DEGRADATION OF MORPHOLINE BY AN ENVIRONMENTAL MYCOBACTERIUM STRAIN INVOLVES A CYTOCHROME-P-450

A Mycobacterium strain (RP1) was isolated from a contaminated activate d sludge collected in a wastewater treatment unit of a chemical plant, It was capable of utilizing morpholine and other heterocyclic compoun ds, such as pyrrolidine and piperidine, as the sole source of carbon, nitrogen, and energy, The use of in situ H-1 nuclear magnetic resonanc e (H-1 NMR) spectroscopy allowed the determination of two intermediate s in the biodegradative pathway, 2-(2-aminoethxyy)acetate and glycolat e, The inhibitory effects of metyrapone on the degradative abilities o f strain RP1 indicated the involvement of a cytochrome P-450 in the bi odegradation of morpholine. This observation was confirmed by spectrop hotometric analysis and H-1 NMR, Reduced cell extracts from morpholine -grown cultures, but not succinate-grown cultures, gave rise to a carb on monoxide difference spectrum with a peak near 450 nm, which indicat ed the presence of a soluble cytochrome P-450, H-1 NMR allowed the dir ect analysis of the incubation medium containing metyrapone, a specifi c inhibitor of cytochrome P-450. The inhibition of morpholine degradat ion was dependent on the morpholine/metyrapone ratio. The heme-contain ing monooxygenase was also detected in pyrrolidine-and piperidine-grow n cultures, The abilities of different compounds to support strain gro wth or the induction of a soluble cytochrome P-450 were assayed. The r esults suggest that this enzyme catalyzes the cleavage of the C-N bond of the morpholine ring.