Changes in Proteolysis in Fermented Milk Produced by Streptococcus thermophilus in Co-Culture with Lactobacillus plantarum or Bifidobacterium animalis subsp. lactis During Refrigerated Storage
Abstract
:1. Introduction
2. Results and Discussion
2.1. Changes in pH
2.2. Proteolytic Activity
2.3. Protease Activity
2.4. Aminopeptidase Activity
2.5. Free Amino Acid Content
2.6. Electrophoresis Analysis
3. Materials and Methods
3.1. Microbial Strains and Their Activation
3.2. Fermented Milks Preparation
3.3. pH Determination
3.4. Proteolysis Evaluation
3.5. Crude Enzyme Extraction
3.6. Protease Assays
3.7. Aminopeptidase Assays
3.8. Free Amino Acid Analysis
3.9. Polyacrylamide Gel Electrophoresis
3.10. Statistical Processing
4. Conclusions
Author Contributions
Funding
Conflicts of Interest
References
- Mills, O.E.; Thomas, T.D. Nitrogen sources for growth of lactic streptococci in milk. N. Z. J. Dairy Sci. Technol. 1981, 16, 43–55. [Google Scholar]
- Abu-Tarboush, H.M. Comparison of associative growth and proteolytic activity of yoghurt starters in whole milk from camels and cows. J. Dairy Sci. 1996, 79, 366–371. [Google Scholar] [CrossRef]
- Shihata, A.; Shah, N.P. Proteolytic profiles of yogurt and probiotic bacteria. Int. Dairy J. 2000, 10, 401–408. [Google Scholar] [CrossRef]
- Nielsen, S.S. Plasmin system and microbial proteases in milk: Characteristics, roles and relationship. J. Agric. Food Chem. 2002, 50, 6628–6634. [Google Scholar] [CrossRef] [PubMed]
- Caira, S.; Ferranti, P.; Gatti, M.; Fornasari, M.E.; Barone, F.; Lilla, S.; Mucchetti, G.; Picariello, G.; Chianese, L.; Neviani, E.; et al. Synthetic peptides as substrate for assaying the proteolytic activity of Lactobacillus helveticus. J. Dairy Res. 2003, 70, 315–325. [Google Scholar] [CrossRef]
- Wohlrab, Y.; Bockelmann, W. Purification and characterization of a dipeptidase from Lactobacillus delbrueckii subsp. bulgaricus. Int. Dairy J. 1992, 2, 345–361. [Google Scholar] [CrossRef]
- Yang, M.; Fu, J.; Li, L. Rheological characteristics and microstructure of probiotic soy yogurt prepared from germinated soybeans. Food Technol. Biotechnol. 2012, 50, 73–80. [Google Scholar]
- Farahat, A.M.; EI-Batawy, O.I. Proteolytic activity and some properties of stirred fruit yoghurt made using some fruits containing proteolytic enzymes. World J. Dairy Food Sci. 2013, 8, 38–44. [Google Scholar]
- Kunji, E.R.S.; Mierau, I.; Hagting, A.; Poolman, B.; Konings, W.N. The proteolytic systems of lactic acid bacteria. Antonie van Leeuwenhoek 1996, 70, 187–221. [Google Scholar] [CrossRef]
- Fox, P.F. Developments in biochemistry of cheese ripening. 25th International Dairy Congress, Aarhus, Denmark, Sept. 21-24. Progress of Dairy Science and Technology, 1998, 11-37.
