Substrate selectivity of Novozym 435 in the esterification of glycerol with an equimolar mixture of linoleic, conjugated linoleic, and pinolenic acids

Abstract

The selectivity of Novozym 435, an immobilized Candida antarctica lipase B toward linoleic (LA), conjugated linoleic (CLA), and pinolenic acids (PLA) was investigated in the esterification of glycerol with an equimolar (27.3 mol% each) mixture of the fatty acids (FAs) in a solvent‐free system to prepare triacylglycerols (TAGs) with anti‐obesity effects. The maximal TAG content of 98.9 wt% was achieved under the following conditions: temperature, 70°C; reaction time, 24 h; glycerol‐to‐FA mixture molar ratio, 1:3; enzyme loading, 10 wt% of total substrates; vacuum level, 0.4 kPa. During the initial phase of the reaction, Novozym 435 showed pronounced selectivity in the order of PLA > CLA > LA; however, no significant difference in the selectivity of Novozym 435 toward the three FAs was observed after the reaction plateaued. Novozym 435 showed the significant selectivity toward PLA over CLA and LA at the sn‐2 position of TAGs throughout the reaction. The resulting TAGs had the same total content (∼27 mol% each) of the three FAs. However, the LA, CLA, and PLA contents at the sn‐2 position were 27.7, 24.7, and 30.3 mol%, respectively, indicating that unlike LA, CLA, and PLA were unevenly distributed on the glycerol backbone of the TAGs. Practical applications: CLA and PLA have received much attention because of their anti‐obesity effects. Structured TAGs containing both CLA and PLA not only maintain the health benefits of each but also may provide several advantages (e.g., enhanced oxidative stability, reduced incidence of off‐flavor) to both for food applications compared to the free FA forms. The selectivity of a lipase toward FAs used as substrates for the lipase‐catalyzed synthesis of structured TAGs affects the composition and positional distribution of FAs in the resulting structured TAGs. The present study reports a new finding about the selectivity of Novozym 435 toward LA, CLA, and PLA which are all C18‐unsaturated FAs, in the esterification of glycerol with the FAs to prepare structured TAGs containing both CLA and PLA. Novozym 435‐catalyzed esterification of glycerol with an equimolar mixture of linoleic, conjugated linoleic, and pinolenic acids.

A Simple Enzymatic Process to Produce Functional Lipids From Vegetable and Fish Oil Mixtures

“…For the lipase Novozyme 435, the increase of DHA content might be related to its random behavior and, also, to the 2‐selectivity toward fatty acids with higher degree of unsaturation. Effectively, Woo et al described that Novozyme 435 showed a tendency to esterify the most unsaturated fatty acids at the sn ‐2 when the susbtrate was composed of fatty acids with the same number of carbons but different unsaturation degree. Regarding to TL IM, it might be feasible that, in such a low water content and being 1,3‐DAG the main substrate, the esterification in the central position also takes place.…”

A Structured Pine Nut Oil Has Hypocholesterolemic Activity by Increasing LDLR Gene Expression in the Livers of Obese Mice

“…An FFA fraction containing ≈13 wt% PLA was prepared by saponification of the NPNO according to the method described in our previous study . NPNO (100 g) was saponified with 400 mL of 10 wt% NaOH in 75 vol% ethanol.…”

“…An FFA fraction containing ≈44 wt% PLA was prepared via urea crystallization of the FFA containing ≈13 wt% PLA obtained above, following the method described in our previous study . The FFA fraction (50 g) containing ≈13 wt% PLA and urea (150 g) were both dissolved in methanol (500 mL), and the resulting mixture was refluxed at 250 °C until the urea was dissolved completely.…”

Preparation of Pinolenic Acid Concentrates from Pine Nut Oil Fatty Acids by Solvent Fractionation

“…From the experimental results in sections 3.2 and 3.3, we concluded that the optimal conditions for the preparation of PLA concentrates from the FFA fraction obtained from pine nut oil are a temperature of 85 , an n-hexane-to-FFA ratio of 30:1, and a fractionation time of 36 h. Under these conditions, PLA concentrates containing 69.8 wt PLA were obtained with a yield of 77.4 wt based on the initial weight of PLA in the FFA fraction. Woo et al 5 used urea crystallization to prepare PLA concentrates containing 47 wt PLA with a yield of PLA of 77 wt from the FFA fraction obtained from Korean pine nut oil with a PLA content of 13 wt . Lee et al 6 and Zhao et al 7 found that ethyl ester forms of PLA concentrates can be prepared by C. antarctica lipase B-catalyzed ethanolysis of pine nut oil.…”

“…Finally, as shown in Table 1, with the exception of 20:0, 11 different species of FA contained in the original FFA fraction remained in the PLA concentrates. Woo et al 5 reported that PLA concentrates prepared by urea crystallization from Korean pine nut oil containing 13 different species of FA only contained 7 species of FA, despite the relatively low PLA content of 47 wt . In particular, all SFA i.e., 16:0, 18:0, and 20:0 were eliminat-ed from the PLA concentrates.…”