Cellobiose dehydrogenase (CDH) is an extracellular hemoflavoprotein catalyzing the oxidation reaction of β-1,4-glycosidic-bonded sugars (lactose or cellobiose), which results in the formation of aldobionic acids and hydrogen peroxide as a byproduct. The biotechnological application of CDH requires the immobilization of the enzyme on a suitable support. As a carrier of natural origin used for CDH immobilization, chitosan seems to increase the catalytic potential of the enzyme, especially for applications as packaging in the food industry and as a dressing material in medical applications. The present study aimed to immobilize the enzyme on chitosan beads and determine the physicochemical and biological properties of immobilized CDHs obtained from different fungal sources. The chitosan beads with immobilized CDHs were characterized in terms of their FTIR spectra or SEM microstructure. The most effective method of immobilization in the proposed modification was the covalent bonding of enzyme molecules using glutaraldehyde, resulting in efficiencies ranging from 28 to 99%. Very promising results, compared to free CDH, were obtained in the case of antioxidant, antimicrobial, and cytotoxic properties. Summarizing the obtained data, chitosan seems to be a valuable material for the development of innovative and effective immobilization systems for biomedical applications or food packaging, preserving the unique properties of CDH.

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.

Lactobionic acid, widely used in the pharmaceutical, cosmetic, and food industries, is produced through chemical and biological methods, each with distinct advantages and challenges. This review examines the key approaches to its production, highlighting the chemical oxidation of lactose and biotechnological processes using microbial and enzymatic systems. Chemical methods offer high yields and rapid production but are often hindered by environmental concerns and lower product purity. In contrast, biological methods provide eco-friendly alternatives with superior product quality, albeit with limitations in scalability and higher initial costs. The comparison in this overview focuses on productivity, cost efficiency, purity, and environmental impacts, underscoring the potential of biological methods to align with sustainability goals. This review highlights further advances in biological approaches and explores hybrid solutions that combine the strengths of both methods for optimal lactobionic acid production.

Enzymes produced by white rot fungi are involved in the synthesis of secondary metabolites with valuable biotechnological properties. One of these metabolites is lactobionic acid (LBA). The aim of this study was to characterize a novel enzyme system consisting of a cellobiose dehydrogenase from Phlebia lindtneri (PlCDH), a laccase from Cerrena unicolor (CuLAC), a redox mediator (ABTS or DCPIP), and lactose as a substrate. We used quantitative (HPLC) and qualitative methods (TLC, FTIR) to characterise the obtained LBA. The free radical scavenging effect of the synthesised LBA was assessed with the DPPH method. Bactericidal properties were tested against Gram-negative and Gram-positive bacteria. We obtained LBA in all the systems tested; however, the study showed that the temperature of 50 °C with the addition of ABTS was the most advantageous condition for the synthesis of lactobionic acid. A mixture with 13 mM LBA synthesised at 50 °C with DCPIP showed the best antioxidant properties (40% higher compared with the commercial reagent). Furthermore, LBA had an inhibitory effect on all the bacteria tested, but the effect was better against Gram-negative bacteria with growth inhibition no lower than 70%. Summarizing the obtained data, lactobionic acid derived in a multienzymatic system is a compound with great biotechnological potential.

Lactobionic acid (LBA) is a compound that, in the last decade, has become critically important due to its potential applications in the food, chemical, pharmaceutical, and cosmetic industries. Enzymatic biosynthesis in the presence of a redox mediator is one method of producing LBA biologically. Cellobiose dehydrogenase (CDH) oxidizes the lactose to lactobionic acid, while laccase (LAC) enables the regeneration of the redox mediator (ABTS), which acts as an electron acceptor for CDH. The aim of this study was to develop an effective immobilized enzymatic system for the production of LBA. Two enzymes were used in the experiment: CDH from Phanerodontia chrysosporium (PchCDH) and LAC from Cerrena unicolor (CuLAC), which were immobilized on precipitated silica (Sipernat 22) activated by APTES and PEI. The immobilization process increased enzyme stability, improved the efficiency of LBA synthesis, and reduced costs, particularly in the context of using Sipernat 22 silica, which is inexpensive and widely used across various industries. The co-immobilization of both enzymes on the carrier proved to be the most effective approach, achieving a 90% conversion of lactose to lactobionic acid after ten cycles of synthesis. Comprehensive biochemical characterization, including protein loading, catalytic activity, and optimal pH, is provided in the main text.