Immobilisation of Cellobiose Dehydrogenase and Laccase on Chitosan Particles as a Multi-Enzymatic System for the Synthesis of Lactobionic Acid
Source abstract
Abstract
Lactobionic acid (LBA) is a bioactive compound that has become increasingly popular in medicine in recent years due to its unique properties. This chemical can be formed via the enzymatic oxidation of lactose using fungal oxidoreductive enzymes. This study aimed to intensify the synthesis of LBA using immobilised enzymes (cellobiose dehydrogenase from Phanerochaete chrysosporium (PchCDH) and laccase from Cerrena unicolor (CuLAC)) on chitosan microspheres. We used three different crosslinking agents: genipin, glutaraldehyde, and polyethyleneimine to activate the chitosan. The FTIR and CellDrop techniques were used to characterise the activated microspheres. Quantitative (HPLC) and qualitative (TLC) methods were used to determine the obtained LBA. The results show that the type of activator used influences the efficiency of the binding of the enzyme to the matrix. Furthermore, the amount of LBA formed depends on the type of system used. The use of a system in which one of the enzymes is immobilised on a PEI-activated carrier (PchCDH) and the other is free (CuLAC) proved to be the most optimal, as it yielded almost 100% conversion of lactose to lactobionic acid. Summarising the data obtained the following: lactobionic acid immobilised on chitosan microspheres has great potential for medical applications.
Source: crossref · Quality: baseline
Research context
OpenAlex topics: Enzyme Catalysis and Immobilization · Electrochemical sensors and biosensors · Enzyme-mediated dye degradation
SciVX discovery tags are terminology-based. OpenAlex topics are source-provided classifications.
Affiliations
Department of Biochemistry and Biotechnology, Institute of Biological Sciences, Maria Curie-Sklodowska University, 19 Akademicka St., 20-033 Lublin, Poland
Department of Plant Physiology and Biophysics, Institute of Biological Sciences, Maria Curie-Sklodowska University, 19 Akademicka St., 20-033 Lublin, Poland
Core Facility of Biospectroscopy, Institute of Biological Sciences, Maria Curie-Sklodowska University, 19 Akademicka St., 20-033 Lublin, Poland
Department of Cell Biology, Institute of Biological Sciences, Maria Curie-Sklodowska University, 19 Akademicka St., 20-033 Lublin, Poland
References (78)
Show source-provided references
1. Bio-production of lactobionic acid: Current status, applications and future prospects10.1016/j.biotechadv.2013.04.010 ↗
2. Lactobionic acid as a potential food ingredient: Recent studies and applications10.1111/1750-3841.14686 ↗
3. Organ preservation10.1146/annurev.med.46.1.235 ↗
4. Lactobionic acid: A high value-added lactose derivative for food and pharmaceutical applications10.1016/j.idairyj.2012.05.003 ↗
5. Lactobionic and gluconic acid complexes of FeII and FeIII; control of oxidation pathways by an organ transplantation preservant10.1016/0162-0134(93)80046-C ↗
6. Clinical and cosmeceutical uses of hydroxyacids10.1016/j.clindermatol.2009.06.023 ↗
7. Evaluation of microencapsulated synbiotic preparations containing lactobionic acid10.1007/s12010-021-03622-9 ↗
8. Substrate specificity of cellobiose dehydrogenase from Phanerochaete chrysosporium10.1016/S0167-4838(97)00180-5 ↗
9. Continuous enzymatic regeneration of redox mediators used in biotransformation reactions employing flavoproteins10.1016/S1381-1177(00)00034-5 ↗
10. Kinetic modeling of a bi-enzymatic system for efficient conversion of lactose to lactobionic acid10.1002/bit.22165 ↗
11. Evaluation of cellobiose dehydrogenase and laccase containing culture fluids of Termitomyces sp. OE147 for degradation of Reactive blue 2110.1016/j.btre.2016.10.002 ↗
12. Laccases–enzymes with an unlimited potential
13. Enhanced penetration and cytotoxicity of metformin and collagenase conjugated gold nanoparticles in breast cancer spheroids10.1016/j.lfs.2019.116545 ↗
14. Enzyme-based autophagy in anti-neoplastic management: From molecular mechanisms to clinical therapeutics10.1016/j.bbcan.2020.188366 ↗
15. Smart chemistry of enzyme immobilization using various support matrices—A review10.1016/j.ijbiomac.2021.09.006 ↗
16. Lindeque, R.M., and Woodley, J.M. (2019). Reactor selection for effective continuous biocatalytic production of pharmaceuticals. Catalysts, 9.10.3390/catal9030262 ↗
17. Industrial applications of immobilized enzymes—A review10.1016/j.mcat.2019.110607 ↗
18. Preparation of chitosan particles suitable for enzyme immobilization10.1016/j.jprot.2007.11.005 ↗
19. Enzyme immobilization: An update10.1007/s12154-013-0102-9 ↗
20. Chitosan activated with divinyl sulfone: A new heterofunctional support for enzyme immobilization. Application in the immobilization of lipase B from Candida antarctica10.1016/j.ijbiomac.2019.02.145 ↗
21. Effect of Additional Amino Group to Improve the Performance of Immobilized Lipase From Aspergillus niger by Adsorption-Crosslinking Method10.3389/fenrg.2021.616945 ↗
22. Immobilization of laccase from Trametes versicolor on chitosan macrobeads for anthracene degradation10.1080/00032719.2017.1282504 ↗
23. Design of alpha mangostin-loaded chitosan/alginate controlled-release nanoparticles using genipin as crosslinker10.1016/j.jddst.2018.05.029 ↗
24. A physico-chemical approach to the study of genipin crosslinking of biofabricated peptide hydrogels10.1016/j.procbio.2018.04.005 ↗
