Synthesis of 1,3‐dicapryloyl‐2‐docosahexaenoylglycerol by a combination of nonselective and <i>sn</i>‐1,3‐selective lipase reactions

TAG (MLM) with medium-chain FA (MCFA) at the 1,3-positions and long-chain FA (LCFA) at the 2-position, and TAG (LMM) with LCFA at the 1(3)-position and MCFA at 2,3(1)-positions are a pair of TAG regioisomers. Large-scale preparation of the two TAG regioisomers was attempted. A commercially available FFA mixture (FFA-CLA) containing 9-cis,11-trans (9c,11t)-and 10t,12c-CLA was selected as LCFA, and caprylic acid (C 8 FA) was selected as MCFA. The MLM isomer was synthesized by acidolysis of acylglycerols (AG) containing two CLA isomers with C 8 FA: A mixture of AG-CLA/C 8 FA (1:10, mol/mol) and 4 wt% immobilized Rhizomucor miehei lipase was agitated at 30°C for 72 h. The ratio of MLM to total AG was 51.1 wt%. Meanwhile, LMM isomer was synthesized by acidolysis of tricaprylin with FFA-CLA: A mixture of tricaprylin/FFA-CLA (1:2, mol/mol) and 4 wt% immobilized R. miehei lipase was agitated at 30°C for 24 h. The ratio of LMM to total AG was 51.8 wt%. MLM and LMM were purified from 1,968 and 813 g reaction mixtures by stepwise short-path distillation, respectively. Consequently, MLM was purified to 92.3% with 49.1% recovery, and LMM was purified to 93.2% with 52.3% recovery. Regiospecific analyses of MLM and LMM indicated that the 2-positions of MLM and LMM were 95.1 mol% LCFA and 98.3 mol% C 8 FA, respectively. The results showed that a process comprising lipase reaction and short-path distillation is effective for large-scale preparation of high-purity regiospecific TAG isomers.Much attention has been focused on the effect of position of FA in TAG on physiological activities. Two TAG regioisomers, which have the same FA composition and carry a functional FA at the 1(3)-and 2-position, are necessary in order to study this subject. The two regioisomers have been synthesized by transesterification of long-chain TAG (LLL) with medium-chain FA (MCFA) using a 1,3-position-specific lipase and by transesterification of medium-chain TAG (MMM) with long-chain FA (LCFA) (1-9). The products can reportedly be purified by HPLC (10,11). The process comprising lipase reaction and HPLC is effective for preparing high-purity TAG regioisomers, and the products have been used for examining the intestinal absorption of TAG (12). However, animal tests involving oral administration require larger amounts of TAG regioisomers, and the purification process by HPLC is not suitable for large-scale preparation. Xu and colleagues (7,8) carried out large-scale preparation of TAG regioisomers by short-path distillation, but the resulting preparations were mixtures of MMM, MLM, and MLL because the distillation removed only FFA. Negishi et al. (9) purified MLM from the reaction mixture, but the content of MLM was only 76%. For these reasons, a process for preparing larger amounts of pure TAG regioisomers (purity >90%) is strongly desired.TAG (MLM) with MCFA at the 1,3-positions and LCFA at the 2-position, and TAG (LMM) with LCFA at the 1(3)-position and MCFA at the 2,3(1)-positions, represent a pair of TAG regioisomers. To evaluate nutritional effects ...

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confidence: 99%

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confidence: 99%

Preparation of regioisomers of structured TAG consisting of one mole of CLA and two moles of caprylic acid

Kawashima

Nagao

Watanabe

et al. 2004

“…60% of TAG-CLA with lipase RMP alone. Lipase QLM alone did not catalyze the esterification, producing only 3% TAG-CLA after 30 h. We speculate that the notable improvement obtained by using a blend of two lipases was due to the reactivity of these lipases (14). We also speculate that lipase QLM converted 1(3)-MAG and 1,3-DAG to 2-MAG and 1,2-DAG by interesterification.…”

Section: Resultsmentioning

confidence: 89%

“…Previously, we discussed the application of powdered lipase in interesterification reaction systems (12)(13)(14). We showed that it was possible to use powdered lipase in such reaction systems and to synthesize TAG consisting of mediumand long-chain FA, phytosterol esters, and structured lipids containing PUFA.…”

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confidence: 99%

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Synthesis of triacylglycerol containing conjugated linoleic acid by esterification using two blended lipases

Hirose

Yamauchi-Sato

Arai

et al. 2006

Self Cite

We have developed an efficient esterification for the synthesis of triacylglycerol (TAG) containing conjugated linoleic acids (CLA) using a blend of two powdered lipases. Two pairs of blended lipases promoted the esterification. Rhizomucor miehei lipase plus Alcaligenes sp. lipase and Penicillium camembertii MAG and DAG lipase plus Alcaligenes sp. lipase were used. At the optimal ratio of two lipases, the content of TAG containing CLA (TAG-CLA) in all glycerols reached 82-83% after 47 h using 1 wt% of lipases. With R. miehei lipase plus Alcaligenes sp. lipase, the reaction time to obtain ca. 60% of TAG-CLA was one-third of that needed with R. miehei lipase alone. The optimal ratio of two lipases differed between these two pairs. The optimal ratio was 70-80 wt% of R. miehei lipase in the last stage of the reaction, whereas it was over a wide range of 10-90 wt% for P. camembertii lipase. In the blend of R. miehei lipase plus Alcaligenes sp. lipase, activity remained very high after 10 cycles of esterification (every 47 h) and could be used in the industrial production of TAG-CLA.

Enzymatic Interesterification of Palm Stearin and Coconut Oil by a Dual Lipase System

Ibrahim

Guo

2007

Enzymatic interesterification of palm stearin with coconut oil was conducted by applying a dual lipase system in comparison with individual lipase-catalyzed reactions. The results indicated that a synergistic effect occurred for many lipase combinations, but largely depending on the lipase species mixed and their ratios. The combination of Lipozyme TL IM and RM IM was found to generate a positive synergistic action at all test mixing ratios. Only equivalent amount mixtures of Lipozyme TL IM with Novozym 435 or Lipozyme RM IM with Novozym 435 produced a significant synergistic effect as well as the enhanced degree of interesterification. The interesterification catalyzed by Lipozyme TL IM mixed with thermally inactivated immobilized lipase preparations indicated that the carrier property may play an important role in affecting the interaction of two mixed lipases and the subsequent reactions. A dual enzyme system, consisting of immobilized lipases and a non-immobilized one (Lipase AK), in most cases apparently endows the free lipase with a considerably enhanced activity. 70% Lipase AK mixed with 30% immobilized lipase (Lipozyme TL IM, RM IM and Novozym 435) can achieve an increase in activity greater than 100% over the theoretical value when the reaction proceeds for 2 h. The co-immobilization action of the carrier of the immobilized lipases towards the free lipase was proposed as being one of the reasons leading to the synergistic effect and this has been experimentally verified by a reaction catalyzed by a Lipase AK-inactivated preparation. No apparently synergistic effect of the combinations of Lipozyme TL IM and RM IM was observed when the dual enzyme systems applied to the continuous reaction performed in a packed bed reactor.In brief, this work demonstrated the possibility of increasing the reaction rate or enhancing the degree of conversion by employing a dual lipase system as a biocatalyst.