Search Results (86)

Hyaluronic acid (HA) is a linear heteropolysaccharide that naturally occurs in vertebrates. Thanks to its unique physico-chemical properties, it is involved in many key processes in living organisms. These biological activities provide the basis for its broad applications in cosmetics, medicine, and the food industry. The molecular weight of HA might vary significantly, as it can be less than 10 kDa or reach more than 6000 kDa. There is a strong correlation between variations in its molecular weight and bioactivities, as well as with various pathological processes. Consequently, monodispersity is a crucial requirement for HA production, together with purity and safety. Common industrial approaches, such as extraction from animal sources and microbial fermentation, have limits in fulfilling these requests. Research and protein engineering with hyaluronic acid synthases can provide a strong tool for the production of monodisperse HA. One-pot multi-enzyme reactions that include in situ nucleotide phosphate regeneration systems might represent the future of HA production. In this review, we explore the current knowledge about HA, its production, hyaluronic synthases, the most recent stage of in vitro enzymatic synthesis research, and one-pot approaches. Full article

Background/Objectives: Trace minerals (TMs), both essential and toxic, are integral to human physiology, participating in enzymatic reactions, oxidative balance, immune function, and the modulation of chronic disease risk. Despite their importance, imbalances due to deficiencies or toxic exposures are widespread globally. While low-income countries often face overt deficiencies and environmental contamination, middle- and high-income populations increasingly deal with subclinical deficits and chronic toxic metal exposure. This review aims to explore the relevance of serum as a matrix for evaluating TM status across diverse clinical and epidemiological, geographic, and demographic settings. Methods: A narrative literature review was conducted focusing on the physiological roles, health impacts, and current biomarker approaches for key essential (e.g., zinc, copper, selenium) and toxic (e.g., lead, mercury, cadmium, arsenic) trace elements. Particular emphasis was placed on studies utilizing serum analysis and on recent advances in multi-element detection using inductively coupled plasma mass spectrometry (ICP-MS). Results: Serum was identified as a versatile and informative matrix for TM assessment, offering advantages in terms of clinical accessibility, biomarker reliability, and capacity for the simultaneous quantification of multiple elements. For essential TMs, serum levels reflect nutritional status with reasonable accuracy. For toxic elements, detection depends on instrument sensitivity, but serum can still provide valuable exposure data. The method’s scalability supports applications ranging from public health surveillance to individualized patient care. Conclusions: Serum trace mineral analysis is a practical and scalable approach for nutritional assessment and exposure monitoring. Integrating it into clinical practice and public health strategies can improve the early detection of imbalances, guide interventions such as nutritional supplementation, dietary modifications, and exposure mitigation efforts. This approach also supports advanced personalized nutrition and preventive care. Full article

Creatinine serves as a crucial diagnostic biomarker for assessing kidney disease. This work developed portable non-enzymatic and multienzyme-modified electrochemical biosensors for the detection of creatinine based on commercial screen-printed carbon electrodes (SPCEs). The non-enzymatic creatinine sensor was constructed by the electrochemical deposition of AuNPs onto the surface of a pre-activated SPCE by electrochemical activation, followed by the surface modification of a Nafion membrane. The developed AuNPs/SCPE exhibited excellent reproducibility, and the proposed Nafion/AuNPs/SPCE sensor showed excellent detection sensitivity and selectivity toward creatinine. In comparison, the enzymatic creatinine biosensor was gradually established by the electrodeposition of a Prussian blue (PB) membrane on the optimal AuNPs/SCPE surface, followed by multi-enzyme cascade modification (which consisted of creatinine amidohydrolase (CA), creatine oxidase (CI) and sarcosine oxidase (SOx)) and drop-casting the Nafion membrane to stabilize the interface. The introduction of a PB interlayer acted as the redox layer to monitor the generation of hydrogen peroxide (H₂O₂) produced by the enzymatic reaction, while the Nafion membrane enhanced the detection selectivity toward creatine, and the multi-enzyme cascade modification further increased the detection specificity. Both non-enzymatic and enzymatic creatinine sensors could detect the lowest concentrations of less than or equal to 10 μM. In addition, the efficiency and reproducibility of the proposed composite biosensor were also confirmed by repetitive electrochemical measurements in human serum, which showed a positive linear calibration relation of peak currents versus the logarithm of the concentration between 10 μM and 1000 μM, namely, i_p (μA) = −7.06 lgC (μM) −5.30, R² = 0.996. This work offers a simple and feasible approach to the development of enzymatic and non-enzymatic creatinine biosensors. Full article

