Search Results (399)

The non-monotonic behavior of amperometric enzyme-based biosensors under uncompetitive and noncompetitive (mixed) substrate inhibition is investigated computationally using a two-compartment model consisting of an enzyme layer and an outer diffusion layer. The model is based on a system of reaction–diffusion equations that includes a nonlinear term associated with non-Michaelis–Menten kinetics of the enzymatic reaction and accounts for the partitioning between layers. In addition to the known effect of substrate inhibition, where the maximum biosensor current differs from the steady-state output, it has been determined that external diffusion limitations can also cause the appearance of a local minimum in the current. At substrate concentrations greater than both the Michaelis–Menten constant and the uncompetitive substrate inhibition constant, and in the presence of external diffusion limitation, the transient response of the biosensor, after immersion in the substrate solution, may follow a five-phase pattern depending on the model parameter values: it starts from zero, reaches a global or local maximum, decreases to a local minimum, increases again, and finally decreases to a steady intermediate value. The biosensor performance is analyzed numerically using the finite difference method. Full article

Background: N^ε-(carboxymethyl)-lysine (CML) is formed in the human body by non-enzymatically driven reactions including glycation, oxidation, and lipoxidation. CML is a ubiquitous product of normal physiology, but its levels are increased under disease conditions like chronic kidney disease (CKD) and diabetes mellitus (DM). Free CML is eliminated from the human body mainly through kidney excretion, and its accumulation in the kidney tissue is linked to CKD pathogenesis. Aim: The main goal of this study was to evaluate the relative contribution of CKD and Type 2 DM (T2DM) to the accumulation of CML in patients’ sera. Methods: The study included 22 patients with CKD without DM, 55 with CKD and comorbid T2DM, and 21 with T2DM without CKD. Serum CML levels were measured by ELISA. The Kruskal-Wallis test was used to detect differences among groups. Spearman correlation analysis was performed, and the one-tailed Dunn test was considered to indicate statistical significance at p < 0.05. Results: The median serum CML levels (CKD, 658.4 ± 434.3 ng/mL; CKD + T2DM, 431.3 ± 327.9 ng/mL; T2DM, 273.9 ± 134.2 ng/mL) differed significantly (p < 0.05) among the three patient groups. A positive correlation was observed between serum CML and microalbuminuria (p = 0.004; r = 0.58), proteinuria (p = 0.002; r = 0.6), and age (p = 0.007; r = 0.52) only in the CKD patients. In all T2DM patients, independent of CKD status, serum CML correlated negatively (p < 0.05) with postprandial glucose and duration of diabetes, while its correlation with fasting glucose and HbA1c was negative only in the T2DM cohort without CKD. Conclusions: In patients with CKD, higher levels of CML were observed compared to those with T2DM. Serum CML correlated positively with proteinuria, albuminuria, and patient age in non-diabetic CKD patients, and negatively with blood glucose, HbA1c, and DM duration of T2DM in patients without CKD. Full article

Background: This study is the first to investigate the association between colorectal cancer (CRC) risk and the hMLH1 −93G>A and hMSH2 1032G>A polymorphisms of mismatch repair (MMR) genes in the Azerbaijani population. Methods: Peripheral blood samples containing EDTA were collected from the study subjects (134 patients and 137 controls), and genomic DNA was extracted using the non-enzymatic salting-out method. Genotypes were determined by polymerase chain reaction–restriction fragment length polymorphism (PCR-RFLP), and the results were visualized through agarose gel electrophoresis. Results: Overall, no statistically significant correlation was observed between CRC risk and the hMLH1 −93G>A polymorphism in the heterozygous GA (OR = 0.760; 95% CI = 0.374–1.542; p = 0.446), the mutant AA (OR = 1.474; 95% CI = 0.738–2.945; p = 0.270), or the A allele (OR = 1.400; 95% CI = 0.984–1.995; p = 0.062). However, in contrast to the dominant model, a statistically significant association was found between the recessive model and an increased CRC risk, with an odds ratio of 1.788 (95% CI = 1.102–2.900; p = 0.018). The hMLH1 −93G>A polymorphism was identified at a significantly higher frequency across the TNM stages, with the distribution showing statistical significance (p < 0.05). Additionally, no statistically significant association was observed between the hMSH2 1032G>A polymorphism and CRC risk. Conclusions: Although no overall association was observed for hMLH1 −93G>A, our findings suggest a potential link with increased colorectal cancer risk under the recessive model in the Azerbaijani population. Further studies are warranted to confirm this model-specific association and investigate the underlying biological mechanisms. 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

