Adsorption of lipases on porous silica-based materials for esterification in a solvent-free system.

This study deals with lipase immobilization on micro- and mesoporous silica-based materials. The effects of the type of support (silica MCM-41, zeolite HZSM-5 (SAR 25), zeolite HZSM-5 (SAR 280), and the silica-aluminas Siral 10, Siral 20, and Siral 40) were investigated on the immobilization of lipase B from Candida antarctica (CALB) and lipase from Rhizomucor miehei (RML). The supports that allowed the highest immobilization efficiencies for the CALB were Siral 40 (91.4%), HZSM-5 (SAR 280) (90.6%), and MCM-41 (89.4%). Siral 20 allowed the highest immobilization efficiency for RML (97.6%), followed by HZSM-5 (SAR 25) (77.1%) and HZSM-5 (SAR 280) (62.7%). The effect of protein concentration on lipase immobilization was investigated, and the results adjusted well on the Langmuir isotherm model (R2 > 0.9). The maximum protein adsorption capacity of the support determined by the Langmuir model was equal to 10.64 and 20.97 mgprotein gsupport-1 for CALB and RML, respectively. The effects of pH (pH 7.0 and pH 11.0) and phosphate buffer solution concentration (5 and 100 mmol L-1) were also investigated on lipase immobilization. The immobilization efficiency for both lipases was similar for the different pH values. The use of 100 mmol L-1 phosphate buffer decreased the lipase immobilization efficiency. The biocatalysts (CALB-Siral 40 and RML-Siral 20) were tested in the ethyl oleate synthesis. The conversion of 61.7% was obtained at 60 °C in the reaction catalyzed by CALB-Siral 40. Both heterogeneous biocatalysts showed increased thermal stability compared with their free form. Finally, the reuse of the biocatalysts was studied. CALB-Siral 40 and RML-Siral 20 maintained about 30% of the initial conversion after 3 batches of ethyl oleate synthesis. Silica-aluminas (Siral 20 and 40) proved to be a support that allowed a high efficiency of immobilization of lipases and activity for esterification reaction.

The influence of polycyclic aromatic hydrocarbons in protein profile of Medicago sativa L.

Medicago sativa L. (alfalfa) are studied as potential phytoremediation agents of priority pollutants like polycyclic aromatic hydrocarbons (PAH). However, elucidation of the biochemical mechanisms involved in phytoremediation is a topic to be explored with knowledge gaps. This study aims to identify and classify proteins expressed in the aerial parts of laboratory-cultivated alfalfa in the presence and absence of pyrene, anthracene, and phenanthrene. Soil samples were amended with 100 mg.kg-1 of each PAH (total concentration of 300 ppm) and cultivated with alfalfa plants for 20 days. After this, aerial parts of cultivated plants from each condition were collected for qualitative proteomic analysis (ESI-Q/TOF). The results showed a significant increase (Student's t-test p < 0.05) of 41.7% in the concentration of proteins from plants grown in PAH-amended substrates, changes in the protein profile, with intense protein bands observed at 40-55, 34, 28, and 15 kDa when compared to the control. A total of 504 proteins were identified and classified into 12 functional categories, highlighting the identification of 11 phytoremediation-related proteins candidates in plants grown in the presence of PAH, with biological functions related to diverse metabolisms involved in the xenobiotics biodegradation (included PAH), glutathione and response to stress.

Phytoremediation of polycyclic aromatic hydrocarbons (PAH) by cv. Crioula: A Brazilian alfalfa cultivar.

This work aimed to evaluate the phytoremediation capacity of the alfalfa cultivar Crioula in soils contaminated with polycyclic aromatic hydrocarbons (PAHs), primary pollutants with mutagenic and carcinogenic potential. Alfalfa was grown from seed for 40 days on soil amended with anthracene, pyrene, and phenanthrene. Soil and plant tissue was collected for biometric assay, dry mass analysis, and PAH analysis by liquid chromatography. Increased total PAH concentration was associated with decreases in plant biomass, height, and internode length. The Crioula cultivar had a satisfactory phytoremediation effect, reducing total PAH concentration (300 ppm) in the experimental soil by 85% in 20 days, and by more than 95% in 40 days. The PAH showed a tendency to be removed in the temporal order: phenanthrene before pyrene before anthracene, and the removal ratio was influenced by the initial soil concentration of each PAH.

Detection of polycyclic aromatic hydrocarbons (PAHs) in Medicago sativa L. by fluorescence microscopy.

Green technologies, such as phytoremediation, are effective for removing organic pollutants derived from oil and oil products, including polycyclic aromatic hydrocarbons (PAHs). Given the increasing popularity of these sustainable remediation techniques, methods based on fluorescence microscopy and multiphoton microscopy for the environmental monitoring of such pollutants have emerged in recent decades as effective tools for phytoremediation studies aimed at understanding the fate of these contaminants in plants. However, little is known about the cellular and molecular mechanisms involved in PAH uptake, responses and degradation by plants. Thus, the present study aimed to detect the location of pyrene, anthracene and phenanthrene using fluorescence microscopy techniques in shoots and roots of Medicago sativa L. (alfalfa) plants grown in artificially contaminated soil (150ppm PAHs) for 40days. Leaflet and root samples were then collected and observed under a fluorescence microscope to detect the presence of PAHs in various tissues. One important finding of the present study was intense fluorescence in the glandular secreting trichomes (GSTs) of plants grown in contaminated soil. These trichomes, with a previously unknown function, may be sites of PAH conjugation and degradation.

Immobilization of lipases on hydrophobic supports involves the open form of the enzyme.

The lipases from Thermomyces lanuginosus and Pseudomonas cepacia have been immobilized on octyl and cyanogen bromide (CNBr) agarose beads. The immobilization on octyl-agarose is slowed with increasing ionic strength, while the immobilization on CNBr is not significantly affected by the ionic strength. The inhibition of the immobilized preparations with diethyl p-nitrophenylphosphate (D-pNPP) was analyzed. The inhibition was more rapid using octyl-lipase preparations than using covalent preparations, and the covalent preparations were much more sensitive to the reaction medium. The addition of detergent increased the inhibition rate of the covalent preparation while an increase on the ionic strength produced a slowdown of the inhibition rate by D-pNPP for both lipases. The effect of the medium on the activity versus fully soluble substrate (methyl mandelate) was in the same direction. The octyl preparations presented a slight decrease in activity when comparing the results using different concentrations of sodium phosphate buffer (between 0.025 and 1M), while the CNBr preparations suffered drastic drops in its activity at high ionic strength. The results confirm that the lipases immobilized on octyl agarose presented their open form stabilized while the covalent preparation maintains a closing/opening equilibrium that may be modulated by altering the medium.