Continuous production of L-carnitine with NADH regeneration by a nanofiltration membrane reactor with coimmobilized L-carnitine dehydrogenase and glucose dehydrogenase

L-Carnitine dehydrogenase (CDH) was partially purified from Pseudomonas put ida IAM12014 for the stereospecific reduction of 3-dehydrocarnitine to L-ca rnitine. CDH and glucose dehydrogenase (GDH) were coimmobilized in a nanofi ltration membrane bioreactor (NFMBR) for the continuous production of L-car nitine from 3-dehydrocarnitine with NADH regeneration. In the NFMBR, NAD wa s partially immobilized through rejection by the nanofiltration membrane an d effectively regenerated by the conjugation reaction of CDH and GDH. Since 3-dehydrocarnitine was unstable at neutral pH, it was maintained under aci dic conditions (pH 0.7) and supplied to the NFMBR separately from the other substrates, glucose and coenzyme NAD. As 50 mM 3-dehydrocarnitine in HCl s olution, 0.05 mM NAD, and 100 mM glucose in 0.5 M Tris buffer (pH 8) were c ontinuously supplied to the NFMBR with immobilized CDH (200 U/ml) and GDH ( 200 U/ml) at the retention time of 80 min and temperature of 25 degrees C, the maximum conversion, reactor productivity, and NAD regeneration number w ere 78%, 113 g/l/d, and 780, respectively. The half-life of the NFMBR was l onger than 500 h.