Four new minor monosulfated triterpene penta- and hexaosides, cladolosides S (1), S₁ (2), T (3), and T₁ (4), were isolated from the Vietnamese sea cucumber Cladolabes schmeltzii (Sclerodactylidae, Dendrochirotida). The structures of the compounds were established based on extensive analysis of 1D and 2D NMR spectra as well as HR-ESI-MS data. Cladodosides S (1), S₁ (2) and T (3), T₁ (4) are two pairs of dehydrogenated/hydrogenated compounds that share identical carbohydrate chains. The oligosaccharide chain of cladolosides of the group S is new for the sea cucumber glycosides due to the presence of xylose residue attached to C-4 Xyl1 in combination with a sulfate group at C-6 MeGlc4. The oligosaccharide moiety of cladolosides of the group T is unique because of the position of the sulfate group at C-3 of the terminal sugar residue instead of the 3-O-Me group. This suggests that the enzymatic processes of sulfation and O-methylation that occur during the biosynthesis of glycosides can compete with each other. This can presumably occur due to the high level of expression or activity of the enzymes that biosynthesize glycosides. The mosaicism of glycoside biosynthesis (time shifting or dropping out of some biosynthetic stages) may indicate a lack of compartmentalization inside the cells of organism producers, leading to a certain degree of randomness in enzymatic reactions; however, this also offers the advantage of providing chemical diversity of the glycosides. Analysis of the hemolytic activity of a series of 26 glycosides from C. schmeltzii revealed some patterns of structure–activity relationships: the presence or absence of 3-O-methyl groups has no significant impact, hexaosides, which are the final products of biosynthesis and predominant compounds of the glycosidic fraction of C. schmeltzii, are more active than their precursors, pentaosides, and the minor tetraosides, cladolosides of the group A, are weak membranolytics and therefore are not synthesized in large quantities. Two glycosides from C. schmeltzii, cladolosides D (18) and H₁ (26), display selectivity of cytotoxic action toward triple-negative breast cancer cells MDA-MB-231, while remaining non-toxic in relation to normal mammary cells MCF-10A. Quantitative structure–activity relationships (QSAR) were calculated based on the correlational analysis of the physicochemical properties and structural features of the glycosides and their hemolytic and cytotoxic activities against healthy MCF-10A cells and cancer MCF-7 and MDA-MB-231 cell lines. QSAR highlighted the complexity of the relationships as the cumulative effect of many minor contributions from individual descriptors can have a significant impact. Furthermore, many structural elements were found to have different effects on the activity of the glycosides against different cell lines. The opposing effects were especially pronounced in relation to hormone-dependent breast cancer cells MCF-7 and triple-negative MDA-MB-231 cells. Full article

During processing and storage, vegetables often experience undesirable color changes, including fading, lightening, or yellowing and softening, due to browning (enzymatic and non-enzymatic) and chlorophyll degradation. These changes diminish commercial and nutritional value. Therefore, it is necessary to maintain vegetable color and improve the quality of vegetable-based dishes. To address these issues, it is a scientific and practical necessity to summarize and discuss existing strategies and innovative techniques. This review first highlights the mechanisms of vegetable browning. This review then provides a comprehensive overview of recent advances in methods for color preservation, focusing on underlying mechanisms and techniques for inhibiting color changes from physical, chemical, and biological perspectives. A review of innovative technologies suggests that effective color preservation in vegetables is achieved by inhibiting the conditions that lead to three unfavorable color change reactions: enzymatic browning, non-enzymatic browning, and chlorophyll degradation. Current research frequently employs combined approaches that integrate two or more techniques to mitigate these adverse color changes. Moreover, most of these methods could simultaneously inhibit the three reaction processes. Future research directions are proposed for in-depth investigations into the molecular mechanisms of color changes in vegetables and the impact of treatments on the nutritional value. 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

The “11th International Congress on Biocatalysis (biocat2024)” was part of a biennial series that unites the fields of biology and chemistry, attracting researchers from the life sciences, engineering, and computer science. This international forum provides an opportunity for scientists worldwide to connect, seek collaboration for future projects, and gain insights into contemporary topics and innovative techniques. Biocat covers a range of compelling subjects and recent advancements in biocatalysis, including enzyme discovery, evolution, and applications. This congress focused on six key topics: AI and computational methods, structure–function analysis and enzyme engineering, enzymatic and whole-cell biotransformations, reaction cascades (electro-, chemo-, and photoenzymatic synergies), bioprocess engineering and the design of smart reactors, and facing climate change through sustainability and a circular bioeconomy. In 2024, we welcomed 344 expert delegates alongside 21 internal attendees, including 154 women and 1 non-binary participant, bringing the total number of participants to an impressive 365. Established researchers and emerging scientists from academia and industry delivered a total of 119 presentations, comprising 59 standard lectures, 60 lightning talks, and 195 posters. Six industry exhibitors showcased their latest products and services, providing an excellent opportunity to strengthen the connection between science and industry. Furthermore, the biocat award, recognized as one of the most prestigious honors in biotechnology, was presented for the eleventh time in the categories of “Science in Academia”, “Lifetime Achievement,” and “Industry”. Full article

