Search Results (122)

Space conditions offer new insights into fundamental biological and molecular mechanisms. The study aimed to evaluate the enzymatic activity of proteinase K (PK) under extreme conditions relevant to space environments: simulated microgravity, hypergravity, and gamma radiation. PK activity was tested using azocasein (AZO) as a chromogenic substrate, with enzymatic reactions monitored spectrophotometrically at 450 nm. A rotating wall vessel (RWV) simulated microgravity, centrifugation at 1000× g (3303 rpm) generated hypergravity, and gamma radiation exposure used cesium-137 as the ionizing source. PK activity showed no remarkable changes under microgravity after 16 or 48 h; however, higher absorbance values after 96 h indicated enhanced AZO proteolysis compared to 1 g (Earth gravity) controls. In hypergravity, low PK concentrations exhibited slightly increased activity, while higher concentrations led to reduced activity. Meanwhile, gamma radiation caused a dose-dependent decline in PK activity; samples exposed to deep-space equivalent doses showed reduced substrate degradation. PK retained enzymatic activity under all tested conditions, though the type and duration of stress modulated its efficiency. The results suggest that enzyme-based systems may remain functional during space missions and, in some cases, exhibit enhanced activity. Nevertheless, their behavior must be evaluated in a context-dependent manner. These findings may be significant to advance biotechnology, diagnostics, and the development of enzyme systems for space applications. Full article

The reusability of enzymes is a fundamental aspect of sustainable biotechnology and the development of biosensors. This study presents one of the first quantitative evaluations of DNA polymerase reusability by utilizing integrated quartz crystal microbalance (QCM) kinetics and real-time monitoring of exonuclease activity. The results showed that immobilized T7 DNA polymerase retained approximately 50% of its initial activity after three 90-min cycles and around 20% after five cycles. Significantly lower activities were observed for shorter, 45-min cycles. This indicates an unexpected time-dependent enhancement in stability for longer reaction times. The findings suggest a promising trend in enzyme stability and reusability, establishing a quantitative relationship between reaction duration and enzyme performance. This relationship offers a scalable pathway for the regeneration of biosensors and for sustainable enzymatic catalysis. Additionally, the work provides a transferable framework that can be applied to other DNA-processing enzymes, which supports long-term biosensor performance and industrial biocatalysis. The demonstrated approach offers a transferable and scalable methodology for the development of reusable polymerase-based biosensors and sustainable biocatalytic systems. Full article

Alginate is a natural polysaccharide extracted from brown algae and is commonly used as a biomaterial scaffold in tissue engineering. In this study, we performed phenol functionalization of sodium alginate based on chemical modification methods using 1-ethyl-(3-dimethylaminopropyl)carbodiimide/N-hydroxybutanediimide/2-(N-morpholino) ethanesulfonic acid (EDC/NHS/MES) and tyramine. The presence of phenol groups was confirmed by spectrophotometry and Fourier Transform Infrared. We successfully prepared hydrogels using a horseradish peroxidase/hydrogen peroxide (HRP/H₂O₂) enzymatic system as well as an sodium persulfate (SPS)/ruthenium light-crosslinking system. Optimization identified 1 mM ruthenium and 4 mM SPS as the most effective photo crosslinking conditions. At the same time, 1 mM H₂O₂ and 10 U/mL HRP are considered optimal conditions for the enzyme-linked reaction. Rheological measurements monitored the gelation process, revealing that the viscosity, storage modulus, and loss modulus of the material increased by at least one hundredfold after crosslinking. Thixotropy results demonstrated excellent recovery of the material. Texture analysis indicated that the crosslinked material possessed notable strength and toughness, highlighting its potential applications in tissue engineering after 3D bioprinting. Full article