In this study, a simple, mild, and eco-friendly cold plasma-solution interaction method is employed to rapidly prepare gold colloids. Through modification with multi-walled carbon nanotubes (MWCNTs), a non-enzymatic glucose-sensing electrode material is successfully fabricated. The prepared electrode material is characterized via X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), and transmission electron microscopy (TEM). The results show that compared with the chemically reduced AuNPs-C-MWCNTs, the plasma-prepared AuNPs-P-MWCNTs exhibits enhanced glucose catalytic performance with a higher sensitivity of 73 μA·mM⁻¹·cm⁻² (approximately 3.2 times that of AuNPs-C-MWCNTs), lower response time of 2.1 s, and ultra-low detection limit of 0.21 μM. It also demonstrates excellent selectivity, reproducibility (RSD = 4.37%), repeatability (RSD = 3.67%), and operational stability (RSD = 4.51%). This improvement can be attributed to the smaller particle size and better dispersion of plasma-derived AuNPs on the surface of MWCNTs. Furthermore, the AuNPs-P-MWCNTs surface is enriched with oxygen-containing functional groups, which is conducive to the enhancement of the hydrophilicity of the electrode surface. These synergistic effects facilitate the AuNPs-catalyzed glucose oxidation reaction, ultimately leading to superior glucose catalytic performance. Full article

Phenolic acids, as widely distributed secondary metabolites in plants, possess significant biological activities, such as antioxidant and anti-inflammatory effects. However, their practical applications are limited by low absorption rates and poor bioavailability. Biotransformation technology, with its advantages of strong substrate specificity and mild reaction conditions, has become an effective strategy for the directional modification of phenolic acid molecular structures and the preparation of high-value-added derivatives. Among the various methodologies, enzymatic methods stand out due to their high selectivity and specificity, establishing them as a key approach for phenolic acid biotransformation. The research indicates that coordinated multi-pathway approaches, including decarboxylation, reduction, and hydrolysis, can effectively enhance the efficiency of phenolic acid biotransformation. This review systematically examines the structure and mechanism of action of the key enzymes involved in the phenolic acid biotransformation process. It also proposes innovative pathways and future development directions for existing technologies. Furthermore, it provides an in-depth analysis of the specific application potential of these key enzymes within the food sector. The objective of this review is to furnish a theoretical foundation and technical support for the efficient application of enzymatic methods in phenolic acid biotransformation, thereby accelerating their practical implementation. Full article

Nicotinamide mononucleotide (NMN) has emerged as a promising non-natural cofactor with significant potential to transform biocatalysis, synthetic biology, and therapeutic applications. By modulating NAD⁺ metabolism, NMN offers unique advantages in enzymatic reactions, metabolic engineering, and regenerative medicine. This review provides a comprehensive analysis of NMN’s biochemical properties, mechanisms of action, and diverse applications. Emphasis is placed on its role in addressing challenges in multi-enzyme cascades, biofuel production, and the synthesis of high-value chemicals. The paper also highlights critical research gaps, including the need for scalable NMN synthesis methods, improved integration into enzymatic systems, and comprehensive toxicity studies for therapeutic use. Emerging technologies such as AI-driven enzyme design and CRISPR-based genome engineering are discussed as transformative tools for optimizing NMN-dependent pathways. Furthermore, the synergistic potential of NMN with synthetic biology innovations, such as cell-free systems and dynamic regulatory networks, is explored, paving the way for precise and modular biotechnological solutions. Looking forward, NMN’s versatility as a cofactor positions it as a pivotal tool in advancing sustainable bioprocessing and precision medicine. Addressing current limitations through interdisciplinary approaches will enable NMN to redefine the boundaries of metabolic engineering and therapeutic innovation. This review serves as a roadmap for leveraging NMN’s potential across diverse scientific and industrial domains. Full article

