Abstract
In this study, Lacticaseibacillus casei NCIM 5752, a new isolate has been explored for probiotic properties and has shown significant bile salt hydrolase activity and cholesterol-reducing activity (56.7 ± 0.27%) in the presence of bile salts. It also tested negative for the production of lecithinase and gelatinase, indicating its non-pathogenic nature. The test strain was able to tolerate pH of 2.0 and 3.0 with 63.42 and 94.7% of the cells survived after 3 h. L. casei showed auto-aggregation of 85.3% and surface hydrophobicity of 22.5% in xylene and 19.4% in hexane. Paneer whey was explored as a basic raw material for alternative media formulation for growing lactic acid bacteria. Paneer whey was found to contain lactose (4.15%), protein (0.42%), and rich in mineral content. Response surface methodology was employed to optimize the medium composition with three independent variables yeast extract (X1), dextrose (X2), and dipotassium hydrogen phosphate (X3), and the response-Y was set to biomass obtained in terms of log CFU/ml. They were supplemented to paneer whey medium for growing this strain. The second-order polynomial regression model predicted that the maximum cell mass production of 11.30 ± 0.5 log CFU/ml at optimal composition of 16.22 g/L of yeast extract, 19.31 g/L of dextrose, and 2.12 g/L of dipotassium hydrogen phosphate in paneer whey medium. Experiments were conducted to validate the RSM results, and the biomass achieved was 11.27 ± 0.50 log CFU/ml, which is in close agreement with the yield predicted by the RSM. By applying the fermentation strategy, the biomass was increased to 5.56 ± 0.34 g/L dry cell weight corresponding to 11.58 ± 0.24 log CFU/ml. The newly optimized media was significantly cost-effective and produced 26.45% more biomass than the conventional MRS media. This optimized media may find application for the large-scale biomass production of probiotics.
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References
Aguirre-Ezkauriatza EJ, Aguilar-Yáñez JM, Ramírez-Medrano A, Alvarez MM (2010) Production of probiotic biomass (Lactobacillus casei) in goat milk whey: comparison of batch, continuous and fed-batch cultures. Bioresour Technol 101:2837–2844. https://doi.org/10.1016/j.biortech.2009.10.047
Anderson TM, Bodie EA, Goodman N, Schwartz RD (1986) Inhibitory effect of autoclaving whey-based medium on propionic acid production by Propionibacterium shermanii. Appl Environ Microbiol 51:427–428. https://doi.org/10.1128/aem.51.2.427-428.1986
Archer AC, Halami PM (2015) Probiotic attributes of Lactobacillus fermentum isolated from human feces and dairy products. Appl Microbiol Biotechnol 99:8113–8123. https://doi.org/10.1007/s00253-015-6679-x
Bernárdez PF, Amado IR, Castro LP, Guerra NP (2008) Production of a potentially probiotic culture of Lactobacillus casei subsp. casei CECT 4043 in whey. Int Dairy J 18(10–11):1057–1065
Bielecka M (2006) Probiotics in food. Chemical and functional properties of food components, 3rd edn. CRC Press, Boca Raton, pp 413–426
Birri DJ, Brede DA, Tessema GT (2013) Bacteriocin production, antibiotic susceptibility and prevalence of haemolytic and gelatinase activity in faecal lactic acid bacteria isolated from healthy Ethiopian infants. Microb Ecol 65:504–516. https://doi.org/10.1007/s00248-012-0134-7
Chang CP, Liew SL (2013) Growth medium optimization for biomass production of a probiotic bacterium, Lactobacillus rhamnosus ATCC 7469. J Food Biochem 37:536–543. https://doi.org/10.1111/jfbc.12004
De Vuyst L, Leroy F, De Vuyst L, Leroy F (2007) Bacteriocins from lactic acid bacteria: production, purification, and food applications. J Mol Microbiol Biotechnol 13:194–199. https://doi.org/10.1159/000104752
