Date of Award

Spring 1989

Document Type

Thesis

Degree Name

Master of Science in Chemical Engineering - (M.S.)

Department

Chemical Engineering, Chemistry and Environmental Science

First Advisor

Samir S. Sofer

Second Advisor

Piero M. Armenante

Third Advisor

Dekang Shen

Abstract

An investigation of the properties of calcium alginate immobilized hog liver microsomes, rich in FAD-containing mixed function oxidase, was performed. Studies involving catalyst storage, immobilization of cofactors within the matrix, and alginate gel concentration were performed using a 1.8 ml mixed flow micro assay reactor. A recirculation flow reactor was designed and used for the biochemical production of 2,2,4,4-tetramethyl-1,3-oxazolidine-l-oxyl, a potentially useful MRI contrast agent.

The activity of the immobilized microsomes was found to increase over time when stored in CaCl2 or in buffer at 4-8°C. The increase in activity is a result of the removal of inherent inhibitors present in the microsomal preparation. The cofactors NADP+ and G6P and coenzyme G6PDH can be immobilized within the alginate matrix, however significant losses will result from diffusion of the cofactors during the catalyst preparation. The rate of diffusion through the matrix is a function of molecular weight, NADP+ being the largest cofactor is retained the most. The alginate gel produces a uniform resistance to mass transfer between 0.5 and 1.5 % gel. The rate of oxygen consumption is found to be a linear function of the enzyme concentration. The immobilized enzyme has an effective Vmax of 10.4 nmole/min * mg protein.

A recirculation flow reactor was designed for the use of immobilized microsomes. The nitroxide 2,2,4,4- tetramethyl-1, 3-oxazolidine-l-oxyl was produced enzymatically using 2,2,4,4-tetramethyl-1,3-oxazolidine as substrate. The product was compared to the same compound synthesized organically, using UV/VIS and HPLC. The formation of the nitroxide free radicals is verified by ESR spectroscopy.

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