Main Article Content
Abstract
This study aimed to isolate and identify the Bacillus licheniformis bacteria that produces the enzyme collagenase. Due to the selective nature of this enzyme in breaking down and degrading collagen, it was used to tenderize aged buffalo meat. The bacterial isolates were screened using a selective medium and then grown on collagen-agar medium to identify enzyme-producing isolates. Thirteen enzyme-producing isolates were obtained based on the formation of a transparent corona around the colony. Biochemical assessments of the bacterial isolates were carried out using the ABIS online program. The isolates BL1–BL9, BL12 and BL13 were found to be related to B. licheniformis with a percentage match of 98%, while BL10 and BL11 were related to Bacillus pumilus and Bacillus subtilis with a percentage match of 95% and 95.3%, respectively. The enzyme activity of the isolates varied, with the highest activity reaching 200.71 units.ml-1 for isolate BL12. The enzyme of this isolate was chosen to complete the other tests. The optimal time required to obtain the highest enzymatic activity was found to be 20 h, which amounted to 210.25 units.ml-1, with a specific activity of 135.64 units.ml-1. The enzyme was partially purified using ammonium sulphate at a saturation rate of 20–70% as it showed an enzymatic activity and specificity of 355.81 units.ml-1 and a specific activity of 423.58 units.ml-1. Partially purified collagenase was used to tenderize aged buffalo meat. The results showed that treating aged buffalo meat with the enzyme solution for different immersion durations improved the pH, water-holding capacity (WHC) and cooking yield, with an increase in the concentration of hydroxyproline and a significant percentage of soluble collagen; the treated samples showed the best properties with a 60-min immersion duration.
Keywords
Article Details
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.
References
- Abdeldaim, M. H., & Ali, H. G. M. (2013). Tenderization of camel meat by using fresh ginger (Zingiber officinale) extract. Food Science and Quality Management, 21, 12-25.
- https://iiste.org/Journals/index.php/FSQM/article/view/10470
- Adinarayana, K., Ellaiah, P., & Prasad, D. S. (2003). Purification and partial characterization of thermostable serine alkaline protease from a newly isolated Bacillus subtilis PE-11. An Official Journal of the American Association of Pharmaceutical Scientists, 4(4), E56.
- https://link.springer.com/article/10.1208/pt040456
- Al-Abadi, H. H. M., & Al-Temimi, W. K. A. (2022). Production of hydrolyzed collagen from common carp Cyprinus carpio L. scales by synergistic enzyme systems. Basrah Journal of Agricultural Sciences, 35(1), 94-105.
- https://doi.org/10.37077/25200860.2022.35.1.08
- Al-Temimi, W. K. A. (2014). Purification and Characterization of Collagenase Extracted from Fish Stomach by Products and Study it’s Tenderizing Meat. Ph. D. Thesis, Univ. Basrah: 151pp. (In Arabic, English abstract).
- Anandh, M. A. (2013). Effect of Different Tenderizers on Tenderness and Quality of Buffalo Tripe. Asian Journal. Dairy and Food Research, 32(2), 144-148.
- https://arccjournals.com/journal/asian-journal-of-dairy-and-food-research/ARCC347
- Antonio, B. (2005). Buffalo Production and Research. FAO. 00100 Rome, 316pp.
- AOAC. (2016). Official methods of anaylsis. Retrieved from http://www.eoma.aoac.org/metho ds/info.asp?ID=16264
- Badr, H. M. (2008). Tenderness properties and microbial safety of spent hen meat treated by papain and gamma irradiation. Journal of Radiation Research and Applied Sciences, 1(2), 443-462.
- https://www.sciencedirect.com/journal/journal-of-radiation-research-and-applied-sciences
- Banoon, S. R., & Ali, Z. M. (2018). Identification the local thermophilic strains of Bacillus licheniformis by VITEK-2 system and Gyrase B gene. Al-Kufa University Journal for Biology, 10(2), 157-167.