Periodontitis is a biofilm-driven inflammatory disease in which conventional indices (probing depth, clinical attachment level, and radiographs) quantify tissue destruction without capturing the biology of infection. In this review, we synthesized microbiological diagnostics, from chairside tools to omics. We outline sampling strategies and emphasize the quantitative monitoring of bacterial load. Enzymatic assays (e.g., N-benzoyl-DL-arginine-2-naphthylamide hydrolysis assay test) measure functional activity at the point of care. Immunological methods include rapid immunochromatography for Porphyromonas gingivalis and enzyme-linked immunosorbent assay for the high-throughput measurement of bacterial antigens. Molecular platforms encompass quantitative polymerase chain reaction (qPCR) (TaqMan, SYBR, multiplex panels; propidium monoazide quantitative-qPCR for viable cells), checkerboard DNA–DNA hybridization for semi-quantitative community profiling, loop-mediated isothermal amplification (LAMP)/molecular beacon-LAMP for portable isothermal detection, and microarrays. Complementary modalities such as fluorescent in situ hybridization, next-generation sequencing, and Fourier transform infrared spectroscopy provide spatial, ecological, and biochemical resolutions. We discuss the limitations of current approaches, including sampling bias, presence–activity discordance, semi-quantitation, method biases, limited strain/function resolution, low-biomass artifacts, and lack of validated cutoffs. To address these challenges, we propose a pragmatic hybrid strategy: site-specific quantitative panels combined with activity and host-response markers interpreted alongside clinical metrics under standardized quality assurance/quality control. Priorities include outcome-linked thresholds, strain-aware/functional panels, robust point-of-care chemistry, and harmonized protocols to enable personalized periodontal care. Full article

Collagen peptides derived from fish skin may be limited in food applications due to undesirable flavors. To investigate the effects of Maillard reaction modification on their physicochemical and flavor properties, collagen peptides from tilapia skin were prepared via enzymatic hydrolysis, followed by the Maillard reaction with four reducing sugars (xylose, ribose, glucose and glucosamine) through a combined procedure involving simultaneous enzyme inactivation and Maillard reaction at 100 °C. The resultant Maillard reaction products (MRPs) were characterized by analyzing free amino groups, peptide size distribution and color difference, while the reaction progression was monitored using UV absorption and fluorescence spectroscopy. The flavor profile of MRPs was analyzed through quantitative descriptive sensory evaluation and GC-MS coupled with principal component analysis. Among the four reducing sugars tested, glucosamine-induced Maillard reaction products exhibited the most pronounced physicochemical and sensory improvements. Specifically, glucosamine-MRPs showed the greatest reduction in free amino groups (0.69 μmol/L) and a notable decrease in high-molecular-weight peptides (3.31%), accompanied by an increase in low-molecular-weight fractions. Colorimetric analysis revealed a marked color change (ΔE = 31.78), and spectral analysis further confirmed intensified UV absorbance and fluorescence intensity in the glucosamine group, indicating advanced reaction progression. Sensory evaluation demonstrated a significant reduction in bitterness and enhancement of umami and saltiness. Moreover, GC-MS analysis revealed that the glucosamine-treated group exhibited the most favorable volatile profile, characterized by an increase in aromatic compounds and a substantial decrease in undesirable odorants. This study provides a theoretical basis for controlling the undesirable flavor of collagen peptides through low-extent Maillard reactions by different reducing sugars. Full article

The valorization of agricultural by-products represents a sustainable strategy to reduce waste and create high-value biotechnological products. This review highlights Colombian plant-derived peroxidases (PODs) obtained from Guinea grass, royal palm, African oil palm, lemongrass, sleepy plant, and sweet potato. These enzymes catalyze oxidative reactions and show potential in biosensing, polymer synthesis, environmental remediation, and health monitoring. We summarize extraction and purification strategies while addressing current challenges such as operational stability, scalability, and cost. Special emphasis is given to applications like cross-linked enzymatic aggregates (CLEAs) and electrochemical biosensors, where Colombian PODs demonstrate superior stability and sensitivity compared to horseradish peroxidase (HRP). This review frames these advances within the circular bioeconomy, presenting insights into waste reduction and CO₂ savings. By integrating local biodiversity into innovative processes, Colombian PODs can drive sustainable technologies and provide industrial and environmental solutions. Full article

