The aim of this study was to investigate a new process of enzyme/enzyme hydroesterification for biodiesel
production using a low-cost acid oil (10.5 wt.% acidity) from macauba (Acrocomia aculeata) pulp as
raw material. The ethyl esters were produced by the hydrolysis of the macauba oil using vegetable
enzyme (VE) obtained from dormant castor seeds followed by esterification of the released free fatty
acids (FFAs) with ethanol catalyzed by fermented and dry babassu cake with lipase activity from Rhizomucor
miehei. The vegetable enzyme-catalyzed hydrolysis produced 99.6% of FFAs after 6 h in a medium
with high oil concentration (50% v/v) and without organic solvent and emulsifier. For the esterification
reaction, the best result was attained with an ethanol:FFA molar ratio of 2:1 and 15.1 U of dry fermented
solid per g of FFAs at 40 C, which yielded 91% of conversion after 8 h in a solvent-free system. In order to
confirm the potential of the fermented solid as biocatalyst, it was confronted with the best commercial
lipases and was also evaluated for its reuse. Similar conversions were obtained with the commercial
lipases Novozym 435 and Lipozyme RM IM and the fermented solid. The fermented solid was reused
in successive 6-h batches for esterification reactions and conversions of over 60% were maintained for
eight cycles. After two consecutive esterification reactions the resulting biodiesel met important Brazilian
standards such as: density (ASTM D4052), viscosity kinematic (ASTM D445), flash point (ASTM D93), carbon
residue (ASTM D4530), free glycerol and total glycerol, monoglycerides and triglycerides (ASTM
D6584). The ester content was of 96.7% (esters of fatty acids of 8–18 carbons). To the best of our knowledge,
this is the first time that an enzyme/enzyme hydroesterification process using low cost biocatalysts
obtained from vegetable and microorganism using solvent-free media in both reactions is described for
the conversion of an acid and low value oil into biodiesel.