Enzymatic Synthesis and Structural Modeling of Bio-Based Oligoesters as an Approach for the Fast Screening of Marine Biodegradation and Ecotoxicity.

Given the widespread use of esters and polyesters in products like cosmetics, fishing nets, lubricants and adhesives, whose specific application(s) may cause their dispersion in open environments, there is a critical need for stringent eco-design criteria based on biodegradability and ecotoxicity evidence. Our approach integrates experimental and computational methods based on short oligomers, offering a screening tool for the rapid identification of sustainable monomers and oligomers, with a special focus on bio-based alternates. We provide insights into the relationships between the chemical structure and properties of bio-based oligomers in terms of biodegradability in marine environments and toxicity in benchmark organisms. The experimental results reveal that the considered aromatic monomers (terephthalic acid and 2,5-furandicarboxylic acid) accumulate under the tested conditions (OECD 306), although some slight biodegradation is observable when the inoculum derives from sites affected by industrial and urban pollution, which suggests that ecosystems adapt to non-natural chemical pollutants. While clean seas are more susceptible to toxic chemical buildup, biotic catalytic activities offer promise for plastic pollution mitigation. Without prejudice to the fact that biodegradability inherently signifies a desirable trait in plastic products, nor that it automatically grants them a sustainable "license", this study is intended to facilitate the rational design of new polymers and materials on the basis of specific uses and applications.

Synthesis, Characterization, and Antiproliferative Properties of New Bio-Inspired Xanthylium Derivatives.

Xanthylium derivatives are curcumin analogs showing photochromic properties. Similarly, to anthocyanins, they follow the same multistate network of chemical species that are reversibly interconverted by external stimuli. In the present work, two new asymmetric monocarbonyl analogues of curcumin, 4-(4-hydroxy-3-metoxybenzylidene)-1,2,3,4-tetrahydroxanthylium chloride (compound 3) and 4-(4-hydroxybenzylidene)-6-methoxy-1,2,3,4-tetrahydroxanthylium chloride (compound 4) were synthesized, and their photochromic and biological properties were investigated. The UV-Vis spectroscopy and the direct and reverse pH-jumps studies confirmed the halochromic properties and the existence of different molecular species. A network of chemical reactions of these species was proposed. Furthermore, the antiproliferative properties of both compounds were evaluated using P19 murine embryocarcinoma cells and compared with each other. The results demonstrate that both new xanthylium derivatives modify the progression through the cell cycle of P19 cells, which translates into a significant antiproliferative effect. The effect of the methoxy group position is discussed and several checkpoint proteins are advanced as putative targets.

Enzymatic route for selective glycerol oxidation using covalently immobilized laccases.

Glycerol is an important starting material for the synthesis of many chemical compounds and its selective oxidation represents an efficient way to produce value-added compounds. Glyceric acid, one of these selective oxidation products, is an important intermediate in the food, medicine, cosmetics, and light industries. In this work, four commercially available native laccases were screened for glycerol oxidation using different initiators, and the two most efficient biocatalysts were covalently immobilized on functionalized magnetic and polymethacrylate (Lifetech™) solid supports. Apart from the mostly employed Fe3O4 magnetic particles, in this work Ni-Zn or Ni-Zn-Co spinel ferrite (MFe2O4) microparticles were used. Particularly, the utilization (for the first time for laccase immobilization) of Ni-Zn ferrite support Ni0.7Zn0.3Fe2O4 functionalized with 3-aminopropyl-trimethoxysilane, via crosslinking by glutaraldehyde and reduction with NaBH4 led to excellent biocatalytic efficiency and stability. These results confirm the feasibility of Trametes versicolor laccase for covalent bonding, as presumed by computational modelling. The resulted enzymatic preparations were characterized in detail in terms of stability and reusability, demonstrating enhanced storage, pH and thermal stability compared to the native enzymes. The most active biocatalysts (790.93 [U/g]) were successfully used for glycerol oxidation and the specific conversion in glyceric acid exceeded 50%.

