| 1. |
- Adlercreutz, Dietlind, et al.
(författare)
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An enzymatic method for the synthesis of mixed-acid phosphatidylcholine
- 2004
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Ingår i: Journal of the American Oil Chemists Society. - : Wiley. - 0003-021X. ; 81:6, s. 553-557
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Tidskriftsartikel (refereegranskat)abstract
- The enzymatic synthesis of PC with decanoic acid in the sn-1 and hexanoic acid in the sn-2 position is described. The procedure comprises the following enzymatic steps: (i) treatment of egg yolk with phospholipase A(2) (PLA(2)) to hydrolyze egg yolk PC to 1-acyl lysophosphaticlylcholine (LPC) (ii) esterification of I-acyl LPC with hexanoic acid catalyzed by PLA(2) to yield PC with hexanoic acid in the sn-2 position; (iii) removal of the FA in the sn-1 position by lipase-catalyzed ethanolysis to yield 2-hexanoyl LPC; and finally (iv) introduction of decanoic acid in this position by lipase-catalyzed esterification of 2-hexanoyl LPC to yield 1-decanoyl-2-hexanoyl-PC. Two egg yolks with a weight of 16 g were required to obtain 160 mg of the desired product. The chemical purity of the PC product and the positional purity of the FA were around 99%. The method is applicable for the synthesis of other mixed-acid PC species as well.
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| 2. |
- Adlercreutz, Dietlind, et al.
(författare)
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Synthesis of phosphatidylcholine with defined fatty acid in the sn-1 position by lipase-catalyzed esterification and transesterification reaction.
- 2002
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Ingår i: Biotechnology and Bioengineering. - : Wiley. - 1097-0290 .- 0006-3592. ; 78:4, s. 403-411
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Tidskriftsartikel (refereegranskat)abstract
- The incorporation of caproic acid in the sn-1 position of phosphatidylcholine (PC) catalyzed by lipase from Rhizopus oryzae was investigated in a water activity-controlled organic medium. The reaction was carried out either as esterification or transesterification. A comparison between these two reaction modes was made with regard to product yield, product purity, reaction time, and byproduct formation as a consequence of acyl migration. The yield in the esterification and transesterification reaction was the same under identical conditions. The highest yield (78%) was obtained at a water activity (a(w)) of 0.11 and a caproic acid concentration of 0.8 M. The reaction time was shorter in the esterification reaction than in the transesterification reaction. The difference in reaction time was especially pronounced at low water activities and high fatty acid concentrations. The loss in yield due to acyl migration and consequent enzymatic side reactions was around 16% under a wide range of conditions. The incorporation of a fatty acid in the sn-1 position of PC proved to be thermodynamically much more favorable than the incorporation of a fatty acid in the sn-2 position.
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| 3. |
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| 4. |
- Persson, Mattias, et al.
