*Rhizomucor miehei* **Lipase Supported on Inorganic Solids, as Biocatalyst for the Synthesis of Biofuels: Improving the Experimental Conditions by Response Surface Methodology**

**Juan Calero 1, Diego Luna 1,\*, Carlos Luna 1, Felipa M. Bautista 1, Beatriz Hurtado 1, Antonio A. Romero 1, Alejandro Posadillo 2 and Rafael Estevez 1**


Received: 12 February 2019; Accepted: 26 February 2019; Published: 2 March 2019

**Abstract:** Two inorganic solids have been evaluated as supports of Lipozyme RM IM, a *Rhizomucor miehei* lipase immobilized on a macroporous anion exchange resin, in order to improve its application as a biocatalyst in the synthesis of biofuels. The experimental conditions have been optimized to ge<sup>t</sup> the selective transesterification of sunflower oil, by using a multi-factorial design based on the response surface methodology (RSM). In this way, the effects of several reaction parameters on the selective ethanolysis of triglycerides to produce Ecodiesel, a biodiesel-like biofuel constitute by one mole of monoglyceride (MG) and two moles of fatty acid ethyl ester (FAEE), have been evaluated. Thus, it was obtained that a 6:1 oil/ethanol molar ratio, 0.215 g of biocatalyst supported in silica-gel (0.015 g Lipase/0.2 g silica-gel), 50 μL of 10 N NaOH, together with previous optimized reaction parameters, 35 ◦C reaction temperature and 120 min of reaction time, gave the best results (conversions around 70%; selectivity around 65%; kinematic viscosities about 9.3 mm2/s) in the reaction studied. Besides, Lipozyme RM IM, supported on silica-gel, biocatalyst exhibited a very good stability, remaining its activity even after 15 cycles.

**Keywords:** biodiesel; Ecodiesel; selective ethanolysis; sunflower oil; Lipozyme RM IM; *Rhizomucor miehei*; ANOVA method; response surface methodology
