Supercritical carbon dioxide extraction of hydrocarbons from the microalgaBotryococcus braunii

Samples of the microalgaBotryococcus braunii were submitted to supercritical fluid extraction with carbon dioxide at 40 °C and pressures of 12.5, 20.0 and 30.0 MPa. The extraction yield and the fraction of the hydrocarbons in the extracts both increased with pressure and at 30 MPa these compounds were obtained rapidly. This behaviour is associated with the localization of the hydrocarbons outside the cell wall. In the extracts, which are fluid, golden and limpid, chlorophyll and phospholipids were not detected.Author for correspondence     

An ultrafiltration membrane bioreactor for the lipolysis of olive oil in reversed micellar media

The enzymatic hydrolysis of olive oil using Chromobacterium viscosum lipase B encapsulated in reversed micelles of dioctyl sodium sulfosuccinate (AOT) in isooctane was investigated in an ultrafiltration ceramic membrane reactor of tubular type, operating in a batch mode. Water concentration was found to be a critical parameter in the enzyme kinetics and hydrolysis yield of the reaction. The size of micelles, recirculation rate, and substrate concentration were found to be the major factors affecting the separation process. A correlation that enables the prediction of final conversion degrees in this bioreactor from the initial reaction conditions was established. (c) 1993 Wiley & Sons, Inc.     

Comparison of glucose fermentation by suspended and gel-entrapped yeast cells: An in vivo nuclear magnetic resonance study

Phosphorus-31 nuclear magnetic resonance ((31)P NMR) was used to compare the anaerobic metabolism of glucose by suspended and gel-entrapped Saccharomyces bayanus cells. The fermentation of glucose was carried out in a reaction system with continuous circulation through the NMR sample tube. The intracellular pH and the levels of some phosphorylated compounds were the levels of some phosphorylated compounds were noninvasively monitored by (31)P NMR while glucose, fermentation products, and biomass were determined by analytic techniques comparisons showed that no significant differences are observed in the relative concentrations in the spectra, but distinct profiles for the variation of both intracellular and extracellular pH are found. The internal pH of immobilized cells is maintained at a constant value throughout the fermentation as opposed to freely suspended cells for which a steady decrease in the internal pH occurs. A faster and stronger acidification is also observed in the external medium of the assays with suspended cells. Furthermore, higher yields for ethanol and biomass production and lower yields of fermentation by-products are obtained with immobilized cells. It is concluded that the higher intracellular pH achieved in the presence of the gel matrix had a regulatory effect on the metabolism which favored the ethanol production pathway. (c) 1993 John Wiley & Sons, Inc.     

Fusarium solani pisi recombinant cutinase partitioning in peg/potassium phosphate aqueous two-phase systems

Summary An aqueous two-phase system of polyethylene glycol (PEG) and potassium phosphate was developed for extraction of a cutinase from cell debris of a recombinant Escherichia coli strain. Basic studies to identify the primary factors which affect cutinase partition, namely the influence of polymer molecular weight, polymer concentration and pH were carried out using a purified preparation of the cutinase. The enzyme partition coefficient was enhanced with decreasing PEG molecular weight, increasing tie-line length and pH.     

Liquid-liquid extraction of a recombinant cutinase from fermentation media with AOT reversed micelles

This work reports the extraction and back-extraction of an intracellular recombinant cutinase from complex biological media using AOT reversed micelles in isooctane. Cutinase was recovered from different complex media namely, fermentation broths and supernatants after cell disruption by osmotic shock and sonication. The application of the AOT reversed micellar system to the extraction of cutinase allowed activity yields and purification factors ranging from about 5% to 50% and 1.2 to 10.2, respectively, depending on the biological medium.Maria das Graças Carneiro da Cunha, from ITEP-Instituto Tecnológico do Estado de Pernambuco, acknowledges a Ph.D fellowship from Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) and Centro de Pesquisa Aggeu Magalhães, Recife — PE — Brasil. E. P. Melo thanks Junta Nacional de Investigação Científica, Lisboa, Portugal, for providing a Ph.D. fellowship. The scientific support given by Prof. Sílvia M. B. Costa for the spectroscopic data and further discussions are particularly acknowledged.This work was partly financed by the BRIDGE and BIOTECHNOLOGY Programmes (Contracts BIOT-CT91-0274(DTEE) and BIOT 2 CT-943016).     

