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Lactose hydrolysis in packed-and fluidized-bed reactors using a recombinant β-galactosidase immobilized on magnetic core-shell capsules

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Abstract

The objective of this study was to develop a bioprocess for lactose hydrolysis in diverse dairy matrices, specifically skim milk and cheese whey, utilizing column reactors employing a core–shell enzymatic system featuring β-galactosidase fused to a Cellulose Binding Domain (CBD) tag (β-galactosidase-CBD). The effectiveness of reactor configurations, including ball columns and toothed columns operating in packed and fluidized-bed modes, was evaluated for catalyzing lactose hydrolysis in both skim milk and cheese whey. In a closed system, these reactors achieved lactose hydrolysis rates of approximately 50% within 5 h under all evaluated conditions. Considering the scale of the bioprocess, the developed enzymatic system was capable of continuously hydrolyzing 9.6 L of skim milk while maintaining relative hydrolysis levels of approximately 50%. The biocatalyst, created by immobilizing β-galactosidase-CBD on magnetic core-shell capsules, exhibited exceptional operational stability, and the proposed bioprocess employing these column reactors showcases the potential for scalability.

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References

  1. Rueda N, Albuquerque TL, Bartolome-Cabrero R et al (2016) Reversible immobilization of lipases on heterofunctional octyl-amino agarose beads prevents enzyme desorption. Molecules 21:646. https://doi.org/10.3390/MOLECULES21050646

    Article  PubMed  PubMed Central  Google Scholar 

  2. Vera C, Guerrero C, Aburto C et al (2020) Conventional and non-conventional applications of β-galactosidases. Biochim Biophys Acta Proteins Proteom 1868:140271. https://doi.org/10.1016/J.BBAPAP.2019.140271

    Article  CAS  PubMed  Google Scholar 

  3. Bellé AS, Hackenhaar CR, Spolidoro LS et al (2018) Efficient enzyme-assisted extraction of genipin from genipap (Genipa americana L.) and its application as a crosslinker for chitosan gels. Food Chem 246:266–274. https://doi.org/10.1016/J.FOODCHEM.2017.11.028

    Article  PubMed  Google Scholar 

  4. Balthazar CF, Pimentel TC, Ferrão LL et al (2017) Sheep milk: physicochemical characteristics and relevance for functional food development. Compr Rev Food Sci Food Saf 16:247–262. https://doi.org/10.1111/1541-4337.12250

    Article  CAS  PubMed  Google Scholar 

  5. Duan F, Sun T, Zhang J et al (2022) Recent innovations in immobilization of β-galactosidases for industrial and therapeutic applications. Biotechnol Adv 61:108053. https://doi.org/10.1016/J.BIOTECHADV.2022.108053

    Article  CAS  PubMed  Google Scholar 

  6. Shafi A, Husain Q (2022) Immobilization of β-galactosidase on concanavalin a modified silica-coated titanium dioxide nanocomposite and its interaction with monovalent and divalent cations. Mater Today Commun 32:103828. https://doi.org/10.1016/J.MTCOMM.2022.103828

    Article  CAS  Google Scholar 

  7. Lima PC, Gazoni I, de Carvalho AMG et al (2021) β-galactosidase from Kluyveromyces lactis in genipin-activated chitosan: an investigation on immobilization, stability, and application in diluted UHT milk. Food Chem 349:129050. https://doi.org/10.1016/J.FOODCHEM.2021.129050

    Article  CAS  PubMed  Google Scholar 

  8. Argenta AB, Nogueira A, de Scheer PA (2021) Hydrolysis of whey lactose: Kluyveromyces lactis β-galactosidase immobilisation and integrated process hydrolysis-ultrafiltration. Int Dairy J 117:105007. https://doi.org/10.1016/J.IDAIRYJ.2021.105007

    Article  CAS  Google Scholar 

  9. Hackenhaar CR, Rosa CF, Flores EEE et al (2022) Development of a biocomposite based on alginate/gelatin crosslinked with genipin for β-galactosidase immobilization: performance and characteristics. Carbohydr Polym 291:119483. https://doi.org/10.1016/J.CARBPOL.2022.119483

