Long chain capsaicin analogues synthetized by CALB-CLEAs show cytotoxicity on glioblastoma cell lines.

Glioblastoma is one of the most lethal tumors, displaying striking cellular heterogeneity and drug resistance. The prognosis of patients suffering from glioblastoma after 5 years is only 5%. In the present work, capsaicin analogues bearing modifications on the acyl chain with long-chain fatty acids showed promising anti-tumoral activity by its cytotoxicity on U-87 and U-138 glioblastoma multiforme cells. The capsaicin analogues were enzymatically synthetized with cross-linked enzyme aggregates of lipase B from Candida antarctica (CALB). The catalytic performance of recombinant CALB-CLEAs was compared to their immobilized form on a hydrophobic support. After 72 h of reaction, the synthesis of capsaicin analogues from linoleic acid, docosahexaenoic acid, and punicic acid achieved a maximum conversion of 69.7, 8.3 and 30.3% with CALB-CLEAs, respectively. Similar values were obtained with commercial CALB, with conversion yields of 58.3, 24.2 and 22% for capsaicin analogues from linoleic acid, DHA and punicic acid, respectively. Olvanil and dohevanil had a significant cytotoxic effect on both U-87 and U-138 glioblastoma cells. Irrespective of the immobilization form, CALB is an efficient biocatalyst for the synthesis of anti-tumoral capsaicin derivatives. KEY POINTS: • This is the first report concerning the enzymatic synthesis of capsaicin analogues from docosahexaenoic acid and punicic acid with CALB-CLEAs. • The viability U-87 and U-138 glioblastoma cells was significantly affected after incubation with olvanil and dohevanil. • Capsaicin analogues from fatty acids obtained by CALB-CLEAs are promising candidates for therapeutic use as cytotoxic agents in glioblastoma cancer cells.

Computational insights of the molecular recognition between volatile molecules and odorant binding proteins from the red palm weevil Rhynchophorus ferrugineus.

The red palm weevil Rhynchophorus ferrugineus (Coleoptera: Curculionidae) is one of the most harmful pests for palm trees, causing serious economic damage worldwide. The present work aims to model and study the 3D structures of highly expressed odorant binding proteins from R. ferrugineus (RferOBPs) and identify possible binding modes and ligand release mechanism by docking and molecular dynamics. Highly confident 3D structures of a total of 11 odorant binding proteins (OBPs) were obtained with AlphaFold2. All 3D RferOBPs modeled structures displayed six characteristic α-helices, except for RfeOBP7 and RfeOBP10, which had an extra terminal α-helix. Among the eleven modeled RferOBPs, RferOBP4 was highly expressed in the antennae and subsequently selected for further analyses. Molecular docking analyses demonstrated that ferruginol, α-pinene, DEET, and picaridin can favorably bind the RferOBP4 cavity with low affinity energies. Molecular dynamic simulations of RferOBP4 bound to ferruginol at different pH values showed that low pH environments dictate a structural change into an apo-state that modifies the number of tunnels where the ligand can coexist, further triggering ligand release by a pH-dependent mechanism. This is the first report concerning the modelling and study of ligand binding modes and release mechanism of R. ferrugineus OBPs.Communicated by Ramaswamy H. Sarma.

Thermoplastic Starch Biocomposite Films Reinforced with Nanocellulose from Agave tequilana Weber var. Azul Bagasse.

