Microglial activation can release free radicals and various pro-inflammatory cytokines, which implicates the progress of a neurodegenerative disease. Therefore suppression of microglial activation can be an appropriate strategy for combating neurodegenerative diseases. Betanin is a red food dye that acts as free radical scavenger and can be a promising candidate for this purpose. In this study, purification of betanin from red beetroots was carried out by normal phase colum chromatography, yielding 500 mg of betanin from 100 g of red beetroot. The purified betanin was evaluated by TLC, UV-visible, HPLC, ESI-MASS, FT-IR spectroscopy. Investigation on the inhibitory effect of betanin on activated microglia was performed using primary microglial culture. The results showed that betanin significantly inhibited lipopolysaccharide induced microglial function including the production of nitric oxide free radicals, reactive oxygen species, tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and interleukin-1 beta (IL-1β). Moreover, betanin modulated mitochondrial membrane potential, lysosomal membrane permeabilization and adenosine triphosphate. We further investigated the interaction of betanin with TNF-α, IL-6 and Nitric oxide synthase (iNOS or NOS2) using in silico molecular docking analysis. The docking results demonstrated that betanin have significant negative binding energy against active sites of TNF-α, IL-6 and iNOS.

Aldehyde-functionalized silica and silica nanoparticles (SBA-15) were prepared as a matrix system for enzyme immobilization. Immobilization of Rhizomucor miehei lipase (RML) on these supports was performed via a multicomponent reaction under extremely mild conditions (25 °C, pH 7). Investigation on the mechanism of this reaction confirmed the Ugi four-component immobilization approach. The loading capacity of the supports and specific activity of the immobilized derivatives were interestingly improved. The results revealed very rapid immobilization of 10 and 60 mg of RML on 1 g of aldehyde-functionalized silica and SBA-15 after 10 and 30 minutes, respectively. Leaching experiments were performed by incubation of the immobilized derivatives in 1 M NaCl solution. The lack of the free lipase in the solution confirmed the covalent nature of the linkage. The thermal stability and co-solvent stability of the derivatives in the presence of three polar organic solvents (1-propanol, 2-propanol and dioxane) were greatly improved compared to the soluble enzyme. Both the derivatives were also used to catalyze the transesterification of colza oil with methanol to produce fatty acid methyl esters (FAMEs). In the case of RML immobilized on SBA-15 (SBA-RML), the presence of 40% of tert-butanol (v/v) as solvent in the reaction medium largely improved the conversion yield.

This paper presents a novel strategy for the simple immobilization of biomolecules on epoxy-functionalized supports via a one-pot three component reaction. Lipase B from Candida antarctica (CALB) and Rhizomucor miehei lipase (RML) as model enzymes were immobilized on epoxy-functionalized silica and mesoporous silica nanoparticles (SBA-15). Investigation of the mechanism of this reaction confirmed the participation of three functional groups including carboxylic acid groups from the enzyme surface, epoxy groups from the support and isocyanide from the reaction medium. The results revealed very rapid immobilization of 10 and 40 mg of RML on 1 gr of the supports shortly after 30 minutes of incubation. The loading capacity of the supports was also dramatically improved with the maximum loading of 158 mg of CALB and 77 mg of RML on SBA-15 and silica, respectively. Silica-epoxy-RML showed an enantiomeric excess (ee) of 92% and an E-value of 29.9 in the enantioselective esterification of (R,S)-ibuprofen.