    Article  CAS  PubMed  Google Scholar 

  10. Estevinho BN, Samaniego N, Talens-Perales D et al (2018) Development of enzymatically-active bacterial cellulose membranes through stable immobilization of an engineered β-galactosidase. Int J Biol Macromol 115:476–482. https://doi.org/10.1016/J.IJBIOMAC.2018.04.081

    Article  CAS  PubMed  Google Scholar 

  11. Tavares GF, Xavier MR, Neri DFM, de Oliveira HP (2016) Fe3O4@polypyrrole core-shell composites applied as nanoenvironment for galacto-oligosaccharides production. J Chem Eng 306:816–825. https://doi.org/10.1016/J.CEJ.2016.08.018

    Article  CAS  Google Scholar 

  12. Zhao W, Yang RJ, Qian TT et al (2013) Preparation of novel Poly(hydroxyethyl methacrylate-co-glycidyl methacrylate)-grafted core-shell magnetic chitosan microspheres and immobilization of lactase. Int J Mol Sci 14:12073–12089. https://doi.org/10.3390/IJMS140612073

    Article  PubMed  PubMed Central  Google Scholar 

  13. Wu Z, Wang Z, Guan B et al (2013) Improving the properties of β-galactosidase from Aspergillus oryzae via encapsulation in aggregated silica nanoparticles. New J Chem 37:3793–3797. https://doi.org/10.1039/C3NJ00685A

    Article  CAS  Google Scholar 

  14. Ishiguro T, Obata A, Nagata K et al (2022) Core–shell fibremats comprising a poly(AM/DAAM)/ADH nanofibre core and nylon6 shell layer are an attractive immobilization platform for constructing immobilised enzymes. RSC Adv 12:34931–34940. https://doi.org/10.1039/D2RA06620C

    Article  ADS  PubMed  PubMed Central  Google Scholar 

  15. Galogahi FM, Zhu Y, An H, Nguyen NT (2020) Core-shell microparticles: generation approaches and applications. J Sci-Adv Mater Dev 5:417–435. https://doi.org/10.1016/J.JSAMD.2020.09.001

    Article  CAS  Google Scholar 

  16. Kurian M, Thankachan S (2021) Structural diversity and applications of spinel ferrite core - shell nanostructures- a review. Open Ceramics 8:100179. https://doi.org/10.1016/J.OCERAM.2021.100179

    Article  CAS  Google Scholar 

  17. Zhou Y, Li Y, Hou Y et al (2023) Core-shell catalysts for the elimination of organic contaminants in aqueous solution: a review. Chem Eng J 455:140604. https://doi.org/10.1016/J.CEJ.2022.140604

    Article  CAS  Google Scholar 

  18. Bolivar JM, López-Gallego F (2020) Characterization and evaluation of immobilized enzymes for applications in flow reactors. Curr Opin Green Sustain Chem 25:100349. https://doi.org/10.1016/J.COGSC.2020.04.010

    Article  Google Scholar 

  19. Kowalczykiewicz D, Szymańska K, Gillner D, Jarzębski AB (2021) Rotating bed reactor packed with heterofunctional structured silica-supported lipase. Developing an effective system for the organic solvent and aqueous phase reactions. Micropor Mesopor Mat 312:110789. https://doi.org/10.1016/J.MICROMESO.2020.110789

    Article  CAS  Google Scholar 

  20. Hama S, Tamalampudi S, Yoshida A et al (2011) Enzymatic packed-bed reactor integrated with glycerol-separating system for solvent-free production of biodiesel fuel. Biochem Eng J 55:66–71. https://doi.org/10.1016/J.BEJ.2011.03.008

    Article  CAS  Google Scholar 

  21. da Natividade SJ, Matte CR, Charqueiro DS et al (2017) Directed immobilization of CGTase: The effect of the enzyme orientation on the enzyme activity and its use in packed-bed reactor for continuous production of cyclodextrins. Process Biochem 58:120–127. https://doi.org/10.1016/J.PROCBIO.2017.04.041

    Article  Google Scholar 

  22. Van Zessen E, Tramper J, Rinzema A, Beeftink HH (2005) Fluidized-bed and packed-bed characteristics of gel beads. Chem Eng J 115:103–111. https://doi.org/10.1016/J.CEJ.2005.08.016