This study investigates the changes in mechanically harvested ‘Leccino’ olives stored under cold and room-temperature conditions from harvest up to 23 days of storage during two consecutive seasons. Variations in quality parameters, including maturity index, weight, firmness, and colour, were monitored throughout the storage period. In addition, the phenolic profile of the olives was analysed using HPLC, and the fatty acid composition was determined by GC–MS. These analyses enabled a comparison of changes across different storage durations, seasons, and storage conditions. Results show that fruit ripeness at harvest differed notably between the two seasons. In the second season, the olives displayed a higher maturity index, lower firmness, and lower content of certain individual phenolic compounds, indicating a more advanced stage of ripening compared to that of the previous year. These initial differences strongly influenced the subsequent development of fruit quality and biochemical characteristics during storage. Storage temperature had a significant effect on the quality parameters and metabolism. As expected, olives stored at room temperature lost their firmness and weight more quickly than those stored under cold conditions. The most abundant phenolic in olive fruit, oleuropein, degraded more rapidly at room temperature, resulting in a quicker accumulation of its derivatives. Fatty acids were more stable than phenolic compounds during storage, likely due to their lower susceptibility to enzymatic degradation and oxidative reactions under the tested conditions. While saturated fatty acids remained largely unchanged, a slight increase in unsaturated fatty acids was observed after 23 days of cold storage, possibly as a result of adaptations of the membrane to cold storage conditions. Full article

Myoglobin, a heme protein involved in oxygen storage and transport, also exhibits pseudo-peroxidase activity by catalyzing the breakdown of hydrogen peroxide. While this enzymatic function is well-documented, the potential for substrate inhibition at high hydrogen peroxide concentrations remains underexplored. In this study, we aimed to investigate the kinetic properties of myoglobin’s peroxidase-like activity, focusing on substrate inhibition over time. We employed spectrophotometric assays to monitor reaction rates of myoglobin exposed to increasing hydrogen peroxide concentrations. Our results show that myoglobin activity begins to decline within 3 min of exposure to hydrogen peroxide and reaches full inhibition after approximately 30 min. This progressive inhibition suggests that myoglobin undergoes a delayed inactivation process rather than an immediate loss of function. Additionally, we extended the investigation to hemoglobin, comparing its response to hydrogen peroxide, and preliminary data suggest differences in substrate inhibition dynamics. These findings provide new insights into the regulation of myoglobin’s oxidative function and suggest potential physiological implications for oxidative stress and redox balance, especially in relation to other heme proteins like hemoglobin. Full article

Isoquercitrin, a monoglucoside form of quercetin, exhibits superior antioxidant, anti-inflammatory, and cardiovascular protective effects in comparison to its precursor, rutin. However, its natural abundance is limited. This study aimed to increase the functional value of Diospyros lotus leaf extract through enzymatic conversion of rutin to isoquercitrin using α-l-rhamnosidase and to evaluate the changes in biological activities after conversion. A sugar-free D. lotus leaf extract was prepared and subjected to enzymatic hydrolysis with α-l-rhamnosidase under optimized conditions (pH 5.5, 55 °C, and 0.6 U/mL). Isoquercitrin production was monitored via high-performance liquid chromatography. Antioxidant and anti-inflammatory activities were assessed using the 2,2-diphenyl-1-picrylhydrazyl radical scavenging and lipoxygenase (LOX) inhibition assays, respectively. The enzymatic reaction resulted in complete conversion of 30 mM rutin into isoquercitrin within 180 min, increasing isoquercitrin content from 9.8 to 39.8 mM. The enzyme-converted extract exhibited significantly enhanced antioxidant activity, with a 48% improvement in IC₅₀ value compared with the untreated extract. Similarly, LOX inhibition increased from 39.2% to 48.3% after enzymatic conversion. Both extracts showed higher inhibition than isoquercitrin alone, indicating synergistic effects of other phytochemicals present in the extract. This study is the first to demonstrate that α-l-rhamnosidase-mediated conversion of rutin to isoquercitrin in D. lotus leaf extract significantly improves its antioxidant and anti-inflammatory activities. The enzymatically enhanced extract shows potential as a functional food or therapeutic ingredient. 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

Chlorothalonil and its toxic metabolite, 4-hydroxy-chlorothalonil, pose significant environmental and health risks. However, their simultaneous and accurate detection remains challenging due to their differing ionization efficiencies in mass spectrometry and the interference caused by enzymatic reactions in sulfur-rich vegetables. This study developed a UHPLC-MS/MS method for simultaneous detection of chlorothalonil and 4-hydroxy-chlorothalonil, using an atmospheric pressure chemical ionization (APCI) source, optimizing the probe temperature to 600 °C and a set of optimal chromatography parameters. A low-temperature and acidification synergistic enzyme inhibition strategy was developed, involving refrigerating samples and extraction reagents, acidifying with citric acid before sample homogenization, and extracting with formic acid/acetonitrile, significantly improving chlorothalonil recovery. Method validation demonstrated limits of detection (LOD) and quantification (LOQ) of 0.003 mg/kg and 0.01 mg/kg, respectively, with recoveries of 76.5–91.1% for chlorothalonil and 87.6–96.7% for 4-hydroxy-chlorothalonil. The method was successfully applied in monitoring the residue risks in sulfur-rich vegetables. Full article

