Reaction-separation coupling process to purify α-linolenic acid based on the hydratase catalytic kinetics differentiation of flaxseed oil fatty acids

α-Linolenic acid (ALA) is an important omega-3 polyunsaturated fatty acid with wide-ranging applications across various fields, where purity requirements are the key target. This work has investigated a catalytic specific hydratase for different unsaturated fatty acids --- BbMCRA2 from Bifidobacterium breve NCIMB 702258, which could achieve different reaction kinetics for various unsaturated fatty acids to split and purify α-linolenic acid. The reaction conditions (pH, reaction temperature, lyophilized enzyme powder loading and buffer addition) of fatty acids mixtures were optimized in combination with the construction of the resolution mathematical model. Furthermore, under the optimal hydration conditions, a validation of separation under the scaled-up system for flaxseed oil fatty acids was conducted combining a column chromatography. The purity of ALA could be increased from 59% to 92.51%, and the corresponding yield was 79.55% (yield for ALA: 90.28%). The enzymatic method effectively reduces the separation difficulty of ALA in the mixture of unsaturated fatty acids, and could provide an excellent yield and purity of ALA on a larger scale. From this process, a green and efficient bio-enzymatic purification process for ALA with industrial application potential has been proposed.