Production of glycerides from glycerol and fatty acid by immobilized lipases in non-aqueous media

Immobilized Mucor miehei lipase catalyzes synthesis reactions between glycerol and oleic acid. No organic solvent is necessary to solubilize the substrates, which allows for the use of a reaction medium solely composed of the necessary substrates. Water produced in the reaction evaporates due to the high temperature used for the process. A conversion of 86% of oleic acid into triolein is obtained when using the substrates in stoichiometric amounts. Varying the ratio of glycerol over oleic acid allows for the preferential synthesis of one of the glycerides. Some batch reactors have been set up using different means of removing the water: spontaneous evaporation, molecular sieves, vacuum, and dry air bubbling.     

Ester synthesis in aqueous media in the presence of various lipases

Summary The ability of seven lipase preparations to catalyse methyl ester synthesis in aqueous media was compared and the synthesis reaction (esterification or alcoholysis) determined. Three behaviours were observed: three enzymes catalysed ester synthesis by esterification of free fatty acids and one enzyme catalysed alcoholysis but the other three lipases did not catalyse a net ester synthesis under the conditions tested. The three groups also differed by the influence of methanol on the hydrolysis reaction. The first group was not significantly inhibited up to the highest methanol concentration tested (5 M). Hydrolysis in the presence of the enzyme of the second group was increasingly inhibited with increasing methanol concentrations. In the presence of the third group, hydrolysis was 40 to 50% inhibited for all the concentrations tested (0.2–5 M).     

Synthesis and spectral properties of DNA capped CdS nanoparticles in aqueous and non-aqueous media

Synthesis of novel cadmium sulphide nanoparticles has been carried out in aqueous and non-aqueous media. DNA has been added during the synthesis of the nanoparticles, which results into cadmium-rich nanoparticles forming a stable complex with DNA. These particles exhibit strong fluorescence, spectral nature of which depends upon the medium in which the particles are synthesized. When interacted with proteins, fluorescence peak intensity of CdS nanoparticles increases considerably. It is possible that such CdS nanoparticles would be useful as a protein sensor.     

Production of 10(S)-hydroxy-8(E)-octadecenoic acid mono-estolides by lipases in non-aqueous media

In this study, Novozym 435, a lipase B from Candida antarctica, was used for fatty acid polymerization. For the first time, an apolar reaction media, n-hexane, was used to synthesize in vitro estolides from trans-hydroxy-fatty acids derived from the biotransformation of oleic acid by Pseudomonas aeruginosa 42A2 NCIMB 40045. We studied the effects of the substrate, the enzyme ratio, the enzyme stability and the reusability of the biocatalyst. To determine the structure of the oligomers formed, both liquid chromatography mass spectrometry and MALDI-TOF mass spectrometry, with a DHB matrix neutralized with lithium hydroxide, were used to obtain simpler mass spectra. Estolides composed of two units of (10S)-HOME were synthesized with a reaction yield of 30%. Finally, various lipases were screened, and another apolar organic solvent, iso-octane, was assayed to try to increase the reaction yield.     

Methods for detection and characterization of lipases: A comprehensive review

Microbial lipases are very prominent biocatalysts because of their ability to catalyze a wide variety of reactions in aqueous and non-aqueous media. The chemo-, regio- and enantio-specific behaviour of these enzymes has caused tremendous interest among scientists and industrialists. Lipases from a large number of bacterial, fungal and a few plant and animal sources have been purified to homogeneity. This article presents a critical review of different strategies which have been employed for the detection, purification and characterization of microbial lipases.     

Reactions of 6,6-dimethyl-2,2-bipyridyl with iron(II) in aqueous and non-aqueous media

Reaction of anhydrous FeCl₂ with 6,6^′-dimethyl-2,2^′-bipyridyl (dmby) in non-aqueous media gives the yellow, high spin, tetrahedral complex FeCl₂(dmby), which is characterized crystallographically, magnetically and by ¹H NMR spectroscopy. In contrast, reaction of FeCl₂ · 4H₂O with dmby in 0.1 M hydrochloric acid, the method of choice for preparing 3:1 and 2:1 iron(II) complexes of 2,2^′-bipyridyl, gives [H₂dmby][FeCl₄] and [Hdmby][FeCl₄], in which the dmby has been protonated. These complexes are also characterized crystallographically.     

