. (2015) Cheese Whey Fermented with Kefir Micro-Organisms: Antagonism against Salmonella and Immunomodulatory Capacity. International Journal of Dairy Technology, 68, 118-126.
[23] Correa Franco, M., Golowczyc, M.A., De Antoni, G.L., Perez, P.F., Humen, M. and Serradell, M.A. (2013) Administration of Kefir-Fermented Milk Protects Mice against Giardia Intestinalis Infection. Journal of Medical Microbiology, 62, 1815-1822.
[24] Londero, A., León, A., Diosma, G., De Antoni, G., Abraham, A. and Garrote, G. (2014) Fermented Whey as Poultry Feed Additive to Prevent Fungal Contamination. Journal of the Science of Food and Agriculture, 94, 3189-3194.
[25] Caro, A. and León, A. (2014) Fungal Growth Inhibition of Aspergillus ochraceus with “Panela” Fermented with Water Kefir Grains. Vitae Colombia, 21, 191-200.
[26] Speranza, J. (2011) Ecosuero con valor agregado. Noticiero Tecnológico de la Región Centro. INTI. Ministerio de Industria. Presidencia de la Nación N. 16. Octubre de 2011.
[27] Feijoo, G., Moreira, M.T., Roca, E. and Lema, J.M. (1999) Use of Cheese Whey as a Substrate to Produce Manganese Peroxidase by Bjerkandera sp BOS55. Journal of Industrial Microbiology, 23, 86-90.
[28] Mann, E.J. (1980) Alcohols from Whey. Dairy Industries International, 3, 47-48.
[29] Maullu, C., Lampis, G., Basile, T., Infianni, A., Rossolini, G.M. and Pompei, R. (1999) Production of Lysozyme-Enriched Biomass from Cheese Industry By-Products. Journal of Applied Microbiology, 86, 182-186.
[30] Mehaia, M.A., Cheryan, M. (1986) Lactic Acid from Acid Whey Permeate in a Membrane Recycle Bioreactor. Enzyme and Microbial Technology, 8, 289-292.
[31] Rech, R., Cassini, C.F., Secchi, A. and Ayub, M.A.S. (1999) Utilization of Protein-Hydrolyzed Cheese Whey for Production of B-Galactosidase by Kluyveromyces marxianus. Journal of Industrial Microbiology and Biotechnology, 23, 91-96.
[32] Horton, B. (1996) Wheys of Recovery. Dairy Industry International, 61, 39-40.
[33] Molina, M. and Giannuzzi, L. (1999) Combined Effect of Temperature and Propionic Acid Concentration on the Growth of Aspergillus parasiticus. Food Research International, 32, 677-682.
[34] León, A., Quintero, E., Serna, A., Gamba, R., De Antoni, G. and Giannuzzi, L. (2012) Inhibitory Activity of Lactic and Acetic Acid on Aspergillus flavus Growth for Food Preservation. Food Control, 24, 177-183.
[35] Gutierrez-Ruiz, M.C., Quiroz, S., Souza, B., Bucio, L., Hernandez, E., Olivares, I.P., Llorente, L., Vargas-Vorackova, F. and Kershenobich, D. (1999) Cytokines, Growth Factors, and Oxidative Stress in HepG2 Cells Treated with Ethanol, Acetaldehyde and LPS. Toxicology, 134, 197-207.
[36] Ou, C.C., Chiu, Y.H., Lin, S.L., Chang, Y.J., Huang, H.Y. and Lin, M.Y. (2012) Hepatoprotective Effect of Lactic Acid Bacteria in the Attenuation of Oxidative Stress from Tert-Butyl Hydroperoxide. Journal of Food and Drug Analysis, 20, 101-110.
[37] López-Malo, A., Barreto-Valdivieso, J., Palou, E. and San Martín, F. (2007) Aspergillus flavus Growth Response to Cinnamon Extract and Sodium Benzoate Mixtures. Food Control, 18, 1358-1362.
[38] Horner, K. and Anagnostopoulos, G. (1973) Combined Effects of Water Activity, pH and Temperature on the Growth and Spoilage Potential of Fungi. Journal of Applied Bacteriology, 36, 427-436.
[39] Cuppers, H., Oomes, S. and Brulm, S. (1999) A Model for the Combined Effects of Temperature and Salt Concentration on Growth Rate of Food Spoilage Molds. Applied and Environmental Microbiology, 63, 3764-3769.
[40] Mosmann, T. (1983) Rapid Colorimetric Assay for Cellular Growth and Survivals: Application to Proliferation and Cytotoxicity Assays. Journal of Immunological Methods, 65, 55-63.
[41] Kakisu, E., Irigoyen, A., Torre, P., De Antoni, G.L. and Abraham, A.G. (2011) Physicochemical, Microbiological and Sensory Profiles of Fermented Milk Containing Probiotic Strains Isolated from Kefir. Journal of Dairy Research, 78, 456-463.
[42] Toba, T. (1987) Fermented Milks Produced with Mesophilic Lactic Acid Bacteria. Kefir and Scandinavian Ropy Sour Milk. Japan Journal of Dairy Science, 36, 235-244.
[43] Ismaiel, A., Ghaly, M. and El-Naggar, A. (2011) Milk Kefir: Ultrastructure, Antimicrobial Activity and Efficacy on Aflatoxin B1 Production by Aspergillus flavus. Current Microbiology, 62, 1602-1609.
[44] Eklund, T. (1980) Inhibition of Growth and Uptake Processes in Bacteria by Some Chemical Food Preservatives. Journal of Bacteriology, 48, 423-432.
[45] AOAC (1984) Association of Official Analytical Chemistry, Official Methods of Analysis. 14th Edition, Washington DC.
[46] Park, D.L., Miller, B., Nesheim, S., Truckless, M.W., Vekich, A., Bidigare, B., McVey, J. and Brown, L.H. (1989) Visual and Semiquantitative Spectrophotometric ELISA Screening Method for Aflatoxin B1 in Corn and Peanut Products: Follow up Collaborative Study. Journal of the Association of Official Analytical Chemists, 72, 638-643.
[47] Neogen Corporation (2008) Veratox(x) HS Quantitative Aflatoxin High Sensitivity Test. USA/Canada.
[48] Rodríguez Velasco, M.L., Calonge Delso, M.M. and Ordónez Escudero, D. (2003) ELISA and HPLC Determination of the Occurrence of Aflatoxin M1 in Raw Cow’s Milk. Food Additives and Contaminants, 20, 276-280.
[49] Mc Kean, C., Tang, L., Tang, M., Billam, M., Wang, Z., Theodorakis, C., Kendall, R. and Wang, J. (2006) Comparative Acute and Combinative Toxicity of Aflatoxin B1 and Fumonisin B1 in Animals and Human Cells. Food and Chemical Toxicology, 44, 868-876.
[50] Fuch, S., Sontag, G., Stidl, R., Ehrlich, V., Kundi, M. and Knasmüller, S. (2008) Detoxification of Patulin and Ochratoxin A, Two Abundant Mycotoxins, by Lactic Acid Bacteria. Food and Chemical Toxicology, 46, 1398-1407.

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