Nucleoside phosphorylases (NPs) are pivotal enzymes in the salvage pathway, catalyzing the reversible phosphorolysis of nucleosides to produce nucleobases and α-D-ribose 1-phosphate. Due to their efficiency in catalyzing nucleoside synthesis from purine or pyrimidine bases, these enzymes hold significant industrial importance in the production of nucleoside-based drugs. Given that the thermodynamic equilibrium for purine NPs (PNPs) is favorable for nucleoside synthesis—unlike pyrimidine NPs (PyNPs, UP, and TP)—multi-enzymatic systems combining PNPs with PyNPs, UPs, or TPs are commonly employed in the synthesis of nucleoside analogs. In this study, we report the first development of two engineered bifunctional fusion enzymes, created through the genetic fusion of purine nucleoside phosphorylase I (PNP I) and thymidine phosphorylase (TP) from Thermus thermophilus. These fusion constructs, PNP I/TP-His and TP/PNP I-His, provide an innovative one-pot, single-step alternative to traditional multi-enzymatic synthesis approaches. Interestingly, both fusion enzymes retain phosphorolytic activity for both purine and pyrimidine nucleosides, demonstrating significant activity at elevated temperatures (60–90 °C) and within a pH range of 6–8. Additionally, both enzymes exhibit high thermal stability, maintaining approximately 80–100% of their activity when incubated at 60–80 °C over extended periods. Furthermore, the transglycosylation capabilities of the fusion enzymes were explored, demonstrating successful catalysis between purine (2′-deoxy)ribonucleosides and pyrimidine bases, and vice versa. To optimize reaction conditions, the effects of pH and temperature on transglycosylation activity were systematically examined. Finally, as a proof of concept, these fusion enzymes were successfully employed in the synthesis of various purine and pyrimidine ribonucleoside and 2′-deoxyribonucleoside analogs, underscoring their potential as versatile biocatalysts in nucleoside-based drug synthesis. Full article

Ethyl (S)-4-chloro-3-hydroxybutyrate ((S)-CHBE) is an important chiral intermediate in the synthesis of the cholesterol-lowering drug atorvastatin. Studying the use of SpyTag/SpyCatcher and SnoopTag/SnoopCatcher systems for the asymmetric reduction reaction and directed coupling coenzyme regeneration is practical for efficiently synthesizing (S)-CHBE. In this study, Spy and Snoop systems were used to construct a double-enzyme directed fixation system of carbonyl reductase (BsCR) and glucose dehydrogenase (BsGDH) for converting 4-chloroacetoacetate (COBE) to (S)-CHBE and achieving coenzyme regeneration. We discussed the enzymatic properties of the immobilized enzyme and the optimal catalytic conditions and reusability of the double-enzyme immobilization system. Compared to the free enzyme, the immobilized enzyme showed an improved optimal pH and temperature, maintaining higher relative activity across a wider range. The double-enzyme immobilization system was applied to catalyze the asymmetric reduction reaction of COBE, and the yield of (S)-CHBE reached 60.1% at 30 °C and pH 8.0. In addition, the double-enzyme immobilization system possessed better operational stability than the free enzyme, and maintained about 50% of the initial yield after six cycles. In summary, we show a simple and effective strategy for self-assembling SpyCatcher/SnoopCatcher and SpyTag/SnoopTag fusion proteins, which inspires building more cascade systems at the interface. It provides a new method for facilitating the rapid construction of in vitro immobilized multi-enzyme complexes from crude cell lysate. Full article

Carbohydrate-based surfactants are amphiphilic compounds containing hydrophilic moieties linked to hydrophobic aglycones. More specifically, carbohydrate esters are biosourced and biocompatible surfactants derived from inexpensive renewable raw materials (sugars and fatty acids). Their unique properties allow them to be used in various areas, such as the cosmetic, food, and medicine industries. These multi-applications have created a worldwide market for biobased surfactants and consequently expectations for their production. Biobased surfactants can be obtained from various processes, such as chemical synthesis or microorganism culture and surfactant purification. In accordance with the need for more sustainable and greener processes, the synthesis of these molecules by enzymatic pathways is an opportunity. This work presents a state-of-the-art lipase action mode, with a focus on the active sites of these proteins, and then on four essential parameters for optimizing the reaction: type of lipase, reaction medium, temperature, and ratio of substrates. Finally, this review discusses the latest trends and recent developments, showing the unlimited potential for optimization of such enzymatic syntheses. Full article