- Moslehishad, M.; Ehsani, M.R.; Salami, M.; Mirdamadi, S.; Ezzatpanah, H.; Niasari-Naslaji, A.; Moosavi-Movahedi, A.A. The comparative assessment of ACE-inhibitory and antioxidant activities of peptide fractions obtained from fermented camel and bovine milk by Lactobacillus rhamnosus PTCC 1637. Int. Dairy J. 2013, 29, 82–87. [Google Scholar] [CrossRef]
- El-Fattah, A.A.; Sakr, S.; El-Dieb, S.; Elkashef, H. Angiotensin-converting enzyme inhibition and antioxidant activity of commercial dairy starter cultures. Food Sci. Biotechnol. 2016, 25, 1745–1751. [Google Scholar] [CrossRef] [PubMed]
- Oberman, H.; Libudisz, Z. Fermented milks. In Microbiology of Fermented Foods, 2nd ed.; Wood, B.J.B., Ed.; Blackie Academic and Professional: London, UK, 1998; Volume 1, pp. 308–350. [Google Scholar]
- Ávila, M.; Garde, S.; Medina, M.; Nuñez, M. Effect of milk inoculation with bacteriocin-producing lactic acid bacteria on a Lactobacillus helveticus adjunct cheese culture. J. Food Prot. 2005, 68, 1026–1033. [Google Scholar] [PubMed]
- Lourens-Hattingh, A.; Viljeon, C.B. Yoghurt as probiotic carrier food. Int. Dairy J. 2001, 11, 1–17. [Google Scholar] [CrossRef]
- Klaver, F.A.M.; Kingma, F.; Weerkamp, A.H. Growth and survival of bifidobacteria in milk. Neth. Milk Dairy J. 1993, 47, 151–164. [Google Scholar]
- Settachaimongkon, S.; van Valenberg, H.J.F.; Gazi, I.; Robert Nout, M.J.; van Hooijdonk, T.C.M.; Zwietering, M.H.; Smid, E.J. Influence of Lactobacillus plantarum WCFS1 on post-acidification, metabolite formation and survival of starter bacteria in set-yoghurt. Food Microbiol. 2016, 59, 14–22. [Google Scholar] [CrossRef] [PubMed]
- Casarotti, S.N.; Monteiro, D.A.; Moretti, M.M.S.; Pennaauthor, A.L.B. Influence of the combination of probiotic cultures during fermentation and storage of fermented milk. Food Res. Int. 2014, 59, 67–75. [Google Scholar] [CrossRef]
- Li, S.N.; Tang, S.H.; He, Q.; Gong, J.X.; Hu, J.X. Physicochemical, textural and volatile characteristics of fermented milk co-cultured with Streptococcus thermophilus, Bifidobacterium animalis or Lactobacillus plantarum. Int. J. Food Sci. Technol. 2019. [Google Scholar] [CrossRef]
- Macedo, A.C.; Tavares, T.G.; Malcata, F. Purification and characterization of an intracellular aminopeptidase from a wild strain of Lactobacillus plantarum isolated from traditional Serra da Estrela cheese. Enzyme Microb. Technol. 2003, 32, 41–48. [Google Scholar] [CrossRef]
- Merry, R.J.; Winters, A.L.; Thomas, P.I.; Mueller, M.; Mueller, T. Degradation of fructans by epiphytic and inoculated lactic acid bacteria and by plant enzymes during ensilage of normal and sterile hybrid ryegrass. J. Appl. Bacteriol. 1985, 79, 583–591. [Google Scholar] [CrossRef]
- Tian, H.; Shen, Y.; Yu, H.; Chen, C. Effects of 4 probiotic strains in coculture with traditional starters on the flavor profile of yogurt. J. Food Sci. 2017, 82, 1693–1701. [Google Scholar] [CrossRef]
- Kailasapathy, K. Survival of free and encapsulated probiotic bacteria and their effect on the sensory properties of yoghurt. LWT-Food Sci. Technol. 2006, 39, 1221–1227. [Google Scholar] [CrossRef]
- Khalid, N.M.; Marth, E.H. Proteolytic activity by strains of Lactobacillus plantarum and Lactobacillus casei. J. Dairy Sci. 1990, 73, 3068–3076. [Google Scholar] [CrossRef]
