Statistical optimization of fermentation media for postbiotic metabolite production from Lactobacillus plantarum LG138 of primate origin


  • Reena Kumari Reenu Department of Biotechnology, HPU,Shimla-5
  • Kiran Bala Sharma Department of Biotechnology, HPU,Shimla-5
  • Prem Lata Department of Biotechnology, HPU,Shimla-5
  • shailja Rangra
  • Dr Savitri


Exopolysaccharide, Lactic acid bacteria (LAB), Response Surface Methodology (RSM), Lactobacillus plantarum, optimization


The probiotic  metabolites  called postbiotics  synthesized by consist of range of molecules namely organic acids, antimicrobial peptides, short-chain fatty acids, exopolysaccharide (EPS), cofactors, vitamins, immune-modulating compounds, enzymes, neurotransmitters etc. LABs are widely recognized as an efficient EPS producer. Hence, the goal of the current study is to statistically maximize the media components for maximum exopolysaccharide production by Lactobacillus plantarum LG138 from primate feces. The de Man, Rogosa and Sharpe (MRS) medium is used for optimization of production process. Batch culture system is used for optimization of exopolysaccharide production in MRS medium using Lactobacillus plantarum LG138. The optimization process of EPS production improved its yield by 2.7-folds (from 12.00 mg/ml to 32.88 mg/ml).The enhanced EPS yield was achieved after optimization of different media components such as sucrose (5%), ammonium sulfate (1.2%), temperature (32.5⁰C), incubation time (22 h) and pH (6.5) using Response Surface Methodology. The actual experimental value (32.88 mg/ml) was comparable to the predicted maximum EP by Lactobacillus plantarum LG138.S production (32.87mg/ml) under optimal conditions for L. plantarum LG138.The in vitro antioxidant assays (free radical scavenging ability and reducing power) revealed  antioxidant properties of EPS. These significant  activities  recommend the  possible potential use of EPS  in animal food and feed applications. 

Author Biographies

Kiran Bala Sharma, Department of Biotechnology, HPU,Shimla-5

Department of Biotechnology

Prem Lata, Department of Biotechnology, HPU,Shimla-5

Department of Biotechnology


Aguilar-Toala, J. E., Garcia-Varela, R., Garcia, H. S., Mata-Haro, V., Gonzalez-Cordova, A. F.,Vallejo-Cordoba, B., Hernández-Mendoza, A. (2018). Postbiotics: An evolving term within the functional foods field. Trends in Food Science and Technology.75:105–114.

Barros, C.P., Guimaraes, J.T., Esmerino, E. A., Duarte, M. C. K. H., Silva, M. C., Silva, R., Cruz, A.G. (2020). Paraprobiotics and postbiotics: Concepts and potential applications in dairy products. Current Opinion in Food Science.32, 1-8.

Badel, S., Bernardi,T., Michaud P. (2011). New perspectives for Lactobacilli exopolysaccharides. Biotechnology Advances. 29:54–66. https://doi: 10.1016/j.biotechadv.2010.08.011.

Wu, M., Pan, T., Wu, Y., Chang, S., Chang, M., Hu, C. (2010). Exopolysaccharide activities from probiotic bifidobacterium :Immunomodulatory effects (on J774A1 macrophages) and antimicrobial properties. International Journal of Food Microbiology.144 (1):104–110. https://doi: 10.1016/j.ijfoodmicro.2010.09.003.

Lynch, K.M., Zannini, E., Coffey, A. , Arendt, E.K. 2018. Lactic acid bacteria exopolys accharides in foods and beverages: isolation, properties, characterization, and health benefits. Annual Review of Food Science and Technology.9(1):155–176.

Gu, D., Jiao Y., Wu J., Liu Z., Chen Q. (2017). Optimization of EPS production and characterization by a halophilic bacterium, Kocuria rosea ZJUQH from Chaka Salt Lake with response surface methodology. Molecules. 2(5):814. https :// 10.3390/molecules 22050814.

Tallon, R., Bressollier, P., Urdaci, M.C.(2003).Isolation and characterization of two exopolysaccharides produced by Lactobacillus plantarum EP56. Research in Microbiology. 154(10):705-712.