The Maillard reaction is a complex chemical reaction that occurs between nucleophilic groups, such as thiolates or amino groups primarily from amino acids, peptides, proteins, and carbonyl groups, particularly from reducing sugars. The pH value of the medium is a key parameter controlling the kinetics of the Maillard reaction, as it influences the concentration of nucleophilic groups. Other specific conditions of reaction medium such as temperature, reaction time (or residence time in a process), and water activity also significantly influence the Maillard reaction. Understanding the impact of these parameters is essential for optimizing the Maillard reaction to enhance sensory attributes, nutritional qualities, and product stability during the storage and distribution of the final products. The Maillard reaction is responsible for the formation of desirable sensory qualities such as flavor, aroma, color, and texture in cooked and thermally processed foods, in addition to the improvement of nutritional value and shelf life of foods. In contrast, there are limitations in its industrial applications, as it can also generate harmful compounds such as acrylamide, N(6)-carboxymethyllysine, furans, and heterocyclic amines, as well as undesired changes in the nutritional value of the food. This review provides an overview of the Maillard reaction’s mechanism, influencing parameters, pros and cons, as well as some food industrial applications. Full article

Industrial biotechnology offers a potential ecological solution for PET recycling under relatively mild reaction conditions via enzymatic degradation, particularly using the leaf branch compost cutinase (LCC) quadruple mutant ICCG. To improve the efficient downstream processing of this biocatalyst after heterologous gene expression with a suitable production host, protein crystallization can serve as an effective purification/capture step. Enhancing protein crystallization was achieved in recent studies by introducing electrostatic (and aromatic) interactions in two homologous alcohol dehydrogenases (Lb/LkADH) and an ene reductase (NspER1-L1,5) produced with Escherichia coli. In this study, ICCG, which is difficult to crystallize, was utilized for the application of crystal contact engineering strategies, resulting in ICCG mutant L50Y (ICCGY). Previously focused on the Lys-Glu interaction for the introduction of electrostatic interactions at crystal contacts, the applicability of the engineering strategy was extended here to an Arg-Glu interaction to increase crystallizability, as shown for ICCGY T110E. Furthermore, the rationale of the engineering approach is demonstrated by introducing Lys and Glu at non-crystal contacts or sites without potential interaction partners as negative controls. These resulting mutants crystallized comparably but not superior to the wild-type protein. As demonstrated by this study, crystal contact engineering emerges as a promising approach for rationally enhancing protein crystallization. This advancement could significantly streamline biotechnological downstream processing, offering a more efficient pathway for research and industry. Full article

Glycation influences skin aging through non-enzymatic reactions between reducing sugars and proteins, forming advanced glycation end-products (AGEs) that accelerate skin deterioration. This study evaluates the curative anti-glycation effects of an injectable formulation combining dual-molecular-weight hyaluronic acid (low and mid/high) with trehalose in methylglyoxal-induced glycation in human skin explants. Thirty-six human skin explants were allocated across five experimental groups in a 12-day study. Glycation was induced using methylglyoxal (500 μM) on days 1 and 4, followed by curative product administration on day 5. CML (Nε-(carboxymethyl)lysine) immunohistochemistry was performed to assess glycation levels in the reticular dermis at days 6, 8, and 12, with quantitative analysis conducted through standardized image analysis. The formulation significantly reduced CML formation by 60% on day 6 compared to untreated controls (p < 0.001). Under methylglyoxal-induced glycation stress the product showed sustained curative effects, with CML reductions of 69% on day 6 (p = 0.008), 68% on day 8 (p = 0.012), and 61% on day 12 (p = 0.033) compared to methylglyoxal treatment alone. Cell viability remained unaffected throughout the study period across all experimental conditions. The tested injectable formulation exhibits significant and sustained curative anti-glycation properties in human skin explants for 12 days, effectively counteracting methylglyoxal-induced glycation damage without affecting cell viability. These findings advance anti-aging skin interventions, offering a novel approach to address glycation-induced skin damage with potential applications in clinical dermatology and aesthetic medicine. Full article