- https://journal.uokufa.edu.iq/index.php/ajb/article/view/8125
- Bassetto, F., Maschio, N., Abatangelo, G., Zavan, B., Scarpa, C., & Vindigni, V. ( 2016). Collagenase from Vibrio alginolyticus cultures: Experimental study and clinical perspectives. Annals of Surgical Innovation and Research, 23(6), 557-562.
- https://doi.org/10.1177/1553350616660630
- Bergeyer, H. U. (1974). Methods of Enzymatic Analysis, Vol. 2, Second Edition, Academic Press, Elsevier Inc., 682pp.
- https://www.sciencedirect.com/book/9780120913022/methods-of-enzymatic-analysis
- Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72, 248-254.
- https://doi.org/10.1016/0003-2697(76)90527-3
- Dumitru, M., Vodnar, D. C., Elemer, S., Ciurescu, G., Habeanu, M., Sorescu, I., Georgescu, S. E., & Dudu, A. (2021). Evaluation of non-encapsulated and microencapsulated lactic acid bacteria. Applied Science, 11(9867), 1-15.
- https://doi.org/10.3390/app11219867
- Feng, Y. H., Zhang, S. S, Sun, B. Z., Wen, P. X. K. X., & Xu, C. C. (2020). Changes in physical meat traits, protein solubility, and the microstructure of different beef muscles during post-mortem aging. Foods, 806(9), 1-10.
- https://doi.org/10.3390/foods9060806
- Gecgel, U., Yilmaz, I., Soysal, M. I., Gurcan, E. K., & Kok, S. (2019). Investigating proximate composition and fatty acid profile of Longissimus dorsi from Anatolian water Buffaloes (Bubalus bubalis) raised in similar conditions. Food Science and Technology, 39(4), 830-836.
- https://doi.org/10.1590/fst.08918
- Geles, K., Poschl, E., & Aiagner, T. (2003). Collagens-structure, function, and biosynthesis. Advanced Drug Delivery Reviews, 28, 1531-1546.
- https://doi.org/10.1016/j.addr.2003.08.002
- Habtu, E., Mekonnen, B., Kiros, H., Fesseha, H., & Getachew, B. (2020). Meat tenderization of efficiency of papain, bromelain and Zingiber officinale on old aged beef carcass of local zebu cattle. Journal of Trends in Technical & Scientfic Research, 4(1), 9-15.
- https://doi.org/10.19080/TTSR.2020.04.555628
- Irurueta, M., Cadoppi, A., Langman, L., Grigioni, G., & Carduza, F. (2008). Effect of aging on meat characteristics from water buffalo grown in delta del Paraná in Argentina. Meat Science, 79(3), 529-533.
- https://doi.org/10.1016/j.meatsci.2007.12.010
- Jin, P., Zhang, L., Yuan, P., Kang, Z., Du, G., & Chen, J. (2016). Efficient biosynthesis of polysaccharides chondroitin and heparosan by metabolically engineered Bacillus sublitis. Carbohydrate Polymers, 140, 424-432.
- https://doi.org/10.1016/j.carbpol.2015.12.065
- Kandeepan, G., Anjaneyulu, A. S. R., Kondaiah, N., Mendiratta, S. K., & Lakshmanan, V. (2009). Effect of age and gender on the processing characteristics of buffalo meat. Meat Science, 83, 10-14.
- https://doi.org/10.1016/j.meatsci.2009.03.003
- Liu, L., Ma, M., Cai, Z., Yang, X., & Wang, W. (2010). Purification and properties of a collagenolytic protease produced by Bacillus cereus MBL13 strain. Food Technology and Biotechnology, 48(2), 151-160.
- https://hrcak.srce.hr/53624
- Moon, S. S. (2018). Effect of proteolytic enzymes and ginger extract on tenderization of M. pectoralis profundus from Holstein steer. Korean Journal for Food Science of Animal Resources, 38(1), 143-151.
