Biocatalysis of linoleic acid to conjugated linoleic acid

Abstract

CLA refers to a group of geometrical and positional isomers of linoleic acid (LA) with conjugated double bonds. CLA has been reported to have diverse health benefits and biological properties. Traditional organic synthesis is highly capital-intensive and results in an isomeric mixture of CLA isomers. Biotechnology presents new alternatives to traditional lipid manufacturing methods. The objective of this study was to examine the effect of protein isolation procedures on linoleate isomerase (LAI) recovery from microbial cells and biocatalysis of LA to CLA. Protein isolation experiments were carried out using Lactobacillus acidophilus L1 and two strains of Lactobacillus reuteri (ATCC 23272 and ATCC 55739). Under the same assay conditions, ATCC 55739 had the highest LAI activity among the microbial cultures examined in this study. Efficiency of cell lysis methods, which included various combinations of lysozyme and mutanolysin treatments in combination with sonication and osmotic rupture of cells with liquid nitrogen, was very low. Although treatment of cell material with a detergent (octylthioglucoyranoside) freed a significant amount of LAI activity into the solution, it was not sufficient to recover all the LAI activity from the residual cells. Crude LAI preparations produced mainly the cis-9,trans-11 CLA isomer. Time and substrate/protein ratio had a significant effect on biocatalysis of LA to CLA. It appears that the mechanism and kinetics of enzymatic conversion of LA to CLA are quite complex and requires further research using pure LAI preparations.

Isomers of conjugated linoleic acids are synthesized via different mechanisms in ruminal digesta and bacteria

“…To complete the reaction, a hydrogen atom is then abstracted from water. The proposed mechanism may explain why the reaction kinetics of RA formation are so unusual (49,51) and why a cofactor is required (E. Devillard, unpublished data). It is also possible that the formation of other 9,11 conjugated isomers is marginally less energetically favorable than RA for the stabilization of the radical intermediate.…”

“…The mechanism responsible for RA formation is much less clear, partly because of the difficulty of purifying enzymes from B. fibrisolvens (16) and Lactobacillus species (49). A complicated mechanism based on a hydration-dehydration reaction and two successive cis-trans isomerizations was proposed for RA synthesis by L. acidophilus based on the accumulation of 10-hydroxy-cis-12-18:1 during incubations with LA (17,44).…”

Conjugated Linoleic Acid in Milk and Fermented Milks: Variation and Effects of the Technological Processes

“…The linoleic acid ingested in the diet is firstly isomerized to rumenic acid (C18:2 cis9, trans11) by the cis-12, trans-11 isomerase enzyme, and then biohydrogenated to vaccenic acid (C18:1 trans11) in the rumen by the bacteria Butyrivibrio fibrisolvens (10). This anaerobic bacteria is also responsible of the formation of the rumenic and vaccenic acids from the α-linolenic (C18:3 cis9, cis12, cis15) and ɣ-linolenic (C18:3 cis6, cis9, cis12) acids (25). If the initial isomerization of the C18:2 cis9, cis12 acid involves the cis12 double bond, the isomer C18:2 cis9, trans11 is produced, whereas if the initial double bond isomerized is the cis9, the isomer C18:2 trans10, cis12 is mainly produced (11).…”

Conjugated Linoleic and Linolenic Acid Production by Bacteria: Development of Functional Foods

“…Other authors isolated and characterized the LAI from L. reuteri MRS8 [86], showing a molecular weight of more than 100 kDa. In this study, the optimal activity of the enzyme was in the pH range of 4.7 to 5.4.…”