(författare)
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Effects of solvent, water activity and temperature on lipase and hydroxynitrile lyase enantioselectivity
- 2002
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Ingår i: Enzyme and Microbial Technology. - 0141-0229. ; 30:7, s. 916-923
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Tidskriftsartikel (refereegranskat)abstract
- The influence of the reaction conditions on the enantioselectivity of reactions catalysed by lipases or hydroxynitrile leases (HNLs) in organic solvents was investigated. The lipases catalysed kinetic resolution of chiral secondary alcohol, or chiral carboxylic acids and the HNLs catalysed asymmetric addition of hydrogen cyanide to aldehydes. The temperature effects on enantioselectivity, were studied in detail. From measurements of the enantiomeric ratio (C) at different temperatures the activation parameters DeltaDeltaH(#) and DeltaDeltaS(#) were determined. In the lipase-catalysed reactions the enthalpic and entropic effects on E always counteracted, while in a few of the HNL-catalysed reactions, DeltaDeltaH(#) and DeltaDeltaS(#) had opposite sign, and therefore the effects cooperated to give high E values (-RTInE = DeltaDeltaG(#) = DeltaDeltaH(#) - TDeltaDeltaS(#)). In all the HNL-catalysed reactions and most of the lipase-catalysed ones, the enantioselectivity increased with decreasing reaction temperature. However, in one of the lipase-catalysed reactions, the enantioselectivity decreased with decreasing temperature. The theoretical background of these observations wars discussed. In the HNL-catalysed reactions, the enantioselectivity increased with increasing water content up to water saturation, while in the lipase-catalysed reactions the opposite trend was found in one case and in the others no significant effect was observed. Solvent mixtures of diisopropylether and hexane were used to obtain solvents with different log P values. The log P value of the solvent did not influence the enantioselectivity in the HNL-catalysed reactions. while the enantioselectivity increased with increasing log P value in two of the lipase-catalysed reactions. The reaction temperature was shown to be a very useful way to influence enzyme selectivity and the effects obtained could be rationalised. The influence of the reaction medium (solvent and water activity) is much more difficult to rationalise and predict. (C) 2002 Elsevier Science Inc. All rights reserved.
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| 5. |
- Persson, Mattias, et al.
(författare)
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Factors governing the activity of lyophilised and immobilised lipase preparations in organic solvents
- 2002
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Ingår i: ChemBioChem. - 1439-4227. ; 3:6, s. 566-571
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Tidskriftsartikel (refereegranskat)abstract
- Active site titration and activity measurements were performed in hexane on lyophilised lipase preparations containing different amounts of phosphate buffer and lipase immobilised on porous polypropylene. Lyophilisation of Thermomyces lanuginosus lipase with large quantities of phosphate salts (200 mm) increased the specific activity fourfold, and the number of rapidly titratable active sites increased to 50% from the 73% observed when smaller amounts of phosphate buffer were used (20 mm) during lyophilisation. The phosphate buffer worked as an immobilisation matrix for the lipase, and the increase in specific activity was at least portly due to decreased mass transfer limitations. When lipase was immobilised on porous polypropylene, the specific activity was 770 times higher than that of the best freeze-dried preparation. At optimal enzyme loading, 93% of the enzyme molecules were titrated at a high rate; this indicates that this adsorption on a hydrophobic surface was a very efficient means of reducing moss transfer limitations and of immobilising the enzyme in its active conformation for use in organic solvents. The variation in specific activity with water activity was found to correlate very well with the variation in titratable active sites when lipases from Burkholderia cepacia and Thermomyces lanuginosus were immobilised on porous polypropylene. The catalytic activity per competent active site was thus constant over the whole range of water activities.
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| 6. |
- Persson, Mattias, et al.
(författare)
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Preparation of lipases for use in organic solvents
- 2002
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Ingår i: Enzyme and Microbial Technology. - 0141-0229. ; 31:6, s. 833-841
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Tidskriftsartikel (refereegranskat)abstract
- The efficiency of different preparations of lipases was evaluated in organic solvents. Lipases from Humicola lanuginosa, Candida rugosa, Rhizomucor miehei and Pseudomonas cepacia were adsorbed onto the surfactant sorbitan monostearate (Span 60) and the specific activities were compared in hexane to crude powder (used straight from the bottle) and lipase freeze-dried from buffer solution. Lipases adsorbed on the surfactant were "activated" 1.9- to 150-fold compared to the crude lipase. The solubility of the lipase-surfactant preparation in the reaction media was extremely low and the preparation contained aggregates of micrometer size. In further comparisons lipase from H. lanuginosa was freeze-dried in the presence of KCl, crown ethers, immobilised by entrapment into a sol-gel and immobilised on porous polypropylene support (Accurel EP-100). Addition of potassium chloride before freeze-drying of the lipase increased the activity up to 46-fold compared to crude powder. The additive probably worked as an immobilisation matrix for the lipase. When 18-crown-6 was added to the lipase before freeze-drying a 40-fold increase in activity was achieved. In this case a low amount of additive (0.4 mg crown ether/g protein) was needed for activation indicating that specific interactions were involved in the activation. In order to obtain maximal activity, immobilisation on Accurel EP-100 and entrapment into a sol-gel were the best methods to use. The activities were 400- and 320-fold better than that of crude powder. The high activities obtained were due to an improved dispersion of the catalyst in the organic media. The protein loading that could be used when the lipase was adsorbed onto Accurel EP-100 was much higher than what could be used when the lipase was entrapped into a sol-gel. This makes the adsorption technique the best for practical applications. (C) 2002 Elsevier Science Inc. All rights reserved.