In this work, cellulose nanocrystals (CNCs), bleached cellulose nanofibers (bCNFs), and unbleached cellulose nanofibers (ubCNFs) isolated by acid hydrolysis from Agave tequilana Weber var. Azul bagasse, an agro-waste from the tequila industry, were used as reinforcements in a thermoplastic starch matrix to obtain environmentally friendly materials that can substitute contaminant polymers. A robust characterization of starting materials and biocomposites was carried out. Biocomposite mechanical, thermal, and antibacterial properties were evaluated, as well as color, crystallinity, morphology, rugosity, lateral texture, electrical conductivity, chemical identity, solubility, and water vapor permeability. Pulp fibers and nanocelluloses were analyzed via SEM, TEM, and AFM. The water vapor permeability (WVP) decreased by up to 20.69% with the presence of CNCs. The solubility decreases with the presence of CNFs and CNCs. The addition of CNCs and CNFs increased the tensile strength and Young's modulus and decreased the elongation at break. Biocomposites prepared with ubCNF showed the best tensile mechanical properties due to a better adhesion with the matrix. Images of bCNF-based biocomposites demonstrated that bCNFs are good reinforcing agents as the fibers were dispersed within the starch film and embedded within the matrix. Roughness increased with CNF content and decreased with CNC content. Films with CNCs did not show bacterial growth for Staphylococcus aureus and Escherichia coli. This study offers a new theoretical basis since it demonstrates that different proportions of bleached or unbleached nanofibers and nanocrystals can improve the properties of starch films.

Structural Behavior of Amphiphilic Triblock Copolymer P104/Water System.

A detailed study of the different structural transitions of the triblock copolymer PEO27-PPO61-PEO27 (P104) in water, in the dilute and semi-dilute regions, is addressed here as a function of temperature and P104 concentration (CP104) by mean of complimentary methods: viscosimetry, densimetry, dynamic light scattering, turbidimetry, polarized microscopy, and rheometry. The hydration profile was calculated through density and sound velocity measurements. It was possible to identify the regions where monomers exist, spherical micelle formation, elongated cylindrical micelles formation, clouding points, and liquid crystalline behavior. We report a partial phase diagram including information for P104 concentrations from 1 × 10-4 to 90 wt.% and temperatures from 20 to 75 °C that will be helpful for further interaction studies with hydrophobic molecules or active principles for drug delivery.

Characterization, comparative, and functional analysis of arylacetamide deacetylase from Gnathostomata organisms.

BACKGROUND: Arylacetamide deacetylase (AADAC) is a lipolytic enzyme involved in xenobiotic metabolism. The characterization in terms of activity and substrate preference has been limited to a few mammalian species. The potential role and catalytic activities of AADAC from other organisms are still poorly understood. Therefore, in this work, the physicochemical properties, proteomic analysis, and protein-protein interactions from Gnathostomata organisms were investigated. RESULTS: The analysis were performed with 142 orthologue sequences with ~ 48-100% identity with human AADAC. The catalytic motif HGG[A/G] tetrapeptide block was conserved through all AADAC orthologues. Four variations were found in the consensus pentapeptide GXSXG sequence (GDSAG, GESAG, GDSSG, and GSSSG), and a novel motif YXLXP was found. The prediction of N-glycosylation sites projected 4, 1, 6, and 4 different patterns for amphibians, birds, mammals, and reptiles, respectively. The transmembrane regions of AADAC orthologues were not conserved among groups, and variations in the number and orientation of the active site and C-terminal carboxyl were observed among the sequences studied. The protein-protein interaction of AADAC orthologues were related to cancer, lipid, and xenobiotic metabolism genes. CONCLUSION: The findings from this computational analysis offer new insight into one of the main enzymes involved in xenobiotic metabolism from mammals, reptiles, amphibians, and birds and its potential use in medical and veterinarian biotechnological approaches.

Mechanical and Physicochemical Properties of 3D-Printed Agave Fibers/Poly(lactic) Acid Biocomposites.

In order to provide a second economic life to agave fibers, an important waste material from the production of tequila, filaments based on polylactic acid (PLA) were filled with agave fibers (0, 3, 5, 10 wt%), and further utilized to produce biocomposites by fused deposition modeling (FDM)-based 3D printing at two raster angles (-45°/45° and 0°/90°). Differential scanning calorimetry, water uptake, density variation, morphology, and composting of the biocomposites were studied. The mechanical properties of the biocomposites (tensile, flexural, and Charpy impact properties) were determined following ASTM international norms. The addition of agave fibers to the filaments increased the crystallinity value from 23.7 to 44.1%. However, the fibers generated porous structures with a higher content of open cells and lower apparent densities than neat PLA pieces. The printing angle had a low significant effect on flexural and tensile properties, but directly affected the morphology of the printed biocomposites, positively influenced the impact strength, and slightly improved the absorption values for biocomposites printed at -45°/45°. Overall, increasing the concentrations of agave fibers had a detrimental effect on the mechanical properties of the biocomposites. The disintegration of the biocomposites under simulated composting conditions was slowed 1.6-fold with the addition of agave fibers, compared to neat PLA.