A subject of increasing fundamental and technological interest is the techno- and bio-functionality of functional foods and nutraceuticals in high-solid gels. This encompasses the diffusion of natural bioactive compounds, prevention of oxidation of essential fatty acids, minimization of food browning, and the prevention of malodorous flavour formation in enzymatic and non-enzymatic reactions, to mention but a few. Textural and sensory considerations require that these delivery/encapsulating/entrapping vehicles are made with natural hydrocolloids and co-solutes in a largely amorphous state. It is now understood that the mechanical glass transition temperature is a critical consideration in monitoring the performance of condensed polymer networks that incorporate small bioactive compounds. This review indicates that the metastable properties of the rubber-to-glass transition in condensed gels (as opposed to the thermodynamic equilibrium in crystalline lattices) are a critical parameter in providing a fundamental quality control of end products. It appears that the “sophisticated synthetic polymer research” can provide a guide in the design of advanced biomaterials for targeted release or the prevention of undesirable byproducts. Such knowledge can assist in designing and optimizing functional foods and nutraceuticals, particularly those including vitamins, antioxidants, essential fatty acids, stimulants for performance enhancement, and antimicrobials. Full article

Advanced glycation end products (AGEs) are a group of compounds formed through non-enzymatic reactions between reducing sugars and proteins, lipids, or nucleic acids. AGEs can be generated in the body or introduced through dietary sources and smoking. Recent clinical and animal studies have highlighted the significant role of AGEs in various health conditions. These compounds accumulate in nearly all mammalian tissues and are associated with a range of diseases, including diabetes and its complications, cardiovascular disease, and neurodegeneration. This review summarizes the major diseases linked to AGE accumulation, presenting both clinical and experimental evidence. The pathologies induced by AGEs share common mechanisms across different organs, primarily involving oxidative stress, chronic inflammation, and direct protein cross-linking. Interventions targeting AGE-related diseases focus on inhibiting AGE formation using synthetic or natural antioxidants, as well as reducing dietary AGE intake through lifestyle modifications. AGEs are recognized as significant risk factors that impact health and accelerate aging, particularly in individuals with hyperglycemia. Monitoring AGE level and implementing nutritional interventions can help maintain overall health and reduce the risk of AGE-related complications. Full article

The Maillard reaction (MR) between sugars and amino acids, peptides, or proteins is understood to occur gradually during the production and aging of sparkling wines, where it contributes to caramel, roasted, and toasted aromas. Divalent metal ions can accelerate the MR, although this has not been previously reported in wine or wine-like conditions. In this work, the effect of calcium (Ca) and magnesium (Mg) ions on the concentration of 10 Maillard reaction-associated products (MRPs) was measured in modified sparkling base wine during accelerated aging at 50 °C for four weeks. Chardonnay base wine was modified by the addition of fructose (0.02 M) and a single amino acid (lysine, glycine, cysteine; 0.01 M) in combination with Ca²⁺ or Mg²⁺ at zero, low (10 mg/L), or high (50 mg/L) dose levels. MRPs were quantified by headspace solid-phase microextraction coupled with gas chromatography–mass spectrometry (HS-SPME-GC/MS), sugar concentration was measured by enzymatic assay, and amino acids and free metal ions were monitored by capillary electrophoresis. Fructose levels did not substantially decrease during aging despite increases in all MRPs, suggesting that trace sugars or α-dicarbonyl species present in the wine matrix likely play a greater role in MRP formation than fructose. Aging duration and amino acid content had a greater effect than metal addition on the composition of the MRPs. Treatments containing cysteine and 50 mg/L Ca²⁺ had elevated concentrations of benzaldehyde and furfural ethyl ether following 4 weeks of accelerated aging. This work identified key MRPs that increase during base wine accelerated aging and informs future research on the relationship between wine composition and aging markers. Full article