Journal of food science and technology(Iran)

Journal of food science and technology(Iran)

Isolation and diagnosis of high lipolytic activity’s Enterococcus faecium and using its lipase product in beef meat degradation

Document Type : Original Research

Authors
zainab amin 1
Abdulla Mohammed 2
Hasanain Abbood 3
1 university of Busrah
2 university of Basrah
3 University of Misan
10.48311/fsct.2026.115674.0
Abstract
Microbial lipases are a very important group of enzymes used in many biotechnological industries, such as flavor enhancement technology in foods, especially meat products. Lipases are produced alone or in combination with esterase by a variety of microorganisms. In the present study, Bacterial colonies isolated from various food samples were screened for lipolytic activity by producing a bright orange halo around them on tributyrin-MRS agar and a clear zone diameter on Tributyrin agar. Of the ten colonies observed of the lipolytic activity, colony number 6 achieved the significant and highest lipase activity, maximum 23mm lipolytic zone and 46U/ml lipolytic activity. The morphological, biochemical, and genetic examinations had diagnosed the colony as Enterococcus faecium, which was registered in the gene bank under the accession number PV299116 with 99.89% identification. Lipase was produced from E. faecium in MSM medium then purified by acetone precipitation and gel chromatography. The application for adding extracted lipase to beef meat burger showed significant improvement in the meat sensory properties (appearance, colour, flavor, and overall acceptance) with values of 7.7, 7.1, 7.2, and 7.3 respectively for the longest storage time (4 weeks) at p≤ 0.001.
 
Keywords
Enterococcus faecium
lipolytic activity
meat degradation

Subjects

]1] Edwards M, Mohiuddin SS. 2023. Biochemistry, Lipolysis. In: Stat Pearls [Internet]. Treasure Island (FL): Stat Pearls Publishing; 2025 Jan–. PMID: 32809399.

[2] García-Cano I, Rocha-Mendoza D, Ortega-Anaya J, Wang K, Kosmerl E, Jiménez-Flores R. 2019. Lactic acid bacteria isolated from dairy products as potential producers of lipolytic, proteolytic, and antibacterial proteins. Applied Microbiology and Biotechnology 103:pp.5243-5257. doi.org/10.1007/s00253-019-09844-6.

[3] Ali, Z.A., Niamah, A.K. and Salih, H.W., 2021 . Isolation, preparation and characterization of polylactic acid film reinforced with nano silica. In Journal of Physics: Conference Series (Vol. 2063, No. 1, p. 012028). IOP Publishing. doi:10.1088/1742-6596/2063/1/012028.

[4] Cerk, I. K., Wechselberger, L., & Oberer, M. 2018. Adipose triglyceride lipase regulation: an overview. Current Protein and Peptide Science, 19(2), 221-233. doi: 10.2174/1389203718666170918160110.

[5] Ramakrishnan, V., Narayan, B., & Halami, P. M. 2019. Lipase of Lactic Acid Bacteria: Diversity and Application. In Microbes for sustainable development and bioremediation (pp. 313-323). CRC Press. Doi: 10.1201/9780429275876-18.

[6]  Sizova, E. A., & Ryazantseva, K. V. 2022. Fats and emulsifiers in feeding broiler chickens. Sel'skohozyajstvennaya biologiya= Agricultural biology, 57(4), 664-680. Doi: 10.15389/agrobiology.2022.4.664eng.

[7]  Tanasupawat S, Phoottosavako M, Keeratipibul S. 2015. Characterization and lipolytic activity of lactic acid bacteria isolated from Thai fermented meat. J. App. Pharm. Sci. 5(03):006–012. doi.org/10.7324/JAPS.2015.50302.

[8] Graham, K., Stack, H., & Rea, R. 2020. Safety, beneficial and technological properties of enterococci for use in functional food applications–a review. Critical Reviews in Food Science and Nutrition, 60(22), 3836-3861.doi: 10.1080/10408398.2019.1709800.

