Preparation of diisononyl adipate in a solvent-free system via an immobilized lipase-catalyzed esterification.

Diisononyl adipate is a plasticizer which has excellent property of low temperature resistance and is permitted in food contact materials. In this study, diisononyl adipate was synthesized from adipic acid and isononyl alcohol in a solvent-free system via immobilized lipase-catalyzed esterification. Liquid Eversa lipase from Thermomyces lanuginosus was immobilized on Lewatit VP OC 1600 carrier. The effects of temperature, substrate molar ratio, water activity of enzyme, vacuum, and enzyme loading on the reaction efficiency were investigated. Application of vacuum played a key role to achieve a 100% conversion. The optimal temperature, molar ratio (adipic acid to isononyl alcohol), water activity of enzyme, vacuum were 50 °C, 1:3, 0.75, 13.3 kPa, and 10% (based on weight of total substrate). Under these conditions, 100% conversion was achieved within 6 h.

Production of stearidonic acid-rich triacylglycerol via a two-step enzymatic esterification.

The aim of this study was to synthesize stearidonic acid (SDA)-rich triacylglycerol (TAG) via a two-step lipase-catalyzed esterification under vacuum. SDA-rich fatty acid, which was prepared from echium oil via Candida rugosa lipase-catalyzed selective esterification, was used as the substrate. Two different immobilized lipases, Novozym 435 from Candida antarctica and Lipozyme TL IM from Thermomyces lanuginosus, were employed for the synthesis of SDA-rich TAG. In the first step, Novozym 435-catalyzed esterification of the SDA-rich fatty acid with glycerol was carried out for 2 h. In the second step, Lipozyme TL IM-catalyzed esterification of the reaction mixture from the first step was performed for an additional 10 h. The optimal reaction conditions for the second step were a temperature of 65 °C, an enzyme loading of 20%, and a vacuum of 0.7 kPa. Consequently, the maximum TAG conversion of ca. 86.4 wt% was obtained after 12 h via a two-step lipase-catalyzed esterification.

Preparation of High Purity Δ5-Olefinic Acids from Pine Nut Oil via Repeated Lipase-Catalyzed Esterification.

Δ5-Olefinic acids have been characterized in gymnosperm plants and have been reported to have several biological health benefits. Δ5-Olefinic acids from pine nut oil were effectively concentrated by repeated lipase-catalyzed esterification. The pine nut oil contained three major Δ5-olefinic acids, namely taxoleic acid (C18:2 Δ5,9), pinolenic acid (C18:3 Δ5,9,12), and sciadonic acid (C20:3 Δ5,11,14). The fatty acids present in pine nut oil were selectively esterified with ethanol using Lipozyme RM IM from Rhizomucor miehei as a biocatalyst. The Δ5-olefinic acids were concentrated in the unesterified fatty acid fraction. The optimum molar ratio of the substrates (fatty acid:ethanol), temperature, the enzyme loading, and the reaction time were 1:7, 25°C, 5% of total substrate weight, and 6 h, respectively. There was no significant effect in the concentration of Δ5-olefinic acids when water was added in the reaction mixture. The same protocol and optimum conditions were employed for two times repeated lipase-catalyzed esterifications. In first lipase-catalyzed esterification, the Δ5-olefinic acids content in the pine nut oil increased from 17 mol% to 51 mol% with a yield of 40 mol%. In a second lipase-catalyzed esterification, with the Δ5-olefinic acids-concentrated fatty acids obtained from the first reaction as the substrate, the Δ5-olefinic acids content increased to 86 mol% with a yield of 15 mol%. Finally, a maximum Δ5-olefinic acids content of ca. 96 mol% with a yield of 6 mol% was obtained via a third lipase-catalyzed esterification.

Synthesis of Fatty Acid Methyl Esters Using Mixed Enzyme in a Packed Bed Reactor.

