Evelin A. Manoel

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.

Nanomaterials for biocatalyst immobilization – state of the art and future trends

Nanotechnology is an area that has been growing over the years, being possible nowadays to find numerous materials constructed at nanoscale. In addition, many applications have been attributed to these “new” materials. In this review is presented a brief overview of nanoparticles used for the immobilization of enzymes. Considering the extensive universe of immobilization in nanoparticles, some were chosen to be exposed here, such as chitosan, graphene, silica, polymers, magnetic, nanoflowers, among others. Advantages, disadvantages and limitations of nanoimmobilization also be discussed. Some applications of nanoimmobilized enzymes are presented, like as biodiesel, flavor synthesis ester and biosensors. The purpose of this paper is to provide an overview of what is being studied in relation to nanoparticles for enzymes immobilization, and some discussions about them, aimed at assisting researchers in future studies and reviews.

Design of a core–shell support to improve lipase features by immobilization

Different core–shell polymeric supports, exhibiting different morphologies and composition, were produced through simultaneous suspension and emulsion polymerization, using styrene (S) and divinylbenzene (DVB) as co-monomers. Supports composed of polystyrene in both the core and the shell (PS/PS) and the new poly(styrene-co-divinylbenzene) support (PS-co-DVB/PS-co-DVB) were used for the immobilization of three different lipases (from Rhizomucor miehie (RML), from Themomyces lanuginosus (TLL) and the form B from Candida antarctica, (CALB)) and of the phospholipase Lecitase Ultra (LU). The features of the new biocatalysts were evaluated and compared to the properties of commercial biocatalysts (Novozym 435 (CALB), Lipozyme RM IM and Lipozyme TL IM) and biocatalysts prepared by enzyme immobilization onto commercial octyl-agarose, a support reported as very suitable for lipase immobilization. It was shown that protein loading and stability of the biocatalysts prepared with the core–shell supports were higher than the ones obtained with commercial octyl-agarose or the commercial lipase preparations. Besides, it was shown that the biocatalysts prepared with the core–shell supports also presented higher activities than commercial biocatalysts when employing different substrates, encouraging the use of the produced core–shell supports for immobilization of lipases and the development of new applications.

Evaluation of the performance of differently immobilized recombinant lipase B from Candida antarctica preparations for the synthesis of pharmacological derivatives in organic media

This paper shows the production of lipase B from Candida antarctica (LIPB) after cloning the gene that encoded it in Pichia pastoris using PGK as a constitutive promoter. The production of the lipase is lower using this strategy but it avoids the use of inducers like methanol. The performance of this enzyme was compared with that of the commercial enzyme (CALB) after immobilization on different supports in different reactions. As supports, we used Accurel 1000, and three core–shell supports (poly(methyl methacrylate) on the core and on the shell – PMMA/PMMA; poly(methyl methacrylate-co-divinylbenzene) on the core and on the shell – PMMA-co-DVB/PMMA-co-DVB; and poly(styrene-co-divinylbenzene) on the core and on the shell – PS-co-DVB/PS-co-DVB). The popular Novozym 435 was also utilized to assess the features of the new biocatalysts. All these supports adsorbed lipases via interfacial activation of the open form of the lipase on the hydrophobic surface of the supports. The studied reactions were esterification of oleic acid and ethanol in a solvent-free medium, resolution of (±)-1,3,5-O-benzyl-myo-inositol via acylation using vinyl acetate in hexane and resolution of (±)-1,2-O-isopropylidene-3,6-di-O-benzyl-myo-inositol via acylation using vinyl acetate (solvent free system). The results varied depending on the employed supports and on the studied reactions, but some general trends may be observed, pointing to better behavior of LIPB compared to CALB. The use of 4 different supports gave more strength to these differences, as it did not depend on a specific difference between a single support/enzyme pair, but it is more general. Thus, LIPB seems to have some advantages compared to the commercial enzyme on all the reactions assayed in this paper. PS-co-DVB/PS-co-DVB-LIPB is in general the most active preparation (even 50% higher activity was observed). Further investigations are in development to determine the structural reasons for these differences.

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.

A New Tree Species of Strychnos Sect. Longiflorae (Loganiaceae)

Abstract A new species Strychnos dantaensis was discovered in lowland forest of Poço das Antas Biological Reserve, and is the unique tree species of the genus Strychnos in Rio de Janeiro State. The new species is related to S. trinervis (Vell.) Mart., but differs by its arborescent growth form, absence of tendrils on branches, ovate calyx lobes, and oblong anthers. Strychnos dantaensis should be treated as endangered under IUCN red list criteria due to its restricted geographical distribution in the Atlantic Rain Forest, an ecosystem in continuous decline.

Strychnos jacarepiensis, a new species of Loganiaceae from Brazil

SummaryStrychnos jacarepiensis E. A. Manoel & E. F. Guim. is described and illustrated as a new species from Rio de Janeiro state, Brazil. It differs from S. gardneri by the terminal inflorescence and shorter corolla tube, and from S. brasiliensis by the connate and amplexicaul interpetiolar stipule, velutinous calyx lobes, villous filaments and discoid seeds.

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

ABSTRACT 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. GRAPHICAL ABSTRACT Extracts containing PAH were obtained from soils cultivated with Medicago sativa L. (cv. Crioula) in the presence and absence of 150, 300, and 450 ppm total of PAH (phenanthrene, anthracene, and pyrene). After ultrasonication steps and purification of PAH by silica adsorption chromatography, samples containing PAH were quantified by high-performance liquid chromatography in reverse phase. At the end of this study, it was determined that the cv. Crioula showed a satisfactory phytoremediation potential. It presented the reduction of the total of PAH at around 85% in 20 days.

Loganiaceae no estado do Rio de Janeiro: chave para os gêneros e taxonomia de Spigelia

Resumo Loganiaceae engloba cerca de 350 especies em 13 generos com distribuicao pantropical. Spigelia e um dos maiores generos da familia, com cerca de 90 especies e distribuicao neotropical. No Brasil ocorrem 54 especies, o que torna o pais o centro de diversidade do genero. A morfologia dos frutos de Spigelia ja foi anteriormente abordada, entretanto, pouca enfase foi dada a morfologia da porcao basal diferenciada do fruto, o carpoatlas. O presente trabalho apresenta a chave para os generos da flora fluminense, e o tratamento taxonomico de Spigelia no estado do Rio de Janeiro, dando enfase ao carpoatlas. Nove especies sao aceitas para o estado do Rio de Janeiro, sendo S. reflexicalyx reestabelecida no presente trabalho. Estas, sao descritas, ilustradas e sao apresentados comentarios, mapa de distribuicao e chaves de identificacao. Tanto os carpoatlas quanto os forames e os estigmas, mostraram-se com formas e medidas diferenciadas entre as especies.

CHAPTER 9:Lipases in Enantioselective Syntheses: Evolution of Technology and Recent Applications

Lipases are the most widely employed biocatalysts in enantioselective syntheses. In this review, we discuss, with a historical perspective, the advances in the technology behind lipase catalysis. The different classes (resolutions, deracemizations, desymmetrizations) of lipase-catalysed processes towards enantioselective synthesis are briefly discussed. The relevance of such biocatalysts and the practicality of their use in this context are discussed with regards to features such as medium engineering, immobilization, protein engineering, reactor technology and new media (e.g., ionic liquids), whose advances we discuss herein.