Demirci TPA (2004) Evaluation of agar diffusion bioassay for nisin quantification. Appl Microbiol Biotechnol 65:268–272. https://doi.org/10.1007/s00253-004-1579-5
Eyahmalay J, Elsayed EA, Dailin DJ et al (2020) Statistical optimization approaches for high cell biomass production of Lactobacillus casei. J Sci Ind Res (India) 79:216–221
Franz CMAP, Specht I, Haberer P, Holzapfel WH (2001) Bile salt hydrolase activity of enterococci isolated from food: screening and quantitative determination. J Food Prot 64:725–729. https://doi.org/10.4315/0362-028X-64.5.725
Goyal N, Gandhi DN (2009) Comparative analysis of Indian paneer and cheese whey for electrolyte whey drink. World J Dairy Food Sci 4:70–72
Hongpattarakere T, Cherntong N, Wichienchot S et al (2012) In vitro prebiotic evaluation of exopolysaccharides produced by marine isolated lactic acid bacteria. Carbohydr Polym 87:846–852. https://doi.org/10.1016/j.carbpol.2011.08.085
Huang S, Cauty C, Dolivet A et al (2016) Double use of highly concentrated sweet whey to improve the biomass production and viability of spray-dried probiotic bacteria. J Funct Foods 23:453–463. https://doi.org/10.1016/j.jff.2016.02.050
**i R, Swapna HC, Rai AK et al (2011) Isolation and characterization of potential lactic acid bacteria (LAB) from freshwater fish processing wastes for application in fermentative utilisation of fish processing waste. Braz J Microbiol 42:1516–1525. https://doi.org/10.1590/S1517-83822011000400039
Jones ML, Tomaro-duchesneau C, Martoni CJ (2013) Cholesterol lowering with bile salt hydrolase-active probiotic bacteria, mechanism of action, clinical evidence, and future direction for heart health applications. Expert Opin Biol Ther 13:631–642
Kolev P, Rocha-Mendoza D, Ruiz-Ramírez S et al (2022) Screening and characterization of β-galactosidase activity in lactic acid bacteria for the valorization of acid whey. JDS Commun 3:1–6. https://doi.org/10.3168/jdsc.2021-0145
Kosin B, Rakshit SK (2006) Microbial and processing criteria for production of probiotics: a review. Food Technol Biotechnol 44:371–379
Kumar M, Nagpal R, Kumar R et al (2012) Cholesterol-lowering probiotics as potential biotherapeutics for metabolic diseases. Exp Diabetes Res. https://doi.org/10.1155/2012/902917
Lye HS, Rahmat-Ali GR, Liong MT (2010) Mechanisms of cholesterol removal by lactobacilli under conditions that mimic the human gastrointestinal tract. Int Dairy J 20:169–175. https://doi.org/10.1016/j.idairyj.2009.10.003
Mathara JM, Schillinger U, Guigas C et al (2008) Functional characteristics of Lactobacillus spp. from traditional Maasai fermented milk products in Kenya. Int J Food Microbiol 126:57–64. https://doi.org/10.1016/j.ijfoodmicro.2008.04.027
Nithya V, Halami PM (2013) Evaluation of the probiotic characteristics of Bacillus species isolated from different food sources. Ann Microbiol 63:129–137. https://doi.org/10.1007/s13213-012-0453-4
Oh S, Rheem S, Sim J et al (1995) Optimizing conditions for the growth of Lactobacillus casei YIT 9018 in tryptone-yeast extract-glucose medium by using response surface methodology. Appl Environ Microbiol 61:3809–3814. https://doi.org/10.1128/aem.61.11.3809-3814.1995
Omar S, Sabry S (1991) Microbial biomass and protein production from whey. J Islamic World Acad Sci 4:170–172
Ooi LG, Liong MT (2010) Cholesterol-lowering effects of probiotics and prebiotics: a review of in vivo and in vitro findings. Int J Mol Sci 11:2499–2522. https://doi.org/10.3390/ijms11062499