Saadat, Y.R., Khosroushahi, A.Y., Gargari, B.P. (2019). A comprehensive review of anticancer, immunomodulatory and health beneficial effects of the lactic acid bacteria exopolysaccharides. Carbohydrate Polymers . 217: 79–89.

Zaghloul, E.H. and Ibrahim M.I.A. (2022). Poduction and characterization of exopolysaccharide from newly isolated marine probiotic Lactiplantibacillus plantarum E16 with in vitro wound healing activity. Frontiers in Microbiology. 13:903363.

Banerjee, A., Mohammed Breig, S.J., Gómez, A., Sánchez-Arévalo, I., González-Faune, P., Sarkar, S., Bandopadhyay, R., Vuree, S., Cornejo, J., Tapia, J., Bravo, G., Carbrera-Barjas, G. (2022). Optimization and Characterization of a Novel Exopolysaccharide from Bacillus haynesii CamB6 for Food Applications. Biomolecules. 12(6): 834. /10.3390 biom12060834.

Derdak, R., Sakoui, S., Pop, O.L., Cristian Vodnar, D., Addoum Elmakssoudi, B., Errachidi, F., Suharoschi, R., Soukri, A., El Khalfi, B. (2022). Screening, Optimization and characterization of exopolysaccharides produced by novel strains isolated from Moroccan raw donkey milk. Food Chemistry: X. 14:100305.

Mingchen, X., Zhang, S., Shen, L., Yu, R., Liu, Y., li, J., Wu, X., Chen, M., Qiu, G., Zeng, W. (2022).Optimization and characterization of an antioxidant exopolysaccharide produced by Cupriavidus pauculus 1490. Journal of Polymer and the Environment. 30(5):2077-2086.

Cirrincione, S, Breuer, Y., Mangiapane, E., Mazzoli, R., Pessione, E.(2018).’Ropy’ phenotype, exopolysaccharides and metabolism: study on food isolated potential probiotics LAB. Microbiological Research 214: 137-145.

Mahapatra S, Banerjee D. (2013). Fungal exopolysaccharide: production, composition and applications. Microbiology Insights 29(6):1-16.

Sharma, K., Sharma, N., Handa, S., Pathania, S. (2020). Purification and characterization of novel exopolysaccharides produced from Lactobacillus paraplantarum KM1 isolated from human milk and its cytotoxicity. Journal of Genetic Engineering and Biotechnology.18(1):56.

Angelaalincy, M., Senthilkumar, N., Karpagam, R., Kumar, G.G., Kumar, A. B., Varalakshmi, P.(2017). Enhanced extracellular polysaccharide production and self-sustainable electricity generation for PAMFCs by Scenedesmus sp.SB1. ACS Omega. 2(7):3754-3765.

Niknezhad, S.V., Kianpour, S., Jafarzadeh, S., Alishahi, M., Najafpour Darzi, G., Morowvat, M. H., Ghasemi, Y, Shavandi, A.(2022). Biosynthesis of exopolysaccharide from waste molasses using Pantoea sp. BCCS 001 GH: a kinetic and optimization study. Scientific Reports. 12(1): 1-14.

Chen L, Gu Q, Zhou T. (2022). Statistical optimization of novel medium to maximize the yield of exopolysaccharide from Lacticaseibacillus rhamnosus ZFM216 and its Immunomodulatory activity. Frontiers in Nutrition. 9:924495.

Suryawanshi, N., Naik, S., Eswari, J. S. (2019). Extraction and optimization of exopolysaccharide from Lactobacillus sp. Using response surface methodology and artificial neural networks. Preparatory Biochemistry and Biotechnology. 49(10):987-996.

Wang, B., Xu, Y., Chen, L., Zhao, G., Mi, Z., Lv, D., Niu, J.(2020). Optimizing the extraction of polysaccharides from Bletilla ochracea Schltr. using response surface methodology (RSM) and evaluating their antioxidant activity. Processes. 8(3):341.

Liang, C., Qing, G., Zhou Tao, Z. (2022). Statistical optimization of novel medium to maximize the yield of exopolysaccharide from Lacticaseibacillus rhamnosus ZFM216 and its immunomodulatory activity. Frontiers in Nutrition.9.