25. Genipin blues: An alternative non-toxic crosslinker for heart valves?
26. In vitro evaluation of the genotoxicity of a naturally occurring crosslinking agent (genipin) for biologic tissue fixation10.1002/1097-4636(200010)52:1<58::AID-JBM8>3.0.CO;2-0 ↗
27. Hyaluronic acid cryogels with non-cytotoxic crosslinker genipin
28. ‘Genipin’–the natural water soluble cross-linking agent and its importance in the modified drug delivery systems: An overview10.2174/15672018113106660059 ↗
29. Tacias-Pascacio, V.G., García-Parra, E., Vela-Gutiérrez, G., Virgen-Ortiz, J.J., Berenguer-Murcia, Á., Alcántara, A.R., and Fernandez-Lafuente, R. (2019). Genipin as an emergent tool in the design of biocatalysts: Mechanism of reaction and applications. Catalysts, 9.10.3390/catal9121035 ↗
30. Polymeric microspheres for medical applications10.3390/ma3063537 ↗
31. Natural biopolyester microspheres with diverse structures and surface topologies as micro-devices for biomedical applications10.1016/j.smaim.2022.07.004 ↗
32. Chitosan microspheres in novel drug delivery systems
33. A study of the characteristics and in vitro permeation properties of CMC/chitosan microparticles as a skin delivery system for vitamin E10.1590/S1516-93322006000100007 ↗
34. Piątek-Gołda, W., Sulej, J., Grąz, M., Waśko, P., Janik-Zabrotowicz, E., and Osińska-Jaroszuk, M. (2023). Multi-Enzymatic Synthesis of Lactobionic Acid Using Wood-Degrading Enzymes Produced by White Rot Fungi. Metabolites, 13.10.3390/metabo13040469 ↗
35. Sulej, J., Osińska-Jaroszuk, M., Jaszek, M., Olszewska, A., Belcarz, A., and Piątek-Gołda, W. (2023). Chitosan as a Promising Support of a CDH Activity Preservation System for Biomedical and Industrial Applications. Int. J. Mol. Sci., 24.10.3390/ijms24054535 ↗
36. Antimicrobial and antioxidative potential of free and immobilised cellobiose dehydrogenase isolated from wood degrading fungi10.1016/j.funbio.2019.09.007 ↗
37. A simple assay for measuring cellobiose dehydrogenase activity in the presence of laccase10.1016/S0167-7012(99)00022-6 ↗
38. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding10.1016/0003-2697(76)90527-3 ↗
39. Wlizło, K., Polak, J., Kapral-Piotrowska, J., Grąz, M., Paduch, R., and Jarosz-Wilkołazka, A. (2020). Influence of carrier structure and physicochemical factors on immobilisation of fungal laccase in terms of Bisphenol a removal. Catalysts, 10.10.3390/catal10090951 ↗
40. Novel application of porous and cellular materials for covalent immobilization of pepsin10.1590/0104-6632.20160332s20140111 ↗
41. Production of lactobionic acid using an immobilized cellobiose dehydrogenase/laccase system on magnetic chitosan spheres10.1016/j.procbio.2020.09.024 ↗
42. Involvement of Acetobacter orientalis in the production of lactobionic acid in Caucasian yogurt (“Caspian Sea yogurt”) in Japan10.3168/jds.2008-1081 ↗
43. Enzyme immobilisation: Fundamentals and application10.1039/c3cs90042h ↗
44. Federsel, H.-J., Moody, T.S., and Taylor, S.J. (2021). Recent trends in enzyme immobilization—Concepts for expanding the biocatalysis toolbox. Molecules, 26.10.3390/molecules26092822 ↗
45. Immobilization and stabilization of different β-glucosidases using the glutaraldehyde chemistry: Optimal protocol depends on the enzyme10.1016/j.ijbiomac.2019.02.057 ↗
46. Immobilization of fungal laccase on glutaraldehyde cross-linked chitosan beads and its bio-catalytic potential to degrade bisphenol A10.1016/j.bcab.2019.101174 ↗
47. Porous biochar/chitosan composites for high performance cellulase immobilization by glutaraldehyde10.1016/j.enzmictec.2020.109561 ↗
48. Chronic toxicity of glutaraldehyde: Differential sensitivity of three freshwater organisms10.1016/j.aquatox.2004.12.001 ↗
49. From sub cellular to community level: Toxicity of glutaraldehyde to several aquatic organisms10.1016/j.scitotenv.2013.09.054 ↗
