Improvement of activity and stability of Rhizomucor miehei lipase by immobilization on nanoporous aluminium oxide and potassium sulfate microcrystals and their applications in the synthesis of aroma esters

YILDIRIM D. , Baran E., Ateş S., Yazici B. , TÜKEL S. S.

BIOCATALYSIS AND BIOTRANSFORMATION, vol.37, no.3, pp.210-223, 2019 (Journal Indexed in SCI) identifier identifier

  • Publication Type: Article / Article
  • Volume: 37 Issue: 3
  • Publication Date: 2019
  • Doi Number: 10.1080/10242422.2018.1530766
  • Page Numbers: pp.210-223


In this study, Rhizomucor miehei lipase (RML) was immobilized on the hexagonally-ordered nanoporous aluminium oxide membranes (RML-Al2O3-NP) by adsorption and as protein-coated microcrystals (RML-PCMCs) by simultaneously precipitating RML on micron-sized potassium sulfate crystals (K2SO4) in pre-chilled acetone. The hydrolytic activities of immobilized lipase preparations were investigated in terms of p-nitrophenyl palmitate hydrolysis and their esterification activities were examined for the synthesis of some aroma esters such as butyl acetate, isoamyl acetate, hexyl acetate, heptyl acetate, and geranyl acetate. The immobilization yields were 33.8 and 25.1%, respectively for RML immobilized on Al2O3-NP membranes and potassium sulfate crystals. The catalytic efficiency ratios of RML-Al2O3-NP and RML-PCMCs were 2.3- and 3.9-fold higher than that of the free lipase, respectively in terms of hydrolytic activity. The free lipase was stabilized as 4.1- and 10.5-fold, respectively at 40 and 50 degrees C when immobilized on Al2O3-NP. The corresponding stabilization factors were 4.6- and 12.8-fold higher for RML-PCMCs. RML-Al2O3-NP and RML-PCMCs maintained 84 and 86% of their initial hydrolytic activities, respectively after 10 reuses. Of the synthesized aroma esters, the highest yield was obtained for the geranyl acetate. After 4 h reaction time, no geraniol was detected in the preparative-scale (196 g/L) synthesis of geranyl acetate for both the immobilized lipases when the initial geraniol amount, vinyl acetate amount, RML-PCMCs amount, and reaction temperature values were 1 mmol, 3 mmol, 100 mg (or 300 mg RML-Al2O3-NP), and 50 degrees C, respectively. These results show that the immobilization of R. miehei lipase by adsorption on nanoporous aluminium oxide and as protein-coated microcrystals leads to the obtention of highly stable, catalytically more active, and reusable lipase preparations.