Abhini, K.N., Rajan, A.B., Fathumathu Zuhara, K., Sebastian, D. (2022). Response surface methodological optimization of L-asparaginase production from the medicinal plant endophyte Acinetobacter baumannii ZAS1. Journal of Engineering and Biotechnology.20(22);

Gangalla, R., Sampath, G., Beduru, S., Kasarla, S., Govindrajan, R.K., Ameen, F., Alwakeel, S.S., Thampu, R. (2021). Optimization and characterization of exopolysaccharide produced by Bacillus aerophilus rk1 and its in vitro antioxidant activities. Journal of King Saud University- Science. 33 (5): 101470.

Okoro, O.V., holipour, A.R., Sedighi, F., Shavandi,. A, Hamidi, M.(2021). Optimization of exopolysaccharide (EPS) production by Rhodotorula mucilaginosa sp. GUMS16. Chem Engineering. 5(3):39.

Nguyen, P.T., Nguyen, T.T., Bui, D.C., Hong, P.T., Hoang, Q.K., Nguyen, H.T. (2020). Exopolysaccharide production by lactic acid bacteria: the manipulation of environmental stresses for industrial applications. AIMS Microbiology. 6(4):451-469.

Fashogbon, R.O., Adebayo-Tayo,B., Sanusi, J. (2021). Optimization of Extracellular Polysaccharide Substances from Lactic Acid Bacteria Isolated from Fermented Dairy Products. Microbiology Journal, 11: 1-11.

Dubois, M.K., Gilles, K.A., Hamilton, J.K., Rebers, P. T., Smith F.(1956). Colorimetric method for determination of sugars and related substances, Analytical Chemistry, 28(3), 350-356.

Yin, J., Heo, S. I., & Wang, M. H. (2008). Antioxidant and antidiabetic activities of extracts from Cirsium japonicum roots. Nutrition Research and Practice. 2(4): 247-251.

Vijayalakshmi, M. and Ruckmani, K. (2016). Ferric reducing anti-oxidant power assay in plant extract. Bangladesh Journal of Pharmacology. 11(3): 570-572 .

Onilude, A.A., Olaoye, O., Fadahunsi, I. F., Owoseni, A., Garuba, E. O., Atoyebi, T.(2013). Effects of cultural conditions on dextran production by Leuconostoc spp. International Food Research Journal. 20(4): 1645-1651.

Tao, J., Huang, X., Ling, F., Yu, Zhou, B., X., Shen Q., Sagratin, I, G. (2021).Immobilization of lactic acid bacteria for production of extracellular polysaccharides. Food Science Technology. 42, e99021.

Wang, X., Xu, P., Yuan, Y., Liu, C., Zhang, D., Yang, Z., Yang, C., Ma, C. (2006). Modeling for gellan gum production by Sphingomonas paucimobilis ATCC 31461 in a simplified medium. Applied and Environmental Microbiology. 72(5): 3367–3374.

Kimmel, S.A., Roberts, R.F., and Ziegler, G.R. 1998. Optimization of exopolysaccharide production by Lactobacillus delbrueckii subsp. bulgaricusRR grown in a semidefined medium. Applied and Environmantal Microbiology. 64: 659- 664.

Shivakumar, S., Vijayendra, S.V.N. (2006).Production of exopolysaccharides by Agrobacterium sp. CFR-24 using coconut water - A byproduct of food industry. Letters in Applied Microbiology. 42: 477-482.

Bejar, C., Calvo, J., Moliz, F., Diaz-Martine, F., Quesada, E. (1996). Effect of growth conditions on the rheological properties and chemical composition of Volcaniella eurihalina exopolysaccharide. Applied Biochemistry and Biotechnology. 59(1): 77-86.

Si, T.Z., Liu, C.J., Qin, X.M., Li, X.R., Luo, Y.Y., Yang, E. (2017). Optimization of biosynthesis conditions for the production of exopolysaccharides by Lactobacillus plantarum YM-2. Food Science. 38:24–30.

Patil, S. V., Salunkhe, R. B., Patil, C. D.. Patil, D. M., Salunke, B. K. (2010). Bioflocculant exopolysaccharide production by Azotobacter indicus using flower extract of Madhuca latifolia L. Applied Biochemistry and Biotechnology.162: 1095-1099.