Microfluidic devices have attracted much attention in the current day owing to the unique advantages they provide. However, their application for industrial use is limited due to manufacturing limitations and high cost. Moreover, the scaling-up process of the microreactor has proven to be difficult. Three-dimensional (3D) printing technology is a promising solution for the above obstacles due to its ability to fabricate complex structures quickly and at a relatively low cost. Hence, combining the advantages of the microscale with 3D printing technology could enhance the applicability of microfluidic devices in the industrial sector. In the present work, a 3D-printed single-channel immobilized enzyme microreactor with a volume capacity of 30 μL was designed and created in one step via the fused deposition modeling (FDM) printing technique, using polylactic acid (PLA) as the printing material. The microreactor underwent surface modification with chitosan, and β-glucosidase from Thermotoga maritima was covalently immobilized. The immobilized biocatalyst retained almost 100% of its initial activity after incubation at different temperatures, while it could be effectively reused for up to 10 successful reaction cycles. Moreover, a multi-channel parallel microreactor incorporating 36 channels was developed, resulting in a significant increase in enzymatic productivity. Full article

Phosphorus is an essential nutrient for all life on earth and has a major impact on plant growth and crop yield. The forms of phosphorus that can be directly absorbed and utilized by plants are mainly HPO₄²⁻ and H₂PO₄⁻, which are known as usable phosphorus. At present, the total phosphorus content of soils worldwide is 400–1000 mg/kg, of which only 1.00–2.50% is plant-available, which seriously affects the growth of plants and the development of agriculture, resulting in a high level of total phosphorus in soils and a scarcity of available phosphorus. Traditional methods of applying phosphorus fertilizer cannot address phosphorus deficiency problems; they harm the environment and the ore material is a nonrenewable natural resource. Therefore, it is imperative to find alternative environmentally compatible and economically viable strategies to address phosphorus scarcity. Phosphorus-solubilizing bacteria (PSB) can convert insoluble phosphorus in the soil into usable phosphorus that can be directly absorbed by plants, thus improving the uptake and utilization of phosphorus by plants. However, there is no clear and systematic report on the mechanism of action of PSB. Therefore, this paper summarizes the discovery process, species, and distribution of PSB, focusing on the physiological mechanisms outlining the processes of acidolysis, enzymolysis, chelation and complexation reactions of PSB. The related genes regulating PSB acidolysis and enzymatic action as well as genes related to phosphate transport and the molecular direction mechanism of its pathway are examined. The effects of PSB on the structure and abundance of microbial communities in soil are also described, illustrating the mechanism of how PSB interact with microorganisms in soil and indirectly increase the amount of available phosphorus in soil. And three perspectives are considered in further exploring the PSB mechanism in utilizing a synergistic multi-omics approach, exploring PSB-related regulatory genes in different phosphorus levels and investigating the application of PSB as a microbial fungicide. This paper aims to provide theoretical support for improving the utilization of soil insoluble phosphorus and providing optimal management of elemental phosphorus in the future. Full article

Gold nanoparticles (AuNPs) can be described as nanozymes, species that are able to mimic the catalytic activities of several enzymes, such as oxidase/peroxidase, reductase, or catalase. Most studies in the literature focus on the colloidal suspension of AuNPs, and it is obvious that their immobilization could open the doors to new applications thanks to their increased stability in this state. This work aimed to investigate the behavior of surfaces covered by immobilized AuNPs (iAuNPs). Citrate-stabilized AuNPs (AuNPs-cit) were synthesized and immobilized on glass slides using a simple dip coating method. The resulting iAuNPs were characterized (surface plasmon resonance, microscopy, quantification of immobilized AuNPs), and their multi-enzymatic-like activities (oxidase-, peroxidase-, and catalase-like activity) were evaluated. The comparison of their activities versus AuNPs-cit highlighted their added value, especially the preservation of their activity in some reaction media, and their ease of reuse. The huge potential of iAuNPs for heterogeneous catalysis was then applied to the degradation of two model molecules of hospital pollutants: metronidazole and methylene blue. Full article