    Article  Google Scholar 

  23. Costa-Silva TA, Carvalho AKF, Souza CRF et al (2021) Enhancement lipase activity via immobilization onto chitosan beads used as seed particles during fluidized bed drying: application in butyl butyrate production. Appl Catal A Gen 622:118217. https://doi.org/10.1016/J.APCATA.2021.118217

    Article  CAS  Google Scholar 

  24. Bianchi P, Williams JD, Kappe CO (2020) Oscillatory flow reactors for synthetic chemistry applications. J Flow Chem 10:475–490. https://doi.org/10.1007/S41981-020-00105-6/FIGURES/17

    Article  CAS  Google Scholar 

  25. Tafete GA, Habtu NG (2023) Reactor configuration, operations and structural catalyst design in process intensification of catalytic reactors: a review. Chem Eng Process 184:109290. https://doi.org/10.1016/J.CEP.2023.109290

    Article  CAS  Google Scholar 

  26. Gennari A, Simon R, de Andrade BC et al (2021) Production of beta-galactosidase fused to a cellulose-binding domain for application in sustainable industrial processes. Bioresour Technol 326:124747. https://doi.org/10.1016/J.BIORTECH.2021.124747

    Article  CAS  PubMed  Google Scholar 

  27. Gennari A, Mobayed FH, Da Rolt NB et al (2019) Immobilization of β-galactosidases on magnetic nanocellulose: textural, morphological, magnetic, and catalytic properties. Biomacromol 20:2315–2326. https://doi.org/10.1021/ACS.BIOMAC.9B00285

    Article  CAS  Google Scholar 

  28. Gennari A, Simon R, de Moura Sperotto ND et al (2022) One-step purification of a recombinant beta-galactosidase using magnetic cellulose as a support: rapid immobilization and high thermal stability. Bioresour Technol 345:126497. https://doi.org/10.1016/J.BIORTECH.2021.126497

    Article  CAS  PubMed  Google Scholar 

  29. Rech R, Cassini CF, Secchi A, Ayub MAZ (1999) Utilization of protein-hydrolyzed cheese whey for production of β-galactosidase by Kluyveromyces marxianus. J Ind Microbiol Biotechnol 23:91–96. https://doi.org/10.1038/SJ.JIM.2900692

    Article  CAS  PubMed  Google Scholar 

  30. Shafi A, Ahmed F, Husain Q (2021) β-Galactosidase mediated synthesized nanosupport for the immobilization of same enzyme: Its stability and application in the hydrolysis of lactose. Int J Biol Macromol 184:57–67. https://doi.org/10.1016/J.IJBIOMAC.2021.06.034

    Article  CAS  PubMed  Google Scholar 

  31. Mateo C, Palomo JM, Fernandez-Lorente G et al (2007) Improvement of enzyme activity, stability and selectivity via immobilization techniques. Enzyme Microb Technol 40:1451–1463. https://doi.org/10.1016/J.ENZMICTEC.2007.01.018

    Article  CAS  Google Scholar 

  32. Gennari A, Simon R, de Moura Sperotto ND et al (2022) Application of cellulosic materials as supports for single-step purification and immobilization of a recombinant β-galactosidase via cellulose-binding domain. Int J Biol Macromol 199:307–317. https://doi.org/10.1016/J.IJBIOMAC.2022.01.006

    Article  CAS  PubMed  Google Scholar 

  33. Pender K, Yang L (2019) Investigation of catalyzed thermal recycling for glass fiber-reinforced epoxy using fluidized bed process. Polym Compos 40:3510–3519. https://doi.org/10.1002/PC.25213

    Article  CAS  Google Scholar 

  34. Lorenzoni ASG, Aydos LF, Klein MP et al (2015) Continuous production of fructooligosaccharides and invert sugar by chitosan immobilized enzymes: comparison between in fluidized and packed bed reactors. J Mol Catal B Enzym 111:51–55. https://doi.org/10.1016/J.MOLCATB.2014.11.002

    Article  CAS  Google Scholar 

  35. Hussain A, Minh LQ, Lee M (2017) Intensification of the ethylbenzene production process using a column configured with a side reactor. Chem Eng Process 122:204–212. https://doi.org/10.1016/J.CEP.2017.10.003