- https://doi.org/10.5851/kosfa.2018.38.1.143
- Naveena, B. M., Mendiratta, S. K., & Anjaneyulu, A. S. R. (2004). Tenderization of buffalo meat using plant proteases from Cucumis trigonus roxb (Kachri) and Zingiber officinale roscoe (Ginger rhizome). Meat Science, 68, 363-369.
- https://doi.org/10.1016/j.meatsci.2004.04.004
- Naveena, B. M., Kiran, M., Sudhakar, K. R., Ramkrishna, C., Vaithiyanathan, S., & Devetkal, S. K. (2011). Effect of ammonium hydroxide on ultrastructure and tenderness of buffalo meat. Meat Science, 88, 727-732.
- https://doi.org/10.1016/j.meatsci.2011.03.005
- Parte, A. C. (2018). LPSN-list of prokaryotic names with standing in nomenclature (bactrio.net), 20 years on. International Journal of Systematic and Evolutionary Microbiology, 68(6), 1825-1829. https://10.1099/ijsem.0.002786
- Quaglia, G. B., & Gennaro, L. (2003). Enzymes: Uses in Food Processing. Pp., 2125-2139. In Encyclopedia of Food Sciences and Nutrition. Second Edition.
- https://doi.org/10.1016/B0-12-227055-X/00420-X
- Sartika, D., Natsir, H., Dali, S., & Leliani, L. (2019). Production and characterization of collagenase from Bacillus sp. 6-2 isolated from fish liquid waste. Jurnal Akta Kimia Indonesia, 2(1), 58-66.
- https://doi.org/10.20956/ica.v12i1.5924
- Savita, K., & Arachana, P. (2015). Production of collagenase by Bacillus KM369985 isolated from leather sample. International Journal of Research in Biosciences, 4(4), 81-87.
- http://www.ijrbs.in/index.php/ijrbs/article/view/177
- Schroeter, R., Hoffmann, T., Voigt, B., Meyer, H., Bleisteiner, M., Muntel, J., Jurgen, B., Albrecht, D., Becher, D., Lalk, M., Evers, S., Bongaerts, J., Maurer, K. H., Putzer, H., Hecker, M., Schweder, T., & Bremer, E. (2013). Stress responses of the industrial workhorse Bacillus licheniformis to osmotic challenges. PLOS One, 8(11), 1-22.
- https://doi.org/10.1371/journal.pone.0080956
- Singh, P. K., Ahlawat, S. S., Sharma, D. P., & Pathera, A. (2018). Carcass characteristics of male buffalo calf & meat quality of its veal. Buffalo Bulletin, 37(2), 129-144.
- https://kuojs.lib.ku.ac.th/index.php/BufBu/article/view/134
- Sorushanova, A., Delgado, L. M., Wu, Z., Shologu, N., Kshirsagar, A., Raghunath, R., Mullen, A. M., Bayon, Y., Pandit, A., Raghunath, M., & Zeugolis, D. I. (2018). The collagen suprafamily: From biosynthesis to advanced biomaterial development. Advanced Materials, 31(1), 1801651-1801690.
- https://doi.org/10.1002/adma.201801651
- Stoica, C., & Sorescu, I. (2017). ABIS online – Advanced Bacterial Identification Software, an original tool for phenotypic bacterial identification. Regnum Prokaryotae.
- https://www.tgw1916.net/bacteria_Bacillus_input.php
- Tran, L. H., & Nagano, H. (2002). Isolation and characteristics of Bacillus subtilis CN2 and its collagenase production. Journal of Food Science, 67(3), 1184-1187.
- https://doi.org/10.1111/j.1365-2621.2002.tb09474.x
- Wanapat, M., Kang, S., & Phesatcha, K. (2013). Enhancing buffalo production efficiency through rumen manipulation and nutrition. Buffalo Bulletin, 32(1), 258-275.