Ordered mesoporous carbon CMK-3 sieves with a hexagonal structure and uniform pore size have recently emerged as promising materials for applications as adsorbents and electrodes. In this study, using sucrose as the sustainable carbon source and SBA-15 as a template, CMK-3 sieves are synthesized to form bioelectrocatalytic immobilization matrices for enzymatic biofuel cell (EFC) electrodes. Their electrochemical performance, capacitive features, and the stability of enzyme immobilization are analyzed and compared to commercially available multi-walled carbon nanotubes (MWCNT) using cyclic voltammetry and electrochemical impedance spectroscopy (EIS). The anodic reaction in the presence of glucose oxidase (GOx) and ferrocene methanol (FcMeOH) on the sustainably sourced CMK-3-based electrodes produces bioelectrocatalytic current responses at 0.5 V vs. saturated calomel electrode (SCE) that are twice as high as on the MWCNT-based electrodes under saturated glucose conditions. For the cathodic reaction, the MWCNT-based cathode performs marginally better than the CMK-3-based electrodes in the presence of bilirubin oxidase (BOD) and 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS²⁻). The CMK-3-based EFCs assembled from the GOx anode and BOD cathode results in a power output of 93 μW cm⁻². In contrast, the output power of MWCNT-based EFCs is approximately 53 μW cm⁻². The efficiency of CMK-3 as a support material for biofuel cell applications is effectively demonstrated. Full article

The objective of this study is to create a reliable predictive model for the electrochemical performance of self-powered biosensors that rely on urea-based biological energy sources. Specifically, this model focuses on the development of a human energy harvesting model based on the utilization of urea found in sweat, which will enable the development of self-powered biosensors. In the process, the potential of urea hydrolysis in the presence of a urease enzyme is employed as a bioreaction for self-powered biosensors. The enzymatic reaction yields a positive potential difference that can be harnessed to power biofuel cells (BFCs) and act as an energy source for biosensors. This process provides the energy required for self-powered biosensors as biofuel cells (BFCs). To this end, initially, the platinum electrodes are modified by multi-walled carbon nanotubes to increase their conductivity. After stabilizing the urease enzyme on the surface of the platinum electrode, the amount of electrical current produced in the process is measured. The optimal design of the experiments is performed based on the Taguchi method to investigate the effect of urea concentration, buffer concentration, and pH on the generated electrical current. A general equation is employed as a prediction model and its coefficients calculated using an evolutionary strategy. Also, the evaluation of effective parameters is performed based on error rates. The obtained results show that the established model predicts the electrical current in terms of urea concentration, buffer concentration, and pH with high accuracy. Full article

Neurodegenerative diseases affect millions of people worldwide. The failure of the enzymatic degradation, the oxidative stress, the dyshomeostasis of metal ions, among many other biochemical events, might trigger the pathological route, but the onset of these pathologies is unknown. Multi-target and multifunctional molecules could address several biomolecular issues of the pathologies. The tripeptide GHK, a bioactive fragment of several proteins, and the related copper(II) complex have been largely used for many purposes, from cosmetic to therapeutic applications. GHK derivatives were synthesized to increase the peptide stability and improve the target delivery. Herein we report the synthesis of a new biotin–GHK conjugate (BioGHK) through orthogonal reactions. BioGHK is still capable of coordinating copper(II), as observed by spectroscopic and spectrometric measurements. The spectroscopic monitoring of the copper-induced ascorbate oxidation was used to measure the antioxidant activity Cu(II)-BioGHK complex, whereas antiglycant activity of the ligand towards harmful reactive species was investigated using MALDI-TOF. The affinity of BioGHK for streptavidin was evaluated using a spectrophotometric assay and compared to that of biotin. Finally, the antiaggregant activity towards amyloid-β was evaluated using a turn-on fluorescent dye. BioGHK could treat and/or prevent several adverse biochemical reactions that characterize neurodegenerative disorders, such as Alzheimer’s disease. Full article