Papamanoli E, Tzanetakis N, Kotzekidou P (2003) Characterization of lactic acid bacteria isolated from a Greek dry-fermented sausage in respect of their technological and probiotic properties. Meat Sci 65:859–867. https://doi.org/10.1016/S0309-1740(02)00292-9
Pescuma M, de Valdez GF, Mozzi F (2015) Whey-derived valuable products obtained by microbial fermentation. Appl Microbiol Biotechnol 99:6183–6196. https://doi.org/10.1007/s00253-015-6766-z
Prajapati JB, Hati S, Sreeja V, Trivedi J (2017) Deproteinated cheese whey medium for biomass production of probiotic Lactobacillus helveticus MTCC 5463. Int J Curr Microbiol Appl Sci 6:174–187
Ron EZ, Rosenberg E (2016) Consequences of microbial interactions with hydrocarbons, oils, and lipids: production of fuels and chemicals. Springer, Cham
Son S, Lewis BA (2002) Free radical scavenging and antioxidative activity of caffeic acid amide and ester analogues: structure-activity relationship. J Agric Food Chem 50:468–472. https://doi.org/10.1021/jf010830b
Terpstra M, Ahrens M, Smith S (2001) Effect of manganese on Lactobacillus casei fermentation to produce lactic acid from whey permeate. Process Biochem 36:671–675. https://doi.org/10.1016/S0032-9592(00)00265-X
Thapa N, Pal J, Tamang JP (2004) Microbial diversity in ngari, hentak and tungtap, fermented fish products of North-East India. World J Microbiol Biotechnol 20:599–607. https://doi.org/10.1023/B:WIBI.0000043171.91027.7e
Tomás MSJ, Bru E, Wiese B, Nader-Macías MEF (2010) Optimization of low-cost culture media for the production of biomass and bacteriocin by a urogenital Lactobacillus salivarius Strain. Probiotics Antimicrob Proteins 2:2–11. https://doi.org/10.1007/s12602-010-9037-4
Zall RR (1984) Trends in whey fractionation and utilization, a global perspective. J Dairy Sci 67:2621–2629. https://doi.org/10.3168/jds.S0022-0302(84)81623-9
Zarnowski R, Connolly PA, Wheat LJ, Woods JP (2007) Production of extracellular proteolytic activity by Histoplasma capsulatum grown in Histoplasma-macrophage medium is limited to restriction fragment length polymorphism class 1 isolates. Diagn Microbiol Infect Dis 59:39–47. https://doi.org/10.1016/j.diagmicrobio.2007.03.020
Zotta T, Cuatrin A, Lavari L et al (2015) Growth of Lactobacillus rhamnosus 64 in whey permeate and study of the effect of mild stresses on survival to spray drying. LWT - Food Sci Technol 63:322–330
Gama R, Petrides D (2019) Production of Industrial Enzymes Modeling and Evaluation with by Simulation, Design, and Scheduling Tools. Intelligen, Inc 1–40
Acknowledgments
The authors gratefully thank the Director of CSIR-CFTRI, Mysore, for providing facilities to carry out this work. One of the authors, MN, gratefully acknowledges RCB-DBT, New Delhi, for the award of DBT-JRF fellowship - DBT/JRF/15/AL/232. The author also thanks Mega-dairy, Mysuru, for providing paneer whey for experiments.
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This work was supported by the RCB-DBT (DBT/JRF/15/AL/232) Faridabad, India.
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MN: Investigation, writing, editing, and reviewing, NKR: RSM experimental design and statistical analysis of the RSM data, SD: conceptualization, supervision, editing, and reviewing. All the authors read and approved the final manuscript.
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Nanjaiah, M., Rastogi, N.K. & Devappa, S. Study of the probiotic properties of Lacticaseibacillus casei subsp. casei NCIM 5752 and the optimization of whey-based media for the production of its biomass using response surface methodology. 3 Biotech 14, 49 (2024). https://doi.org/10.1007/s13205-023-03899-z
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DOI: https://doi.org/10.1007/s13205-023-03899-z