Microbial consumption and production of volatile organic compounds at the soil-litter interface

Substantial amounts of volatile organic compounds (VOCs) can be released during decomposition and these compounds can affect atmospheric chemistry, belowground processes, and the structure of microbial communities in litter and soil. However, we have a limited understanding of the types, quantities and ecological impacts of VOCs emitted from litter. Here we used a closed flow-through system and proton transfer reaction mass spectrometry (PTR-MS) to characterize VOC emissions from soil and two litter types (Pinus taeda and Acer rubrum) over a 72-day incubation period. Microbial respiration rates were measured throughout the incubation, and the soils were harvested at the end of the incubation to determine how litter VOCs influenced soil C dynamics, N mineralization rates, and bacterial communities. Using the PTR-MS we identified over 100 VOCs, with 10 VOCs making up the majority of emissions. VOCs accounted for up to 2.5% of the C flux from litter. Soil was a net sink of litter VOCs, absorbing up to 80% of VOCs released by litter, and exposure of soil to litter VOCs increased microbial respiration rates in soil by up to 15%. However, we observed negligible impacts of litter VOCs on soil nutrient levels and bacterial community structure, suggesting that soils must be exposed to higher concentrations of VOCs than observed in our study, to cause effects on these soil characteristics. Overall, VOCs appear to have an important influence on C dynamics at the soil-litter interface and VOC emissions from decomposing litter may represent an understudied component of biosphere–atmosphere interactions.     

Enhancement of Candida rugosa lipase activity in non-aqueous media by co-lyophilization with amphiphiles from aqueous solution

Modified Candida rugosa lipase was co-lyophilized with two gemini-type amphiphiles, l- and d-2-(3-bis-[3-(2,3,4,5,6-pentahydroxy-hexanoylamino)-propyl]-carbamoyl -propionylamino)-pentanedioic acid didodecyl ester or dodecanoic acid 2-[(3-bis-[3-(2,3,4,5,6-pentahydroxy-hexanoylamino)-propyl]-carbamoyl -propionyl)-(2-dodecanoyloxy-ethyl)-amino]-ethyl ester. Enzymatic activities of the modified lipases in the transesterification between racemic 2,2-dimethyl-1,3-dioxolane-4-methanol and vinyl butyrate in cyclohexane were enhanced as much as by 37-78, 1.5–5- and 41–83-fold of magnitude relative to that of native enzyme, respectively. The lack of significant enhancement of the enzymatic activity, only in the case of the d-isomeric amphiphile-modified lipase, was considered from the topological view of the amphiphile.     

Studies on the mechanism of the lipase reaction. II. Comparative studies on the adsorption of lipases and various proteins at the air-water interface.

Adsorption of lipases (EC 3.1.1.3) and various proteins at the air-water interface has been investigated in relation to the mechanism of lipase reaction. Aqueous solutions of lipases and denaturated proteins show surface activity as strong as that of synthetic detergents. However, ths surface activity of esterases and various other proteins is little or none. By foam fractionation it was shown that lipases were adsorbed at the air-water interface and the adsorption followed the equation of Langmuir's adsorption isotherm. The properties of lipase at the interface are discussed in relation to the mechanism of lipase reaction and the differences from the esterase reaction.     

Phospholipase A2 at the bilayer interface.

Interfacial catalysis is a necessary consequence for all enzymes that act on amphipathic substrates with a strong tendency to form aggregates in aqueous dispersions. In such cases the catalytic event occurs at the interface of the aggregated substrate, the overall turnover at the interface is processive, and it is influenced the molecular organization and dynamics of the interface. Such enzymes can access the substrate only at the interface because the concentration of solitary monomers of the substrate in the aqueous phase is very low. Moreover, the microinterface between the bound enzyme and the organized substrate not only facilitates formation of the enzyme-substrate complex, but a longer residence time of the enzyme at the substrate interface also promotes high catalytic processivity. Binding of the enzyme to the substrate interface as an additional step in the overall catalytic turnover permits adaptation of the Michaelis-Menten formalism as a basis to account for the kinetics of interfacial catalysis. As shown for the action of phospholipase A2 on bilayer vesicles, binding equilibrium has two extreme kinetic consequences. During catalysis in the scooting mode the enzyme does not leave the surface of the vesicle to which it is bound. On the other hand, in the hopping mode the absorption and desorption steps are a part of the catalytic turnover. In this minireview we elaborate on the factors that control binding of pig pancreatic phospholipase A2 to the bilayer interface. Binding of PLA2 to the interface occurs through ionic interactions and is further promoted by hydrophobic interactions which probably occur along a face of the enzyme, with a hydrophobic collar and a ring of cationic residues, through which the catalytic site is accessible to substrate molecules in the bilayer. An enzyme molecule binds to the surface occupied by about 35 lipid molecules with an apparent dissociation constant of less than 0.1 pM for the enzyme on anionic vesicles compared to 10 mM on zwitterionic vesicles. Results at hand also show that aggregation or acylation of the protein is not required for the high affinity binding or catalytic interaction at the interface.     

Studies on the mechanism of crown-ether-induced activation of enzymes in non-aqueous media.