- https://kukrdb.lib.ku.ac.th/journal/BuffaloBulletin/search_detail/result/286523
- Watanabe, K. (2004). Collagenolytic proteases from bacteria. Applied Microbiology and Biotechnology, 63, 520-526.
- https://link.springer.com/article/10.1007/s00253-003-1442-0
- Zhao, G.Y., Zhou, M. Y., Zhao, H. L., Chen, X. L., Xie, B. B., Zhang, X. Y., He, H. L., Zhou, B. C., & Zhang, Y. Z. (2012). Tenderization effect of cold-adapted collagenolytic protease MCP-01 on beef meat at low temperature and its mechanism. Food Chemistry, 134, 1738-1744.
- https://doi.org/10.1016/j.foodchem.2012.03.118
- Ziauddin, S. K., Mahendrakar, N. S., Rao, D. N., Ramesh, B. S., & Aml, A, B. L. (1994). Observations on some chemical and physical characteristics of buffalo meat. Meat Science, 37, 103-113.
- https://doi.org/10.1016/0309-1740(94)90148-1
References
Abdeldaim, M. H., & Ali, H. G. M. (2013). Tenderization of camel meat by using fresh ginger (Zingiber officinale) extract. Food Science and Quality Management, 21, 12-25.
https://iiste.org/Journals/index.php/FSQM/article/view/10470
Adinarayana, K., Ellaiah, P., & Prasad, D. S. (2003). Purification and partial characterization of thermostable serine alkaline protease from a newly isolated Bacillus subtilis PE-11. An Official Journal of the American Association of Pharmaceutical Scientists, 4(4), E56.
https://link.springer.com/article/10.1208/pt040456
Al-Abadi, H. H. M., & Al-Temimi, W. K. A. (2022). Production of hydrolyzed collagen from common carp Cyprinus carpio L. scales by synergistic enzyme systems. Basrah Journal of Agricultural Sciences, 35(1), 94-105.
https://doi.org/10.37077/25200860.2022.35.1.08
Al-Temimi, W. K. A. (2014). Purification and Characterization of Collagenase Extracted from Fish Stomach by Products and Study it’s Tenderizing Meat. Ph. D. Thesis, Univ. Basrah: 151pp. (In Arabic, English abstract).
Anandh, M. A. (2013). Effect of Different Tenderizers on Tenderness and Quality of Buffalo Tripe. Asian Journal. Dairy and Food Research, 32(2), 144-148.
https://arccjournals.com/journal/asian-journal-of-dairy-and-food-research/ARCC347
Antonio, B. (2005). Buffalo Production and Research. FAO. 00100 Rome, 316pp.
AOAC. (2016). Official methods of anaylsis. Retrieved from http://www.eoma.aoac.org/metho ds/info.asp?ID=16264
Badr, H. M. (2008). Tenderness properties and microbial safety of spent hen meat treated by papain and gamma irradiation. Journal of Radiation Research and Applied Sciences, 1(2), 443-462.
https://www.sciencedirect.com/journal/journal-of-radiation-research-and-applied-sciences
Banoon, S. R., & Ali, Z. M. (2018). Identification the local thermophilic strains of Bacillus licheniformis by VITEK-2 system and Gyrase B gene. Al-Kufa University Journal for Biology, 10(2), 157-167.
https://journal.uokufa.edu.iq/index.php/ajb/article/view/8125
Bassetto, F., Maschio, N., Abatangelo, G., Zavan, B., Scarpa, C., & Vindigni, V. ( 2016). Collagenase from Vibrio alginolyticus cultures: Experimental study and clinical perspectives. Annals of Surgical Innovation and Research, 23(6), 557-562.
https://doi.org/10.1177/1553350616660630
Bergeyer, H. U. (1974). Methods of Enzymatic Analysis, Vol. 2, Second Edition, Academic Press, Elsevier Inc., 682pp.
https://www.sciencedirect.com/book/9780120913022/methods-of-enzymatic-analysis
Bradford, M. M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72, 248-254.
