Reduction of Prochiralic Ketones by NAD(H)- dependent Alcohol Dehydrogenase in Membrane Reactor

Chiral alcohols are intermediates of organic synthesis protocols for attaining flavors, pheromone, phytohormone and therapeutic molecules. This work dealt with the conversion of acetophenone and hexanone into1-phenyl ethanol and 2-hexanol, respectively, by NAD(H)-dependent alcohol dehydrogenase (ADH) in a membrane reactor (MR) operated in a continuous regimen, there being at the bottom a nanofiltration membrane (MWCO 500Da). The conversions, in which the ethanol was employed as a co-substrate, were carried out at 100rpm, a feeding rate of 5mL/h, ADH (150U), temperature (30ºC), pH (8.8), initial substrate concentration (30mM) and β-NAD/β-NADH (0.9mM). The ADH activity as well as the factors affecting it (temperature and initial substrate concentration) was measured through the consumption or formation of β-NADH at 340nm. The optimum temperature for 5min of reaction was 45oC. The Michaelis-Menten constant related to the action of ADH on NADH, acetophenone and 2-hexanone were (KM)NADH = 0.177mM, (KM)acetophenone = 1.04mM and (KM)2hexanone = 1.44mM. Along the reaction, the NAD/NADH regeneration occurred and the membrane used retained both the ADH and the coenzymes (NAD; NADH) inside the reactor. Finally, the acetophenone/1-phenylethanol and 2-hexanone/2-hexanol conversions through the MR were 60% and 30%, respectively.