Fatty acid methyl esters were synthesized from palm fatty acid distillate (PFAD) and methanol in a packed bed reactor via lipase-catalyzed esterification. The PFAD consisted of 91 wt% of free fatty acids, 2 wt% monoacylglycerides, 3 wt% diacylglycerides, and 4 wt% triacylglycerides. t-Butanol was employed as a reaction medium and a mixed enzyme consisting of Lipozyme TL IM from Thermomyces lanuginosus and Novozym 435 from Candida antarctica was employed as the biocatalyst. The effect of mixed enzyme was investigated and the optimum blending ratio (w/w) of Novozym 435 to Lipozyme TL IM was 5:95. Using the mixed enzyme, the optimum molar ratio (PFAD to methanol) and temperature were determined to be 1:6 and 30°C, respectively. Under the optimized conditions, the maximum yield of ca. 96% was achieved.

Production of Phytosteryl Ester from Echium Oil in a Recirculating Packed Bed Reactor Using an Immobilized Lipase.

The synthesis of phytosteryl ester via the lipase-catalyzed esterification of phytosterol with fatty acid from echium oil was performed in a recirculating packed bed reactor. A commercial lipase from Candida rugosa was immobilized and then used to optimize the reaction conditions for the synthesis of the phytosteryl ester. The parameters investigated were temperature, molar ratio of phytosterol to fatty acid, retention time, and amount of solvent. The optimum conditions were determined as the temperature of 40℃, the molar ratio of 1:3 (phytosterol to fatty acid), the retention time of 3 min, and the solvent amount of 40 mL. The maximum conversion of phytosteryl ester was ca. 90 mol% under the optimum conditions. The major fatty acid in phytosteryl ester synthesized was α-linolenic acid (ca. 38 mol%).

Synthesis of α-linolenic acid-rich triacylglycerol using a newly prepared immobilized lipase.

An α-linolenic acid (ALA)-rich triacylglycerol (TAG) was synthesized from an ALA-rich fatty acid (FA) from perilla oil and glycerol, using a newly prepared immobilized lipase under vacuum. The ALA-rich FA (purity >90wt%) used as the substrate was prepared by urea complexation from perilla oil FAs. Liquid Lipozyme TL 100L lipase from Thermomyces lanuginosus was used for immobilization. Nine different hydrophilic and hydrophobic carriers for immobilization were tested, and Duolite A568, which is a hydrophilic resin, was selected as the best carrier. This immobilized lipase was used to synthesize TAG by direct esterification under vacuum. The parameters investigated were temperature, enzyme loading, and vacuum level. The optimum reaction conditions were a temperature of 60°C, an enzyme loading of 15% (based on the total weight of the substrate), and a vacuum of 0.7kPa, respectively. The maximum conversion to TAG of ca. 88wt% was obtained in 12h under the optimum conditions.

Production of Biodiesel from Acid Oil via a Two-Step Enzymatic Transesterification.

A two-step enzymatic transesterification process in a solvent-free system has been developed as a novel approach to the production of biodiesel using acid oil from rice bran oil soapstock. The acid oil consisted of 53.7 wt% fatty acids, 2.4 wt% monoacylglycerols, 9.1 wt% diacylglycerols, 28.8 wt% triacylglycerols, and 6.0 wt% others. Three immobilized lipases were evaluated as potential biocatalysts, including Novozym 435 from Candida antarctica, Lipozyme RM IM from Rhizomucor miehei, and Lipozyme TL IM from Thermomyces lanuginosus. The effects of molar ratio of acid oil to ethanol, temperature, and enzyme loading were investigated to determine the optimum conditions for the transesterification with the three immobilized lipases. The optimum conditions of the three immobilized lipases were a molar ratio of 1:5 (acid oil to ethanol), the temperature range of 30-40°C, and the enzyme loading range of 5-10%. The two-step transesterification was then conducted under the optimum conditions of each lipase. The stepwise use of Novozym 435 and Lipozyme TL IM or Lipozyme RM IM and Lipozyme TL IM resulted in similar or higher levels of yield to the individual lipases. The maximum yields obtained in both stepwise uses were ca. 92%.