[9] Ali, Z.A.,2025. Isolation and genetic diagnosis of local isolate Enterococcus faecium with evaluation of inhibitory against bacterial pathogens. 11(2), DOI:10.36582/97ksxq66.

[10] Rabbani, M., Bagherinejad, M.R., Sadeghi, H.M., Shariat, Z.S., Etemadifar, Z., Moazen, F., Rahbari, M., (...), Zaghian, S. 2013. Isolation and characterization of novel thermophilic lipase-secreting bacteria. Brazilian J. of Microbiol., 44 (4), pp. 1113-1119. doi: 10.1590/S1517-83822013000400013 http://www.scielo.br/pdf/bjm/v44n4/13.pdf.

[11] Dincer, E. And Kivanc, M. 2018. Lipolytic activity of lactic acid bacteria isolated from turkish pastirma. Anadolu univ. J. Of sci. And technology c – life sci. And biotech. 7 (1), pp. 12 - 19. DOI: 10.18036/aubtdc.306292.

[12] Zouaoui, B, Bouziane, A, Rahoghalem, B. 2012. Production, optimization, and purification of lipase from Pseudomonas aeruginosa. Afr. J. Microbiol. Res., 6 (20), pp. 4417-4423. https://doi.org/10.5897/AJMR11.1567.

[13] Mohammed,A.A.; Hussain,N.A.; Niamah,A.K. 2020. Antibacterial Spectrum of produced reuterin from new isolates of lactobacillus reuteri. J microbiol biotech food sci., 10 (1) pp.134-139. Doi: 10.15414/jmbfs.2020.10.1.134-139.

[14]  Duan, X. Zhai, W. Li, X. Wu, S. Wang, Y. Wang, L. Basang, W. Zhu, Y. and Gao,Y. 2023. Preparation, purification, and biochemical of fat degrading bacterial enzymes from pig carcass compost and its application. BMC Biotechnology. 23(48), pp.14. https://doi.org/10.1186/s12896-023-00818-1.

[15] Kulkarni,A.G. and Vedamurthy,A.B. 2017. Isolation, Characterization And Optimization Of Lipase Produced From Bacteria. International. J. Res. Public. Vol.7(3),pp.20-37.ISSN2251-1563.

 [16] Al-Mossawi, L.H.M. Mahdi, N.Z. Zwain, L.A. 2016. Higher production of lipase enzyme from different microorganisms grown in local natural culture media. Int. J. Adv. Res. Biol. Sci., 3(5): pp.232-239. ISSN: 2348-8069. SOI: http://s-o-i.org/1.15/ijarbs-2016-3-5-33.

[17] Mandepudi, D.; Mandepudi, B.; Chowdary, G.V. and Gajbhiye, A. 2012. Screening, Isolation and Biochemical Characterization of Novel Lipase Producing Bacteria from Soil Samples. International J. of Biol. Eng., 2(2): pp.18-22. DOI: 10.5923/j.ijbe.20120202.03.

[18] Alkabee , H. J. J. and AlHamdani, M. A. 2015. Purification and Characterization of Lipase produced by Pseudomonas aeruginosa isolated from some soils of Basrah Governorate. Al-Kufa University J. for Biology. vol.7 (2), pp.42-49. Print ISSN: 2073-8854. Online ISSN: 2311-6544.

[19]  Yousif, A.Y. and Muhssen, E.H. 2019. The Effect of Adding Probiotics on Microbial and Organoleptic Characteristics of Beef Burger Product.J.of Global Pharma. Technol. vol. 11(9), pp.152-157. ISSN: 0975 -8542.

[20]  Muhssen,E.H. 2016. The Use of Sumac (Rhus glabra) Plantin Prolonging the Period of Beef Meat Storage. J. of Basrah Agric.Sci. vol.29 (1), pp.26-34.

[21] SPSS. (2019). IBM Statistics 26 step by step. 16th Edition. https://doi.org/10.4324/9780429056765. 