- Moslehishad, M.; Mirdamadi, S.; Ehsani, M.R.; Ezzatpanah, H.; Moosavi-Movahedi, A.A. The proteolytic activity of selected lactic acid bacteria in fermenting cow’s and camel’s milk and the resultant sensory characteristics of the products. Int. J. Dairy Technol. 2013, 66, 279–285. [Google Scholar] [CrossRef]
- Pritchard, G.G.; Coolbear, T. The physiology and biochemistry of the proteolytic system in lactic acid bacteria. FEMS Microbiol. Rev. 1993, 12, 179–206. [Google Scholar] [CrossRef]
- Zakharov, A.; Carchilan, M.; Stepurina, T.; Rotari, V.; Wilson, K.; Vaintraub, I. A comparative study of the role of the major proteinases of germinated common bean (Phaseolus vulgaris L.) and soybean (Glycine max (L.) Merrill) seeds in the degradation of their storage proteins. J. Exp. Bot. 2004, 55, 2241–2249. [Google Scholar] [CrossRef]
- Ohmiya, K.; Sato, Y. Studies on the proteolytic action of dairy lactic acid bacteria. Agric. Biol. Chem. 1969, 33, 1628–1635. [Google Scholar]
- Gilbert, C.; Atlan, D.; Blanc, B.; Protaleir, R. Proline iminopeptidase form Lactobacillus delbrueckii subsp. bulgaricus CNRZ397: purification and characterization. Microbiology 1994, 140, 537–542. [Google Scholar] [CrossRef]
- Cheng, C.C.; Nagasawa, T. Effect of peptides and amino acids produced by Lactobacillus casei in milk on the acid production of bifidobacteria. Jpn. J. Zootech. Sci. 1984, 55, 339–349. [Google Scholar]
- Damir, A.A.; Salama, A.A.; Mohamed, M.S. Acidity, microbial, organic and free amino acids development during fermentation of skimmed milk Kishk. Food Chem. 1992, 43, 265–269. [Google Scholar] [CrossRef]
- Tongnual, P.; Nanson, N.J.; Fields, M.L. Effect of proteolytic bacteria in the natural fermentation of corn to increase its nutritive value. J. Food Sci. 1981, 46, 100–109. [Google Scholar] [CrossRef]
- Savijoki, K.; Ingmer, H.; Varmanen, P. Proteolytic systems of lactic acid bacteria. Appl. Microbiol. Biotechnol. 2006, 71, 394–406. [Google Scholar] [CrossRef] [PubMed]
- González-Olivares, L.G.; Añorve-Morga, J.; Castañeda-Ovando, A.; Contreras-López, E.; Jaimez-Ordaz, J. Peptide separation of commercial fermented milk during refrigerated storage. Food Sci. Technol. 2014, 34, 674–679. [Google Scholar] [CrossRef] [Green Version]
- Ghosh, D.; Chattoraj, D.K.; Chattopadhyay, P. Studies on changes in microstructure and proteolysis in cow and soy milk curd during fermentation using lactic cultures for improving protein bioavailability. J. Food Sci. Technol. 2013, 50, 979–985. [Google Scholar] [CrossRef] [PubMed]
- Chang, C.H.; Kong, B.H.; Zhao, X.H.; Zhao, X.H. Quality attributes of the set-style yoghurt from whole bovine milk as affected by an enzymatic oxidative cross-linking. CyTA–J. Food 2014, 12, 249–255. [Google Scholar] [CrossRef]
- Amirdivani, S.; Baba, A.S. Changes in yogurt fermentation characteristics, and antioxidant potential and in vitro inhibition of angiotensin-1 converting enzyme upon the inclusion of peppermint, dill and basil. LWT-Food Sci. Technol. 2011, 44, 1458–1464. [Google Scholar] [CrossRef] [Green Version]