Optimization of the Obtaining of Cellulose Nanocrystals from Agave tequilana Weber Var. Azul Bagasse by Acid Hydrolysis.

A multilevel factorial design of 23 with 12 experiments was developed for the preparation of cellulose nanocrystals (CNC) from Agave tequilana Weber var. Azul bagasse, an agro-industrial waste from tequila production. The studied parameters were acid type (H2SO4 and HCl), acid concentration (60 and 65 wt% for H2SO4, 2 and 8N for HCl) temperature (40 and 60 °C for H2SO4, 50 and 90 °C for HCl), and hydrolysis time (40, 55 and 70 min for H2SO4; and 30, 115 and 200 min for HCl). The obtained CNC were physical and chemically characterized using dynamic light scattering (DLS), atomic force microscopy (AFM), Fourier-transform infrared spectroscopy (FT-IR), X-ray photoelectron spectroscopy (XPS), and X-ray diffraction (XDR) techniques. The maximum CNC yield was 90 and 96% for HCL and H2SO4, respectively, and the crystallinity values ranged from 88-91%. The size and morphology of A. tequilana CNC strongly depends on the acid type and hydrolysis time. The shortest CNC obtained with H2SO4 (65 wt%, 40 °C, and 70 min) had a length of 137 ± 68 nm, width 33 ± 7 nm, and height 9.1 nm, whereas the shortest CNC obtained with HCl (2 N, 50 °C and 30 min) had a length of 216 ± 73 nm, width 69 ± 17 nm, and height 8.9 nm. In general, the obtained CNC had an ellipsoidal shape, whereas CNC prepared from H2SO4 were shorter and thinner than those obtained with HCl. The total sulfate group content of CNC obtained with H2SO4 increased with time, temperature, and acid concentration, exhibiting an exponential behavior of CSG=aebt.

Cross-linked enzyme aggregates of recombinant Candida antarctica lipase B for the efficient synthesis of olvanil, a nonpungent capsaicin analogue.

Despite the proven therapeutic role of capsaicin in human health, its usage is still hampered by its high pungency. In this sense, nonpungent capsaicin analogues as olvanil are a feasible alternative to the unpleasant sensations produced by capsaicin while maintaining a similar pharmacological profile. Olvanil can be obtained by a lipase-catalyzed chemoenzymatic process. In the present work, recombinant Candida antarctica lipase B (CALB) was expressed in Pichia pastoris and subsequently immobilized by cross-linked enzyme aggregate (CLEA) methodology for the synthesis of olvanil. The CALB-CLEAs were obtained directly from the fermentation broth of P. pastoris without any purification step in order to assess the role of the contaminant proteins of the crude extract as co-feeders. The CALB-CLEAs were also bioimprinted to enhance the catalytic performance in olvanil synthesis. When CALB was precipitated with isopropanol, the obtained CALB-CLEAs exhibited the highest activity in the synthesis of olvanil, regardless of the glutaraldehyde concentration. The maximum product synthesis was found at 72 hr obtaining 6.8 g L-1 of olvanil with a reaction yield of 16%. When CALB was bioimprinted with olvanil, the synthesis was enhanced 1.3 times, reaching 10.7 g L-1 of olvanil at 72 hr of reaction with a reaction yield of 25%. Scanning electron microscopy images indicated different morphologies of the CLEAs depending on the precipitating agent and the template used for bioimprinting. Recombinant CALB-CLEAs obtained directly from the fermentation broth are a suitable alternative to commercial enzymatic preparations for the synthesis of olvanil in organic medium.