[22] Duza, M. B. and Mastan, D. S. 2014. Optimization of lipase production from Bacillus thuringiensis (TS11BP), Achromobacter xylosoxidans J2 (TS2MCN)-isolated from soil sediments near oilseed farm.IOSR J. Pharm. Biol. Sci, 9 (2), pp. 66-76.

[23] Pitt, J I, Hocking, A D. 2009. Spoilage of stored, processed, and preserved foods. Fungi and Food Spoilage, (3rd ed., Springer, Boston, MA) pp. 401-421.

[24] Rambaut A. FigTree v1.4.2. 2014. Available at: http:// tree.bio.ed.ac.uk/ software/figtree.

[25] Sabir, S.F., Alwatar, W.M.A., Matti, B.F. 2021. Impact of Programmed Death 1 Expression in Chronic myeloid Leukemia at Different Molecular Responses to imatinib Mesylate Treatment. Biochemical and Cellular Archives, 21 (1), pp. 671-678. http://www.connectjournals.com/toc1.php?bookmark=CJ-033216WWW.CONNECTJOURNALS.COM/BCA.

 [26] Sagar, K, Bashir, Y, Phukan, M M, Konwar, B K. 2013. Isolation of lipolytic bacteria from waste contaminated soil: A study about process optimization for lipase. J. Sci. Technol. Res, 2 (10), pp. 214-218.

[27] Sharma, S., Kanwar, S.S. 2014. Organic solvent tolerant lipases and applications (2014) The Scientific World Journal, art. No. 625258. doi: 10.1155/2014/625258.

[28] Lee, L.P., Karbul, H.M., Citartan, M., Gopinath, S.C.B., Lakshmipriya, T., Tang, T. H. 2015. Lipase-secreting Bacillus species in an oil-contaminated habitat: Promising strains to alleviate oil pollution. BioMed Research International, 2015, art. no. 820575. http://www.hindawi.com/journals/biomed/ Doi: 10.1155/2015/820575.

[29] Kim E., Kim D., Yang S., Kim H. 2022. The accurate identification and quantification of six Enterococcus species using quantitative polymerase chain reaction based novel DNA markers. LWT - Food Science and Technology, 166, 113769. https://doi.org/10.1016/j.lwt.2022.113769.  

[30] Narasimha,G.; Kumar,A.P.; Subramanyam D. 2011. Production and optimization of lipase enzyme by Pseudomonas sps. BTAIJ, 5(1), pp.36-42. ISSN: 0974 – 7435.

[31] Padmapriya, B.; Rajeswari, T.; Noushida, E.; Sethupalan, D.G. and Venil, C.K. 2011. Production of Lipase Enzyme from Lactobacillus spp. and Its Application in the Degradation of Meat. World Appl. Sci. J., 12 (10): pp.1798-1802. ISSN 1818-4952.

