Study in UF-membrane reactor on activity and stability of nitrile hydratase from Microbacterium imperiale CBS 498-74 resting cells for propionamide production

The bioconversion of propionitrile to propionamide was catalysed by nitrile hydratase (NHase) using resting cells of Microbacterium imperiale CBS 498-74 (formerly, Brevibacterium imperiale). This microorganism, cultivated in a shake flask, at 28 degreesC, presented a specific NHase activity of 34.4 U mg(DCW)(-1) (dry cell weight). The kinetic parameters, K-m and V-max tested in 50 mM sodium phosphate buffer, pH 7.0, in the propionitrile bioconversion was evaluated in batch reactor at 10degreesC and resulted 21.6 mM and 11.04 mumol min(-1) mg(DCW)(-1), respectively. The measured apparent activation energy, 25.54 kJ mol(-1), indicated a partial control by mass transport, more likely through the cell wall. UF-membrane reactors were used for kinetic characterisation of the NHase catalysed reaction. The time dependence of enzyme deactivation on reaction temperature (from 5 to 25 degreesC), on substrate concentrations (from 100 to 800 mM), and on resting cell loading (from 1.5 to 200 mug(DCW) ml(-1)) indicated: lower diffusional control (E-a = 37.73 kJ mol(-1)); and NHase irreversible damage caused by high substrate concentration. Finally, it is noteworthy that in an integral reactor continuously operating for 30 h, at 10 degreesC, 100% conversion of propionitrile (200 mM) was attained using 200 mug(DCW) ml(-1) of resting cells, with a maximum volumetric productivity of 0.5 g l(-1) h(-1). (C) 2004 Elsevier B.V. All rights reserved.