Habibi, N., Soleimanian-Zad, S., Mohammad, S. Z. (2011).Exopolysaccharides produced by pure culture of Lactobacillus, Lactococccus and Yeast isolated from kefir grain by Microtiter Plate Assay: Optimization and comparision. World Applied Sciences Journal. 12: 742-750.

Kanmani, P., Suganya, K., Kumar, R.S., Yuvaraj, N., Pattukumar, V., Paari, K. A. Arul,V.(2013).Synthesis and functional characterization of antibiofilm exopolysaccharide produced by Enterococcus faecium MC13 isolated from the gut of fish.

Applied Biochemistry and Biotechnology.169:1001-1015.

Shi, T., Aryantini, N.P.D., Uchida, K., Urashima, T., Fukuda, K. (2014). Enhancement of exopolysaccharide production of Lactobacillus fermentum TDS030603 by modifying culture conditions. Bioscience Microbiota, Food and Health. 33: 85.

Sirajunnisa, A.R., Vijayagopal, V., Sivaprakash, B, Viruthagiri, T., Surendhiran, D. (2016). Optimization, kinetics and antioxidant activity of exopolysaccharide produced from rhizosphere isolate, Pseudomonas fluorescens CrN6. Carbohydrate Polymer. 135:35–43.

Duboc, P., Mollet, B. (2001). Applications of exopolysaccharides in the dairy industry. International Dairy Journal. 11:759–768. 1016/S0958-6946(01)00119-4.

Yeesang, C., Chanthachum, S., Cheirsilp, B. (2008). Sago starch as a low-cost carbon source for exopolysaccharide production by Lactobacillus kefiranofaciens. World Journal of Microbiology and Biotechnology.24:1195–1201.

Mende, S., Krzyzanowski, L., Weber, J., Jaros, D., Rohm, H.( 2012).Growth and exopolysaccharide yield of Lactobacillus delbrueckii ssp. bulgaricus DSM 20081 in batch and continuous bioreactor experiments at constant pH. Journal of Bioscience and Bioengineering. 113:185–191.

Van den Berg, D.J.C., Smits, A, Pot, B, Ledeboer, A.M., Kersters, K., Verbakel. J.M.A., and Verrips, C.T. (1995).Isolation, screening and identification of lactic acid bacteria from traditional food fermentation processes and culture collections. Food Biotechnology, 7(3): 189-205.

De Vuyst. (1998). Diversity of heteropolysaccharide producing lactic acid bacterium strains and their biopolymers. Applied and Environmental Microbiology, 72 (6): 4431–4435.

Kuntiya A., Hanmoungjai, P., Techapun, C., Sasaki, K., Seesuriyachan, P. (2010). Influence of pH, sucrose concentration and agitation speed on exopolysaccharide production by Lactobacillus confusus TISTR 1498 using coconut water as a raw material substitute. Maejo International Journal of Science and Technology, 4(2), 318-330.

Sonawdekar, S., Gupte, A. (2016). Production and characterization of exopolysaccharide produced by oil emulsifying bacteria. International Journal of Current Microbiology and Applied Sciences. 5(2): 254-262.

Salman, J. A. S., Salim, M. Z. (2016). Production and characterization of dextran from leuconostoc mesenteroides ssp. mesenteroides isolated from iraqi fish intestine. European Journal of Biomedical and Pharmaceutical sciences. 3 (8):62-69.

Sarwat, F., Qader, S. A. U., Aman, A. and Ahmed, N. (2008). Production and characterization of a unique dextran from an indigenous Leuconostoc mesenteroides CMG713. International Journal of Biological Sciences. 2 4(6): 379-386. 10.7150/ijbs.4.379.

Ragavan, M.L., Das, N. (2019). Optimization of exopolysaccharide production by probiotic yeast Lipomyces starkeyi VIT-MN03 using response surface methodology and its applications. Annals of Microbiology. 69, 515–530.

Zhang, L., Liu, C., Li, D., Zhang, X., Zeng, X., Yang, Z., Li, S.(2013). Antioxidant activity of an exopolysaccharide isolated from Lactobacillus plantarum C88.International Journal of Biological Macromolecules. 54:270-275.

Pan, D., Mei, X. (2010).Antioxidant activity of an exopolysaccharide purified from Lactococcus lactis subsp. lactis 12. Carbohydrate Polymers. 80 (3):908-914.