50. Ehrmann, A. (2021). Non-toxic crosslinking of electrospun gelatin nanofibers for tissue engineering and biomedicine—A review. Polymers, 13.10.3390/polym13121973 ↗
51. Characterization and Optimization of Injectable in situ Crosslinked Chitosan-Genipin Hydrogels10.1002/mabi.202200505 ↗
52. Comparison of the properties of lipase immobilized onto mesoporous resins by different methods10.1007/s12010-010-9157-z ↗
53. Immobilization of a keratinolytic protease from Purpureocillium lilacinum on genipin activated-chitosan beads10.1016/j.procbio.2014.04.016 ↗
54. Facile and green fabrication of biocatalytic chitosan beads by one-step genipin-mediated β-glucosidase immobilization for production of bioactive genistein10.1016/j.scp.2019.100187 ↗
55. Cellular and molecular barriers to gene transfer by a cationic lipid10.1074/jbc.270.32.18997 ↗
56. Poly (aspartate-g-PEI800), a polyethylenimine analogue of low toxicity and high transfection efficiency for gene delivery10.1016/j.biomaterials.2007.07.043 ↗
57. The targeted co-delivery of DNA and doxorubicin to tumor cells via multifunctional PEI-PEG based nanoparticles10.1016/j.biomaterials.2012.12.038 ↗
58. In silico study of PEI-PEG-squalene-dsDNA polyplex formation: The delicate role of the PEG length in the binding of PEI to DNA10.1039/D1BM00973G ↗
59. Polyethyleneimine in immobilization of biocatalysts10.1016/0141-0229(91)90101-F ↗
60. New insights into the gelation behavior of polyethyleneimine cross-linking partially hydrolyzed polyacrylamide gels10.1021/ie301818f ↗
61. Studies on a novel nanofiltration membrane prepared by cross-linking of polyethyleneimine on polyacrylonitrile substrate10.1016/j.memsci.2013.10.003 ↗
62. Effect of crosslinking on the CO2 adsorption of polyethyleneimine-impregnated sorbents10.1016/j.cej.2016.09.005 ↗
63. Immobilization of urease on vapour phase stain etched porous silicon10.1016/j.procbio.2006.08.008 ↗
64. Screen-printed electrodes modified with HRP-zirconium alcoxide film for the development of a biosensor for acetaminophen detection10.2478/s11532-010-0070-7 ↗
65. Preparation and evaluation of chitosan microspheres containing nicorandil10.4103/2230-973X.127738 ↗
66. Genipin-crosslinked gelatin microspheres as a strategy to prevent postsurgical peritoneal adhesions: In vitro and in vivo characterization10.1016/j.biomaterials.2016.04.012 ↗
67. Jha, R., and Mayanovic, R.A. (2023). A Review of the Preparation, Characterization, and Applications of Chitosan Nanoparticles in Nanomedicine. Nanomaterials, 13.10.3390/nano13081302 ↗
68. Gelatin microspheres crosslinked with genipin for local delivery of growth factors10.1002/term.267 ↗
69. Takahashi, T. (2020). Potential of an Automated-and Image-Based Cell Counter to Accelerate Microalgal Research and Applications. Energies, 13.10.3390/en13226019 ↗
70. Non-invasive cell counting of adherent, suspended and encapsulated mammalian cells using optical density10.2144/btn-2019-0052 ↗
71. Applicability of automated cell counter with a chlorophyll detector in routine management of microalgae10.1038/s41598-018-23311-8 ↗
72. Comparison of two automatic cell-counting solutions for fluorescent microscopic images10.1111/jmi.12272 ↗
73. Stabilization of enzymes via immobilization: Multipoint covalent attachment and other stabilization strategies10.1016/j.biotechadv.2021.107821 ↗
74. Structural changes in the crosslinking process of a protein bioadhesive
75. FTIR-ATR determination of protein content to evaluate whey protein concentrate adulteration10.1016/j.lwt.2018.09.079 ↗
76. Laccase production by Pleurotus ostreatus and its application in synthesis of gold nanoparticles10.1016/j.btre.2014.11.001 ↗
77. Fourier transform infrared spectroscopic analysis of protein secondary structures10.1111/j.1745-7270.2007.00320.x ↗
78. Determination of protein secondary structure by Fourier transform infrared spectroscopy: A critical assessment10.1021/bi00053a001 ↗