https://doi.org/10.1016/0003-2697(76)90527-3
Dumitru, M., Vodnar, D. C., Elemer, S., Ciurescu, G., Habeanu, M., Sorescu, I., Georgescu, S. E., & Dudu, A. (2021). Evaluation of non-encapsulated and microencapsulated lactic acid bacteria. Applied Science, 11(9867), 1-15.
https://doi.org/10.3390/app11219867
Feng, Y. H., Zhang, S. S, Sun, B. Z., Wen, P. X. K. X., & Xu, C. C. (2020). Changes in physical meat traits, protein solubility, and the microstructure of different beef muscles during post-mortem aging. Foods, 806(9), 1-10.
https://doi.org/10.3390/foods9060806
Gecgel, U., Yilmaz, I., Soysal, M. I., Gurcan, E. K., & Kok, S. (2019). Investigating proximate composition and fatty acid profile of Longissimus dorsi from Anatolian water Buffaloes (Bubalus bubalis) raised in similar conditions. Food Science and Technology, 39(4), 830-836.
https://doi.org/10.1590/fst.08918
Geles, K., Poschl, E., & Aiagner, T. (2003). Collagens-structure, function, and biosynthesis. Advanced Drug Delivery Reviews, 28, 1531-1546.
https://doi.org/10.1016/j.addr.2003.08.002
Habtu, E., Mekonnen, B., Kiros, H., Fesseha, H., & Getachew, B. (2020). Meat tenderization of efficiency of papain, bromelain and Zingiber officinale on old aged beef carcass of local zebu cattle. Journal of Trends in Technical & Scientfic Research, 4(1), 9-15.
https://doi.org/10.19080/TTSR.2020.04.555628
Irurueta, M., Cadoppi, A., Langman, L., Grigioni, G., & Carduza, F. (2008). Effect of aging on meat characteristics from water buffalo grown in delta del Paraná in Argentina. Meat Science, 79(3), 529-533.
https://doi.org/10.1016/j.meatsci.2007.12.010
Jin, P., Zhang, L., Yuan, P., Kang, Z., Du, G., & Chen, J. (2016). Efficient biosynthesis of polysaccharides chondroitin and heparosan by metabolically engineered Bacillus sublitis. Carbohydrate Polymers, 140, 424-432.
https://doi.org/10.1016/j.carbpol.2015.12.065
Kandeepan, G., Anjaneyulu, A. S. R., Kondaiah, N., Mendiratta, S. K., & Lakshmanan, V. (2009). Effect of age and gender on the processing characteristics of buffalo meat. Meat Science, 83, 10-14.
https://doi.org/10.1016/j.meatsci.2009.03.003
Liu, L., Ma, M., Cai, Z., Yang, X., & Wang, W. (2010). Purification and properties of a collagenolytic protease produced by Bacillus cereus MBL13 strain. Food Technology and Biotechnology, 48(2), 151-160.
Moon, S. S. (2018). Effect of proteolytic enzymes and ginger extract on tenderization of M. pectoralis profundus from Holstein steer. Korean Journal for Food Science of Animal Resources, 38(1), 143-151.
https://doi.org/10.5851/kosfa.2018.38.1.143
Naveena, B. M., Mendiratta, S. K., & Anjaneyulu, A. S. R. (2004). Tenderization of buffalo meat using plant proteases from Cucumis trigonus roxb (Kachri) and Zingiber officinale roscoe (Ginger rhizome). Meat Science, 68, 363-369.
https://doi.org/10.1016/j.meatsci.2004.04.004
Naveena, B. M., Kiran, M., Sudhakar, K. R., Ramkrishna, C., Vaithiyanathan, S., & Devetkal, S. K. (2011). Effect of ammonium hydroxide on ultrastructure and tenderness of buffalo meat. Meat Science, 88, 727-732.
