Modification and characterization of clinoptilolite for the co-immobilization of formate dehydrogenase and glycerol dehydrogenase enzymes

In the last decades there is a rising concern for the increasing concentration of carbon dioxide, considered the major responsible of Global Warming. A solution to this critical issue is the catalytic conversion of CO2 into high value-added products. Among the different strategies that could be applied, the enzymatic process of CO2 reduction to methanol, employing a sequence of three enzyme-catalyzed reactions, seems to be very promising. The simultaneous employment of formate dehydrogenase and glycerol dehydrogenase allows to reduce CO2 to formic acid, the first of the sequential reactions, and at the same time regenerate the nicotinamide cofactor, that is very expensive. To reuse enzymes, with a consequential reduction of cost, and increased their stability, they can be immobilized on a proper support. In this context, porous materials, such as zeolites, present appropriate features to be suitable for enzymes immobilization. In particular, they are well suited for the covalent immobilization technique due to the fact that they can be functionalized with different functional groups. Natural zeolites, like Clinoptilolite have the advantage to be low-cost materials largely diffused in different part of the world. Clinoptilolite was subjected to dealumination-desilication treatments to modify the zeolite’s morphology, increasing its specific surface area. According to the literature, the dealumination procedure was done with sequential acid attacks using HCl solutions. Instead, for the subsequent desilication process NaOH solution is required. The effects of desilication-dealumination treatments were investigated through complementary techniques such as N2 physisorption at -196 °C, XRD and SEM. The analysis revealed that the Clinoptilolite specific surface area increased by 400% following the dealumination-desilication procedure; at the same time the XRD analysis shows that the processed Clinoptilolite has the same main peaks of the unmodified one. Finally, the retained activity and the stability of the immobilized enzymes were evaluated, the results show that these aspects were enhanced by the modification through acid-alkaline attacks of the Clinoptilolite

Simultaneous CO2 reduction and NADH regeneration using formate and glycerol dehydrogenase enzymes co-immobilized on modified natural zeolite

In this work, the co-immobilization of formate dehydrogenase (FDH) and glycerol dehydrogenase (GlyDH) enzymes is proposed to reduce CO(2) into formic acid, an important chemical intermediate. The reduction of carbon dioxide is carried out by FDH to obtain formic acid, simultaneously, the GlyDH regenerated the nicotinamide cofactor in the reduced form (NADH) by the oxidation of glycerol into dihydroxyacetone. Natural zeolite was selected as immobilization support given its good properties and low cost. The natural zeolite was modified with subsequent acid-alkaline attacks to obtain a mesostructurization of the clinoptilolite. The two enzymes were co-immobilized on clinoptilolite, previously hetero-functionalized with amino and glyoxyl groups. The distribution of the enzymes was confirmed by fluorescence microscopy analysis. Furthermore, a great increase in the retained activity for the formate dehydrogenase enzyme was noted, passing from 18% to 89%, when the mesostructured clinoptilolite was used as support. The immobilization yield of formate dehydrogenase and glycerol dehydrogenase is around 100% with all the supports studied. The promising results suggest a possible development of this procedure in enzyme immobilization and biocatalysis. The biocatalysts were characterized to find the optimal pH and temperature. Furthermore, a thermal stability test at 50 °C was carried out on both enzymes, in free and immobilized forms. Finally, it was shown that the biocatalyst is effective in reducing CO(2), both by using the cofactor in the reduced form (NADH) or the oxidized form (NAD(+)), obtaining NADH through the regeneration with glycerol in this latter case

Preparation of a Mesoporous Biosensor for Human Lactate Dehydrogenase for Potential Anticancer Inhibitor Screening

Cancer is the second leading cause of death worldwide, with a dramatic impact due to the acquired resistance of cancers to used chemotherapeutic drugs and treatments. The enzyme lactate dehydrogenase (LDH-A) is responsible for cancer cell proliferation. Recently the development of selective LDH-A inhibitors as drugs for cancer treatment has been reported to be an efficient strategy aiming to decrease cancer cell proliferation and increase the sensitivity to traditional chemotherapeutics. This study aims to obtain a stable and active biocatalyst that can be utilized for such drug screening purposes. It is conceived by adopting human LDH-A enzyme (hLDH-A) and investigating different immobilization techniques on porous supports to achieve a stable and reproducible biosensor for anticancer drugs. The hLDH-A enzyme is covalently immobilized on mesoporous silica (MCM-41) functionalized with amino and aldehyde groups following two different methods. The mesoporous support is characterized by complementary techniques to evaluate the surface chemistry and the porous structure. Fluorescence microscopy analysis confirms the presence of the enzyme on the support surface. The tested immobilizations achieve yields of ≥80%, and the best retained activity of the enzyme is as high as 24.2%. The optimal pH and temperature of the best immobilized hLDH-A are pH 5 and 45 °C for the reduction of pyruvate into lactate, while those for the free enzyme are pH 8 and 45 °C. The stability test carried out at 45 °C on the immobilized enzyme shows a residual activity close to 40% for an extended time. The inhibition caused by NHI-2 is similar for free and immobilized hLDH-A, 48% and 47%, respectively. These findings are significant for those interested in immobilizing enzymes through covalent attachment on inorganic porous supports and pave the way to develop stable and active biocatalyst-based sensors for drug screenings that are useful to propose drug-based cancer treatments

Copper-manganese oxide catalysts prepared by solution combustion synthesis for total oxidation of VOCs

A set of Cu-Mn oxides was prepared through the simple and effective solution combustion synthesis method by varying the relative amount of copper and manganese. The physico-chemical properties of the samples were investigated through complementary techniques such as N2 physisorption at − 196 °C, XRD, HR-TEM, Raman spectroscopy, temperature programmed analyses (H2-TPR, O2-TPD, and NH3-TPD), and XPS. The prepared catalysts were tested for the total oxidation of volatile organic compounds (ethylene, propylene, and toluene). The best performances, in terms of total VOC oxidation, were achieved with a copper content ranging from 15 at% to 45 at%. The catalytic test outcomes demonstrate the beneficial effect of acidic sites, oxygen mobility, and redox ability. In particular, ethylene oxidation is mainly favored by oxygen vacancies and redox properties, while propylene and toluene oxidation is mostly enhanced by acidic sites. All the catalysts prepared can totally oxidize the examined pollutants examined below 310 °C. Moreover, the binary oxides exhibit good catalytic stability over a time-on-stream of 7 h and low water vapor inhibition (5 vol% H2O in the gas stream)