An evaluation of the biocatalytic ability and the potential utilization of Calcium-alginate immobilized hepatic microsomes (beads) for the biosynthesis of amine oxides has been performed, To accomplish this task, the following three areas Were investigated: immobilization, catalytic biooxidation ability, and reactor configurations.

Immobilization was found to result in higher yields over free microsomes at the expense of higher cofactor requirements. Refrigerated drying of immobilized micro-somes appears to be a better storage technique than wet storage in buffer. Smaller beads resulted in higher rates than larger beads, but size did not affect the overall yield.

Chlorpromazine (CPZ), diphenylamine (DPA) and 4-hydroxy-2,2,6,6-tetramethylpiperidine (TMP) were used as substrates to demonstrate the catalytic ability of microsomal enzymes to biooxidize tertiary and hindered secondary amines. The biooxidation of all three compounds to their oxides, by hog liver microsomes, required the presence of NADPH and oxygen. CPZ was converted to its N-oxide while, DPA and TMP were converted to nitroxide free radicals.

Optimal reaction requirements for each substrate were established, NADPH is a reaction limiting factor for the oxidation of all three compounds. Biooxidation of the substrates increased, up to two-fold, by addition of n-tylamine, suggesting that mixed function amine oxidase (MFAO) is responsible for the N-oxidation of the amines.

The progress of the previously described reactions was followed by means of oxygen uptake and HPLC. The identification of CPZ N-oxide was done using NMR, HPLC and TLC. ESR spectroscopy was used to verify the formation of nitroxide free radicals.

A recirculation flow reactor is recommended for large scale production with immobilized whole microsomes. This configuration helps to maintain the physical integrity of the biocatalyst over longer reaction periods. The recirculation flow reactor also gives higher yields than a batch reactor.