© 2019 Institution of Chemical Engineers 3-Hydroxyisobutyric acid is an important intermediate in the biosynthesis of methacrylic acid. Its biocatalytic synthesis can be performed by aldolase-catalyzed aldol addition of propanal to formaldehyde followed by an enzymatic oxidation of the resulting 3-hydroxy-2-methylpropanal to 3-hydroxyisobutyric acid. In this work, D-fructose-6-phosphate aldolase D6Q variant was investigated as a key step for the biocatalytic preparation of 3-hydroxy-2-methylpropanal, a commercially unavailable precursor of 3-hydroxyisobutyric acid. The kinetic model of this step was developed for the purpose of reactor selection and process optimization. It was found that enzyme operational stability decay is co-dependent on the initial formaldehyde concentration. Thus, the choice of the initial conditions is crucial for a successful process set-up. It was concluded that fed-batch was the best reactor choice for this reaction due to enzyme inhibition by formaldehyde and propanal, and its operational stability decay. At the optimal process conditions, the product concentration, product yield, and volume productivity after 5.5 h were 72 g L−1, 88.5% and 313.7 g L−1 d−1, respectively. Enzymatic oxidation of 3-hydroxy-2-methylpropanal to the corresponding acid was performed as a proof of concept using an aldehyde dehydrogenase in the presence of NAD+, regenerated by water-forming NADH oxidase, and 2.5 g L−1 (24 mM) of 3-hydroxyisobutyric acid was obtained.
- Aldol addition
- D-fructose-6-phosphate aldolase
- Mathematical model
- Methacrylic acid