Frozen Microemulsions for MAPLE Immobilization of Lipase

Candida rugosa lipase (CRL) was deposited by matrix assisted pulsed laser evaporation (MAPLE) in order to immobilize the enzyme with a preserved native conformation, which ensures its catalytic functionality. For this purpose, the composition of the MAPLE target was optimized by adding the oil phase pentane to a water solution of the amino acid 3-(3,4-dihydroxyphenyl)-2-methyl-l-alanine (m-DOPA), giving a target formed by a frozen water-lipase-pentane microemulsion. Fourier transform infrared (FTIR) spectroscopy and atomic force microscopy (AFM) were used to investigate the structure of MAPLE deposited lipase films. FTIR deconvolution of amide I band indicated a reduction of unfolding and aggregation, i.e., a better preserved lipase secondary structure in the sample deposited from the frozen microemulsion target. AFM images highlighted the absence of big aggregates on the surface of the sample. The functionality of the immobilized enzyme to promote transesterification was determined by thin layer chromatography, resulting in a modified specificity