- Church, F.C.; Swaisgood, H.E.; Porter, D.H.; Catignani, G.L. Spectrophotometric assay using o-phthaldialdehyde for determination of proteolysis in milk and isolated milk proteins. J. Dairy Sci. 1983, 66, 1219–1227. [Google Scholar] [CrossRef]
- Ramchandran, L.; Shah, N.P. Proteolytic profiles and angiotensin-I converting enzyme and α-glucosidase inhibitory activities of selected lactic acid bacteria. J. Food Sci. 2008, 73, M75–M81. [Google Scholar] [CrossRef]
- Li, L.; Yang, Z.Y.; Yang, X.Q.; Zhang, G.H.; Tang, S.Z.; Chen, F. Debittering effect of actinomucor elegans peptidases on soybean protein hydrolysates. J. Ind. Microbiol. Biotechnol. 2008, 35, 41–47. [Google Scholar] [CrossRef]
- Fernandez-Espla, M.D.; Martin-Hernandez, M.C.; Fox, P.F. Purification and characterization of a prolidase from Lactobacillus casei subsp. casei IFPL 731. Appl. Environ. Microbiol. 1997, 63, 314–316. [Google Scholar]
- Das, P.R.; Kim, Y.; Hong, S.J.; Eun, J.B. Profiling of volatile and non-phenolic metabolites—Amino acids, organic acids, and sugars of green tea extracts obtained by different extraction techniques. Food Chem. 2019, 296, 69–77. [Google Scholar] [CrossRef]
- Laemmli, U.K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 1970, 227, 680–685. [Google Scholar] [CrossRef] [PubMed]
- Bradford, M.M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 1976, 72, 248–254. [Google Scholar] [CrossRef]
Sample Availability: Not available. |
Starter Cultures | Day 1 | Day 7 | Day 14 | Day 21 |
---|---|---|---|---|
St | 4.38 ± 0.01 aA | 4.27 ± 0.01 bA | 4.21 ± 0.01 cA | 4.19 ± 0.00 dA |
StBa | 4.32 ± 0.01 aB | 4.23 ± 0.01 bB | 4.17 ± 0.01 cB | 4.14 ± 0.01 dB |
StLp | 4.30 ± 0.02 aB | 4.21 ± 0.01 bBC | 4.15 ± 0.01 cBC | 4.11 ± 0.01 dC |
StBaLp | 4.30 ± 0.01 aB | 4.19 ± 0.01 bC | 4.14 ± 0.01 cC | 4.10 ± 0.01 dC |
Starter Cultures | Day 1 | Day 7 | Day 14 | Day 21 |
---|---|---|---|---|
St | 0.23 ± 0.01 dBC | 0.37 ± 0.01 cB | 0.49 ± 0.00 bB | 0.51 ± 0.01 aB |
StBa | 0.21 ± 0.02 cC | 0.39 ± 0.02 bA | 0.52 ± 0.01 aA | 0.53 ± 0.02 aB |
StLp | 0.25 ± 0.01 dB | 0.41 ± 0.01 cA | 0.52 ± 0.01 bA | 0.55 ± 0.01 aA |
StBaLp | 0.27 ± 0.01 dA | 0.41 ± 0.01 cA | 0.53 ± 0.01 bA | 0.56 ± 0.00 aA |
Amino Acids | St Treatment | StBa Treatment | StLp Treatment | StBaLp Treatment | ||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|
Day 1 | Day 21 | Increment | Day 1 | Day 21 | Increment | Day 1 | Day 21 | Increment | Day 1 | Day 21 | Increment | |
Asp | 1.94 ± 0.01 c | 1.14 ± 0.09 d | −0.80 | 1.92 ± 0.02 c | 1.37 ± 0.05 d | −0.55 | 2.52 ± 0.08 ab | 2.26 ± 0.35 bc | −0.26 | 2.68 ± 0.19 a | 2.46 ± 0.17 ab | −0.22 |
Thr | 1.91 ± 0.06 ab | 1.21 ± 0.11 c | −0.70 | 2.11 ± 0.03 a | 1.13 ± 0.04 cd | −0.98 | 1.78 ± 0.18 b | 1.30 ± 0.22 c | −0.48 | 1.30 ± 0.05 c | 0.84 ± 0.19 d | −0.46 |
Ser | 0.85 ± 0.00 ab | 0.70 ± 0.03 bc | −0.15 | 0.95 ± 0.03 a | 0.60 ± 0.06 cd | −0.35 | 0.73 ± 0.10 bc | 0.48 ± 0.11 de | −0.25 | 0.64 ± 0.04 c | 0.44 ± 0.08 e | −0.20 |
Glu | 2.32 ± 0.07 cd | 1.38 ± 0.17 f | −0.94 | 2.38 ± 0.02 bc | 1.15 ± 0.06 g | −1.23 | 2.60 ± 0.09 b | 2.12 ± 0.14 de | −0.48 | 3.11 ± 0.05 a | 2.07 ± 0.09 e | −1.04 |
Gly | 0.06 ± 0.00 d | 0.14 ± 0.11 d | 0.08 | 0.09 ± 0.01 d | 0.17 ± 0.02 d | 0.08 | 1.09 ± 0.02 bc | 1.73 ± 0.21 a | 0.64 | 1.06 ± 0.02 c | 1.33 ± 0.18 b | 0.27 |
