Pilot‐scale development of core–shell polymer supports for the immobilization of recombinant lipase B from Candida antarctica and their application in the production of ethyl esters from residual fatty acids

Abstract

A recombinant lipase B from Candida antarctica (LipB) in Pichia pastoris was synthesized through submerged fermentation using crude glycerin as substrate. The immobilization of this enzyme on the core–shell polymeric supports is an effective alternative for its application. The supports with distinct levels of hydrophobicity were produced through combined suspension and emulsion polymerization in pilot scale. Particles with distinct compositions were synthesized (PMMA/PMMA; PMMA‐co‐DVB/PMMA‐co‐DVB; and PS‐co‐DVB/PS‐co‐DVB) and employed on the immobilization of the produced lipase (LipB) and the commercial enzyme (CalB). The morphological properties (specific area, average pore diameter, specific volume of pores, and hydrophobicity level) and the influence of the polymerization conditions on the morphology of the supports were studied. The thermal stability of such biocatalysts was also investigated in the presence of calcium cation (Ca+2), maintained 100% of the activity after 3 h at 50°C when the PMMA‐co‐DVB/PMMA‐co‐DVB was employed. The synthesized enzyme and supports manufactured in pilot scale were employed successfully for production of esters using residual fatty acids as substrates, adding value to these raw materials and increasing the ranges of possible applications.

Production and optimization of isopropyl palmitate via biocatalytic route using home‐made enzymatic catalysts

“…On the other hand, CALB‐PMMA/PMMA exhibited the lowest conversion of ester, achieving 14.2% of ester after 4 h of reaction. In the esterification reaction of oleic acid and ethanol in solvent‐free medium, the biocatalyst CALB‐P(S‐ co ‐DVB)/P(S‐ co ‐DVB) also presented the highest ester conversion …”

“…So, the distinct activities of the biocatalysts were a result of the supports hydrophobicity. The higher hydrophobic degree of P(S‐co‐DVB)/P(S‐co‐DVB) supports, for example, probably contributes to higher exposure of the active site of the lipase after the immobilization (contact angle 92.3° vs 56.6° for PMMA–PMMA) . Besides, the higher hydrophobic feature may also contribute to the mass transfer of the substrates and product during the reaction, resulting in an increase in the ester conversion.…”

“…Briefly, the core–shell supports were produced by a combined suspension and emulsion polymerization process: first, a suspension polymerization was carried out for synthesis of the cores; after this step, the emulsion constituents were added to the reactor for synthesis of the porous shell. Therefore, particles of PMMA/PMMA; P(S ‐co‐ DVB)/P(S ‐co‐ DVB) (25 wt% of DVB) and P(MMA ‐co‐ DVB)/P(MMA ‐co‐ DVB) (25 wt% of DVB) were synthesized …”

“…Based on this procedure, CAL‐B is physically adsorbed onto hydrophobic supports . The synthesis of such home‐made biocatalysts ((CALB‐PMMA/PMMA) – CALB immobilized on core–shell supports of poly(methyl methacrylate); (CALB‐P(MMA ‐co‐ DVB)/P(MMA ‐co‐ DVB)) – CALB immobilized on core–shell supports of poly(methyl methacrylate‐co‐ divinylbenzene); (CALB‐P(S‐ co ‐DVB)/P(S‐ co ‐DVB)) – CALB immobilized on core–shell supports was described in previous work …”

“…This polymerization technique can be very advantageous since it allows the production of micrometric porous particles and permits functionalization of the core and the shell of the polymer carriers obtained, without additional reaction steps . The use of these porous core–shell supports has been reported in some studies for the immobilization of the recombinant lipase B from Candida antarctica (CALB) . Such biocatalysts were used in some standard esterification reactions, using oleic acid and ethanol as substrates, for example .…”