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| 7. |
- Wehtje, Ernst, et al.
(författare)
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Stabilization of Adsorbed Enzymes Used as Biocatalysts in Organic Solvents
- 1992. - C
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Ingår i: Progress in Biotechnology. - 0921-0423. ; 8:C, s. 377-382
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Bokkapitel (refereegranskat)abstract
- Enzymes immobilized by deposition onto a solid support at low enzyme loadings were shown to be stabilized by additives, such as proteins and PEG. The additive protects the enzyme from deactivation during immobilization. The amount of additive required to obtain full stabilization was dependent on the type of support material used and corresponded approximately to a monolayer coverage of the additive on the surface of the support. The stabilizing effect of the additive was less pronounced at high enzyme loadings.
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| 8. |
- Wehtje, Ernst, et al.
(författare)
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Water activity and substrate concentration effects on lipase activity.
- 1997
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Ingår i: Biotechnology and Bioengineering. - 1097-0290. ; 55:5, s. 798-806
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Tidskriftsartikel (refereegranskat)abstract
- Catalytic activity of lipases (from Rhizopus arrhizus, Canadida rugosa, and Pseudomonas sp. was studied in organic media, mainly diisopropyl ether. The effect of water activity (a(w)) on V(max) showed that the enzyme activity in general increased with increasing amounts of water for the three enzymes. This was shown both for esterification and hydrolysis reactions catalyzed by R. arrhizus lipase. In the esterification reaction the K(m) for the acid substrate showed a slight increase with increasing water activities. On the other hand, the K(m) for the alcohol substrate increased 10-20-fold with increasing water activity. The relative changes in K(m) were shown to be independent of the enzyme studied and solvent used. The effect was attributed to the increasing competition of water as a nucleophile for the acyl-enzyme at higher water activities. In a hydrolysis reaction the K(m) for the ester was also shown to increase as the water activity increased. The effect of water in this case was due to the fact that increased concentration of one substrate (water), and thereby increased saturation of the enzyme, will increase the apparent K(m) of the substrate (ester) to be determined. This explained why the hydrolysis rate decreased with increasing water activity at a fixed, low ester concentration. The apparent V(max) for R. arrhizus lipase was similar in four of six different solvents that were tested; exceptions were toulene and trichloroethylene, which showed lower values. The apparent K(m) for the alcohol in the solvents correlated with the hydrophobicity of the solvent, hydrophobic solvents giving lower apparent K(m). (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 798-806, 1997.
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| 9. |
- Adlercreutz, Patrick, et al.
(författare)
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Enzymatic conversions of polar lipids. Principles, problems and solutions
- 2001
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Ingår i: Journal of Molecular Catalysis - B Enzymatic. - 1381-1177. ; 11:4-6, s. 173-178
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Forskningsöversikt (refereegranskat)abstract
- This text provides a brief overview of the principles of enzymatic lipid conversion and some recent advances in the enzymatic conversion of glycerophospholipids and galactolipids. Lipases and phospholipases are used to exchange fatty acids or the polar group in the lipids. The reactions can be carried out either as hydrolysis-esterification sequences or as one-step transferase reactions. The scope and limitations of the different methods are discussed.
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| 10. |
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