https://doi.org/10.1016/j.meatsci.2011.03.005
Parte, A. C. (2018). LPSN-list of prokaryotic names with standing in nomenclature (bactrio.net), 20 years on. International Journal of Systematic and Evolutionary Microbiology, 68(6), 1825-1829. https://10.1099/ijsem.0.002786
Quaglia, G. B., & Gennaro, L. (2003). Enzymes: Uses in Food Processing. Pp., 2125-2139. In Encyclopedia of Food Sciences and Nutrition. Second Edition.
https://doi.org/10.1016/B0-12-227055-X/00420-X
Sartika, D., Natsir, H., Dali, S., & Leliani, L. (2019). Production and characterization of collagenase from Bacillus sp. 6-2 isolated from fish liquid waste. Jurnal Akta Kimia Indonesia, 2(1), 58-66.
https://doi.org/10.20956/ica.v12i1.5924
Savita, K., & Arachana, P. (2015). Production of collagenase by Bacillus KM369985 isolated from leather sample. International Journal of Research in Biosciences, 4(4), 81-87.
http://www.ijrbs.in/index.php/ijrbs/article/view/177
Schroeter, R., Hoffmann, T., Voigt, B., Meyer, H., Bleisteiner, M., Muntel, J., Jurgen, B., Albrecht, D., Becher, D., Lalk, M., Evers, S., Bongaerts, J., Maurer, K. H., Putzer, H., Hecker, M., Schweder, T., & Bremer, E. (2013). Stress responses of the industrial workhorse Bacillus licheniformis to osmotic challenges. PLOS One, 8(11), 1-22.
https://doi.org/10.1371/journal.pone.0080956
Singh, P. K., Ahlawat, S. S., Sharma, D. P., & Pathera, A. (2018). Carcass characteristics of male buffalo calf & meat quality of its veal. Buffalo Bulletin, 37(2), 129-144.
https://kuojs.lib.ku.ac.th/index.php/BufBu/article/view/134
Sorushanova, A., Delgado, L. M., Wu, Z., Shologu, N., Kshirsagar, A., Raghunath, R., Mullen, A. M., Bayon, Y., Pandit, A., Raghunath, M., & Zeugolis, D. I. (2018). The collagen suprafamily: From biosynthesis to advanced biomaterial development. Advanced Materials, 31(1), 1801651-1801690.
https://doi.org/10.1002/adma.201801651
Stoica, C., & Sorescu, I. (2017). ABIS online – Advanced Bacterial Identification Software, an original tool for phenotypic bacterial identification. Regnum Prokaryotae.
https://www.tgw1916.net/bacteria_Bacillus_input.php
Tran, L. H., & Nagano, H. (2002). Isolation and characteristics of Bacillus subtilis CN2 and its collagenase production. Journal of Food Science, 67(3), 1184-1187.
https://doi.org/10.1111/j.1365-2621.2002.tb09474.x
Wanapat, M., Kang, S., & Phesatcha, K. (2013). Enhancing buffalo production efficiency through rumen manipulation and nutrition. Buffalo Bulletin, 32(1), 258-275.
https://kukrdb.lib.ku.ac.th/journal/BuffaloBulletin/search_detail/result/286523
Watanabe, K. (2004). Collagenolytic proteases from bacteria. Applied Microbiology and Biotechnology, 63, 520-526.
https://link.springer.com/article/10.1007/s00253-003-1442-0
Zhao, G.Y., Zhou, M. Y., Zhao, H. L., Chen, X. L., Xie, B. B., Zhang, X. Y., He, H. L., Zhou, B. C., & Zhang, Y. Z. (2012). Tenderization effect of cold-adapted collagenolytic protease MCP-01 on beef meat at low temperature and its mechanism. Food Chemistry, 134, 1738-1744.
https://doi.org/10.1016/j.foodchem.2012.03.118
Ziauddin, S. K., Mahendrakar, N. S., Rao, D. N., Ramesh, B. S., & Aml, A, B. L. (1994). Observations on some chemical and physical characteristics of buffalo meat. Meat Science, 37, 103-113.