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By using pig pancreatic lipase (EC 3.1.1.3 or PPL) as a biocatalyst, covalently immobilized on amorphous AlPO4 support, a new second generation biodiesel was obtained in the transesterification reaction of sunflower oil with ethanol. The resulting biofuel is composed of fatty acid ethyl esters and monoglycerides (FAEE/MG) blended in a 2:1 molar ratio. This novel product, which integrates glycerol as monoacylglycerols (MG) into the biofuels composition, has similar physicochemical properties as conventional biodiesel and also avoids the removal step of the by-product by washing of the biodiesel with water. Immobilization of PPL was achieved by covalent attachment of the ε-amino group of the lysine residues of PPL with the aldehyde groups of p-hydroxybenzaldehyde linked on a hybrid organic-inorganic functionalized AlPO4 surface. With this procedure, the PPL biocatalyst was strongly fixed to the inorganic support surface (94.3%). Nevertheless, the efficiency of the immobilized enzyme was relatively lower compared to that of the free PPL, but it showed a remarkable stability as well as a great capacity of reutilization (25 reuses) without a significant loss of its initial catalytic activity. Therefore, this enzymatic method allows the production of a biodiesel which integrates the glycerol, allows a more efficient fabrication method and minimizes the waste production as compared to the conventional alkali-catalyzed process.