Understanding Marine Biodegradation of Bio-Based Oligoesters and Plasticizers.

The study reports the enzymatic synthesis of bio-based oligoesters and chemo-enzymatic processes for obtaining epoxidized bioplasticizers and biolubricants starting from cardoon seed oil. All of the molecules had MW below 1000 g mol-1 and were analyzed in terms of marine biodegradation. The data shed light on the effects of the chemical structure, chemical bond lability, thermal behavior, and water solubility on biodegradation. Moreover, the analysis of the biodegradation of the building blocks that constituted the different bio-based products allowed us to distinguish between different chemical and physicochemical factors. These hints are of major importance for the rational eco-design of new benign bio-based products. Overall, the high lability of ester bonds was confirmed, along with the negligible effect of the presence of epoxy rings on triglyceride structures. The biodegradation data clearly indicated that the monomers/building blocks undergo a much slower process of abiotic or biotic transformations, potentially leading to accumulation. Therefore, the simple analysis of the erosion, hydrolysis, or visual/chemical disappearance of the chemical products or plastic is not sufficient, but ecotoxicity studies on the effects of such small molecules are of major importance. The use of natural feedstocks, such as vegetable seed oils and their derivatives, allows the minimization of these risks, because microorganisms have evolved enzymes and metabolic pathways for processing such natural molecules.

Rational Guidelines for the Two-Step Scalability of Enzymatic Polycondensation: Experimental and Computational Optimization of the Enzymatic Synthesis of Poly(glycerolazelate).

The lipase-catalyzed polycondensation of azelaic acid and glycerol is investigated according to a Design-of-Experiment approach that helps to elucidate the effect of experimental variables on monomer conversion, Mn and regioselectivity of acylation of glycerol. Chemometric analysis shows that after 24 h the reaction proceeds regardless of the presence of the enzyme. Accordingly, the biocatalyst was removed after a first step of synthesis and the chain elongation continued at 80 °C. That allowed the removal of the biocatalyst and the preservation of its activity: pre-requites for efficient applicability at industrial scale. The experimental study, combined with docking-based computational analysis, provides rational guidelines for the optimization of the regioselective acylation of glycerol. The process is scaled up to 73.5 g of monomer. The novelty of the present study is the rigorous control of the reaction conditions and of the integrity of the immobilized biocatalyst, which serve to avoiding any interference of free enzyme or fines released in the reaction mixture. The quantitative analysis of the effect of experimental conditions and the overcoming of some major technical bottlenecks for the scalability of enzymatic polycondensation opens new scenarios for industrial exploitation.

Experimental and Computational Studies on Bio-Inspired Flavylium Salts as Sensitizers for Dye-Sensitized Solar Cells.

Six new bio-inspired flavylium salts were synthesized and investigated by a combined computational and experimental study for dye-sensitized solar cell applications. The compounds were characterized by FT-IR, UV-Vis, NMR spectroscopy, and LC-MS spectrometry techniques. The pH-dependent photochromic properties of the flavylium dyes were investigated through a UV-Vis spectroscopy study and revealed that they follow the same network of chemical reactions as anthocyanins upon pH changes. The structural and electronic properties of the dyes were investigated using density functional theory (DFT) and time-dependent density functional theory (TD-DFT). Geometry optimization calculation revealed that all dyes, regardless of the specie, flavylium cations or quinoidal bases, present a planar geometry. The photovoltaic performances of the dyes, in both flavylium and quinoidal base forms, were evaluated by the HOMO and LUMO energies and by calculating the light-harvesting efficiencies, the free energy change of electron injection, and the free energy change regeneration. The MO analysis showed that all dyes can inject electrons into the conduction band of the TiO2 upon excitation and that the redox couple can regenerate the oxidized dyes. The results obtained for the free energy change of electron injection suggest that the quinoidal bases should inject electrons into the semiconductor more efficiently than the flavylium cations. The values for the free energy change regeneration showed that the redox electrolyte can easily regenerate all dyes. Dipole moment analysis was also performed. DSSCs based on the dyes, in both flavylium and quinoidal base forms, were assembled, and their photovoltaic performances were evaluated by measuring the open-circuit voltage, the short circuit current density, the fill factor, and the energy conversion efficiency. Results obtained by both experimental and computational studies showed that the overall performances of the DSSCs with the quinoidal forms were better than those obtained with the flavylium cations dyes.

