The Amano AK lipase from Pseudomonas fluorescens was immobilized in fibroin-alginate spheres (FA-LPf) and used to catalyze the transesterification reaction of soybean oil under different conditions (solvents, time, amounts of immobilized lipase, and temperature) for the production of biodiesel (fatty acid ethyl esters, FAEE). The FAEE was quantified by GC-FID analysis, and tricaprin was used as an internal standard. The maximum FAEE yield (63%) was attained after 96 h at 32 A degrees C and 400 rpm in 30% (v/v) n-hexane, which used 20% (w/w) of the immobilized lipase. However, it is noteworthy that with a period of 96 h, the spheres of FA-LPf were partially destroyed by friction from the magnetic stirring, and the long time for the transesterification reactions is impractical in these conditions. For this reason, the reaction time was reduced to 48 h, which resulted in 42% FAEE and assures better reuse conditions for four consecutive cycles. This study shows the efficacy of an enzyme immobilized in a biodegradable FA-LPf support. Graphical Abstract [GRAPHICS] .