Efficient immobilization of lipases by entrapment in hydrophobic sol‐gel materials

The commercial application of lipases as biocatalysts for organic synthesis requires simple but efficient methods to immobilize the enzyme, yielding highly stable and active biocatalysts which are easy to recover. In this study, we present a novel method to achieve lipase immobilization by entrapment in chemically inert hydrophobic silica gels which are prepared by hydrolysis of alkyl‐substituted silanes in the presence of the enzyme. A typical immobilization procedure uses: an aqueous solution of lipase; sodium fluoride as a catalyst; and additives like polyvinyl alcohol or proteins and alkoxysilane derivatives like RSi‐(OMe)3 with R = alkyl, aryl, or alkoxy as gel precursors. The effect of various immobilization parameters like stoichiometric ratio of water, silane, type and amount of additive, type and amount of catalyst, and type of silane has been carefully studied. The new method is applicable for a wide variety of lipases, yielding immobilized lipases with esterification activities enhanced by a factor of up to 88, compared to the commercial enzyme powders under identical conditions. Studies on the stability of sol‐gel immobilized lipases under reaction conditions or storage (dry, in aqueous or organic medium) revealed an excellent retention of enzymatic activity. The possible reasons for the increased enzyme activities are discussed. © 1996 John Wiley & Sons, Inc.

[1]  Bruce Dunn,et al.  Sol-gel encapsulation methods for biosensors , 1994 .

[2]  M. Reetz,et al.  Efficient Heterogeneous Biocatalysts by Entrapment of Lipases in Hydrophobic Sol–Gel Materials , 1995 .

[3]  W. Stöber,et al.  Controlled growth of monodisperse silica spheres in the micron size range , 1968 .

[4]  L. Thim,et al.  A model for interfacial activation in lipases from the structure of a fungal lipase-inhibitor complex , 1991, Nature.

[5]  J. Bosley,et al.  Blueprint for a lipase support: Use of hydrophobic controlled‐pore glasses as model systems , 1994, Biotechnology and bioengineering.

[6]  C. H. Amundson,et al.  Immobilized lipase reactors for modification of fats and oils—A review , 1990 .

[7]  L. Sarda,et al.  Action de la lipase pancréatique sur les esters en émulsion , 1958 .

[8]  B. Mattiasson,et al.  On the importance of the support material for bioorganic synthesis. Influence of water partition between solvent, enzyme and solid support in water-poor reaction media. , 1988, European journal of biochemistry.

[9]  E. Ruckenstein,et al.  Lipase immobilized on hydrophobic porous polymer supports prepared by concentrated emulsion polymerization and their activity in the hydrolysis of triacylglycerides , 1993, Biotechnology and bioengineering.

[10]  P. Halling,et al.  Use of salt hydrates to buffer optimal water level during lipase catalysed in synthesis in organic media: a practical procedure for organic chemists , 1992 .

[11]  B. Mattiasson,et al.  Improved activity retention of enzymes deposited on solid supports , 1993, Biotechnology and bioengineering.

[12]  David Avnir,et al.  Enzymes and Other Proteins Entrapped in Sol-Gel Materials , 1994 .

[13]  V. V. Mozhaev,et al.  Strategy for Stabilizing Enzymes Part One: Increasing Stability of Enzymes via their Multi-Point Interaction with a Support , 1990 .

[14]  W. Boland,et al.  Esterolytic and Lipolytic Enzymes in Organic Synthesis , 1992 .