Cryptic coloration and choice of escape microhabitats by grasshoppers (Orthoptera: Acrididae)

Cryptic coloration is found in many Orthoptera, especially in Acrididae, showing a great variety of forms. In a grasshopper assemblage in southeastern Brazil, preferences for escape places were detected; grasshoppers tended to escape to backgrounds in which they seem to be more cryptic. Coloration was measured using the Simpson diversity index, to quantify 'aspect diversity' (diversity of colours and shapes of patches along the insect's body). A weak positive correlation was found between grasshoppers' aspect diversity and diversification in use of escape places (use of many backgrounds to escape). Grasshoppers with similar colour patterns tended to use the same structures (leaves, sandy soil, stones) to escape.     

Surface immobilization and entrapping of enzymes on glutaraldehyde crosslinked gelatin particles

A mild and reproducible method has been developed for the surface-immobilization of enzymes on glutaraldehyde crosslinked gelatin beads. In this method glutaraldehyde is used in a dual capacity, as crosslinking agent and as the enzyme coupling agent. Glucoamylase (exo-α-1,4-d-glucosidase, EC 3.2.1.3), β-d-fructofuranosidase (invertase, EC 3.2.1.26) and β-d-glucoside (cellobiase, β-d-glucoside glucohydrolase, EC 3.2.1.21) have been successfully immobilized by this method, on the surface of the crosslinked gelatin particles. The method can be combined with the existing technology for the production of gelatin-entrapped enzymes. Thus, dual immobilized enzyme conjugates of glucoamylase and invertase have been prepared using this method, by entrapment of one enzyme in, and surface-binding of the other to, the gelatin matrix. The coupling of glucoamylase onto cross-linked gelatin particles by precipitation with poly(hexamethylenebiguanide hydrochloride) was also tested.     

Release of poly a(+) messenger RNA from rat liver rough microsomes upon disassembly of bound polysomes

Several procedures were used to disassemble rat liver rough microsomes (RM) into ribosomal subunits, mRNA, and ribosome-stripped membrane vesicles in order to examine the nature of the association between the mRNA of bound polysomes and the microsomal membranes. The fate of the mRNA molecules after ribosome release was determined by measuring the amount of pulse-labeled microsomal RNA in each fraction which was retained by oligo-dT cellulose or by measuring the poly A content by hybridization to radioactive poly U. It was found that ribosomal subunits and mRNA were simultaneously released from the microsomal membranes when the ribosomes were detached by: (a) treatment with puromycin in a high salt medium containing Mg++, (b) resuspension in a high salt medium lacking Mg++, and (c) chelation of Mg++ by EDTA or pyrophosphate. Poly A-containing mRNA fragments were extensively released from RM subjected to a mild treatment with pancreatic RNase in a medium of low ionic strength. This indicates that the 3' end of the mRNA is exposed on the outer microsomal surface and is not directly bound to the membranes. Poly A segments of bound mRNA were also accessible to [(3)H] poly U for in situ hybridization in glutaraldehyde-fixed RM. Rats were treated with drugs which inhibit translation after formation of the first peptide bonds or interfere with the initiation of protein synthesis. After these treatments inactive monomeric ribosomes, as well as ribosomes bearing mRNA, remained associated with their binding sites in microsomes prepared in media of low ionic strength. However, because there were no linkages provided by nascent chains, ribosomes, and mRNA, molecules were released from the microsomal membranes without the need of puromycin, by treatment with a high salt buffer containing Mg++. Thus, both in vivo and in vitro observations are consistent with a model in which mRNA does not contribute significantly to the maintenance of the interaction between bound polysomes and endoplasmic reticulum membranes in rat liver hepatocytes.     

Immobilization of Lipase from Mucor Miehei Onto Poly (Ethylene) Based Graft Copolymers

Lipase from Mucor miehei was immobilized covalently onto hydrolyzed poly(ethylene)-g.co-hydroxyethyl methacrylate (PE-HEMA). This hydrolysis of the copolymer was achieved using 0.1 M NaOH over different periods of time, under controlled conditions. The graft copolymers and their hydrolyzed equivalents were characterized by scanning electron microscopy (SEM) and by differential scanning calorimetry analysis (DSC). Water sorption studies were undertaken to provide a measure of relative hydrophobicity of the samples.

The lipase immobilization reaction was studied in order to assess the effects of controlling various important parameters. These include the nature of the buffering medium, the time over which the immobilization was allowed to occur, the concentration of the activating and coupling agent used (CMC) and the concentration of enzyme employed during attempts at effective immobilization. The immobilized lipase was used in the hydrolysis of triolein (glycerol trioleate). From this study, the apparent K_M, the optimum pH for hydrolysis and the optimum temperature for hydrolysis were revealed.

The suitability of hydrolyzed poly(ethylene)-g.co-HEMA as a support in the immobilization of lipase was assessed by determination of the amount of lipase coupled to the support and by assessment of the retention of activity of the immobilized lipase after its exposure to the immobilization reagents, procedure and conditions.