Studies on the mechanism of crown-ether-induced activation are described in this paper. Michaelis Menten kinetics of -chymotrypsin in toluene in the presence and absence of 18-crown-6 showed that only V_max is increased upon crown ether treatment. Parallel Lineweaver–Burk plots indicate that crown ethers do not activate the enzyme by specific interactions in the active site, such as transition state stabilization or facilitated transport of water molecules. Increased V_max values of crown-ether-treated enzyme most probably originate from conformational changes, which alter k_cat as well as the amount of catalytically active enzyme.     

森林凋落物的微生物分解

森林凋落物的分解是森林生态系统中物质循环和能量流动的一个重要环节,而微生物在这一过程中起着重要作用。本文系统介绍了森林凋落物微生物分解的过程及其生态学意义,并从参与凋落物分解的微生物多样性、凋落物分解过程中的微生物数量动态及群落演替、影响微生物分解的因素及微生物分解酶学等方面综述了森林凋落物的微生物分解研究概况,探讨了未来研究的方向。     

Microbial decolouration of azo dyes: A review

The textile industry is a substantial consumer of water and produces enormous volumes of contaminated water; the most important contaminants are azo dyes. Microbial processes for the treatment of textile wastewater have the advantage of being cost-effective and environmentally friendly and producing less sludge. The most promising microorganisms for wastewater treatment are those isolated from sites contaminated with dyes or from the sludge of treatment plants because they have adapted to survive in adverse conditions. The mechanism of microbial decolouration occurs from adsorption, enzymatic degradation or a combination of both. Both reductases and oxidases are involved in the microbial degradation process. The goal of microbial treatment is to decolourise and detoxify the dye-contaminated effluents. In this review, we summarise the methodologies used to evaluate the toxicity of azo dyes and their degradation products. Recent studies on the decolouration or degradation of azo dyes using algae, yeast, filamentous fungi and bacteria, genetically modified microorganisms, microbial consortia and microbiological systems combined with Advanced Oxidation Processes (AOPs) and Microbial Fuel Cells (MFCs) are discussed in this review.     

Microbial transglutaminase and its application in the food industry. A review

The extremely high costs of manufacturing transglutaminase from animal origin (EC 2.3.2.13) have prompted scientists to search for new sources of this enzyme. Interdisciplinary efforts have been aimed at producing enzymes synthesised by microorganisms which may have a wider scope of use. Transglutaminase is an enzyme that catalyses the formation of isopeptide bonds between proteins. Its cross-linking property is widely used in various processes: to manufacture cheese and other dairy products, in meat processing, to produce edible films and to manufacture bakery products. Transglutaminase has considerable potential to improve the firmness, viscosity, elasticity and water-binding capacity of food products. In 1989, microbial transglutaminase was isolated from Streptoverticillium sp. Its characterisation indicated that this isoform could be extremely useful as a biotechnological tool in the food industry. Currently, enzymatic preparations are used in almost all industrial branches because of their wide variety and low costs associated with their biotechnical production processes. This paper presents an overview of the literature addressing the characteristics and applications of transglutaminase.     

Microbial processes at the aerobic-anaerobic interface in the deep-water zone of the black sea

Chemical and key microbiological processes (assimilation of carbon dioxide, oxidation and formation of methane, and sulfate reduction) occurring at the aerobic-anaerobic interface in the deep-water zone of the Black Sea were investigated. Measurements were taken at depths from 90 to 300 m at intervals of 5–10 m. The integral rate of the dark assimilation of carbon dioxide varied from 120 to 207 mg C/(m² day) with a maximum at the boundary of cyclonic currents. The organic matter (OM) formed from methane comprised less than 5% of the OM formed from carbon dioxide. A comparison between the rates of methane oxidation and methane production suggests that methane that is oxidized at depths from 100 to 300 m was formed in deeper water horizons. The maximum rate of sulfate reduction (1230 mg S/(m₂ day)) was observed in the western halistatic region, and the minimum rate (490 mg S/(m₂ day)), in the eastern halistatic region. The average rate of hydrogen sulfide production measured at three deep-sea stations amounted to 755 mg S/(m₂ day), or 276 g S/(m₂ year).     

Enantioselective recognition mechanism of secondary alcohol by surfactant-coated lipases in nonaqueous media.

The enantioselective recognition mechanism of secondary alcohol by lipases originated from Candida rugosa and Pseudomonas cepacia was elucidated on the basis of the kinetic study of the esterification of alcohol with lauric acid in isooctane. To obtain inherent kinetic parameters, we utilized a surfactant-coated lipase whose conformation is considered to be an "open" form in a homogeneous organic solvent. Based on the experimental results, the enantioselectivity of lipases was found to be derived from the difference in the V(max) values between the two enantiomers. The same result was observed when lipases of different origin and substrates with different molecular structures were applied. © 1999 John Wiley & Sons, Inc.