Biotransformation of 2,4,6-Trinitrotoluene by a cocktail of native laccases from Pycnoporus sanguineus CS43 under oxygenic and non-oxygenic atmospheres.

2,4,6-Trinitrotoluene (TNT) is a highly toxic nitroaromatic explosive known for its environmental consequences, contaminating soil and groundwater throughout its life cycle, from production to disposal. Therefore, the urgency of developing innovative and ecological strategies to remedy the affected areas is recognized. This study reports, for the first time, the enzymatic biotransformation of TNT by a cocktail of native laccases from Pycnoporus sanguineus CS43. The laccases displayed efficient TNT conversion under both oxygenic and non-oxygenic conditions, achieving biotransformation rates of 80% and 87% within 48 h at a temperature of 60 °C and pH 7. Preliminary kinetic constants were calculated with the laccase cocktail, being a Vmax of 1.133 µM min-1 and 0.2984 µM min-1, and the Km values were 1586 µM and 458 µM, in an oxygenic and non-oxygenic atmosphere, respectively. High-performance liquid chromatography-mass spectrometry (HPLC/MS) confirmed the formation of amino dinitrotoluene isomers and hydroxylamine isomers as biotransformation products. In summary, this study suggests the potential application of laccases for the direct biotransformation of recalcitrant compounds like TNT, offering an environmentally friendly approach to address contamination issues.

Metal-organic frameworks for enzyme immobilization and nanozymes: A laccase-focused review.

Laccases are natural catalysts with remarkable catalytic activity. However, their application is limited by their lack of stability. Metal-organic frameworks (MOFs) have emerged as a promising alternative for enzyme immobilization. Enzymes can be immobilized in MOFs via two approaches: postsynthetic immobilization and in situ immobilization. In postsynthetic immobilization, an enzyme is embedded after MOF formation by covalent interactions or adsorption. In contrast, in in situ immobilization, a MOF is formed in the presence of an enzyme. Additionally, MOFs have exhibited intrinsic enzyme-like activity. These materials, known as nanozymes when they have the ability to replace enzymes in certain catalytic processes, have multiple key advantages, such as low cost, easy preparation, and large surface areas. This review presents a general overview of the most recent advances in both enzyme@MOF biocatalysts and MOF-based nanozymes in different applications, with a focus on laccase, which is one of the most widely investigated enzymes with excellent industrial potential.

Microscopical Evaluation of the Effects of High-Pressure Processing on Milk Casein Micelles.

The effect of different high-pressure processing (HPP) treatments on casein micelles was analyzed through scanning electron microscopy (SEM) and a particle size distribution analysis. Raw whole and skim milk samples were subjected to HPP treatments at 400, 500 and 600 MPa for Come-Up Times (CUT) up to 15 min at ambient temperature. Three different phenomena were observed in the casein micelles: fragmentation, alterations to shape and agglomeration. The particle size distribution analysis determined that, as pressure and time treatment increased, the three phenomena intensified. First, the size of the casein micelles began to decrease as their fragmentation occurred. Subsequently, the casein micelles lost roundness, and their shape deformed. Finally, in the most intense treatments (higher pressures and/or longer times), the micelles fragments began to agglomerate, which resulted in an increase in their average diameter. Homogenization and defatting had no significant effect on the casein micelles; however, the presence of fat in whole milk samples was bioprotective, as the effects of the three phenomena appeared faster in treated skim milk samples. Through this study, it was concluded that the size and structure of casein micelles are greatly altered during high-pressure treatment. These results provide information that broadens the understanding of the changes induced on casein micelles by high-pressure treatments at room temperature.

Effect of High Hydrostatic Pressure and Pulsed Electric Fields Processes on Microbial Safety and Quality of Black/Red Raspberry Juice.

Black and red raspberries are fruits with a high phenolic and vitamin C content but are highly susceptible to deterioration. The effect of high hydrostatic pressure (HHP 400−600 MPa/CUT-10 min) and pulsed electric fields (PEF, frequency 100−500 Hz, pulse number 100, electric field strength from 11.3 to 23.3 kV/cm, and specific energy from 19.7 to 168.4 kJ/L) processes on black/red raspberry juice was studied. The effect on the inactivation of microorganisms and pectin methylesterase (PME) activity, physicochemical parameters (pH, acidity, total soluble solids (°Brix), and water activity (aw)), vitamin C and phenolic compounds content were also determined. Results reveal that all HHP-treatments produced the highest (p < 0.05) log-reduction of molds (log 1.85 to 3.72), and yeast (log 3.19), in comparison with PEF-treatments. Increments in pH, acidity, and TSS values attributed to compounds' decompartmentalization were found. PME activity was partially inactivated by HHP-treatment at 600 MPa/10 min (22% of inactivation) and PEF-treatment at 200 Hz/168.4 kJ/L (19% of inactivation). Increment in vitamin C and TPC was also observed. The highest increment in TPC (79% of increment) and vitamin C (77% of increment) was observed with PEF at 200 Hz/168.4 kJ/L. The putative effect of HHP and PEF on microbial safety, enzyme inactivation, and phytochemical retention is also discussed in detail. In conclusion, HHP and PEF improve phytochemical compounds' content, microbial safety, and quality of black/red raspberry juice.

High Hydrostatic Pressure Induced Changes in the Physicochemical and Functional Properties of Milk and Dairy Products: A Review.

High-pressure processing (HPP) is a nonthermal technology used for food preservation capable of generating pasteurized milk products. There is much information regarding the inactivation of microorganisms in milk by HPP, and it has been suggested that 600 MPa for 5 min is adequate to reduce the number of log cycles by 5-7, resulting in safe products comparable to traditionally pasteurized ones. However, there are many implications regarding physicochemical and functional properties. This review explores the potential of HPP to preserve milk, focusing on the changes in milk components such as lipids, casein, whey proteins, and minerals, and the impact on their functional and physicochemical properties, including pH, color, turbidity, emulsion stability, rheological behavior, and sensory properties. Additionally, the effects of these changes on the elaboration of dairy products such as cheese, cream, and buttermilk are explored.

Potentialities of active membranes with immobilized laccase for Bisphenol A degradation.

Herein, we report the development of immobilized laccase based membrane bioreactor as a novel bio-catalytic system for the degradation of emerging endocrine disruptor i.e., Bisphenol A. Two laccase forms i.e. (1) in-house isolated and purified from an indigenous white-rot fungi Pycnoporus sanguineus (CS43) and (2) Trametes versicolor (commercial laccase from Sigma-Aldrich®) were immobilized on a multi-channel ceramic membrane (1.4µm in diameter) using 4% glutaraldehyde as a cross-linking agent. The immobilization yield and bisphenol A degradation activities of immobilized laccases were recorded at various pH levels. The surface topographies of immobilized-laccase membranes were accessed by scanning electron microscopy. In this study, 100% degradation of bisphenol A (20mg/L) was achieved in less than 24h in the presence of laccase from P. sanguineus (CS43) (620.55±14.85U/L) and T. versicolor (620.55±14.85U/L). The enzymes showed an optimal activity at pH 5 and 5.4 with a degradation rate of 204.8±1.8 and 79.0±0.1µmol/min/U for P. sanguineus (CS43) and T. versicolor, respectively. In conclusion, the highest immobilization of unit per square centimeter and efficient degradation potentiality strongly recommend the newly developed immobilized laccase based membrane bioreactor as a novel tool to tackle emerging contaminants degradation issues.