Background: Mesenchymal stem cell-based therapy may be indicated in ischaemic heart disease. The use of autologous adipose-derived mesenchymal stromal cells (AdMSCs) offers regenerative potential due to their paracrine effects. The aim of this study was to expand and characterise adult human thymus-derived MSCs harvested during open heart surgery with respect to their stem cell and paracrine properties. Methods: Enzymatically and non-enzymatically isolated human thymic AdMSCs (ThyAdMSCs) were cultured in xeno-free media containing pooled human platelet lysate (pPL). MSC characterisation was performed. Ex vivo expanded ThyAdMSCs were differentiated into three lineages. Proliferative capacity and immunomodulatory properties were assessed by proliferation assays and mixed lymphocyte reaction, respectively. Gene expression analysis was performed by qPCR. Results: Both isolation methods yielded fibroblast-like cells with plastic adherence and high proliferation. Flow cytometry revealed distinct expression of MSC markers in the absence of haematopoietic cell surface markers. Ex vivo expanded ThyAdMSCs could be differentiated into adipocytes, osteocytes, and chondrocytes. Activated peripheral blood mononuclear cells were significantly reduced when co-cultured with ThyAdMSCs, indicating their ability to inhibit immune cells in vitro. Gene expression analysis showed significantly less IFNγ and TNFα, indicating an alteration of the activated and pro-inflammatory state in the presence of ThyAdMSCs. Conclusions: These results demonstrate an efficient method to generate AdMSCs from human thymus. These MSCs have a strong immunomodulatory capacity and are, therefore, a promising cell source for regenerative medicine. The culture conditions are crucial for cells to proliferate in culture. Further research could explore the use of ThyAdMSCs or their secretome in surgical procedures. Full article

Estimating confidence intervals in small or noisy datasets is a recurring challenge in biomolecular research, particularly when data contain outliers or exhibit high variability. This study introduces a robust statistical method that combines a hybrid bootstrap procedure with Steiner’s most frequent value (MFV) approach to estimate confidence intervals without removing outliers or altering the original dataset. The MFV technique identifies the most representative value while minimizing information loss, making it well suited for datasets with limited sample sizes or non-Gaussian distributions. To demonstrate the method’s robustness, we intentionally selected a dataset from outside the biomolecular domain: a fast-neutron activation cross-section of the ¹⁰⁹Ag(n, 2n)^108mAg reaction from nuclear physics. This dataset presents large uncertainties, inconsistencies, and known evaluation difficulties. Confidence intervals for the cross-section were determined using a method called the MFV–hybrid parametric bootstrapping (MFV-HPB) framework. In this approach, the original data points were repeatedly resampled, and new values were simulated based on their uncertainties before the MFV was calculated. Despite the dataset’s complexity, the method yielded a stable MFV estimate of 709 mb with a 68.27% confidence interval of [691, 744] mb, illustrating the method’s ability to provide interpretable results in challenging scenarios. Although the example is from nuclear science, the same statistical issues commonly arise in biomolecular fields, such as enzymatic kinetics, molecular assays, and diagnostic biomarker studies. The MFV-HPB framework provides a reliable and generalizable approach for extracting central estimates and confidence intervals in situations where data are difficult to collect, replicate, or interpret. Its resilience to outliers, independence from distributional assumptions, and compatibility with small-sample scenarios make it particularly valuable in molecular medicine, bioengineering, and biophysics. Full article

Surimi gel, a type of hydrocolloidal food, is formed through the gelation of fish meat proteins. Myosin heavy chain (MHC), a key myofibrillar protein, plays a crucial role in the formation of the gel network via both transglutaminase (TGase)-catalyzed and non-enzymatic polymerization. Gel disintegration in surimi is primarily attributed to the proteolytic degradation of MHC. This study focused on golden threadfin bream Nemipterus virgatus, a species characterized by low TGase activity and high protease activity at elevated temperatures. We investigated the competition between non-enzymatic polymerization and proteolytic degradation of MHC and their effects on gel mechanical properties using a mathematical model. A mathematical model based on kinetic reactions accurately reflected the changes in MHC observed through SDS-PAGE analysis during N. virgatus gel disintegration. Our results indicate that not only unpolymerized but also polymerized MHC was significantly degraded, which substantially contributed to the reduction in the mechanical properties of the N. virgatus surimi. Mathematically understanding the dynamics of MHC in surimi during heating helps promote the utilization of noncommercial fish species for surimi processing by enabling better control over surimi gel properties. Full article