Ala | 4.05 ± 0.14 bcd | 3.81 ± 0.76 cd | −0.24 | 3.97 ± 0.05 cd | 3.57 ± 0.03 d | −0.40 | 4.76 ± 0.13 b | 5.79 ± 0.15 a | 1.03 | 4.56 ± 0.16 bc | 5.61 ± 0.33 a | 1.05 |
Cys | 1.21 ± 0.01 cd | 1.00 ± 0.22 d | −0.21 | 1.16 ± 0.06 cd | 1.10 ± 0.11 d | −0.06 | 1.40 ± 0.14 bc | 1.98 ± 0.02 a | 0.58 | 1.61 ± 0.07 b | 2.20 ± 0.13 a | 0.59 |
Met | 1.20 ± 0.03 ab | 0.93 ± 0.11 bc | −0.27 | 1.26 ± 0.08 a | 1.10 ± 0.04 abc | −0.16 | 1.10 ± 0.18 abc | 0.83 ± 0.13 cd | −0.27 | 0.41 ± 0.02 e | 0.60 ± 0.21 de | 0.19 |
Val | 0.22 ± 0.01 c | 0.30 ± 0.07 b | 0.08 | 0.20 ± 0.01 c | 0.42 ± 0.02 a | 0.22 | 0.07 ± 0.01 d | 0.23 ± 0.04 bc | 0.16 | 0.07 ± 0.01 d | 0.26 ± 0.04 bc | 0.19 |
Ile | 1.40 ± 0.04 a | 0.66 ± 0.04 cd | −0.74 | 1.54 ± 0.01 a | 1.06 ± 0.01 b | −0.48 | 0.86 ± 0.20 bc | 0.52 ± 0.16 de | −0.34 | 0.18 ± 0.01 f | 0.42 ± 0.06 e | 0.24 |
Leu | 0.50 ± 0.01 a | 0.31 ± 0.02 bc | −0.19 | 0.57 ± 0.01 a | 0.48 ± 0.07 ab | −0.09 | 0.27 ± 0.09 cd | 0.41 ± 0.13 abc | 0.14 | 0.12 ± 0.00 d | 0.41 ± 0.12 abc | 0.29 |
Tyr | 0.19 ± 0.01 cd | 0.04 ± 0.01 e | −0.15 | 0.18 ± 0.00 cd | 0.03 ± 0.01 e | −0.15 | 0.26 ± 0.08 c | 0.51 ± 0.06 a | 0.25 | 0.11 ± 0.01 de | 0.38 ± 0.04 b | 0.27 |
Phe | 1.15 ± 0.01 a | 0.92 ± 0.26 a | −0.23 | 1.18 ± 0.01 a | 0.89 ± 0.11 a | −0.29 | 0.92 ± 0.07 a | 0.71 ± 0.17 a | −0.21 | 0.61 ± 0.22 a | 0.54 ± 0.12 a | −0.07 |
Lys | 1.40 ± 0.00 ab | 1.51 ± 0.05 a | 0.11 | 1.42 ± 0.06 ab | 1.47 ± 0.02 a | 0.05 | 0.93 ± 0.05 d | 1.35 ± 0.09 b | 0.42 | 0.92 ± 0.05 d | 1.06 ± 0.06 c | 0.14 |
His | 4.73 ± 0.06 c | 4.67 ± 0.54 c | −0.06 | 4.52 ± 0.11 c | 5.67 ± 0.47 b | 1.15 | 5.03 ± 0.04 bc | 7.46 ± 0.30 a | 2.43 | 4.87 ± 0.21 c | 7.85 ± 0.04 a | 2.98 |
Arg | 0.75 ± 0.08 d | 1.34 ± 0.05 b | 0.59 | 0.91 ± 0.08 cd | 1.65 ± 0.11 a | 0.74 | 0.84 ± 0.01 cd | 0.95 ± 0.06 c | 0.11 | 0.97 ± 0.10 c | 1.17 ± 0.03 b | 0.20 |
Pro | 11.04 ± 0.12 cd | 6.02 ± 0.95 f | −5.02 | 11.67 ± 0.06 bc | 7.52 ± 0.08 e | −4.15 | 12.57 ± 0.35 a | 10.71 ± 0.01 d | −1.86 | 12.40 ± 0.11 ab | 10.19 ± 0.04 d | −2.21 |
Total | 34.89 ± 0.04 d | 26.02 ± 1.22 f | −8.87 | 36.00 ± 0.12 bcd | 29.35 ± 1.11 e | −6.65 | 37.68 ± 1.13 abc | 39.31 ± 0.76 a | 1.63 | 35.58 ± 0.35 bcd | 37.77 ± 1.26 ab | 2.19 |
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Li, S.; Tang, S.; He, Q.; Hu, J.; Zheng, J. Changes in Proteolysis in Fermented Milk Produced by Streptococcus thermophilus in Co-Culture with Lactobacillus plantarum or Bifidobacterium animalis subsp. lactis During Refrigerated Storage. Molecules 2019, 24, 3699. https://doi.org/10.3390/molecules24203699
Li S, Tang S, He Q, Hu J, Zheng J. Changes in Proteolysis in Fermented Milk Produced by Streptococcus thermophilus in Co-Culture with Lactobacillus plantarum or Bifidobacterium animalis subsp. lactis During Refrigerated Storage. Molecules. 2019; 24(20):3699. https://doi.org/10.3390/molecules24203699
Chicago/Turabian StyleLi, Sining, Shanhu Tang, Qiang He, Jiangxiao Hu, and Jing Zheng. 2019. "Changes in Proteolysis in Fermented Milk Produced by Streptococcus thermophilus in Co-Culture with Lactobacillus plantarum or Bifidobacterium animalis subsp. lactis During Refrigerated Storage" Molecules 24, no. 20: 3699. https://doi.org/10.3390/molecules24203699