]1] Edwards M, Mohiuddin SS. 2023. Biochemistry, Lipolysis. In: Stat Pearls [Internet]. Treasure Island (FL): Stat Pearls Publishing; 2025 Jan–. PMID: 32809399.
[2] García-Cano I, Rocha-Mendoza D, Ortega-Anaya J, Wang K, Kosmerl E, Jiménez-Flores R. 2019. Lactic acid bacteria isolated from dairy products as potential producers of lipolytic, proteolytic, and antibacterial proteins. Applied Microbiology and Biotechnology 103:pp.5243-5257. doi.org/10.1007/s00253-019-09844-6.
[3] Ali, Z.A., Niamah, A.K. and Salih, H.W., 2021 . Isolation, preparation and characterization of polylactic acid film reinforced with nano silica. In Journal of Physics: Conference Series (Vol. 2063, No. 1, p. 012028). IOP Publishing. doi:10.1088/1742-6596/2063/1/012028.
[4] Cerk, I. K., Wechselberger, L., & Oberer, M. 2018. Adipose triglyceride lipase regulation: an overview. Current Protein and Peptide Science, 19(2), 221-233. doi: 10.2174/1389203718666170918160110.
[5] Ramakrishnan, V., Narayan, B., & Halami, P. M. 2019. Lipase of Lactic Acid Bacteria: Diversity and Application. In Microbes for sustainable development and bioremediation (pp. 313-323). CRC Press. Doi: 10.1201/9780429275876-18.
[6]  Sizova, E. A., & Ryazantseva, K. V. 2022. Fats and emulsifiers in feeding broiler chickens. Sel'skohozyajstvennaya biologiya= Agricultural biology, 57(4), 664-680. Doi: 10.15389/agrobiology.2022.4.664eng.
[7]  Tanasupawat S, Phoottosavako M, Keeratipibul S. 2015. Characterization and lipolytic activity of lactic acid bacteria isolated from Thai fermented meat. J. App. Pharm. Sci. 5(03):006–012. doi.org/10.7324/JAPS.2015.50302.
[8] Graham, K., Stack, H., & Rea, R. 2020. Safety, beneficial and technological properties of enterococci for use in functional food applications–a review. Critical Reviews in Food Science and Nutrition, 60(22), 3836-3861.doi: 10.1080/10408398.2019.1709800.
[9] Ali, Z.A.,2025. Isolation and genetic diagnosis of local isolate Enterococcus faecium with evaluation of inhibitory against bacterial pathogens. 11(2), DOI:10.36582/97ksxq66.
[10] Rabbani, M., Bagherinejad, M.R., Sadeghi, H.M., Shariat, Z.S., Etemadifar, Z., Moazen, F., Rahbari, M., (...), Zaghian, S. 2013. Isolation and characterization of novel thermophilic lipase-secreting bacteria. Brazilian J. of Microbiol., 44 (4), pp. 1113-1119. doi: 10.1590/S1517-83822013000400013 http://www.scielo.br/pdf/bjm/v44n4/13.pdf.
[11] Dincer, E. And Kivanc, M. 2018. Lipolytic activity of lactic acid bacteria isolated from turkish pastirma. Anadolu univ. J. Of sci. And technology c – life sci. And biotech. 7 (1), pp. 12 - 19. DOI: 10.18036/aubtdc.306292.
[12] Zouaoui, B, Bouziane, A, Rahoghalem, B. 2012. Production, optimization, and purification of lipase from Pseudomonas aeruginosa. Afr. J. Microbiol. Res., 6 (20), pp. 4417-4423. https://doi.org/10.5897/AJMR11.1567.
[13] Mohammed,A.A.; Hussain,N.A.; Niamah,A.K. 2020. Antibacterial Spectrum of produced reuterin from new isolates of lactobacillus reuteri. J microbiol biotech food sci., 10 (1) pp.134-139. Doi: 10.15414/jmbfs.2020.10.1.134-139.
[14]  Duan, X. Zhai, W. Li, X. Wu, S. Wang, Y. Wang, L. Basang, W. Zhu, Y. and Gao,Y. 2023. Preparation, purification, and biochemical of fat degrading bacterial enzymes from pig carcass compost and its application. BMC Biotechnology. 23(48), pp.14. https://doi.org/10.1186/s12896-023-00818-1.
[15] Kulkarni,A.G. and Vedamurthy,A.B. 2017. Isolation, Characterization And Optimization Of Lipase Produced From Bacteria. International. J. Res. Public. Vol.7(3),pp.20-37.ISSN2251-1563.