Azelaic Acid: A Bio-Based Building Block for Biodegradable Polymers.

Azelaic acid is a dicarboxylic acid containing nine C atoms, industrially obtained from oleic acid. Besides its important properties and pharmacological applications, as an individual compound, azelaic acid has proved to be a valuable bio-based monomer for the synthesis of biodegradable and sustainable polymers, plasticizers and lubricants. This review discusses the studies and the state of the art in the field of the production of azelaic acid from oleic acid, the chemical and enzymatic synthesis of bio-based oligo and polyester and their properties, including biodegradability and biocompostability.

Co-Entrapment of Sorafenib and Cisplatin Drugs and iRGD Tumour Homing Peptide by Poly[ε-caprolactone-co-(12-hydroxystearate)] Copolymer.

The drug-loaded nanocarriers have overcome various challenges compared with the pure chemotherapeutic drug, such as limited bioavailability, multiple drug resistance, poor patient compliance, and adverse drug reactions, offering advantages such as protection from degradation in the blood stream, better drug solubility, and improved drug stability. One promising group of controlled and targeted drug delivery systems is polymer-based nanoparticles that can sustain the release of the active agent by diffusion and their degradation. Sorafenib is the only drug that can prolong the life of patients suffering from hepatocellular carcinoma. Cisplatin remains one of the most widely used broad-spectrum anticancer drugs for the treatment of a variety of solid tumours. Nanoformulations can exert a synergistic effect by entrapping two drugs with different modes of action, such as sorafenib and cisplatin. In our study, polymeric nanoparticles were prepared with a good production yield by an improved double emulsion solvent evaporation method using the copolymer of 12-hydroxystearic acid with ε-caprolactone (12CL), a biocatalytically synthesised biocompatible and biodegradable carrier, for the co-entrapment of sorafenib and cisplatin in nanotherapeutics. A bovine serum albumin (BSA) model compound was used to increase the cisplatin incorporation; then, it was successfully substituted by a iRGD tumour penetrating peptide that might provide a targeting function of the nanoparticles.

Enzymatic synthesis of short-chain flavor esters from natural sources using tailored magnetic biocatalysts.

Immobilized lipases are excellent biocatalysts for the enzymatic synthesis of short- and medium-chain fatty esters used as food flavor compounds. Herein a new approach for a magnetic core-shell biocatalyst by immobilization of Candida antarctica B lipase is reported, coating single-core magnetic nanoparticles with an organic shell, preferably poly(benzofurane-co-arylacetic acid), followed by the covalent attachment of the enzyme and embedment of the primary biocatalyst in a silica layer. Although covalent and sol-gel immobilization were efficient on their own, their combination can ensure additional operational stability through multi-point linkages. Moreover, silanes holding glycidoxy groups, which can also form covalent linkages, have been successfully used as precursors for the silica coating layer. The structural, magnetic and morphological characteristics were assessed by TEM, SEM-EDX, X-ray photoelectron spectroscopy and vibrating sample magnetometry. The new biocatalysts demonstrated high catalytic efficiency in the solventless synthesis of isoamyl esters of natural carboxylic acids, also in multiple reaction cycles.

Biodegradable Oligoesters of ε-Caprolactone and 5-Hydroxymethyl-2-Furancarboxylic Acid Synthesized by Immobilized Lipases.

