Immobilizing white-rot fungi laccase: Toward bio-derived supports as a circular economy approach in organochlorine removal.

Despite their high persistence in the environment, organochlorines (OC) are widely used in the pharmaceutical industry, in plastics, and in the manufacture of pesticides, among other applications. These compounds and the byproducts of their decomposition deserve attention and efficient proposals for their treatment. Among sustainable alternatives, the use of ligninolytic enzymes (LEs) from fungi stands out, as these molecules can catalyze the transformation of a wide range of pollutants. Among LEs, laccases (Lac) are known for their efficiency as biocatalysts in the conversion of organic pollutants. Their application in biotechnological processes is possible, but the enzymes are often unstable and difficult to recover after use, driving up costs. Immobilization of enzymes on a matrix (support or solid carrier) allows recovery and stabilization of this catalytic capacity. Agricultural residual biomass is a passive environmental asset. Although underestimated and still treated as an undesirable component, residual biomass can be used as a low-cost adsorbent and as a support for the immobilization of enzymes. In this review, the adsorption capacity and immobilization of fungal Lac on supports made from residual biomass, including compounds such as biochar, for the removal of OC compounds are analyzed and compared with the use of synthetic supports. A qualitative and quantitative comparison of the reported results was made. In this context, the use of peanut shells is highlighted in view of the increasing peanut production worldwide. The linkage of methods with circular economy approaches that can be applied in practice is discussed.

Elucidating solvent effects on lipase-catalyzed peroxyacid synthesis through activity-based kinetics and molecular dynamics.

Peroxyacid synthesis is the first step in Prilezhaev epoxidation, which is an industrial method to form epoxides. Motivated by the development of a kinetic model as a tool for solvent selection, the effect of solvent type and acid chain length on the lipase-catalyzed peroxyacid synthesis was studied. A thermodynamic activity-based ping-pong kinetic expression was successfully applied to predict the effect of the reagent loadings in hexane. The activity-based reaction quotients provided a prediction of solvent-independent equilibrium constants. However, this strategy did not achieve satisfactory estimations of initial rates in solvents of higher polarity. The lack of compliance with some assumptions of this methodology could be confirmed through molecular dynamics calculations i.e. independent solvation energies and lack of solvent interaction with the active site. A novel approach is proposed combining the activity-based kinetic expression and the free binding energy of the solvent with the active site to predict kinetics upon solvent change. Di-isopropyl ether generated a strong interaction with the enzyme's active site, which was detrimental to kinetics. On the other hand, toluene or limonene gave moderate interaction with the active site rendering improved catalytic yield compared with less polar solvents, a finding sharpened when peroctanoic acid was produced.

An alternative approach for quantification of glyceraldehyde and dihydroxyacetone as trimethylsilyl derivatives by GC-FID.

A method for quantification of glyceraldehyde (GA), dihydroxyacetone (DHA) and glycerol (GLY) by gas chromatography coupled to a flame ionization detector (GC-FID) involving one-step derivatization into trimethylsilyl ethers is presented. In pyridine, DHA and GA showed predominant peaks assigned to dimeric structures and smaller peaks corresponding to the monomers. The later were identified by GC-MS as their completely derivatized molecules and were useful for construction of calibration curves with high linear correlation. On the other hand, DHA dimers were completely dissociated in water but GA dimers remained whereas with both, intermediates peaks arose which were associated to hydrated trymethyil silyl species. A calibration approach involving the sum of areas of most relevant peaks associated to aqueous solutions of GA and DHA was developed. Replicates measurements of a problem solution were in accordance with the results obtained by a well stablished HPLC technique. The coefficient of variation was below 5% for GLY and below 12% for GA and DHA. Compared with the HPLC method, the new GC-FID method presented a similar limit of quantification in the case of GA whereas for GLY and DHA a one-order-of-magnitude increase of sensitivity was achieved. TMS derivatives of GA and DHA without prior oximation enable a useful technique to study the equilibrium of the different tautomeric forms in solution.

Dynamic response of the expression of hxt1, hxt5 and hxt7 transport proteins in Saccharomyces cerevisiae to perturbations in the extracellular glucose concentration.

Glucose transport in Saccharomyces cerevisiae relies on a multi-factorial uptake system. The modulation of its efficiency depends on the differential expression of various sets of hexose transport-related proteins whose glucose affinity differs considerably. The expression of three different glucose transport proteins (HXT1, HXT5 and HXT6/7 with low-, intermediate- and high-affinity, respectively) was monitored as a result of modified extracellular glucose concentrations. Cultivation at glucose-limited (continuous) conditions was instantly replaced by a batch (and thus, non-limited) mode. Further, to mimic concentration gradients in large-scale production bioreactors, multiple and rapid transient glucose pulses were applied to chemostat cultivation. Antibodies against the HXT-proteins were used to monitor the proteins' expression levels prior to and after perturbing the external glucose concentrations. HXT5 and HXT6/7 were either expressed during the starvation-like steady-state phases in the chemostat cultivations, whereas HXT1 could not be detected at all. HXT1, however, is subsequently expressed during the excess of glucose in the batch mode, while the HXT5 and HXT6/7 transporters were at least found to decline. These findings coincide well with the transporters' affinity profiles. As a result of repeated and rapid transient glucose pulses during continuous fermentation, especially HXT6/7 pointed out to alter the protein expression pattern.