 [16] Al-Mossawi, L.H.M. Mahdi, N.Z. Zwain, L.A. 2016. Higher production of lipase enzyme from different microorganisms grown in local natural culture media. Int. J. Adv. Res. Biol. Sci., 3(5): pp.232-239. ISSN: 2348-8069. SOI: http://s-o-i.org/1.15/ijarbs-2016-3-5-33.
[17] Mandepudi, D.; Mandepudi, B.; Chowdary, G.V. and Gajbhiye, A. 2012. Screening, Isolation and Biochemical Characterization of Novel Lipase Producing Bacteria from Soil Samples. International J. of Biol. Eng., 2(2): pp.18-22. DOI: 10.5923/j.ijbe.20120202.03.
[18] Alkabee , H. J. J. and AlHamdani, M. A. 2015. Purification and Characterization of Lipase produced by Pseudomonas aeruginosa isolated from some soils of Basrah Governorate. Al-Kufa University J. for Biology. vol.7 (2), pp.42-49. Print ISSN: 2073-8854. Online ISSN: 2311-6544.
[19]  Yousif, A.Y. and Muhssen, E.H. 2019. The Effect of Adding Probiotics on Microbial and Organoleptic Characteristics of Beef Burger Product.J.of Global Pharma. Technol. vol. 11(9), pp.152-157. ISSN: 0975 -8542.
[20]  Muhssen,E.H. 2016. The Use of Sumac (Rhus glabra) Plantin Prolonging the Period of Beef Meat Storage. J. of Basrah Agric.Sci. vol.29 (1), pp.26-34.
[21] SPSS. (2019). IBM Statistics 26 step by step. 16th Edition. https://doi.org/10.4324/9780429056765. 
[22] Duza, M. B. and Mastan, D. S. 2014. Optimization of lipase production from Bacillus thuringiensis (TS11BP), Achromobacter xylosoxidans J2 (TS2MCN)-isolated from soil sediments near oilseed farm.IOSR J. Pharm. Biol. Sci, 9 (2), pp. 66-76.
[23] Pitt, J I, Hocking, A D. 2009. Spoilage of stored, processed, and preserved foods. Fungi and Food Spoilage, (3rd ed., Springer, Boston, MA) pp. 401-421.
[24] Rambaut A. FigTree v1.4.2. 2014. Available at: http:// tree.bio.ed.ac.uk/ software/figtree.
[25] Sabir, S.F., Alwatar, W.M.A., Matti, B.F. 2021. Impact of Programmed Death 1 Expression in Chronic myeloid Leukemia at Different Molecular Responses to imatinib Mesylate Treatment. Biochemical and Cellular Archives, 21 (1), pp. 671-678. http://www.connectjournals.com/toc1.php?bookmark=CJ-033216WWW.CONNECTJOURNALS.COM/BCA.
 [26] Sagar, K, Bashir, Y, Phukan, M M, Konwar, B K. 2013. Isolation of lipolytic bacteria from waste contaminated soil: A study about process optimization for lipase. J. Sci. Technol. Res, 2 (10), pp. 214-218.
[27] Sharma, S., Kanwar, S.S. 2014. Organic solvent tolerant lipases and applications (2014) The Scientific World Journal, art. No. 625258. doi: 10.1155/2014/625258.
[28] Lee, L.P., Karbul, H.M., Citartan, M., Gopinath, S.C.B., Lakshmipriya, T., Tang, T. H. 2015. Lipase-secreting Bacillus species in an oil-contaminated habitat: Promising strains to alleviate oil pollution. BioMed Research International, 2015, art. no. 820575. http://www.hindawi.com/journals/biomed/ Doi: 10.1155/2015/820575.
[29] Kim E., Kim D., Yang S., Kim H. 2022. The accurate identification and quantification of six Enterococcus species using quantitative polymerase chain reaction based novel DNA markers. LWT - Food Science and Technology, 166, 113769. https://doi.org/10.1016/j.lwt.2022.113769.  
[30] Narasimha,G.; Kumar,A.P.; Subramanyam D. 2011. Production and optimization of lipase enzyme by Pseudomonas sps. BTAIJ, 5(1), pp.36-42. ISSN: 0974 – 7435.
[31] Padmapriya, B.; Rajeswari, T.; Noushida, E.; Sethupalan, D.G. and Venil, C.K. 2011. Production of Lipase Enzyme from Lactobacillus spp. and Its Application in the Degradation of Meat. World Appl. Sci. J., 12 (10): pp.1798-1802. ISSN 1818-4952.