Following the latest developments, bio-based polyesters, obtained from renewable raw materials, mainly carbohydrates, can be competitive for the fossil-based equivalents in various industries. In particular, the furan containing monomers are valuable alternatives for the synthesis of various new biomaterials, applicable in food additive, pharmaceutical and medical field. The utilization of lipases as biocatalysts for the synthesis of such polymeric compounds can overcome the disadvantages of high temperatures and metal catalysts, used by the chemical route. In this work, the enzymatic synthesis of new copolymers of ε-caprolactone and 5-hydroxymethyl-2-furancarboxylic acid has been investigated, using commercially available immobilized lipases from Candida antarctica B. The reactions were carried out in solvent-less systems, at temperatures up to 80 °C. The structural analysis by MALDI TOF-MS, NMR, and FT-IR spectroscopy confirmed the formation of cyclic and linear oligoesters, with maximal polymerization degree of 24 and narrow molecular weight distribution (dispersity about 1.1). The operational stability of the biocatalyst was explored during several reuses, while thermal analysis (TG and DSC) indicated a lower thermal stability and higher melting point of the new products, compared to the poly(ε-caprolactone) homopolymer. The presence of the heterocyclic structure in the polymeric chain has promoted both the lipase-catalyzed degradation and the microbial degradation. Although, poly(ε-caprolactone) is a valuable biocompatible polymer with important therapeutic applications, some drawbacks such as low hydrophilicity, low melting point, and relatively slow biodegradability impeded its extensive utilization. In this regard the newly synthesized furan-based oligoesters could represent a "green" improvement route.

Biocatalytic Route for the Synthesis of Oligoesters of Hydroxy-Fatty acids and ϵ-Caprolactone.

Developments of past years placed the bio-based polyesters as competitive substitutes for fossil-based polymers. Moreover, enzymatic polymerization using lipase catalysts has become an important green alternative to chemical polymerization for the synthesis of polyesters with biomedical applications, as several drawbacks related to the presence of traces of metal catalysts, toxicity and higher temperatures could be avoided. Copolymerization of ϵ-caprolactone (CL) with four hydroxy-fatty acids (HFA) from renewable sources, 10-hydroxystearic acid, 12-hydroxystearic acid, ricinoleic acid, and 16-hydroxyhexadecanoic acid, was carried out using commercially available immobilized lipases from Candida antarctica B, Thermomyces lanuginosus, and Pseudomonas stutzeri, as well as a native lipase. MALDI-TOF-MS and 2D-NMR analysis confirmed the formation of linear/branched and cyclic oligomers with average molecular weight around 1200 and polymerization degree up to 15. The appropriate selection of the biocatalyst and reaction temperature allowed the tailoring of the non-cyclic/cyclic copolymer ratio and increase of the total copolymer content in the reaction product above 80%. The catalytic efficiency of the best performing biocatalyst (Lipozyme TL) is evaluated during four reaction cycles, showing excellent operational stability. The thermal stability of the reaction products is assessed based on TG and DSC analysis. This new synthetic route for biobased oligomers with novel functionalities and properties could have promising biomedical applications.

Biocatalytic synthesis of δ-gluconolactone and ε-caprolactone copolymers.

The biodegradability and biocompatibility properties of ε-caprolactone homopolymers place it as a valuable raw material, particularly for controlled drug delivery and tissue engineering applications. However, the usefulness of such materials is limited by their low hydrophilicity and slow biodegradation rate. In order to improve polycaprolactone properties and functionalities, copolymerization of ε-caprolactone with δ-gluconolactone was investigated. Since enzymatic reactions involving sugars are usually hindered by the low solubility of these compounds in common organic solvents, finding the best reaction medium was a major objective of this research. The optimal copolymerization conditions were set up by using different organic media (solvent and solvents mixtures), as well as solvent free systems that are able to dissolve (completely or partially) sugars, and are nontoxic for enzymes. Native and immobilized lipases by different immobilization techniques from Candida antarctica B and Thermomyces lanuginosus have been used as biocatalyst at 80°C. Although the main copolymer amount was synthesized in DMSO:t-BuOH (20:80) medium, the highest polymerization degrees, up to 16 for the copolymer product, were achieved in solventless conditions. The products, cyclic and linear polyesters, have been characterized by FT-IR and MALDI-TOF MS analysis. The reaction product analysis revealed the formation of cyclic products that could be the major impediment of further increase of the chain length.