    Article  CAS  Google Scholar 

  36. Gilbert EM, Agrawal S, Schwartz T et al (2015) Comparing different reactor configurations for partial nitritation/anammox at low temperatures. Water Res 81:92–100. https://doi.org/10.1016/J.WATRES.2015.05.022

    Article  CAS  PubMed  Google Scholar 

  37. Dal Magro L, Pessoa JPS, Klein MP et al (2021) Enzymatic clarification of orange juice in continuous bed reactors: fluidized-bed versus packed-bed reactor. Catal Today 362:184–191. https://doi.org/10.1016/J.CATTOD.2020.02.003

    Article  Google Scholar 

  38. Uytdenhouwen Y, Meynen V, Cool P, Bogaerts A (2020) The potential use of core-shell structured spheres in a packed-bed DBD plasma reactor for CO2 conversion. Catalysts 10:530. https://doi.org/10.3390/CATAL10050530

    Article  CAS  Google Scholar 

  39. Szab E, Szakos D, Kasza G, Ózsvari L (2021) Analysis of the target group of lactose-free functional foods for product development. Acta Aliment 50:153–161. https://doi.org/10.1556/066.2020.00168

    Article  Google Scholar 

  40. Sainz-García SH, López GL, Alvarado VM et al (2022) Adaptive control for narrow bandwidth input and output disturbance rejection for a non-isothermal CSTR system. Mathematics 10:1–29

    Article  Google Scholar 

  41. Shu S, Vidal D, Bertrand F, Chaouki J (2019) Multiscale multiphase phenomena in bubble column reactors: a review. Renew Energy 141:613–631. https://doi.org/10.1016/J.RENENE.2019.04.020

    Article  CAS  Google Scholar 

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Acknowledgements

We would like to thank the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) (grant no. 308515/2020-0, no. 306010/2021-6,) and Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul (FAPERGS) for the scholarships, and Universidade do Vale do Taquari – Univates, CNPq (grant no. 405688/2021-0) and FAPERGS (grant no. 19/2551-0001740-5, no. 20/2551-0000524-0, no. 22/2551-0000397-4 (Bioprocess and Biotechnology for Food Research Center—Biofood)) for the financial support granted for this research paper. We also thank Quatro G Pesquisa & Desenvolvimento Ltda for their support in the experiment.

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Authors and Affiliations

  1. Laboratório de Biotecnologia de Alimentos, Universidade Do Vale Do Taquari - Univates, Av. Avelino Talini, Lajeado, RS, 171, ZC 95914-014, Brazil

    Adriano Gennari, Renate Simon & Claucia Fernanda Volken de Souza

  2. Programa de Pós-Graduação Em Biotecnologia, Universidade Do Vale Do Taquari - Univates, Lajeado, RS, Brazil

    Adriano Gennari & Claucia Fernanda Volken de Souza

  3. Quatro G Pesquisa & Desenvolvimento Ltda, Porto Alegre, RS, Brazil

    Gaby Renard & Jocelei Maria Chies

  4. Instituto Federal de Educação, Ciência e Tecnologia Do Rio Grande Do Sul - IFRS, Campus Porto Alegre, Porto Alegre, RS, Brazil

    Giandra Volpato

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  1. Adriano Gennari
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  2. Renate Simon
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  3. Gaby Renard
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  4. Jocelei Maria Chies
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  6. Claucia Fernanda Volken de Souza
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Contributions

AG: Conceptualization, investigation, methodology, formal analysis, writing—original draft preparation, and writing—reviewing and editing. RS: Investigation, and methodology. GR: Conceptualization, methodology, formal analysis, writing—original draft preparation, and writing—reviewing and editing. JMC: Resources, and supervision. GV: Conceptualization, formal analysis, resources, supervision, writing—original draft preparation, and writing—reviewing and editing. CFVS: Conceptualization, investigation, methodology, formal analysis, resources, supervision, writing—original draft preparation, and writing—reviewing and editing.

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Correspondence to Claucia Fernanda Volken de Souza.

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Gennari, A., Simon, R., Renard, G. et al. Lactose hydrolysis in packed-and fluidized-bed reactors using a recombinant β-galactosidase immobilized on magnetic core-shell capsules. Bioprocess Biosyst Eng 47, 263–273 (2024). https://doi.org/10.1007/s00449-023-02960-8

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