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• α-chymotrypsin-assisted synthesis of methotrexate oligomers; • Biosynthesis of oligomers up to 6 units; • Enzymatic oligomerization is reduced with the size of the substrate. The enzymatic synthesis of methotrexate (MTX) catalysed by α -chymotrypsin was studied for the first time. The proteolytic enzyme displayed activity for the synthesis of MTX oligomers composed by 6 repeating units (DP avg = 1.5). For longer oligomers, molecular dynamics simulations confirmed that as the oligomeric chain grows its accommodation in the enzymes' active site is hindered, which is evidenced by a decrease of the binding energy associated. The full characterization of the oligomers produced was performed by nuclear magnetic resonance (NMR, 1H and 13C), matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF), electrospray ionization (ESI) and differential scanning calorimetry (DSC). [ABSTRACT FROM AUTHOR]

Polyoxyethanyl-α-tocopheryl sebacate (PTS) is an amphiphilic compound with self-emulsifying properties known to form micelles. In this work, we report the production of PTS micelles for the encapsulation and delivery of a hydrophobic derivative of methotrexate, MTX di-ethylated (MTX-OEt). We optimized the micelles production by testing two different techniques: auxiliary solvent and sonication. Small and homogeneous micelles (≈40 nm) were obtained through the auxiliary solvent method performed at 30 °C and using 15 mg/mL of PTS. The produced micelles with the most promising physicochemical properties did not induce cytotoxicity when tested in normal human cells (BJ5ta cells), being considered for the encapsulation of MTX-OEt. This prodrug was achieved by Fisher esterification using ethanol, being isolated in good yield (η = 68%). MTX-OEt was efficiently encapsulated onto the produced micelles which preserved their physicochemical properties. The PTS micelles loaded with MTX-OEt, free MTX-OEt and free unmodified MTX revealed similar biological effect against cancer cells (Caco-2 cells). These results demonstrated that the biological activity of MTX is not altered after modification. The developed PTS micelles revealed a promising intracellular delivery performance with potentiality for cancer therapy. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • Laccase polymerization of phenolics by high-pressure homogenization. • Colouration of fabrics by enzymatically produced polymers. • Conductive fabrics by surface coating with new polymers. Abstract Coloured and conductive fabrics were obtained through "in situ" laccase polymerization of catechol and p -phenylenediamine under high-pressure homogenization. Both monomers, catechol and p -phenylenediamine, were polymerized by different laccase forms, namely native, PEGylated and Epoxy-PEGylated. All the catalysts were placed inside a textile fabric bag which served simultaneously as enzyme support and as substrate for coating with the newly produced polymers. The PEGylated laccase forms gave rise to a higher amount of oligomers/polymers and higher colouration level of polyethylene terephthalate (PET), cotton and wool fabrics compared to native laccase. Both functional polymers were able to confer conductivity to the substrates however in a different extent. Fabrics coated with poly(p -phenylenediamine) present higher conductivity, rather due to its polymerized structure than to the amount of polymer produced by enzyme catalysis. Herein a green approach was presented to produce polyphenols with increased fixation onto different textile substrates. These substrates reach high levels of colouration and good fastness behaviour after washing. [ABSTRACT FROM AUTHOR]

Abstract Laccase from Myceliophthora thermophila was used to catalyze the polymerization of catechol under high-pressure homogenization for the green coloration of textile substrates. The oxidation reactions were conducted using different forms of laccase, namely native laccase, PEGylated laccase and PEGylated laccase immobilized onto an epoxy resin. The three enzyme forms were deposited inside a polyester fabric bag during the experiments. The amount of polymer obtained was similar when using the three enzyme forms and its dispersion in water/DMSO mixture lead to powder particles of about 30–60 nm. The immobilized and PEGylated enzymes lead to poly(catechol) with 13 and 10 units, respectively, while the native form gave rise to shorter polymers (DP = 8). We have shown that the oxidation of catechol conducted under high-pressure homogenization can be an efficient methodology for the in situ coloration of textiles. The polymers produced by this methodology stained strongly the textile container, revealing this experimental set-up as a promising greener coloration/coating methodology involving milder conditions than the normally used in textile processes. Graphical abstract Image 1 Highlights • Laccase-assisted polymerization of catechol by high-pressure homogenization. • Polyethylene terephthalate (PET) bag as enzyme container. • Green coloration of PET fabric with enzymatically produced poly(catechol). [ABSTRACT FROM AUTHOR]

Several polyesters were synthesized from ethylene glycol, glycerol and adipate, succinate dimethyl esters. Immobilized Candida antarctica lipase B was used as catalyst for 6 h under vacuum at 70 °C without any further solvents. The highest conversion rate of 88.5% occurred for the polymerization of poly(ethylene adipate), evaluated by 1 H NMR. MALDI-TOF analysis indicated that most of the oligomers formed were dimers or trimers. After successfully synthesize the polyesters we set-up the optimal conditions for their in-situ coating onto cotton substrates with a soluble lipase from Thermomyces lanuginosus. This work presents a novel bio-approach to impart hydrophobic properties to coated cotton-based fibre materials. [ABSTRACT FROM AUTHOR]

Poloxamer 407 (P407)-based nanoparticles were produced by the high pressure homogenization method for the encapsulation and delivery of methotrexate (MTX), aiming intravenous therapeutic applications. The surface of these nanoparticles was functionalized by conjugation of P407 with folic acid (FA) or with MTX, which served as targeting ligand agents. MTX-P407 conjugate was also developed to increase the final drug cargo. Two hydrophobic derivatives of MTX, MTX di-ethylated ester (MTX-OEt) and the ionic complex MTX-dimethyldioctadecylammonium bromide (MTX-DODAB) were produced and entrapped onto P407-based nanoparticles. All formulations developed revealed a monodisperse character comprising small and narrow nanoparticles (<100 nm). P407 nanoparticles (functionalized with FA) and MTX-P407 nanoparticles, both loaded with MTX-OEt, demonstrated a slow drug release profile. The effect of lipase from Aspergillus oryzae on the hydrolysis of the linkage between the P407 and MTX, and consequent MTX release profile, was also evaluated. We observed a controlled and slow release of MTX (<50% of release after 11 days) in the presence of enzyme. These MTX-P407 nanoparticles loaded with MTX-OEt induced a great effect against Caco-2 cancer cells (≈40% of cell death after 72 h of incubation), demonstrating higher efficiency than the free MTX at the same concentration. [ABSTRACT FROM AUTHOR]